From bench to clinic: Emerging therapies for corneal scarring.

Corneal diseases are one of the leading causes of moderate-to-severe visual impairment and blindness worldwide, after glaucoma, cataract, and retinal disease in overall importance. Given its tendency to affect people at a younger age than other blinding conditions such as cataract and glaucoma, corneal scarring poses a huge burden both on the individuals and society. Furthermore, corneal scarring and fibrosis disproportionately affects people in poorer and remote areas, making it a significant ophthalmic public health problem. Traditional medical strategies, such as topical corticosteroids, are not effective in preventing fibrosis or scars. Corneal transplantation, the only effective sight-restoring treatment for corneal scars, is curbed by challenges including a severe shortage of tissue, graft rejection, secondary conditions, cultural barriers, the lack of well-trained surgeons, operating rooms, and well-equipped infrastructures. Thanks to tremendous research efforts, emerging therapeutic options including gene therapy, protein therapy, cell therapy and novel molecules are in development to prevent the progression of corneal scarring and compliment the surgical options currently available for treating established corneal scars in clinics. In this article, we summarise the most relevant preclinical and clinical studies on emerging therapies for corneal scarring in recent years, showing how these approaches may prevent scarring in its early development.

Do Corneal Tissue Providers Inform Their Community That They Export Corneas? An Audit of Publicly Available Sector Websites.

AIM: The exportation of corneas from one nation to another, for transplantation services, is responsible for 23% of all global transplants. Global allocation is possible because of the end-of-life donations from citizens and residents of export nations. To date, there is no information indicating if export nation donors are aware that their corneas may be exported, nor if organizations that export provide information regarding their export engagement to their community. To ascertain if and how exporters inform their community, we audited known export organization public websites. MATERIALS AND METHODS: We designed and conducted a double-blind audit of known exporting eye banks, eye tissue sharing and distributor organization websites. RESULTS: We audited 79 websites, from 9 nations. Of the 79, 46 (58.2%) did not mention corneal tissue exportation, 17 (21.5%) implied exportation, and 16 (10.2%) explicitly mentioned it. Of the 16 that mentioned they exported, 75% (12/16) provided information regarding their export license, and 12.5% (2/16) indicated partnership with a third party. We could not locate information explaining how organizations decided on how and to whom they export. DISCUSSION: Organizations that export corneal tissue across national borders do not share sufficient information regarding their export activities on their website. The general public and donors within export nations may not be aware that this practice occurs or could occur with their donation. Export organizations and the eye tissue sector must evaluate their communication strategies and collaborate, preferably nationally, to develop publicly appropriate information regarding corneal tissue exportation.

Successful Treatment of Mycobacterium chelonae Keratitis Within a Corneal Transplant Using Intrastromal Amikacin Injections-A Case Report Demonstrating the Fundamental Principles and Challenges of Infective Keratitis Management and Novel Therapeutic Approaches.

Mycobacterium chelonae keratitis is rare and difficult to treat. This is the first known case worldwide of effective treatment using intrastromal amikacin injections in a corneal transplant recipient who had metastatic breast cancer. The challenges and principles of management, applicable to other causes of infective keratitis, are reviewed.

Postoperative detection of unusual pathology in donor corneal tissue.

We present a series of three patients with previously undetected corneal pathology in grafted corneal tissue following keratoplasty for keratoconus. Postoperatively, a faint layer of anterior stromal haze involving the graft was observed in each patient upon slit lamp examination. Anterior segment optical coherence tomography (AS-OCT) confirmed the presence of anterior stromal scarring across the transplanted cornea. However, the ocular and systemic medical histories of the donors were unremarkable. As the suboptimal donor corneal tissue may escape the standard screening protocols, eye banks should consider adding AS-OCT imaging for screening donor corneal tissue before transplantation.

A single-cell transcriptome atlas of the adult human retina.

The retina is a specialized neural tissue that senses light and initiates image processing. Although the functional organization of specific retina cells has been well studied, the molecular profile of many cell types remains unclear in humans. To comprehensively profile the human retina, we performed single-cell RNA sequencing on 20,009 cells from three donors and compiled a reference transcriptome atlas. Using unsupervised clustering analysis, we identified 18 transcriptionally distinct cell populations representing all known neural retinal cells: rod photoreceptors, cone photoreceptors, Müller glia, bipolar cells, amacrine cells, retinal ganglion cells, horizontal cells, astrocytes, and microglia. Our data captured molecular profiles for healthy and putative early degenerating rod photoreceptors, and revealed the loss of MALAT1 expression with longer post-mortem time, which potentially suggested a novel role of MALAT1 in rod photoreceptor degeneration. We have demonstrated the use of this retina transcriptome atlas to benchmark pluripotent stem cell-derived cone photoreceptors and an adult Müller glia cell line. This work provides an important reference with unprecedented insights into the transcriptional landscape of human retinal cells, which is fundamental to understanding retinal biology and disease.

Ethical Issues in Transnational Eye Banking.

PURPOSE: To review ethical issues that may arise in the setting of transnational eye banking activities, such as when exporting or importing corneal tissue for transplantation. METHODS: A principle-based normative analysis of potential common dilemmas in transnational eye banking activities was performed. RESULTS: Transnational activities in eye banking, like those in other fields involving procurement and use of medical products of human origin, may present a number of ethical issues for policy makers and professionals. Key ethical concerns include the potential impact of export or import activities on self-sufficiency of corneal tissue supply within exporting and importing countries; potential disclosure requirements when obtaining consent or authorization for ocular tissue donation when donations may be exported; and difficulties inherent in assuring equity in the allocation of tissues available for export and in establishing and respecting standards of safety and quality across different jurisdictions. CONCLUSIONS: Further analysis of specific ethical issues in eye banking is necessary to inform development of guidelines and other governance tools that will assist policy makers and professionals to support ethical practice.

The influence of Australian eye banking practices on corneal graft survival.

OBJECTIVE: To identify eye banking practices that influence corneal graft survival. DESIGN, SETTING AND PARTICIPANTS: Prospective cohort study of records of 19,254 followed corneal grafts in 15160 patients, submitted to the Australian Corneal Graft Registry between May 1985 and July 2012. MAIN OUTCOME MEASURES: Influence of corneal preservation method (organ culture, moist pot, Optisol, other); death-to-enucleation, death-to-preservation and enucleation-to-graft times; transportation by air; graft era; and indication for graft on probability of graft survival at most recent follow-up. RESULTS: In multivariate analysis, 919 penetrating grafts performed using corneas transported interstate by air exhibited worse survival than 14,684 grafts performed using corneas retrieved and used locally (hazard ratio [HR], 1.44; 95% CI, 1.21-1.73; P = 0.001). This was also the case for traditional lamellar grafts (64 corneas transported by air and 813 used locally; HR, 1.69; 95% CI, 1.03-2.78; P = 0.038). Indication for graft influenced survival of penetrating grafts (4611 keratoconus, 727 emergency or high-risk, 10,265 other indication; global P < 0.001) and traditional lamellar grafts (65 keratoconus, 212 emergency or high-risk, 600 other indication; global P < 0.001). The preservation medium in which corneas used for traditional lamellar grafts were stored exerted a marginal influence on graft survival (global P = 0.047). CONCLUSIONS: Donor corneas transported interstate exhibited poorer survival after transplantation than those retrieved and grafted locally. Higher proportions of emergency procedures involving transported corneas did not account for this difference. Where possible, efforts to avoid transportation of corneal tissue by air freight within Australia may be warranted.

Localized changes in stromal reflectivity after corneal collagen cross-linking observed with different imaging techniques.

PURPOSE: To report changes observed in the corneal stroma using confocal microscopy and Scheimpflug imaging following epithelium-off collagen cross-linking (CXL) in cases of progressive keratoconus. METHODS: Fifteen eyes of 14 patients were examined before and after CXL using slit-lamp biomicroscopy, Scheimpflug imaging, and confocal microscopy. A subset of patients also had optical coherence tomography imaging performed. RESULTS: After CXL, confocal microscopy revealed a discrete layer of increased reflectivity in the mid to deep stroma. This layer was visible from as early as 1 week following treatment with patchy changes persisting until 12 months. This posterior reflective zone was found to consist of numerous linear or striate reflective structures (intrastromal striate reflections [ISRs]). ISRs were not observed in any of the eyes before the CXL procedure. High-resolution Scheimpflug and optical coherence tomography images also demonstrated a narrow zone of increased reflectivity at a similar depth. A layer-by-layer match was possible between the confocal and Scheimpflug images. The location of the ISR layer appears to correlate with a zone of increased reflectivity visible on postoperative slit-lamp examinations. CONCLUSION: Altered stromal reflectivity after CXL can be observed with modern diagnostic imaging technologies. These findings seem to correlate not only among the different devices but also with biomicroscopic observations and could potentially provide a non-invasive tool to monitor the cross-linking effect in individual corneas.

Histopathological evaluation of anterior lamellar corneal tissue-on/-off storage conditions on DSAEK donor tissue after storage in organ culture.

PURPOSE: To compare the quality of corneal endothelium of precut Descemet-stripping automated endothelial keratoplasty (DSAEK) tissue when transported with and without the anterior lamellar corneal tissue (ALCT) when organ-culture corneal storage methods are used. METHODS: After microkeratome-assisted excision of anterior corneal lamella, five pairs of corneas (10 eyes) were stored either with the ALCT on the stroma (five eyes) or with ALCT off the stroma (five eyes) in organ-culture medium. Three pairs (six matched corneas) were left in the transport medium for 24 h prior to the microkeratome cut. Two pairs (four matched corneas) were left in the transport medium for 48 h prior to the microkeratome cut. All cuts were performed using a 300 (four eyes) or 350 (six eyes) microns head. A vital dye assay (alizarin red S and trypan blue) was used to identify devitalized and necrotic endothelial cells. RESULTS: In all matched cases, there was no difference between the endothelial cell appearance with or without the anterior corneal lamella. In all cases, there was no evidence for trypan blue stained cells beyond that normally seen on acceptable transplantable corneas and there was no evidence of loss of cells or any lifting of Descemet's membrane. CONCLUSIONS: There is no difference between the quality of the donor endothelial cell appearance with ALCT-on or -off when the donor cornea is stored using the organ-culture system of corneal storage. Organ-culture storage system is a safe and effective system in regard to preparation and transport of donor lenticules for DSAEK.

In vivo effects of fluoroquinolones on rabbit corneas.

PURPOSE: The use of topical fluoroquinolones to treat microbial keratitis is associated with an increased incidence of corneal perforation compared to other standard treatments. This study examined the effects of topical fluoro-quinolones on corneal collagen and keratocytes in intact rabbit corneas and corneas with an epithelial defect. METHODS: Studies consisted of one group of intact corneas and one group of corneas where a 6-mm epithelial defect was created with a surgical scrape. Within each group, eyes were randomly assigned to one of four topical medications (0.3% ciprofloxacin, 0.3% ofloxacin, fortified antibiotics (1.36% tobramycin, 5% cefrazolin) or Tears Naturale (Alcon Laboratories, Frenchs Forest, NSW, Australia). Two drops were instilled hourly for 48 h and then 2-hourly for an additional 48 h. At 96 h the corneas were removed and processed for light microscopy, immunohistology for collagen IV, V and VI, and apoptosis staining. RESULTS: In intact rabbit corneas there was no demonstrable difference between treatment groups. In corneas with an epithelial defect, both fluoroquinolones delayed epithelial healing when compared to fortified antibiotics or tears. Keratocyte loss was seen in all groups and was greatest in the ofloxacin group. Median stromal thickness with keratocyte loss were: ofloxacin 30%; ciprofloxacin 10%; fortified antibiotics 7.5%; and tears 15% (ofloxacin vs tears, Mann-Whitney = 16.0, P = 0.09). Keratocyte loss did not correlate with the amount of demonstrable apoptosis. Collagens IV, V and VI showed no differences between treatments. CONCLUSIONS: These results suggest that ofloxacin is potentially cytotoxic to corneal keratocytes. Such an effect could lead to the observed increased incidence of corneal perforation in microbial keratitis.

TrkB receptor signaling regulates developmental death dynamics, but not final number, of retinal ganglion cells.

We investigated the effects of endogenous neurotrophin signaling on the death-survival of immature retinal ganglion cells (RGCs) in vivo. Null mutation of brain-derived neurotrophic factor [(BDNF) alone or in combination with neurotrophin 4 (NT4)] increases the peak rate of developmental RGC death as compared with normal. Null mutation of NT4 alone is ineffective. Null mutation of the full-length trkB (trkBFL) receptor catalytic domain produces a dose-dependent increase in the peak RGC death rate that is negatively correlated with retinal levels of trkBFL protein and phosphorylated (activated) trkBFL. Depletion of target-derived trkB ligands by injection of trkB-Fc fusion protein into the superior colliculus increases the peak rate of RGC death compared with trkA-Fc-treated and normal animals. Adult trkBFL+/- mice have a normal number of RGCs, despite an elevated peak death rate of immature RGCs. Thus, target-derived BDNF modulates the dynamics of developmental RGC death through trkBFL activation, but BDNF/trkB-independent mechanisms determine the final number of RGCs.

Complexity in the modulation of neurotrophic factor mRNA expression by early visual experience.

The expression of mRNA for brain-derived neurotrophic factor (BDNF) is regulated by early visual experience. In this study, we sought to determine whether other neurotrophic factor mRNAs are similarly regulated. We reared pigmented rats from birth to postnatal day 21 in a normal light cycle, constant light (LR) or constant darkness (DR). In the retina, superior colliculus (SC), primary visual cortex (V1), hippocampus (HIPP) and cerebellum (CBL), using a ribonuclease protection assay (RPA), we examined expression of the mRNAs for nerve growth factor (NGF), BDNF, NT3, NT4, ciliary neurotrophic factor (CNTF) and glial cell line-derived neurotrophic factor (GDNF). LR or DR alter the expression of the mRNAs for NGF, BDNF and NT3 and CNTF within the visual system. LR also upregulated BDNF mRNA expression within the cerebellum. In all of the structures examined, NT4 mRNA expression was unaltered by LR or DR and GDNF mRNA was undetectable. Notably, the same rearing condition could induce changes of opposite sign in the mRNA for a single factor in different structures or for different factors in the same structure. Thus, during developmental stages when sensory experience and neuroelectric activity are important in the shaping of visual circuitry, vision regulates the expression of multiple neurotrophic factor mRNAs and each mRNA has a unique profile with respect to the locus and sign of activity-induced changes.