Fig. 3

Mtb-induced cell death is independent of caspase activation. a Western blot analysis of proteolytic cleavage of caspase 3 in Mtb-infected J774.2 Mφ (MOI 5). Equal amounts of protein from infected J774.2 Mφ treated with or without dexamethasone (5 µM) were subjected to immunoblot analysis for cleaved caspase 3 following 24 and 30 h of infection. β-Actin was used as a loading control and staurosporine (1 µM) as a positive control. Images are representative of two individual experiments. b MRC-5 lung fibroblasts were infected with Mtb (MOI 10) and treated with dexamethasone (5 µM), doramapimod (10 µM), or Z-VAD-FMK (10 µM). After 48 h caspase 3 and caspase 7 activity was assessed using a luminescent probe. Uninfected and staurosporine (1 µM)-treated cells were used as controls. c, d Effect of caspase 3 and caspase 7 inhibition using the pan-caspase inhibitor Z-VAD-FMK (10 µM) on survival of infected MRC-5 lung fibroblasts (c) and human Mφ from healthy donors (d). Viable fibroblasts were detected using Prestoblue and Mφ were quantified by DAPI staining. Representative data from two experiments with multiple replicates are shown in b–d. Results are expressed as mean ± SEM. Analysis was done using unpaired t-test (ns not significant; **p ≤ 0.01; ***p ≤ 0.001)