IGF-1 can protect cardiomyocytes from apoptosis through BTEB and CYPs pathway
Author(s): ZHANG Jian-kai, DING Bi-lan, XU Xiao-lin, LI Tao, WU Zhu-guo
Pages: 64-71
Year: 2015 Issue: 1
Journal: Chinese Journal of Clinical Anatomy
Keyword: Cardiomyocytes; Apoptosis; Basic transcription element binding protein(BTEB); Cytochromes P450(CYPs); Insulin-like growth factor-1(IGF-1);
Abstract: Objective To investigate gene regulation mechanism of the anti-apoptotic effect of the insulin-like growth factor 1 on rat cardiomyocytes. Methods Primary neonatal rat cardiomyocytes (NRCMs) were cultured in vitro,the transcription and translation levels of BTEB and its downstream genes Cytochrome P450 family members (CYP1A1, 2E1, 3A11 and 11A1) were measured by RT-PCR and Western-Blot. NRCMs were treated with 100umol/L Hydrogen peroxide (H2O2) to induce apoptosis. CYP1A1, 2E1, 3A11 and 11A1 specific siRNA were transfected into the cells by Lipofectamine 2000. Myocardial cells apoptosis was detected by Annexin V-FITC/PI dual staining,Caspase-3 activity assay,Hoechst33258 staining and Tunel method. The mitochondrial function was measured by JC-1 mitochondrial membrane potential assay. The mitochondrial morphology was observed by transmission electron microscopy. Results The mRNA and protein expression levels of BTEB gene in NRCMs were down-regulated significantly after IGF-1 had stimulated for 60 minutes with concomitant attenuation of CYP1A1, 2E1, 3A11 and 11A1 expression levels 48 hours later (all P<0.01). Artificial inhibition of BTEB gene expression with BTEB specific siRNA could down-regulate CYP1A1, 3A11 mRNA and protein expression obviously (P<0.01), but CYP2E1, 11A1 mRNA 2E1, 3A11 and 11A1 genes silencing on NRCMs was similar with that of IGF-1 treatment. And the fall rates of mitochondrial membrane potential in IGF-1 group and CYP1A1, 2E1, 3A11 and 11A1 specific siRNA groups declined significantly (all P<0.05 vs control group), mitochondrial morphology obviously improved. Conclusion IGF-I protects rat cardiomyocytes from apoptosis through down-regulation of Cytochrome P450 family members (CYP1A1, 2E1, 3A11 and 11A1) expression pathways and improving mitochondrial function, of which CYP1A1 and 3A11 were regulated by BTEB.
Pages: 64-71
Year: 2015 Issue: 1
Journal: Chinese Journal of Clinical Anatomy
Keyword: Cardiomyocytes; Apoptosis; Basic transcription element binding protein(BTEB); Cytochromes P450(CYPs); Insulin-like growth factor-1(IGF-1);
Abstract: Objective To investigate gene regulation mechanism of the anti-apoptotic effect of the insulin-like growth factor 1 on rat cardiomyocytes. Methods Primary neonatal rat cardiomyocytes (NRCMs) were cultured in vitro,the transcription and translation levels of BTEB and its downstream genes Cytochrome P450 family members (CYP1A1, 2E1, 3A11 and 11A1) were measured by RT-PCR and Western-Blot. NRCMs were treated with 100umol/L Hydrogen peroxide (H2O2) to induce apoptosis. CYP1A1, 2E1, 3A11 and 11A1 specific siRNA were transfected into the cells by Lipofectamine 2000. Myocardial cells apoptosis was detected by Annexin V-FITC/PI dual staining,Caspase-3 activity assay,Hoechst33258 staining and Tunel method. The mitochondrial function was measured by JC-1 mitochondrial membrane potential assay. The mitochondrial morphology was observed by transmission electron microscopy. Results The mRNA and protein expression levels of BTEB gene in NRCMs were down-regulated significantly after IGF-1 had stimulated for 60 minutes with concomitant attenuation of CYP1A1, 2E1, 3A11 and 11A1 expression levels 48 hours later (all P<0.01). Artificial inhibition of BTEB gene expression with BTEB specific siRNA could down-regulate CYP1A1, 3A11 mRNA and protein expression obviously (P<0.01), but CYP2E1, 11A1 mRNA 2E1, 3A11 and 11A1 genes silencing on NRCMs was similar with that of IGF-1 treatment. And the fall rates of mitochondrial membrane potential in IGF-1 group and CYP1A1, 2E1, 3A11 and 11A1 specific siRNA groups declined significantly (all P<0.05 vs control group), mitochondrial morphology obviously improved. Conclusion IGF-I protects rat cardiomyocytes from apoptosis through down-regulation of Cytochrome P450 family members (CYP1A1, 2E1, 3A11 and 11A1) expression pathways and improving mitochondrial function, of which CYP1A1 and 3A11 were regulated by BTEB.
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