Hypoxia inducible factor-1 regulates spermine oxidase leading to acrolein generation in Müller Glial cells

Abstract

Purpose : We previously reported that hypoxia induces the expression of spermine oxidase (SMOX) that catalyzesthe oxidation of spermine to spermidine, followed by hydrogen peroxide (H₂O₂) generation in rat retinal glial cells. The purpose of this study was to further investigate the production regulatory mechanism and the role of SMOX in retinal glial cells under hypoxic condition.

Methods : TR-MUL5 cells, immortalized rat Müller cell line, were cultured under normoxic (20% O₂) or hypoxic (1% O₂) condition. The gene and protein levels of SMOX were quantified by real-time PCR and enzyme-linked immunosorbent assay, after treated with echinomycin, an inhibitor of hypoxia inducible factor-1 (HIF-1), or vehicle. The localization of SMOX was detected in TR-MUL5 cells using immunofluorescence under normoxic or hypoxic condition. In addition, level of acrolein-conjugated protein (FDP-Lys) was measured in TR-MUL5 cells treated with or without SMOX inhibitor MDL72527.

Results : Hypoxia significantly increased the mRNA (1.9-fold) and protein (1.5-fold) levels of SMOX in TR-MUL5 (p<0.01), while echinomycin treatment abrogated the SMOX synthesis (p<0.01). The SMOX positive signals were apparently increased in the cytoplasm after hypoxic stimulation compared with normoxic condition. MDL72527 significantly suppressed the FDP-Lys production, which showed 3.0-fold increase in TR-MUL5 stimulated with hypoxia.

Conclusions : The current data indicate that HIF-1 regulates SMOX production, leading to acrolein generation in glial cells under hypoxic condition.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.