Advancements in Functional Polymeric Scaffolds for Scar-Free Skin Regeneration

Abstract

Skin tissue engineering offers a promising approach to improving wound healing outcomes, with polymeric scaffolds playing a critical role in guiding tissue regeneration and minimizing scar formation. This review summarizes recent advancements in polymeric scaffold design for skin tissue engineering, specifically emphasizing strategies that promote functional regeneration rather than fibrotic repair. Key aspects discussed include scaffold architecture, polymer selection, and fabrication techniques that influence cellular behavior, extracellular matrix (ECM) deposition, and mechanical integration with the host tissue. The review highlights how natural, synthetic, and composite polymer systems can be engineered to modulate inflammation, support cell adhesion and differentiation, and facilitate organized tissue remodeling. Translational considerations are examined, including cell-seeding approaches and the role of cellular self-organization in supporting the regeneration of skin appendages and restoring the native tissue architecture. Currently available skin substitutes are critically evaluated, revealing persistent limitations in achieving a scar-free healing. Emerging fabrication technologies, particularly three-dimensional printing, electrospinning, and artificial intelligence-guided design, are all discussed for their potential to enhance scaffold precision and regenerative performance. Overall, this review underscores the importance of continued innovation in polymer chemistry, materials engineering, and bioprinting to enable the development of next-generation skin scaffolds that support scar-free skin regeneration.

1. Introduction

The human skin is the largest organ in the body by area, covering up to 2 square meters and accounting for approximately one-sixth, or 15%, of the total body weight. The skin acts as a barrier between the human body and its surroundings, serving multiple functions, including protection, sensation, temperature regulation, metabolic and biochemical processes, and immune system support. In addition to protecting against physical, chemical, and biological threats, the skin regulates the body’s water content, preventing excessive evaporation, which helps maintain thermal balance.

Wounds are often a clear indication of a disruption in the integumentary system. Clinicians describe wound healing as a complex and highly organized process involving various molecular and cellular mechanisms. As shown in Figure , wounds heal in an instructed manner, progressing through four distinct but overlapping stages to achieve closure: hemostasis, inflammation, proliferation, and remodeling. Hemostasis, which involves the formation of a blood clot to stop bleeding, occurs immediately after an injury. Following injury, immune cells such as neutrophils and macrophages are attracted to the area to clear debris and combat infection, initiating the process known as inflammation. Subsequently, during the proliferation phase, fibroblasts and endothelial cells proliferate, leading to the formation of granulation tissue and re-epithelialization of the wound bed. Lastly, the remodeling phase involves the maturation and restructuring of newly formed tissues, restoring their strength and function. Understanding the different types of wounds and their respective healing processes is crucial to developing effective therapeutic approaches. Additionally, wounds can be classified according to their depth, ranging from full-thickness dermal or subcutaneous wounds to minor epidermal injuries. Acute wounds result from sudden trauma or injury and typically heal through the four-stage process. Examples include abrasions, cuts, and surgical incisions. On the other hand, chronic wounds take longer to heal and are often caused by underlying medical conditions such as pressure ulcers, diabetes, venous insufficiency, or burns. , Clinicians face numerous challenges when treating chronic wounds, including diabetic foot ulcers and venous leg ulcers, due to their complex pathophysiology and tendency to develop complications such as infection and delayed healing. The biggest challenge with chronic wounds is their inability to close on their own, as numerous factors can obstruct one or more stages of the four-stage healing process, leading to improper or compromised healing. Contributing factors include age, gender, lack of oxygen, infection, stress, diabetes, obesity, medications, alcoholism, and smoking. Chronic wounds are estimated to affect about 2% of the US population and can significantly reduce quality of life while raising numerous complications and exorbitant medical costs. Thus, addressing this issue is crucial for public health and well-being, especially as the nation experiences an increase in cases of chronic wounds.

1.

While acute wounds successfully transition from an initial inflammatory response to tissue remodeling, chronic wounds are defined by a pathological “persistent inflammatory phase.” This arrest prevents essential proliferation and remodeling stages, resulting in failure to achieve functional tissue closure.

As summarized in Figure , while scar tissue forms naturally during wound healing, skin-substitute-supported tissue regeneration offers several important advantages. The resulting regenerated tissue closely recapitulates the native skin in both structure and function, restoring essential properties, such as barrier integrity, sensibility, and elasticity. In contrast, scar tissue lacks these capabilities since it is primarily composed of densely aligned collagen fibers, resulting in reduced mechanical compliance, impaired sensory function, and limited biological functionality. Aesthetically, enhanced tissue regeneration often leads to minimal or scarless healing, preserving the natural appearance and mitigating excessive fibrosis, unlike the unsightly appearance of the scar tissue. Regenerated tissue also recovers mechanical strength, enabling it to withstand normal levels of mechanical stress, deformation, and pressure, whereas scar tissue is weaker and more functionally restricted. Furthermore, regenerated tissue maintains proper cellular organization, which is imperative for coordinated function, unlike that of scar tissue. Effective tissue regeneration reduces the risk of complications such as contractures and impaired organ function, thereby supporting long-term health and helping to prevent chronic diseases.

2. Ideal Scaffold Properties for Skin Regeneration

Skin tissue engineering is a fascinating emerging field that integrates principles from engineering, biology, material science, and regenerative medicine to construct artificial skin that replicates the structure and function of natural skin. It offers a promising solution by providing personalized scaffolds loaded with cells that promote tissue regeneration and wound closure. In addition to serving as temporary wound dressings, these engineered skin substitutes promote the body’s natural healing processes, ultimately improving patient outcomes. The ideal for wound healing is lofty, with “scarless” or “scar-free” conditions in regenerated skin highly sought after in both experimental procedures and commercial applications. This optimistic designation can be clarified by its resolution into three crucial aspects. Firstly, the aesthetics of a regenerated wound site play an important role in restoring quality of life post-injury while ensuring complete, first-rate healing. The flatness and smoothness of a scar indicate a balanced and coordinated deposition of Type I and III collagen fibers, along with elastin fibers, in the regenerating site. Similarly, the absence of wrinkles and raised nodules signify a well-regulated process of wound contraction and closure. Moreover, a naturally colored scar area belies a superb degree of wound remodeling, as melanogenesis and re-pigmentation have been documented to be executed either by newly synthesized melanocytes produced by regenerated hair follicles or by migratory melanocytes originating from the wound edges. Secondly, various biological criteria must be met in a satisfactory manner in the healed injury site. Broadly, these include (1) vital histological conditions such as the Types I and III collagen balance (in healthy skin the ratio of Types I to III collagen is 2:1–3:1), the appropriate modulation of vital fibroblast activity markers, especially of alpha-smooth muscle actin (α-SMA) and transforming growth factor-beta (TGF-β), and the reformation of an anisotropic microstructure containing aligned collagen and elastin fibers within the newly formed ECM; (2) the activation and regulation of signaling pathways dictating the progression of fibrosis platelet-produced expressions of TGF-β1 and TGF-β3 are relayed to fibroblasts via SMAD signaling proteins, triggering their differentiation into myofibroblasts that secrete ECM components, while mechanotransductive integrins and the mechanosensitive Yes-associated protein/transcriptional co-activator with PDZ-binding motif (YAP/TAZ) pathway may pose a corresponding influence upon sensing an increase in stiffness in the surrounding tissue as collagen deposition occurs, perpetuating the generation of fibrotic tissues; and lastly, (3) the mitigation of potent immunogenic phenomena, especially the polarization of macrophages from their pro-inflammatory M1 phenotype into their pro-regenerative M2 phenotype at a pivotal point during the stages of healing. Consequently, analyzing the expression of various biomarkers expressed during the progression of these intricate processes is crucial for validating the production of a near-scarless or scar-free outcome. Thirdly, the definition of scarless/scar-free healing can be expanded to include the restoration of the functions and mechanical properties of the native skin, along with the regrowth of fundamental appendages such as nerves, hair follicles, and sweat and sebaceous glands, since these are hallmarks of advanced regeneration levels. The recovery of the skin’s functions also significantly contributes to quality of life in terms of long-term comfort, as the skin facilitates sensation, perspiration/thermoregulation, and the formation of an antibacterial acid mantle. Hereafter, essential scaffold properties and components (Figure ) will be briefly discussed for their role in promoting an optimal standard of wound healing through their interaction with the biological elements outlined above.

2.

Mechanistic pathways of scaffold properties in driving wound healing outcomes. Ideal scaffold properties (top) facilitate pro-regenerative cellular environments, including healthy angiogenesis and ordered collagen assembly, for “scarless” recovery. In contrast, inappropriate scaffold features (bottom) hinder neuro-angiogenesis and drive excessive fibrotic responses, leading to nonfunctional, scarred tissue.

2.1. Biocompatibility

Biocompatibility is the most critical criterion for scaffold design, constituting the relation of a biomaterial with its biological environment. Intrinsic requirements for the biocompatibility of engineered skin grafts include the ability of cells to adhere and proliferate as well as the absence of immunological rejection. Mitigating the host immune response is vital for preventing inflammation, scarring, or other complications that would interfere with optimal skin regeneration. Macrophage activity drives the immunological response to injury, and the expression of their pro-inflammatory M1 phenotype is both natural and necessary within the first few days after a burn or wound is sustained. M1 macrophages secrete various cytokines, such as tumor necrosis factor-alpha (TNF-α), Interleukins-1 beta and 6 (IL-1β and IL-6), and reactive oxygen species (ROS), that expedite the removal of cell and ECM debris. Subsequently, IL-3 and IL-4 mediate the transition of the wound environment into the proliferative stage of wound healing and initiate polarization toward the expression of the M2 macrophage phenotype, which in turn secretes pro-regenerative cytokines as TGF-β and vascular endothelial growth factor (VEGF) to recruit collagen-producing fibroblasts and commence the formation of new blood vessels (angiogenesis), respectively. Another antifibrotic cytokine, IL-10, also plays a major role in both the inflammatory and remodeling phases by suppressing pro-inflammatory cytokines and hindering excessive collagen deposition. However, the M1-to-M2 polarization may be prevented by aging, diabetes, autoimmune diseases, or the perception of a biomaterial construct as a foreign element due to a mechanical, biological, or chemical mismatch, resulting in the persistence of chronic inflammation and delaying wound reconstruction. Additional catalysts for immune rejection may include other intrinsic properties of a biomaterial itself, involving its potential for carrying or generating excessive amounts of ROS buildup on its surface, impairing its function, and disrupting convalescent work in the surrounding injured tissues. − Therefore, adequate replication of the fundamental structural, mechanical, biological, and bioelectrochemical properties of the ECM within native skin tissue is the hallmark of successful skin tissue engineering endeavors. Additionally, a biomaterial’s potential for promoting nonspecific fouling (the unwanted accumulation of excessive proteins, cells, or bacteria on the surface of an implant or wound dressing) must be efficiently reduced through strategies such as the implementation of antifouling coatings or creation of surface modifications, which have been described in recent work. Superior biocompatibility is exhibited in vivo by most naturally occurring biomaterials, including collagen, gelatin, hyaluronic acid, and elastin, while synthetic and conductive biomaterials require some characteristic modifications to increase their acceptability to the body and reduce the likelihood of an immune response. Naturally occurring biomoleculesi.e., polyphenols, tannins, enzymes have proven themselves to be excellent neutralizing agents of ROS through redox reactions and other biochemical means, producing scarless outcomes when incorporated into dressings applied to wounds. Therefore, it often becomes necessary to combine natural and synthetic/conductive biomaterials into an appropriately balanced mixture to manufacture a suitable scaffold that meets the conditions for biocompatibility. Additionally, the use of various immunomodulatory cytokines in engineered skin substitutes has been reported. For instance, Nie et al. (2026) investigated the use of interferon-γ (IFN-γ) in combination with urine-derived stem cells and encapsulated in a decellularized small intestine submucosa hydrogel to induce M2 macrophage polarization through modulation of the IL-17 pathway. Zou et al. (2025) reported the application of apoptotic extracellular vesicles derived from IL-10-treated fibroblasts in a collagen scaffold to inhibit the fibrogenetic Hedgehog signaling pathway. Furthermore, Chen et al. (2021) utilized IL-10 nanoparticles in an electrospun hyaluronic acid scaffold to polarize macrophages into their M2 phenotype.

2.2. Biodegradability

The biodegradability of a grafted biomaterial is a vital factor in the regeneration process of a skin wound. The rate of scaffold degradation and the rate of tissue growth must be synchronized to prevent gaps and the formation of fibrotic tissue and sustain the structural and biological integrity of regenerating tissue during the healing process. Overly rapid degradation can hinder the formation of a robust and complete tissue, while overly slow degradation can result in incomplete or imperfect fibrous tissue formation around the scaffold, impeding the regeneration process. , Thus, the selection of biomaterials used in the manufacture of a skin substitute is a key factor in regulating biodegradation. Some soft hydrogels have been shown to be susceptible to enzymatic/oxidative degradation, exhibiting increased degradation rates and rapid loss of mechanical performance, , while others degrade stably over long-term observation periods, constitute stable, advanced platforms for drug delivery and immunotherapy that closely mimic the microstructure and mechanical properties of the implementation site while promoting enhanced regeneration through their composition and design. , Comparatively, scaffolds constructed from synthetic-natural polymer blends demonstrate tunable, controlled degradation rates over extended periods, ensuring excellent cell adhesion, migration, and proliferation throughout the duration of wound closure. −

2.3. Microstructure

Porosity is another important bulk property for cell infiltration and nutrient exchange. A porous structure is crucial to scaffold integration, providing voids and interconnected channels that facilitate the migration of fibroblasts, endothelial cells, and keratinocytes into the scaffold, as well as the diffusion of oxygen and nutrients into the scaffold from the surrounding native tissues. − A study involving the use of a tri-layered nanofiber scaffold with 0.3 μm internal pores and 2–4 μm inter-fiber pores showed promising in vitro fibroblast adhesion and NIH-3T3 cell viability while also demonstrating high antibacterial resistance. Another tri-layered construct with an average pore size of 1.5–2.2 μm presented a high saturation of ECM, keratinocytes, fibroblasts, and hair follicle bulge-derived stem cells after a 14-day culturing period. These studies and others , prove that porous, multilayered scaffolds with varying pore sizes according to dermal layers are a successful development in skin tissue engineering. Larger average pore sizes of 93 μm and 200–400 μm , were also reported to be successfully implemented in experiments, featuring good cell activity, ECM deposition, and angiogenesis. Therefore, it has been concluded for skin tissue engineering applications that smaller pores (≤2 μm) promote cell adhesion through close mimicry of the epidermal topography, while intermediate pore sizes (2–12 μm) allow for proper cell/nutrient migration and ECM deposition, with larger pores (≤100 μm) being better suited for structural integrity and hair follicle support. Lastly, scaffold microstructure has also been proven to be a significant determinant of timely revascularization and anastomosis, and has been used with cell-rich hydrogel layers to further enhance graft/scaffold integration and angiogenic properties supportive of accelerated skin regeneration. ,

2.4. Mechanical Properties/Anisotropy/Stiffness

Furthermore, engineered scaffolds should replicate the mechanical properties of the native skin tissue to maintain structural integrity, foster cell and macrophage activity, support long-term functionality, and prevent biomechanical disturbance. In vivo biomechanical examination of human skin layers demonstrates a pronounced stiffness gradient, with the epidermis displaying the greatest Young’s modulus, followed by the dermis, and the hypodermis, which is notably softer. A study utilizing broadband Rayleigh-wave optical coherence elastography (OCE) reveals that the epidermis, encompassing the stratum corneum, possesses a Young’s modulus of roughly 4 MPa. In contrast, the dermis and hypodermis have lower values of 40 and 15 kPa, respectively. It is worth noting that the anatomical site and the testing methods significantly affect the Young’s modulus values. For instance, tensile testing commonly generates maximum values, such as 83.3 MPa on the posterior side. Indentation techniques often yield significantly lower stiffness values, ranging from 10.4 to 89.4 kPa in the lower leg, because they predominantly measure thickness-mode responses. Suction techniques result in intermediate values, ranging from 130 to 260 kPa on the cheek, owing to their efficacy in both in-plane and conventional loading conditions. Additionally, human skin is an anisotropic, , nonlinear complex composite that behaves very stiffly when subjected to tension but offers very little load when subjected to compression. This anisotropy stems from a property known as Langer’s Lines, which describes the orientation of collagen fiber (75% of dry skin weight) and elastin fiber (0.1% of dry skin weight) in the dermis. Thus, the functional mechanical properties of skin, such as elasticity, strength, and resistance to deformation, are dictated by this intricate subcutaneous microarchitecture. , Consequently, restoration of this mechanical anisotropy by replicating the structure of the original ECM is a required factor in the regeneration of functional tissue. It has been observed that scaffolds with anisotropically arranged fibers simultaneously decreased concentrations of cluster of differentiation 86 (CD86), a surface marker for inflammatory M1 macrophages, and increased concentrations of cluster of differentiation 163 (CD163), a surface marker for anti-inflammatory M2 macrophages, while regulating the presence of the Nod-Like Receptor Family Pyrin Domain Containing 3 (NLRP3) inflammasome, influencing a gradual increase in the polarized expression of the M2 macrophage phenotype. Moreover, anisotropy in a micropatterned chitosan film covered with (30 μm × 30 μm) grooves induced alignment in skin precursor-derived Schwann cells and expression of various Schwann cell-associated genes, including c-Jun, neurotrophin receptor p75 (p75NTR), and neural cell adhesion molecule (NCAM), along with 8 pro-neuroregenerative cytokines, galectin-3, eotaxin, Notch-2, neurophilin-2, galectin-1, ciliary neurotrophic factor (CNTF), vascular endothelial growth factor A (VEGF-A), and β nerve growth factor (β-NGF), in skin precursor-derived Schwann cells, including those associated with axon extension and neuron development. Furthermore, the application of the film to damaged motoneurons resulted in the expression of motoneuron-specific markers Neurofilament 200 kDa (NF200) and Choline Acetyltransferase (ChAT), with axon sprouts, neurites, and nerve branches observable after 24 h due to contact between the damaged motoneuron nerves and the scaffold’s anisotropic microtopography. Additionally, other recent studies demonstrated that fiber anisotropy within scaffolds contributed to faster wound closure rates, − to deterring bacterial/microbial presence, , to increasing fibroblast adhesion and proliferation, and to reducing blood loss, all of which are extremely beneficial to wound healing. Finally, parity between the stiffness of skin and the stiffness of a biomaterial construct is required for a wound environment conducive to scar-free healing. There is a rapidly increasing burden of proof that stiffness and tension in the skin drive the progression of fibrosis through various mechanisms involving the FAPα-AKT pathway, the Piezo1-Wnt2/Wnt11-CCL24 mechanotransduction feedback loop, and ELK1 and ERK signaling. Understanding these principles, Lin et al. (2026) created a silk fibroin methacryloyl/β-sheet-rich silk nanofiber hydrogel with pH-tunable mechanical properties (at pH values of 10.5, 9.9, 9.2, 9.0, 8.9, and 7.4, the Young’s moduli were 1.79 ± 0.94, 5.44 ± 4.21, 14.20 ± 8.99, 42.12 ± 24.45, 68.18 ± 25.31, and 139.04 ± 41.62 kPa, respectively). It was observed that the hydrogel with the stiffness value of 1.79 ± 0.94 kPa enhanced re-epithelialization rates, produced minimal scarring visually, and facilitated organized ECM remodeling with a collagen Type I/Type III ratio of 1.42 ± 0.06 and Young’s modulus of 10.03 ± 4.41 kPa in the regenerated skin. Additionally, the hydrogel enhanced the angiogenetic process in vivo (confirmed through CD31 and α-SMA staining), expedited skin appendage regeneration (validated through extensive proteomic/protein expression analysis), and promoted early polarization to the M2 macrophage phenotypeM2 markers CD45, F4/80, CD11b, and CD206 increased in concentration and peaked within 7 days after implementationwith expressions of the M2 phenotype noted to increase as the stiffness of the hydrogel decreased. Most importantly, evaluation of the mechanotransductive YAP and Peizo1 pathways through Western Blotting and immunofluorescence staining revealed that low-stiffness hydrogels significantly downregulated expressions of activity in both pathways while greatly increasing phosphorylated YAP expressions, confirming that ECM/skin substitute stiffness plays a crucial role in modulating fibrosis. Various studies that reported the stiffness of implemented biomimetic skin substitutes alongside scarless healing are reported in Table .

1. Mechanical Properties of Select Engineered Skin Constructs and Their Biological Contributions to Scarless Healing.

Biomaterial Construct	Construct Properties	Biological Impact	Wound Healing Results	Reference
3D-printed gelatin methacrylate (GelMA) hydrogels	Young’s modulus: 3 kPa, Porosity: 23–70.4%	α-SMA, fibronectin 1, Type III collagen expressions significantly downregulated regardless of TGF-β1 presence.	In vitro testing only; potential for scarless regeneration noted.
Electrospun curcumin-loaded poly (octanediol-co-citric acid) gelatin nanofibrous mats	Young’s modulus: 57.12 kPa	Inhibited Extracellular Matrix Metalloproteinase Inducer expression; modulated fibroblast activity; facilitated organized laminin, collagen deposition.	In vitro, in vivo (chronic wound mouse model, hypertrophic scarring porcine model) testing reported; promoted scarless healing through biomimetic mechanical properties; wound area ≤5% by day 15.
Freeze-dried oxidized hyaluronic acid/alginate hydrogels loaded with Interleukin-10	Young’s modulus: 12.5 ± 1.8 kPa, Pore size: 280–400 μm2	α-SMA, TGF-β1, FAK, YAP expressions downregulated; TGF-β3 expressions upregulated; mechanotransduction gene expressions greatly downregulated; facilitated balanced, organized Types I, III collagen deposition.	In vitro, in vivo (rabbit ear wound model) testing reported; higher wound healing/closure rate; Scar Evaluation Index value = 1 (normotrophic/near-scarless appearance) by day 25.
Hyaluronan/vitamin E/dopamine/β-cyclodextrin wearable biomimetic film	Tensile strength: 0.13–2.08 MPa, Tensile modulus: 0.72 ± 0.03 MPa	α-SMA, TGF-β1, TNF-α expressions downregulated; modulated fibroblast activity; reduced inflammatory response; facilitated moderate and organized, Types I, III collagen deposition.	In vitro, in vivo (full-thickness skin defect rat model) testing reported; full wound closure by day 14; normotrophic appearance by day 30.
Silk fibroin methacryloyl/β-sheet-rich silk nanofiber hydrogel	Young’s modulus: 1.79 ± 0.94 kPa	Activity of YAP and Peizo1 pathways significantly downregulated; facilitated Types I, III collagen deposition with a ratio of 1.42 ± 0.06; influenced M2 macrophage polarization.	In vitro, in vivo (C57BL/6J mouse model) testing reported; promoted scarless healing with closure rates of 14% closure by day 3, 51% by day 7, and complete re-epithelialization by day 14.
GelMA/liposome-encapsulated tetrahydro curcumin/liposome-encapsulated hepatocyte growth factor hydrogel	Young’s modulus: 4.32 kPa, Compression modulus: 0.39 MPa	α-SMA, TGF-β, TGF-β/Smad, P-Smad3/Smad3, fibronectin 1, HMGB1/RAGE/NF-kB, COL1A1 expressions significantly downregulated; facilitated organized collagen deposition.	In vitro, in vivo (full-thickness wound rat model) testing reported; wound healing rate = 95.83% and wound area of 0.06 ± 0.02 cm2 by day 14.

2.5. Thermal Properties

The skin plays a role as the primary heat transfer medium in and out of the human body and has been proven to exhibit temperature fluctuations of 0.2–0.5 °C at deep tissue injury sites. This necessity for thermal stability in engineered skin tissue scaffolds has been met by novel use of various nanoscale biomaterials. − Beyond this, recent publications have demonstrated that scaffolds composed of thermoresponsive nanofibers and inorganic biomaterials with photothermal properties were used in combination with external temperature-based conditions to respectively promote fibroblast differentiation, marked by upregulated TGF-β and α-SMA gene expression, cytoskeletal remodeling (marked by mRNA expression of Myosin II and RhoA), and eliminate bacteria while promoting vascularization, speeding up the healing process of skin wounds in both instances. Additionally, thermoresponsive hydrogels have been shown to promote angiogenesis and accelerate wound closure, marked by expressions of HIF-1α, CD31, and VEGF, through gradual temperature-responsive contractile deformation that mechanically facilitates wound apposition and closure. Summarily, these exploitations of the thermal properties of biomaterials present an interesting development for the field of skin tissue engineering.

2.6. Bioelectric Properties

Similarly, effective synthetic skin grafts should possess a certain degree of electrical conductivity. Post-injury, hypoxia creates an ionic flow gradient at the wound site, generating an endogenous electric field that fluctuates throughout the healing process, which functions as a biological “battery” that guides cells and molecules to facilitate wound regeneration and closure. Recent work reports the value of this voltage gradient to be between 10 and 60 mV. Upon injury to the skin, a flux of 1–10 μA/cm² is created over the wound, generating a field of intensity 40–200 mV/mm. This skin wound bioelectric field regulates regeneration in a multiplicity of ways, orienting cell migration of fibroblasts, keratinocytes, and macrophages, distributing collagen cations, glycosaminoglycan anions, and other charged molecules through charge balances, maintaining microcirculation, and promoting angiogenesis. Therefore, harnessing the potency of the bioelectric phenomenon has become a point of increasing interest in the world of regenerative tissue engineering, with intrinsically piezoelectric or charged biomaterials used to fabricate constructs that either generate their own electrochemical voltage or integrate favorably with the native skin’s electroconductive gradient. Fascinatingly, an electrospun polydioxanone (PDX)/poly­(hydroxybutyrate-co-valerate) (PHBV) scaffold at a 20/80 ratio was demonstrated to contribute to scarless skin tissue regeneration by its zeta potential (piezoelectric activity-generated surface charge) values, which fell between the thresholds of 0.168 ± 0.27 mV for pure PHBV scaffolds and 17.4 ± 1.6 mV for pure PDX scaffolds. When assessed in vitro under diabetic and nondiabetic conditions with regard to myofibroblasts, L929 fibroblasts, and reticular fibroblasts, which expedite fibrosis, it was revealed that the PDX/PHBV scaffolds facilitated high levels of fibroblast proliferation through their hydrophilic nature, while their sturdy elastic moduli (71.9 ± 6.0 MPa) resisted the fibroblast-induced contraction/deformation of the injury site that is responsible for uneven scar tissue formation. The surface charges of the scaffold also participated in contraction resistance, while strategically distributing adsorbed proteins and ions and piezoelectrically responding to fibroblast movement to create net dipoles that influenced fibroblast behavior to favor elongation. Additionally, PDX/PHBV scaffolds greatly augmented fibroblast migration for accelerated ECM deposition, as well as angiogenesis and neovascularization when cultured in tandem with endothelial Ea-hy926 cells, with the former secreting VEGF-A, TGF-β, and platelet-derived growth factors (PDGF), leading the endothelial cells to adopt “tip” and “stalk” phenotypes and commence building a substantial vascular network between the cell population. In addition, in vivo testing of the PDX/PHBV scaffolds on Wistar rats displayed excellent outcomes, exhibiting superb biocompatibility, biodegradability, and conduciveness to cell infiltration, leading to the rapid formation of a thick layer of fibroblasts, macrophages, and collagen. Consequently, scarless, fully healed wounds were noted by day 17, with near-natural skin thicknesses, appropriately dense collagen fibrils, and the presence of follicles and sebaceous glands in the regenerated injury sites reported by day 28. Besides piezoelectric scaffolds, other skin substitutes imbued with passive concentration-dependent bioelectric properties have also been used to produce noteworthy outcomes through the electrochemical recruitment of vital molecules and modulation of cellular behavior. A polypyrrole (PPY) hydrogel developed by Xiong et al., possessing a voltage potential range of roughly 0.5–0.6 V when tested with the application of a 100–200 mA current range, was shown to promote platelet adhesion and increase the attraction of Fe³⁺ ions to injury sites, enhancing neovascularization and hair follicle regeneration during the skin healing process. When coupled with external “hot-spring” thermal conditions (30–45 °C), the hydrogel was further stated to have promoted angiogenesis and fibroblast activity. Another PPY hydrogel of conductivity 14–36 mS/cm greatly decreased hemolysis ratios and supported coagulation activity, its conductive nature influencing hemodynamics around the injury site. Other conductive polymers, such as polyaniline (PANI) and poly­(3,4-ethylenedioxythiophene) (PEDOT), used in different constructs, are also capable of successfully propagating the bioelectric phenomenon in wound healing in a similar manner, proving their effectiveness in cases of infected wounds , and diabetic wounds. − By augmenting proliferation and remodeling at a range of different conductivities, with 3.33 × 10^–5 S/m from a PANI-incorporated hydrogel reported by Liu et al. to produce scarless healing of burn wounds through polarization of antifibrotic Th-1 cells, approximately 1.0–2.2 × 10^–2 S/cm from a poly­(deep eutectic solvent) hydrogel reported by Liu et al., 0.754 ± 0.04 S/m from a trimethylammonium chloride-based bio-ionic liquid-incorporated hydrogel reported by Cao et al. to significantly reduce scarring by promoting the M2 macrophage phenotype and accelerating collagen deposition, a range of 0.22–2.53 mS/cm from a polyaniline-grafted quaternized chitosan-incorporated hydrogel reported by Bi et al., and 0.14 ± 0.02 S/cm from an MXene nanosheet scaffold reported by Liu et al. From these experiments, it is evident that the incorporation of a bioelectric element into engineered skin substitutes to cooperate with or enhance the body’s natural endogenous electric field through simple conductance or piezoelectric activity is productive of an enhanced level of wound healing through tunable and minimally invasive methods, thereby presenting great potential for the improvement of available biomedical technologies. ,

2.7. Antibacterial Properties

The acid mantle of native human skin (pH 4.5–5.5), produced by the sebaceous glands, is vital for regulating the bacterial microbiome on the surface of the skin. Thus, while not necessarily through acidity, an engineered skin graft must prevent microbial colonization and bacterial infection at the injury site until wound closure is complete, since bacteria exacerbate inflammatory conditions and produce tissue-degrading enzymes. Besides antibiotics, various metallic nanoparticles are used frequently in regenerative tissue engineering, with their intrinsic electrochemical, piezoelectric, or photocatalytic characteristics imbuing skin substitute constructs with powerful bactericidal properties. In recent studies, silver (Ag) nanoparticles incorporated into nanofibrous scaffolds via electrospinning significantly hindered the growth of Gram-positive and Gram-negative pathogens, Escherichia coli (E. coli), and Staphylococcus aureus (S. aureus). , Zinc oxide (ZnO) nanoparticles have also been shown to have similar effects, even while their mechanism of action on microbes remains unquantified. , Likewise, a titanium oxide (TiO₂) nanoparticle scaffold induced high levels of oxidative stress in bacteria without demonstrating cytotoxicity to the native surroundings. In another study, a hydrogel cross-linked with cyanuric chlorine proved bactericidal when tested on E. coli-infected wounds as the cationic nature of the chlorine led to ruptures in the negatively charged lipopolysaccharide membrane of E. coli cells, contributing to a faster decrease of wound area after application. Still another publication detailed the incorporation of divalent cupric cations (Cu²⁺) into a photo-cross-linked gelatin methacrylate (GelMA)/sodium alginate (SA)-based hydrogel, which facilitated the sustained release of charged Cu²⁺ particles into infected rat wounds. The presence of the Cu²⁺ ionic particles was observed to induce increased membrane permeability in bacteria, resulting in a 92.2% bactericide rate in S. aureus populations and contributing to the overall success of the hydrogel’s performance in promoting angiogenesis, anti-inflammatory macrophage polarization, and accelerated regeneration rates. ,

2.8. Biochemical Signaling

Growth factors are bioactive peptides/polypeptides that serve as pivotal signaling molecules in regulating wound healing and have been frequently used in combination with other enzymes to produce remarkable results in the skin’s regeneration process. To direct cellular responses, growth factors bind to specific receptors in target cells, triggering a series of intracellular signals. Growth factors can be incorporated into a scaffold through several strategies: physical adsorption, chemical bonding with cross-linking agents, encapsulation into biocompatible materials-based microspheres or nanoparticles, hydrogel matrix coating, or layer-by-layer assembly techniques. Some crucial growth factors are TGF-β and its TGF-β1, β2, and β3 isoforms; VEGF, which induces vascularization/angiogenesis and promotes increased cell growth on grafts; PDGF and fibroblast growth factor (FGF), which attract and promote cell and fibroblast activity. In summary, the application of these growth factors possesses great potential to enhance the natural skin healing process through faster wound closure, improved blood flow, and better tissue formation, , resulting in scarless healing. The mechanical stiffness-driven aspect of fibrosis has also been addressed through methods based on biochemical signaling. Chen et al. (2022) employed a strategy that produced minimal scarring in split-thickness skin-grafting porcine models through the implementation of a pullulan hydrogel that delivered focal adhesion kinase inhibitor (FAKI) in the form of VS-6062. FAKI, which disrupts the mechanotransduction pathways of cells, was delivered to the wound in a controlled release via the sustained biodegradation rate of the pullulan hydrogel, resulting in a significant decrease in wound contraction (30% wound contraction at 28 days compared to 70% in wounds treated with split-thickness grafts) while reducing stiffness (average stiffness readings taken with an indentation cutometer averaged at a value of 1.5 as compared to an approximate average value of 1.8 for normal skin and an approximate average value of 0.9 in wounds treated with split-thickness grafts) of in the regenerated wound. Fibrosis inhibition was further confirmed extensively through histological analysis (collagen was found to be less densely deposited and more naturally arranged in the FAKI/hydrogel-treated wounds), single-cell RNA-sequencing (scRNA-seq) analysis (treatment with FAKI downregulated expressions of COL1A1, COL3A1, and FN1 for ECM remodeling while upregulating expressions of CXCL10, IP-10, SOCS3, WP4495, WP1835, and GOBP:0006958 for inflammation and fibroblast response to inflammation), RNA velocity analysis, APOE, CLEC3B, CD34, and PPARG proteins were upregulated in lipofibroblasts in response to FAKI, while the presence of decorin and TGF-β1 significantly increased and decreased, respectively, and immunofluorescent staining, which last provided visual confirmation of all the preceding in terms of apolipoprotein, CD34, and APOE expressions.

2.9. Appendage Recovery Support (Hair Follicles)

The restoration of various cutaneous appendages, such as nerves, hair follicles, and the eccrine, apocrine, and holocrine glands, is crucial for the recovery of the skin’s vital functions, which include aesthetics, regeneration, thermoregulation, moisture retention, surface acid barrier maintenance, and facilitation of the somatosensory process. Hair follicle regeneration has been achieved in various studies, in which natural polymer-based scaffolds comprised of silk fibroin (SF), SA, collagen/chitosan, and strategically selected cells contributed to the regrowth of these appendages. In this regard, Kang et al. utilized advanced 3D bioprinting to fabricate a multilayered gelatin-SA scaffold that replicated the structures of the dermis and epidermis and simulated the native hair follicle environment through encapsulation of spheroids containing fibroblasts, human umbilical vein endothelial cells (HUVECs), dermal papilla cells, and epidermal cells. Besides producing significant in vitro results, which included expressions of hair-inducing markers alkaline phosphatase (ALP), β-catenin, and α-SMA in hair follicle-like structures, promising in vivo results were also obtained when the bioprinted scaffolds were implanted into nude immunodeficient mice. Substantial hair growth was reported after 6–8 weeks, with the engineered hair follicles remarkably assuming a natural hair growth orientation. Similar results, albeit using a bilayered dermal-epidermal composite comprising type I rat tail collagen and human neonatal foreskin keratinocytes to encapsulate the dermal papilla cells. Scarless healing was also reported in tandem with hair follicle regeneration by Dou et al., with the use of an SF/SA scaffold loaded with human umbilical cord mesenchymal stem cells. The reduction of scar formation was demonstrated by an increased presence of TGF-β1, TGF-β3, and types I and III collagen and a wound healing rate of 95.9% by day 20 in Sprague Dawley rat models, while increased expressions of the CD31 angiogenetic marker contributed to the progression of hair follicle regrowth. Ji et al. likewise documented an outcome in which artificial hair follicle seeding and scar-free healing was accomplished with the use of a scaffold composed of hydrogel microspheres served a dual function as (1) liposome nanodrug delivery system conveying medical drugs (Tideglusib and Tamibarotene) into an injury site to chemically reprogram fibroblasts into dermal papilla cells while (2) displaying a porous ECM-like three-dimensional microstructure and a cationic polyamide-incorporated chemical composition that exploited the wound site bioelectric effect to allow for electrostatic wound adhesion. When applied to full-thickness wounds in mice, it was found that, by day 6, wound sizes had dropped to an average size of 8.15 ± 0.3 mm² from an average original size of 47.07 ± 3.4 mm², and that by day 12, the wounds had completely healed. Additionally, on day 21, a gene profiling study revealed that expressions of the profibrotic growth factor TGF-βR2 were significantly decreased as compared to other treatments, while cytokeratin 14 immunostaining revealed densely packed, mature hair follicles with long hair shafts within the regenerated area.

2.10. Appendage Recovery Support (Glands)

Regrowing and restoring the function of important glands, particularly sweat glands and sebaceous glands, is part of the “gold standard” for regenerative skin tissue engineering due to the important roles these glands play in skin hydration, heat dissipation, thermoregulation, and maintaining the antibacterial/antioxidant acid mantle. Nevertheless, to date, very few studies have successfully accomplished the regeneration of both gland types in situ with the application of a single, multifunctional engineered skin substitute, usually being singly focused on either sweat gland regeneration − or sebaceous gland regeneration. However, a 2024 study conducted by Xiong et al. detailed the design and implementation of a programmable bilayered artificial skin containing ECM-like DNA supramolecular hydrogels that fostered the regrowth of multiple functional skin appendage types. The lower skin-contacting layer of the construct comprised liposomes encapsulating Verteporfin, while its upper layer was a double-network hydrogel that supported the construct and replicated the mechanical and rheological properties of the surrounding skin. The Verteporfin-incorporated composition of the construct was found to greatly modulate inflammation, amplifying arginase-1 expressions toward M2 macrophage polarization while inhibiting expressions of IL-1β, IL-6, and TNF- α, which indicate M1 macrophage polarization. Superb outcomes were also observed after the construct was implemented in the treatment of dorsal wounds in C57BL/6 mice, with newly formed hair follicles (confirmed by the presence of cytokeratin 14, cytokeratin 5, fatty acid synthase, and leucine-rich G-protein-coupled receptor 5 markers), functional sebaceous glands (confirmed by the presence of cytokeratin 14, fatty acid synthase, stearoyl-CoA desaturase 1, and peroxisome proliferator-activated receptor-γ), and flat, near-scarless skin morphology clearly displayed by day 30. Additionally, gene expression analysis confirmed the significant upregulation of regenerative genes (i.e., DKK2, ADGRD1, NEGR1, FHL1) and appendage repair genes (i.e., PPARγ, PPARα, PRRX1, KDR) and the downregulation of pro-fibrosis genes (i.e., CXCL14, RUNX1, MYC, SOX9), ECM deposition genes (i.e., COL9A3, ADAMTS3, ACAN, COL11A1), and mechanotransduction genes (i.e., YAP1, RUNX1, HES1, JUND), demonstrating the antifibrotic capabilities of the graft. Exceptionally, functional sweat gland regeneration was also accomplished by day 14 after artificial skin implementation, which was verified by a starch-iodine sweat test in addition to a detailed histological analysis.

2.11. Appendage Recovery Support (Nerves)

Beyond glands and hair follicle restoration, sufficiently and speedily innervating engineered skin after its implantation to enable significant sensation recovery remains extremely difficult in the modern era of regenerative medicine due to the sheer delicacy and complexity of the somatosensory nervous system and its regeneration. The main skin sensory nerve fiber types include A-δ fibers, unmyelinated C group fibers, and A-α fibers, which play finely interrelated roles in the perception of mechanical stimuli, thermal stimuli, pressure, and pain. After damage is sustained from deep burns or cuts, macrophages populate the damaged area to clear away axonal and myelin debris from the crushed/severed nerve stumps, after which Schwann cells arrive to secrete and synthesize laminin and nerve growth and neurotropic factors, and to form the Bands of Büngner, a connective conduit that directs the growth of new nerve fibers. Optimally, these conditions produce a nerve fiber growth rate of roughly 1 mm/day. However, because post-wounding fibrosis drastically transforms the microstructure and composition of the injury site through the deposition of dense ECM for rapid wound contraction, the progress of innervation is greatly hindered and ultimately causes the loss of sensation in scar tissue. Therefore, various innervated 3D skin models have been constructed to foster nerve outgrowth and extension through multifaceted dermo-mimetic systems, − while other skin substitutes have promoted neurogenesis through scaffold anisotropy, as discussed earlier, or through the agency of the bioelectric phenomenon. A remarkable strategy featured in a 2022 publication by Tan et al. involved the use of a self-powered conductive “smart patch” composed of polyvinylidene fluoride (PVDF), reduced graphene oxide, doped methacrylate gelatin, ginseng-derived exosome-like vesicles, and the CXCL12 chemokine. During tests, the PVDF within the patch demonstrated exceptional piezoelectric and conductive properties, generating electric potentials of approximately ± 35 mV and +350 to −650 mV under 1 and 4% bending conditions, respectively. Consequently, when tested in vitro with bone marrow mesenchymal stem cells (BMSCs), the patch directed neural differentiation of the BMSCs, as demonstrated by significant expressions of nestin and β-tubulin, a significant inward Ca²⁺ flux, pronounced gene enrichment of the Pi3K-Akt and Ras neurotrophic signaling pathways, and regulation/expression of 83 neural differentiation-related genes, including Cortactin, Pik3cb, and Erbb3. Moreover, when applied to BMSCs in a skin full-excision rat model, the patch similarly amplified the expressions of CD90, p-PI3K, p-Akt, and the cyclic AMP response element binding (CREB) protein, which resulted in alleviated fibrosis, reduced scarring, and the observation of a dense network of nerve-like structures running through an organized collagen matrix closely resembling pre-wound conditions in the healed skin.

3. Biomaterials for Skin Regeneration

When selecting a biomaterial for skin scaffolding, several factors, including biocompatibility, biodegradability, porosity, mechanical properties, and anisotropy, must be considered to replicate the behavior and strength of the real skin.

3.1. Natural Polymers

Natural polymers such as collagen, gelatin, and hyaluronic acid are valued for their high biocompatibility, hydrophilicity, and biodegradability, as they closely resemble the natural ECM. Hydrophilic polymers, which constitute most natural polymers, are generally more effective for cell development because they attract and retain water, creating a moist environment that promotes cellular adhesion and proliferation. Their polar functional groups facilitate protein adsorption, which is essential for cell attachment, and enhance interactions with cell membrane receptors. , However, natural polymers can be challenging to process due to their complex structures and may exhibit poorer mechanical properties compared to synthetic polymers (Figure ). ,

3.

Advantages and disadvantages of natural polymers. Natural polymers offer excellent cell interaction but lack the necessary mechanical properties that would make them sufficiently durable for skin tissue engineering applications.

3.1.1. Collagen

Collagen is the most abundant protein in mammals, contributing to about 30% of the human body and providing strength in connective tissues, including skin and bone. Collagen is biodegradable and biocompatible, has a low immunogenicity, and is often combined with other natural polymers to improve its mechanical properties and control biodegradation rates. , Currently, 29 collagen types have been identified, with collagen types I, II, and III constituting 90% of the total collagen content in the human body. Because of its prominent role in contributing to the anisotropy, ECM microstructure, and mechanical properties of the skin, the synthesis and deposition of fresh collagen in an injury site is crucial to the wound healing process. It is generally understood that Type III collagen, in the form of thin fibrils, is the first collagen type to be created by the body in a wound area, with Type I collagen synthesis following soon after to promote angiogenesis, ECM remodeling, and provide mechanical support to regrowing tissues. Thereafter, a balanced ratio of Types I and III collagen in the regenerating wound is maintained. However, this balance can be disrupted due to the degradation of collagen by bacterial infection, diabetic conditions, or persistent inflammation, creating the need for scaffolds/grafts/hydrogels to possess bactericidal and anti-inflammatory properties. Additionally, collagen fiber concentration and orientation contribute to the formation of hypertrophic and keloid scars, with a high content of linearly oriented collagen fibers (as compared to a less-concentrated, “basket weave”-patterned structure in normal skin) observed to result in stiff, significant scar areas. Thus, delivering apposite collagen types and amounts to an injury site is a crucial design factor to be considered in the fabrication of a collagen-based scaffold. Xu et al. investigated the limitations of current therapies for chronic wounds, particularly diabetic foot ulcers, which often fail due to impaired cellular function during the wound healing process. To address this challenge, recombinant human collagen III (rhCol III) was used as a delivery matrix because of its excellent biocompatibility, high water solubility, structural stability, and reduced risk of pathogenic contamination compared to animal-derived collagen. These properties enabled a targeted and sustainable platform for extracellular vesicle (EV) delivery. The rhCol III hydrogel was fabricated by cross-linking rhCol III with transglutaminase (TGase) derived from Streptomyces mobaraensis, resulting in a stable hydrogel capable of protracted EV release. This delivery system effectively modulated the inflammatory response by promoting macrophage polarization from the pro-inflammatory M1 phenotype to the pro-healing M2 phenotype. In vitro studies further demonstrated enhanced migration of L929 fibroblasts and increased angiogenic activity in HUVECs. In a diabetic rat skin wound model, treatment with the rhCol III-EV hydrogel significantly reduced inflammation and promoted cell migration and neovascularization within the wound tissue. These combined effects led to accelerated and improved wound healing, as a 94% wound recovery rate was reported by day 14 after the hydrogel was applied. Overall, this study highlights the promising potential of rhCol III-based EV hydrogels as a novel therapeutic strategy for chronic wound treatment by restoring key cellular functions essential for effective tissue regeneration. The application of Type I bovine collagen-based hydrogels (Collost Gel) bearing 7% or 15% collagen concentrations depending on the size and locations of wounds resulted in the long-term reduction of pruritus, scar stiffness/tightness, and vascular and pigment indices when applied to individual cases of traumatic facial wounds, blepharoplasty, abdominoplasty, and revision mammoplasty. To treat the traumatic facial wound, the hydrogel was injected intradermally into the wound margins on the same day as, and on days 2 (during the inflammatory phase), 17 (during the proliferative phase), and 34 (during the remodeling phase) after wounding occurred, resulting in the documentation of a healthy normotrophic scar with a Vancouver scar score scale of 1 about four and a half months later. These two previously referenced studies imply a dichotomy of long-term outcomes produced by the respective application of Type I and Type III collagen in wound healing endeavors. Nevertheless, the broader implementation of collagen in the treatment of wounds/burns in combination with other regenerative biomaterials is recognized to produce positive results, as demonstrated in a 2020 publication by Kalirajan et al., who applied a collagen-incorporated scaffold combined with oxidized jackfruit seed polysaccharide and silver catechin toward the scarless healing of infected diabetic burn wounds, with full healing reported by 24 days. By 30 days, a scar evaluation index value of 1.3 ± 0.1 was observed in the treated wounds. Furthermore, histological analysis revealed a regenerated epidermis containing hair follicles and sebaceous glands. It was noted that the scaffold’s collagen composition facilitated the rapid placement of well-aligned collagen fibrils in the wound site, and also that its silver catechin component efficiently mitigated collagen over-deposition, consequently reducing scarring significantly. Similarly, collagen-incorporated hydrogels implemented in two different studies facilitated comparable full wound closure times of 21 days for diabetic rats with minimal scarring. , Collectively, these studies emphasize the potency of collagen as an expediting agent in the wound regeneration process, attracting important matrix metalloproteinases, fibroblasts, and macrophages and signaling angiogenesis as a wound transitions from one healing phase to another. Thus, collagen remains an extremely popular choice as a biomaterial for skin tissue engineering with optimal outcomes in wound regeneration. However, ensuring that the right collagen types and amounts are delivered to an injury site to ensure that the body restores a Type I/Type III ratio in a moderately dense “weave” characteristic of natural skin persists as a principal challenge encountered in the use of collagen.

3.1.2. Gelatin

Gelatin is produced through the degradation of insoluble collagen by disintegration or denaturation obtained via acid or alkaline hydrolysis of collagen. , Generally, gelatin is a degraded form of collagen extracted from animal bones, skin, and tendons, existing in two types based on the hydrolysis process: partial acid hydrolysis produces type A gelatin, and alkaline hydrolysis produces type B gelatin. Gelatin is biocompatible, biodegradable, and contains large amounts of arginine-glycine-aspartic acid (RGD) sequences that support cell adhesion and growth. Moreover, gelatin molecules contain chemically modifiable functional groups, allowing them to be modulated or loaded with drugs, making them effective carriers for drug delivery applications. Thus, gelatin has been recently used in various engineered skin substitutes to mitigate hemostasis through activating platelets and stimulating coagulation, reduce inflammation caused by ROS due to intrinsic free radical-scavenging properties, − deliver potent angiogenic and immunomodulatory peptides, , and support scarless or near-scarless healing in the epidermis at elevated rates. , Elaborating on this last, Wen et al. crafted a fibrous membrane from gelatin, poly­(lactic-co-ε-caprolactone), and salvianolic acid B that demonstrated to possess excellent cytocompatibility (indicated >70% viability in L929 fibroblasts), angiogenetic (indicated lumen network formation and PI3K/Akt pathway activation in HUVECs), M2 macrophage polarization (indicated by upregulation of CD206 and downregulation of CD86 and IL-1β markers in RAW 264.7 cells), and antifibrotic capabilities (indicated by fibrotic gene markers CD36, α-SMA, and Type I collagen inhibitions) in vitro. These results became more pronounced during in vivo testing, with treated wound sites exhibiting significantly thinner fibrotic envelopes at days 7 and 14, accompanied by well-ordered collagen fibers and decreased presence of Types I and III collagen and the pro-fibrotic cytokine TGF-β1, higher levels of angiogenesis and vascular network formation, and the recovery of hair follicles documented. Additionally, a wound healing rate of 96.6 ± 1.5% was reported by day 14, with a mean scar length of 0.63 ± 0.25 mm. Corresponding results have also been attributed to the use of GelMA (gelatin bonded to methacrylic groups) when used in the fabrication of a microsphere delivery system bearing epigallocatechin gallate and triamcinolone acetonide by Luo et al. in 2025. In this study, the sustained biodegradation rate of the GelMA microspheres created sequential drug delivery in harmony with the phases of the wound healing process, which was highlighted as a key feature that contributed to the success of the experiment. When applied to 6 mm full-thickness skin defects in C57BL/6 mice, the microsphere system achieved a wound closure rate of 99.62% by the tenth day after implementation, featuring rapid re-epithelialization, significant inflammation reduction, notable M2 macrophage polarization and angiogenetic activity, and rapid but controlled collagen deposition. A more positive response occurred when the microsphere system was used on hypertrophic scar models in rabbits, producing fully regenerated wounds with average scar evaluation index values of 1.39 and average collagen value fractions of 49.77% (comparable to a fraction of 46.43% in normal skin) by the fourth week. Sirius Red staining further revealed that the ratio of Types I and II collagen was well balanced, while immunofluorescence staining detected significantly low levels of α-SMA, which indicated the suppression of excessive myofibroblast activity and consequent uncontrolled collagen deposition, highlighting the potential of the GelMA microsphere system for scarless wound healing. The cost-effectiveness of gelatin due to its being freely available, along with its other advantages of good biocompatibility, biodegradability, and low likelihood for triggering immune rejectionare offset by its low thermal stability near the body’s temperature, as well as by its poor mechanical properties, exhibiting brittleness when dry and excessive softness when hydrated, requiring that it be cross-linked or combined with other polymers to increase the strength and plasticity of its matrix. However, these methods increase manufacturing complexity and present the risk of introducing cytotoxicity, thus counterweighing the appeal of gelatin for use in skin substitutes.

3.1.3. Alginate

Alginate is a polysaccharide derived from natural brown seaweeds such as Laminaria and Lessonia and is made up of (1–4)-linked β-D-mannuronic acid (M) and α-l-guluronic acid (G). M residues exhibit greater biocompatibility, whereas G residues provide increased rigidity. The source from which alginate is procured determines the number and sequence of the M and G residues, affecting the physical and chemical properties. Due to its biocompatibility, hydrophilicity, and stability, alginate is helpful in various tissue engineering applications, drug delivery, and wound healing. As an example of the base regenerative power of these properties, Toumaj et al. developed an alginate/chitosan hydrogel loaded with Obestatin and evaluated collagen type I, II, and TGF- β gene expression using an in vitro rat model to study the wound healing process in the skin. When exposed to phosphate-buffered saline (PBS), which acted as a substitute for liquid present in the tissue, the Alg/Cs and Alg/Cs/Obes showed increased degradation and swelling, demonstrating their ability to absorb extracellular fluid and facilitate cell migration and proliferation. In vitro results showed that the Alg/Cs/Obes hydrogel with 100 μM of Obestatin had the highest proliferation (200% after 72 h in 3T3 fibroblast populations) and wound closure rate (just under 80% by 7 days, and roughly 90–95% by day 14), thus increasing wound healing. Furthermore, alginate-based hydrogels act as effective wound dressings for tissue engineering due to their mechanical strength, low water swelling, and high compatibility with living tissue in vivo. Beyond this, the potency of alginate for producing scar-free outcomes has been validated by various studies, as a porous alginate/gelatin/aloe vera hydrogel (average pore size 163.33 ± 14.45 μm, Young’s modulus 17.43 ± 1.41 MPa) produced a significantly low scar evaluation index value of 1.17 ± 0.04, signifying negligible scarring, when applied to wounds in Wistar rat models through excellent improvement of wound bed angiogenesis and accelerated re-epithelialization rates, while another bioprinted SA/gelatin hydrogel greatly inhibited scar formation by attracting large numbers of myofibroblasts, noting minimal injury site contraction and the lack of cicatricial formation. In the former publication, alginate was credited with enhancing tissue regeneration and angiogenesis, while the latter remarked that its excellent printability motivated its selection as a material, consequently producing average micropore sizes of 102.9 μm, which appear to have hindered the excessive adhesion of fibrosis-inducing myofibroblasts to the site. Similarly to gelatin, alginate is at its most advantageous when combined with other polymers to mitigate its high viscosity and naturally poor degradability/solubility. Such combinations render alginate highly malleable, making it a prime candidate for the fabrication of highly tunable, multifunctional, and porous constructs that are extremely hospitable to regenerative cell behaviors.

3.1.4. Hyaluronic Acid

Hyaluronic acid (HA) is a naturally occurring anionic disaccharide composed of N-acetylglucosamine and glucuronic acid molecules, and is secreted in the ECM surrounding skin, joints, eyes, and nervous system, acting as an autocrine or paracrine signaling molecule in different biological scenarios. Upon injury to the skin, HA binds to various isoforms of CD44, a nonkinase transmembrane glycoprotein expressed as a cell surface receptor in many epidermal cells, including neutrophils, macrophages, keratinocytes, fibroblasts, and endothelial cells, activating countless intracellular signaling cascades that regulate cell proliferation, differentiation, migration, and expression throughout all phases of wound healing and scar formation. , A study conducted by Winters et al. featured hydrogels composed of a combination of HA, acrylic acid (AA), Poly­(3,4-ethylenedioxythiophene) (PEDOT) nanoparticles, and lignin, which demonstrated remarkable biocompatibility besides excellent mechanical properties and a well-arranged microstructure. Seeded cultures of NIH-3T3 fibroblasts showed high proliferation rates of 92.5 ± 10.3% over 48 h after being subjected to an Alamar Blue assay, while a Live/Dead cell assay revealed that the concentration of live cells was 80% higher than the concentration of dead cells, with the highest overall percentage of viable cells being recorded at 89.7 ± 2.6%. Similarly, another 2024 study described biomimetic skins created from a mixture of sulfated HA and collagen that were applied in vivo to diabetic mouse models, resulting in increased expressions of VEGF-A and TNF-α, signaling the activation of angiogenetic protocols via the notch signaling pathway. Furthermore, diabetic wounds that were treated with HA-based biomimetic skins demonstrated regenerated blood vessels and maturing hair follicles and sebaceous glands 10 days after application as compared to significantly lesser manifestations in the control group. Furthermore, HA incorporation into scaffolds has been proven to greatly alleviate scarring over extended wound healing periods by providing moist, bioactive environments, , and accelerating re-epithelialization through the maturation and balancing of types I and III collagen, resulting in smoother, naturally colored scar sites with low elevation and nodule counts. ,− Further establishing its benefits for the skin, HA is widely used outside of skin tissue engineering in the dermatological industry as a topically applied agent for skin hydration, rejuvenation (for cases of UV radiation damage or acne scarring), and to counteract the visual effects of aging. Complications surrounding the procurement of HA are the most significant caveats to its promise. HA is primarily obtained from animal sources, requiring a lengthy purification process that increases its cost and restricts its experimental use through financial factors. Furthermore, purification methods vary, leading to discrepancies in exhibited biological properties. Thus, great room for improvement remains for the use of HA in biomedical studies.

3.1.5. Elastin

Elastin, synthesized primarily during the prenatal stage through high expressions of the ELN gene and referenced earlier with regard to its contributions to skin anisotropy, differs from collagen in that it is the primary modulator of the skin’s elasticity through an intricate arrangement of fibers that run both parallel and perpendicular to the skin, while collagen regulates its mechanical strength and firmness. Thus, the disruption/degradation of the elastin network due to wounding, aging, or chronic UV/solar radiation exposure makes the skin stiff, creating profibrotic feedback circuits via mechanotransductive integrins and the YAP/TAZ pathway in fibroblasts, which sense increased ECM stiffness due to the over-deposition of collagen and release TGF-β1, activating myofibroblasts and increasing expression levels of TGF-β1 and α-SMA in the cellular milieu, thus exacerbating the progression of fibrosis through the creation of ECM components in excess, as alluded to earlier. Injury to the skin triggers the expression of Tropoelastin, an asymmetrical molecule with alternating hydrophilic–hydrophobic domains encoded by the ELN gene, by TGF-β, TNF-α, Interleukin-1β, Vitamin D, and insulin-like growth factor-1. After synthesis of its monomer form in the endoplasmic reticulum of fibroblasts and its secretion and self-assembly into a polymer on the fibroblast cell surface. Tropoelastin then binds to other vital elastic fiber proteins, including Fibrillins 1 and 2, Latent TBF-β-Binding Proteins (LTBPs) 1, 2, and 4, Fibulins 2, 3, 4, and 5, and Microfibril-Associated Proteins 1, 3, and 4, whose expressions also increase when the skin sustains a burn or wound. The coalescence and interaction of these tropoelastin molecules and glycoproteins with various mechanotransductive and cell adhesion integrins (αvβ3, α5β1, αvβ6, α8β1, αvβ1, αvβ5), fibronectin, and the elastin-binding protein result in the formation of new, cross-linked elastin fibers throughout the ECM during the process of elastogenesis. Subsequently, these fibers bind TGF-β through their Fibrillin-1 content, influencing the differentiation of fibroblasts into myofibroblasts to promote wound contraction and closure. Simultaneously, the maturation of the new elastin fibers prevents over-contraction in the regenerating wound, attempting to regain their native arrangement and restore the skin’s original mechanical and elastic properties. However, because elastogenesis occurs relatively late (after the deposition of Type I collagen) and to a limited degree during the wound healing process, the full regeneration of the elastin fiber network is greatly disrupted, preventing the healed skin from demonstrating the elasticity and appearance of its uninjured state. The strategic delivery of elastin to a wound site via a collagen-elastin hydrolysate scaffold manufactured through freeze drying and its potential for scar-free wound healing was investigated by Krymchenko et al. in a 2025 pre-clinical trial. The scaffold, which was reported to be porous and of a dermo-mimetic stiffness due to its near-natural collagen-elastin ratio of 95:5, was observed to produce the least wound contraction in comparison to the outcomes observed in untreated wounds and wounds treated with a similar scaffold with a collagen-elastin ratio of 97:3. While the presence of elastin was noted to significantly increase the time to complete wound closure to 35 days (compared to 14 days to complete wound closure in untreated wounds), it also facilitated the neovascularization, enhanced ECM remodeling, and organization of extensively deposited collagen into finer fibrils while producing reduced expressions of α-SMA, ensuring the resolution of the wound into a linear, normotrophic scar. Elastin-like recombinamers, SKS (sequences contain serine and lysine) and VKV (sequences contain valine, proline, glycine, and lysine), have been used to create thin, porous bioactive membranes with pore sizes of 2–5 μm, an elastic modulus of 1–10 kPa, and a biodegradation rate of approximately 22 days. When tested in vivo using C57BL/6J murine models, the membranes were shown to augment wound healing by promoting angiogenesis, modulating inflammation, and guiding the alignment of newly synthesized collagen fibers, while preventing wound contraction and preventing the formation of visible scarring. 7 weeks after implementation, the membranes were observed to have been fully incorporated into the newly formed skin. Beyond these publications, the application of elastin-based scaffolds/hydrogels , or other constructs containing elastin-like polypeptides/recombinamers − has been investigated in various studies with promising regenerative outcomes. However, due to the sheer difficulty of synthesizing mature elastin fiber from its precursors, obtaining purified elastin, or producing biomaterials that exhibit similar mechanical behavior, the use of elastin in engineered skin substitutes that expedite true scarless healing of the skin remains regrettably limited.

3.2. Synthetic Polymers

On the other hand, synthetic polymers, such as polylactic acid (PLA), polyglycolic acid (PGA), and polycaprolactone (PCL), offer greater control over the mechanical properties and degradation rates. These polymers can be precisely tailored to achieve the desired porosity and anisotropy through advanced production techniques. However, since most synthetic polymers are hydrophobic, they may face challenges with biocompatibility and cell adherence, often requiring surface modifications or the incorporation of bioactive compounds to enhance biological responses. , Figure highlights the advantages and disadvantages of synthetic polymers.

4.

Advantages and disadvantages of synthetic polymers. Most synthetic polymers have the advantage of possessing mechanical strength and good tunability for manufacturing but require biochemical modifications through combination with natural polymers to better their interactions with the human body.

3.2.1. Polylactic Acid

Starch and starch-derived biomaterials are attractive for tissue engineering experiments due to their ease of manufacture and workability into the human body to accomplish delicate repair. PLA is a hydrophobic, biodegradable polymer composed of lactic acid derived from corn starch or other natural resources through fermentation processes and additionally by polymerization as a petrochemical derivative. , PLA exists as three forms: poly-L-lactic acid (PLLA), poly-D-lactic acid (PDLA), and poly-D,L-lactic acid (PDLLA). Furthermore, degradation of L-lactic acid is harmless to the human body compared to the harmful D-lactic acid. PLA has wide applications in tissue engineering due to its good mechanical properties, biodegradability, and biocompatibility. However, PLA degrades slowly in the human body and produces acids, which can lead to inflammatory reactions. Therefore, PLA should be modified to enhance bioactivity and improve healing. Cai et al. neutralized the acidic components of PLA that hinder the wound healing process through combining PLA with a mildly alkaline plant, which accelerated wound healing and confirmed an optimal environment for wound healing. This pH-regulated wound dressing was fabricated through an emulsion mixed from grass and wood ash, PLA powder, and a polymer surfactant, which was frozen to reduce solubility and to facilitate precipitation of the dressing. During the degradation of the resulting nanofiber dressing over 2 weeks, a stable pH value of 7–8 was maintained, which was close to that of natural skin, while demonstrating excellent hydrophilicity and mitigating inflammatory responses, making it extremely suitable for tissue regeneration. Besides this, Zhao et al. designed a tri-layered PLA/silk fibroin/retinoic acid scaffold that prevented epidermal hyperplasia when applied to wounds inflicted in rabbits. This intervention significantly reduced scarring after 28 days of observation in burn and hypertrophic scar models. Additionally, histological analyses further revealed dense, well-aligned deposits of collagen and fibroblasts, signifying that the scaffold had augmented the orientation and proliferation of both these regenerative components. Surprisingly, PLA has also been demonstrated by Yang et al. in early 2026 to have piezoelectric properties that contributed to scarless wound healing when used in combination with GelMA to fabricate a bilayered scaffold that delivered a sustained release of lactic acid and bone morphogenic protein 4 to polarize regulatory T cells, secreting the GATA3 transcription factor and the IL-4 and IL-3 cytokines. These three powerful biological agents suppressed the profibrotic immune responses and the persistent over-proliferation of ECM-depositing myofibroblasts. Moreover, electrostatic 3D printing was used to create a grid-like arrangement of PLA fibers with average sizes of 50 μm, and exhibited voltage generation capabilities of roughly 20 volts in response to an applied force range of 0.01–0.1 N. The PLA membrane exhibited a fascinating electrochemical interaction with the negatively charged GelMA hydrogel layer that was subsequently 3D-printed upon it. The generated piezoelectric field accelerated the rate of GelMA degradation and consequently increased the drug release rate into the injury site through regulating electro-osmotic flow and altering the hydrogel’s microstructure through the exertion of electrostatic force on the GelMA polymer chains, consequently increasing the scaffold’s porosity and facilitating efficient diffusion of oxygen and nutrients. In addition, in vivo tests also presented exceptional outcomes, as the piezoelectrically generated stretching force effectively resisted fibrotic wound contraction, producing skin tissue with a normotrophic appearance by day 28. On days 14 and 21, collectively, hematoxylin, eosin, and Masson staining revealed even, well-organized tissue composed of a near-natural ratio of Types I and III collagen (amounts of Type III collagen were noted to be higher), enclosing restored hair follicles and sebaceous glands. Furthermore, flow cytometry and transcriptome profile analyses demonstrated overwhelming evidence that profibrotic genes (Wnt3a, Actg1, Col1a1, Myc, Tgfb1, Fgr1) were greatly downregulated, while skin tissue genes (Col2a1, Col9a3) and skin appendage genes (Sox9, Notch1, Krt17, Krt40) had been upregulated significantly, confirming the superb level of healing. Finally, when applied to a rabbit ear wound model, the bilayered PLA-incorporated scaffold was observed to promote near-complete wound closure by day 15, with light-colored, smooth, and naturally appearing sites. While pure PLA is limited by its brittleness, stiffness, and hydrophobicity, its excellent workability with other polymers creates versatile, multifunctional wound dressings that possess PLA’s mechanical strength and shape-memory properties along with more appropriate biocompatibility and biodegradation characteristics, as discussed. Thus, the current work involves investigating novel, PLA-incorporated biomaterial blends to maximize their usefulness in tissue engineering endeavors and increase the likelihood of optimal regenerative outcomes.

3.2.2. Polycaprolactone

PCL is a popular component choice for tissue engineering for its excellent compatibility with a wide variety of polymers, good flexibility, low immunogenicity, and malleability during manufacturing processes. PCL was used with gelatin and ε-polylysine in a recent work by Ghomi et al. to construct a nanofibrous (fiber diameters of 267 ± 7 nm to 331 ± 8 nm) scaffold by means of electrospinning, which resulted in the near-complete migration of seeded human dermal fibroblasts and immortalized keratinocytes across a 500 μm gap after a period of 24 h due to the highly aligned microstructure on the nanofibers. Additionally, fibroblasts were observed to assume their natural elongated morphologies along the nanofiber orientation. Moreover, the incorporation of ε-polylysine into the scaffold imbued it with potent bactericidal properties, eliminating various resilient bacterial strains, including MRSA 700699, EC 25922 (E. coli), and SA 15981 (S. aureus) during a disk diffusion assay. PCL scaffolds have also been demonstrated to contribute to scar-free wound healing through regulating disoriented deposition of Types I and III collagen and the differentiation of fibroblasts into α-SMA-positive myofibroblasts, even when applied to diabetic chronic wound models. PCL is arguably one of the most widely used synthetic polymers for skin tissue engineering applications, as it combines well with many other biomaterials, imparting its strength, tunability, excellent workability, and sustained biodegradation rate to mechanically weaker materials such as collagen, alginate, and hyaluronic acid, as discussed. These polymeric blends also benefit PCL, which is naturally hydrophobic due to its aliphatic backbone and the general absence of polar side groups in its polymer chain, by improving the wettability of the resulting construct and creating a healthy hydrophobic/hydrophilic balance that is both conducive for protein adsorption and attractive for cell populations, respectively. Due to these qualities, PCL is commonly featured in tissue engineering endeavors, with novel biomaterial combinations and outcomes highly sought after to achieve the goal of truly scarless wound regeneration.

3.2.3. Poly­(lactic-co-glycolic Acid)

Poly­(lactic-co-glycolic acid) (PLGA), the copolymer of PLA and PGA, is an attractive component for tissue engineering applications due to its rapid biodegradation rate due to hydrolysis and good biocompatibility as an ester-linked polymer, making this synthetic polymer a popular candidate for use in tissue scaffolds and biomimetic drug delivery systems. As with PCL, PLGA is generally favored greatly for use in the fabrication of nanofibrous, interconnectedly porous electrospun constructs with mechanical properties and microstructural features that promote cell adhesion and infiltration in accordance with the contact guidance phenomenon. It has been reported that a bilayered skin graft was fabricated by combining an electrospun PLGA membrane with a fibrin hydrogel film seeded with fibroblasts. In a subsequent in vivo study using a rat full-thickness wound model, the PLGA–fibrin–fibroblast scaffold significantly accelerated type III collagen deposition for up to 14 days and enhanced granulation tissue formation. Granulation tissue thickness reached 1046.5 ± 242.5 μm at day 14 and 1378.2 ± 324.0 μm at day 21, compared with the negative control group, which exhibited thicknesses of 886.9 ± 196.9 μm at day 14 and 919.9 ± 352.5 μm at day 21. Furthermore, during the proliferative phase of wound healing, elevated expression levels of anti-inflammatory markers IL-10 (101.7 ± 15.1 pg/mL) and TGF-β1 (230.1 ± 17.6 pg/mL) were observed, along with an increased concentration of epidermal growth factor (EGF) (55.8 ± 4.1 pg/mL). These findings indicate an enhanced anti-inflammatory response and augmented re-epithelialization. Remarkably, PLGA microcapsules delivering TGF-β1 inhibitor (SB-431542) embedded into an o-Nitrobenzene-modified HA hydrogel produced healed skin wounds with flat surfaces and low scarring, disfigurement, and modularity in porcine and rabbit models. Experimental analysis revealed that the PLGA microcapsules exhibited excellent biodegradation rates that complemented wound regeneration rates, providing controlled release of TGF-β1 inhibitor and resulting in reduced phosphorylation of phospho-SMAD2, and phospho-SMAD3 to modulate the progression of fibrosis. A documented problem encountered in the use of PLGA as a drug delivery platform is its propensity for demonstrating an initially high burst release profile, which speaks to its rapid, somewhat erratic biodegradation rate. Therefore, a small layer of complexity is added during the incorporation of PLGA into experimental skin substitutes to modify this characteristic toward healthy contribution within a regenerative environment for favorable outcomes. Looking past this drawback, the combination of PLGA with other biomaterials seems promising for an elevated quality of wound healing.

3.2.4. Poly­(3,4-ethylenedioxythiophene)

Recognizing the important role of bioelectrical properties in the wound healing process, − synthetic, conductive polymers like PEDOT are frequently used in tissue engineering experiments to create constructs that are capable of harnessing the bioelectric phenomenon to amplify skin regeneration. The intrinsic conductive property of PEDOT arises from the presence of polarons/bipolarons in the atomic structure of thiophene monomers, and can be altered due to oxidation or mixture with another polymer during the manufacturing technology. A study reported the development of a conductive, self-healing hydrogel composed of SA, calcium, and PEDOT:Poly (sodium 4-styrenesulfonate) (PSS). The hydrogel featured an interconnected porous architecture with average pore sizes of 50 to 200 μm, an electrical conductivity of 0.0035 S/cm, and a maximum compressive strength of 22.23 kPa. Notably, the material exhibited robust self-healing behavior, recovering 92 to 95% of its original shape following repeated deformation to 500% strain. When co-cultured with human dermal fibroblasts and human umbilical vein endothelial cells and exposed to external electrical stimulation at 3 V for 1 h per day, cell viability increased from approximately 130% on day 1 to 160% by day 3 in both cell types. Scratch assays further demonstrated 79.7% wound closure within 24 h. Applied electrical stimulation (ES) significantly modulated the inflammatory response, as indicated by downregulation of the pro-inflammatory cytokines CD80, TNF-α, and IL-1β by 3.8-fold, 4.5-fold, and 2.9-fold, respectively, alongside upregulation of the anti-inflammatory markers Arg1, CD206, and IL-10 by 5.7-fold, 6.1-fold, and 7.2-fold, respectively. In addition, angiogenic activity in endothelial cells was enhanced, with increased expression of CD31 by 3.2-fold, FGF-2 by 2.5-fold, and VEGF-A by 2.3-fold, collectively indicating the establishment of a pro-regenerative microenvironment conducive to wound healing. Moreover, when applied in combination with ES to an in vivo rat model of full-thickness burn wounds, the hydrogel demonstrated further therapeutic efficacy. Thus, wounds achieved 96.46% closure by day 10 and nearly 100% closure by day 14. Histological analyses at day 14 revealed the formation of a continuous epidermal layer approximately 150 μm thick containing hair follicles, along with collagen deposition covering approximately 50% of the wound area, as confirmed by multiple staining methods. Additionally, Immunohistochemical analysis further demonstrated significant polarization of macrophages toward the M2 phenotype at the same time point, indicating a pro-regenerative and anti-inflammatory wound microenvironment. As with PLGA, the main challenges associated with the use of PEDOT stem from its natural rigidity and hydrophobicity, as well as its extremely high biodegradation rate, characteristics which create the potential for mechanical mismatch with the surrounding skin and immune rejection. Moreover, it is also challenging to implement balanced PEDOT concentration in a biomaterial-based skin substitute that is both sufficiently conductive and minimally cytotoxic. Therefore, while the use of PEDOT has garnered significant attention for the fabrication of scaffolds/hydrogels that interface with the natural bioelectrics of the human body, a greater level of application is still out of reach due to the drawbacks presented by its synthetic nature.

4. Manufacturing Methods

In skin tissue engineering, selecting the appropriate manufacturing methods and materials is also essential for creating functional scaffolds that replicate the structure and function of the real skin. Biomimetic scaffold fabrication involves various processes, including solvent casting/particle leaching, freeze drying, electrospinning, 3D printing, and gas foaming. Each approach offers distinct advantages and challenges regarding mechanical properties, porosity, and anisotropy, all of which are critical factors for effective tissue regeneration. This section examines each of these manufacturing technologies and how they contribute to the creation of scaffolds that are ideal for skin tissue engineering applications.

4.1. Solvent Casting/Particle Leaching

This manufacturing process involves dissolving a polymer in a solvent, adding porogen particles, pouring the mixture into a mold, evaporating the solvent, and lastly, leaching off the porogen to produce porous scaffolds (Figure ). − Although it permits great porosity, it may lead to residual solvent retention and poor mechanical qualities. , Solvent-cast chitosan/gelatin scaffolds showed promise for cell-friendly environments by encouraging fibroblast attachment and proliferation as the gelatin content was raised. Nevertheless, cross-linking is necessary to stop rapid degradation and maintain gelatin’s long-term stability in aqueous conditions, which can make fabrication more difficult and compromise biocompatibility. Another study demonstrated the biocompatibility and mechanical strength of a solvent-cast biocomposite film, revealing a water absorption rate of 451.1 ± 6.02% and a tensile strength of 8.9 ± 0.30 MPa. In vivo tests revealed that the film had antimicrobial and antioxidant properties as well as accelerated healing and reduced scarring. These results highlight the outstanding versatility of solvent casting for creating bioactive wound dressings, but it must be noted that difficulty remains in achieving optimal mechanical properties and degradation rates. Moreover, starch/TMPyP-based antimicrobial films that exhibited strong photodynamic activity against E. coli have also been constructed via solvent casting. These films demonstrated high potential for infection control through reducing bacteria populations by 99.99% (4.0log₁₀) after 30 min of exposure to white light (400–800 nm). However, the dependence of these scaffolds on light activation limits their use in low-light or light-inaccessible wound environments.

5.

Solvent casting is a highly feasible method for tissue engineering since it does not require the use of expensive equipment. Simple leachable particulates have the potential to create a porous microarchitecture that is hospitable for cells.

4.2. Electrospinning

Electrospinning creates ECM-mimetic nanofibrous scaffolds with interconnected porosity that enhances cell adhesion and supports proliferation (Figure ). − However, drawbacks of electrospinning include difficulties in obtaining desirable scaffold mechanical strength and anisotropy and fostering adequate levels of cell infiltration. , The combination of electrospun PCL nanofibers with a chitosan hydrogel film dramatically increased fibroblast infiltration and proliferation by 150 ± 20%, according to a study on bilayered skin-like nanopads. The scaffold’s wound closure rates also outperformed those of traditional wound dressings in vivo, reaching 60% closure by day 7 and 87% by day 10. Even so, neat PCL nanofibers have limited bioactivity and low degradation rates, necessitating functional modifications to maximize cellular interactions. The use of electrospun silk fibroin nanofibers stratified upon a human amniotic membrane (HAM) to constitute a bilayered skin substitute demonstrated superior dermal and epidermal regeneration in a full-thickness wound model in mice. Keratinocyte attachment and ECM development were facilitated by the electrospun fibers’ large surface area and interconnected pores, which improved healing. However, structural reinforcement was required for long-term stability due to HAM’s poor mechanical properties and enzymatic susceptibility. Another study investigated the use of a drug-loaded bilayered electrospun scaffold for cutaneous squamous cell carcinoma (cSCC) postoperative wound healing. Both infection control and tumor suppression were combined in the scaffold’s antibacterial PCL-enrofloxacin (ENR) nanofiber outer layer and anticancer PCL/gelatin-bleomycin (BLM) nanofiber inner layer. Despite its functional benefits, PCL’s hydrophobicity hindered its early cell interactions. Consequently, gelatin was utilized to increase the scaffold’s hydrophilicity and bioactivity. In conclusion, while electrospun scaffolds efficiently promote wound healing and fibroblast proliferation, addressing their mechanical, bioactive, and biodegradation limitations for clinical translation remains an imperative.

6.

Capability of electrospinning to create biomimetic nanofibers is its most appealing feature. Factors contributing to the creation of these fibers are the electrospinner’s voltage, distance from the collector, and flow rate with which the polymeric solution is delivered.

4.3. Freeze Drying

Freeze drying produces porous biomaterials by freezing the aqueous phase of a polymeric compound and removing it via ice crystal sublimation. Although this improves cell infiltration and maintains bioactivity, it produces poor mechanical characteristics. Anisotropy and pore size can be regulated by varying the freezing and drying conditions. One study found that freeze drying aided in the creation of highly porous halloysite nanotube (HNT)-enhanced hydrogel scaffolds that mimicked the natural ECM for cell adhesion and proliferation. The pore sizes of scaffolds ranged from 15 to 60 μm, allowing for cell migration and proliferation. Nevertheless, incorporating HNT slightly reduced overall porosity, affecting permeability, while improving structural stability. Similarly, in another study, freeze drying was critical for creating rapidly curable HA-catechol hydrogels for wound healing applications. The process helped to improve water absorption and swelling control, both of which are important for wound hydration. However, the gelation process required additional enzymatic cross-linking to achieve mechanical strength, increasing the complexity of fabrication. Additionally, pectin-based hydrogels made by freeze drying showed improved swelling and mechanical properties, making them suitable for long-term wound coverage. The technique ensured homogeneous polymer distribution; however, variable degradation rates remained a challenge, necessitating further material optimization.

4.4. Gas Foaming

Gas foaming creates a porous structure without the use of organic solvents by introducing gas bubbles into a polymer matrix (Figure ). , But, while this method produces scaffolds with a range of pore sizes, resulting constructs frequently lack adequate mechanical strength and consistent anisotropic structures. , In a study, a bilayer wound dressing made of GelMA hydrogel and poly­(vinyl alcohol)-carboxymethyl cellulose (PVA-CMC) foam was created. Gas foaming was used to improve porosity and structural integrity. The gas-generated porous structure increased wound exudate absorption, promoted gas exchange, and encouraged cell adhesion. However, an observed constraint was the difficulty in sustaining mechanical strength, which may necessitate further cross-linking techniques to boost durability. Another study explored an anti-inflammatory hydrogel dressing made from HA and hydroxyethyl starch, using a gas foaming technique to create porosity. The resulting structure improved water retention and facilitated medication release. However, the study found that excessive porosity could reduce the hydrogel’s mechanical stability, resulting in rapid deterioration under physiological settings. A study found that a multifunctional hydrogel made of oxidized HA, ε-polylysine, and quaternized chitosan (QCS) can be highly porous and injectable by incorporating gas foaming. This design facilitated rapid hemostasis, cell proliferation, and self-healing. While the porous structure significantly improved tissue integration, the study found that the difficulty in optimizing pore size distribution to balance mechanical strength and biodegradability persists.

7.

Gas foaming creates highly porous constructs that are excellent at fostering cell infiltration. However, cross-linking is often required to increase construct mechanical strength. The freeze-drying process is almost identical to the gas foaming process, except that its pores are produced by directly freezing the polymeric solution at extreme temperatures.

4.5. Three-Dimensional Printing and Bioprinting

Scaffolds are built layer by layer using 3D printing, giving exact control over mechanical properties, geometry, and anisotropy. , It is more costly and time-consuming than other approaches, despite being very customizable. A study using digital light processing (DLP)-based 3D printing created functional living skin (FLS) constructs with GelMA/HA-NB/LAP bioink. The resulting scaffold had high cell viability, rapid gelation, and tunable mechanical properties, promoting dermal regeneration and vascularization. While DLP-based printing improved resolution and structural complexity, material degradation rates and mechanical mismatches are still concerns for long-term applications. Another study investigated solvent exchange deposition modeling (SEDM) for a bilayered flexible skin substitute composed of PLGA and EGF. In a rat model, the nano-/micro-structured scaffold improved fibroblast adhesion and tissue ingrowth, resulting in significantly faster wound healing. However, PLGA’s slow degradation rate and the need for bioactive surface modifications were noted as limitations. Another study looked at 3D-printed SA/polyethylene glycol (SA/PEG) scaffolds infused with Satureja cuneifolia extract for diabetic wound healing. These scaffolds demonstrated excellent antibacterial properties, biocompatibility, and controlled drug release, indicating potential for infection prevention and wound healing. However, optimizing mechanical strength while maintaining porosity was a challenge that was noted to require further material refinement.

Bioprinting is a promising technique developed based on 3D printing technologies (Figure ). By adapting conventional 3D printing principles to biological systems, bioprinting enables the spatially controlled deposition of cells, biomaterials, and bioactive factors to fabricate complex, tissue-like structures. Three-dimensional and four-dimensional bioprinting methods have recently gained significant traction for their ability to quickly and accurately fabricate complex biological microstructures from bioinks and natural polymers while simultaneously depositing live cells, antioxidants, and growth factors into a construct during manufacture. − Extrusion-based 3D bioprinting was used to create a decellularized ECM (dECM)/Gel/QCS scaffold with an average pore size of 90.66 ± 8.62 μm and an E-modulus of 74.31 ± 26.26 MPa, which exhibited excellent hemocompatibility (hemolysis ratio of 3.04 ± 0.47%), antibacterial properties (near-100% bactericide rate in E. Coli and S. aureus populations after culturing for 24 h), and revealed high L929 fibroblast cell densities and ECM secretion levels marked by expression of type-I collagen, fibronectin (21.43 ± 0.41 ng/mL), and decorin (13.57 ± 0.49 ng/mL) after 7 days of incubation. Another bioprinting method, laser-based 3D bioprinting, has been used to fabricate full-thickness skin substitutes composed of Hs27 fibroblasts encapsulated in Cellendes hydrogel bioink (maleimide-functionalized dextran with polyethylene glycol cross-linker) to form a complex vascular construct in which high levels of vimentin and types I and IV collagen were expressed after 41 days of culturing. Four-dimensional bioprinting has also recently emerged as the next phase of skin tissue engineering, capable of creating smart, dynamic, temperature and stimuli-responsive constructs. A light-responsive skin patch composed of Gel/Alg//HA/polyphenylene vinylene (PPV) with a 300 μg/mL PPV concentration demonstrated an 89.4% toxicity toward an S. aureus culture when irradiated with 60 mW/cm² light, while another GelMA-composed hydrogel coated with 1% concentration tetrapodal-ZnO and VEGF exhibited a controlled release of these elements in vivo when exposed to light treatment (547 nm), resulting in increased angiogenesis (marked by a 10.35 ± 0.46%/ 0.01 mm² expression of CD31), reduced inflammation (marked by decreased IL-6 cytokine levels and increased IL-10 cytokine levels), and wound reduction to 24.9 ± 4.21% and 3.48 ± 6.03% after 1 and 2 weeks, respectively. Similarly, pH-responsive, thermoresponsive, , and humidity-responsive hydrogels have all been shown to greatly augment wound healing in various ways, proving the immense future potential of 4D bioprinting for skin tissue engineering.

8.

Bioprinting is a state-of-the-art manufacturing process for tissue engineering, as it can produce complex bioactive constructs that foster enhanced wound regeneration without the need for additional modifications.

4.6. Artificial Intelligence-Guided Design

Anticipating the future, artificial intelligence (AI) is shaping up to play a significant contributing role to skin tissue engineering endeavors (Figure ). , A study using AI to optimize bioprinting conditions resulted in ALG-Gel scaffolds containing a highly aligned grid-shaped network with skin-compatible elastic moduli of (0.043–0.639 MPa) and porosities of (69.3–79.5%) that demonstrated a biocompatibility of >90% for human epidermal keratinocytes after a 48-hour incubation period. AI models have also been used recently to successfully predict polymer-based scaffold degradation rates and fibroblast scaffold performance using a correlation between parameters including fiber and pore diameters, mechanical properties, water contact angle, and MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay results. A machine learning model has also been used predictively to analyze the results of an scRNA-seq procedure through specific application to biomarker, diversity, plasticity, immune response, spatial arrangement, cell–cell communication, and regeneration trajectory data procured from various skin fibroblast subpopulations to mitigate the progression of fibrosis and the formation of keloid scars. Results from the experiment were promising, as the machine learning model was able to accurately predict the healing trajectories of several fibroblast types when compared to existing knowledge on fibroblast activity during the regeneration process. While some discrepancies between machine learning-predicted trajectories and known trajectories were noted, the accuracy of such technology is anticipated to be refined over time, allowing for the future development of wound healing treatments tailored to individuals and wound types that provide a clear framework for strategic decisions to be made regarding the implementation of biomimetic skin substitutes. Another study by Lu et al. (2025), while not directly implementing a skin substitute, illustrates the great potential for artificial intelligence-regulated external stimuli to be used in combination with external stimuli-responsive scaffolds/hydrogels. In the experiment, a machine learning model was used to regulate the intensity of an endogenous electric field exerted over, as well as dosages of fluoxetine administered to a porcine full-thickness wound model in response to being presented with images of the inflicted wounds every 2 h over the course of 7 days. The electric field was applied over the first day, while fluoxetine was administered for the rest of the 7 days. This AI-controlled setup, which used visual indicators of regeneration to regulate the application of external therapies, was reported to accelerate the wound healing process by 17.71% as compared to untreated wounds. Given these results, these studies illustrate the nascent potential for the use of AI as a tool for design and fine-tuning the manufacturing process for effective next-generation skin substitutes and reducing the magnitude of biological encounters in the implementation of biomaterial-based regenerative constructs. Nonetheless, several challenges hinder the integration of AI and machine learning into tissue engineering. A primary challenge is the lack of clear concepts and standardized terminology in biology. Intricate terms like “biocompatibility” lack precise, universally accepted definitions, which impedes the interpretability and efficacy of machine learning models. Additionally, the implementation is limited by problems with data quality, ethical issues, and a myriad of regulatory problems. , Therefore, future research should focus on improving model interpretability to ensure that clinicians and materials scientists can understand and trust AI-driven decision-making. Moreover, addressing ethical considerations in personalized scaffold design requires the adoption of differential privacy technologies to protect data confidentiality and the use of fairness algorithms to reduce algorithmic bias. Furthermore, the development of technologies such as physical information neural networks (PINNs) and federated learning, combined with the integration of artificial intelligence and traditional materials science expertise as well as enhanced multicenter data collaboration, is expected to substantially increase the precision and scalability of AI applications in tissue engineering. Table shows studies using different manufacturing methods in the application of skin tissue engineering and wound healing.

9.

Artificial intelligence-guided design can assimilate existing experimental data and subsequently perform predictive or regulative functions with regard to wound healing, facilitating an optimized level of repair through analysis of regenerative biological factors.

2. Summary of Various Manufacturing Methods Applied to Different Polymers.

Method	Materials	Cells	Results	Wound Healing Outcome	Source
Solvent casting/particle leaching	Collagen (Col) and chitosan (CS) in a 7:3 ratio	Human dermal fibroblasts (HDFs)	Improved re-epithelialization by 69.7 ± 9.6% on day 21. Limited mechanical flexibility with elongation at break of 2.4 ± 0.6%.	Enhanced wound healing.
Solvent casting/particle leaching	CS and gelatin (Gel) with varying compositions	NIH-3T3 fibroblasts and S5Y5 neuroblastoma cells	CS/Gel scaffolds with higher gelatin content resulted in higher adhesion. Long-term stability of gelatin in aqueous environments requires cross-linking.	Enhanced fibroblast attachment and proliferation.
Solvent casting/particle leaching	Tropoelastin (TE) and polyglycerol sebacate (PGS)	In vivo: BALB/c mice	TE-PGS Scaffold performed at an accelerated rate for wound closure, performing superiorly to the Single TE experiment and the PGS scaffold.	The reduced inflammation promoted a regenerative environment.
Solvent casting/particle leaching	CS, grade-1 gum kondagogu (GKG), Zinc Oxide nanoparticles (ZnO NP), and Barbaloin (BB)	In vivo: Wistar adult rats	The combined biocomposite film displayed a tensile strength of 8.9 ± 0.30 MPa, a water absorption rate of 451.1 ± 6.02%, and good biocompatibility. The film showed antioxidant effects as well as antimicrobial properties.	In vivo accelerated healing capabilities and minimal scarring.
Solvent casting/particle leaching	Starch and TMPyP (5,10,15,20-tetrakis (1-methylpyridinium-4-yl) porphyrin)	Ex vivo: pig abdominal skin	TMPyP Enhanced mechanical properties such as flexibility and increased elasticity and promoted endothelial cell migration.	Enhanced wound healing through antibacterial activity.
Electrospinning	Polycaprolactone (PCL) and CS	Fibroblasts	The skin-like nanopad (SLN) made from chitosan and PCL showed properties of high flexibility. SLN blocked bacterial penetration from Staphylococcus Aureus and Pseudomonas Aeruginosa.	Wound healing capabilities of the scaffold showed a 60% wound closure by day 7.
Electrospinning	Degummed silk fibers from Bombyx mori silk cocoons.	Keratinocytes, fibroblasts, endothelial cells	The bilayered scaffold with silk fibroin enhanced epidermal and dermal regeneration, fibroblast maturation, and vascularization.	Scar formation was minimized and the scaffold demonstrated organized epidermal and dermal layers.
Electrospinning	Polyether ether ketone (PEEK) and Aloe Vera powder	Mouse fibroblasts (3T3), immortalized human epidermal keratinocytes (HaCaT)	The PEEK nanofiber scaffold exhibited high porosity (85%) and enhanced hydrophilicity. It also improved cell adhesion and proliferation. Live/Dead assays confirmed high cell viability.	Scarless wound healing with complete regeneration.
Electrospinning	Polyvinyl alcohol (PVA) and konjac glucomannan (KGM)	3T3 fibroblasts, flood samples of Sprague Dawley (SD) rats	PVA/KGM resulted in a mechanical strength of 2.5 MPa and a faster wound closure. In addition, at a (5:5) ratio of KGM/PVA, the wound was healed by 98% after 21 days. The combined scaffold had higher fibroblast growth.	Enhanced wound healing via enhanced fibrinogen absorption.
Electrospinning	PCL, Gel, and anticancer drug bleomycin (BLM)	HaCaT and human epidermoid carcinoma cells (A431)	PCL-Gel-BLM had an elongation of 18.53%, a tensile strength of 15.25 MPa, and an elongation of 26.35%. Tests on A431 cancer cells showed reduced proliferation, showing promise for anticancer treatment.	Enhanced wound healing through antibacterial and anticancer activity.
Freeze drying*	Sodium alginate (SA) and phosphate-buffered saline (PBS)	In vitro: human fibroblast cells, in vivo: NMRI mice	SA scaffolds had lower compressive strength than standard calcium alginate, and greater than 80% cell viability.	Organized collagen deposition with approximately 90% wound healing.
Freeze drying	N-hydroxy succinimide (NHS) and 1-(3 dimethylaminopropyl)-3-ethyl-carbodiimide hydrochloride (EDC)	In vitro: rat fibroblast Cells (L929)	The adhesion strength ranged from 5 to 16.96 kPa, reduced blood loss by 74% compared to controls, and achieved 97% wound closure.	Enhanced wound healing and hemostasis.
Freeze drying	Gelatin methacryloyl (GelMA), PCL, and PVA	Mammalian cells (murine dorsal skin cells and liver tissue), bacterial strains (E. coli)	The nanosheet reduced clotting time to 4 minutes with Ca2+ promoting coagulation as well as reduced blood loss.	Wound healing in the dorsal skin cells and liver models showed efficient healing and wound closure.
Freeze drying	Tripeptide glycyl-l-histidyl-l-lysine (GHK), flaxseed gum (FSG), and low methoxyl amidated (LMA)	In vivo: Wistar adult rats	LMA pectin hydrogel had higher elastic properties, FSG retained water and had a highly porous structure, LMA had lamellar microstructure, FSG hydrogel showed faster and more sustained drug release.	Accelerated wound healing with improved tissue repair.
Freeze drying	Col I and SA	Human foreskin fibroblast cells (HFFs), human keratinocytes	The hydrogel had improved mechanical strength and structural integrity, enhanced cell adhesion in both fibroblasts and keratinocytes, and accelerated wound healing.	Enhanced skin remodeling and re-epithelialization.
3D printing	Poly(lactide-co-glycolide) (PLGA), DOPA-modified epidermal growth factor (EGF), and bovine serum albumin (BSA)	NIH-3T3 fibroblasts, Escherichia coli (E. coli), in vivo: SD adult rats	The scaffold exhibited a porous and rough surface. Tensile strength increased by 57.07% and acted as a barrier against bacteria. Moreover, it increased fibroblast proliferation and growth.	Enhanced wound healing rate, achieving 98.88% wound closure by day 21.
3D printing	SA, polyethylene glycol (PEG), and Satureja cuneifolia (SC) extract	L929 mouse fibroblast cells, bacterial strains such as E. coli	SA/PEG scaffold showed high porous structure inciting fibroblast cell adhesion. PEG improved the flexibility and elongation of the scaffold.	Enhanced wound healing with antibacterial activity.
3D printing	GelMA, N-(2-aminoethyl)-4-(4-(hydroxymethyl)-2-methoxy-5-nitrosophenoxy) Butanamide (HA-BN)	Human skin Fibroblasts (HSF), human umbilical vein endothelial cells (HUVEC), in vivo: SD adult rats, Bama miniature pigs	The hydrogel scaffold showed rapid gelation ensuring mechanical strength, dense upper layer that mimicked the epidermis. HSF and HUVEC showed 95% viability, DNA content doubled for HUVEC and tripled for HSF.	Enhanced skin regeneration and the wounds showed 98.8% closure by day 21.
3D printing	CS, AA, Polyzwitterion-based Copolymer, and Poly(sulfobetaine-co-acrylic acid) (PBCP)	No biological cells were used	Enhanced elongation at break of approximately 800% and a self-healing efficiency of 95.4%. It also exhibited strong adhesion to diverse surfaces and maintained properties after multiple peeling cycles.	Not reported
3D printing	CS and PLA	Mouse fibroblast cells (L929)	Enhanced mechanical properties of the scaffold were documented, as well as good biocompatibility with no signs of cytotoxicity.	Not reported
Gas foaming	Diazirine-modified gelatin (GelDZ), Silver Nitrate (AgNO3)	Mouse 3T3 fibroblasts, BALB/c mice	GelDZ hydrogel exhibited strong adhesive properties, and supported 3T3 fibroblasts viability and proliferation. Additionally, antibacterial zone of inhibition was significantly larger for GelDZ-Ag NPs.	Infected wounds treated with GelDZ-Ag NPs healed significantly faster than untreated wounds.
Gas foaming	Quaternized chitosan (QCS) and EGF	L929, HUVECs	The hydrogel demonstrated enhanced mechanical properties, burst pressure, and gelation time. Nearly 100% bacterial death against S. aureus and E. coli. Liver bleeding reduced from 783 mg (control) to 256 mg (Hydrogel).	A complete wound closure within 10 days.
Gas foaming	Oxidized hydroxyethyl starch (OHES), Hyaluronic Acid-Adipic Dihydrazide (HA-ADH), and Xanthatin (XT)	RAW 264.7 macrophages, SD adult rats	The hydrogel exhibited strong adhesion, high swelling ratio, low hemolysis (0.8%), and high cell viability (> 90% RAW 264.7 macrophages).	In vivo wound healing showed accelerated healing up to 89.1% by day 20.
Gas foaming	GelMA, Poly(ethylene oxide) (PEO), and Polyethylene Glycol (PEG-DBCO)	HUVECs, mouse mesenchymal stem cells (MSC), macrophages (CD68), regulatory T cells, mice (C57BL/6)	The hydrogel resulted in a 4x density of blood vessels, more uniform endothelial and smooth muscle cell integration, and 10x reduction in pro-inflammatory M1 macrophages.	Scarless wound healing with hair follicle formation.
Gas foaming	Gel, Silver nanoparticles (Ag NPs), QCS, and Oxidized Pullulan	L929 Mouse fibroblasts, RAW 264.7 macrophages, SD adult rats	The hydrogel’s properties showed rapid self-healing, controlled degradation over 14 days, and swelling ratio exceeded 95%. The hydrogel inhibited 99.3% of S. aureus and 98.7% of E. coli growth.	82.6% in vivo wound closure by day 10
Bioprinting	Decellularized ECM (dECM), Gel, and QCS	L929 fibroblasts	The extrusion-printed dECM/Gel/QCS scaffold strongly promoted fibroblast proliferation and ECM production, exhibiting high L929 cell density, and antibacterial activity.	Robust expression of type-I collagen, fibronectin, and decorin.
Bioprinting	Maleimide-functionalized Dextran, and PEG	Hs27 fibroblasts	The laser-printed full-thickness skin substitute supported long-term fibroblast viability and maturation.	Sustained expression of vimentin and collagen types I and IV after 41 days of culture.
Bioprinting	Gel, Alginate (Alg), HA, and polyphenylene vinylene (PPV)	Staphylococcus aureus for antibacterial assay	The light-responsive Gel/Alg/HA/PPV hydrogel enhanced wound-relevant antimicrobial performance by achieving high phototoxicity against S. aureus.	Enhanced wound healing with antibacterial activity.
Bioprinting	GelMA, Tetrapodal zinc particles (1% concentration), and vascular endothelial growth factor (VEGF)	No cells were used	The light-activated GelMA hydrogel with tetrapodal-ZnO particles and VEGF by promoting angiogenesis and reducing inflammation.	Significantly accelerated wound healing in vivo, achieved near-complete wound closure within 2 weeks.

5. Current Status of Experimental and Commercial Skin Substitutes

As of 2020, more than 70 commercially available skin substitutes had been identified by the National Institutes of Health (NIH). While many of these products have entered mainstream clinical use following rigorous scientific validation and industrial development, a substantial number remain investigational. In the United States, the Food and Drug Administration (FDA) has established a series of regulatory pathways that skin tissue substitutes must follow after passing the experimental stage and prior to commercialization (Figure ). This framework incorporates a risk-based classification system for emerging medical technologies, in which class I (low-risk) and class II (moderate-risk) devices are typically reviewed through the 510­(k) premarket notification pathway. In contrast, Class III (high-risk) products, including novel or insufficiently tested technologies and those incorporating human- or animal-derived biological components, are generally subject to more stringent review through the Premarket Approval or Biological License Application pathways, depending on product composition and manufacturing processes. Clinical trials hinder the translation of many skin substitutes between the experimental and commercial realms through the prolonged, complex nature of the process itself, which may be beyond the means of research groups designing and fabricating novel constructs. Additionally, the recruitment of sample populations for testing of a skin substitute is extremely challenging, as an experimental skin substitute may be tailored toward specific injury types and may not be implemented in individuals with comorbidities or preexisting medical conditions. Consequently, the recruitment phase of a clinical trial may become exhaustively drawn out, and when added to a necessary post-treatment phase for long-term observation of wound healing outcomes, delaying the procurement of reliable results and incurring drastic costs to ensure that all biosafety and medical care requirements are met.

10.

General commercial approval process for skin substitutes is depicted. The duration and costs associated with all combined phases are extreme, preventing many experimental skin substitutes from successfully achieving commercialization.

5.1. Promising Clinical Trials of Experimental Skin Substitutes (2021–2025)

A 2021-updated clinical study (NCT02870816) presented a detailed publication comparing the effectiveness of AmnioBand, a dehydrated human amnion/chorion allograft, and Apligraf, a tissue-engineered skin substitute, on patients suffering from stubborn diabetic foot ulcers. For sample sizes of 30 patients per treatment group, it was reported that 77% of ulcers treated with AmnioBand and 23% of ulcers treated with Apligraf were closed at 6 weeks. At 12 weeks, 90% of ulcers treated with AmnioBand and 40% of ulcers treated with Apligraf were closed, with the respective mean time to heal in each group reported as 32 days and 63 days. Another Phase 1 clinical trial (NCT02145130) conducted a long-term evaluation of the performance of denovoSkin, a hydrogel infused with a patient’s cultured dermal and epidermal cells, on 10 patients. Over a period of 5 years, 7 out of 10 injury sites proved stable, resulting in average Patient and Observer Scar Assessment Scale (POSAS) scores of 3.44 ± 2.03 for vascularity, 3.21 ± 2.33 for pigmentation, 2.47 ± 1.16 for pliability, and 2.15 ± 1.23 for relief. Another commercial skin substitute, Hyalomatrix, a hyaluronic acid scaffold layered onto a silicon membrane, was tested in a clinical study that was most recently updated in 2023 (NCT03107546). However, pending the publication of results, the study refers to another 2014 paper describing a similar study testing the use of Hyalomatrix on 23 children who had undergone a syndactyly release surgery. Although wound closure times were not explicitly reported, the authors noted that the wound sites exhibited near-normal pigmentation and pliability, along with normal vascularity and no reported discomfort over an extended follow-up period. Glyaderm, a glycerolized acellular dermis graft implemented in a single-stage bilayer skin reconstruction procedure, was evaluated in a 2024-updated clinical study (NCT06249971) sponsored by the University of Ghent over its applicability to full-thickness burns. The use of Glyaderm combined with an autologous split-thickness skin graft was compared to the use of an autologous split-thickness skin graft on its own, with various characteristics evaluated at 3, 6, 9, and 12 months after intervention surgeries. Wounds treated with the Glyaderm graft saw an average time to full wound closure of (1.58 ± 0.95 months), and a mean graft take of (96.22% ± 5.4%) with no adverse effect due to infection or a lack of vascularization. At 12 months post-surgery, injury sites that had received the Glyaderm graft had a mean hydration value of (30.91 ± 13.75), a mean trans-epidermal water loss value of (13.01 ± 6.53), and erythema and melanin index values of (368.68 ± 86.7) and (215.68 ± 127.37), respectively. The publication of these results further reports that there was no significant difference in these qualities exhibited by the wounds that had received the Glyaderm treatment and wounds that had received the split-thickness skin graft, noting, however, that the Glyaderm treatment produced an increased long-term scar quality. A randomized clinical study (NCT06873867) by the Adichunchanagiri Institute of Medical Sciences completed in November 2025, compared the performance of Helicoll, a high-purity (>97%) type 1 collagen (HPTC) skin substitute and dehydrated human amnion/chorion membranes (dHACM) in the treatment of patients with diabetic foot ulcers. Over a 4-week study period, the study reported that patients receiving the HPTC treatment exhibited a mean wound size reduction of 86.48% (5.48 ± 2.23 cm²) with full wound closure occurring in 10 out of 14 patients, as compared to a mean wound size reduction of 77.7% (4.3 ± 3.21 cm²) with full wound closure occurring in 7 out of 14 patients receiving the dHACM treatment. Additionally, the average ulcer size in the HPTC treatment group shrank to 5.33 cm² as compared to 22 cm² in the dHACM treatment group by the end of the study, significantly underscoring the superior wound healing capability of Helicoll. Still another clinical study (NCT05944250) sponsored by Stanford University, updated in July 2025, evaluated the efficacy of Spincare Matrix, an electrospun nanofibrous polymer-based skin substitute delivered by a handheld device, on skin wounds in patients with Recessive Dystrophic Epidermolysis Bullosa (RDEB). The published results acknowledged that there was no significant difference in wound closure times or areas between a 21-patient group receiving the Spincare Matrix treatment and a 21-patient control group over a 4-month study period but pointed out that the Spincare Matrix notably produced no adverse effects whatsoever. In January 2025, Ohio State University’s Wexner Medical Center described the successful pioneer use of Cohealyx Collagen Dermal Matrix (AVITA Medical) to accelerate dermal regeneration of burns and deep cuts in combination with an autologous skin graft through a two-step procedure, reporting graft times of 2–4 weeks depending on the utilized treatment option. Currently, AVITA Medical is recruiting for a clinical trial (NCT06787690) to confirm these results, with an expected study completion date set for June 2026. Similarly, another technology known as Genetically Modified Epidermolysis Bullosa Self-Assembled Skin Substitute (GMEB-SASS) is involved in a recently begun clinical trial seeking recruits (NCT07193134). This skin graft, which is engineered from a sample of a patient’s skin and contains living fibroblasts and keratinocytes meant to produce Type VII Collagen, is anticipated to assist patients with Recessive Dystrophic Epidermolysis to achieve wound healing through permanent integration. The study is expected to be completed in 2035. Table summarizes the recent clinical studies associated with commercial skin substitutes.

3. Summary of Recently Updated Clinical Studies Involving Commercial Skin Substitutes.

Study/Trial ID	Product Description	Wound Type	Results	Most Recent Update
NCT07193134	Genetically modified Epidermolysis bullosa self-assembled skin substitute (GMEB-SASS)	Skin wounds in patients with Recessive Dystrophic Epidermolysis Bullosa (RDEB)	Expected to increase production of Type VII collagen through engineered patient-derived fibroblasts, and keratinocytes.	Expected 2035
NCT06787690	Cohealyx (AVITA Medical)	Burns, deep cuts	Reported graft times of 2–4 weeks, faster than other similar products (published by Ohio State University, Wexner Medical Center)	Expected 2026
NCT06873867	HeliColl; high-purity >97% Type-1 Bovine collagen skin substitute (EnColl Corp)	Diabetic foot ulcers	Mean wound size reduction of 86.48% (5.48 ± 2.23 cm2), average final wound size of 5.33 cm2, full wound closure in 10/14 patients in treatment group over 4-week study period.	November 2025
NCT05944250	Spincare Matrix; electrospun nanofibrous polymer-based film delivered by handheld device (Nanomedic Technologies Ltd.)	RDEB skin wounds	No significant difference in wound closure times and areas with control group; no adverse effects reported over a 4-month study period in 21 patients.	July 2025
NCT06249971	Glyaderm; glycerolized acellular dermis, single-stage grafting procedure (Euro Skin Bank)	Full-thickness skin burns	Average time to full wound closure of (1.58 ± 0.95 months), mean graft take of (96.22% ± 5.4%); reported comparatively increased long-term scar quality.	February 2024
NCT03107546	Hyalomatrix; hyaluronic acid matrix (Anika Industries, Inc.)	Syndactyly release surgery wounds in children	Wound closure times not reported; almost normal pigmentation, pliability, vascularity; no complaints of discomfort.	March 2023
NCT02870816	AmnioBand; dehydrated human amnion/chorion allograft (MTF Biologics)	Nonhealing diabetic foot ulcers	90% of AmnioBand-treated ulcers were closed at 12 weeks, with mean time to heal = 32 days.	December 2021
	Apligraf; tissue-engineered skin substitute (Organogenesis)		40% of Apligraf-treated ulcers were closed at 12 weeks, with mean time to heal = 63 days.
NCT02145130	denovoSkin; hydrogel infused with autologous dermal, epidermal cells (CUTISS)	Acute cases, reconstructive cases	Wound closure times not reported; stable sites in 7/10 patients; POSAS scores are 3.44 ± 2.03 for vascularity, 3.21 ± 2.33 for pigmentation, 2.47 ± 1.16 for pliability, and 2.15 ± 1.23 for relief.	March 2021

5.2. Summary Evaluation of Key Brands

Commercially available skin substitutes (Figure ) are categorized based on the source of their components and the design principles they employ. Because the microstructure and properties of native skin tissue are of paramount importance for facilitating complication-free wound healing, human and animal tissue sources are used almost ubiquitously in the construction of scaffolds/grafts for skin burn/wound healing. However, other medical industry tentpoles, which are fabricated from strategic blends of biomaterials, have also proved efficacious. Additionally, various skin substitutes are prepared to contain live cells and growth factors to further enhance regeneration, while still other skin substitutes display optimal performance as acellular constructs. Nevertheless, a common limitation of many commercial skin substitutes appears to be their sole focus on achieving rapid wound closure rather than facilitating appendage restoration and scar-free wound healing. Furthermore, a primary limitation remains the industry’s focus on rapid wound closure over functional appendage restoration or scar-free healing. While current designs emphasize porosity and anisotropy to mimic native ECM, they often overlook thermal or bioelectric properties. Moreover, skin substitutes are often very highly priced, with 4 in² (2 in × 2 in) Integra sheet for fingertip wounds/burns priced at $1978 as of 2012, while a 2013 review of hospital invoices in a certain locality revealed that a 44 cm² Apligraf sheet cost $1704, a 10.5 cm² Oasis wound matrix cost $91, and a 38 cm² Dermagraft sheet cost $1590 with successful healing rates of only 40, 55, and 50%, respectively. These figures were reported without considering the prices of the duration of care. Thus, it is evident that commercial skin substitute efficacy is not necessarily guaranteed, even at high costs. The following sections will briefly enumerate choice examples to present a solid perspective on the different design principles and materials that are currently used.

11.

Skin substitute categories based on design and component sources. While current trends seem to favor naturally sourced components, procurement and purification are complicated, expensive processes that ultimately drive up the cost of skin substitutes.

5.2.1. Human Skin-Derived Acellular Skin Substitutes

Products in this category include AlloPatch, AlloSkin, Coll-e-derm, DermACELL, DermaPure, DermaSpan, FlowerDerm, GammaGraft, GraftJacket, hMatrix, InteguPly, Matrix HD, and Theraskin.

These donated skin allografts from human cadavers are purified from cells and lipids, producing soft tissue scaffolds that are extremely conducive to biological repopulation and highly capable of catalyzing the skin regeneration process without immunogenic responses. − anisotropy, In example, DermaPure (human acellular dermal matrix purified from 99% donor DNA) closed hard-to-heal-wounds (mean presence before treatment 33 ± 7 weeks) with large sizes (13 ± 24 cm²) on an average of 8 ± 21 weeks faster compared to other treatments; DermACELL (human acellular dermal matrix purified from 97% nucleic acid presence) demonstrated wound closure percentages of 65% after 12 weeks (compared to 41.1% closure from standard care) and 82.5% after 16 weeks (compared to 48.1% closure from standard care) after a single application to diabetic foot ulcers, and resulted in a well-healed injury site with little scar formation after observation periods of 30 and 180 days when applied to second and third-degree burns; AlloPatch Pliable (human acellular dermal matrix retaining amounts of collagen types I, III, IV, VI, elastin, fibronectin, laminin) successfully healed a diabetic foot ulcer in 2 weeks in one case (44 year-old male with Type 2 diabetes; final HgA1c level 9.9%), in 1 week for another case (72 year-old male with Type 2 diabetes; final HgA1c level 8.2%), and in 5 weeks for a third case (61 year-old male with Type 2 diabetes; final HgA1c level 10.2%); TheraSkin (human acellular dermal matrix containing extracellular matrix and growth factors VEGF, IGF-1, FGF-2, TGF-β1), applied to diabetic foot ulcers (mean wound size 6.2 ± 11.8 cm²) and veinous leg ulcers (mean wound size 11.8 ± 22.5 cm²), had closed 60.38% of the former and 60.77% of the latter at week 12, and 74.1% of the former and 74.6% of the latter at week 20.

5.2.2. Human Placenta/Amnion/Chorion-Derived Acellular Skin Substitutes

Notable examples within this classification are AlloWrap, AltiPlast, AmnioBand, AmnioExcel, AmnioFill, AmnioFix, Amniomatrix, Artacent, BioDFactor, BioDFence, Biovance, Cellesta Amniotic Membrane, Cygnus, Dermavest, Plurivest, EpiCord, EpiFix, FlowerAmnioPatch/FlowerAmnioFlo, Genesis Amniotic Membrane, Integra BioFix, Interfyl, Matrion, Neox, NuShield, PalinGen, Restorigin, WoundEx Membrane/WoundEx Flow, and Xwrap.

Similarly, grafts composed of decellularized human placenta/amnion/chorion tissues possess qualities that provide them with excellent biocompatibility but possess the added advantage of retaining a myriad of glycoproteins, cytokines, cells, and growth factors, which can be transferred to an injury site. − Besides AmnioBand, which was featured in a clinical trial mentioned in the previous section, another 2025 study involved the use of Matrion, a full-thickness acellular placental trophoblast membrane retaining a high concentration of growth factors (TGF-β1 (114 ± 49 pg/mL), basic fibroblast growth factor (BFGF) (1385 ± 323 pg/mL), EGF (193 ± 14 pg/mL), PDGF (202 ± 19 pg/mL), IGF-1 (846 ± 186 pg/mL), VEGF (87 ± 36 pg/mL)), applied to a modified-intention-to-treat population and a per-protocol population with diabetic foot ulcers, resulting in 39% of ulcers closed in the first population and 48% of ulcers closed in the second population at 12 weeks period as compared to 26 and 27% of wounds closed at 12 weeks from standard of care in both populations, respectively. Other noteworthy clinical studies have evaluated the regenerative capability of EpiFix (NCT01693133), AmnioClear (NCT02120755), Neox (NCT02166294), and AmnioExcel (NCT02209051) within similar contexts. Additionally, other publications have evaluated the successful use of AmnioFix and Biovance, demonstrating the ability of decellularized human placenta/amnion/chorion scaffolds to treat severe, nonhealing wounds and ulcers.

5.2.3. Animal-Derived Acellular Skin Substitutes

Representatives of this group are Architect, Cytal, EZ Derm, DermaGide, Helicoll, Integra Matrix, Kerecis, Wound Dressing, MicroMatrix, MiroDerm, OASIS, Puracol, PuraPly, and Talymed. Animal-derived tissues are also widely used in commercial skin substitutes because they are readily available and provide significant support for wound healing. Additionally, since the regeneration process for injuries in pigs (porcine), sheep (ovine), and cows (bovine) has been determined to be similar to that occurring in the human body, using scaffolds composed of biological materials obtained from these species has been identified to be both beneficial and safe. From the above list, OASIS is derived from the small intestinal submucosa of pigs, MiroDerm from pig livers, and Cytal and EZ Derm from pig skins. OASIS was determined to be superior Regranex becaplermin gel in the treatment of diabetic ulcers, closing 53% of wounds as compared to 35% closed by Regranex at 12 weeks, and superior to Hyaloskin (topical hyaluronic acid) when used to mitigate arterial/venous ulcers, achieving full closure of 82 ± 6% of treated ulcers at 16 weeks compared to 46 ± 2% of ulcers closed after treatment with Hyaloskin at 16 weeks. Helicoll, a skin substitute with a 97% content of high-purity Type 1 Bovine-derived collagen, was briefly considered in the previous section of clinical trials and exhibited greater efficacy than a graft composed of decellularized human amnion/chorion matrix. Besides porcine, bovine, and ovine-derived substitutes, Kerecis, a 2019-approved scaffold composed of omega-3 oil-rich fish skin has garnered recent attention for its significant bearing on wound healing. A 2025 study investigated the outcomes of the use of Kerecis Omega-3 on enzymatically debrided deep burn wounds and determined a mean wound healing time of 16.77 ± 6.14 days due to its close resemblance to human extracellular matrix and secretion of omega-3, which is highly likely to be anti-inflammatory and antimicrobial.

5.2.4. Biomaterials-Based Acellular Skin Substitutes

This category is exemplified by Hyalomatrix, Integra Dermal Regeneration Template (IDRT) engineered skin grafts, composed of natural and synthetic polymers, which are a prevalent alternative to foster wound healing because they combine the best characteristics of various elements into complex, multifaceted constructs through controlled manufacturing processes, producing a proven record of optimal outcomes. In addition to the study involving its application to syndactyly release surgeries, Hyalomatrix (HYAFF 11/esterified hyaluronan on silicone membrane) was used in a separate study to treat 12 cases of serious wounds where muscle, tendons, or bone were exposed and necrotic tissue was present. The results indicated that 75% of cases achieved complete healing with a mean healing time of 37.9 days, while the other 25% of cases required intervention but ultimately also achieved complete healing after a mean time of 42 days. Of all commercially available skin substitutes, the IDRT, “a bilayered microporous scaffold composed of Type 1 Bovine collagen, glycosaminoglycan, and chondroitin-6-sulfate matrix”, has arguably received the most scientific backing in terms of academic publications after FDA approval in 1996. Recent work affirming the efficacy of the IDRTs, including a 2025 publication, in which the template was applied to 13 cases of skin reconstruction after the removal of cutaneous tumors (mean defect size 105.9 cm²), resulted in a take rate of 92.3% with a mean grafting period of 20 days. Moreover, earlier publications concern the efficacy of the IDRTs to treat deep burn wound healing, − severe wounds and tissue defects, − and remarkable scar management capabilities.

5.2.5. Living Cellular Skin Substitutes

Key representatives of this category consist of Dermagraft, Affinity, Apligraf, SkinTE, and Grafix. A recent trend in the world of regenerative medicine involves combining certain types of living cells with an ECM-like scaffold or hydrogel to further enhance the speed, quality, and completeness of the regeneration of an injury site. Therefore, several skin substitutes, including Dermagraft, a biodegradable polyglactin membranous scaffold holding a culture of neonatal-derived fibroblasts, and Apligraf, a bilayered graft composed of human keratinocytes and fibroblasts in a type 1 bovine collagen matrix, are popular, effective choices for wound treatment. A 2016 study reported that out of 13,193 cases of treatment for diabetic foot ulcers between 2011 and 2014, Dermagraft was used to treat 41.9% of those cases and closed 58% of treated ulcers at 90 days, while Apligraf was used to treat 37.3% of cases, also closing 58% of treated ulcers at day 90. The capability of Apligraf has been further described in other publications since the early 2000s, with a 2000 paper documenting that the application of Apligraf to thermal and chemical burn wounds in 38 patients resulted in wound closure in 89% of patients by week 2 and >75% wound closure in 95% of patients by 1 month, with 47% of patients displaying normal vascularity and 61% of patients exhibiting normal pliability in the grafted sites by 24 months. The capability of Apligraf has been further described in subsequent publications since the early 2000s, with a 2000 study reporting that the application of Apligraf to thermal and chemical burn wounds in 38 patients resulted in wound closure in 89% of patients by week 2 and greater than 75% wound closure in 95% of patients by 1 month. At 24 months, 47% of patients exhibited normal vascularity and 61% demonstrated normal pliability at the grafted sites.

6. Conclusion and Future Perspectives

In conclusion, the complexity of the skin, with its diverse cell types, multilayered structure, and functional appendages, presents a significant challenge in the development of effective skin substitutes. Replicating native skin functions, including protection, temperature regulation, and fluid control, requires careful consideration of the biomaterial selection, mechanical properties, and fabrication strategies. Natural polymers provide bioactive cues that promote cell adhesion and proliferation but often lack structural stability, whereas synthetic polymers offer tunable mechanical properties and degradation profiles but may require surface modification to enhance their biocompatibility. Therefore, composite systems represent a promising strategy for integrating the advantages of both material classes. In addition, the incorporation of conductive polymers and nanoengineered components introduces bioelectrical and nanoscale cues that regulate cellular behavior, enhance angiogenesis and nerve regeneration, and improve cell-material interactions. Additionally, fabrication techniques such as electrospinning, freeze drying, gas foaming, and 3D printing enable control over scaffold architecture and function, although challenges remain in achieving optimal mechanical strength, degradation behavior, and clinical scalability.

Ultimately, achieving scar-free skin regeneration requires a shift from prioritizing rapid wound closure toward restoring native tissue architecture and function. Scar-free healing is governed by the coordinated regulation of inflammation, controlled collagen deposition, and balanced mechanotransduction signaling. Advanced scaffold designs that integrate biomimetic microstructures, immunomodulatory properties, and dynamic mechanical and bioelectrical cues have demonstrated the ability to suppress excessive myofibroblast activity and promote regenerative healing over fibrotic responses. Furthermore, successful regeneration of skin appendages, including hair follicles, glands, and nerves, is essential for restoring full skin functionality. Despite significant progress, current clinical skin substitutes remain limited in their ability to consistently achieve scar-free outcomes, often prioritizing wound closure over complete regeneration. Based on current design trends discussed in this work, the skin substitutes of the future that accomplish the full realization of “scarless” or “scar-free” healing will most likely be living, bioactive multilayered constructs with anisotropic, porous microstructures that possess appropriate stiffnesses, elasticities, and conductivities, fabricated through advanced high-precision manufacturing technologiese.g., 3-dimensional and 4-dimensional bioprintingthat allow for the construction of intricate structures from cells, growth factors, and other powerful biological drivers of wound healing. Artificial intelligence/machine learning has the potential to play a major role in the design and optimization of these constructs through material selection and manufacturing process optimization, as well as in predicting wound healing trajectories and applying external stimuli to accelerate and improve regeneration via the interaction between these stimuli and an implemented responsive biomaterial construct. Thus, current work to develop “next level” wound dressings with increased effectiveness for inhibiting scar formation while remaining accessible and affordable may benefit from (1) doubling down on collaboration-heavy efforts to ensure that the mechanical properties, biochemistry, and microarchitecture of the native skin are replicated with a greater degree of accuracy through (2) increased incorporation of natural materials such as elastin and collagen, along with ethically sourced extracellular matrices, proteins, cytokines, interleukins, and growth factors, which have been exhaustively proven to be powerful agents for scar-free healing, and (3) the improvement of currently available manufacturing and refinement technologies to allow for greater ease of biomaterial synthesis and procurement, greater levels of biomimetic microarchitectural complexity, and a more straightforward and tunable method for the incorporation of the mentioned biological elements while speeding up fabrication times, all of which are crucial for ensuring widespread availability and price reduction for skin substitutes.

The authors declare no competing financial interest.