Differential innate immune response and replication of SARS-CoV-2 in human diabetic corneal epithelial cells

ABSTRACT

Purpose: Diabetes predisposes an individual to severe COVID-19. Diabetic cornea is also known to have impaired wound healing, increasing the chances of infection. Earlier, we reported the ability of SARS-CoV-2 to infect conjunctival cells, and the presence of viral RNA and proteins was also detected in the corneas of COVID-19 donors. In this study, we evaluated the effect of diabetes on corneal innate immune response during SARS-CoV-2 infection and sought to determine the underlying mechanisms. Methods: Human primary corneolimbal epithelial cells (HCECs) were isolated from the corneas of three diabetic and three non-diabetic donors. In vitro studies were performed by infecting HCECs with SARS-CoV-2-USA-WA1/2020 strain at MOI 0.5. Viral replication was assessed by viral genome copy number. RNAseq analysis was performed to determine genes/pathways altered by diabetic vs non-diabetic HCECs. qPCR was used to assess the expression of innate inflammatory and antiviral genes. Western blot was performed to detect the protein expression of antiviral signaling molecules. Results: The primary HCECs were found permissive to SARS-CoV-2 infection, as evidenced by increased viral replication which peaked at day 3 p.i. along with an induction of pSTAT1. Interestingly, HCECs from diabetic cornea had higher viral RNA on all three days post-infection. SARS-CoV-2 infected HCECs exhibited induced expression of inflammatory genes and their levels were relatively higher in diabetic cells. RNA-seq analysis revealed significant differences in diabetic vs. non-diabetic SARS-CoV-2 infected cells with alteration in genes regulating viral response, inflammation, and injury. The most affected down-regulated genes are related to lipid metabolism, ferroptosis, and oxidative stress. Conclusions: Our study demonstrates increased SARS-CoV-2 replication and differential innate antiviral and inflammatory response in HCECs from diabetic corneas. These results indicate that diabetes is a potential risk for enhanced infectivity of SARS-CoV-2 for the ocular surface.