Mono-(2-ethylhexyl) phthalate (MEHP) affects the intercellular junctions of Sertoli cell: a potential role of oxidative stress.

SCHTEINGART HELENA FEDORA; SOBARZO CM; NOGUEIRA DE MORAIS, R; LUSTIG L; DENDUCHIS B

San Francisco

Congreso; ENDO 2013, 95th Annual Meeting & Expo of the Endocrine Society.; 2013

Endocrine Society

MEHP, an active metabolite of di-(2-ethylhexyl) phthalate (DEHP), is recognized as a reproductive toxicant, causing testicular atrophy and disruption of cellular redox state. Environmental toxicants trigger oxidative stress in the testis; increasing reactive oxygen species that led to lipid peroxidation (LPO) of cell membranes. We previously described (1) high levels of Glutathione (GSH) in Sertoli cell (SC), essential for cell protection against oxidative stress. Glutathione-S-Transferases (G-S-T), is present in the testis and catalyze conjugation of GSH with toxic substrates, such MEHP. We previously observed (2) that DEHP induces in vivo changes in the expression of specific junction proteins in rat testis. The aim of this study is to analyze MEHP effect on the integrity of intercellular junctions on SC primary cultures and to evaluate oxidative stress and GSH role. Sertoli cells were isolated from 20-day-old male rats and cultured for 5 days. Cells were exposed to MEHP (200 µM) alone or MEHP plus N-Acetyl-cysteine (NAC - 1mM) during the last 24 h of culture. The localization and expression of adherens (N-cadherin, α, ß and p120-catenins), tight (occludin, claudin-11, zonula occludens-1(ZO-1)), and gap (connexin 43 (Cx-43)) junction proteins were evaluated by immunofluorescence (IF) and Western blot (Wb) (2). Intracellular GSH level and G-S-T activity were determined by spectrophotometry (2). Alternatively, SC were treated only the last hour, and LPO levels were quantified by the FOX2 method (3). Results were expressed as percentage of control value, ANOVA and Tukey?s test. As an indicative of oxidative stress, LPO concentrations (161±17%, P