Effects of boric acid on cell death and oxidative stress of mouse TM3 Leydig cells in vitro

Background: Boron (B) is an abundant element on earth and presents at physiological pH in the form of boric acid (BA). It has both positive and negative effects on biological systems. BA and sodium borates have been considered as being toxic to the reproduction system in animal experiments. Unfortunately, the molecular mechanism underlying the toxic effects of BA is not fully understood. Methods: Here, we demonstrate the influence of BA on mouse TM3 Leydig cells which are male reproductive system cells targeted by BA exposure. The cytotoxicity was evaluated by MTT and NRU assays. Annexin V-FITC/PI double staining kit, mitochondria membrane potential (Delta psi m) assay kit with JC-1 and caspase-3 colorimetric assay kit were used to indicate the cell death pathway. To estimate the role of oxidative stress in BA induced toxicity, glutathione (GSH) level, catalase (CAT) and superoxide dismutase (SOD) activities were measured manually. Results: The cell viability assays showed that BA was not cytotoxic within the tested concentrations up to 1000 mu M. Sub-toxic concentrations were used for detecting oxidative stress status. BA exposure was significantly reduced GSH level at 1000 mu M and CAT activity in a concentration-dependent manner. However, SOD activity was increased at the tested concentrations (100-1000 mu M). Moreover, Delta psi m was significantly decreased at 500 and 1000 mu M of BA, while caspase-3 activity was not changed apparently. Conclusion: These findings demonstrated that BA is not cytotoxic and apoptotic but may slightly induces oxidative stress in TM3 Leydig cells at higher concentrations.