Sphingosine 1 Phosphate Receptor-1 (S1PR1) Signaling Protects Cardiac Function By Inhibiting Cardiomyocyte Autophagy

AIMS:Sphingosine 1 phosphate(SIP)has an inhibitory effect on the development of cardiac hypertrophy and could inhibit autophagy of various cells.We aimed to investigate the role of sphingosine-1-phosphate(SIP)and its receptors in cardiomyocyte autophagy,cardiomyocyte hypertrophy and cardiac function.Methods:Cardiomyocytes were isolated from neonatal Vista rats.Autophagy and hypertrophy of cardiomyocytes were induced via starvation culture and phenylephrine(PE),respectively,and SIP was used to treat the cardiomyocytes.The effect of SIP on cardiomyocyte autophagy was evaluated using the number of autophagosomes,the expression of autophagy-related proteins and autophagic marker genes in cardiomyocytes.The effect of SIP on cardiomyocyte hypertrophy was evaluated by examining the surface area of cardiomyocytes and the expression of hypertrophic genes.Subsequently,different siRNAs were used to knockdown the expression of the three types of SIP receptors on cardiomyocytes and to analyze the type of receptor that mediates S1P signaling in cardiomyocytes.Finally,sphingosine 1 phosphate receptor-1(S1PR1)was knocked down in the mouse cardiomyocytes using the Cas9 technique.The effect of S1PR1 on cardiac autophagy and cardiac hypertrophy was examined by assessing cardiomyocyte autophagy,cardiomyocyte hypertrophy and cardiac function.Results:Starvation-induced cardiomyocyte autophagy and PE-induced cardiomyocyte hypertrophy were significantly attenuated by S1P.The results showed that the formation of autophagosomes was decreased,the autophagy-associated protein LC3Ⅱ、Ⅰ and the expression of autophagic marker genes Atg5,Atgl2,Beclinl and LC3B decreased after S1P treatment.The surface area of the cardiomyocytes was decreased,and the expression of hypertrophic genes,such as atrial natriuretic factor(ANF),brain natriuretic peptide(BNP),myosin heavy chain(β-MHC)and skeletal muscle and cardiac actin(SKA)decreased after SIP treatment.The autophagy and hypertrophy of cardiomyocytes in the S1PR1 knocked-down group was significantly increased compared to those in the control group,the S1PR2 and the S1PR3 knocked-down groups.In vivo,the knockout of S1P1 in cardiomyocytes exacerbated stress-induced cardiac autophagy,cardiac hypertrophy and the impairment of cardiac function.Conclusion:SIP could inhibit cardiomyocyte autophagy,thereby inhibiting cardiomyocyte hypertrophy and protecting cardiac function by activating S1PR1 in pressure-overloaded cardiomyocytes in mice.