Protective effects of moringa oleifera aqueous leaf extract on Tm3 Leydig cells against hydrogen peroxide-induced oxidative stress

Abstract

Introduction: Moringa oleifera (M. oleifera) is a medicinal plant rich in dietary antioxidants that may protect reproductive cells from oxidative stress-induced damage and male fertility problems. This study investigated M. oleifera aqueous leaf extracts (ALE) effects on TM3 Leydig cells exposed to hydrogen peroxide (H2O2). Methods: M. oleifera ALE and ascorbic acid (10−1000 µg/mL) were assessed for antioxidant activity. TM3 Leydig cells were exposed to various concentrations of M. oleifera ALE (0−1000 µg/mL) or α-tocopherol (50 µM) for 24 h, alongside 200 µM H2O2, with or without hCG (4 ng/mL). Cell viability, testosterone production, mitochondrial membrane potential (MMP), total protein and oxidative stress parameters (reactive oxygen species [ROS], superoxide dismutase [SOD], reduced glutathione [GSH], total antioxidant capacity [TAC] and lipid peroxidation [MDA]) were assessed. Results: Ascorbic acid demonstrated superior DPPH radical scavenging and FRAP activity compared to M. oleifera ALE at most concentrations (p < 0.0001). H2O2 (200 µM) exposure significantly reduced cell viability, MMP, SOD, GSH and TAC levels, while increasing ROS and MDA levels (p < 0.05). Treatment with M. oleifera ALE or α-tocopherol significantly increased cell viability, total protein, SOD activity and GSH levels while reducing ROS and MDA compared to H2O2 control (p < 0.05). However, testosterone levels remained unchanged (p > 0.05). Conclusions: M. oleifera ALE effectively protects the TM3 Leydig cells from H2O2-induced oxidative stress by restoring the redox balance, though unchanged testosterone levels suggest limitations in using TM3 Leydig cells for assessing testosterone synthesis.