GW24-e2123 Effect of Sini Decoction on the expression of Sirt-1 and eNOS system in EAhy926 cells injured by homocysteine

Objectives To detect the effect of Sini Decoction on the expression of Sirt-1 and eNOS in EAhy926 cell injured by homocysteine. Methods Model of EAhy926 cell injured by homocysteine was made,the protection on the EAhy926 cell of Sini Decoction with different dosages were observed. NO concentration of cell culture fluid was detected by nitrate reductase method, effect of Sini Decoction on the expression of protein of Sirt-1 and eNOS in EAhy926 cell were observed by Western-blot, and effect of Sini Decoction on the expression of mRNA of Sirt-1 and eNOS in EAhy926 cell were observed by fluorescent quantitation PCR. Results After Model of EAhy926 cell injured by homocysteine was made, we found that cultured with 0.5,1.0,2.0,4.0,8.0 μmol/L homocysteine, cells grew less than cultured with normal culture medium, and with the increase of homocysteine concentration, the number of attached cell grew downwards obviously, as culturing with homocysteine 4.0 μmol/L for 24h did lower damage to cells and could induce effective cell injuring, it was made to be the model of injury. To detect the effect of Sini Decoction on EAhy926 cell injured by homocysteine, well growing EAhy926 cells were cultured in culture plate. 24 h later, cells were cultured with DMEM medium containing 2% fetal calf serum for 8 hours to make cells hungry, then cultured with medium containing Sini Decoction 0, 0.25, 0.5, 1.0g/ml respectively for 30 minutes,then cultured with medium containing homocysteine 4.0 μmol/L for 24 h. It was found that, compared with control group, attached cells in Sini Decoction groups grew better, and attached cells in Sini Decoction 1.0g/ml plus homocysteine 4.0 μmol/L group grew best. 4.0 μmol/L. Detected by nitrate reductase method, it was found that compared with control group, there was no obvious change of NO concentration of cell culture fluid in Sini Decoction 1.0 g/ml group, but in homocysteine 4.0 μmol/L medol group, NO concentration of cell culture fluid decreased oberviously, and in Sini Decoction groups, NO concentration of cell culture fluid increased, and in Sini Decoction 1.0g/ml plus homocysteine 4.0 μmol/L group it was the most obvious (p < 0.05). Detected by Western-blot, it was found that,campared with control group, there was no obervious change of protein of Sirt-1 and eNOS in Sini Decoction 1.0 g/ml group, but in homocysteine 4.0μmol/L model group, expression of Sirt-1 and eNOS protein weakened obviously, and in Sini Decoction groups, expression of Sirt-1 and eNOS protein enhanced, and in Sini Decoction 1.0g/ml plus homocysteine 4.0 μmol/L group it was the most obvious (p < 0.05). Detected by fluorescent quantitation, it was found that, compared with control group, there was no obvious change of mRNA of Sirt-1 and eNOS in Sini Decoction 1.0g/ml group, but in homocysteine 4.0 μmol/L model group, expression of Sirt-1 and eNOS mRNA weakened obviously, and in Sini Decoction groups, expression of Sirt-1 and eNOS mRNA enhanced, and in Sini Decoction 1.0 g/ml Plus homocysteine 4.0 μmol/L group, it was the most obvious (p < 0.05). Conclusions Homocysteine may injure EAhy926 cell by suppressing the expression of Sirt-1 then suppressing the expression of eNOS system, while Sini Decoction may protect EAhy926 cell by enhancing the expression of caveolin-1 then enhancing the expression of eNOS system.