Effect of natural-based biological hydrogels combined with growth factors on skin wound healing

2.1 The structure and function of normal skin tissue

Normal skin tissue consists of the epidermis and dermis. The epidermis layer is located on the outside of the skin. More than 95% of the cells in the epidermis are keratinocytes, with the rest made up of melanocytes, Merkel cells, and Langerhans cells [15]. As shown in Figure 1, the epidermis is divided into a corneous layer, transparent layer, granular layer, spinous layer, and basal layer from outside to inside [16]. The structural integrity and activity of the cells in each layer increased from the outside to the inside, and the cells in the inner layer gradually transformed to the outer layer over time [17]. The cells of the corneum have been completely keratinized, and the desmosomes between the cells are loose and shed to form dandruff [18]. The lucidum layer consists of 2–3 layers of flattened cells with fuzzy cell boundaries and the absence of nuclei and organelles [19]. The cells in the granular layer are mainly spindle cells, with nuclei and organelles that begin to degenerate, and there are many lamellar granules and keratohyalin granules in the cells [20]. There are a large number of spinous cells in the stratum spinosum, which have a strong protein synthesis function and can synthesize a large number of keratin filaments and lamellar granules [21]. At the same time, the desmosomes between the spinous cells are closely connected, which can prevent external substances from penetrating the epidermis and has the function of protection and isolation [22]. The basal layer is the most important part of the epidermis. The self-renewal and repair of the epidermis mainly depends on the basal cells attached to the basal membrane in the basal layer [23]. These basal cells are the stem cells in the epidermis with the ability to proliferate and differentiate [24]. In the process of proliferation, basal cells can separate from the basal membrane and differentiate into spinous cells, which can replenish the number of spinous layer cells and play a role in self-renewal [21]. After skin injury, the number and activity of basal cells have an important influence on the healing effect.

Figure 1

Normal structure of skin. Normal skin tissue consists of the epidermis and dermis. The epidermis is divided into a corneous layer, transparent layer, granular layer, spinous layer, and basal layer from outside to inside.

The dermis is a dense connective tissue located below the epidermis, which is divided into the papillary layer and the reticulated layer. There are abundant capillaries in the papillary layer, which protrude to the epidermis to form papillary structures, increasing the contact and connection between the epidermis and dermis and facilitating the epidermis to obtain nutrients from the dermis [25]. The reticulated layer contains a large number of collagen fibers, which interweave to form a network that provides skin with toughness and elasticity [26]. Under normal conditions, the basal cells in the basal layer of the epidermis obtain nutrients from the dermis, and while proliferating and renewing themselves, some of the basal cells differentiate into spinous cells to supplement the number of spinous cells and maintain normal epidermal functions [27]. As time passes, the cells in the lateral layers gradually shift to the more lateral layers until they fall off as keratinocytes.

2.2 Physiology of wound healing

The normal skin tissue structure can protect the internal tissues and organs of the body from damage by external harmful substances and is the first barrier to maintain homeostasis of the human body environment. When the skin is injured by external factors such as surgery, external forces, chemical reagents, heat, and electric current, the normal physiological structure of the skin is damaged, and the protective effect of the body weakens or disappears. The process of skin wound healing can be divided into four continuous stages, namely, hemostasis, inflammation, proliferation, and remodeling, as shown in Figure 2 [28]. When injury occurs, the blood vessels in the damaged area rupture, the subcutaneous matrix in the blood vessels is exposed, and the glycoproteins on the platelet membrane bind with the collagen of the basement membrane, making the platelets adhere to and be activated [29]. Activated platelets release endogenous ADP and TXA2, which in turn promote irreversible aggregation of platelets, resulting in platelet thrombosis [30]. In addition to hemostasis, platelets can also release cytokines and growth factors such as interleukin (IL)-1β, tumor necrosis factor (TNF)-α, fibroblast growth factor (FGF), and platelet derivation growth factor (PDGF) together with the cells in the injured area and promote the migration of neutrophils and macrophages in the wound area [31]. In the early stage of inflammation, neutrophils first accumulate in the wound area and phagocytose and release reactive oxygen species (ROS), antimicrobial peptides, proteolytic enzymes, and other substances to remove necrotic tissues and pathogens [3,32]. Mast cells and macrophages then arrive at the damaged area to remove remaining cell debris and neutrophils [33].

Figure 2

The stages of wound repair and their major cellular components [28]. The process of skin wound healing can be divided into four continuous stages, namely, hemostasis, inflammation, proliferation, and remodeling.

At the later stage of inflammation, macrophages develop an anti-inflammatory, profibrotic phenotype that promotes the proliferation of fibroblasts, keratinocytes, and endothelial cells by secreting a variety of growth factors and cytokines [34,35]. Under the action of FGF, fibroblasts provide the skeleton structure for cell migration, proliferation, and growth to the wound area through the synthesis of type III collagen, proteoglycan, and fibronectin [36]. At the same time, endothelial cells at the wound site proliferate and migrate under the stimulation of factors such as VEGF and angiopoietin, forming a new capillary network, which is necessary for tissue regeneration and granulation tissue formation and transportation in the injured area [37]. Keratinocytes from near the injured area migrate to the injured area under the stimulation of growth factors and promote the epithelialization of the epidermal tissue through proliferation and differentiation [38]. When wound repair enters the remodeling stage, the remaining temporarily transplanted cells in the damaged area are apoptotic, and fibroblasts synthesize type I collagen, elastin, and other extracellular matrices [39]. Type III collagen in the extracellular matrix (ECM) of the injured area was gradually replaced by type I collagen, the collagen fibers rearranged into a lattice-like structure, and the skin tissue in the injured area regained strength and toughness similar to normal tissue [40].

2.3 Factors affecting wound healing

Wound healing involves many factors, and it takes a long time from the occurrence of injury to the completion of remodeling. Figure 3 illustrates common factors that affect skin healing. The factors affecting wound healing are mainly divided into endogenous factors and exogenous factors. Age is one of the endogenous factors affecting wound healing and repair. With increasing age, the thickness of the cuticle decreases, the activity of macrophages decreases, and the release of growth factors and cytokines decreases [41]. These factors lead to a prolonged proliferation period and an increase in the time required for wound healing. In addition, the body’s hormone level also has a certain effect on wound healing. For example, estrogen can promote the re-epithelialization of keratinocytes and the angiogenesis of endothelial cells in the process of wound healing, which has a certain promoting effect on wound healing [42]. Glucocorticoids and cortisol can inhibit the body’s own immune inflammatory response, which restricts the migration and aggregation of white blood cells to the injured area during inflammation [43]. The healing process can also be influenced by genetic factors. Studies have shown that keloids are a wound repair abnormality with a strong genetic tendency. In such people, excessive collagen deposition leads to the formation of a large number of fibrous scars on the wound surface, resulting in scar healing [44].

Figure 3

Factors affecting wound healing.

In addition to the influence of the body’s own factors on wound healing, exogenous factors are also the main factors affecting wound healing. Local infection after skin injury is an important factor affecting wound healing. When the wound is infected by S. aureus or other Streptococcus, a large number of white blood cells migrate and gather to the injured area, and the bacteria are removed by phagocytosis of white blood cells. The process of removing bacteria will cause the release of bacterial endotoxin, resulting in necrosis and inflammation of the local tissues [2]. In addition, due to the increase in proinflammatory cytokines, the number of released growth factors decreased, which inhibited the growth of cells in the proliferation stage, resulting in delayed wound healing or unhealing [45]. The nutritional status of the body is another important factor affecting wound healing. In the process of wound healing, a variety of trace elements, vitamins, fatty acids, and proteins are necessary, and the loss of any nutrients will cause the delay of wound healing [46,47,48]. In addition, many chronic diseases affect wound healing by interfering with platelet activation, the inflammatory response, epithelial formation, and matrix remodeling. Diabetes, for example, affects the migration and activation of white blood cells to the damaged area and increases the release of proinflammatory cytokines, leading to chronic inflammation [49]. Diabetes also causes abnormalities in microvascular formation that inhibit keratinocyte- and fibroblast-mediated epithelial reformation and ECM remodeling [50]. Drugs and unhealthy lifestyle habits such as smoking and drinking can also affect wound healing. Smoking can increase the aggregation and adhesion of platelets, raising the risk of blood clots [51]. Alcohol inhibits the body’s immune response and reduces collagen formation and matrix metalloproteinases (MMP) concentrations [52].

3.1 Role of PDGF

PDGF is a basic protein stored in the platelet α particles. It is a dimer glycoprotein formed by two peptide chains linked by disulfide bonds [53]. According to the difference of two peptide chains, PDGF can be divided into 5 types: PDGF–AA, PDGF–BB, PDGF–CC, PDGF–DD, and PDGF–AB [54]. A variety of cells, such as macrophages, fibroblasts, and endothelial cells, can secrete and synthesize PDGF [55,56]. PDGF is involved in the whole process of wound healing and plays an important role in promoting wound healing. In the inflammatory stage, PDGF can promote the migration of neutrophils and macrophages to the injured site [57]. In addition, PDGF can stimulate macrophages to secrete synthetic TGF-β, further enhancing the chemotactic effect on inflammatory cells [55]. PDGF can also promote the proliferation and differentiation of fibroblasts and promote the synthesis of ECM of fibroblasts [53,58]. PDGF can promote the migration and proliferation of vascular endothelial cells at the injury site and promote the formation of new blood vessels [59]. During remodeling, PDGF can stimulate the differentiation of fibroblasts into myofibroblasts, upregulate the expression, synthesize MMP, and regulate the degradation and remodeling of ECM [60].

3.2 Role of FGF

There are 23 types of FGF families, among which FGF–2, FGF–7, and FGF–10 are related to wound repair. FGF is produced by keratinocytes, fibroblasts, endothelial cells, smooth muscle cells, chondrocytes, and mast cells. When injury occurs, the concentration of FGF increases sharply in the injury area and stimulates the migration of macrophages, white blood cells, and neutrophils to the injury area [61]. FGF can directly promote the proliferation and differentiation of fibroblasts, vascular endothelial cells, and smooth muscle cells, and promote the formation of granulation tissues [62,63]. In addition, FGF can also promote epithelial mesenchymal transformation by binding with TGF–β1 [64]. FGF can also promote the secretion of EGF, promote the proliferation of epidermal cells, regulate the differentiation of fibroblasts to myofibroblasts, and accelerate wound contraction activity, which is conducive to early wound coverage [62]. FGF can inhibit terminal differentiation of myofibroblasts and reduce glial scar hyperplasia during wound remodeling [65]. bFGF can also reduce collagen synthesis and scar formation by increasing procollagen mRNA degradation and inhibiting mRNA transcription [66].

3.3 Role of VEGF

VEGF is a family that includes VEGF–A, VEGF–B, VEGF–C, VEGF–D, VEGF–E, and placental growth factor (PGF). When injury occurs, activated platelets release VEGF to improve the lumen permeability of residual vessels, which is conducive to the migration and aggregation of inflammatory cells to the injured area [67–69]. VEGF promotes the proliferation, migration, and differentiation of vascular endothelial cells and the formation and functionalization of new capillaries during the proliferation phase [70,71]. In addition, VEGF can also promote lymphatic vessel formation by activating VEGFR–2 [72]. VEGF is also associated with scar tissue formation during remodeling. Wilgus T. A. found that injection of recombinant VEGF into wounds can increase scar formation [73].

3.4 Role of EGF

In the EGF family, EGF, heparin-binding EGF (HB–EGF), and TGF–α are mainly related to wound repair [74,75]. Platelet granules can release EGF during hemostasis, and epithelial cells migrate from the wound to the wound surface [76]. In the inflammatory stage, EGF can chemotaxis macrophages and fibroblasts to the wound surface and promote the inflammatory response [77]. EGF can stimulate the proliferation and division of endothelial cells and epithelial cells so that the epithelial cells migrate to the surface and promote wound shrinkage [78]. During remodeling, EGF inhibits the inflammatory response, reduces TGF–β1 expression, mediates collagen formation, and reduces scar tissue formation [79].

3.5 Role of TGF–β

TGF–β mainly includes three types: TGF–β1, TGF–β2, and TGF–β3, among which TGF–β1 plays a dominant role in wound healing [80]. TGF–β1 can promote the aggregation of inflammatory cells such as neutrophils and macrophages during inflammation [81,82]. TGF–β1 promotes the synthesis of collagen, elastic fibers, and fibronectin in fibroblasts and the formation of granulation tissue [83]. TGF–β1 regulates epithelial mesenchymal transformation and promotes cell migration, angiogenesis, and fibrous tissue proliferation [84]. TGF–β1 is involved in the synthesis of type I and III collagen during wound remodeling, inhibits the synthesis of MMP, reduces the degradation of collagen, and promotes the formation of scar tissue [85,86]. TGF–β1 and TGF–β2 had similar effects on wound healing, while TGF–β3 had opposite effects. TGF–β3 inhibits inflammation and protein synthesis, reducing scar formation by inhibiting collagen formation [87,88].

3.6 Role of NGF

In addition to playing a role in the nervous system, NGF also plays an important role in wound healing. NGF can promote the proliferation and migration of fibroblasts and keratinocytes and improve the synthesis level of ECM [89]. NGF also promotes angiogenesis and myofibroblast differentiation and promotes wound contraction by promoting collagen synthesis [90].

4.1 Chitosan

Chitosan is the product of the natural polysaccharide chitin removing part of the acetyl group, which has good biocompatibility and degradability. Meanwhile, chitosan also has antibacterial and antitumor properties, which make it very suitable for the preparation of wound dressings [94]. Chitosan can also participate in wound healing. Chitosan can enhance the expression of GPIIB–IIIA on the platelet membrane and promote platelet adhesion to the vascular wall [96]. In addition, positively charged chitosan can also interact with a large number of negatively charged substances on the surface of activated platelets to promote platelet aggregation [97]. Through the above mechanisms, chitosan can promote wound hemostasis, induce RBC aggregation, and inhibit fibrinolysis. In addition, chitosan can increase the secretion of IL-8 in fibroblasts, accelerate the inflammatory process, and promote angiogenesis [98,99]. Chitosan with a low molecular weight can also promote the proliferation of fibroblasts, enhance the synthesis and secretion of collagen, and promote the growth of granulation tissue [98]. Chitosan also has an inhibitory effect on bacterial growth, which may be achieved by destroying the bacterial cell wall and membrane, chelating with metal cations, interacting with intracellular targets, and deposition on the bacterial surface [100–102]. Table 2 listed some combination of chitosan and growth factors in skin wound healing.

The hydrogel form of chitosan applied in wound repair can retain the properties of chitosan itself and has better flexibility, moisture retention, adsorption capacity, and air permeability. On this basis, hydrogel-like chitosan can be further modified and combined with growth factors to construct a more suitable composite dressing for wound healing. Yao et al. prepared a chitosan-PAA-pHEMA hydrogel (CSPAH) by UV polymerization grafting poly(acrylic acid) (PAA) and poly(hydroxyethyl methacrylate) (pHEMA) and loaded EGF into the hydrogel to promote wound healing. The results showed that the hydrogel had good thrombosis ability and antibacterial activity. At the same time, CSPAH can effectively delay the release of EGF, which can be as long as 7 days or even longer, enhancing the activity of EGF in the wound site. Animal experimental results shown in Figure 4 illustrated that CSPAH loaded with EGF could significantly improve the wound healing rate. HE staining results showed that epithelial migration, inflammatory cell migration and neovascularization were more obvious in the EGF group [103]. Choi and Yoo prepared a Pluronic/chitosan hydrogel containing EGF and showed that chitooligosaccharide and rhEGF in the hydrogel significantly enhanced epidermal differentiation during wound healing [104]. Mariia et al. prepared a cellulose nanocrystal (CNC)-enhanced chitosan hydrogel (CS-U-CNC-EGF) loaded with EGF by the freeze-drying method. The results showed that granulation tissue formation in the wound area was significantly accelerated, collagen deposition was increased, and the wound healing rate was significantly improved in the treatment group [105].

Figure 4

Representative images and HE staining results of wound surface healing over time in CSPAH, CSPAH-incorporating EGF, commercial dressings, and the control at days 10 and 20. HEMA: hydroxyethyl methacrylate; CSPAH: chitosan-PAA-pHEMA; EGF: epidermal growth factor; PAA: poly(acrylic acid) [103].

In addition to EGF, chitosan can also be combined with VEGF and PDGF to improve wound angiogenesis. Yang et al. prepared visible light curable GC hydrogels capable of continuously releasing VEGF and PDGF to promote angiogenesis in the injured area, accelerate wound reepithelialization, and promote wound healing by continuously releasing VEGF and PDGF [110]. Huang et al. prepared a tunable sequential drug delivery system based on chitosan/hyaluronic acid (HA) and PLGA microspheres, which promoted angiogenesis by carrying VEGF and vancomycin, reduced the inflammatory response caused by the bacterial growth, and promoted wound healing [111]. Xu et al. prepared a thermosensitive chitosan hydrogel containing PLGA microspheres and loaded it with PDGF for the treatment of skin injury. The results showed that hydrogels containing PDGF could promote the formation of granulation tissue and collagen, reduce the level of autophagy and promote wound healing. Fibroblasts play an important role in wound healing and proliferation [112]. Obara and others used ultraviolet radiation crosslinking chitosan (Az-CH-LA) in aqueous solution containing FGF, prepared flexible water insoluble gel, and applied water gel to diabetic mouse skin defects. The results showed that hydrogels containing FGF raised wound granulation tissue, blood capillaries, and epithelization levels, significantly induced wound contraction, and accelerated wound closure [113].

4.2 Alginate

Alginate is a kind of natural anionic polysaccharide compound extracted from the cell wall of brown algae and some bacterial strains, such as nitrogen-fixing bacteria and Pseudomonas. They are long chain polymers composed of 1,4-β-crosslinked d-mannuronic acid and 1,4-α-crosslinked guluronic acid [116]. Alginate from algae is insoluble in water, but its sodium salts are solvable. Adding Ca²⁺ to alginate solution can precipitate alginate in a gel state [117]. Alginate can be divided into the M region (mannuronic acid-rich region), G region (guluronic acid-rich region), and MG region (rich in both uronic acids) [118]. The proportion of MG in alginate from different sources is quite different, and the different proportion of MG determines the different properties of alginate [119]. For example, alginate gels with high G content have greater hardness, while alginate saltwater gels with high M content are softer and more elastic [120]. Alginate is nondegradable in mammals due to the absence of alginase [121]. Some studies believe that alginate has certain immunogenicity, which can cause toxicity or immune response in the body, but this is not caused by alginate itself. Heavy metals and impurities in the extraction and preparation of alginate are the causes of this problem [122,123]. Alginate has certain antibacterial and antiviral properties [124,125]. In addition, alginate can promote hemostasis by activating platelets and promoting thrombin clot formation [126]. Dry alginate is not suitable for wound surfaces because dry alginate is highly absorbent and can cause wound adhesion [127]. However, the alginate hydrogel showed good water retention, which is conducive to keeping the wound environment moist and promoting wound healing [128]. At the same time, alginate hydrogel can ensure good adhesion to the wound and will not adhere to the wound tissue, so the removal will not cause secondary injury to the wound [129]. These properties make alginate hydrogels have the potential to be used in wound dressings. Table 3 lists some combination of alginate and growth factors in skin wound healing.

Since alginate brine gel itself cannot promote cell migration and cell adhesion, it has good application prospects to load growth factors into alginate hydrogels or modify hydrogels with other substances to obtain a better effect. As shown in Figure 5, Zhu et al. constructed a zinc-doped bioactive glass (ZBG)/succinyl chitosan (SCS)/oxidized alginate (OAL) composite hydrogel (GEL–ZBG) and embedded EGF into the hydrogel to improve cell proliferation and tissue remodeling in the wound bed. The results showed that the hydrogel had good antibacterial properties and that the addition of EGF improved the early healing speed of wounds. Histological results showed that GEL–ZBG loaded with EGF could promote the migration and enrichment of fibroblasts to the wound area, promote the growth of granulation tissue and neovascularization, and increase the deposition level of collagen and collagen fibers [130]. Liu et al. prepared a HAAL hydrogel composed of HA and sodium alginate by ion and covalent crosslinking, loaded it with EGF in the hydrogel, and found that the EGF-loaded HAAL hydrogel showed enhanced cell proliferation and adhesion [131]. Rezaei et al. constructed a dual drug delivery system in which EGF and pH-sensitive silk fibroin/alginate nanoparticles were embedded in PNIPAM hydrogel for the treatment of severe infectious burn wounds. The results showed that the system could significantly reduce the incidence of wound infection and promote the proliferation and growth of fibroblasts [132].

Figure 5

(a and b) TEM image for BG and ZBG. SEM images of (c) Gel and (d) Gel–ZBG composite hydrogels. (e) Schematic representation of the composite hydrogels as wound dressing for wound closure of SD rats [128].

To rapidly control inflammation and accelerate wound healing, Cui et al. developed a two-drug delivery system in which micron alginate fibers, FGF, and penicillin were encapsulated in instant self-assembling peptide hydrogels. The results showed that this two-drug delivery system could continuously release FGF and suddenly release antibiotics. Animal studies have shown that hydrogels can quickly reduce wound inflammation and speed up wound healing [133]. Azarpira et al. mixed alginate oxide containing VEGF and bFGF into acellular collagen-alginate complex hydrogels and found that it could significantly improve blood vessel generation and promote endothelial cell migration and growth [134]. Chou et al. constructed a thermosensitive and reversible IPN hydrogel by treating alginate with Pluronic F127 (PF127), which was equipped with VEGF to further promote tissue regeneration and wound healing rate [135]. In recent years, PDGF and alginate hydrogels have been combined to promote wound healing. Babavalian et al. prepared alginate sulfate hydrogels by photocrosslinking and applied them in the treatment of rat skin injury by loading PDGF. The results showed that hydrogels containing PDGF could significantly promote angiogenesis and fibroblast proliferation and promote wound recovery [136]. Zhang et al. synthesized an injectable hydrogel using sodium alginate and glucan as a delivery system to deliver PDGF and BMSCs simultaneously in the wound to promote wound healing. The results showed that the PDGF/SA/Dex hydrogel could promote angiogenesis by activating the PDGF-BB/PDGFR-β-mediated PI3K/Akt/eNOS pathway, thus accelerating BMSC-mediated skin wound healing [137].

4.3 Collagen

Collagen is the most abundant protein in mammals, accounting for 25–30% of the total protein. Collagen is widely found in various connective tissues. Approximately 29 kinds of collagen have been discovered thus far, among which type I collagen is the most important type used [120]. Collagen is a major component of the ECM and plays a dominant role in maintaining the biological and structural integrity of the ECM. Collagen is resistant to protease hydrolysis and has good biocompatibility as a protein produced from the organism itself [138]. Although the use of collagen between species may cause an immune response, this problem can be avoided to some extent by heterologous expression in animals or in E. coli [139,140]. Collagen stimulates cell migration and contributes to the development of new tissue. Collagen stimulates and recruits corresponding cell types, such as macrophages and fibroblasts, according to the healing cascade order, promoting cell aggregation in the wound area [141]. Due to the chemotactic effect of collagen on fibroblasts, exogenous collagen can stimulate the aggregation of fibroblasts in the wound and the deposition of endogenous collagen, forming a local healing environment. Collagen also has a good moisturizing effect and adhesion to the wound and is easy to replace without damaging the wound surface. Collagen can also form 3D structures with high tensile strength and stability through crosslinking and self-aggregation [142]. Such excellent mechanical properties make collagen suitable as a matrix material for wound dressings. Table 4 gives the summary of collagen loaded with growth factors in wound healing.

Collagen is widely used in tissue regeneration and tissue repair due to its excellent biological properties. At present, commercial collagen products such as gelatin sponges have been used in tissue repair and other fields. Using other materials to modify collagen and load factors to make it more suitable for wound repair is a research hotspot.Thones et al. constructed a HA/collagen hydrogel containing HA sulfate to promote wound healing through continuous release of heparin binding EGF-like growth factor, as shown in Figure 6. In vitro experiments showed that hydrogels containing HB–EGF induced keratinocyte migration, EGFR signaling, and HGF expression in dermal fibroblasts. In a porcine skin organ culture model, it was found that the growth of the epithelial tip could be significantly improved [143]. Kondo et al. constructed a HA collagen sponge dressing made of HA and collagen of different molecular weights and added EGF to it. They found that EGF-loaded dressings significantly increased the release of VEGF and HGF in fibroblasts, promoting wound healing and granulation tissue formation [144]. Niiyama and Kuroyanagi constructed a new wound dressing composed of HA and collagen loaded with vitamin C and EGF and found that the new wound dressing promoted granulation tissue and angiogenesis and accelerated epithelial formation in SD rats and diabetic mice [145]. Cheng et al. prepared a hybrid lyophilized dressing composed of EGF and recombinant human collagen and found that this dressing significantly enhanced fibroblast proliferation adhesion and migration to the wound. Immunohistochemical results showed that the wound healing rate, epithelial regeneration, and orderly arrangement and deposition of collagen were significantly accelerated in rats [146].

Figure 6

Schematic diagram of HE-EGF-combined hyaluronan/collagen hydrogels as wound dressings for wound healing [143].

Wang et al. constructed a chitosan crosslinked collagen sponge containing FGF and applied it in diabetic rat wound healing. The results showed that the wound healing time was shortened, collagen generation was the fastest, and TGF expression was the earliest, which significantly promoted the wound healing of diabetes [115]. Pandit et al. treated full-thickness skin defects in rabbits by delivering FGF via collagen scaffolds, and the results showed that FGF collagen scaffolds increased angiogenesis and epithelial formation and promoted wound healing [147]. Chakrabarti et al. found that FGF-loaded collagen-AgSD hydrogels accelerated burn healing in animal models, alleviated burn inflammation by activating ERK and TRK pathways, and promoted fibroblast proliferation [148]. Juzith et al. developed a novel complex hydrogel that combines collagen, chitosan, and PDGF, which increased the levels of antioxidants and lipid peroxides, increased collagen synthesis and granulation tissue production, and significantly shortened the wound healing time in the treated group than in the control group [57]. Lin et al. constructed a collagen scaffold with PDGF targeted binding for wound repair and found that such collagen targeted binding PDGF had a longer release cycle, significantly increased fibroblast proliferation and accelerated wound granulation tissue formation [58]. Tateshita et al. applied a collagen matrix containing TGF-β1 to the surface of wounds, and the results showed that the application of TGF-β1-treated collagen matrix is effective for preventing contraction producing the so-called “neodermis” in treating a delayed healing type model and may be highly beneficial for treating chronic wounds [149].

4.4 Fibrin

Fibrin is a natural polymer formed during wound healing and is formed by cleavage of fibrinogen by serine protease thrombin during wound healing and hemostasis. Fibrin, along with platelets, is involved in wound hemostasis and provides the basis for subsequent repair. During the proliferative stage, fibrin and fibronectin can promote the proliferation and migration of fibroblasts to the wound site and promote the formation and deposition of collagen [150]. As the wound repair process progresses, fibrin clots eventually become deabsorbed by plasmin and participate in the formation of ECM. Fibrin can also serve as a reservoir of growth factors and cytokines during wound repair, and a variety of cytokines, such as EGF, FGF, and PDGF, can be found in fibrin clots [151]. Growth factors coated with fibrin can avoid rapid release and degradation in the body and prolong the action time of growth factors in wound repair. This property makes fibrin particularly suitable for the delivery of growth factors and cells during wound healing and has the potential to become a wound dressing [152]. However, the rapid degradation of fibrin has always been the main problem restricting its clinical transformation. Because fibrin can be easily degraded by plasmin in the body, its ability to maintain growth factors and cells is significantly reduced [153]. To solve this problem, there are a variety of modification methods for fibrin, such as adding heparin to fibrin and combining it with other biological matrix materials [154]. These methods prolong the degradation cycle of fibrin to a certain extent and increase the effect and feasibility of fibrin in clinical applications.

The schematic diagram made by Rahman et al. in Figure 7 constructed a platelet-rich and fibrin-rich hydrogel from expired platelet components that contained physiological concentrations of TGF–β1, PDGF, IGF–1, FGF, and EGF. Application of the hydrogel to skin wounds in SKH–1 mice significantly enhanced angiogenesis in the wound area. High levels of IL–6 and KC also lasted longer in treated wounds than in untreated wounds. The epithelialization and healing of the wound area were significantly accelerated [156]. Certelli et al. constructed a system capable of instantaneous delivery of engineered VEGF and PDGF–BB in fibrin hydrogels. On applying this system to a diabetic mouse model of skin injury, it was found that this combination of treatments could establish stable angiogenesis in the injured area and promote wound growth and healing [157]. Tan et al. constructed a 3D fibrin gel embedded with BMSCs and VEGF, and BMSCs in the gel differentiated into CD31 + and vWF + ECs induced by VEGF. Animal studies have shown that fibrin gel can stop bleeding quickly without secondary bleeding. Fibrin gels containing BMSCs have shown the potential to promote neovascularization and wound healing [155].

Figure 7

Schematic diagram of biofunctionalized fibrin gel co-embedded with BMSCs and VEGF for accelerating skin injury repair [155].

4.5 Gelatin

Gelatin has homology with collagen and is the product of partial hydrolysis of collagen. The structure of gelatin is similar to that of the ECM, and it can simulate the structure and morphology of normal tissues well, which is conducive to cell migration and growth [158]. Gelatin has temperature reversibility. By adjusting the temperature, hydrogel materials with high tensile strength, dermal structure, and good permeability and water can be prepared [159]. Gelatin molecules also contain a large number of functional groups, which can be chemically modified or modified to carry different drugs or growth factors into the molecules to improve their performance or therapeutic effect. In addition, as a hydrolysate of collagen, gelatin has significantly lower immunogenicity and better histocompatibility than collagen [159]. However, gelatin itself is a hydrophilic protein that usually needs to be modified to increase its mechanical properties and stability when it is used in the preparation of wound dressings or hydrogels [6]. In addition, gelatin itself lacks certain antibacterial properties, and simple gelatin easily causes bacteria to grow on its surface, so in the application, it should be combined with antibacterial substances to enhance its antibacterial properties [160].

Augustine et al. constructed a poly(3-hydroxybutyrate-co-3-hydroxyvalerate)/gelatin methacryloyl hybrid patch using GelMA and PHBV, in which EGF was loaded for the treatment of diabetic wound healing. GelMA shows good moisture retention and water absorption, while the PHBV film provides the hydrogel with the right mechanical strength. EGF-loaded patches can promote cell migration, cell proliferation, and angiogenesis and thereby, the rapid healing of diabetic wounds [161] (Figure 8). Huang et al. constructed a functional bilayer skin substitute for wound repair using tissue-engineered ECM and gelatin hydrogel doped with microspheres. It was found that the bionic skin improved healing in the wound area, showing early epithelial regeneration in the wound area [162]. Jin et al. prepared a foam dressing for skin regeneration using gelatin, HA, and carboxymethyl chitosan containing FGF and found that wound healing was significantly accelerated in animal experiments [163]. Wan et al. constructed a 3D double-layer hydrogel scaffold loaded with PDGF and Ag, which can significantly reduce wound surface infection, results showed great potential for promoting diabetic wound healing and fighting bacterial infection [164].

Figure 8

Schematic diagram of growth factor-loaded in situ photocrosslinkable poly(3-hydroxybutyrate-co-3-hydroxyvalerate)/gelatin methacryloyl hybrid patch for diabetic wound healing [159].