Study On The Apoptosis Of Germ Cells In Testis Of Kidney-Yang Deficiency Syndrome Mice Model

Objective:This experimental study focused on the mechanism of germ cell apoptosisin the testis of Kidney-Yang deficiency syndrome mice model.Meanwhile,the effects ofJinguishenqi bolus on decreasing the germ cell apoptotic index were explored.Methods:In this study,a mice model of Kidney-yang deficiency syndrome was usedas experimental animal which had been established by us before.The method used todevelop Kidney-yang deficiency syndrome mice model based on principle of Chinesemedicine classical theory.The mice model was caused by overfatigue and sexualoverstrain.The morphological changes of mice testis tissue were observed after HEstaining.Ultramicrostructure were observed with transmission electron microscopy aswell.The concentrations of follicle-stimulating hormone and testosterone in the testis ofmice were measured with radioimmunoassay method.Then the mRNA levels of certainapoptosis related genes including Bcl-2,Bax,Bcl-XL,Fas,FasL and P53 were assessed byRT-PCR.Furthermore,the expression of Fas,FasL and Caspase3 protein in mice testiswere determined by immunohistochemical method.In addition,the germ cell apoptosisratios were detected by TUNEL.Results:The testicular germ cell apoptotic index of Kidney-Yang deficiencysyndrome mice model caused by overfatigue and sexual overstrain increased significantly.And typical ultrastructural changes of apoptosis cell such as cell shrinkage,roughmembrane,nuclei shrinkage and margination of chromatin in model mice testis could beseen under ultramicroscope.While intratesticular testosterone concentration in mice modelwas much higher than normal mice,the content of follicle-stimulating hormone in mice model showed no change compared with normal mice.RT-PCR results showed thatmRNA levels of Fas and FasL in model mice testis were much higher than that in normalmodel.And abnormalities of location and expression of Fas and FasL protein in modelmice testis were detected by immunohistochemical method.And there was a higherexpression of Fas activated apoptopic signal pathway downsteam enzyme Caspase3 inmodel mice testis.When the model mice were treated with Jinguishenqi bolus ormethyltestosterone,the content of intratesticular testosterone in mice testis increasedobviously.And Fas,FasL and Caspase3 expression were downregulated by Jinguishenqibolus or methyltestosterone too,and so the apoptopic index.And there was also mRNAexpression change of one of Bcl-2 family protein named Bcl-XL in model mice testis.Itseemed that Jinguishenqi bolus or methyltestosterone had no any effect on this diversity.Conclusion:Intratesticul testosterone plays an important roles on regulatingapoptosis of male germ cell.Kidney-yang deficiency syndrome model mice caused byoverfatigue and sexual overstrain have a low level of intratesticul testosteroneconcentration.The low content of intratesticul testosterone may upregulate theexpression Fas and FasL and then initiate a Fas dependent apoptotic pathway.Caspase3which has also been regulated works as a major executer in the Fas dependent apoptoticpathway.Jinguishenqi bolus can induce the apoptopic index of germ cells of model malemice.The mechanism of Jinguishenqi bolus on germ cell apoptosis may mainly dependon increasing intratesticul testosterone.In this way,the Fas and FasL expression havebeen depressed and so does Caspase3.Furhermore,Bcl-XL is also involved in theapoptosis process of the Kidney-Yand deficiency model mice.