High Glucose Induced Mouse Glomerular Podocytes Injury And Effects Of Hydrogen Sulfide On Podocytes

ObjectiveTo observe the expressions of cystathionine-y-lyase(CSE), zona occludens-2(ZO-2), Nephrin and β-catenin in high glucose-induced mouse podocytes injury and the changes of ZO-2, Nephrin and β-catenin after treated by hydrogen sulfide, to investigate the mechanisms of high glucose-induced podocytes injury, effects of hydrogen sulfide on Diabetic Nephropathy and its most possible protective mechanisms underlying.MethodsSubculture of mouse podocytes(MPCs) in vitro were randomly divided into4groups:(1)high glucose(HG group):after treated by non-serum media for24h, each group was respectively cultured in high glucose(30mmol/L)1640media for0,12,24,48h;(2)normal glucose(NG) group;(3) normal glucose+DL-propargylglycine (PPG) group:each group was cultured for24h by serum starvation and then different concentrations of PPG intervened for48h;(4) high glucose+NaHS (HG+NaHS):each group were cultured by serum starvation for24h, the cells were disposed to1640media with30mmol/L glucose and100μmol/L NaHS for48h. Mannitol were used to adjust osmotic pressure to30mmol/L in all the groups to avoid the interference of different osmotic pressure.After treating for a certain time, the cells were collected for further detection. The expression of ZO-2, Nephrin, β-catenin and cystathionine-y-lyase(CSE) were determined by western blot.Results1. Compared with HG Oh group, the expressions of Nephrin, ZO-2and CSE were significantly reduced in the cells treated with high glucose for12,24,48h (P<0.05), while the levels of P-catenin were elevated obviously (P<0.05), four proteins changed in a time-dependent manner.2. Compared with NG group, the levels of ZO-2, Nephrin were decreased significantly in normal glucose groups cultured with different concentrations of PPG (P<0.05). In contrast, the levels of β-catenin elevated significantly (P<0.05). Three proteins presented greater variation with different concentration of PPG.3. Compared with HG group, high glucose-induced podocytes treated by NaHS induced a more significant expressions of ZO-2and Nephrin (P<0.01), whereas there was a severe reduction of β-catenin in HG+NaHS group (P<0.01). Relative to NG group, the expressions of ZO-2and Nephrin in HG+NaHS group were decreased obviously (P<0.01), the level of β-catenin in HG+NaHS group was higher than that in NG group (P<0.01)ConclusionsOur results suggested high glucose induced mouse podocytes injury by reducing ZO-2expression, excessive activation of Wnt/β-catenin pathway and decreasing the level of Nephrin in a time-dependent manner. On the other side, downregulation of CSE contributes to hyperglycemia induced podocyte injury also in a time-dependent way. That suggested the reduction of endogenous H2S triggered by the CSE decline probably is one of mechanisms in high glucose induced podocytes injury. Exogenous H2S protects against hyperglycemia induced podocyte injury possibly through upregualtion of ZO-2and inhibition of Wnt/β-catenin pathway, and then reducing the loss of Nephrin.