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Clinical exam progression of LiQD Cornea in Göttingen mini-pigs. (A) Pachymetry showing corneal thickness measured by OCT, showing no significant differences in thickness compared to controls. There was a normal increase in corneal thickness in unoperated controls as the pigs matured. (B) Intraocular pressures were similar in all three groups, showing a slight overall increase over the normal aging process of the pigs. (C) Central corneal haze measured using a modified McDonald-Shadduck scoring system on a scale from 0 to 4. An increase of haze corresponds to the period of in-growth of stromal cells into the cell-free implants. By 12 months after operation, the cells appeared to have attained quiescence. (D) Corneal neovascularization was seen in the LiQD Cornea, mainly from the animal that sustained an unintended perforation. (E) Corneal blink response measured by Cochet-Bonnet esthesiometry showed no significant differences among the three groups. (F) Corneal nerve density in the LiQD Cornea group was significantly lower than the unoperated corneas during months 3 to 9 after operation when the severed nerves were regenerating. (G) Schirmer's tear test showed similar responses in all three groups tested. (H) Expression of high-molecular weight collagens (HMW, , and ), type V collagen, and type I collagen (1 and 2) in the central portion of the cornea. Figures (A), (B), and (E) to (H) were assessed using a mixed-effects model with a Tukey post hoc test for multiple comparisons. Figures (C) to (D) were analyzed using a Mann-Whitney U test for ordinal data. *P ≤ 0.05 for LiQD Cornea to unoperated, †P ≤ 0.05 for LiQD Cornea to syngeneic graft, and ‡P ≤ 0.05 syngeneic graft to unoperated. All data are plotted as mean ± SEM or mean with individual values.
Source publication
Transplantation with donor corneas is the mainstay for treating corneal blindness, but a severe worldwide shortage necessitates the development of other treatment options. Corneal perforation from infection or inflammation is sealed with cyanoacrylate glue. However, the resulting cytotoxicity requires transplantation. LiQD Cornea is an alternative...
Contexts in source publication
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... Cornea was applied as for the rabbits. Figure S2A shows the progress of repair and regeneration of all pigs receiving the LiQD Cornea compared to syngeneic grafts and healthy unoperated controls. At 12 months, the application of LiQD Cornea was successful in all pigs (Fig. 1F), although in all cases, the surgeon applied the LiQD Cornea at least twice, removing the first material before reapplication to achieve the desired curvature. ...
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... LiQD Cornea at least twice, removing the first material before reapplication to achieve the desired curvature. One pig, which received four attempts at LiQD Cornea application, underwent full corneal perforation and was given a suture to bridge the unintended gape. Postsurgical optical coherence tomography (OCT) of the LiQD Cornea application ( fig. S2B) showed that although the initial LiQD Cornea fills were imperfect, the anterior corneal surfaces of all four pigs were smooth and followed the contours of the host tissue by 3 months after operation. These results also show that an easy-to-use point-of-care (POC) delivery device ( fig. S3) is merited for future clinical ...
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... follow-up showed that at 1-month follow-up, all pigs had successfully reepithelialized. At 3 months after surgery, pachymetric analyses showed that the standard corneal thickness was restored in LiQD Cornea animals ( Fig. 3A and fig. S2B). Intraocular pressure was normal at all postsurgical exams, indicating that the LiQD Cornea successfully sealed the surgical site (Fig. 2B). The LiQD Cornea pigs showed more significant haze and neovascularization than syngeneic grafts at all postsurgical time points, but haze was reduced in three of four animals at 12 months after ...
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... at 1-month follow-up, all pigs had successfully reepithelialized. At 3 months after surgery, pachymetric analyses showed that the standard corneal thickness was restored in LiQD Cornea animals ( Fig. 3A and fig. S2B). Intraocular pressure was normal at all postsurgical exams, indicating that the LiQD Cornea successfully sealed the surgical site (Fig. 2B). The LiQD Cornea pigs showed more significant haze and neovascularization than syngeneic grafts at all postsurgical time points, but haze was reduced in three of four animals at 12 months after operation (Fig. 2, C and D). The fourth pig had poor surgical results with iritis and formation of peripheral anterior synechiae (attachment of ...
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... S2B). Intraocular pressure was normal at all postsurgical exams, indicating that the LiQD Cornea successfully sealed the surgical site (Fig. 2B). The LiQD Cornea pigs showed more significant haze and neovascularization than syngeneic grafts at all postsurgical time points, but haze was reduced in three of four animals at 12 months after operation (Fig. 2, C and D). The fourth pig had poor surgical results with iritis and formation of peripheral anterior synechiae (attachment of the iris to the cornea) and infiltration of a large blood vessel into the surgical site, resulting in a hazy cornea at 12 months after operation. This pig had inadvertently received a full-thickness corneal perforation ...
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... was reattached with a suture that trekked in a large blood vessel. This pig, nevertheless, showed full regeneration of corneal tissue and nerves. Esthesiometry performed to determine touch sensitivity showed the restoration of the corneal blink response in all operated corneas, indicating the presence of regenerated nerves within the graft site (Fig. 2E). Analysis of the density of corneal nerves over time showed that by 12 months after operation, there were no statistically significant differences in the nerve density (Fig. 2F) between the LiQD Table 1. Optical, physical, and mechanical properties of LiQD Cornea ...
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... touch sensitivity showed the restoration of the corneal blink response in all operated corneas, indicating the presence of regenerated nerves within the graft site (Fig. 2E). Analysis of the density of corneal nerves over time showed that by 12 months after operation, there were no statistically significant differences in the nerve density (Fig. 2F) between the LiQD Table 1. Optical, physical, and mechanical properties of LiQD Cornea ...
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... (3012.8 ± 1613.7 m/mm 2 ), syngeneic (2205.3 ± 1162.4 m/mm 2 ), and unoperated control (4800.4 ± 1964.9 m/mm 2 ). However, it is clear that the unoperated controls had a higher nerve density. There were no marked differences in tear production in the three treatment groups at any time point, as indicated by Schirmer's test (Fig. ...
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... content analysis of the central cornea demonstrated significantly lower levels of high-molecular weight, -, -, 1(V)-, and 1(I)-type collagen in the LiQD Cornea pigs, as compared to the syngeneic grafts and unoperated eyes ( Fig. 2H and table ...
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... in the healthy unoperated control corneas (Fig. 5F). Nerves in the syngeneic grafts were not as well defined in their configuration (Fig. 5E). From 3 to 9 months, reflective keratocytes indicative of in-growing cells were seen within the matrix. The presence of reflective cells corresponded with the increased haze seen by slit lamp biomicroscopy (Fig. 2C). At 12 months, keratocytes grew into the cell-free matrix to reconstitute the stroma (Fig. 5G). Most of these keratocytes were not reflective and resemble keratocytes in the syngeneic grafts (Fig. 5H) and untreated controls (Fig. 5I). The decrease in reflectivity likely corresponds to the decrease in haze in Fig. ...
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... by slit lamp biomicroscopy (Fig. 2C). At 12 months, keratocytes grew into the cell-free matrix to reconstitute the stroma (Fig. 5G). Most of these keratocytes were not reflective and resemble keratocytes in the syngeneic grafts (Fig. 5H) and untreated controls (Fig. 5I). The decrease in reflectivity likely corresponds to the decrease in haze in Fig. ...
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... pig study confirmed that LiQD Cornea allowed regeneration of the corneal epithelium, stroma, and nerves. Even if the material does not achieve the desired perfectly smooth surface directly after application, OCT results showed that the corneal thickness and curvature were restored to those matching the syngeneic grafts and unoperated controls (fig. S2B). The primary difference in the clinical performance of the LiQD Cornea and the syngeneic grafts was increased haze in the surgical site between 3 to 6 months after operation, during the period of rapid keratocyte in-growth into the cell-free matrix. Syngeneic grafts were already populated with donor cells, so no rapid in-growth of ...
Citations
... Different hydrogels-with or without cells-have been demonstrated in several studies to be an effective option for stromal replacement using donor tissue. [33][34][35][36]. Exosomes [37], anti-TGF- [6,7,38], anti-PDGF [7,39,40], and HGF [41,42] have all been shown to play a role in either preventing or reversing corneal scars. ...
Corneal opacification or scarring is one of the leading causes of blindness worldwide. Human limbus-derived stromal/mesenchymal stem cells (hLMSCs) have the potential of clearing corneal scarring. In the current preclinical studies, we aimed to determine their ability to heal the scarred corneas, in a murine model of corneal scar, and examined their ocular and systemic toxicity after topical administration to rabbit eyes. The hLMSCs were derived from human donor corneas and were cultivated in a clean room facility in compliance with the current good manufacturing practices (cGMP). Before the administration, the hLMSCs were analyzed for their characteristic properties including immunostaining, and were further subjected to sterility and stability analysis. The corneas (right eye) of C57BL/6 mice (n = 56) were stripped of their central epithelium and superficial anterior stroma using a rotary burr (Alger Brush® II). Few mice were left untreated (n = 8), while few (n = 24) were treated immediately with hLMSCs after debridement (prophylaxis group). The rest (n = 24, scar group) were allowed to develop corneal scarring for 2 weeks and then treated with hLMSCs. In both groups, the treatment modalities included encapsulated (En+) and non-encapsulated (En−) hLMSCs and sham (vehicle) treatment. The follow-up (4 weeks) after the treatment or debridement included clinical photography, fluorescein staining, and optical coherence tomography at regular intervals. All the images and scans were analyzed using ImageJ software to assess the changes in corneal haze, scar area, and the reflectivity ratio of the epithelium to the stroma. The scar area and the scar intensity were found to be decreased in the groups that received hLMSCs. The reflectivity of the stroma was found to be normalized to the baseline levels before the debridement in the eyes that were treated with hLMSCs, relative to the untreated. In the safety study, the central corneas of the left eye of 18 New Zealand rabbits were scraped with a needle and then treated with En+ hLMSCs, En− hLMSCs, and the sham (n = 6 each). Rabbits were then followed up for 4 weeks, during which blood and tear samples were collected at regular intervals. These rabbits were then assessed for changes in the quantities of inflammatory markers (TNF-α, IL-6, and IgE) in the sera and tears, changes in the ocular surface observations such as intraocular pressure (IOP), and the hematological and clinical chemistry parameters. Four weeks later, the rabbits were euthanized and examined histopathologically. No significant changes in conjunctival congestion, corneal clarity, or IOP were noticed during the ophthalmic examination. The level of inflammatory molecules (TNF-α and IL-6 TNF-α) and the hematological parameters were similar in all groups without any significant changes. Histological examination of the internal organs and ocular tissues did not reveal any abnormalities. The results of these studies summarize that the En+ and En− hLMSCs are not harmful to the recipient and potentially restore the transparency of debrided or scarred corneas, indicating that hLMSCs can be assessed for clinical use in humans.
... Refractive index 1.373, similar to human cornea [66,67]. ...
... The biofabricated corneal tissue should withstand the equivalent (minimum) or higher (preferred) liquid pressure than the IOP of native human cornea, which ranges between 11-21 mmHg or ~2.5 kPa [67]. ...
... Despite genes, stem cells, gels and drops have been developed, human donor corneal transplant is the most effective treatment for corneal blindness and restoring patients' vision (with a cure rate of more than 80%) over a century. In 116 countries, approximately 185,000 corneal transplants are performed each year, yet 13 million people are still waiting for corneal transplants [130][131][132]. Therefore, there is an urgent need to find novel strategy to promote corneal regeneration. ...
Tissue injury is a common clinical problem, which may cause great burden on patients' life. It is important to develop functional scaffolds to promote tissue repair and regeneration. Due to their unique composition and structure, microneedles have attracted extensive attention in various tissues regeneration, including skin wound, corneal injury, myocardial infarction, endometrial injury, and spinal cord injury et al. Microneedles with micro-needle structure can effectively penetrate the barriers of necrotic tissue or biofilm, therefore improving the bioavailability of drugs. The use of microneedles to deliver bioactive molecules, mesenchymal stem cells, and growth factors in situ allows for targeted tissue and better spatial distribution. At the same time, microneedles can also provide mechanical support or directional traction for tissue, thus accelerating tissue repair. This review summarized the research progress of microneedles for in situ tissue regeneration over the past decade. At the same time, the shortcomings of existing researches, future research direction and clinical application prospect were also discussed.
... Collagen-based artificial vascular scaffolds or hydrogels also display great potential in promoting cardiovascular regeneration [20]. In addition, collagen-based bioactive materials also have good application perspectives in gynecology, reproductive medicine [8,21], plastic repair [22,23], cornea regeneration [24][25][26] among others. Collagen scaffolds provide a bionic microenvironment to maintain the structural integrity of regenerated tissues [27], which has an important influence on the cell behavior and tissue repair. ...
In tissue engineering, bioactive materials play an important role, providing structural support, cell regulation and
establishing a suitable microenvironment to promote tissue regeneration. As the main component of extracellular
matrix, collagen is an important natural bioactive material and it has been widely used in scientific research and clini-
cal applications. Collagen is available from a wide range of animal origin, it can be produced by synthesis or through
recombinant protein production systems. The use of pure collagen has inherent disadvantages in terms of physico-
chemical properties. For this reason, a processed collagen in different ways can better match the specific require-
ments as biomaterial for tissue repair. Here, collagen may be used in bone/cartilage regeneration, skin regeneration,
cardiovascular repair and other fields, by following different processing methods, including cross-linked collagen,
complex, structured collagen, mineralized collagen, carrier and other forms, promoting the development of tissue
engineering. This review summarizes a wide range of applications of collagen-based biomaterials and their recent
progress in several tissue regeneration fields. Furthermore, the application prospect of bioactive materials based on
collagen was outlooked, aiming at inspiring more new progress and advancements in tissue engineering research.
... 20 Recent efforts have been aimed at developing in situ-forming corneal substitutes. 21,22 Collagen type I is a particularly promising biomaterial for corneal regeneration since it is the most abundant protein in the native stromal extracellular matrix (ECM). 23 Our laboratory has recently developed in situ-forming hydrogels composed of collagen type I and bioinert polyethylene glycol (PEG) using N-hydroxysuccinimide (NHS) ester chemistry that improved wound healing and reduced scar formation in rabbits with lamellar injury. ...
Purpose:
Millions worldwide suffer vision impairment or blindness from corneal injury, and there remains an urgent need for a more effective and accessible way to treat corneal defects. We have designed and characterized an in situ-forming semi-interpenetrating polymer network (SIPN) hydrogel using biomaterials widely used in ophthalmology and medicine.
Methods:
The SIPN was formed by cross-linking collagen type I with bifunctional polyethylene glycol using N-hydroxysuccinimide ester chemistry in the presence of linear hyaluronic acid (HA). Gelation time and the mechanical, optical, swelling, and degradation properties of the SIPN were assessed. Cytocompatibility with human corneal epithelial cells and corneal stromal stem cells (CSSCs) was determined in vitro, as was the spatial distribution of encapsulated CSSCs within the SIPN. In vivo wound healing was evaluated by multimodal imaging in an anterior lamellar keratectomy injury model in rabbits, followed by immunohistochemical analysis of treated and untreated tissues.
Results:
The collagen-hyaluronate SIPN formed in situ without an external energy source and demonstrated mechanical and optical properties similar to the cornea. It was biocompatible with human corneal cells, enhancing CSSC viability when compared with collagen gel controls and preventing encapsulated CSSC sedimentation. In vivo application of the SIPN significantly reduced stromal defect size compared with controls after 7 days and promoted multilayered epithelial regeneration.
Conclusions:
This in situ-forming SIPN hydrogel may be a promising alternative to keratoplasty and represents a step toward expanding treatment options for patients suffering from corneal injury.
Translational relevance:
We detail the synthesis and initial characterization of an SIPN hydrogel as a potential alternative to lamellar keratoplasty and a tunable platform for further development in corneal tissue engineering and therapeutic cell delivery.
... Refractive index 1.373, similar to human cornea [66,67]. ...
... The biofabricated corneal tissue should withstand the equivalent (minimum) or higher (preferred) liquid pressure than the IOP of native human cornea, which ranges between 11-21 mmHg or ~2.5 kPa [67]. ...
Corneal diseases are the third most prevalent cause of blindness after cataract and glaucoma. It is estimated that about 5 million people in the world are affected by bilateral corneal blindness with an additional 23 million with unilateral blindness. Cornea transplantation is the standard practice for the management of various cornea related pathologies like fibrosis, ulcers, keratitis, etc. The high transplant cost, increased risk of graft failure/rejection, and long waiting list due to limited availability of good quality donor cornea imposes a huge clinical burden. Recently, biofabrication technologies are gaining a lot of attention because of their potential to direct hierarchical assembly of three-dimensional (3D) biological structures for tissue construction for various biomedical and clinical applications. In this regard, 3D bioprinting, which involves layer-by-layer deposition of acellular or cell-laden bioink in a specific pattern corresponding to the organotypic morphology of tissues/organs, has been extensively investigated for the fabrication of corneal substitutes. In addition to this methodology, novel biofabrication techniques have been explored for the fabrication of corneal tissues using bioinks with optical and mechanical performances comparable to native cornea tissue. In this review, we highlight the recent advances and offer future perspectives in the fabrication of corneal tissue equivalents that can be potentially employed for effective clinical repair, reconstruction, and regeneration of the cornea.
... A number of alternative treatments have been published as alternatives to treat corneal perforations including using collagen-based fillers, 15 fibrin glueassisted amniotic membranes, 16 and LiQD corneas. 17 These studies have demonstrated successful sealing results. The LiQD corneas also contained fibrinogen and thrombin, similar to the fibrin glue-assisted amniotic membrane approach to allow for in situ gelation. ...
Purpose:
Corneal perforation is a clinical emergency that can result in blindness. Currently corneal perforations are treated either by cyanoacrylate glue which is toxic to corneal cells, or by using commercial fibrin glue for small perforations. Both methods use manual delivery which lead to uncontrolled application of the glues to the corneal surface. Therefore, there is a need to develop a safe and effective alternative to artificial adhesives.
Methods:
Previously, our group developed a transparent human platelet lysate (hPL)-based biomaterial that accelerated corneal epithelial cells healing in vitro. This biomaterial was further characterized in this study using rheometry and adhesive test, and a two-component delivery system was developed for its application. An animal trial (5 New Zealand white rabbits) to compare impact of the biomaterial and cyanoacrylate glue (control group) on a 2 mm perforation was conducted to evaluate safety and efficacy.
Results:
The hPL-based biomaterial showed higher adhesiveness compared to commercial fibrin glue. Treatment rabbits had lower pain scores and faster recovery, despite generating similar scar-forming structure compared to controls. No secondary corneal ulcer was generated in rabbits treated with the bio-adhesive.
Conclusions:
This study reports an in situ printing system capable of delivering a hPL-based, transparent bio-adhesive and successfully treating small corneal perforations. The bio-adhesive-treated rabbits recovered faster and required no additional analgesia.
Translational relevance:
The developed in situ hPL bio-adhesives treatment represents a new format of treating corneal perforation that is easy to use, allows for accurate application, and can be a potentially effective and pain relief treatment.
... However, cyanoacrylate has low biocompatibility 17 and causes inflammation 19 . Incomplete polymerization leaves behind toxic cyanoacrylate monomers that can undergo hydrolysis to release potentially toxic compounds such as formaldehyde and alkyl cyanoacrylate 20 . These can induce further corneal injury through scarring and neovascularization 20 . ...
... Incomplete polymerization leaves behind toxic cyanoacrylate monomers that can undergo hydrolysis to release potentially toxic compounds such as formaldehyde and alkyl cyanoacrylate 20 . These can induce further corneal injury through scarring and neovascularization 20 . Many patients treated with cyanoacrylate eventually require full corneal transplantation. ...
... Other crosslinkers, e.g., glutaraldehyde and hexamethylene diisocyanate, have also been used to produce collagen-based implants; however, modification of important functional groups of the parent biomaterial and low biocompatibility remain limitations. Recently, light-induced crosslinking of ECM components in the presence of photo initiators has become popular 29,35 , although patients with corneal inflammation are highly light-sensitive and it can be difficult for them to tolerate intense visible light application for any length of time without anaesthesia 20 . ...
Development of an artificial cornea can potentially fulfil the demand of donor corneas for transplantation as the number of donors is far less than needed to treat corneal blindness. Collagen-based artificial corneas stand out as a regenerative option, having promising clinical outcomes. Collagen crosslinked with chemical crosslinkers which modify the parent functional groups of collagen. However, crosslinkers are usually cytotoxic, so crosslinkers need to be removed from implants completely before application in humans. In addition, crosslinked products are mechanically weak and susceptible to enzymatic degradation. We developed a crosslinker free supramolecular gelation strategy using pyrene conjugated dipeptide amphiphile (PyKC) consisting of lysine and cysteine; in which collagen molecules are intertwined inside the PyKC network without any functional group modification of the collagen. The newly developed collagen implants (Coll-PyKC) are optically transparent and can effectively block UV light, are mechanically and enzymatically stable, and can be sutured. The Coll-PyKC implants support the growth and function of all corneal cells, trigger anti-inflammatory differentiation while suppressing the pro-inflammatory differentiation of human monocytes. Coll-PyKC implants can restrict human adenovirus propagation. Therefore, this crosslinker-free strategy can be used for the repair, healing, and regeneration of the cornea, and potentially other damaged organs of the body.
... 12 Although keratoplasty is widely used to replace the diseased cornea, the supply of donor corneas is significantly short, meeting only one-seventieth of the global need. 13 Tissue engineering approaches have been developed in an effort to construct artificial corneas with biomaterials, such as hydrogel-based scaffolds and gelatin nanofibers. 14 Biosynthetic implants made from recombinant human collagen are low risk. ...
Purpose
To propose improved stem cell-based therapeutic strategy for limbal stem cell deficiency (LSCD) treatment.
Design
Experimental, randomized or parallel-grouped animal study.
Participants
Fifty adult male New Zealand white rabbits.
Method
Human limbal stem/progenitor cells (LSCs) and limbal stromal stem/progenitor cells (LSSCs) were cultured in serum-free condition, and further differentiated into corneal epithelial cells (CECs) and keratocytes, respectively. All types of cells were characterized with lineage-specific markers. Gene expression analysis was performed to identify the potential function of LSSCs in corneal regeneration. Two LSCD models of rabbits for transplantation experiments were used: transplantation was performed at the time of limbal and corneal epithelial excision (LSCD model), and transplantation was performed after clinical signs were induced in an LSCD model (pLSCD model). The pLSCD model better mimics the pathological changes and symptoms of human LSCD. Rabbit models were receiving LSC or LSC-LSSC treatment. Corneal epithelial defects, neovascularization and opacity were assessed every 3 weeks for 24 weeks. ZsGreen-labeled LSSCs were used for short-term tracking in vivo.
Main Outcome Measures
Rates of corneal epithelial defect area; corneal neovascularization and opacity scores; graft survival rate; immunofluorescence staining of specific markers.
Results
Both LSC transplantation and LSC-LSSC co-transplantation could effectively repair the corneal surface in the LSCD model. These two strategies showed no significant differences in terms of the graft survival rate or epithelial repair. However, corneal opacity was observed in the LSC group (in 3 of 8 rabbits) but not in the LSC-LSSC group. Notably, when treating LSCD rabbit models with distinguishable stromal opacification and neovascularization, co-transplantation of LSCs and LSSCs exhibited significantly better therapeutic effects than LSCs alone, with graft survival rates of 87.5% and 37.5%, respectively. The implanted LSSCs could differentiate into keratocytes during the wound-healing process. RNA sequencing (RNA-seq) analysis shown that the stromal cells produced not only a collagen-rich extracellular matrix to facilitate reconstruction of the lamellar structure, but also niche factors that accelerated epithelial cell growth and inhibited angiogenesis and inflammation.
Conclusions
Our findings highlight the support of stromal cells in niche homeostasis and tissue regeneration, providing LSC-LSSC co-transplantation as a new treatment strategy for corneal blindness.
... Collagen, the most abundant fibrous protein, is found within hard and soft tissues, such as connective tissue [126], skin [127][128][129], tendon [130], cornea [131], and cartilage [132][133][134][135], and is a crucial constituent of the ECM [136]. In this regard, collagen derived from the bovine [137], porcine [138], fish [139], marine sponge [140], shellfish [141], and jellyfish [142] is largely explored, so as to be utilized as a biocompatible material in various fields [143]. ...
The successful design of a hydrogel for tissue engineering requires a profound understanding of its constituents’ structural and molecular properties, as well as the proper selection of components. If the engineered processes are in line with the procedures that natural materials undergo to achieve the best network structure necessary for the formation of the hydrogel with desired properties, the failure rate of tissue engineering projects will be significantly reduced. In this review, we examine the behavior of proteins as an essential and effective component of hydrogels, and describe the factors that can enhance the protein-based hydrogels’ structure. Furthermore, we outline the fabrication route of protein-based hydrogels from protein microstructure and the selection of appropriate materials according to recent research to growth factors, crucial members of the protein family, and their delivery approaches. Finally, the unmet needs and current challenges in developing the ideal biomaterials for protein-based hydrogels are discussed, and emerging strategies in this area are highlighted.
