Conjunctival epithelial cells resist productive SARS-CoV-2 infection.

Conjunctival epithelial cells, which express viral-entry receptors angiotensin-converting enzyme 2 (ACE2) and transmembrane protease serine type 2 (TMPRSS2), constitute the largest exposed epithelium of the ocular surface tissue and may represent a relevant viral-entry route. To address this question, we generated an organotypic air-liquid-interface model of conjunctival epithelium, composed of basal, suprabasal, and superficial epithelial cells, and fibroblasts, which could be maintained successfully up to day 75 of differentiation. Using single-cell RNA sequencing (RNA-seq), with complementary imaging and virological assays, we observed that while all conjunctival cell types were permissive to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genome expression, a productive infection did not ensue. The early innate immune response to SARS-CoV-2 infection in conjunctival cells was characterised by a robust autocrine and paracrine NF-κB activity, without activation of antiviral interferon signalling. Collectively, these data enrich our understanding of SARS-CoV-2 infection at the human ocular surface, with potential implications for the design of preventive strategies and conjunctival transplantation.

Use of biomaterials in corneal endothelial repair.

Human corneal endothelium (HCE) is a single layer of hexagonal cells that lines the posterior surface of the cornea. It forms the barrier that separates the aqueous humor from the rest of the corneal layers (stroma and epithelium layer). This layer plays a fundamental role in maintaining the hydration and transparency of the cornea, which in turn ensures a clear vision. In vivo, human corneal endothelial cells (HCECs) are generally believed to be nonproliferating. In many cases, due to their nonproliferative nature, any damage to these cells can lead to further issues with Descemet's membrane (DM), stroma and epithelium which may ultimately lead to hazy vision and blindness. Endothelial keratoplasties such as Descemet's stripping automated endothelial keratoplasty (DSAEK) and Descemet's membrane endothelial keratoplasty (DEK) are the standard surgeries routinely used to restore vision following endothelial failure. Basically, these two similar surgical techniques involve the replacement of the diseased endothelial layer in the center of the cornea by a healthy layer taken from a donor cornea. Globally, eye banks are facing an increased demand to provide corneas that have suitable features for transplantation. Consequently, it can be stated that there is a significant shortage of corneal grafting tissue; for every 70 corneas required, only 1 is available. Nowadays, eye banks face long waiting lists due to shortage of donors, seriously aggravated when compared with previous years, due to the global COVID-19 pandemic. Thus, there is an urgent need to find alternative and more sustainable sources for treating endothelial diseases, such as utilizing bioengineering to use of biomaterials as a remedy. The current review focuses on the use of biomaterials to repair the corneal endothelium. A range of biomaterials have been considered based on their promising results and outstanding features, including previous studies and their key findings in the context of each biomaterial.

In the eye of the storm: SARS-CoV-2 infection and replication at the ocular surface?

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) first emerged in December 2019 and spread quickly causing the coronavirus disease 2019 (COVID-19) pandemic. Recent single cell RNA-Seq analyses have shown the presence of SARS-CoV-2 entry factors in the human corneal, limbal, and conjunctival superficial epithelium, leading to suggestions that the human ocular surface may serve as an additional entry gateway and infection hub for SARS-CoV-2. In this article, we review the ocular clinical presentations of COVID-19 and the features of the ocular surface that may underline the overall low ocular SARS-CoV-2 infection. We critically evaluate the studies performed in nonhuman primates, ex vivo organ culture ocular models, stem cell derived eye organoids and the differences in infection efficiency observed in different parts of human ocular surface epithelium. Finally, we highlight the additional work that needs to be carried out to understand the immune response of the ocular surface to SARS-CoV-2 infection, which can be translated into prophylactic treatments that may be applied to other organ systems.

Co-expression of SARS-CoV-2 entry genes in the superficial adult human conjunctival, limbal and corneal epithelium suggests an additional route of entry via the ocular surface.

PURPOSE: The high infection rate of SARS-CoV-2 necessitates the need for multiple studies identifying the molecular mechanisms that facilitate the viral entry and propagation. Currently the potential extra-respiratory transmission routes of SARS-CoV-2 remain unclear. METHODS: Using single-cell RNA Seq and ATAC-Seq datasets and immunohistochemical analysis, we investigated SARS-CoV-2 tropism in the embryonic, fetal and adult human ocular surface. RESULTS: The co-expression of ACE2 receptor and entry protease TMPRSS2 was detected in the human adult conjunctival, limbal and corneal epithelium, but not in the embryonic and fetal ocular surface up to 21 post conception weeks. These expression patterns were corroborated by the single cell ATAC-Seq data, which revealed a permissive chromatin in ACE2 and TMPRSS2 loci in the adult conjunctival, limbal and corneal epithelium. Co-expression of ACE2 and TMPRSS2 was strongly detected in the superficial limbal, corneal and conjunctival epithelium, implicating these as target entry cells for SARS-CoV-2 in the ocular surface. Strikingly, we also identified the key pro-inflammatory signals TNF, NFKß and IFNG as upstream regulators of the transcriptional profile of ACE2+TMPRSS2+ cells in the superficial conjunctival epithelium, suggesting that SARS-CoV-2 may utilise inflammatory driven upregulation of ACE2 and TMPRSS2 expression to enhance infection in ocular surface. CONCLUSIONS: Together our data indicate that the human ocular surface epithelium provides an additional entry portal for SARS-CoV-2, which may exploit inflammatory driven upregulation of ACE2 and TMPRSS2 entry factors to enhance infection.

One threat, different answers: the impact of COVID-19 pandemic on cornea donation and donor selection across Europe.

OBJECTIVES: To assess to which extent the COVID-19 pandemic affected corneal transplantation by virtue of donor selection algorithms in different European countries. DESIGN: Survey. SETTING: 110 eye banks in 26 European countries. PARTICIPANTS: 64 eye banks covering 95% of European corneal transplantation activity. INTERVENTIONS: A questionnaire listing the number of corneas procured and distributed from February to May 2018-2020 was circulated to eye banks. MAIN OUTCOME MEASURES: The primary outcome was the number of corneal procurements. Additional outcomes were national algorithms for donor selection, classified according to their stringency (donors with COVID-19 history, suspected for COVID-19, asymptomatic, PCR testing) and the pandemic severity in each country. We calculated Spearman's correlation coefficient to determine, two by two, the relationship between the 3-month decline in eye banking activity (procurement), the stringency of donor selection algorithm and the grading of pandemic severity (cases and deaths). A partial correlation was run to determine the relationship between decline and stringency while controlling for pandemic severity. RESULTS: Procurements decreased by 38%, 68% and 41%, respectively, in March, April and May 2020 compared with the mean of the previous 2 years, while grafts decreased, respectively, by 28%, 68% and 56% corresponding to 3866 untreated patients in 3 months. Significant disparities between countries and the decrease in activity correlated with stringency in donor selection independent of pandemic severity. CONCLUSIONS: Our data demonstrate significant differences between countries regarding donor screening algorithms based on precautionary principles and, consequently, a decrease in the donor pool, already constrained by a long list of contraindications. Fundamental studies are needed to determine the risk of SARS-CoV-2 transmission by corneal transplantation and guide evidence-based recommendations for donor selection to justify their substantial medical and economic impact.