The Correlated Research About Cystathionine Gamma-lyase By Sodium Hydrosulfide And Pioglitazone

Objectives: To study the influence of exogenous sodium hydrosulfide，homocysteine and pioglitazone on cystathionine-γ-lyase in human umbilicalvein endothelial cells (HUVECs) and make clear cardiovascular effects ofendogenous hydrogen sulfide further.Methods:(1) HUVECs were cultured in vitro and divided into thedifferent groups. After being treated with various amounts, Morphologicalchanges in the HUVECs were observed by microscopy and the status ofHUVECs Cell viability was determined by Cell Counting Kit-8assay.(2)Appropriate primers were designed to analyze the mRNA levels of CSEexpression using Real time quantitative PCR.(3) HUVECs were harvestedinto lysate buffer with protease inhibitor PMSF. After determination of theprotein concentration using the BCA protein assay kit, proceeding WesternBlot Analysis.(4) CSE enzyme activity in cell was measured by asulfur-sensitive electrode, essentially as described elsewhere.Results:(1) High concentration Hcy induced cell death.There was aconcentration-dependent decrease in Cell viability after HUVECs wereincubated with Hcy groups no matter24h or48h, compared with control. Itwas accompanied by a significant decrease in the expression of CSE mRNAand protein levels. Moreover, it was accompanied by a decrease in CSEenzyme activity.(2) By the addition of NaHS concentrations (10μM～30μM),There was a increase in Cell viability levels. After HUVECs were incubatedwith NaHS concentrations （50μM～1000μM）, cell viability significantly decreased compared with control.(3) Compared with control groups, whenHUVECs were incubated with30μM NaHS and1000μM Hcy group incommon, and There was a concentration-dependent increase in Cell viability,CSE mRNA expression level and protein expression level. Furthermore, it isincreased in CSE enzyme activity.(4)10μΜ Pioglitazone induced andincrease cellular activity, there is no the same effect in other groups（1μM、100μM）.Similarly, there were an increase in CSE mRNA expression level,protein expression level and enzyme activity.Conclusion:①High homocysteine to result HUVEC damage.②Acertain concentration sodium hydrosulfide or PPARγagonist Pioglitazone canagainst high homocysteine which lead to HUVEC damage.③Sodiumhydrosulfide and Pioglitazone agonistic function together with endogenousH2S generation.④Sodium hydrosulfide and Pioglitazone through increaseintra-cellular CSE Expression level and enzyme activity enhancementendogenous H2S generation.