Corneal Regeneration Research Articles

Vascular Risk Factor Reduction is Associated with Corneal Nerve Regeneration in Patients with TIA and Ischemic Stroke

Vascular Risk Factor Reduction is Associated with Corneal Nerve Regeneration in Patients with TIA and Ischemic Stroke

Bioprinted Membranes for Corneal Tissue Engineering: A Review.

Corneal transplantation is considered a convenient strategy for various types of corneal disease needs. Even though it has been applied as a suitable solution for most corneal disorders, patients still face several issues due to a lack of healthy donor corneas, and rejection is another unknown risk of corneal transplant tissue. Corneal tissue engineering (CTE) has gained significant consideration as an efficient approach to developing tissue-engineered scaffolds for corneal healing and regeneration. Several approaches are tested to develop a substrate with equal transmittance and mechanical properties to improve the regeneration of cornea tissue. In this regard, bioprinted scaffolds have recently received sufficient attention in simulating corneal structure, owing to their spectacular spatial control which produces a three-cell-loaded-dimensional corneal structure. In this review, the anatomy and function of different layers of corneal tissue are highlighted, and then the potential of the 3D bioprinting technique for promoting corneal regeneration is also discussed.

Intra‐corneal implantation of 3D bio‐printed mesenchymal stem cells using femtosecond‐laser‐ assisted intrastromal keratoplasty

AbstractPurpose: Regeneration of the cornea due to trauma or disease is a complex biological process, involving a delicate balance between cell differentiation, migration and a multitude of secreted factors and signalling molecules. We hereby present a tissue‐engineering technique as a possible first step towards getting a bench‐to‐bedside approach to assist corneal regeneration and ultimately, corneal transplantation.Methods: 3D extrusion‐based bioprinting and femtosecond laser‐assisted intrastromal keratoplasty (FLISK) were used in the study. Ex vivo cultured mesenchymal stem cells (MSCs) derived from adipose tissue, bone marrow and corneal stroma were bioprinted in a complex, collagen I‐containing hydrogel. The transparent, cell‐loaded nanocellulose and alginate‐based 3D constructs were then implanted into a pig cornea organ culture model. The 3D printed intrastromal constructs were first cultivated ex vivo, while the de novo extracellular matrix (ECM) synthesis and migration of cells was followed by optical coherence tomography and immunohistochemistry.Results: The ex vivo cultured, MSC‐loaded 3D bioprinted constructs supported cell survival and propagation for all three types of MSCs used. The transparency of the corneal stroma substitute was not compromised by the presence of MSCs in the bioprinted matrix. At day 14, the 3D‐bioprinted scaffolds appeared intact, as analysed by optical coherence tomography, while immunofluorescent staining using ECM and MSCs markers demonstrated the cellular presence in the constructs.Conclusions: Optimization of the bioink composition, cellular content and properties of the bioprinted material to be used in corneal transplantation and regeneration studies are essential in fine‐tuning the technique towards its clinical use.

Cell therapy for corneal regeneration: Current status and future perspectives

AbstractCurrently, the treatment of many serious ocular pathologies that affect the cornea focuses on the repair and regeneration of the affected structures—epithelium and endothelium—through cell therapy. On the ocular surface, there is a population of multipotent progenitor cells (limbal stem cells; LSC), located in the sclerocorneal limbus, which have a high proliferative, differentiating and regenerative capabilities. On the other hand, the corneal endothelium—the innermost corneal layer with little regenerative capacity in vivo—is responsible for controlling corneal homeostasis and transparency. Dysfunctions of both cell types—corneal epithelial and endothelial cells—are the leading cause of recoverable corneal blindness. Mesenchymal stem cells (MSCs) from adult tissue, obtained from adipose tissue and bone marrow aspirates, have the capacity for adipogenesis, osteogenesis, chondrogenesis, myogenesis and neurogenesis in vitro. However, its true potential towards epithelial differentiation seems to be limited, and its great clinical applicability seems to lie in its contribution to the homeostasis of the cellular microenvironment. Recent developments in the field of somatic cell reprogramming by induced pluripotent stem cells (IPSC) provide an exciting possibility for its future clinical application. Pluripotent stem cells can be obtained from different sources of adult tissue, among which IPSC (induced pluripotent stem cells) have excellent plasticity. Its ability to differentiate into corneal epithelial and endothelial phenotypes is still poorly understood. Likewise, in vitro organoid production methods are presented as an option for obtaining compromised and/or partially differentiated cell lines. In the meantime, their clinical use will require specific application and receptor safety so that they can correctly regenerate tissue, with added effort for adaptation of ‘clinical grade’ protocols. In this sense, the ocular tissue is characterized as an excellent candidate for a possible clinical application, since it constitutes a tissue with immune privilege, it is ‘isolated’ (intraocular spaces) from the other components of the body and also had a highly sophisticated avascular (cornea) and highly differentiated (retina and pigment epithelium) tissues. The possibility of establishing specific clinical‐grade protocols for cell differentiation and identifying specific cellular and molecular behaviour would contribute to a better understanding of the mechanisms involved in repair processes. One of the equally relevant aspects in advanced cell‐based therapies applied to corneal regeneration is related to the ideal composition of the extracellular matrix proteins and biocompatible/biomimetic substrates—artificial or natural—that may contribute to better cell differentiation and adaptation. All this would open innovative perspectives for a more rational and effective treatment of different pathologies associated with human corneal blindness, in the field of regenerative medicine through the application of advanced cell therapy.

Use of new biomaterials with amniotic membrane growth factors in corneal regeneration in vivo

AbstractPurpose: To compare the effect on corneal healing of two novel hydrogels releasing proteins obtained from amniotic membrane (AM).Methods: Forty‐four New Zealand White rabbits underwent corneal wounding with NAOH 1 M and received different treatments: Control (group 1), AM transplantation (group 2), model A hydrogel (group 3) and model B hydrogel (group 4). The progression of wound healing was recorded with photographs and assessed by measuring the remaining epithelial defect over time. Histology was performed to examine acute and chronic changes.Results: There were no statistically significant differences between groups in epithelial defect areas (mm2) at days 0, 7, 14, 21 and 28. The percentage of wound closure (%) showed statistically significant differences on day 7 between Group 1 and Group 4 (p = 0.03). The percentage of completely healed corneas showed no statistically significant differences between the control and treated groups on days 14, 21 and 28, but on day 7, there were differences between Group 4 and Control (p = 0.02). On days 14 and 21 the rate of completely healed corneas was higher in Group 3 and Group 4 (55% on day 14 and 30% on day 21) compared to Group 1 (44.4% on day 14 and 11.1% on day 21).Conclusions: The novel biodegradable hydrogels with AM proteins could promote corneal re‐epithelialization. The chosen animal model showed a high rate of wound reopening once closed. Group 4 hydrogels showed low adhesiveness so the implant movement and the fragile regenerated epithelium could justify the higher de‐epithelized areas on day 7. The results show a higher number of wounds with complete epithelialization and a smaller epithelial defect area (mm2) in the treated groups compared to control, even after removal of the AM proteins, which might indicate that proteins secreted in the early stages of epithelialization could enhance corneal wound closure.

Nerve regeneration and corneal wound healing by ophthalmic formulations based on sodium hyaluronate, taurine, vitamin B6 and vitamin B12

AbstractPurpose: To evaluate the pharmacological profile of ocular formulations based on cross‐linked sodium hyaluronate (CL‐SH), taurine (Tau), Vit B6 and Vit B12 using in vitro and in vivo paradigms.Methods: Rabbit corneal epithelial cells (SIRC) were used to assess wound healing and ROS formation by scratch assay and oxidative stress (0.3 mM H2O2; 30 min), respectively w/ or w/o ocular formulations exposure. In vivo studies were carried out on albino rabbits to evaluate corneal nerve regeneration and corneal wound healing with or without treatment with six different formulations. Animals were anaesthetised, the corneal epithelium was removed, and formulations were topically administered (30 μl/eye; 3 times/day for 6 days). Slit‐lamp observation was carried out at the following day: T0, T2 and T5. After 6 days the animals were killed, and corneas were collected to evaluate corneal re‐innervation by immunohistochemistry of selective neuronal marker β‐III tubulin.Results: Formulations containing the concentrations 0.16% or 0.32% of CL‐SH, Tau, Vit B6 and B12 accelerated corneal wound healing. Cells exposed to H2O2 led to significant (p < 0.001) increase of ROS concentration that was significantly (p < 0.001) counteract by formulations containing CL‐SH (0.32%) and Tau w/ or w/o Vit B6 and B12. The extent of re‐innervation, in terms of β‐III tubulin staining, was 5‐fold greater (p < 0.01) in the eye of rabbits treated with formulation containing 0.32% CL‐SH, Tau, Vit B6 and Vit B12 (RenerviX®) compared with the control group (no treatment). Furthermore, re‐innervation with RenerviX® group was significantly greater (p < 0.01) also compared with the groups treated with the formulation containing 0.32% CL‐SH and Tau w/o vitamins and with the formulation containing 0.5% linear SH, Tau, and Vit B12.Conclusions: Among all formulations tested, the new ophthalmic gel RenerviX® was able to contrast oxidative stress, to accelerate corneal re‐epithelization and to promote nerve regeneration.

Ex vivo models of corneal epithelial regeneration in bioreactor: Respective roles of limbal, conjunctival and corneal epithelia

AbstractPurpose: The corneas preserved in bioreactor (BR) had been shown to have not only a better endothelial viability, but also a more differentiated and stratified epithelium than corneas preserved in organoculture. Purpose: By using BR, we would analyse the respective contribution of corneal (C), limbal (L), and conjunctival (Conj) epithelia in corneal epithelial regeneration.Methods: Five pairs of corneas from body donation to Science were used with a death‐to‐collection time < 20 h. A 3‐ to 5‐mm‐wide conjunctival flange was kept intact. Five patterns were set up by fully mechanical removal of 1, 2, or 3 epithelia: C‐L + Conj+, C‐L‐Conj+, C‐L + Conj‐, C + L‐Conj‐, C‐L‐Conj‐ (control) n = 2 for each pattern. The limbus was destroyed by scraping and thermocoagulation. Corneas were then kept in BR (21 mmHg, 2.5 μl/min of Corneamax Eurobio, 31°C) for 3 weeks to allow epithelial regeneration. The epithelium was then analysed using immunofluorescence (IF) on flat mounted cornea by targeting CK12 (corneal epithelium) and CK15 (limbal epithelium). Cell nuclei were counterstained with DAPI. Corneal transparency was quantified using a transparometer.Results: No epithelium was reconstituted in the controls. In the other 4 models including the C‐L‐Conj+ group, the cornea was transparent and covered by a pluristratified corneal epithelium, characterized by CK12 expression.Conclusions: In this BR model, conjunctival epithelial cells allowed the regeneration of a typical corneal epithelium in model C‐L‐Conj+. The corneal epithelium can also migrate to the limbus and conjunctiva. We hypothesize that all 3 ocular surface epithelia (including the conj) contain stem cells or progenitors capable of migrating throughout the cornea and restoring the corneal epithelium independently of each other. The main difference between our ex vivo model and in vivo situation is absence of neovascularization. This suggests that the main cause of limbic insufficiency is due to the loss of the anti‐angiogenic barrier rather than the loss of limbic stem cells.

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.

Corneal epithelial microcysts as side effect of belantamab mafodotin: Management and evolution

AbstractPurpose: Blenrep® (Belantamab mafodotin) is an antibody‐drug conjugate that targets an overexpressed antigen in myeloma plasma cells. This agent has its indication on adults with triple‐class refractory multiple myeloma (MM). The most frequently reported adverse event is the appearance of microcyst‐like epithelial changes in the cornea (MEC), produced by the apoptosis of epithelial cells. Our purpose is to describe these findings and their evolution.Methods: The evaluation of the MM patients treated with Blenrep® is a coordinated labor between ophthalmology and haematology departments. At the beginning of the treatment, a regulated baseline examination including visual acuity (VA) and slit lamp examination is performed and a cold mask and artificial tears are provided. Subsequently examination is repeated every 3 weeks, prior to the administration of the next cycle. The ophthalmological findings determine if the next dose can be administered or if it has to be postponed due to a VA loss. The evolution of three patients undergoing Blenrep® treatment was analysed, using anterior segment photography and OCT.Results: 31–92% of the patients on Blenrep® develop ocular events. In our case, the three patients studied developed epitheliopathy during the follow up. One of them, after 2 months, presented a VA loss due to MEC affecting visual axis that required next dose administration delay following haematologist's recommendation. After that period, the microcysts disappeared and VA was recovered. The other two patients also developed MEC, but did not need modification of the scheme since VA was not affected.Conclusions: Blenrep® has proved to be effective on the treatment of triple‐class refractory MM. Its principal adverse event is the appearance of MEC, which in some cases may impair the VA. This effect is reversible with the disappearance of the microcysts with the corneal epithelial regeneration cycle by prolonging the interval between the doses.

Injectable double-network hydrogel for corneal repair

Injuries to the cornea can cause corneal opacity or even blindness. Especially scald wounds and chemical burns to the cornea can usually lead to loss of corneal epithelium function and deep damage to the corneal stroma. Injectable hydrogel has been widely applied in corneal tissue engineering because it is easy to handle and can completely fill the defect area with minimally invasive surgical procedures. To reduce the risk and complications of penetrating keratoplasty and lamellar keratoplasty and to meet the demand for donated cornea, an injectable double-network (IDN) hydrogel based on methacrylated gelatin (Gel-MA) and oxidized hyaluronic acid (OHA) was constructed for focal corneal defect repairing. The Gel-MA/OHA IDN hydrogel had an interconnected porous microstructure and tunable mechanical properties with a modulus between 3.4 and 176.4 kPa. The hydrogel could promote cell adhesion and proliferation and afford the sustained release of hydrophilic/hydrophobic drugs. In-vivo corneal repair was performed by injecting the precursors into the pre-drilled epithelial-stromal defects of New Zealand White rabbits followed by immediate in-situ UV crosslinking. The results showed that the hydrogel had a strong capacity to promote corneal regeneration. Optical coherence tomography, slip lamp, fluorescein staining, scanning electron microscopy (SEM), and haematoxylin & eosin (H&E) staining were employed to illustrate the effectiveness and safety of corneal tissue regeneration. All the results showed that the optimized Gel-MA/OHA IDN hydrogel possessed excellent biocompatibility and could promote cornea repair in vivo. This study suggests that the injectable biocompatible double network hydrogel may provide a good choice for corneal repair and regeneration.

Oral Peroxisome Proliferator-Activated Receptor-α Agonist Enhances Corneal Nerve Regeneration in Patients With Type 2 Diabetes.

Diabetic corneal neuropathy (DCN) is a common complication of diabetes. However, there are very limited therapeutic options. We investigated the effects of a peroxisome proliferator-activated receptor-α (PPAR-α) agonist, fenofibrate, on 30 patients (60 eyes) with type 2 diabetes. On invivo confocal microscopy evaluation, there was significant stimulation of corneal nerve regeneration and a reduction in nerve edema after 30 days of oral fenofibrate treatment, as evidenced by significant improvement in corneal nerve fiber density (CNFD) and corneal nerve fiber width, respectively. Corneal epithelial cell morphology also significantly improved in cell circularity. Upon clinical examination, fenofibrate significantly improved patients' neuropathic ocular surface status by increasing tear breakup time along with a reduction of corneal and conjunctival punctate keratopathy. Tear substance P (SP) concentrations significantly increased after treatment, suggesting an amelioration of ocular surface neuroinflammation. The changes in tear SP concentrations was also significantly associated with improvement in CNFD. Quantitative proteomic analysis demonstrated that fenofibrate significantly upregulated and modulated the neurotrophin signaling pathway and linolenic acid, cholesterol, and fat metabolism. Complement cascades, neutrophil reactions, and platelet activation were also significantly suppressed. Our results showed that fenofibrate could potentially be a novel treatment for patients with DCN.

Synthetic Heparan Sulfate Mimetic Polymer Enhances Corneal Nerve Regeneration and Wound Healing after Experimental Laser Ablation Injury in Mice.

(1) Background: Abnormal corneal wound healing compromises visual acuity and can lead to neuropathic pain. Conventional treatments usually fail to restore the injured corneal tissue. In this study, we evaluated the effectiveness of a synthetic heparan sulfate mimetic polymer (HSmP) in a mouse model of corneal wound healing. (2) Methods: A surgical laser ablation affecting the central cornea and subbasal nerve plexus of mice was used as a model of the wound-healing assay. Topical treatment with HSmP was contrasted to its vehicle and a negative control (BSS). Corneal repair was studied using immunofluorescence to cell proliferation (Ki67), apoptosis (TUNEL assay), myofibroblast transformation (αSMA), assembly of epithelial cells (E-cadherin) and nerve regeneration (β-tubulin III). (3) Results: At the end of the treatment, normal epithelial cytoarchitecture and corneal thickness were achieved in HSmP-treated animals. HSmP treatment reduced myofibroblast occurrence compared to eyes irrigated with vehicle (p < 0.01) or BSS (p < 0.001). The HSmP group showed 50% more intraepithelial nerves than the BSS or vehicle groups. Only HSmP-treated corneas improved the visual quality to near transparent. (4) Conclusions: These results suggest that HSmP facilitates the regeneration of the corneal epithelium and innervation, as well as restoring transparency and reducing myofibroblast scarring after laser experimental injury.

Corneal stromal repair and regeneration.

The cornea is a specialized, transparent, avascular, immune-privileged, and heavily innervated tissue that affords 2/3rd of refraction to the eye. Ocular injuries, infections, and genetic factors affect corneal function and cause vision impairment. Presently, a variety of laser/non-laser surgeries, immunosuppressants, and/or corneal transplants are predominantly used to revive sight in human patients. The development of novel, precision-guided, and tissue-targeted non-surgical therapies promoting corneal repair and regeneration based on mechanistic understanding is of paramount importance to reduce the impact of global blindness. Research over the past decade revealed that modulation of pathological signaling pathways and factors by a variety of therapeutic delivery methods effectively treats corneal disorders including corneal scar/haze, inflammation, and angiogenesis in various pre-clinical animal models and are primed for human translation. This review discusses recent advances in the areas of corneal repair, restoration, and regeneration. Herein, we provide an overview of evolving approaches and therapeutic modalities that have shown great promise in reviving corneal transparency and function through the use of small drug molecules, gene therapy, nanomedicine, stem cells, trophic factors, exosomes, stromal equivalents, bioengineered stromal scaffolds, tissue adhesives, and 3D bioprinting.

Corneal Epithelial Regeneration: Old and New Perspectives.

Corneal blindness is the fifth leading cause of blindness worldwide, and therapeutic options are still often limited to corneal transplantation. The corneal epithelium has a strong barrier function, and regeneration is highly dependent on limbal stem cell proliferation and basement membrane remodeling. As a result of the lack of corneal donor tissues, regenerative medicine for corneal diseases affecting the epithelium is an area with quite advanced basic and clinical research. Surgery still plays a prominent role in the treatment of epithelial diseases; indeed, innovative surgical techniques have been developed to transplant corneal and non-corneal stem cells onto diseased corneas for epithelial regeneration applications. The main goal of applying regenerative medicine to clinical practice is to restore function by providing viable cells based on the use of a novel therapeutic approach to generate biological substitutes and improve tissue functions. Interest in corneal epithelium rehabilitation medicine is rapidly growing, given the exposure of the corneal outer layers to external insults. Here, we performed a review of basic, clinical and surgical research reports on regenerative medicine for corneal epithelial disorders, classifying therapeutic approaches according to their macro- or microscopic target, i.e., into cellular or subcellular therapies, respectively.

Hybrid nanostructured gadolinium oxide-collagen-dextran polymeric hydrogel for corneal repair and regeneration

A bio composite hydrogel containing collagen, dextran and rare earth metal nanoparticle, gadolinium oxide nanoparticle was prepared for corneal tissue regenerative application. Gadolinium oxide nanoparticles which are widely used in MRI scanning application are not much explored for tissue regenerative applications. The synthesized nanoparticles provided stability and therapeutic potential to collagen-dextran composite for achieving tissue regenerative potential. A good compatibility towards rabbit corneal fibroblast cells (SIRC) was observed with the hydrogel along with anti-proliferation property in aortic explants. The hydrogel down-regulated the VEGF-A gene expression in endothelial cells showing its ability to safe-guard the vision from corneal neovascularization. The expression of genes, ALDH1A1 and Vimentin needed for the well-being and enhanced proliferation of corneal cells was up-regulated upon treatment with the bio composite, while the expression of pro-inflammatory cytokines IL-6 and COX-2 was down regulated. Faster regeneration and migration of corneal cells into abrasion area was observed with the bio composite in vitro. The explant studies confirmed the compatibility of bio composites towards corneal tissue as well as lens crystallin proteins. The results suggest the potential use of rare-earth metal nanoparticles-based bio composite biomaterials in repairing and regenerating corneal cells when used in combination with protein- polysaccharide biomacromolecules.

Corneal Neuro-Regenerative Effect of Transcutaneous Electrical Stimulation in Rabbit Lamellar Keratectomy Model.

PurposeThis study aimed to evaluate the effect of transcutaneous electrical stimulation (TES) on corneal nerve regeneration in rabbits injured from superficial lamellar keratectomy (SLK).MethodsNew Zealand White rabbits were used in this experimental study. To induce corneal nerve damage, SLK was performed using a 7.0-mm trephine. TES was applied for 28 days after the corneal nerve injury. Corneal sensitivity, Western blotting, real-time polymerase chain reaction (PCR), and immunofluorescence were performed to observe changes in the corneal tissue.ResultsIn the 2-Hz and 20-Hz electrical stimulation groups, the degree of corneal wound healing increased by more than 10% compared to the control group, but no significant difference was observed. Conversely, the electrical stimulation (2-Hz or 20-Hz) group showed significantly increased corneal sensitivity compared to the control group. Western blot analysis revealed that small proline-rich protein 1A (SPRR1a), a regeneration-associated protein was significantly increased in the 2-Hz group on days 1 and 7 compared to that in the other groups. Once again, nerve regeneration in the 2-Hz group was supported by the results of PCR, in which a significant increase in the nerve growth factor (NGF) on day 1 was observed compared with the other groups. Moreover, immunofluorescence after 28 days of electrical stimulation showed significant nerve regeneration in the 2-Hz group.ConclusionsTES promoted corneal nerve regeneration in rabbit SLK model. The application of electrical stimulation of 2-Hz frequency was more effective than the 20-Hz frequency, showing potential clinical applications for corneal diseases.Translational RelevanceThis study shows how application of TES to the eyes that exhibit corneal nerve damage can improve corneal nerve regeneration examined by histologic analysis.

Expression Pattern of Tenascin-C, Matrilin-2, and Aggrecan in Diseases Affecting the Corneal Endothelium.

Purpose: The aim of this study was to examine the expression pattern of tenascin-C, matrilin-2, and aggrecan in irreversible corneal endothelial pathology such as pseudophakic bullous keratopathy (PBK) and Fuchs’ endothelial corneal dystrophy (FECD), which most frequently require corneal transplantation. Materials and methods: Histological specimens of corneal buttons removed during keratoplasty were investigated in PBK (n = 20) and FECD (n = 9) and compared to healthy control corneas (n = 10). The sections were studied by chromogenic immunohistochemistry (CHR-IHC) and submitted for evaluation by two investigators. Semiquantitative scoring (0 to 3+) was applied according to standardized methods at high magnification (400x). Each layer of the cornea was investigated; in addition, the stroma was subdivided into anterior, middle, and posterior parts for more precise analysis. In case of non-parametric distribution Mann–Whitney test was applied to compare two groups. Kruskal–Wallis and Dunn’s multiple comparisons tests have been applied for comparison of the chromogenic IHC signal intensity among corneal layers within the control and patient groups. Differences of p < 0.05 were considered as significant. Results: Significantly elevated tenascin-C immunopositivity was present in the epithelium and every layer of the stroma in both pathologic conditions as compared to normal controls. In addition, also significantly stronger matrilin-2 positivity was detected in the epithelium; however, weaker reaction was present in the endothelium in PBK cases. Minimal, but significantly elevated immunopositivity could be observed in the anterior and posterior stroma in the FECD group. Additionally, minimally, but significantly higher aggrecan immunoreaction was present in the anterior stroma in PBK and in the posterior stroma in both endothelial disorders. All three antibodies disclosed the strongest reaction in the posterior stroma either in PBK or in FECD cases. Conclusions: These extracellular matrix molecules disclosed up to moderate immunopositivity in the corneal layers in varying extents. Through their networking, bridging, and adhesive abilities these proteins are involved in corneal regeneration and tissue reorganization in endothelial dysfunction.

Research advances in human corneal endothelial cell regeneration

Human corneal endothelial cells (HCECs) are the prerequisite for maintaining corneal transparency, but HCECs remain arrested at the G1 phase after embryonic development and can not proliferate and regenerate. Thus, the density of HCECs decreases spontaneously with corneal development. Systemic factors, primary corneal disease, refractive factors, glaucoma, inflammation, and trauma all can cause a massive loss of HCECs, lead to corneal edema and turbidity, and ultimately induce blindness. Currently, keratoplasty is the only effective treatment, but the scarcity of donor corneas and the limitation of corneal preservation technology restrict the availability of keratoplasty. Therefore, the most appealing way to tackle the tissue shortage problem is corneal endothelial cell regeneration. In recent years, not only the endogenous regeneration of HCECs mediated by surgery, drugs and gene therapy but also the exogenous regeneration of HCECs mediated by cell therapy have made fruitful progress. Although a number of regeneration strategies have entered the clinical trial stage, the wide clinical application of corneal endothelial regeneration is still far away. This review elaborates the basic research, clinical application and limitation of current strategies of corneal endothelial cell regeneration.

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.