Hydrogen peroxide-induced cellular apoptosis is mediated by TGF-beta2 signaling pathway in cultured human lens epithelial cells.

The objective of this study was to investigate the signaling characteristics of transforming growth factor-beta2 (TGF-beta2) and the Smads (Caenorhabditis elegans, Sma; Drosophila mothers against dpp, Mad) signal pathway of cellular apoptosis induced by hydrogen peroxide with human lens epithelial cells (HLECs). HLECs were starved for 24 h before exposure to 0.1 mumol/ml of hydrogen peroxide in the presence and in the absence of 0.01 mug/ml of AF-302-NA, a monoclonal anti-TGF-beta2 neutralization antibody. Non-stimulated cells served as controls. Cell apoptosis was examined by in situ immunocytochemistry using terminal deoxynucleotidyl transferase dUTP-mediated biotin nick end labeling (TUNEL) and by flow cytometry (FCM) using Annexin V-FITC apoptosis detection. Gene expression was assessed using the reverse transcription-polymerase chain reaction (RT-PCR). Smad-4 localization was observed by immunocytochemistry. Hydrogen peroxide induced the accumulation of Smad-4 in the nucleus of HLECs, and upregulated the expression of TGF-beta receptors (TbetaRs) mRNA in HLECs, as well as upregulated the expression of the apoptotic gene bax, which leads HLECs to apoptosis. AF-302-NA decreased cellular apoptosis induced by hydrogen peroxide in HLECs and inhibited the translocation of Smad-4 from the cytoplasm to the cell nucleus. Moreover, AF-302-NA upregulated the expression of TbetaRs mRNA and downregulated the expression of bax mRNA in HLECs incubated with hydrogen peroxide. Our study demonstrated that the TGF-beta2 signal pathway participated in the apoptotic signal transfer and might be an initiator of cellular apoptosis of HLECs after incubation with hydrogen peroxide. Interruption of the TGF-beta2 signal pathway could partially protect HLECs from apoptosis induced by incubation with hydrogen peroxide.