Research progress and challenges of bioprinting in wound dressing and healing: Bibliometrics-based analysis and perspectives

Abstract

As the body’s largest organ, the skin has important roles in barrier function, immune response, prevention of water loss and excretion of waste. Patients with extensive and severe skin lesions would die due to insufficient graftable skin. Commonly used treatments include autologous skin grafts, allogeneic/allogeneic skin grafts, cytoactive factors, cell therapy, and dermal substitutes. However, traditional treatment methods are still inadequate regarding skin repair time, treatment costs, and treatment results. In recent years, the rapid development of bioprinting technology has provided new ideas to solve the above-mentioned challenges. This review describes the principles of bioprinting technology and research advances in wound dressing and healing. This review features a data mining and statistical analysis of this topic through bibliometrics. The annual publications on this topic, participating countries, and institutions were used to understand the development history. Keyword analysis was used to understand the focus of investigation and challenges in this topic. According to bibliometric analysis, bioprinting in wound dressing and healing is in an explosive phase, and future research should focus on discovering new cell sources, innovative bioink development, and developing large-scale printing technology processes.

1. Introduction

The skin is an important line of defense for the body against external environmental stimuli and hazards. The skin has a complex tissue structure consisting of three main parts: the epidermis, the dermis, and the subcutaneous tissue[1]. The epidermis is a dynamic, continuously self-renewing, multilayered epithelium composed mainly of keratinocytes. Keratinocytes produce keratin and are surrounded by rich lipids that play an important role in the skin’s protective function. The basement membrane connects the epidermis to the adjacent layer of the dermis. The dermis consists mainly of fibroblasts and the fibers and stroma they produce[2]. Owing to the high elastin content in this layer, it provides mechanical support and elasticity to the skin. The dermis is connected to the subcutaneous tissue, and this part mainly plays the role of connection, cushioning mechanical pressure, energy storage, and thermal insulation[3]. The complex tissue structure of the skin makes it crucial in protecting the body from the external environment, regulating the body’s physiological activities, and maintaining the health of the body[4].

The skin is the first barrier that protects the body and must be repaired promptly when it is damaged to form a wound. Repairing damaged skin is an extremely complex and dynamic process divided into a hemostatic phase, an inflammatory response phase, a cell proliferation phase, and a skin remodeling phase[5] (Figure 1). The hemostatic phase usually occurs immediately after the appearance of the wound and is dominated by platelets, which undergo an aggregation reaction. They form blood clots in the wound to fill the tissue gaps, creating a temporary wound matrix. This phase activates inflammatory cells by stimulating the release of interleukin (IL)-6, IL-1, tumor necrosis factor beta (TNF-β), and other factors while reducing blood loss. The inflammatory response phase and the hemostasis phase occur essentially at the same time. Neutrophils and monocytes aggregate at the site of trauma and remove bacteria, pathogens, necrotic tissue, and cells by phagocytosis. At the same time, these cells can secrete growth factors and cytokines that will lead the skin repair process to the next stage. The proliferative phase consists mainly of migration, proliferation and differentiation of fibroblasts, synthesis and deposition of extracellular matrix, neovascularization, and granulation tissue generation. The final stage is remodeling. Some of the mature sarcomeres further differentiate into myofibroblasts. The type Ι collagen fraction in the extracellular matrix increases and becomes the major fibronectin. Myofibroblast-forming cells interact with collagen bundles and growth factors to contract the wound and repair the skin.

Figure 1.

(A) Hemostasis, (B) inflammation, (C) proliferation, and (D) remodeling stages of wound healing.

Damage to the superficial epidermis of the skin can be regenerated by self-renewal of the epidermis, leading to wound repair and healing. However, self-repair is not possible for large areas of deep skin trauma and manual intervention is often required for treatment[6]. For significant skin defects and difficult-to-heal skin wounds, natural or artificial skin substitutes must be grafted on their surfaces. However, the availability of autologous skin for transplantation is limited, and allogeneic or xenogeneic skin grafts have some immune rejection problems[7]. Therefore, there are significant limitations when using traditional methods to treat large skin lesions, and there is an urgent need to create artificial skin that can be used for skin grafts.

Biological three-dimensional (3D) printing technology is the computer-controlled loading and delivery of pre-designed 3D models with precise positioning of bioink to manufacture human tissue[8]. Bioprinting is more compatible with human ergonomic design than tissue engineering culture methods. The concept of bioprinting technology was first introduced by Mironov et al.[9,10] in 2003. During their exploratory studies, they found that the embryonic heart tubes of chicks were cut into separate rings of cardiomyocytes. The rings of cells were arranged densely on a ring-shaped scaffold and could be fused over time in culture to form a tubular structure similar to a blood vessel. They thus proposed for the first time “cell printing,” which accurately stacks cells and materials into 3D bodies with a specific morphological structure, just like printing. The cells grow by proliferation, fusing themselves into tissues or organs and eventually generating tissues. Wilson and Boland[11] performed the printing of bovine aortic vascular endothelial cells and bovine serum proteins labeled by fluorescein in the same year and found that the printed vascular endothelial cells and serum proteins were still active.

There are several bioprinting technologies, divided into three main types: material extrusion (mechanical and pneumatic) bioprinting[12,13], material jetting (inkjet, microvalve, laser-assisted, acoustic) bioprinting[14,15], and vat polymerization (stereolithography, digital light processing, two-photon polymerization)[16,17]. Compared with traditional tissue engineering research methods, the main advantages of bioprinting are as follows:

• (i) The accurate 3D image processing platform can manufacture suitable organs or tissues according to the actual demand and realize personalization in the medical field.

• (ii) Multiple printhead arrays can be used for simultaneous printing. This allows simultaneous use of different types of cells, growth factors and biomaterials, and precise control of the constituent materials and cells during the printing process according to the tissue-specific composition ratio, which is more conducive to the construction of 3D scaffolds.

• (iii) It can precisely locate the extrusion position of bioink and control the amount of ink extrusion, enabling positioning, and quantitative spot printing control. This helps build the internal microstructure of tissues and control the content of growth factors required for the internal growth of tissues, thus enabling local growth and development.

The ability to create biological organs/tissues in a very short period ensures cell viability and promotes the development of regenerative medicine.

Bioprinting occupies a significant position in biomanufacturing, and its concept and technology realization is closely related to the rapid development of life science informatics, material science, manufacturing science, engineering science, and other disciplines. To date, many reviews on bioprinting technologies have been published[18-23], some of which focus specifically on applications in wound dressing and healing[24-27]. Bibliometric analysis is a literature and information mining method based on mathematical statistics. It can reflect research trends and hotspots through clustering relationships of keywords in the literature and has become an important tool for global analysis in various scientific fields[28-36]. Among them, bibliometrics has been applied to the analysis of 3D printing-related topics. For example, Rodríguez-Salvador et al.[37] analyzed scientific publications and patents in the field of 3D printing for the purpose of understanding the state of development of this field and analyzed the institutions and companies that have important role in this field. In another work[38], they used the competitive technology intelligence cycle to perform another scientometric analysis of bioprinting technologies. In this work, they focused on authors and affiliations. Naveau et al.[39] also performed a similar analysis in this area, although they mainly used VOSviewer as the analysis software. In recent years, bibliometrics has been further applied to the analysis of various directions in the field of 3D printing. For example, Rodriguez-Salvador et al. recently analyzed the research dynamics of tissue spheroids[40] and bioinks[41] using bibliometrics. We focused on another application direction of 3D printing.

This current review summarizes the development of bioprinting in wound dressing and healing. Two bibliometrics software were used in this systematic literature review. The first is CiteSpace, developed by Dr. Chaomei Chen, a professor at the Drexel University School of Information Science and Technology[42-45]. CiteSpace 6.1R2 was used to calculate and analyze all documents. COOC is another emerging bibliometrics software[46]. COOC12.6 was used to analyze annual publications and keywords co-occurrence. We used the core collection on Web of Science as a database to assure the integrity and academic quality of the studied material. “Bioprinting wound” or “bioprinting ‘skin tissue engineering’” were used as a “Topic.” The retrieval period was indefinite, and the date of retrieval was May 2022. A total of 196 articles were retrieved, spanning the years 2011 to 2022. All relevant papers were then manually screened by a co-author (from a 3D printing company) to ensure that they all fit into the analysis of the topic.

2. Developments in the research field

2.1. Literature development trends

Figure 2 shows the annual and cumulative number of publications on wound dressing and healing between 2011 and 2021. Based on this figure, the first paper on this topic was published in 2011. Miller et al.[47] inkjet-printed heparin-bound epidermal growth factors in different concentrations on fibronectin substrates and investigated how they direct cell migration to the wound site. This topic did not receive extensive attention until 2018, with less than 10 publications being published on this topic per year. Until 2018, the annual publication of this topic started to enter an upward period. By 2020, the annual publication volume of this topic suddenly rose to 31 articles, which is 55% higher than that of 2019. The upward trend of 2021 was further strengthened with 58 articles. By the end of writing the Introduction section of this paper, the data for this bibliometric analysis were counted through May 2022, so the data for this year cannot be added to Figure 2. However, we believe that the increasing trend of annual publications on this topic will not be changed, as 43 papers have already been published in 6 months. From Figure 3, it can be concluded that bioprinting in wound dressing and healing is undergoing a rapid development phase. This is the most dynamic and breakthrough phase of the topic. Many scientists, resources, and attention will be devoted to this topic to overcome the challenges involved. This phase, if maintained for a more extended time, will help attract the attention of interdisciplinary scholars and further develop the possibilities of this topic. The following content covers the careful analysis of the areas of this topic.

Figure 2.

Annual and accumulated publications from 2011 to 2021 searched in the Web of Science about bioprinting in wound dressing and healing.

Figure 3.

Top 10 journals that published articles on the bioprinting in wound dressing and healing.

2.2. Journals, cited journals, and research subjects

Figure 3 shows the top 10 journals that published the most papers regarding bioprinting in wound dressing and healing. The materials science-related journals occupy a large proportion of them, such as Polymers, Biomedical Materials, Acta Biomaterialia, Bioactive Materials, ACS Applied Materials & Interfaces, Carbohydrate Polymers, and Advanced Functional Materials. Polymers are the most common choice for wound dressings, such as polyurethane, so polymer-related journals appear frequently here. Because bioprinting involves the relationship between materials science and biology, journals in the category of biomaterials also appear with high frequency. On the other hand, Pharmaceutics and Burns & Trauma have also published papers on the topic of wound dressing and healing as it is an important topic in tissue engineering and medicine. As can be seen from the composition of these journals, the content of this topic is mainly related to materials science and medicine.

In addition to the number of papers published by the journal on the topic, the frequency with which the journal papers on this topic have been cited is also an important indicator. Table 1 shows the top 16 cited journals on bioprinting in wound dressing and healing. It can be seen that most of the journals involved, as shown in Figure 3, appear in Table 1, representing that these journals primarily publish papers on this topic and are often cited by papers on this topic. In addition to the journals in Figure 3, some new journals appear in Table 1, which are mainly about materials science. On the other hand, comprehensive journals also appear in Table 1, such as Scientific Reports and PLoS One. In addition, a number of chemistry, biotechnology, and medical journals also appear in Table 1. Combined with the information in Figure 3 and Table 1, the main areas covered by bioprinting in wound dressing and healing include materials, chemistry, medicine, and biotechnology.

Table 1.

Top 16 cited journals on the bioprinting in wound dressing and healing

No.	Citation	Cited journal
1	137	Biomaterials
2	114	Biofabrication
3	113	Acta Biomaterialia
4	101	Advanced Healthcare Materials
5	97	Scientific Reports
6	93	Advanced Materials
7	88	ACS Applied Materials & Interfaces
8	79	Materials Science and Engineering: C
9	73	Carbohydrate Polymers
10	72	International Journal of Biological Macromolecules
11	72	PLoS One
12	71	Nature Biotechnology
13	66	Biomaterials Science
14	64	International Journal of Molecular Sciences
15	63	Tissue Engineering Part A
16	63	Advanced Drug Delivery Reviews

Although the most important journals on this topic and the fields to which they belong can be known from Figure 3 and Table 1, they do not present the most cutting-edge advances. Since the last 2 years have been an explosive period for this topic, analyzing the journals that published on this topic for the first time in these 2 years can give an idea of the most cutting-edge areas involved (Table 2). As can be seen, in addition to material science journals, many biotechnology and bioengineering journals are beginning to publish bioprinting in wound dressing and healing. This represents a gradual shift in the investigation of bioprinting in wound dressing and healing from materials science preparation to specific application analysis. For example, Zhang et al.[48] demonstrated using 3D printing technology that mechanical forces can control vessel growth in the direction of mechanical stimulation without branching. In vivo experiments verified that the vessels successfully functioned as blood vessels with consistent orientation. On the other hand, a growing number of papers on this topic are being published by medical journals, indicating the performance and potential of bioprinting in addressing wound dressing and healing in addition to the positive results of the technique in some model experiments, which are more often attempted in pharmacological experiments and clinical trials. For example, Glover et al.[49] designed a 3D bioprinted scaffold that could specifically deliver levofloxacin to the diabetic foot ulcers and could continuously release drug for 4 weeks.

Table 2.

List of journals published papers regarding the bioprinting in wound dressing and healing for the first time

Year	Journals
2021	3D Bioprinting, APL Bioengineering, Beilstein Journal of Organic Chemistry, Bioactive Materials, Biomaterials Science, Biomedicines, Biotechniques, Biotechnology and Bioengineering, Biotechnology Progress, British Journal of Surgery, Expert Opinion on Therapeutic Patents, Express Polymer Letters, Frontiers in Chemistry, International Journal of Biological Macromolecules, International Journal of Lower Extremity Wounds, International Journal of Pharmaceutics, Journal of Bionic Engineering, Journal of Food Processing and Preservation, Journal of Tissue Engineering, Journal of Tissue Engineering and Regenerative Medicine, Journal of Translational Medicine, Korean Journal of Chemical Engineering, Materials, Materials Science & Engineering C-Materials for Biological Applications, Nature Protocols, New Journal of Chemistry, NPJ Regenerative Medicine, Pharmaceuticals, Progress in Polymer Science, Regenerative Biomaterials, Wound Repair and Regeneration,
2022	Advanced Materials Technologies, Bio-Design and Manufacturing, Bioengineered, Bioengineering & Translational Medicine, Biomedicine& Pharmacotherapy, Cell Stem Cell, Chemical Engineering Journal, Chemistry-An Asian Journal, Drug Delivery and Translational Research, Experimental Dermatology, International Journal of Molecular Sciences, Journal of Applied Polymer Science, Journal of Biomaterials Science-Polymer Edition, Journal of Cleaner Production, Journal of Dermatological Treatment, Journal of Drug Delivery Science and Technology, Journal of Materials Chemistry B, Progress in Chemistry, Research, Starch-Starke

The category of the published paper can reflect the evolution of the topic. Figure 4 shows the evolution of the category of bioprinting in wound dressing and healing over time. This topic started in the categories of Materials Science, Biomaterials and Engineering, and Biomedical. Between 2011 and 2014, this topic began to cover biotechnology-related categories, including Cell & Tissue Engineering, Cell Biology, and Biotechnology & Applied Microbiology. Between 2015 and 2018, this topic began to move into a range of materials science and chemistry areas, representing a phase where wound dressing and healing concerns were focused on experimenting with different material combinations and chemical modulation. In recent years, this field has gradually extended from chemistry and materials science to several new categories. For example, this topic was first published in Computer Science, Theory & Methods in 2020. Alblawi et al.[50] discussed the cellular Potts model and cellular particle dynamics model applied in bioprinting technology. Subsequently, they proposed mechanical sensing of mesenchymal stem cells on thermally responsive polymer surfaces for bioprinting technology by introducing stresses generated by acoustic energy or polymer strain energy functions.

Figure 4.

Time-zone view of research categories for bioprinting in wound dressing and healing.

2.3. Geographic distribution

Figure 5 shows a plot of different countries’ participation levels on this topic. The USA contributed to 25.25% of the papers published on this topic. The second place was claimed by China, which contributed to 23.74% of published papers. Spain, England, Singapore, and India were involved in more than 5% of the published papers. According to the distribution of countries, it is known that this topic attracts the attention of scientists and doctors from all regions of the world, except South America and Africa (only Egypt and South Africa have published 1 paper each). However, as this topic is in its explosive phase, it will continue attracting new academic groups to join the research avenue. For example, the first papers on this topic were published by researchers from United Arab Emirates, Argentina, France, and Egypt in the first 5 months of 2022.

Figure 5.

Rose plot of papers related to bioprinting in wound dressing and healing contributed by different countries.

Figure 6 illustrates the cooperation network between the different institutions on this topic. As can be seen, this topic has only resulted in an influential collaborative network. This cooperation network was mainly led by the university and research institutions from the USA and Singapore, including Harvard University, Harvard Medical School, National University of Singapore, and Agency for Science, Technology and Research (ASTAR). This collaborative network also covers research institutions and universities from Norway, China, and Iran.

Figure 6.

Institution cooperation network for bioprinting in wound dressing and healing.

Figure 7.

Grouping of keywords for bioprinting in wound dressing and healing.

3. Keyword analysis and evolution of the field

Keywords analysis can be used to further understanding the different research directions under a topic. Table 3 lists the top 14 keywords of bioprinting in wound dressing and healing. Some of these high-frequency keywords are directly related to the topic, such as 3D bioprinting, wound healing, and tissue engineering. Other high-frequency keywords reflect the different focus of investigation under this topic. For example, “scaffold” came in second place as a high-frequency keyword. The two most prominent types of bioprinting are scaffold printing and cell printing. The use of 3D bioprinting technology can potentially improve classical tissue engineering methods as it allows the generation of scaffolds with high spatial control of endothelial cell distribution. Cubo et al.[51] developed a modified extrusion-based 3D bioprinter. The extrusion module of this bioprinter consists of four sterile disposable syringes for holding and extruding human blood plasma, fibroblasts, keratinocytes, and calcium chloride solution, respectively. The extruded bioprinter was used to print a two-layer skin substitute containing both dermal and epidermal layers. The most significant breakthrough in their work is that they were able to print artificial skin scaffold replacements in less than 35 min, offering the possibility of producing skin in an automated and standardized manner. The appearance of “in vitro” as a high-frequency keyword does not mean that the investigation of bioprinting technology has so far yielded data from in vitro experiments. Bioprinting-generated skin is a skin-like substitute with natural skin tissue structure, which can be used to prepare in vitro using cells, extracellular matrix components, and bioactive factors. Different types of cells have been used for bioprinting in vitro, so “stem cell” and “mesenchymal stem cell” also appear in Table 3. Stem cells are cells that can renew and differentiate themselves. Stem cells for skin printing are epidermal stem cells, dermal stem cells, adipose stem cells, melanin stem cells, mesenchymal stem cells, etc. Stem cells are an important source of seed cells for skin bioprinting. They can solve problems that existing technologies cannot solve, such as the lack of blood vessels, sensory receptors, skin appendages, etc. Skardal et al.[52] used laser deposition bioprinting techniques to unite stem cells with the potential to differentiate into blood vessels and found that they greatly facilitated wound healing after injury. Growth factor is also one of the high-frequency keywords in Table 3. The most challenging aspect of skin printing is achieving formation of multiple blood vessels, and early attempts to overcome this difficulty involve using vascular endothelial growth factors and keratinized cells or scaffolds, but more research is needed in this area[53]. Therefore, incorporating growth factors in the ink of bioprinting is an essential part of this topic. The keywords “hydrogel,” “drug delivery,” and “extracellular matrix” are also included in Table 3. Hydrogels are highly hydrophilic 3D polymer networks prepared by chemical or physical cross-linking of natural or synthetic polymers[54] that can swirl rapidly upon water absorption and maintain their structural integrity. The physical properties of the dissolved hydrogel include soft texture, high elasticity, and low interfacial tension. It reduces irritation to the surrounding skin tissue when applied as an extracellular matrix substitute for the skin. Its swelling property can promptly absorb tissue exudate, keep the wound clean, reduce wound infection, and thus accelerate wound repair. In addition, it shows excellent physical or structural similarity to natural tissues and can mimic the natural extracellular matrix very well. The porous structure of hydrogels facilitates the transport of oxygen, nutrients, and some growth factors. This provides a suitable spatial microenvironment for cell growth during skin tissue regeneration and acts as a drug delivery pathway[55]. Therefore, biocompatible hydrogels have a wide range of applications in skin tissue engineering and biomedicine.

Table 3.

List of top 14 keywords for bioprinting in wound dressing and healing

No.	Freq	Centrality	Keywords
1	44	0.14	3D bioprinting
2	42	0.23	Scaffold
3	41	0.21	In vitro
4	33	0.12	Wound healing
5	28	0.14	Tissue
6	28	0.17	Stem cell
7	27	0.15	Hydrogel
8	26	0.12	Drug delivery
9	25	0.12	Tissue engineering
10	23	0.08	Fabrication
11	19	0.12	Mesenchymal stem cell
12	18	0.05	Extracellular matrix
13	16	0.04	Design
14	15	0.05	Keratinocyte

The technologies currently used for bioprinting of skin are inkjet printing, laser-assisted printing, and extrusion printing. To gain further insight into the use of these printing techniques, we manually identified papers investigated in this work (excluding review, book chapter, and meeting abstract). Extrusion printing is the most commonly used technique in this topic. More than half of all the papers used extrusion printing technology to print wound dressings. Inkjet printing is also a commonly used technology, accounting for about 35% of all papers. However, inkjet printing is not widely used in wound dressings because of the restriction imposed on the concentration of bioink. Laser-assisted bioprinting has only recently begun to be used to print wound dressings because of its ability to kill cells. In our survey, only nine articles used this technique.

Cluster analysis can help further understand the different directions of investigation in this topic. Figure 8 shows that 13 clusters were formed. On the whole, many clusters have overlapping areas, indicating the similarities of their contents. This reflects that the topic of bioprinting in wound dressing and healing has not spawned many subdivisions. Table 4 describes the clusters and their ID, size (number of papers), silhouette, and respective keywords. The following is a short explanation of each cluster:

Table 4.

Size, silhouette, respective keywords, and references of bioprinting in wound dressing and healing for each cluster

Cluster ID	Size	Silhouette	Keywords	References
0	44	0.897	3D bioprinting, Cross linking, Chitosan, Cell, Wound dressing, Release	[56-59,96-112]
1	26	0.906	Wound healing, Tissue engineering, Mesenchymal stem cell, Antimicrobial activity, Chitosan based hydrogel,	[60-64,95,98,113-121]
2	25	0.850	Design, Fibroblast, Biomaterial, Wound healing, Artificial skin	[24,51,75,80,122-127]
3	24	0.942	Extracellular matrix, Hyaluronic acid, Skin, Chronic wound	[6,70,71,128-134]
4	23	0.861	In vitro, Growth factor, Model, Regenerative medicine	[6,47,72-77,135-139]
5	22	0.917	Keratinocyte, Angiogenesis, Transplantation, Regeneration, Biofabrication, Technology, pH	[26,76,78,96,140-143]
6	21	0.938	Hydrogel, Delivery, Differentiation, Matrix, Burn	[50,52,67,77,79-82,99,144-146]
7	19	0.911	Scaffold, Stem cell, Drug delivery, Tissue, Fabrication, Nanofiber	[6,24,47,56,58,83-85,96,98,115,124,136,141,142,146-155]
8	15	0.844	3D printing, Tissue engineering, Repair, Cellulose nanofiber, Stiffness, Additive manufacturing	[25,82,86-88,145]
9	13	0.764	Mechanical property, Skin regeneration, 3D, Prevascularized tissue, Hydrogel scaffold	[27,83,89-91]
10	11	1.000	Diabetic foot ulcer, Endothelial cell, Clinical trial, Animal model	[135,156,157]
11	10	0.957	Film, Collagen, Honey, 3D bio-printing	[93,94]
12	4	0.986	Composite hydrogel, Drug delivery, Network hydrogel, Tough	[95]

3.1. Hydrogel.

This cluster contains a series of works on bioprinted hydrogels. For example, Maver et al.[56] compared the effectiveness of bioprinted and electrostatically spun carboxymethylcellulose hydrogels as wound dressings. Diclofenac sodium and lidocaine were added to the hydrogel preparation for wound pain relief. They not only characterized the physicochemical, structural, and morphological properties of the hydrogels derived from the two preparations, but also examined their biocompatibility. Abasalizadeh et al.[57] reviewed the hydrogels formed by alginate with inorganic cations and their potential application as bioinks in bioprinting. Although polysaccharide hydrogels have been widely used, it has been challenging to prepare mechanically stable scaffold materials using only hydrogels. Milojević et al.[58] proposed a hydrogel–plastic polymer composite scaffold. This composite scaffold is an intermediate between pure hydrogel and pure thermoplastic polymer, which can provide both gradient surface properties and good biocompatibility. In wound care, collagen is an excellent choice for biological dressings. Bioprinted collagen has great potential for wound dressing and healing. However, soluble collagen fibrillates at neutral pH and 37°C, thus limiting its use in wound dressing and healing. Hartwell et al.[59] proposed the addition of polyvinyl alcohol:borate hydrogels to overcome this challenge. The results showed that adding this hydrogel to the collagen solution improved stability. Cultured cells also exhibited more organized f-actin and reduced abundance of pro-collagen and α-smooth actin.

3.2. Polysaccharide.

The content of this cluster also focuses on the preparation of different hydrogels. However, polysaccharides were focused on in this cluster. In particular, two articles focus on the value of carrageenan in bioprinting[60,61]. Xu et al.[62] summarized the hydrogels synthesized with Schiff bases and polysaccharides and their applications. Graham et al.[63] investigated the properties of a range of thermosensitive polysaccharides and how these properties can be applied to bioprinting. Rastin et al.[64] wanted to improve the antimicrobial properties of polysaccharide hydrogels. They chose to use Ga³⁺ in the bioink formulation design. The Ga³⁺ cross-linked bioink exhibited potent antimicrobial activity against Gram-positive (Staphylococcus aureus) and Gram-negative bacteria (Pseudomonas aeruginosa).

3.3. Skin.

This cluster focuses on the interaction between bioink and skin cells. Bioprinting inks govern the print resolution and the quality of bioprinted tissue. Bioprinted ink materials provide physical and biochemical microenvironmental signals to seed cells that can influence cell polarity and control cell migration. Skin heat dissipation accounts for 90% of the total heat dissipation in the body. The critical role of the skin cooling system is played by the sweat glands, located in the lower dermis, which secrete sweat, excrete waste, and regulate body temperature. Bioprinted skin requires the construction of interconnected ducts between the basal epidermal layers to uniformly excrete waste to achieve the sweat gland function of the skin. The hair follicles are connected to the sebaceous ducts that carry the secretions produced by the body to the surface of the skin for discharge. No studies have successfully induced the production of new human hair follicles due to the lack of hair papilla cells with dermal sensing properties. In order to more realistically simulate the physiological functions of the skin, hair papilla cells should be added to the dermal papilla and epidermal layers. The bioprinted 3D spatial structure can partially restore the hair induction properties and eventually achieve hair regeneration[65]. This cluster contains melanocytes-and fibroblasts-related bioprinting. Melanin secreted by melanocytes can be used to regulate skin color and protect against ultraviolet rays. Min et al.[66] printed melanocytes and keratinocytes on the top of the dermis and observed freckle-like pigmentation at the dermo-epidermal junction. Fibroblasts are found in the dermis of the skin and are responsible for the production of extracellular matrix and non-fibrous components. Shi et al.[67] fabricated a novel dermal replacement scaffold using sodium alginate/gelatin composites and fibroblasts through extrusion molding, which has similar physical and chemical properties to human skin tissue. Won et al.[68] used skin-decellularized extracellular matrix powders and fibroblasts as bioinks and found that gene expression in cells was similar to skin morphologic biology. Ng et al.[69] used three different types of skin cells (keratinocytes, melanocytes, and fibroblasts) to produce pigmented human skin structures and showed light pigmentation similar to that of the skin donor.

3.4. Healing process.

Skin trauma healing includes interaction of multiple cells, growth factors, and cytokines. The articles and reviews in this cluster focus on the functions of growth factors and bioactive molecules in bioprinted skin. For example, Huang et al.[70] created a functional in vitro cell-loaded 3D extracellular matrix mimic. This biological 3D structure effectively creates local ecological niches for epidermal progenitor cells, ensuring unilateral differentiation into sweat cells. Schmitt et al.[71] reported a closed-loop fat-processing system that processes fat aspirates into microfat clusters. The microfat can be mixed with methacrylic acid collagen bioink to generate microfat-rich collagen structures by bioprinting. This collagen structure remains viable and metabolically active in vitro for 10 days.

3.5. Human skin

This cluster contains 13 papers, but it has a silhouette value of only 0.861, which represents not very excellent clustering. This cluster contains papers on the effectiveness of different bioprinted dressings in wound healing[6,72-75] and a series of investigations on bioprinting using stem cells[47,76,77].

3.6. Keratinocyte.

Keratinocyte is a high-frequency keyword in this cluster. Keratinocytes and fibroblasts of the skin can be accumulated using an extrusion technique and cultured over time to form tissue cells with epithelium and dermis. Michael et al.[78] placed fibroblasts and keratinocytes on a stable matrix, and found that printed keratinocytes formed a multi-layered epidermis with beginning differentiation and stratum corneum.

3.7. In situ bioprinting.

In situ bioprinting and handheld instruments are the focus of attention in this cluster. The in situ skin-printing procedure is generally divided into two phases[79,80]. In phase I, fibroblasts and keratinocytes are isolated from the discarded skin fragments. The cells are proliferated and mixed with biopolymers, i.e., fibrin and collagen type I, to prepare the bioink. In phase II, the area to be repaired is placed underneath the portable bioprinter, and the bioink is printed directly onto the wound, which is then left to allow the material to fuse repeatedly. During the printing process, no part of the printer directly interacted with the animal. The results showed that after 8 weeks of in situ printing, a fully differentiated epidermis could be seen, keratinocytes fully proliferated and covered the wound, and the dermis grew to the wound edge. Handheld instruments were also developed with the hope of translating biofabrication into the field of surgery. O’Connell et al.[81] described a handheld bioproduction tool called “biopen.” Cheng et al.[82] also described a handheld instrument to deliver fibronectin-containing mesenchymal stem/stromal cells directly to the trauma surface, improving re-epithelialization, dermal cell regeneration, and neovascularization.

3.8. Biomaterial.

The papers of this cluster mainly emphasize the properties of different biomaterials. For example, the paper by Ma et al.[83] highlights the vascularization-inducing function of strontium silicate microcylinders. Ulusu et al.[84] emphasize the thermal stability and fluidic properties of a polymeric biomaterial called Caf1. Pitton et al.[85] also highlighted the hydrofluidic properties of pectin-cellulose nanofibers.

3.9. Scaffold.

This cluster contains four different scaffolds for tissue engineering. A thermosensitive hydrogel was prepared by Boffito et al.[86] Xia et al.[87] prepared a curcumin-incorporated gelatin methacryloyl hydrogel. A bioactive microgel was synthesized by de Rutte et al.[88] A biomaterial sheet was prepared by Cheng et al.[82]

3.10. Bioink.

The silhouette value of this cluster is only 0.764, which is the lowest among all clusters. It contains four reviews and one research paper. Masri et al.[89] presented a strategy for printability quality improvement of bioprinting for skin regeneration and wound healing. Nie et al.[90] provided a summary and outlook on the bioassembly by microfluidics. Serban et al.[91] presented hyaluronic acid for 3D structural assembly. Wang et al.[27] specifically presented extrusion-based bioprinting for wound dressing and healing.

3.11. Diabetic wound.

The treatment of diabetic wounds is also important for bioprinted skin dressings and healing. The most frequently occurring keyword in this cluster is diabetic wound. Wan et al.[92] prepared a bilayer skin scaffold using gelatin as the matrix material using 3D printing technology. The upper layer of the scaffold consisted of gelatin cryogel loaded with silver nanoparticles, and the lower layer consisted of printed gelatin scaffold loaded with platelet-derived growth factor. The bilayer skin scaffold was shown to promote granulation tissue formation, collagen deposition, and neointima formation in a diabetic mouse wound model. This bilayer skin scaffold accelerates the epithelial re-formation process and shows extraordinary potential for treating diabetic wounds.

3.12. Film.

This cluster contains two articles. A pectin-based bioprinting ink was proposed by Andriotis et al.[93] This ink forms a transparent film after drying and can rapidly decompose after contact with water. Rees et al.[94] prepared a transparent nanocellulose-based film that can provide a moist wound healing environment and form an elastic gel with bioresponsive properties.

3.13. Nanocomposite

This cluster contains only one review. Traditional hydrogels are less likely to be used directly in wound dressings and healing because of their poor physical properties, and work to improve hydrogels with nanoparticles is summarized by Barrett-Catton et al.[95] The composite hydrogels tend to exhibit superior physical and biochemical properties.

Figure 9 shows the frequency of occurrence between keywords. The results in Figure 9 can verify the clustering analysis results in the above. Since the application of bioprinting in wound dressing and healing is a topic in tissue engineering, both “tissue engineering” and “bioprinting/ wound healing” have a high frequency of co-occurrence. The assembly of scaffolds is also a very important direction in this topic, so “scaffolds” and “fabrication” also have a high frequency of co-occurrence. Meanwhile, hydrogels are the most commonly used option in bioprinted wound dressings. Among them, chitosan is one of the most commonly used raw materials for making hydrogel.

Figure 9.

Keywords confusion matrix for bioprinting in wound dressing and healing.

4. Conclusion and perspectives

Bioprinting is an important new technology in wound dressing and healing. This bibliometric-based investigation provides a statistical summary of how the topic has evolved between 2011 and 2022. Bioprinting is a cost-effective and efficient production method that helps address challenges like high production costs and slowing profits in the skin repair material industry and develop products with better performance. Compared with traditional skin tissue engineering technology, bioprinting technology can locate cell precise and produce complex and controllable structure. Based on the above analysis, the following conclusions can be drawn:

• (i) Bioprinting for wound dressing and healing has been published since 2011 but has not attracted much attention for a short period of time. This topic gained traction in 2018 and has continued to grow in the following years. Based on the bibliometric analysis, this trend does not show signs of stagnation. Therefore, this topic will continue to be dynamic for some time.

• (ii) Based on the category’s analysis, it can be concluded that developments in materials science, especially innovations in polymeric materials, are significant for this topic. Biomaterials are essential to ensure the viability and responsiveness of cells in the printed architecture. It must be biocompatible, biodegradable, printable, biologically inert, strong, durable, and malleable. For example, the hydrogel is one of the most commonly used materials to be printed. However, traditional hydrogels have some detrimental properties to wound dressing and healing, such as poor physical properties. These properties can be improved by compounding hydrogels with other materials.

• (iii) The most common type of material used is polymers. They can be divided into natural and synthetic polymers according to their production sources. Natural biopolymers are naturally occurring biopolymers, including proteins (e.g., collagen, gelatin, fibrinogen, etc.) and polysaccharides (e.g., chitosan, alginate, etc.). Synthetic biopolymers are artificial polymers, including polylactic acid, polyglycolic acid, polycaprolactone, etc. Synthetic biopolymers have excellent mechanical properties and can adapt mechanical properties and degradation characteristics to skin grafts by modifying the polymer structure.

• (iv) There is a strong interest in this topic from countries other than South America and Africa. The USA, China, Spain, England, and India contributed the most significant number of papers on this topic. Since the research on this topic is in a very hot state, researchers from some new countries would contribute papers to this field every year. More countries joining the research on this topic is to be expected.

• (v) This topic has resulted in an influential collaborative network. This cooperative network was mainly led by the university and research institutions from the USA and Singapore.

• (vi) The cells are the ultimate source to ensure the durability and function of the patient’s regenerated skin tissue. Currently, the most commonly used cell types in the manufacturing process of wound healing and skin regenerations are keratinocytes and fibroblasts. However, using only these two types of cells is not able to fully develop the various functions of the skin, so other cells and appendages are needed. Among them, stem cells are unique cell populations that are able to self-renew and differentiate. Stem cells can release specific growth factors and cytokines. These growth factors induce the skin to produce proteins, elastic fibers, and new proteins.

• (vii) The leading technologies currently used for bioprinting of skin are inkjet printing, laser-assisted printing, and extrusion printing. In addition, in situ printing and non-invasive in vivo printing are new modalities developed.

Meanwhile, based on the review of this topic, we believe that the following issues regarding the bioprinting in wound dressing and healing need to be investigated:

• (i) Although inkjet printing allows for high-precision deposition of bioinks, it may expose cells and materials to thermal and mechanical stress environments. High-frequency sound waves can affect the viability of cells. Inkjet printing is also limited by the viscosity of the bioink used. Therefore, inkjet printing is used as a printing technology limited to low-viscosity biological inks. Inkjet printing is often used with extrusion printing to achieve vascularized skin printing. Further exploration of the coupling of this technology is necessary.

• (ii) Light-cured printing does not use nozzles, so high cell density printing is possible. However, the irradiation and curing process of UV light will inevitably cause damage to the cells in the bioink. Bioinks need to be mixed with photoinitiators to achieve curing, and the presence of photoinitiators may make bioinks toxic. In addition, it is necessary to develop non-UV light as a curing light source and biological photoinitiator.

• (iii) Laser-assisted printing is also a nozzle-free technology that offers similar benefits to light-cured printing. However, metal particles can contaminate cells when they absorb high-energy laser beams. The high air pressure generated by local evaporation affects cellular activity. The high cost of laser-assisted printing and the low cell survival rate have led to limited research on the application of this technology. There are several challenges of reducing these disadvantages.

• (iv) The main disadvantage of extrusion printing is the low speed and resolution, while the presence of shear stress affects the viability of the cells. Improving the resolution and cell viability is an important direction in this technique.

• (v) It is still a challenge for the current bioprinting technology to fabricate complex skin structures, such as hair follicles and other skin attachments, although it is now able to fabricate microscopic structures such as papillae. Applying advanced micro- and nano-manufacturing technology to skin printing can improve precise and uniform pigment, hair follicle deposition, and printing efficiency.

In situ and non-invasive in vivo printing requires more dexterous and convenient devices for the rapid closure of wounds in specific body parts. To reduce the impact of respiratory amplitude on scanning and printing accuracy in in situ printing, a more advanced real-time feedback system can obtain more accurate scanning data and monitor the patient’s physiology in real time during the printing process to optimize the printing path at any time.

Funding

This work was supported by the National Natural Science Foundation of China (22004026).

Conflict of interest

The authors declare no conflicts of interest.

Author contributions

Conceptualization: Li Fu, Hassan Karimi-Maleh

Investigation: Shuduan Mao, Junjie Man, Jialei Wang, Chengliang Yin

Methodology: Li Fu

Formal analysis: Shuduan Mao, Junjie Man, Jialei Wang, Chengliang Yin

Writing – original draft: Shuduan Mao, Junjie Man, Jialei Wang, Chengliang Yin

Writing – review & editing: Li Fu, Hassan Karimi-Maleh