Abstract

Previously we reported that hydrogen sulfide (H2S) donor can mitigate hyperhomocysteinemia (HHcy)‐mediated cardiomyocyte hypertrophy by upregulating miR‐133a. However, the dose dependent effect of Hcy and H2S on miR‐133a level, the effect of homocysteine metabolizing enzyme cystathionine‐β‐synthase (CBS) on cystathionine‐ϒ‐lyase (CTH) (a precursor enzyme for H2S biosynthesis), and the role of H2S in CBS and CTH regulation in cardiomyocytes, are poorly understood. We hypothesized that H2S and homocysteine (Hcy) influence CBS and CTH expression in a dose dependent manner, and H2S controls CBS‐CTH feedback regulation by inducing miR‐133a and suppressing SP1, a transcriptional activator of CTH promoter, in cardiomyocytes. We treated HL1 cardiomyocytes with 0, 5, 25, 50, 75 and 100 μmL−1 doses of Hcy or H2S donor Na2S for 24 hours. Our results demonstrated that 50 μmL−1 Na2S upregulates miR‐133a and downregulates CTH, whereas 25 μmL−1 Hcy downregulates miR‐133a and upregulates CTH. To determine the in vivo effect of CBS on CTH, we measured the cardiac levels of CBS and CTH in CBS+/− mice, a model for HHcy. We found that in CBS+/− hearts, CBS is downregulated but CTH is upregulated, suggesting a negative feedback regulation between CBS and CTH. To assess the effect of H2S on CTH, we measured the levels of SP1 in Na2S‐treated cardiomyocytes. Our results revealed that Na2S treatment suppresses SP1 in a dose dependent manner, suggesting that H2S may downregulate CTH by inhibiting SP1. Moreover, our luciferase reporter assay on CTH 3′UTR demonstrated that miR‐133a targets CTH. Since H2S upregulates miR‐133a, it could be an indirect mechanism for H2S‐mediated regulation of CTH. In conclusion, H2S and Hcy regulate CBS and CTH in a dose dependent manner. CBS negatively regulates CTH. H2S upregulates CBS but downregulates CTH, possibly by suppressing SP1 and inducing miR‐133a in cardiomyocytes.