TNF neutralizing antibody (1.1%) or pentoxifylline treated

cells (5.5%) also showed a very significant decrease in apoptosis compared to the untreated (20.0%) or nonspecific antibody treated cells (21.0%) (Figure 6). These results demonstrate that that apoptosis of BMDMs induced by nonpathogenic mycobacteria is dependent upon TNF secretion and caspase-3 activation. Figure 6 Macrophage apoptosis induction by a non-pathogenic mycoabcterium is caspase-3 and TNF-dependent. BMDMs from BALB/c mice were left untreated and uninfected (UT) or infected with M. smegmatis and then either left in medium (Msme) or treated with caspase-3 inhibitor (C3I), nonspecific chemical analog (C3I-A) neutralizing TNF antibody (TNF-Ab), nonspecific control Ab (Co-Ab) and TNF synthesis inhibitor pentoxifylline see more (PTX). The percentage of apoptotic cells out of 10,000 total cells was determined after 20 h using the hypodiploid PI flow cytometry assay and a representative histogram of two independent experiments performed in duplicates is shown. The increased cytokine secretion by macrophages upon infection with non-pathogenic M. smegmatis selleck kinase inhibitor versus facultative-pathogenic M. avium has been demonstrated in human and murine macrophages and human neutrophils [15, 34, 35]. Our study builds upon these previous results by extending the analysis

to include PP2 mouse several non-pathogenic versus several facultative-pathogenic mycobacteria. We underscore that the strong pro-inflammatory response elicited by macrophage might be a more general characteristic of non-pathogenic mycobacteria. The increase of TNF secretion induced by M. smegmatis in murine BMDM is dependent upon stimulation of the cAMP/protein kinase A pathway which results in prolonged ERK1/2 activation[15]. Org 27569 Furthermore, M. smegmatis infection leads to increase in TNF and NOS2 promoter activity but not infection with M. avium [15, 36]. The present study also extends upon these previous findings by linking the increase in TNF secretion to pro-apoptotic capacity of the non-pathogenic mycobacteria

(Figure 6) and characterizing this apoptosis pathway as being caspase-dependent (Figure 6). Non-pathogenic mycobacteria do not induce apoptosis in C57Bl/6 BMDM We demonstrated that non-pathogenic mycobacteria induce a strong apoptotic response and TNF secretion in BALB/c macrophages (Figures 1 and 5) when compared to facultative pathogenic mycobacteria. We also demonstrated that TNF plays a role in this apoptotic response (Figure 6). We therefore intended to further clarify the role of TNF by using TNF knock-out mice. Nevertheless, to our surprise we determined that BMDMs from C57Bl/6 wild-type mice, which is the genetic background of the TNF deficient mice, did not undergo apoptosis upon infection with non- and facultative-pathogenic mycobacteria using two different apoptosis detection assays (p > 0.05; Figure 7A and 7B).