Fig. 2
Recombinant ESAT-6 protein inhibits M. tb-induced apoptosis of THP-1(A) via Caspase 9/Caspase 3. A The effects of M. tb and ESAT-6 on macrophages. Cellular apoptosis was evaluated by flow cytometry. THP-1(A) cells (1 × 106) were co-incubated with M. tb or/and different concentrations of ESAT-6 for 24 h. The dose-dependent response of EAST-6 on macrophage apoptosis was detected by the Annexin V FITC/PI method. B Quantitative data of apoptosis in ESAT-6 and/or mycobacteria stimulated macrophages are shown. C THP-1(A) cells (1 × 106) were co-incubated with M. tb or/and different concentrations of ESAT-6 for 24 h. Cells received ESAT-6 and M. tb were stained with JC-1 for mitochondrial membrane potential measurement. A representative group image of stained cells was observed under an inverted fluorescence microscope. D Quantitative data of MFI in ESAT-6 and/or mycobacteria stimulated macrophages are shown. After different concentrations of ESAT-6 were simultaneously added with M. tb on THP-1 (A) cells for 24 h, the cells were analyzed by Western blotting for E Casepase 3, cleaved-Caspase 3, Caspase 9, and cleaved-Caspase 9 protein levels, F PARP and it’s activated form, F BAX, and Bcl-2 were analyzed by Western blotting in THP-1(A) cells challenged with M. tb. (G-I) Densitometric quantification of representative blots relative to β-actin levels. Data are representative of at least three different experiments. Results are mean ± SEM. *p < 0.05,**p < 0.01 and ***p < 0.001, respectively, compared with the M. tb group