[Sirtuin 3 inhibits cardiomyocyte apoptosis by reducing cytochrome C release in myocardiac H9c2 cells with calcium overload].
OBJECTIVE
To investigate the effects of Sirtuin 3 on cytochrome C (CytC) release and caspase-3 activation in myocardiac H9c2 cells with calcium overload.


METHODS
H9c2 cells were randomly divided into four groups: control group, Sirtuin 3 group, ionomycin group, ionomycin & Sirtuin 3 group. Myocyte calcium overload models were established by ionomycin revulsion. The levels of CytC and caspase-3 in mitochondria and cytoplasm were examined by Western blotting. Intracellular caspase-3 activity was examined by spectrophotometry. H9c2 cell apoptosis rate in each group was detected by annexin V-FITC/PI double staining combined with flow cytometry.


RESULTS
Calcium overload models of H9c2 cells were successfully established. Western blotting showed that CytC level in mitochondria of the ionomycin group was significantly lower than that in the control group, and that it was significantly higher in the ionomycin & Sirtuin 3 group than that in the ionomycin group. Changes of CytC level in cytoplasm was just opposite to that in mitochondria. Caspase-3 level in the ionomycin group was significantly higher than that in the control group, and it was significantly lower in the ionomycin & Sirtuin 3 group than that in the ionomycin group. Spectrophotometry showed that caspase-3 activity in the ionomycin group was significantly higher than that in the control group, and that it was obviously lower in the ionomycin & Sirtuin 3 group than that in the ionomycin group. Flow cytometry combined with double staining revealed that the apoptosis rate in the ionomycin group was significantly higher than that in the control group, and that it was significantly lower in the ionomycin & Sirtuin 3 group than that in the ionomycin group.


CONCLUSIONS
Sirtuin 3 could inhibit cell apoptosis by reducing CytC release and inhibiting caspase-3 activation in cardiomyocyte models with calcium overload.
Department of Cardiothoracic Surgery, First Affiliated Hospital, Chongqing Medical University, Chongqing 400016, China.
Lu
L
Ye
Department of Cardiothoracic Surgery, First Affiliated Hospital, Chongqing Medical University, Chongqing 400016, China.
Shuangqiang
S
Yang
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism*; D061207:Calcium Ionophores / Q000494:pharmacology; D053148:Caspase 3 / Q000378:metabolism; D002460:Cell Line; D045304:Cytochromes c / Q000378:metabolism*; D005434:Flow Cytometry; D015759:Ionomycin / Q000494:pharmacology; D033681:Mitochondrial Membrane Transport Proteins / Q000378:metabolism; D000083162:Mitochondrial Permeability Transition Pore; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D056566:Sirtuin 3 / Q000494:pharmacology*
D061207:Calcium Ionophores; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D015759:Ionomycin; D045304:Cytochromes c; D053148:Caspase 3; D056566:Sirtuin 3; D002118:Calcium