The Mechanism of Environmental Endocrine Disruptors (DEHP) Induces Epigenetic Transgenerational Inheritance of Cryptorchidism

Discussion on the role of DEHP in the critical period of gonadal development in pregnant rats (F0), studied the evolution of F1-F4 generation of inter-generational inheritance of cryptorchidism and the alteration of DNA methylation levels in testis. Pregnant SD rats were randomly divided into two groups: normal control group and DEHP experimental group. From pregnancy 7 d to 19 d, experimental group was sustained to gavage DEHP 750 mg/kg bw/day, observed the incidence of cryptorchidism in offspring and examined the pregnancy rate of female rats through mating experiments. Continuous recording the rat's weight and AGD value, after maturation (PND80) recording testis and epididymis' size and weight, detected the sperm number and quality. Subsequently, we examined the evolution morphological changes of testicular tissue for 4 generation rats by HE staining and Western Blot. Completed the MeDIP-sequencing analysis of 6 samples (F1 generation, F4 generation and Control). DEHP successfully induced cryptorchidism occurrence in offspring during pregnancy. The incidence of cryptorchidism in F1 was 30%, in F2 was 12.5%, and there was no cryptorchidism coming up in F3 and F4. Mating experiment shows conception rate 50% in F1, F2 generation was 75%, the F3 and F4 generation were 100%. HE staining showed that the seminiferous epithelium of F1 generation was atrophy and with a few spermatogenic cell, F2 generation had improved, F3 and F4 generation were tend to be normal. The DNA methyltransferase expression was up-regulated with the increase of generations by Real Time-PCR, immunohistochemistry and Western Blot. MeDIP-seq Data Analysis Results show many differentially methylated DNA sequences between F1 and F4. DEHP damage male reproductive function in rats, affect expression of DNA methyltransferase enzyme, which in turn leads to genomic imprinting methylation pattern changes and passed on to the next generation, so that the offspring of male reproductive system critical role in the development of imprinted genes imbalances, and eventually lead to producing offspring cryptorchidism. This may be an important mechanism of reproductive system damage.