H₂S induces dose-dependent changes in inflammatory and apoptotic responses in human corneal stromal fibroblasts in vitro

Abstract

Purpose : Earlier we optimized an in vitro model for studying hydrogen sulfide (H₂S) toxicity to the human cornea. This study examined if H₂S toxicity to corneal stromal fibroblasts is due to (a) changes in inflammatory and apoptotic parameters and (b) whether the effects are concentration-dependent using an established in vitro model.

Methods : Healthy donor human corneas (n=30) were obtained from the eye bank and used to generate primary human corneal stromal fibroblasts (hCSFs) following our standard protocol. Sodium hydrosulfide (NaSH) was used as a source of H₂S. Primary hCSFs were exposed to different concentrations of NaSH (1 mM - 10 mM) for 24h. The changes in inflammatory and apoptotic parameters were assessed using quantitative real-time PCR, mitochondrial membrane potential (ΔΨm), and TUNNEL assays.

Results : H₂S toxicity to hCSFs is dose-dependent. Exposure to lower concentrations (1 - 4 mM) of NaSH showed alteration of inflammatory genes IL-1α, IL-1β, and TNF-α (p≤0.05). However, higher concentrations (5 - 10 mM) of NaSH exposure leads to alterations in inflammatory genes IL-1α, IL-1β, and TNF-α as well as apoptotic genes caspase 8 and FADD and showed loss of mitochondrial membrane potential and significantly increased TUNEL positive cells as compared to the non-H₂S exposed hCSFs (p≤0.05).

Conclusions : H₂S causes dose-dependent toxicity and changes in inflammatory and apoptotic mediaters in hCSFs in vitro. Onging studies will provide additional mechanistic information.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.