FAD Ameliorates Pathological Cardiac Hypertrophy And Fibrosis Through Activating SCAD

Objective Flavin adenine dinucleotide(FAD)is a cofactor for short chain acyl-Co A dehydrogenase(SCAD)and catalyzes electron transfer in redox reactions.Previous studies have shown that SCAD has a negative regulatory effect on pathological myocardial hypertrophy and myocardial fibrosis.Therefore,regulating the activity of SCAD may become one of the important links to interfere with pathological myocardial hypertrophy and myocardial fibrosis.Studies have shown that FAD regulates the biological activity of SCAD in vivo and in vitro.However,it is unclear whether FAD can prevent pathological myocardial hypertrophy and myocardial fibrosis by upregulating SCAD's enzyme activity or expression.This project aims to observe the effects of FAD on pathological myocardial hypertrophy and myocardial fibrosis,and to provide new ideas and theoretical basis for the prevention and treatment of pathological myocardial hypertrophy and myocardial fibrosis.Method1.Cells were pretreated with FAD for 30 min,and then treated by phenylephrine(PE)for 24 h.The surface area of myocardial,SCAD expression were detected by western blot.The m RNA expression of SCAD,ANF,BNP were detected by PCR,and the SCAD enzyme activity,ATP,BNP and free fatty acid content were detected by Eisa kit.2.Cardiac fibroblasts were pretreated with FAD for 30 minutes,and then treated by angiotensin?(Ang?)for 36 h.The protein expression of SCAD,collagen?,collagen ?,and ?-SMA were detected by western blot.The m RNA expression of SCAD,collagen?,collagen ?,and ?-SMA were detected by PCR.The SCAD enzyme activity,ATP,free fatty acid content and cell proliferation rate were detected.3.12-week-old male spontaneously hypertensive rats(SHR)were treated with tail vein injection of FAD for 10 weeks.The blood pressure and heart rate of rats,the protein expressions of SCAD,Collagen?,collagen? and ?-SMA,and the m RNA expression levels of SCAD,ANF,BNP,Collagen?,collagen? and ?-SMA were detected.SCAD enzyme activity,ATP,free fatty acid,hydroxyproline,and reactive oxygen content were measured.The immunofluorescence single-label method was used to further verify the protein expression level of myocardial SCAD,and cardiac morphology and myocardial fibrosis were analyzed by echocardiography and histology.4.Compared with APO,the RMSD of the FAD-SCAD molecular system shows that the SCAD dimer and substrate pocket are more stable,and the Rg value indicates that the FAD-SCAD molecular system is more compact.Further analysis shows that FAD maintains the stability of the catalytic pocket and prevents the collapse of the catalytic pocket by bonding with amino acid residues in the binding site.Result1.Compared with the control group,the cardiomyocytes surface area,the expression of hypertrophic marker genes ANF,BNP m RNA,free fatty acid and BNP content in the PE group were significantly increased,while the m RNA,protein expression,enzyme activity of SCAD,and ATP content were significantly reduced.Compared with the PE group,the cardiomyocytes surface area,the expression of hypertrophic marker genes ANF,BNP m RNA,free fatty acid and BNP content in the PE+FAD group were significantly reduced,while the m RNA,protein expression,enzyme activity of SCAD,and ATP content were significantly increased.2.Compared with the control group,the m RNA,protein expression,enzyme activity of SCAD,and ATP content in cardiac fibroblast induced by Ang? were significantly reduced,while the proliferation rate,fatty acid content,and the m RNA and protein expression of collagen?,collagen?,and ?-SMA were significantly reduced.Compared with the Ang?group,the m RNA,protein expression,enzyme activity of SCAD,and ATP content in cardiac fibroblast induced by Ang?were significantly increased,while the proliferation rate,fatty acid content,and the m RNA and protein expression of collagen?,collagen ?,and ?-SMA were significantly reduced.3.Compared with the control Wistar,the systolic blood pressure and heart rate of the SHR were significantly increased.Echocardiography and morphological analysis showed that significant myocardial hypertrophy and fibrosis were appeared in the hearts.Besides,the ANF,BNP,collagen?,collagen?,?-SMA m RNA and protein expression,and the free fatty acid,BNP and hydroxyproline content were significantly increased,while the m RNA,protein expression,enzyme activity of SCAD,and ATP content were significantly decreased.Compared with the control SHR,with the treatment of FAD,systolic blood pressure and heart rate of SHR were significantly reduced.The myocardial hypertrophy and fibrosis were significantly ameliorated.Besides,the ANF,BNP,collagen?,collagen?,?-SMA m RNA and protein expression,and the free fatty acid,BNP and hydroxyproline content were significantly reduced.while the m RNA,protein expression,enzyme activity of SCAD,and ATP content were significantly decreased.4.Compared with APO,the RMSD and Rg values of FAD-SCAD molecular system show that SCAD dimer and substrate pocket are more stable.Further analysis showed that FAD maintains the stability of the pocket by bonding with amino acid residues in the substrate pocket.Conclusion Down-regulation of SCAD activity negatively regulates myocardial hypertrophy and myocardial fibrosis.FAD may promote cell energy metabolism by activating SCAD,thereby inhibiting pathological myocardial hypertrophy and myocardial fibrosis.