Corneal Stromal Regeneration Research Articles

Fabrication of GelMA - Agarose Based 3D Bioprinted Photocurable Hydrogel with In Vitro Cytocompatibility and Cells Mirroring Natural Keratocytes for Corneal Stromal Regeneration.

The complex anatomy of the cornea and the subsequent keratocyte-fibroblast transition have always made corneal stromal regeneration difficult. Recently, 3D printing has received considerable attentionin terms of fabrication of scaffoldswith precise dimension and pattern. In the current work, 3D printable polymer hydrogels made of GelMA/agarose are formulated and its rheological properties are evaluated. Despite the variation in agarose content, both the hydrogels exhibited G'>G'' modulus. A prototype for 3D stromal model is created using Solid Works software, mimicking the anatomy of an adult cornea. The fabrication of 3D-printed hydrogels is performed using pneumatic extrusion. The FTIR analysis speculated that the hydrogel is well crosslinked and established strong hydrogen bonding with each other, thus contributing to improved thermal and structural stability. The MTT analysis revealed a higher rate of cell proliferation on the hydrogels. The optical analysis carried out on the 14th day of incubation revealed that the hydrogels exhibit transparency matching with natural corneal stromal tissue. Specific protein marker expression confirmed the keratocyte phenotype and showed that the cells do not undergo terminal differentiation into stromal fibroblasts. The findings of this work point to the potential of GelMA/A hydrogels as a novel biomaterial for corneal stromal tissue engineering.

Harnessing Corneal Stromal Regeneration for Vision Restoration: A Comprehensive Review of the Emerging Treatment Techniques for Keratoconus.

Keratoconus is a progressive corneal disorder characterized by thinning and conical protrusion, leading to visual impairment that often necessitates advanced treatment strategies. Traditional management options, including corrective lenses, corneal cross-linking (CXL), and surgical interventions such as corneal transplants and intracorneal ring segments (ICRS), address symptoms but have limitations, especially in progressive or advanced cases. Recent advancements in corneal stromal regeneration offer promising alternatives for enhancing vision restoration and halting disease progression. This review explores emerging techniques focused on corneal stromal regeneration, emphasizing cell-based therapies, tissue engineering, and gene therapy. Cell-based approaches, including corneal stromal stem cells and adipose-derived stem cells, are promising to promote tissue repair and functional recovery. Tissue engineering techniques, such as developing synthetic and biological scaffolds and 3D bioprinting, are being investigated for their ability to create viable corneal grafts and implants. Additionally, gene therapy and molecular strategies, including gene editing technologies and the application of growth factors, are advancing the potential for targeted treatment and regenerative medicine. Despite these advancements, challenges remain, including technical limitations, safety concerns, and ethical considerations. This review aims to provide a comprehensive overview of these innovative approaches, highlighting their current status, clinical outcomes, and future directions in keratoconus management.

Polyvinyl alcohol: Starch based membrane scaffolds for tissue transparency requirements: Fabrication, characterization and cytotoxicity studies

In this study, the characterization and cytocompatibility of polyvinyl alcohol:starch (PVA:ST) based membrane tissue scaffolds developed for tissue transparency requirements in tissue engineering applications were evaluated in the presence of sodium dodecyl sulphate (SDS). In this context, two different amounts of SDS were added to the polymer structure of transparent membrane scaffolds to roughen the surfaces of PVA:ST membranes and to promote cell adhesion. In the light of this aim, three different types of membranes were created: A constant PVA:ST ratio of 9:1 (wt:wt) was used as a control and varying amounts of SDS (2 and 4 wt% of the total polymer amount) were added to the polymer solutions. Membrane tissue scaffolds were formed by freeze-drying method. Scaffold properties such as morphology, transparency, water absorption, chemical structure and cytocompatibility were investigated. In terms of physicochemical properties, it was observed that with the addition of SDS, the membranes had a rougher surface while retaining water retention and transparency. In cytotoxicity studies, the membranes were found to be non-cytotoxic and promoted cell proliferation as assessed by MTT assay. Furthermore, the PVA:ST membrane scaffold with 4% SDS added was able to promote the attachment, migration and proliferation of Mouse Embryonic Fibroblasts (MEFs) more than the control group. Overall, the results indicate that SDS-added PVA:ST scaffold membranes are suitable for human corneal stromal regeneration, wound dressing material and neural tissue engineering applications.

Acellular human amniotic membrane: A safe and stable matrix for corneal stromal regeneration

Amniotic membrane (AM) transplantation is used as a safe and effective method to treat a variety of ocular complications. On the ocular surface, this membrane degrades within a few days. This study investigated the safety and long-term stability of human amniotic membrane (HAM) grafts in the corneal stroma. Three rabbits with healthy eyes underwent unilateral stromal pocketing with implantation of decellularized HAM (dHAM). Examinations of graft bio-integration were performed by corneal imaging using slit lamp and optical coherence tomography (OCT) at 1 week, 3 months, and 9 months. The thickness of the thickest part of the amniotic membrane was 187 μm at the first follow-up, 184 μm at 3 months, and 166 μm at 9 months. Histological evaluation was performed after the last follow-up. The results showed no clinical complications, such as opacity, neovascularization, infection, or rejection. The thickness of the grafts remained stable over time. Histopathological analyses showed the grafts were integrated into the corneal stroma without any inflammation. This study indicated that HAM was a naturally safe, stable, and compatible matrix in the corneal stroma and was considered a promising candidate for corneal stromal regeneration.

3D printing sequentially strengthening high-strength natural polymer hydrogel bilayer scaffold for cornea regeneration.

3D printing of high-strength natural polymer biodegradable hydrogel scaffolds simultaneously resembling the biomechanics of corneal tissue and facilitating tissue regeneration remains a huge challenge due to the inherent brittleness of natural polymer hydrogels and the demanding requirements of printing. Herein, concentrated aqueous solutions of gelatin and carbohydrazide-modified alginate (Gel/Alg-CDH) are blended to form a natural polymer hydrogel ink, where the hydrazides in Alg-CDH are found to form strong hydrogen bonds with the gelatin. The hydrogen-bonding-strengthened Gel/Alg-CDH hydrogel demonstrates an appropriate thickened viscosity and shear thinning for extrusion printing. The strong hydrogen bonds contribute to remarkably increased mechanical properties of Gel/Alg-CDH hydrogel with a maximum elongation of over 400%. In addition, sequentially Ca2+-physical crosslinking and then moderately chemical crosslinking significantly enhance the mechanical properties of Gel/Alg-CDH hydrogels that ultimately exhibit an intriguing J-shaped stress-strain curve (tensile strength of 1.068 MPa and the toughness of 677.6 kJ/m2). The dually crosslinked Gel-Alg-CDH-Ca2+-EDC hydrogels demonstrate a high transparency, physiological swelling stability and rapid enzymatic degradability, as well as suturability. The growth factor and drug-loaded biomimetic bilayer hydrogel scaffold are customized via a multi-nozzle printing system. This bioactive bilayer hydrogel scaffold considerably promotes regeneration of corneal epithelium and stroma and inhibits cornea scarring in rabbit cornea keratoplasty.

A Versatile Hydrogel with Antibacterial and Sequential Drug-Releasing Capability for the Programmable Healing of Infectious Keratitis.

Hydrogels have attracted tremendous attention as favorable corneal substitutes for treating severe infectious keratitis (IK). However, current hydrogel-based corneal substitutes were majorly designed to promote the single stage of corneal regeneration, which falls short in meeting the clinical management needs of severe IK including the multiple phases of corneal wound healing. Herein, we introduce a versatile hybrid hydrogel (SQPV) composed of silk fibroin and chitosan, which exhibits spatiotemporal properties for drug release. The SQPV is fabricated by incorporating verteporfin-loaded poly(lactic-co-glycolic)-polyethylene glycol-o-nitrobenzene micelles into a hydrogel network, which is formed from methacrylate silk fibroin and glycidyl methacrylate functionalized quaternized chitosan containing polydeoxyribonucleotide. This double network approach results in a material with exceptional anti-inflammatory, antibacterial, and proliferative stimulation and tissue remodeling regulation capabilities. Furthermore, SQPV showcases mechanical strength and transparency akin to those of native cornea. Extensive in vitro and in vivo studies validate SQPV's ability to effectively eliminate residual bacteria, mitigate inflammation, foster regeneration of corneal epithelium and stroma, prevent corneal scarring, and ultimately expedite wound healing. In summary, the SF/CS-based hybrid hydrogel may represent a promising substitute for comprehensive corneal repair and regeneration in severe IK.

Corneal stromal filler injection of gelatin-based photocurable hydrogels for maintaining the corneal thickness and reconstruction of corneal stroma

Refractive errors manifest as thinning of the central cornea, which can seriously impair vision. The corneal stromal filler is an innovative concept for treating refractive problems. However, corneal stromal regeneration poses a challenge for the tissue-engineered cornea. Hence, materials for corneal reconstruction with high transparency and biocompatibility are urged to be developed. This study aimed to establish a novel therapeutic approach by injecting gelatin-based photocurable hydrogel fillers into the corneal stroma to maintain corneal thickness and promote corneal stromal regeneration for refractive error. The hydrogel possesses high transparency and suitable mechanical qualities, which degrade slowly in the cornea. Furthermore, the hydrogel presents high biocompatibility for cellular adhesion and proliferation. The hydrogel can also provide space for cells to develop and facilitate cell migration when cells are infused into it. In vivo experiments showed that the hydrogel could effectively fortify corneal thickness and induce corneal stromal regeneration. The photocurable hydrogel showed potential in terms of corneal reconstruction.

Optimization and evaluation of oxygen-plasma-modified, aligned, poly (Є-caprolactone) and silk fibroin nanofibrous scaffold for corneal stromal regeneration

The shortage of human donor corneas for transplantation necessitates the exploration of tissue engineering approaches to develop corneal substitutes. However, these substitutes must possess the necessary strength, transparency, and ability to regulate cell behaviour before they can be used in patients. In this study, we investigated the effectiveness of an oxygen plasma surface-modified poly-ε-caprolactone (PCL) combined with silk fibroin (SF) nanofibrous scaffold for corneal stromal regeneration. To fabricate the electrospun scaffolds, PCL and SF blends were used on a rotating mandrel. The optimization of the blend aimed to replicate the structural and functional properties of the human cornea, focusing on nanofibre alignment, mechanical characteristics, and in vitro cytocompatibility with human corneal stromal keratocytes. Surface modification of the scaffold resulted in improved transparency and enhanced cell interaction. Based on the evaluation, a composite nanofibrous scaffold with a 1:1 blend of PCL and SF was selected for a more comprehensive analysis. The biological response of keratocytes to the scaffold was assessed through cellular adhesion, proliferation, cytoskeletal organization, gene expression, and immunocytochemical staining. The scaffold facilitated the adhesion of corneal stromal cells, supporting cell proliferation, maintaining normal cytoskeletal organization, and promoting increased expression of genes associated with healthy corneal stromal keratocytes. These findings highlight the potential of a surface-modified PCL/SF blend (1:1) as a promising scaffolding material for corneal stromal regeneration. The developed scaffold not only demonstrated favourable biological interactions with corneal stromal cells but also exhibited characteristics aligned with the requirements for successful corneal tissue engineering. Further research and refinement of these constructs could lead to significant advancements in addressing the shortage of corneas for transplantation, ultimately improving the treatment outcomes for patients in need.

Intracorneal Implantation of 3D Bioprinted Scaffolds Containing Mesenchymal Stromal Cells Using Femtosecond-Laser-Assisted Intrastromal Keratoplasty.

Injury of the cornea is a complex biological process. Regeneration of the corneal stroma can be facilitated by the presence of mesenchymal stromal cells (MSCs) and application of tissue equivalents. A new tissue-engineering strategy for corneal stroma regeneration is presented using cellularized 3D bioprinted hydrogel constructs implanted into organ cultured porcine corneas using femtosecond laser-assisted intrastromal keratoplasty. The ex vivo cultured, MSC-loaded 3D bioprinted structures remain intact, support cell survival, and contain de novo synthesized extracellular matrix components and migrating cells throughout the observation period. At day 14 postimplantation, the cellularized tissue equivalents contain few or no cells, as demonstrated by optical coherence tomography imaging and immunofluorescent staining. This study successfully combines a laboratory-based method with modern, patient-care practice to produce a cell-laden tissue equivalent for corneal implantation. Optimal bioink composition and cellularization of tissue equivalents are essential in fine-tuning a method to promote the current technique as a future treatment modality.

Corneal stromal regeneration

AbstractCorneal grafting is one of the most common forms of human tissue transplantation. The corneal stroma is responsible for many characteristics of the cornea. For these reasons, an important volume of research has been made to replicate the corneal stroma in the laboratory to find an alternative to classical corneal transplantation techniques. There is an increasing interest today in cell therapy of the corneal stroma using induced pluripotent stem cells or mesenchymal stem cells since these cells have shown to be capable of producing new collagen within the host stroma and even to improve its transparency. The first clinical experiment on corneal stroma regeneration in advanced keratoconus cases has been reported and included. Fourteen patients were randomized and enrolled into 3 experimental groups: (1) patients underwent implantation of autologous adipose‐derived adult stem cells alone, (2) patients received decellularized donor corneal stroma laminas, and (3) patients received implantation of recellularized donor laminas with adipose‐derived adult stem cells. Clinical improvement was detected with all cases in their visual, pachymetric, and topographic parameters of the operated corneas.

Silk fibroin based interpenetrating network hydrogel for corneal stromal regeneration

There is a need to develop tissue engineering based approaches to address the shortage of donor corneas worldwide for transplantation. To do this a novel approach to fabricate three-dimensional hydrogels using free-radical polymerization was investigated to generate constructs for corneal stromal tissue regeneration. Different ratios of silk fibroin (SF) to polyacrylamide (PA) were used to fabricate semi-interpenetrating hydrogels. Scanning electron micrograph displayed the interconnectivity of pores within the fabricated hydrogels. Pore sizes ranged from 25 to 66 μm. Scaffolds with increasing concentration of SF had enhanced β-sheet structure (verified by Fourier transform infrared spectroscopy). The biological response of human corneal stromal cells to these hydrogels was examined using cellular adhesion, proliferation, cytoskeleton organization, gene expression and immunocytochemical analysis. The fabricated hydrogels possess rapid gelation (∼3 min) at 37 °C, 84 % porosity facilitating keratocyte migration during healing, improved cellular adhesion and no cytotoxicity, indicating their efficiency for in-situ corneal tissue regeneration. Presence of SF in semi-interpenetrating network hydrogel enhanced cellular proliferation, elevated GAG deposition, and increased expression of keratocyte genes, normally associated with healthy corneal stromal tissue. This study acts as an initial step towards fabricating SF based semi-interpenetrating network hydrogels for developing clinically applicable ocular implants.

Application Prospect and Preliminary Exploration of GelMA in Corneal Stroma Regeneration.

Corneal regeneration has become a prominent study area in recent decades. Because the corneal stroma contributes about 90% of the corneal thickness in the corneal structure, corneal stromal regeneration is critical for the treatment of cornea disease. Numerous materials, including deacetylated chitosan, hydrophilic gel, collagen, gelatin methacrylate (GelMA), serine protein, glycerol sebacate, and decellularized extracellular matrix, have been explored for keratocytes regeneration. GelMA is one of the most prominent materials, which is becoming more and more popular because of its outstanding three-dimensional scaffold structure, strong mechanics, good optical transmittance, and biocompatibility. This review discussed recent research on corneal stroma regeneration materials and related GelMA.

Corneal Stroma Regeneration with Collagen-Based Hydrogel as an Artificial Stroma Equivalent: A Comprehensive In Vivo Study.

Restoring the anatomical and functional characteristics of the cornea using various biomaterials is especially relevant in the context of a global shortage of donor tissue. Such biomaterials must be biocompatible, strong, and transparent. Here, we report a Viscoll collagen membrane with mechanical and optical properties suitable for replacing damaged stromal tissue. After removing a portion of the stroma, a Viscoll collagen membrane was implanted into the corneas of rabbits. After 6 months, the active migration of host cells into Viscoll collagen membranes was noted, with the preservation of corneal transparency in all experimental animals. Effective integration of the Viscoll collagen membrane with corneal tissue promoted nerve regeneration in vivo, as confirmed by in vivo confocal microscopy. We also demonstrated the safety and efficacy of the Viscoll collagen membrane for corneal stroma regeneration. Thus, in combination with the proposed packaging format that provides long-term storage of up to 10 months, this material has great potential for replacing and regenerating damaged stromal tissues.

Natural Dual-Crosslinking Bioadhesive Hydrogel for Corneal Regeneration in Large-Size Defects.

Corneal injectable hydrogels represent a promising alternative to alleviate donor shortage and simplify traditional surgeries. However, most hydrogels focus on repairing focal corneal defects (≤3.5mm) and leave many clinical requirements unmet. Herein, a novel ion-activated bioadhesive hydrogel (IonBAH) is designed and its long-term performance of repairing large corneal defects (6mm) is evaluated in rabbits for 6 months. The IonBAH is a dual-network hydrogel composed of natural corneal extracellular matrix and peptide-modified alginate, which enables its desirable transparency and biocompatibility, tunable mechanics, and robust adhesion. Moreover, the IonBAH maintains the secretory phenotype of quiescent keratocytes, while preventing their myofibroblastic differentiation in vitro. Upon application in situ, it rapidly seals the 6mm corneal defect and forms normal curvature through the coverage of a contact lens impregnated with calcium ions. During the 6 months follow-up, the IonBAH promotes rapid regeneration of corneal epithelium, stroma, and nerves with restored transparency, equivalent to the outcome of donor corneal transplantation. In addition, the suitability of IonBAH as an adhesive and patch for various clinical requirements are also evaluated with a pleasing outcome. Collectively, IonBAH may provide a clinically applicable scaffold for corneal surgeries, especially in large defect repair.

Porous hydrogel constructs based on methacrylated gelatin/polyethylene oxide for corneal stromal regeneration

The lack of donor sources for corneal transplantation leads to the development of alternative materials for corneal regeneration and repair. As the stroma occupies the thickest layer of the cornea, biomimetic corneal stromal constructs were widely observed by tissue engineering. In this study, a light-curing porous hydrogel based on the two-phase aqueous emulsion was prepared to induce the regeneration of corneal stroma. Methacrylated gelatin (GelMA) solution containing photoinitiator was mixed with aqueous polyethylene oxide (PEO) solution to form an immiscible two-phase emulsion. We characterized physical and chemical properties, containing the pore size, mechanical properties, degradation properties, and swelling properties, to choose the preparation process of the porous hydrogel. The in vitro studies performed in this study suggested that the porous hydrogels had good biocompatibility and the specific matrix secreted by corneal stromal cells gradually increased, so the porous hydrogels promoted the secretion of extracellular matrix. The hydrogels were inserted in a mid-stromal pocket created by minor surgery to confirm their histocompatibility and application in corneal regeneration and repair.

Oxidation-mediated scaffold engineering of hyaluronic acid-based microcarriers enhances corneal stromal regeneration

The functional design of scaffolding biomaterials with potent capabilities of promoting cell adhesion and proliferation is critically important for tissue repair and regeneration. Here, we exploit the effects of oxidation level of aldehyde hyaluronic acid (oHA) on gelatin microcarriers for repairing corneal injuries. Specifically, high oxidation levels can endow the microcarrier surface with large oHA grafting amount, smooth topography, and strong stiffness, consequently formulating biocompatible scaffolding materials with superior affinities for keratocyte attachment and growth. In a rabbit model of corneal alkali burn injury, single intracorneal injection of keratocytes/functionalized microcarriers with an appropriate oxidation level could effectively reduce corneal swelling (~62-fold improvement), recover ~94% collagen production and ~89% keratocan expression, and repair disordered collagenous stromal architecture after 4 weeks. These findings on the oxidation level effects of the aldehyde polysaccharide show a great potential use in the development of advanced scaffolds for efficient tissue engineering.

Germinal peptide eye drop promotes corneal epithelial and stromal defect healing in rabbit model

ABSTRACT Objective Corneal defect is a common disease in ophthalmology caused by trauma, inflammation, drug toxicity, or surgery. To investigate the effect of germinal peptide eye drop on corneal epithelial and stromal defects after lamellar keratectomy in rabbit model. Methods Eighty-five male New Zealand white rabbits were divided into five groups: Germinal Peptide eye drop at three different concentration groups, normal saline (negative control group), recombinant human epidermal growth factor (rh-EGF) eye drop (positive control group). Corneal epithelial and stromal defects of around 150–200 μm in depth were created with an 8 mm diameter trephine in the center of the right eyes of all animals. Results Germinal peptide eye drop with the concentration of 0.001%, 0.002%, and 0.004% and rh-EGF eye drop were more effective in promoting healing, reducing opacity, and edema during the process of corneal epithelial and stromal defect regeneration compared with 0.9% normal saline. No significant difference was observed among the three different doses of germinal peptide eye drop. Compared with the saline control group, the structures of the regenerated corneas were more orderly and less inflammatory cell infiltration was observed in the germinal peptide eye drop groups and the rh-EGF eye drop group. Conclusion Germinal peptide eye drop (0.001%, 0.002%, and 0.004%) can significantly stimulate the regeneration of corneal epithelia and stroma and reduce corneal opacity and edema. Dose dependency was not observed in the current study.

Exosomes-loaded thermosensitive hydrogels for corneal epithelium and stroma regeneration

Corneal damage forms scar tissue and manifests as permanent corneal opacity, which is the main cause of visual impairment caused by corneal diseases. To treat these diseases, herein, we developed a novel approach based on the exosome derived from induced pluripotent stem cell-derived mesenchymal stem cells (iPSC-MSCs) combined with a thermosensitive hydrogel, which reduces scar formation and accelerates the healing process. We found that a thermosensitive chitosan-based hydrogels (CHI hydrogel) sustained-release iPSC-MSC exosomes can effectively promote the repair of damaged corneal epithelium and stromal layer, downregulating mRNA expression coding for the three most enriched collagens (collagen type I alpha 1, collagen type V alpha 1 and collagen type V alpha 2) in corneal stroma and reducing scar formation in vivo. Furthermore, iPSC-MSCs secrete exosomes that contain miR-432-5p, which suppresses translocation-associated membrane protein 2 (TRAM2), a vital modulator of the collagen biosynthesis in the corneal stromal stem cells to avert the deposition of extracellular matrix (ECM). Our findings indicate that iPSC-MSCs secrete miRNA-containing exosomes to promote corneal epithelium and stroma regeneration, and that miR-432-5p can prevent ECM deposition via a mechanism most probably linked to direct repression of its target gene TRAM2. Overall, our exosomes-based thermosensitive CHI hydrogel, is a promising technology for clinical therapy of various corneal diseases.

Plant Recombinant Human Collagen Type I Hydrogels for Corneal Regeneration

PurposeTo determine feasibility of plant-derived recombinant human collagen type I (RHCI) for use in corneal regenerative implantsMethodsRHCI was crosslinked with 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) to form hydrogels. Application of shear force to liquid crystalline RHCI aligned the collagen fibrils. Both aligned and random hydrogels were evaluated for mechanical and optical properties, as well as in vitro biocompatibility. Further evaluation was performed in vivo by subcutaneous implantation in rats and corneal implantation in Göttingen minipigs.ResultsSpontaneous crosslinking of randomly aligned RHCI (rRHCI) formed robust, transparent hydrogels that were sufficient for implantation. Aligning the RHCI (aRHCI) resulted in thicker collagen fibrils forming an opaque hydrogel with insufficient transverse mechanical strength for surgical manipulation. rRHCI showed minimal inflammation when implanted subcutaneously in rats. The corneal implants in minipigs showed that rRHCI hydrogels promoted regeneration of corneal epithelium, stroma, and nerves; some myofibroblasts were seen in the regenerated neo-corneas.ConclusionPlant-derived RHCI was used to fabricate a hydrogel that is transparent, mechanically stable, and biocompatible when grafted as corneal implants in minipigs. Plant-derived collagen is determined to be a safe alternative to allografts, animal collagens, or yeast-derived recombinant human collagen for tissue engineering applications. The main advantage is that unlike donor corneas or yeast-produced collagen, the RHCI supply is potentially unlimited due to the high yields of this production method.Lay SummaryA severe shortage of human-donor corneas for transplantation has led scientists to develop synthetic alternatives. Here, recombinant human collagen type I made of tobacco plants through genetic engineering was tested for use in making corneal implants. We made strong, transparent hydrogels that were tested by implanting subcutaneously in rats and in the corneas of minipigs. We showed that the plant collagen was biocompatible and was able to stably regenerate the corneas of minipigs comparable to yeast-produced recombinant collagen that we previously tested in clinical trials. The advantage of the plant collagen is that the supply is potentially limitless.

Recombinant human collagen hydrogels with hierarchically ordered microstructures for corneal stroma regeneration

Tissue engineered scaffolds have demonstrated values in fabricating corneal equivalence. Attempts in this area tend to develop functional scaffold with physiochemical properties and structure resembling the native corneal tissue. Herein, a novel recombinant human collagen (RHC) based scaffold with ordered microstructures is presented to induce the regeneration of corneal stroma. As the RHC is modified with methacrylate anhydride (MA) under optimized concentration and is tailored with hierarchically ordered microstructures of aligned microgrooves and inverse opal nano-scale pores, the derived collagen hydrogels (RHCMA) can possess physiochemical properties resembling native cornea. Diverse topological structures (culture dish, RHCMA hydrogel without patterns, MI-RHCMA hydrogel patch with different width of microgrooves) are determined to study their influence on the growth and differentiation of corneal limbal stromal stem cells (LSSCs), and results demonstrate that the novel MI-RHCMA hydrogel patch could induce LSSCs to grow orderly and differentiate into keratocyte in vitro. Rat intrastromal keratoplasty are performed to examine the tissue repair efficacy of the derived collagen hydrogels, and the results indicate that the novel MI-RHCMA hydrogel patch could induce the regeneration of damaged corneal stroma in vivo. These results indicate that our presented scaffolds can be an effective transplantation graft and will find important values for the damaged cornea in the clinic.