A cell model illustrating cardioprotection of EGCg on H
-induced oxidative stress in H9c2 cells. (a) Phase contrast microscopy showing cell morphology of H9c2 cells in the conditions of control (left top), 20 μM EGCg treatment for 30 min (right top), 400 μM H2O2 exposure for 30 min (left bottom), and 20 μM EGCg pre-treatment for 30 min followed by 400 μM H2O2 exposure for 30 min (right bottom). Calibration bar of 200 μm as indicated. (b) MTT assay of cell viability after incubation with 0, 100, 200, and 400 μM H2O2 with or without 0, 10, and 20 μM EGCg for 30 min. (c) Measurements of intracellular ROS formation by DCF-DA in H9c2 cells. The fluorescence changes of DCF-DA-loaded cells were measured every 10 min before and after the addition of 400 μM H2O2 with 0-50 μM EGCg as indicated by fluorescence spectrophotometry. The fluorescence excitation maximum for DCF-DA was 495 nm, and the corresponding emission maximum was 527 nm. (d) Effects of H2O2 and/or EGCg on intracellular Ca2+ levels in H9c2 cells. Cellular Ca2+ levels were measured using the Fura-2 fluorescence ratio (F340/F380) in H9c2 cells cultured in the conditions of control, 20 μM ECGg treatment for 30 min, and 400 μM H2O2 exposure for 30 min with and/or without 20 μM ECGg treatment for 30 min, then during the measurement in PBS for 3 min periods. The F340/F380 ratio was continuously monitored. In b, c and d, the values are the mean ± SEM (n = 6), with *, # indicating a significant difference compared to the untreated cells or the H2O2-treated cells, respectively.