EFFECT OF ELEVATED PERFUSION PRESSURE ON THE PRODUCTION OF HYDROGEN SULFIDE AND PROSTAGLANDINS IN PORCINE ANTERIOR SEGEMENT EXPLANTS

Abstract

Purpose : We have evidence that that elevated perfusion pressure can increase the release of hydrogen sulfide (H₂S) in the perfusate of an ex vivo model of the porcine anterior segment (POAS) explant, a response that was blocked by inhibitors of cyclooxygenase (COX) (Robinson et al. FASEB J. 30:1271.5, 2016). In the present study, we investigated the role of endogenously produced H₂S during elevated perfusion pressure in POAS explants. Furthermore, we also assessed the effect of elevated perfusion pressure on the release of prostaglandins (PGs) in the POAS explant.

Methods : Porcine ocular anterior segment explants were perfused with Dulbecco’s Modified Eagle’s Medium maintained at 37° C and gassed with carbogen, under an elevated pressure of 15 mm Hg for four hours. Perfusates from the anterior segment explants were collected and immediately assayed for their H₂S content using Methylene Blue assay or the concentrations of PGE₂ using an ELISA kit. Explants were perfused with inhibitors of cystathionine β-synthase, CBS/cystathionine-γ-lyase, CSE (aminooxyacetic acid, AOAA) or 3-mercaptopyruvate sulfurtransferase, 3MST (ketobutyric acid, KBA) or with flurbiprofen (30 µM).

Results : Elevating perfusion pressure in POAS explants significantly (p < 0.001) increased H₂S concentrations in the perfusate from 0.4 ± 0.1 to 67.6 ± 3.6 nM/µg protein (n = 6). AOAA (30 µM) significantly (p < 0.001) reduced the effects of elevated pressure on H₂S levels from 67.6 ± 3.6 to 5.7 ± 0.3 nM/µg protein (n = 6) while KBA (1 mM), significantly (p < 0.001) reduced the effects of elevated pressure on H₂S levels to 4.9 ± 0.8 nM/µg protein (n = 6). Elevating perfusion pressure in the POAS explants significantly (p < 0.001) lowered PGE₂ levels in the perfusate by 53%. Both AOAA (30 µM) and KBA (1 mM) had no significant (p > 0.05) effect on PGE₂ levels in the perfusate. On the other hand, flurbiprofen (30 µM) significantly lowered PGE₂ levels in the perfusate of POAS explants under elevated pressure by 30%.

Conclusions : We conclude that the increase in H₂S concentrations in the perfusate of POAS explants under elevated perfusion pressure depends upon the endogenous biosynthesis of this gas. The decrease in the release of PGE₂ in explants under elevated perfusion pressure appears to depend, at least in part, to the biosynthesis of endogenous prostanoids.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.