Deshpande Sharvari S, Nemani Harishankar, Arumugam Gandhimathi, Ravichandran Avinash, Balasinor Nafisa H
Department of Neuroendocrinology, ICMR-National Institute for Research in Reproductive Health, Jehangir Merwanji Street, Parel, Mumbai, 400012, India.
National Institute of Nutrition Animal Facility, ICMR-National Institute of Nutrition, Jamai-Osmania PO, Hyderabad, 500 007, India.
Clin Epigenetics. 2020 Nov 19;12(1):179. doi: 10.1186/s13148-020-00974-7.
Paternal obesity has been associated with reduced live birth rates. It could lead to inheritance of metabolic disturbances to the offspring through epigenetic mechanisms. However, obesity is a multifactorial disorder with genetic or environmental causes. Earlier we had demonstrated differential effects of high-fat diet-induced obesity (DIO) and genetically inherited obesity (GIO) on metabolic, hormonal profile, male fertility, and spermatogenesis using two rat models. The present study aimed to understand the effect of DIO and GIO on DNA methylation in male germline, and its subsequent effects on the resorbed (post-implantation embryo loss) and normal embryos. First, we assessed the DNA methylation enzymatic machinery in the testis by Real-Time PCR, followed global DNA methylation levels in spermatozoa and testicular cells by ELISA and flow cytometry, respectively. Further, we performed Methylation Sequencing in spermatozoa for both the groups. Sequencing data in spermatozoa from both the groups were validated using Pyrosequencing. Expression of the differentially methylated genes was assessed in the resorbed and normal embryos sired by the DIO group using Real-Time PCR for functional validation.
We noted a significant decrease in Dnmt transcript and global DNA methylation levels in the DIO group and an increase in the GIO group. Sequencing analysis showed 16,966 and 9113 differentially methylated regions in the spermatozoa of the DIO and GIO groups, respectively. Upon pathway analysis, we observed genes enriched in pathways involved in embryo growth and development namely Wnt, Hedgehog, TGF-beta, and Notch in spermatozoa for both the groups, the methylation status of which partially correlated with the gene expression pattern in resorbed and normal embryos sired by the DIO group.
Our study reports the mechanism by which diet-induced and genetically inherited obesity causes differential effects on the DNA methylation in the male germline that could be due to a difference in the white adipose tissue accumulation. These differences could either lead to embryo loss or transmit obesity-related traits to the offspring in adult life.
父亲肥胖与活产率降低有关。它可能通过表观遗传机制导致后代代谢紊乱的遗传。然而，肥胖是一种具有遗传或环境原因的多因素疾病。此前我们使用两种大鼠模型证明了高脂饮食诱导的肥胖（DIO）和遗传遗传性肥胖（GIO）对代谢、激素谱、雄性生育力和精子发生的不同影响。本研究旨在了解DIO和GIO对雄性生殖系DNA甲基化的影响，及其对吸收（植入后胚胎丢失）和正常胚胎的后续影响。首先，我们通过实时PCR评估睾丸中的DNA甲基化酶机制，然后分别通过ELISA和流式细胞术评估精子和睾丸细胞中的总体DNA甲基化水平。此外，我们对两组精子进行了甲基化测序。使用焦磷酸测序验证两组精子的测序数据。使用实时PCR评估DIO组所产吸收胚胎和正常胚胎中差异甲基化基因的表达，以进行功能验证。
我们注意到DIO组中Dnmt转录本和总体DNA甲基化水平显著降低，而GIO组中则升高。测序分析显示，DIO组和GIO组精子中分别有16966个和9113个差异甲基化区域。通过通路分析，我们观察到两组精子中参与胚胎生长和发育的通路（即Wnt、Hedgehog、TGF-β和Notch）中的基因富集，其甲基化状态与DIO组所产吸收胚胎和正常胚胎中的基因表达模式部分相关。
我们的研究报告了饮食诱导和遗传遗传性肥胖对雄性生殖系DNA甲基化产生不同影响的机制，这可能是由于白色脂肪组织积累的差异所致。这些差异可能导致胚胎丢失，或在成年期将肥胖相关特征传递给后代。
