Antiapoptotic effects of anthocyanin from the seed coat of black soybean against oxidative damage of human lens epithelial cell induced by H2O2.

PURPOSE

To describe the protective effect of anthocyanin from black soybean in human lens epithelial cell line (HLE-B3) under H2O2-induced oxidative stress.

METHODS

Cytotoxicity of anthocyanin and H2O2 were determined by Cell Counting Kit-8 test. Viability of HLE-B3 cells under various H2O2 concentration (0, 50 and 100 μM) with or without pretreatment of anthocyanin (0, 50, 100 and 200 μg/ml) was measured. After quantifying the percentage of the apoptosis by Annexin V assay and APO-BrdU TUNEL assay, we conducted western blot and immunostaining of apoptosis-related molecules; Bcl2, BAD, BAX, p53 and caspase-3. To confirm the effect of anthocyanin on an ex vivo model, its effect on cultures of the lenses of porcine were examined.

RESULTS

Anthocyanin reduced cell death of HLE-B3 under H2O2-induced oxidative stress in a dose-dependent manner. In Annexin V analysis, anthocyanin protected HLE-B3 cells from apoptosis. H2O2 increased the expression of BAX, BAD, p53 and caspase-3 in a time-dependent manner, those of which anthocyanin significantly decreased. On the other hand, Bcl2 was increased from anthocyanin-treated lens cells. And in anthocyanin-treated lens organ culture, transparency was maintained.

CONCLUSIONS

This study showed that anthocyanin protects HLE-B3 cells under oxidative stress from apoptosis, and the mechanism of the effect is related to the intrinsic pathway of apoptosis. Anthocyanin has a potential in prevention of cataract.