Endocrine Disruptor Nonylphenol Impairs Lipid Metabolism And Meiotic Entry In Prepubertal Mouse Testis Explants

Abstract

Nonylphenol (NP) is an emerging endocrine-disrupting pollutant that can be present as a residual additive in plastics and can also adsorb onto, and later desorb from, microplastics, thereby enhancing its persistence and bioavailability in aquatic environments. According to the PlastChem Project, which has identified over 16,000 chemicals used in plastics, NP is listed among 3,651 hazardous substances on the Red List that remain internationally unregulated and should be subject to restriction. This toxicant has recently been associated with multiple adverse effects on male fertility in both laboratory animal models and wild species. Previously, using a gas–liquid interphase testicular tissue culture model, we demonstrated that NP disrupts steroidogenesis and lipid homeostasis while simultaneously impairing in vitro spermatogenesis. In this study, we focused on alterations in testicular lipid and fatty acid composition and content, as well as on changes in the expression of genes encoding proteins involved in lipid metabolism. Our results show that NP impairs the normal progression of spermatogenesis in vitro by reducing the number of seminiferous tubules (ST) that reach the meiotic stage. Interestingly, after 22 days in culture, NP-exposed testicular tissue exhibited a slight increase in choline glycerophospholipids accompanied by a decrease in sphingomyelin and ethanolamine glycerophospholipids. In addition, there was a significant accumulation of neutral lipids, including triacylglycerols (TAG) and alkyl-diacylglycerols (ADG). This finding was consistent with results obtained using the fluorescent marker Nile Red, which revealed that NP increases the abundance of neutral lipid droplets in the ST and interstitium. While the fatty acid composition of glycerophospholipids remained unchanged, TAG, ADG, and cholesterol esters (CE) contained higher levels of saturated fatty acids (SFA) and lower levels of polyunsaturated fatty acids (PUFA). These alterations suggest that, in the presence of NP, testicular neutral lipids are unable to incorporate PUFA, which are essential for normal spermatogenic progression. Concomitantly, the mRNA levels of genes involved in fatty acid elongation and hydroxylation, such as very-long-chain elongase 2 (Elovl2) and fatty acid 2- hydroxylase (Fa2h) were down-regulated. In contrast, mRNA levels of acyl-CoA:diacylglycerol acyltransferase 1 (Dgat1), involved in lipid droplet biogenesis, were upregulated at 22 days of culture. Moreover, NP-exposed testes accumulated cholesterol, consistent with the observed disruption of steroid hormone production. Together, these results highlight the detrimental impact of NP on prepubertal spermatogenesis and germ cell maintenance, unveiling part of the mechanisms through which this compound exerts its toxic effects, namely alterations in lipid metabolism and steroid hormone synthesis.