| BackgroudsEssential hypertension is a complex genetic disease caused by the combination of lifestyle,genetic factors,environmental factors and many other factors.The situation of prevention and control of hypertension in China is still grim,so clarifying its etiology and pathogenesis is of great significance to the prevention and treatment of hypertension.Epidemiological studies have found that the antihypertensive treatment rate of parental hypertension is inversely proportional to the incidence of hypertension in the offspring.Although there is a theoretical possibility,there is currently little evidence about the intergenerational inheritance of traits in mammals.Most studies of environmental impact on intergenerational genetics are related to the impact of maternal pregnancy.However,these studies are difficult to distinguish whether these adverse effects exert these effects by affecting the development of the fetus in the uterus or by affecting the eggs.Therefore,in order to circumvent this problem,this study uses male rats for research,because the father’s influence on the development of the next generation of embryos is mainly done through the medium of sperm.Clinical studies have shown that urinary sodium excretion dysfunction is common in hypertensive patients.Therefore,the role of the kidney in the study of the regulatory mechanism of hypertension is increasingly concerned and paid attention by researchers.The excessive activation of RAAS system plays an important role in the regulation of urinary sodium excretion in patients with hypertension,mainly through AT1 R to mediate its downstream signaling.ATRAP can specifically bind to the carboxyl terminus of AT1 R,antagonize the transmission of AT1 R signaling pathway by mediating AT1 R internalization,desensitization and phosphorylation,and inhibit its downstream signaling pathway.Epigenetic information at specific gene loci can have lasting intergenerational effects.Epigenetics participates in the regulation of gene expression programs by responding to specific environmental conditions,which helps to explain the cause of hypertension’s "hereditary loss".Sperm is the only genetic link between the father and the offspring.Studying the epigenetic changes in the father’s sperm gene will be a breakthrough to reveal whether the father receives antihypertensive therapy and affects the blood pressure level of the offspring.Therefore,in order to exclude the influence of congenital genes such as spontaneously hypertensive rats,we adopted exogenous AngⅡ intervention method to construct an acquired parental hypertension model,and were given losartan,propranolol and nifedipine three Antihypertensive drugs,to observe whether exogenous factors induce paternally acquired hypertension and whether antihypertensive therapy can affect the blood pressure of the offspring in adulthood,and to explore whether the changes in blood pressure of the offspring are related to abnormal renal urinary sodium excretion function,and Intergenerational genetic mechanism of paternal acquired traits.ObjectObserve the influence of father-acquired hypertension on blood pressure and sodium excretion of adult offspring;whether hypertension-receiving fathers can improve the health of their offspring by taking antihypertensive therapy,and explore its possible epigenetic mechanism.Methods1.Eighteen 8-week-old healthy male C57 BL / 6J adult mice,weighing 18-22 g,were randomly divided into 6 groups: control group(saline),AngⅡ(400ng / kg / min)group,AngⅡ(400ng / kg / min)+ losartan(30mg / kg / d)group,AngⅡ(400ng / kg /min)+ propranolol group(20mg / kg / d),AngⅡ(400ng / kg / min)+ nifediene In the dipine group(20mg / kg / d),AngⅡ(400ng / kg / min)+ Tempol(1.0mmol / L)group,Alzert micro-osmotic pump was implanted into the back to inject AngⅡ or saline at400 ng / kg / min Pump note 28 d.2.After 28 days of intervention,male rats of each group were mated with10-week-old female mice to obtain daughter rats.After rearing to 7 weeks of age under standard conditions,the blood pressure and 24 h urine volume urine sodium of each group of babies were monitored.Through perfusion of AT1 R inhibitor candesartan,we further observed the role of renal AT1R-mediated urinary sodium excretion in the regulation of blood pressure in progeny.3.Real-time fluorescence quantitative PCR and immunoblotting analysis were used to detect the expression of methyltransferase DNMT1,AT1 receptor and ATRAP in the renal cortex of the father’s testis.4.Bisulfite modified sequencing method(BSP)to detect the AT1 receptor and ATRAP methylation level of paternal sperm.Result1.After 4 weeks of intervention,the blood pressure and urinary sodium excretion function of male rats in the AngⅡ group were significantly damaged(p <0.05);after being given losartan,propranolol and nifedipine,respectively,the basic The blood pressure and urinary sodium excretion were significantly improved compared with the AngⅡ group(p <0.05,),and there was no significant difference compared with the control group(p> 0.05),indicating that the intervention models of each group were experimentally feasible.2.The blood pressure of the offspring in the AngⅡ group at 10 weeks of age is significantly higher than that of the control group(p <0.01);the blood pressure of the offspring in each antihypertensive treatment group is significantly lower than that of the AngⅡ group(p <0.01);through renal perfusion Candesartan found that the AT1 receptor function was significantly inhibited in the offspring of the AngⅡ group(p<0.05),and the urine sodium excretion function of the offspring in the antihypertensive treatment group was improved compared with the AngⅡ group of offspring(p <0.05).3.The expression of AT1 receptor in renal cortex in AngⅡ group increased compared with control group(p <0.05,),ATRAP expression decreased(p <0.05);antihypertensive treatment could reverse the expression of AT1 receptor and ATRAP(P <0.05).4.The sperm ATRAP methylation level of male rats in the AngⅡ group was significantly higher than that in the control group(p <0.05);while the ATRAP methylation level of father sperm in the antihypertensive treatment group was lower than that in the AngⅡ group(p <0.05),compared with the control group There was no significant difference(p> 0.05);there was no significant difference in male AT1 R methylation in each group(p> 0.05).5.The oxidative stress in the testicular microenvironment caused by hypertension led to a significant increase in the expression of DNMT1 in the testis tissue of the fathers of the AngⅡ group compared with the control group and the AngⅡ + Tempol group(p <0.05,n = 3).ConclusionsAngⅡ-induced high oxidative stress in the acquired hypertension of the parents caused up-regulation of DNMT1 expression,resulting in hypermethylation of sperm ATRAP,and transmitted this apparent modification information to the offspring through sperm,up-regulated the expression of ATRAP in the offspring kidneys to suppress AT1 R Mediated urinary sodium excretion function;and reasonable antihypertensive treatment can reverse the reprogramming of the father’s sperm ATRAP gene and restore the health of the offspring. |
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