Mechanism Of Ppargc1α Demethylation In Vascular Smooth Muscle Phenotype Regulation Of Hypertensive Male Offspring Induced By Pregnancy Exercise

Aims:Hypertension is influenced by genes and environment.Early life stage is a critical period for embryo growth and development.Changes in intrauterine environment affect the development of offspring and have a lasting impact on their health.Whether pregnancy exercise can improve the vascular function of hypertensive offspring is not clear.The aim of this study was to investigate the effect of pregnancy exercise on the phenotype of mesenteric arterial vascular smooth muscle cell（VSMC）in hypertensive male offspring,and to explore the effect of ten-eleven translocation 2（TET2）-mediated Ppargc1αgene demethylation in regulating the phenotypic switch of VSMC in hypertensive male offspring.Methods:Spontaneously hypertensive rats（SHR）and Wistar-Kyoto（WKY）rats were selected,male and female rats of each strain were used for mating.Pregnant rats were separated into a sedentary group（p-WKY-SED,p-SHR-SED）and an exercise group（p-WKY-EX,p-SHR-EX）at random.Exercise groups were performed a swimming training（60 min/day,6 days/week,the water depth was 40 cm）until the19th day of pregnancy.The food intake and body weight of pregnant rats were monitored every day.The fetuses were collected on the 21st day of pregnancy to detect the body weight,body length and placental weight.HE staining was used to observe the mesenteric artery morphology from male fetuses and 3-month-old（3M）offspring.Caudal artery tail-cuff,femoral venous and arterial cannular surgery,artery contraction myograph,patch-clamp were used to measure blood pressure and heart rate,cardiovascular response,mesenteric vascular tone,whole-cell currents of both Ca_V1.2 channels and BK_Cachannels of 3M offspring,respectively.qPCR,western blot,bisulfite sequencing PCR were used to detect the level of PGC1αmRNA,protein,methylation rate of the mesenteric arteries,respectively.HPLC was used to assess levels of plasmaα-KG and succinate from both fetuses and 3M offspring.In vitro experiment,mesenteric arteries were intervened with fumarate（a TET inhibitor）,then cultured with high glucose.Intervened arteries were used to detect TET2 protein and 5hm C level,methylation rate of Ppargc1αgene,and the markers expression for VSMC phenotype.Results:1)There were no significant differences in food intake,weight gain,abortion rate and litter rate among the four groups（P>0.05）.2)Pregnancy exercise significantly increased the placental efficiency of the male offspring in p-SHR-EX group compared with that in p-SHR-SED offspring（P<0.05）.3)Systolic blood pressure（SBP）,diastolic blood pressure（DBP）and mean arterial pressure（MAP）of3M male offspring in p-SHR-SED were significantly increased（P<0.05）,while DBP and MAP were markedly reduced in p-SHR-EX dams（P<0.05）.4)The ratio of medial layer thickness to lumen diameter was increased,protein ofα-SMA and SM-MHC were decreased,while OPN was increased in p-SHR-SED from 3M male offspring（all P<0.05）,which were effectively inhibited in p-SHR-EX strain（all P<0.05）.5)Pregnancy exercise significantly attenuated NE and AngⅡcontribution to blood pressure regulation and mesenteric vascular tone regulation from 3M male offspring in p-SHR-EX（both P<0.05）.6)The whole-cell currents of Cav1.2 and BK_Cachannels in mesenteric artery smooth muscle cell were increased markedly in p-SHR-SED from 3M male offspring（P<0.05）,while these channels currents were significantly suppressed in p-SHR-EX dams（P<0.05）.7)The expression of PGC1αmRNA and protein in p-SHR-EX from male fetuses and 3M offspring were up-regulated,which were down-regulated in the same age offspring in p-SHR-SED（P<0.05）.8)The methylation rate of Ppargc1αgene in p-SHR-EX from male fetuses and 3M offspring decreased significantly compared with those in p-SHR-SED（P<0.05）.9)Pregnancy exercise inhibited hypertension-associated decrease levels of TET2 protein and 5hm C of mesenteric artery in p-SHR-EX from male fetuses and3M offspring（P<0.05）.10)Blood glucose was increased,AMPKαactivity and plasmaα-KG/succinate ratio were decreased in p-SHR-SED from fetuses（all P<0.05）,and pregnancy exercise significantly inhibited these effects in p-SHR-EX offspring（all P<0.05）.11)During pregnancy,exercise decreased the maternal blood glucose（P=0.053）and makedly increased AMPKαactivity and the ratio ofα-KG/succinate（P<0.05）in p-SHR-EX compared with those in p-SHR-SED.12)Cultured arteries experiment showed that fumarate-mediated TET2 inhibition enhanced high glucose-induced decrease of the demethylation rate of Ppargc1αgene,and reduced the expression of PGC1αmRNA and protein,which led to a switch from a contractile phenotype to a synthetic one in VSMC（P<0.05）.Conclusions:1)Pregnancy exercise can significantly improve the placental efficiency of male fetuses and reduce the blood pressure of 3M male offspring in hypertension.2)Pregnancy exercise can effectively alleviate the pathological changes of the mesenteric artery structure from the 3M hypertensive male offspring,which was attributed to the inhibition of a switch from contractile phenotype to the synthetic one.3)Pregnancy exercise reduces blood glucose in hypertensive male fetuses by decreasing maternal blood glucose.The reduced blood glucose in hypertensive fetuses increase TET2-mediated Ppargc1αgene demethylation,upregulats PGC1αexpression,and inhibits phenotypic switching in VSMC.