The Effect Of Dibutyl Phthalate Exposure During Pregnancy On The Testicular Germ Cell Apoptosis Of Offspring Rats

Prat ? Effects of in utero exposure of di-n-butyl phthalate on testicular cell apoptosis in late embryonic and pubertal male rat offspringObjective:To investigate the effects of in utero exposure to di-n-butyl phthalate(DBP)on testicular cell apoptosis in late embryonic and pubertal male rat offspring.Methods:Twenty pregnant SD rats were divided into two groups.During gestation day(GD)12 to 19,control group was given lml/d of olive oil,and experimental group was given DBP 500mg/kg/d by gavage.On GD 19.5 and PND 45,the testes were removed.Transmission electron microscope and HE staining were done for morphological analysis.Apoptosis was detected by TUNEL.The expression of Bcl-2,Bcl-XL,Bax and p53 was presented by immunohistochemistry and Western blot.Datas of the two groups were compared using t-test by SPSS 20.0.Results:Multinucleated Gonocyte occurred in GD 19.5 rat testis after DBP exposure and increased apoptosis were observed in PND 45 rat testes.Spermatogenetic cells were significantly decreased in DBP exposed pubertal rat testis.The apoptosis index of testes in DBP treated group was significantly lower on GD 19.5 but higher on PND 45 than that of the controls(P<0.01).Immunohistochemistry and Western blot revealed significant overexpression of Bcl-2 in GD 19.5 rat testis and Bax and p53 in PND 45 rat testis after DBP exposure(P<0.05).Conclusions:In utero exposure of DBP resulted in opposite effects on testicular cell apoptosis in late embryonic and pubertal rat offspring.Overexpression of Bcl-2,Bax and p53 might be related to the abnormal apoptosis and finally produce male infertility.Part ? Regulatory role of miRNA in testicular germ cell apoptosis in rat offspring after in utero exposure of di-n-butyl phthalateObjective:To investigate the differencial expression of miRNA in testicular tissue of rat offspring after in utero exposure of di-n-butyl phthalate(DBP)and its regulatory role to germ cell apoptosis.Methods:Twenty pregnant SD rats were divided into two groups.During gestation day(GD)12 to 19,control group was given 1ml/d of olive oil,and experimental group was given DBP 500mg/kg/d by gavage.On GD 19.5 and PND 45,the testes were removed.RNA sequencing was performed to screen the differentially expressed miRNAs and bioinformatic analysis was used to predict the target genes.Quantitative PCR was performed to validate the apoptosis related miRNA expression.Results:In the testes of GD19.5 and PND45 rats,42 and 237 differentially expressed miRNAs were obtained respectively.The KEGG pathway analysis revealed 15 apoptosis-related miRNAs,with 13 up-regulated miRNAs and 2 down-regulated miRNAs.Quantitative PCR revealed that the expression of miR-140-3p,miR-204-3p and let-7c-5p in the testis of PND45 experimental group was significantly higher than that in the control group(P<0.05).Conclusion:The expression of miRNA in the testis of rat offspring after maternal exposure toDBP was significantly changed.MiR-140-3p,miR-204-3p and let-7c-5p were highly expressed in the testis of DBP-exposed offspring in sexual development stage and finally enhance testicular cell apoptosis through regulating multiple genes auch as Bcl-2 family and p53.