Emerging Polysaccharide-Based Biomaterials in 3D Bioprinting: Materials, Biomedical Applications, and Future Perspectives

Abstract

Contemporary healthcare systems require advanced, sustainable, and patient-specific therapeutic solutions. We need advanced technologies, exemplified by 3D printing and biomaterials such as carbohydrates, to produce many 3D-bioprinted in vitro tissues and organ models for drug testing and regenerative medicine. In recent years, polysaccharide-based biomaterials have emerged as promising candidates for three-dimensional (3D) printing applications owing to their intrinsic biocompatibility, biodegradability, tunable physicochemical properties, and structural resemblance to native extracellular matrices (ECMs). In addition, recent progress in biomaterial modification and 3D printing methodologies has broadened their biomedical applications, particularly in tissue engineering, regenerative medicine, organ model development for drug discovery, and in situ bioprinting. This review presents a comprehensive overview of widely utilized polysaccharides in regenerative medicine, 3D printing techniques, bioink formulations, and the biomedical applications of 3D systems. It highlights recent advancements in material design, fabrication strategies, and biological performance and addresses current challenges and future perspectives for translating these materials from laboratory research to clinical practice.

1. Introduction

A range of techniques have evolved to address persistent challenges in healthcare, medical facilities, the pharmaceutical industry, and drug delivery. Technological advances in 3D printing have opened new opportunities for developing tailored biomaterials for complex biomedical applications. 3D printing, or additive manufacturing, enables the precise fabrication of complex biological constructs through the layer-by-layer deposition of cells, biomaterials, and bioactive molecules. Various 3D printing techniques have been employed to fabricate tissue and organ scaffolds. According to the ISO/ASTM 52900 standard, additive manufacturing techniques are categorized into Material Extrusion, Binder Jetting, Powder Bed Fusion, Vat Polymerization, Direct Energy Deposition, Material Jetting, and Sheet Lamination (Figure a). These methods have been applied in biomedical fields such as dentistry, medical devices, tissue engineering, neural regeneration, drug delivery, and organ printing to produce anatomical models, instrumentation, and prosthetics (Figure b). However, the success of 3D printing in tissue engineering hinges on the development of printable biomaterials that support printability, structural integrity, and biological function.

1.

(a) Various additive manufacturing techniques employed for 3D printing. (b) Overview of biomedical applications with the 3D printing technique for tissue engineering applications. (c) Classification chart of polysaccharides based on the monosaccharide units present in the polymers, as homopolysaccharides and heteropolysaccharides. (d) Classification chart of polysaccharides based on their origin as plant-based and animal-based polysaccharides. The author created the figure through BioRender scientific illustration software.

Polysaccharides have garnered significant attention among biomaterials for 3D printing because of their natural abundance, biocompatibility, biodegradability, and structural similarity to components of the native extracellular matrix (ECM). Polysaccharides are high molecular weight carbohydrate polymers composed of repeating monosaccharide units linked by glycosidic bonds. They are classified as homopolysaccharides, containing a single type of monosaccharide (e.g., starch, cellulose, chitin, inulin, and glycogen), or heteropolysaccharides, comprising two or more different monosaccharides (e.g., hyaluronic acid, pectin, peptidoglycans, carrageenan, and chondroitin sulfate) (Figure c). Additionally, polysaccharides are categorized by their source, primarily plant- or animal-based (Figure d). Plant-derived polysaccharides, such as cellulose, gum arabic, starch, alginate, and pectin, are widely used in biomedical, pharmaceutical, and cosmetic applications because of their abundance and biocompatibility. Animal-derived polysaccharides, such as glycogen, hyaluronic acid, chondroitin sulfate, and chitin, are recognized for their unique bioactivity, biodegradability, and therapeutic potential.

In recent years, polysaccharide-based bioinks have broadened the biomedical applications of 3D bioprinting, including skin regeneration, bone repair, cartilage reconstruction, vascular tissue fabrication, controlled drug delivery, and in vitro modeling. Frequently studied polysaccharides, such as alginate, chitosan, hyaluronic acid, cellulose, agarose, dextran, and carrageenan, exhibit tunable physicochemical and rheological properties, making them suitable for 3D bioprinting. The hydrophilic nature of these materials supports the high retention of water and biological fluids, which is critical for maintaining cell viability during and after printing. Among these materials, alginate is particularly widely used due to its mild gelation conditions, rapid ionic crosslinking, and favorable printability. However, its limited cell-adhesive properties may restrict cellular interactions and necessitate functionalization with bioactive components. Chitosan exhibits structural similarity to glycosaminoglycans, making it attractive for wound healing and tissue regeneration applications. Hyaluronic acid provides important biological cues for cell adhesion, migration, and proliferation, and cellulose and its derivatives offer tunable rheological properties. Carrageenan-based materials have also emerged as promising candidates due to their favorable gelation behavior, their ability to undergo chemical modification for covalent crosslinking, and their ability to form stable three-dimensional architectures. However, intrinsic limitations of polysaccharide-based bioinks include low mechanical strength, rapid degradation, a lack of cell interaction sites, reduced print fidelity, and batch-to-batch variability, which hinder broader clinical translation. To overcome these obstacles, ongoing research focuses on hybrid ink formulations, nanocomposite systems, combinations with other polymers, and advanced crosslinking strategies to enhance the performance of polysaccharide-derived bioinks. , Furthermore, understanding the relationships among polysaccharide structure, processing parameters, printability, and biological performance is essential for developing reproducible, clinically translatable polysaccharide-based bioinks for the efficient fabrication of 3D printed scaffolds in the biomedical field.

This review summarizes recent advancements in the 3D bioprinting of polysaccharides, focusing on bioinks, their physicochemical properties, and their principal biomedical applications. It also critically examines the current challenges and future perspectives for the clinical implementation of polysaccharide-based bioprinted constructs.

2. 3D Printing Techniques

In recent years, advances in 3D printing techniques have revolutionized tissue engineering and regenerative medicine by enabling the precise fabrication of complex biological structures. 3D printing manufacturing approaches enable the controlled layer-by-layer deposition of cells, biomaterials, and bioactive molecules to mimic the native architectures of tissues and organs. Several 3D printing technologies have been developed for biomedical applications, and five major techniques are discussed here.

Stereolithography (SLA) is a 3D printing technique that uses the photopolymerization principle to cure light-reactive polymer resins at room temperature. This is an additive manufacturing technique in which each layer of the printed structure is cured with a laser beam, and subsequent layers are printed on top of the previous layer to form a fully developed 3D printed scaffold as the build platform moves. This technique is used to achieve the intricate structures of complex designs developed by researchers in the fields of mechanical engineering, ceramic development, and biomedical devices.

Digital light processing (DLP) is another prominent vat photopolymerization-based 3D printing technique, distinguished from SLA by its light source. DLP employs digital UV or visible light projected onto a photosensitive resin to facilitate layer-by-layer photocuring, enabling the fabrication of high-resolution, high-speed 3D constructs. For light-based printing, polysaccharide-based biopolymers are surface-functionalized with photocurable moieties, such as methacrylic anhydride and glycidyl methacrylate, to prepare photocurable polymers, which are then used for DLP printing in the presence of photoinitiators. DLP is widely used in 3D bioprinting because of its cytocompatibility and enables higher-resolution printing that mimics the intricate structures of organs. In recent years, several polysaccharide-based biopolymers have been explored for DLP printing in tissue regeneration and biomedical devices. ,

In the laser-assisted bioprinting (LAB) technique, a laser serves as the primary source of high-precision ion contact deposition for bioprinting. Laser-induced forward transfer (LIFT) is the most commonly used method, in which a laser (source) deposits tiny droplets of cell-laden bioinks onto a substrate. This method offers high-resolution printing and preserves high cell viability by avoiding mechanical stress associated with nozzle-based techniques. Laser-based bioprinting is particularly useful in creating complex tissue models, including skin, bone, and vascular networks. It supports multimaterial and multicell-type printing, making it suitable for applications in regenerative medicine and drug testing.

Inkjet printing is another 3D printing technique used for bioprinting, in which inks, along with cells, are deposited in drops onto the substrate as needed. These systems, also called drop-on-demand (DOD), use piezoelectric actuators and are based on standard printers. Usually, alginate, chitosan, dextran, agarose, and other low-viscosity polysaccharides are used in this technique.

Extrusion bioprinting is a technique in which pressure plays a major role in printing. This technique uses pressure to continuously extrude the ink through a pneumatic nozzle. It is commonly used for medium- to high-viscosity polysaccharide-based biopolymers, such as hyaluronic acid and κ-carrageenan, as well as in composites. Various polymers have been successfully bioprinted into mechanically stable structures at high cell density without applying pressure to the cells for a range of biomedical applications.

3. Polysaccharides for 3D Bioprinting

In recent years, polysaccharides have been widely used in regenerative medicine due to their unique physicochemical properties, which enable them to serve as promising scaffold materials, drug-delivery carriers, and supports for tissue repair and regeneration, as summarized in Table . Polysaccharides closely mimic the structural and biochemical characteristics of the extracellular matrix (ECM), making them highly promising for regenerative biomaterials. ECM-mimetic polysaccharides provide structural and biochemical support for cell adhesion, signaling, and differentiation. Polysaccharide-based biopolymers have gained further attention in regenerative medicine due to the presence of abundant functional groups, such as hydroxyl, amino, and carboxyl, that facilitate chemical modification, surface functionalization, and diverse crosslinking techniques. The functionalized polymers enable strong cell interactions and cell-specific targeting, support growth factor loading and release, and interact with bioactive molecules to fabricate new, versatile bioscaffolds.

1. Detailed Overview of Major Polysaccharides Used for 3D Printing Techniques, Their Sources, Key Properties, and Biomedical Relevance .

^aThe chemical structures were acquired from PubChem and BioRender. −

3.1. Major Polysaccharides Used in 3D Printing

Alginate, chitosan, glycosaminoglycans, hyaluronic acid, cellulose, and dextran are some of the extensively used biopolymers that exhibit strong intrinsic antimicrobial activity and biodegradability; mimic ECM; support cell signaling, migration, and hydration; and provide high mechanical strength, water retention capacity, hydrophilicity, and biocompatibility. Other important polysaccharide-based biopolymers, such as carrageenan, agarose, and starch, contribute significantly to the gel-forming capability and biological properties. These polymers are widely explored and applied in bone tissue engineering, skin and cartilage regeneration, wound healing, cosmetology, and other tissue-specific applications (Table ).

2. Detailed Overview of Polysaccharide-Based Composite Bioinks for 3D Bioprinting with Their Crosslinking Strategies, Printing Techniques, Mechanical Properties, and Their Cellular Responses − .

Polysaccharide-based composite bioinks	Application	Crosslinking strategy	Printing technique	Mechanical properties	Cellular responses
Nanocellulose/chitosan-based bioinks	Bone regeneration	Physical and Schiff-base (covalent) crosslinking	Extrusion	Young’s modulus from 85.12 ± 4.31 to 132.40 ± 2.55 Pa.	Good cell viability, proliferation, and enhanced osteogenic differentiation of MC3T3-E1 cells
Carboxymethyl chitosan-based bioink	Cartilage tissue engineering	Ionic crosslinking	Extrusion	Young’s modulus from ∼0.6 to 4.0 MPa.	Promoted cell attachment and chondrogenic gene expression in chondrocytes
Alginate-gelatin-diethylaminoethyl cellulose-fibrinogen-based bioink	Skin tissue constructs	Ionic crosslinking	Extrusion	Tensile strengths of 43.83 ± 13.32 kPa,	High cell viability of human primary fibroblasts and keratinocytes in 3D bioprinted constructs
Alginate-based bioinks	Bone grafts	Ionic crosslinking	Extrusion	Storage modulus from 2.6 ± 1.1 to 9.9 ± 4.9 kPa	MSCs in alg-CaCl2 constructs exhibited cell-matrix interactions and increased osteogenic differentiation
Chondroitin and dermatan sulfate bioinks	Cartilage regeneration	Ionic crosslinking	Extrusion	Young’s modulus was approximately 0.13 kPa to, 0.20 kPa	Good cell viability and functionality of encapsulated D1-MSCs-EPO
Methacrylated-κ-carrageenan/hydroxyapatite composite bioinks	Tissue engineering applications	Photo-crosslinking	Extrusion printing	Compressive modulus in the range of 13.35–297.06 Pa	High biocompatibility and successful hASC incorporation in bioprinted bilayer tissue constructs
Nanocellulose-alginate bioink	Cartilage tissue engineering	Ionic crosslinking	Extrusion printing	Compressive stress of bioinks was approximately 24–33 kPa	High chondrocyte viability and proliferation
Xanthan gum-based bioink	Cartilage regeneration	Photo-crosslinking	Extrusion printing	Compressive modulus was reported as 27 ± 3 KPa	BMSC chondrogenic differentiation and collagen deposition, and in vivo data showed enhanced cartilage regeneration
Methacrylated guar gum	Stem cell delivery and cartilage tissue engineering	Photo-crosslinking	Extrusion printing	Young’s modulus in the range of 0.069–0.184 MPa	Exhibited favorable biocompatibility and promoted cell proliferation, cell migration, and chondrogenic differentiation of BMSCs
Pullulan-based bioink	Tissue engineering and tissue model	Photo-crosslinking	Digital light processing 3D printing	Compressive strength up to 6.2 kPa	No apoptosis of L929 cells

3.2. Hydrogel Formation Mechanism

The development of polysaccharide-based hydrogels and their transformation into desired bioinks for 3D printing are crucial for determining printability, structural stability, and biological performance. Crosslinking is crucial for stabilizing printed structures and enhancing the mechanical strength and robustness of polysaccharide-based scaffolds. In general, hydrogels are formed through physical or chemical crosslinking, resulting in a 3D network that holds large amounts of water while maintaining structural integrity (Figure ). Physical crosslinking is a noncovalent strategy that stabilizes 3D printed structures via electrostatic interactions, such as hydrogen bonding, intermolecular interactions, and ionic interactions. For example, alginate has numerous carboxyl groups that bind calcium ions, forming a stable 3D, stiffer hydrogel. In the chemical crosslinking strategy, covalent bond formation among polymers allows a mechanically stable structure. For example, click chemistry, Schiff base chemistry, and free radical polymerization are the most common methods for chemical crosslinking. Click chemistry, a type of chemical crosslinking technology in which crosslinking is achieved via azide–alkyne cycloaddition and thiol–ene reactions, is commonly used for polysaccharides such as cellulose, chitosan, alginate, hyaluronic acid, dextran, and pullulan. For example, in a study, cellulose and carboxymethyl cellulose were modified to enable an azide–alkyne reaction with Cu⁺ as a crosslinker for 3D bioprinting. In another approach using a Schiff base reaction, covalent crosslinking occurs between the aldehyde and amine groups of the polymers to form an imine bond, which is a covalent, dynamic, and reversible bond, yielding stiffer 3D printed structures and making them biocompatible. In another study, sodium alginate was oxidized, polyacrylamide-coacrylic acid with adipic acid dihydrazide, and Fe³⁺ were used in 3D printing to produce various structures. This strategy efficiently crosslinked the two polymers, resulting in rapid gelation, demonstrating good tensile properties and printability, and enabling the printing of complex 3D structures. The use of methacrylic anhydride (MA) and glycidyl methacrylate (GMA) is a newly emerging chemical crosslinking (photo-crosslinking) strategy that falls under the free-radical polymerization technique and is used to functionalize polysaccharides with reactive groups. Upon exposure to light, the synthesized photocurable polymers form stronger covalent bonds, resulting in a mechanically stable 3D printed structure. For example, several methacrylate-functionalized polysaccharides, such as hyaluronic acid and alginate, have been explored for hydrogel formation and have demonstrated good shear-thinning behavior, viscosity, mechanical stability, reduced stress with greater accuracy, and improved cell adhesion and proliferation. Enzymatic crosslinking is another strategy that uses naturally occurring enzymes as crosslinkers, such as horseradish peroxidase (HRP), transglutaminase, laccase, and tyrosinase, to form a stable hydrogel network within a 3D printed structure. For example, HRP is used to crosslink protein-based GelMA with tyramine-modified PEG to form a stable interpenetrating network. They showed that the enzyme-crosslinked 3D structures were mechanically and thermally stable, with 90% viability.

2.

Representation of the main crosslinking strategies, like physical, chemical, and enzymatic techniques, employed for fabricating 3D printed scaffolds, and their core properties, like viscosity, shear-thinning behavior, and the post-printing process for fabricating an efficient 3D printed scaffold. Created by the author through BioRender scientific illustration software.

3.3. Properties and Parameters for Bioprinting

3D bioprinting facilitates the fabrication and replication of complex native tissue structures for a diverse range of biomedical applications. During bioprinting, multiple parameters, including design, bioink formulations, physicochemical profiles, scaffold preparation, crosslinking strategy, biocompatibility, and postprinting maturation, must be carefully considered to mimic native tissue constructs and organs and to promote cell adhesion, invasion, and proliferation (Figure ). All of these factors play a significant role in bioprinting and influence the characteristics of interactions between scaffolds and cells. Bioprinted scaffolds determine the structural, mechanical, and biological properties of the mimetic tissue constructs.

Design is the backbone of successful 3D bioprinting as it is a preliminary and fundamental aspect of the process. 3D design enables precise control over complex architecture, bioink composition, and cytocompatibility, thereby creating a replica of a native tissue or organ. Multiple design parameters, such as geometry, surface topography, surface roughness, and pore size, influence the outcome and scaffold-to-cell interactions. CAD-based, image-based, implicit-surface, and space-filling-curve designs are the most common strategies used in the field. Designing for bioprinting is modeled in CAD from a patient-specific MRI or CT scan of the organ to ensure anatomical fidelity and reduce the risk of rejection.

Viscosity plays a crucial role in defining a bioink’s flow behavior during bioprinting and is the most fundamental rheological parameter to be considered during extrusion-based printing. For optimal printing applications, flow behavior, smooth extrusion, fluid flow through nozzles, and maintenance of structural integrity are essential. Low viscosity leads to shape deformation, loss of structural integrity, reduced crosslinking capability, and poor shape fidelity. High viscosity leads to hindered cell viability, cell wall lysis, cytoskeletal disruption, nozzle clogging, and shear stress, which directly influence printability and post-bioprinting characteristics. The viscosity of an ink determines the extrusion pressure to prevent post-printing collapse, ink spreading, and cell death.

Shear-thinning behavior is the most commonly examined rheological property before 3D bioprinting. This shear-thinning behavior determines the printed structures’ internal microarchitecture, mechanical stability, and flow behavior. In polymer-based bioinks, there are many interactions, and the shear-thinning property reflects the breakdown of networks in polymer solutions, interactions within particles and cells, and supramolecular interactions and noncovalent interactions such as hydrogen bonds, hydrophobic bonds, and other complexes. Shear thinning clearly depicts the behavior of the ink, allowing the bioink to remain in a stable rest phase inside the cartridge, extrude without damaging cells and components, and control the transition from liquid-like to gel-like behavior. This credits the bioink for being deposited on the substrate with structural integrity, nonspreading, and causing no damage to cells and cellular components.

Postprinting processes, such as tissue maturation, cellular interactions, structural reinforcement, and cell viability and functionality, play a critical role in determining the functionality and behavior of the bioconstructs. In this stage, if all the above parameters are addressed, the postprinting process will yield good results. Postprinting, the tissue maturation process is time-dependent and should be conducted under controlled conditions, such as within bioreactors, where cellular interactions influence cell differentiation and can be modulated by various biochemical and mechanical stimuli. Postprinting, additional crosslinkers should be washed to improve the bioconstructs’ cytocompatibility, and excessive crosslinking can lead to cellular and internal material stress. The physicochemical properties, such as swelling and degradation kinetics, must be carefully tuned. Postbioprinting, if the printed constructs have higher swelling or degradation ratios, the bioconstructs will be structurally deformed, mechanically weak, have increased inter-crosslinking distances, and will not support cells for their biochemical modulation. Maintaining high cell viability remains challenging, as cells are exposed to various stresses during fabrication and bioprinting. Still, standardization and application-specific maturation protocols are ongoing to bridge the gap between laboratory bioprinting and translatable tissue constructs.

4. Application of 3D Bioprinted Polysaccharides

In recent years, integrating polysaccharides with advanced 3D printing techniques has revolutionized the field of tissue engineering and regenerative medicine. Researchers have fabricated intricate biomimetic structures, including skin, liver, lungs, kidneys, veins, and heart valves, through 3D bioprinting. For the fabrication of biomimetic scaffolds, the selection of polymers depends on the target site, mechanical strength, degradation profile, printability, gel-forming ability, and overall biocompatibility.

4.1. Skin and Wound Healing

3D bioprinting has evolved to a greater extent in the regeneration of skin tissue. The skin is the primary source of defense and response for every animal, supporting its originality with activity. However, the skin is the first tissue to be damaged in an accident. The skin has three dermal layers: the epidermis, dermis, and hypodermis. Among the dermal layers, the dermis contains a significant amount of glycosaminoglycan (GAG) polymers embedded in the ECM. Major components of the skin ECM are dermatan sulfate, heparin, heparin sulfate, chondroitin sulfate, and keratin sulfate. Henceforth, scaffolds mimicking the ECM characteristics will pave the way for healing of skin injuries. In recent years, polysaccharide-based biomaterials have played a pivotal role in advancing skin tissue engineering because of their biocompatibility and ECM-mimicking characteristics.

In past years, several studies have reported on advanced biomedical constructs resulting from the integration of 3D bioprinting technologies with polysaccharide-based biomaterials for skin repair and regeneration. For example, Ulvan modified with a methacrylate (UlMA) functional group was used to tune the traditional bioink of gelatin methacryloyl (GelMA). The addition of UlMA in the bioink facilitated low stress while printing, and resultant 3D printed structures showed better mechanical strength with a controlled scaffold degradation rate. 3D printed cell-laden scaffolds with human fibroblast dermal cells showed good cell proliferation. Cells were viable with a sparse number of cell deaths. High metabolic activity was observed with the expression of key dermal ECM genes, mimicking the ECM of human fibroblast skin. Liu et al.‘s research team developed a bioink that mimics the full thickness of skin using an Interpenetrating Polymer Network (IPN) of alginate and methacrylated gelatin polymers (Figure a). These composites were 3D printed using phosphosilicate-calcium bio-glasses (PSCs) to validate their angiogenic properties. PSCs incorporated a 3D-bioprinted scaffold showed good cytocompatibility (HUVECs and MSCs). The results showed remarkable activity, including enhanced expression of angiogenic genes, increased vascularization, and rapid wound healing, as demonstrated through in vitro and in vivo analyses in rat models. Lihao et al.’s research team focused on healing diabetic wound ulcers, which are very challenging to treat due to patients’ physiological conditions. Strategically, they used a 3D-bioprinted alginate-gelatin scaffold to mimic human skin and incorporated salvianolic acid B (SAB), which can induce proangiogenic properties and promote healing of diabetic wounds. Both in vitro and in vivo studies showed that 1 wt % of SAB with alginate and gelatin decreased TNF-α, IL-6, and IL-1β. However, it showed increased expression of TGF-β and VEGF, increased collagen deposition, increased expression of cell adhesion molecules, cytocompatibility (HUVECs and RS1s), and a healed diabetic wound within 14 days with 1% SAB compared to other concentrations. The study demonstrated the vital role of SAB in accelerating the healing of diabetic wounds. Furthermore, the composite must mimic natural skin to aid healing. To mimic natural skin, Tavakoli et al.‘s research team used 3D bioprinting to fabricate a bilayer scaffold with a top layer of pectin and polyacrylic acid (PAA) with platelet-rich fibrin and a bottom nanofibrous layer containing pectin, polyacrylic acid, and simvastatin (SIM). Fabricated pectin-based scaffolds were highly biodegradable and biocompatible, and in vitro studies showed good cell viability. In the in vivo rat model, wound closure was observed after 14 days, coinciding with growth-factor release. SIM-aided quick wound healing with higher collagen deposition, angiogenesis, neovascularization, and re-epithelization was observed.

3.

(a) Fabrication of a bioprinted full-thickness skin substitute using a dual-ink approach. The epidermal ink (alginate + gelatin) and the dermal ink incorporating PSC, HUVEC, and MSC cells are co-printed to generate a layered skin construct. The skin substitute is evaluated both in vitro for skin tissue regeneration, reproduced with permission from reference, Copyright 2024 Elsevier. (b) Clinical application of 3D-bioprinted Autologous Minimally Manipulated Homologous Adipose Tissue (AMHAT) for post-excisional skin defect repair. The Dr. INVIVO bioprinter (ROKIT Healthcare, Seoul, Republic of Korea) was used to fabricate customized AMHAT patches from patient-derived adipose tissue, reproduced with permission from reference, open access, MDPI. (c) Overview of the biofabrication workflow. GelMA, AlgMA, and reduced graphene oxide were combined with cardiac cells to formulate bioink. The bioink was bioprinted into ring-shaped cardiac tissue constructs by photo-crosslinking in the visible range (405 nm). Bio rings were adapted for high-throughput multiwell plate formats and heart-on-a-chip platforms, demonstrating rhythmic beating activity for cardiac tissue regeneration, reproduced with permission from reference, Copyright 2024 Elsevier. (d) One-pot synthesis approach with extrusion-based 3D printing, applied in two biological applications like stem cell differentiation and bone defect repair using HAP hybrid scaffolds and validated in a murine calvaria defect model, reproduced with permission from reference, Copyright 2024 Elsevier.

4.2. Adipose-Based Regenerative Therapies

Adipose tissue, known as body fat, functions as connective tissue and plays essential roles in energy storage, insulation, and metabolic regulation. Adipose tissue is distributed throughout the body and is found at various sites, such as beneath the skin, between internal organs, and within the interior cavities of bones. Notable progress in 3D bioprinting has enabled the fabrication of adipose tissue constructs with precise architectural control and enhanced biological functionality for regenerative applications. Reconstruction of the breast is highly recommended for breast cancer patients to improve their mental state in society. In some cases, doctors may advise mastectomy for the breast. Using additive manufacturing, scaffolds of methacrylamide-modified gelatin (GEL-MA) and methacrylate k-carrageenan (Car-MA) were 3D printed with an extrusion-based printer to mimic the native ECM during adipose tissue regeneration in breast cancer. These 3D printed scaffolds of a hydrogel blend showed good cell viability with seeded adipose tissue-derived stem cells (ASCs) and could differentiate into the adipogenic lineage. Additionally, Säljö et al.‘s research team demonstrated 3D bioprinting of ASCs in an alginate/nanocellulose bioink and then implanted the graft into nude mice. The 3D-bioprinted graft showed enhanced mechanical properties, increased adipocyte density, improved angiogenesis and vascularization, in vivo survival, and a promising graft for clinical application. In another approach, 3D bioprinting of autologous minimally manipulated homologous adipose tissue (3D-AMHAT) with protein-based fibrin gel was used to treat diabetic foot ulcers, as adipose tissue contains numerous ECM components that support tissue regeneration. This composite material efficiently reduced the wound size, and skin regenerated within 12 weeks (Figure b).

4.3. Cardiac Tissue Engineering

The heart is the central organ of the cardiovascular system. Worldwide, cardiovascular disease (CVD) has been the leading cause of mortality and morbidity globally. Heart-related diseases like coronary heart disease, cerebrovascular disease, and rheumatic heart disease are the world’s leading health risks, vitally increasing mortality and, especially, aortic valve deformation. As heart failure is a significant global problem, researchers are working to regenerate cardiomyocytes to achieve heart regeneration. The regeneration of heart cells has been well documented in amphibians and fish. However, regeneration of heart cells in the adult human heart is exceedingly rare. Hence, myocardial regeneration is a focus of regenerative medicine research, which could save millions of lives. In cardiac tissue engineering, Duan et al.‘s research team demonstrated the use of 3D bioprinting to fabricate live trileaflet aortic valve conduits using alginate and gelatin hydrogels, incorporating dual cell types to maintain the anatomical structure. The two types of cells encapsulated were SMCs (smooth muscle cells) and VICs (valve leaflet interstitial cells). Hydrogels containing encapsulated SMCs and VICs showed viability of more than 90% and 80% on days 1 and 7, respectively. According to the study, overall projected cell spreading was higher in VICs than in SMCs. Further, after 7 days of cell culture, encapsulated VICs and SMCs showed protein expression of α-SMA and vimentin. In another study by Mousavi et al.‘s research team, a photo-crosslinked ring-shaped cardiac tissue was bioprinted from a gelatin methacryloyl (GelMA) and alginate methacrylate (AlgMA) composition, followed by subsequent encapsulation with electroconductive graphene oxide nanomaterials for cardiac tissue regeneration. The modified bioink showed enhanced biocompatibility, rheological properties, mechanical properties, and a controlled degradation rate. Cardiomyocytes, cardiac fibroblasts, and HL-1 cells were used to prepare the bioink for printing the cardiac BioRings. The prepared bioink demonstrated good cardiac cell viability, proliferation, spreading, elongation, and significant spontaneous cell beating. These developed models can be applied to drug screening and disease modeling (Figure c).

4.4. Bone Regeneration

Bones are important locomotor support structures in the human body that are affected by infection, tumors, and accidents. For many years, bone grafting methods have been used to address bone-related issues. Autografts and allografts have disadvantages in terms of donor-to-patient ratios, and immune rejection and infection are major research problems in osteology-based clinical therapy. To overcome this problem, researchers have begun investigating bone regeneration using 3D bioprinting and 3D printed polysaccharide-based scaffolds. Several research groups have printed polysaccharides such as alginate, guar gum, chitosan, hyaluronic acid, xanthan gum, cellulose, κ-carrageenan, and agarose using various types of 3D printers and have filed patents. ,,, Chrungoo et al.‘s research team developed a bioink composed of BSA (Bovine Serum Albumin) and alginate, with a self-assembling polyelectrolyte complex of gelatin and chitosan (PEC-GC), which was used to mimic bone. The incorporation of PEC-GC improved printability, shape fidelity, and mechanical stability. The in vitro study using 3D-bioprinted scaffolds demonstrated strong adhesion and biomaterial interactions with osteosarcoma cells, with >90% viability, high collagen synthesis, and enhanced osteogenic properties. Further, You et al.‘s research team used a novel bioink composed of liver decellularized ECM, gelatin, alginate, and rat bone mesenchymal stem cells (rBMSCs) to fabricate a cell-laden composite scaffold. The composites showed good biocompatibility, osteogenic properties, and vascularization and accelerated in vivo bone regeneration. Ren et al.‘s research team used a one-pot synthesis method to prepare a bioink composed of methacrylated gelatin, alginate, hydroxyapatite (HAP), and MSCs. The bioink displayed excellent printability, stability, enhanced mechanical strength, in vitro compatibility, and enhanced osteogenic differentiation of mesenchymal stem cells. In vivo findings showed remarkable bone repair in rat calvarial defects (Figure d). Poldervaart et al.‘s research team reported the 3D bioprinting of methacrylated hyaluronic acid (MeHA) with human bone marrow-derived MSCs to support bone-like tissue formation. MeHA-based hydrogels supported good cell survival and osteogenic differentiation of MSCs, and this effect was further enhanced by the addition of bone morphogenetic protein-2 (BMP-2). In another approach, hyaluronic acid (HA), hydroxyethyl acrylate (HEA), and a gelatin-methacryloyl-based gel were evaluated as bioprinting gels. Here, HEA to HA was graft polymerized, followed by radical polymerization of gelatin-methacryloyl. The gel (HA-g-pHEA gelatin) was bioprinted in a lattice form with MC3T3 bone cells that showed excellent cell viability.

4.5. Cartilage Repair

Cartilage is a connective tissue that protects joints and bones and provides structural stability to the body. It maintains balance between the bones, reduces friction, acts as a cushion, helps absorb shocks, and helps maintain flexibility. The human body contains different types of cartilage, including hyaline cartilage, found mainly in the ribs, nose, larynx, and trachea; fibrocartilage, present in ligaments and intervertebral discs; and elastic cartilage, located in the ear, epiglottis, and larynx. Damage to articular cartilage is a significant hallmark of osteoarthritis, a global healthcare concern affecting nearly 250 million people worldwide. Current treatment strategies, such as grafts and stem cell-based therapies, face limitations including immune rejection, donor availability, and tissue-level complications. 3D bioprinted polysaccharide-based scaffolds offer a promising alternative, enabling the fabrication of patient-specific scaffolds with controlled architectures to support effective cartilage tissue regeneration. Antich et al.‘s research team developed a new bioink for 3D bioprinting of cartilage tissue by combining hyaluronic acid (HA) and alginate and co-printing with polylactic acid (PLA) (Figure a). In vitro studies performed with an HA-based biomimetic hybrid scaffold demonstrated cartilage ECM deposition and chondrogenic gene expression, potentially supporting articular cartilage formation. Markstedt et al.‘s research team reported bioprinting human nasoseptal chondrocytes (hNCs) in combination with nanocellulose fibrils (NFC) and alginate (Alg). The 3D bioink constructs showed a significant increase in cell viability after 7 days of 3D cell culture. Lafuente-Merchan et al. developed a 3D-printable bioink of nanocellulose alginate (NC-Alg) in combination with chondroitin sulfate (CS) and dermatan sulfate (DS). These 3D bioprinted scaffolds cultured with MSCs showed potential for cartilage application, as evidenced by gene expression. Zhang et al.‘s research team reported 3D bioprinted scaffolds fabricated from a photopolymerizable bioink comprising recombinant collagen, chitosan, Laponite-XLG nanoclay, and nanoparticles loaded with kartogenin (KGN) for cartilage repair. The 3D printed scaffolds exhibited excellent antimicrobial properties, good cytocompatibility with human bone marrow mesenchymal stem cells (hBMSCs), and enhanced expression of collagen II and glycosaminoglycan. Additionally, cartilage tissue regeneration was demonstrated in a rabbit cartilage defect model. Guan et al. developed another novel photo-crosslinked, printable cartilage ink comprising polyethylene glycol diacrylate (PEGDA), gelatin methacryloyl (GelMA), and chondroitin sulfate methacrylate (CSMA). 3D bioprinted scaffolds showed remarkably enhanced chondrogenic differentiation of BMSCs. In another approach, a polysaccharide-based 3D printing ink was used to prepare a biphasic scaffold containing decellularized natural cartilage (dNC) and nano-hydroxyapatite for hyaline cartilage formation and osteochondral regeneration.

4.

(a) Workflow of bioink preparation and bioprinting with an HA-based ink: HA-ink with a chondrocyte-derived extracellular matrix to fabricate an HA-bioink and 3D bioprinting using the REGENAT bioprinter to fabricate grid-patterned constructs for cartilage regeneration, reproduced with permission from reference, Copyright 2020 Elsevier. (b) Printability assessment of two GelMA/MC bioink formulations: pressure-speed parameter maps for (i) GelMA5/MC8 and (ii) GelMA7/MC8, where green zones indicate under-extrusion, orange zones represent optimal printing conditions, and blue zones correspond to over-extrusion, corresponding photographic images, and (iii) 3D-bioprinted constructs using GelMA5/MC8 and GelMA7/MC8 bioinks demonstrating high-resolution text fidelity, mimicking the native corneal geometry, reproduced with permission from reference, Copyright 2024 Elsevier. (c) 3D printed kidney organ model used for training, which is a multi-material composite organ phantom that mimics realistic tissue appearance and texture, reproduced with permission from reference, open access, Wiley. (d) Graphical representation of the in situ bioprinting technique, reproduced with permission from reference, open access, Springer Nature.

4.6. Eye

The eye is the primary sense organ in the human body and plays a vital role in acquiring visual information through light. Human eye anatomy is complex, with several layers, including the iris, cornea, pupil, sclera, and conjunctiva. 3D bioprinting is paving the way to address eye-related research problems due to its significant potential and could be a technology capable of solving donor-related issues. Human corneal tissue is widely used to treat corneal blindness, but the incidence of blindness remains high, and the availability of corneas for transplantation is limited. Corneas can be printed perfectly with this technology, as they have mechanical stability, the ability to print intricate structures, transparency during printing, and layer-by-layer cell deposition. Puistola et al.‘s research team used a 3D bioprinting strategy to mimic the native microstructure of corneal stroma construction, using a bioink prepared from human adipose tissue-derived stem cells (hASCs) and hyaluronic acid (HA). The findings from the bioprinted composite structures showed improved mechanical stability and cytocompatibility, and an ex vivo porcine corneal model supported corneal tissue engineering. In another study, 3D printed GelMA and methylcellulose hydrogels were used to mimic the corneal stroma (Figure b). The 3D printed hydrogels exhibited good transparency and improved rheological and mechanical properties. Further in vitro studies using goat corneal stromal cells showed good cell adhesion and proliferation, with positive expression of corneal markers, including keratan sulfate and vimentin.

4.7. Application in Developing Organ Models for Drug Discovery

Another important area of research in 3D printing is the printing of organs for use in the surgical, medical, and biological fields as organ or tissue models. In vitro organ and tissue models support drug testing, drug delivery, surgery, molecular-level testing, the development of new therapeutics, the study of tissue morphology, and so on. Mimicking an organ by 3D printing may enhance a precise understanding of a particular organ or tissue. Therefore, 3D printing plays a leading role in developing organs, ranging from millimeters to centimeters in size, by using naturally available or synthetic polymers. Various research groups have fabricated tissues and organs such as skin, vascular structures, the heart, aortic valves, cornea, ear, cardiac tissue, adipose tissue, prostate organ, brain microvasculature, craniofacial, maxillofacial, liver, bone, and cartilage, through standard 3D bioprinting techniques. , The real, physical organ, devoid of biological activity, can also be printed using 3D printing, mimicking mechanical properties and replicating the tissue environment (Figure c). The bioactive tissue model enhances cell responses, in situ detection, drug screening, organ transplantation, and tissue regeneration. In contrast, the physical organ model enhances surgical navigation, preoperative learning, and education and training kits for patients and their students.

Using a 3D fabrication method, an aortic valve 3D model was fabricated from alginate and PEGDA. In this study, 3D printed valve scaffolds were seeded with porcine aortic VICs (Valve leaflet Interstitial Cells), and the results showed good cell adhesion, migration, and high cell viability. In another approach, 3D printing of a bionic ear was demonstrated using an alginate hydrogel matrix containing viable ear chondrocytes, electrically conductive silver nanoparticles, and nonconductive silicone solutions. The 3D printed hybrid ear cultured in chondrocyte media demonstrated good cell viability, structural integrity, and robust auditory sensing for radiofrequency reception. The research team of Zhang et al. fabricated vascular conduits via direct bioprinting of alginate-encapsulated human umbilical vein smooth muscle cells (HUVSMCs). In vitro results depicted good cell viability, proliferation, and deposition of smooth muscle matrix and collagen around the periphery and luminal surface. Lin et al.‘s research team developed a 3D bioprinted osteosarcoma model (3DBPO) using the same cells, with gelatin methacrylamide and hyaluronic acid methacrylate, mimicking the tumor ECM. Results from the ECM-mimicking 3D model showed significant changes, with weaker cell proliferation, metabolism, and adherens junctions, and stronger catabolism, compared to spheroids and 2D models. Further, drug sensitivity and strong autophagy were well established in osteosarcoma 3D models and could be used in osteosarcoma research, serving as a novel strategy for cancer research. Recently, using a gelatin/sodium alginate-based bioink for 3D bioprinting, Sun et al.‘s research team developed a patient-derived preclinical model system for pancreatic cancer and tested chemotherapy drug responses, with correlation to clinical outcomes.

4.8. Application in In Situ Bioprinting

In situ bioprinting is widely used today, in which the prepared therapeutic ink is deposited directly onto the patient or experimental animal to repair defects based on their shape and size. However, because of many challenges in handling, cost, and availability, in situ bioprinting is comparatively less explored than other 3D printing methods. However, the main advantages of in situ bioprinting include ease of handling, reduced contamination, broad applicability to wounds rather than local application, precise cell distribution for healing, and the ability to sterilize. There are two approaches to in situ bioprinting: the robotic-arm approach and the handheld approach. The robotic-arm approach uses CAD and CAM design with minimal human involvement. The handheld approach model includes the handling, cartridge, nozzle, and extrusion system components, as in a 3D printing machine, with additional advantages: reduced time, the ability to stop at any point, portability, low cost, and ease of use.

Albanna et al.‘s research team used in situ 3D bioprinting (inkjet-based bioprinter technology) to deliver autologous or allogeneic dermal fibroblasts and epidermal keratinocytes encapsulated within a protein-based biological hydrogel system composed of fibrinogen and collagen directly into the injured area in a porcine model (Figure d). Over a 6-week study, the bioprinted wounds showed significantly faster healing, with effective wound coverage, increased epithelialization, and reduced scarring compared with the untreated and matrix-control groups. In another approach, a sprayable and injectable hydrogel based on visible-light-crosslinked methacrylated kappa-carrageenan (κCaMA) was developed in situ for soft tissue engineering. In vitro analysis of crosslinked KaMA demonstrated noncytotoxicity, good cell adhesion, and enhanced cell proliferation. Overall, the study supports the feasibility of developing sutureless in situ wound dressings.

5. Challenges and Translational Considerations

Despite notable scientific breakthroughs in polysaccharide-based 3D printing approaches for tissue repair and regeneration, several challenges in current research models need to be examined and addressed before advancing to further clinical applications and trials. This section discusses key challenges, including the mechanical stability of hydrogels, maintenance of aseptic conditions for bioink formulations, the high cost of 3D bioprinting, and, importantly, obtaining regulatory approval.

5.1. Obstacles in Achieving Desired Mechanical and Structural Properties

One of the major challenges with biopolymer-based biomaterials is recapitulating the mechanical and structural properties of native human tissue. Still, polysaccharide-based biomaterials inherently exhibit poor mechanical strength and structural stability. Most hydrogels fabricated from biobased polymers exhibit favorable cell viability but poor mechanical properties, a major hurdle for fabricating scaffolds for load-bearing bone and cartilage tissue regeneration. Limited spatial resolution has been another bottleneck for conventional 3D bioprinting approaches, particularly in achieving intricate tissue structures, complex organ models, and cellular-scale features with reasonable printing volume. Bioinks used for bioprinting should meet necessary physicochemical requirements, such as rheological properties, to ensure proper printability, maintain shape fidelity, and preserve structural integrity during printing and post-bioprinting. Further, preventing scaffolds from collapsing, shrinking, and deforming is even more challenging. Extrusion-based bioprinting, a widely used AM technique, is particularly susceptible to these challenges. This technique has challenges in maintaining cell viability as shear stress can affect cells loaded into the bioink, and 3D-bioprinted constructs show only 70–80% viability. Improving vascularization remains a structural limitation, despite its major role in biochemical transport within engineered organs and tissues. According to the Krogh cylinder model of diffusion, without an adequate vascular network, fabricated tissue or organ constructs thicker than approximately 100–200 μm may not receive sufficient oxygen and nutrients, thereby compromising cell viability and proliferation. Consequently, cells located beyond this diffusion range may experience hypoxia and reduced cell viability, potentially leading to necrotic regions within densely printed constructs. Therefore, when fabricating larger, densely cellularized constructs, this diffusion constraint is an important parameter for bioink selection and formulation. The formulation of bioinks should be carefully optimized to balance printability, mechanical stability, crosslinking density, porosity, mass transport, and biological functionality. Toward this, chemical modifications, polymer functionalization, crosslinking efficiency optimization, and the fabrication of constructs with composite and hybrid polymers are being explored to tailor the physicochemical and transport properties of bioinks. In addition, advanced printing techniques such as DLP, Sacrificial Writing Into Functional Tissues (SWIFT), and Freeform Reversible Embedding of Suspended Hydrogels (FRESH) are being explored to enhance the resolution of printing intricate structures, fabricate complex and vascular-like architectures, improve mechanical behavior and structural integrity, and ensure biochemical transport without compromising the biological functionality of the 3D printed constructs.

5.2. Reproducibility and Scalability

A significant translational barrier in 3D bioprinting is achieving reproducibility, consistency, and scalability across different 3D bioprinting platforms to ensure that the fabricated scaffolds are suitable for clinical samples for biomedical applications. Moreover, variability in polysaccharide sources, extraction methods, and batch-to-batch variation significantly affects the material properties and performance during 3D bioprinting, ink optimization, and ink preparation. Many attempts have been made to standardize bioprinting process parameters, but they have been limited to a single bioink, a single tissue, a single aspect, and a single application. Parameters such as nozzle dimensions, culture media, temperature, speed, pressure, and humidity collectively determine the properties of the postbioprinted tissues, which are very challenging to reproduce. Such an absence in the harmonized protocols critically challenges the reproducibility of the printing process and bioprinted constructs. Additionally, scalability from small-scale lab printing to large-scale printing and manufacturing is a major challenge in 3D bioprinting. Other persistent challenges include maintaining sterility, resolution, consistency, precision, and quality and quantity without compromising the availability and biocompatibility of raw materials. Furthermore, many 3D bioprinting processes are time-consuming, which limits scalability and can compromise cell viability.

In addition to technical challenges, the lack of well-established regulatory benchmarks, standardized assessment parameters, and evaluation criteria constitutes a substantial obstacle to the clinical translation of 3D bioprinting. In the regulatory evaluation of additively manufactured medical products, reproducibility and consistency are important considerations; however, complex interactions among bioink composition, printing parameters, processing conditions, and cellular components limit reproducible outcomes. The U.S. Food and Drug Administration (FDA)’s principal guidance, Technical Considerations for Additive Manufactured Medical Devices, provides recommendations for additively manufactured medical devices but explicitly excludes biological, cellular, and tissue-based products. Bioprinting-specific inquiries are referred to the Center for Biologics Evaluation and Research (CBER) for case-by-case evaluation rather than providing a standardized regulatory protocol. This lack of a unified regulatory pathway complicates the approval process for 3D-bioprinted tissue-engineered products. In 2024, ASTM International introduced F3659-24, the Standard Guide for Bioinks Used in Bioprinting, which provides guidance on bioink formulation, material properties, crosslinking, printability, sterility, cytocompatibility, and post-printing considerations. However, standardized benchmarks for critical bioprinting parameters, including nozzle diameter, printing speed, pressure, temperature, crosslinking conditions, and other process parameters, remain limited, making comparisons between studies and scale-up across different laboratories challenging. Additionally, variations in bioink formulations and printing parameters can significantly affect the mechanical properties, structural stability, and biological performance of printed scaffolds. These challenges are particularly relevant to polysaccharide-based bioinks, which may exhibit batch-to-batch variations in raw material purity, molecular weight, degree of substitution, and physicochemical properties, potentially affecting printability, crosslinking behavior, mechanical performance, and biological outcomes. The development and adoption of standardized material characterization, manufacturing protocols, process controls, and performance benchmarks are therefore essential to enhance reproducibility and facilitate regulatory approval and clinical translation of polysaccharide-based and other biomaterial-based bioprinting strategies.

5.3. Regulatory and Clinical Translation Challenges

Regulatory pathways for 3D bioprinted constructs toward clinical translation are complex and remain challenging, with stringent regulatory requirements to ensure safety, efficacy, and quality. Regulatory agencies such as the FDA and the European Medicines Agency (EMA) require proper evaluation and thorough characterization of polysaccharide-based bioinks used for 3D bioprinting, and they should meet biocompatibility regulations, safety standards, sterility requirements, mechanical properties, manufacturing quality, and reproducibility for biomedical applications. 3D printed constructs should have a slow degradation rate, be compatible with tissue constructs, and not elicit any adverse immunological responses. Bioinks contain living cells, bioactive compounds, and decellularized materials, raising ethical considerations that further complicate obtaining regulatory clearance and require clinical and preclinical validation. This slows the clinical translation of polysaccharide-based 3D bioprinted scaffolds for biomedical applications. 3D bioprinted constructs with cells, biomaterials, and growth factors fall under various categories, including medical devices, advanced therapy medical products (ATMPs), and tissue-engineered products, each of which is subject to distinct regulatory frameworks. In addition, storage, sterilization, and stability hinder the ethical clearance for biomedical applications. Currently, bioprinted designs have not reached the required level of clinical complexity, and there are no clear, precise good manufacturing practice (GMP) rules or regulatory guidelines. This is a major gap to be addressed and a key factor in translating the newly developed technique from the bench to the bedside. On top of this, only 11 clinical bioprinting trials have been registered globally between 2016 and 2023, which shows a very slow pace of translation and regulatory pathways. These interlocking hindrances and barriers can be overcome through collaboration among researchers and physicians, harmonization of standards, and regulatory agencies to establish clear guidelines for translating the technique into bioprinted, patient-specific therapies. ,

6. Future Perspectives

Polysaccharide-based 3D bioprinting is a newly emerging technique that represents a transformative juncture in materials science, biofabrication, and tissue engineering. Future developments focus on enhancing the functionality, adaptability, and clinical translation of bioinks and 3D bioprinted constructs. Emerging concepts, such as 4D printing, artificial intelligence-assisted bioink design, biomimetic microenvironment engineering, and commercial and clinical translation, can be used to deliver patient-specific therapies in biomedical applications and device development (Figure ).

5.

Schematic illustration focuses on future perspectives in polysaccharide-based 3D bioprinting, encompassing four key domains: 4D bioprinting, where external responses to scaffolds undergo time-dependent shape transformation triggered by factors like pH, temperature, moisture, and enzymatic cues; artificial intelligence and machine learning, integrating compositional, rheological, biological, and printability parameters as input data to enable faster development, high precision, and time-efficient bioink optimization; biomimetic microenvironment, utilizing ECM-mimicking polysaccharides with biochemical cues, mechanical gradients, and vascularization to promote cell adhesion, proliferation, differentiation, and tissue regeneration; and clinical prospects, outlining the translational flow from laboratory research to preclinical evaluation and translating the 3D bioprinting technique from bench to bedside application. Created by the author through BioRender scientific illustration software.

6.1. 4D Bioprinting

4D printing extends 3D printed structures, enabling them to undergo dynamic structural changes in response to external stimuli, such as pH, temperature, moisture, or chemicals. Polysaccharides are inherently stimulus-responsive, and their ease of functionalization further supports a 4D printing approach for tissue engineering applications. 4D printing enables the fabrication of diverse scaffolds with intelligent biomaterials, shape memory, self-healing, time-dependent drug release, and next-generation scaffolds for biomedical innovations. Polysaccharides such as alginate, carrageenan, chitosan, hyaluronic acid, agarose, starch, and dextran are potential candidates for next-generation 4D bioprinting due to their dynamic remodeling behavior and their ability to mimic native tissue and organ structures. Polysaccharides, in combination with 4D bioprinting, will enable the development of patient-specific intricate constructs and adaptive, dynamic structures for hard and soft tissue engineering using stimuli-responsive techniques, especially shape-memory-based products, which are widely used to address mechanical challenges. The incorporation of bioactive materials and nanoparticles within the 4D-bioprinted structure will further be a promising approach toward developing organ- and patient-specific scaffolds for transformative applications in cardiac tissue and valves, cartilage regeneration, bone grafts, neural tissues, wound healing, smart implants and prosthetics, biosensors, bioactuators, biorobotics, and dentistry.

6.2. Artificial Intelligence in Bioink Design

Nowadays, AI (artificial intelligence) and ML (machine learning) have taken a new paradigm in many fields, including biomedical. In this way, AI can identify many promising accelerators for the development of polysaccharide-based bioinks. Bioink formulation requires many critical optimization parameters, such as viscosity, gelation kinetics, crosslinking efficiency, crosslinking strategy, cell loading efficiency, printability, and postprinting parameters, which are very challenging criteria to follow, reproduce, and scale. AI and ML can be applied at various stages of 3D bioprinting processes to analyze biomaterial formulations. In addition, ML is playing an important role in the evaluation and printability optimization process by analyzing the influence of inks during printing, printing parameters, predicting postprinting models, and employing strategies to enhance biocompatibility. Furthermore, polysaccharides exhibit batch-to-batch variability in bio-based polymers, such as carrageenan, chitosan, hyaluronic acid, and collagen. To address this real-time challenge, AI-driven platforms can analyze large datasets to compensate for material inconsistencies, rheological properties, and printability, thereby controlling quality. Therefore, AI will be a futuristic technique for developing rational designs for tissue- and organ-specific scaffold fabrication. AI models like random forest (RF), support vector machines (SVMs), convolutional neural networks (CNNs), artificial neural networks (ANNs), generative adversarial networks (GANs), digital twin frameworks, and principal component analysis (PCA) are some of the AI methods for hydrogel additive manufacturing to improve the prototyping of scaffolds summarized by Zhang et al. In recent years, the application of AI in 3D bioprinting of polysaccharides has been a valuable technique for developing designs for tissue- and organ-specific scaffold fabrication. Quavi et al. employed data-driven methodologies, including design of experiments (DoE), multiresponse optimization, and machine learning, to analyze bioinks composed of sodium alginate, gelatin, and a nanoclay reinforcing agent (laponite). These approaches established relationships among the composition, rheological properties, and printability. Artificial neural network (ANN) models accurately predicted bioink printability based on rheological parameters, achieving a maximum mean absolute error (MAE) of 6.3%. The optimized bioinks exhibited high printing fidelity, with less than 5% deformation at extrusion pressures below 30 kPa while maintaining good cell viability. These findings highlight the potential of ANN-assisted optimization for designing polysaccharide-based bioinks for tissue engineering. In another application of a novel machine-learning model, a sodium alginate/cellulose nanocrystal composite bioink was investigated to improve the accuracy of predictions for printing resolution and z-axis position error in suspended bioprinting. The results demonstrated the robust predictive performance of rheology-informed hierarchical machine learning (RIHML) compared with other supervised machine learning approaches, such as support vector regression (SVR), RF, and concentration-dependent machine learning (CDML). Furthermore, the findings address key challenges related to suspended bioprinting and offer a scalable solution to improve printability, reduce cost and time, and improve the precision and reproducibility of tissue-engineered scaffolds, with limited experimental data.

6.3. Biomimetic Microenvironments

The preliminary objective of 3D bioprinting is to mimic the ECM, intricate architecture, microenvironment, and vascularization and to smoothly fabricate tissue- or organ-mimicking scaffolds with cytocompatible biomaterials. Creating the proper microenvironment with biopolymers enhances the properties and functionality of the 3D printed scaffolds. Polysaccharide-based materials have the potential to mimic native tissue architecture. Advancements such as functionalization, the use of hybrid polymers, and the incorporation of growth factors, signaling molecules, drugs, RGD-active components, and binding domains in bioinks help mimic the microenvironment of the specific tissue or organ and promote cell fate, vascularization, and remodeling at the damaged area of the specific tissue or organ. Proper crosslinking mechanisms, self-healing constructs, responsive materials, and smart materials are nascent techniques for fabricating a biomimetic microspatial environment for clinical translational biomaterials in the biomedical field. Polysaccharide-based biomaterial bioinks with tunable properties, decellularized ECM, nanoparticle loading, and compatibility with biological molecules are suitable materials for providing a sufficient microenvironment for tissue or organ constructs using 3D bioprinting in postprinting processes such as cell adhesion, tissue maturation, and organ development.

6.4. Commercial and Clinical Prospects

The 3D printing market has seen substantial growth across the healthcare, biomedical, aerospace, and industrial sectors and is projected to expand further in the coming decade. This continuous market growth is driven by ongoing technological progress and government initiatives that are favorable to 3D printing. The landscape for commercializing 3D bioprinted polysaccharide-based constructs has rapidly expanded over the last few decades in the field of biomedical product development. The use of polysaccharide-based bioprinting is particularly blossoming in regenerative medicine due to the natural availability, nonimmunogenicity, tunable properties, biocompatibility, thermoreversibility, and gelation timing. These properties help tailor patient-specific 3D bioprinted constructs and facilitate their commercialization, thereby increasing the efficacy of biomedical innovations. Although polysaccharide-based 3D bioprinted scaffolds are promising materials in regenerative medicine, they still face challenges in commercialization, scalability, standardization, and sterilization and require numerous approvals and regulatory approvals. Further, only a few companies are active in 3D bioprinting, and only a few have made significant commercial progress, indicating the slow pace of commercializing 3D-bioprinted constructs.

As a notable development, in April 2025, the FDA announced a roadmap to reduce or replace animal testing with new approach methodologies (NAMs). This may contribute to the development and application of modern laboratory techniques, such as organ-on-a-chip platforms, bioprinting, cell-based assays, and computer-based pharmacological models, which are designed to mimic human tissues and organs and reduce reliance on animal studies. Furthermore, companies such as CELLINK, Aspect Biosystems, Organovo, and 3D Systems have been continually advancing to overcome major challenges in addressing critical medical issues, including organ shortages and personalized therapies. With continued advancements and strategic collaborations with researchers, physicians, industry, and the healthcare sector, the polysaccharides and 3D bioprinting approach hold strong potential for personalized therapies and next-generation therapeutic solutions and can be translated from bench to bedside for treatment and biomedical innovation.

7. Conclusion

The integration of 3D bioprinting technologies with polysaccharide-based biomaterials has established a promising platform for regenerative medicine. 3D bioprinting has emerged as a transformative technology, enabling the customization and fabrication of patient-specific scaffolds and functional biomedical constructs. Polysaccharide-based biomaterials are widely utilized in bioink preparation due to their availability, structural similarity to the extracellular matrix, biocompatibility, noncytotoxicity, and versatility.

This review has examined 3D bioprinting technologies applied to polysaccharide-based biomaterials for regenerative applications, including skin, adipose, cardiac, bone, cartilage, eye, and organ models, as well as in situ tissue engineering. Key challenges such as translational and clinical barriers, and strategies for bench-to-bedside translation have been critically analyzed. The development of smart materials through 4D printing, as well as advancements in artificial intelligence and biomimetic environments, has also been discussed. Overall, 3D printing combined with polysaccharide-based biomaterials offers a versatile and transformative platform with significant potential for advanced personalized healthcare applications.

Glossary

Abbreviations

3D

Three Dimensional

ME

Material Extrusion

BJ

Binder Jetting

PBF

Powder Bed Fusion

VP

Vat Polymerization

DED

Direct Energy Deposition

MJ

Material Jetting

SL

Sheet Lamination

ECM

Extracellular Matrix

SLA

Stereolithography

DLP

Digital Light Processing

LAB

Laser-Assisted Bioprinting

LIFT

Laser-Induced Forward Transfer

DOD

Drop-on-Demand;

MA

Methacrylic Anhydride;

GMA

Glycidyl Methacrylate;

HRP

Horseradish Peroxidase

GelMA

Gelatin Methacrylol

CAD

Computed Aided Designing

MRI

Magnetic Resonance Imaging

CT

Computerized Tomography

GAG

Glycosaminoglycan

UlMA

Ulvan Methacrylate

IPN

Interpenetrating Polymer Network

PSC

Phosphosilicate-Calcium Bio Glasses

HUVEC

Human Umblical Vein Endothelial Cell

MSC

Mesenchymal stem cell

SAB

Salvianolic acid B

PAA

Polyacrylic acid

SIM

Simvastatin

Car-MA

Kappa Carrageenan Methacrylated

ASCs

Adipose Tissue-Derived Stem Cells

3D-AMHAT

3D Bioprinting-Autologous Minimally Manipulated Homologous Adipose Tissue

CVD

Cardiovascular Disease

SMC

Smooth Muscle Cells

VIC

Valve Leaflet Interstitial Cells

AlgMA

Alginate Methacrylate

Sr-CSH

Strontium-Substituted Xonolite

BMSCs

Bone Marrow Mesenchymal Stem Cells

PEC-GC

Polyelectrolyte Complex of Gelatin and Chitosan

ACMP

Amorphous Calcium Magnesium Phosphate

rBMSC

Rat Bone Mesenchymal Stem Cell

DPSCs

Dental Pulp Stem Cells

HAP

Hydroxyapatite

MeHA

Methacrylated Hyaluronic Acid

BMP-2

Bone Morphogenetic Protein-2

HA

Hyaluronic Acid

HEA

Hydroxyethyl Acrylate

PLA

Polylactic Acid

hNC

Human Nasoseptal Chondrocytes

NFC

Nanocellulose Fibrils

Alg

Alginate

NC-Alg

Nanocellulose Alginate

CS

Chondroitin Sulfate

DS

Dermatan Sulfate

KGN

Kartogenin

PEGDA

Polyethylene Glycol Diacrylate

CSMA

Chondroitin Sulfate Methacrylate

DCMS

Decellularized Corneal Stroma Matrix

rCFs

Rabbit Corneal Fibroblasts

VICs

Valve Leaflet Interstitial Cells

3DBPO

3D-Bioprinted Osteosarcoma Model

Epi-SCs

Epidermal Stem Cells

ATMPs

Advanced Therapy Medical Products

GMP

Good Manufacturing Practice

4D

Four Dimensional

RF

Random Forest

SVM

Support Vector Machines

CNN

Convolutional Neural Network

ANN

Artificial Neural Network

GAN

Generative Adversarial Network

PCA

Principal Component Analysis

FDA

Food and Drug Administration

SWIFT

Sacrificial Writing into Functional Tissues

FRESH

Freeform Reversible Embedding of Suspended Hydrogels

The manuscript was written by MS and SK. All authors have given approval to the final version of the manuscript.

MS gratefully recognizes the support received through the Teaching cum Research Assistant (TRA) Fellowship. SK gratefully recognizes the financial support provided by the VIT SEED Grant (Sanctioned File No. SG20220120) and the Department of Science and Technology (DST), New Delhi, through the INSPIRE Faculty Fellowship (Faculty Registration No. IFA21-MS179).

The authors declare no competing financial interest.