M. leprae inhibits apoptosis in THP-1 cells by downregulation of Bad and Bak and upregulation of Mcl-1 gene expression.
BACKGROUND
Virulent Mycobacterium leprae interfere with host defense mechanisms such as cytokine activation and apoptosis. The mitochondrial pathway of apoptosis is regulated by the Bcl-2 family of proteins. Expression of Fas ligand and apoptotic proteins is found in leprosy lesions and M. leprae has been shown to activate pro-apoptotic Bcl-2 genes, Bak and Bax. However, the mechanism by which M. leprae modulates apoptosis is as yet unclear. We investigated expression of apoptotic genes in THP-1 monocytes in response to infection by M. leprae and non-pathogenic M. bovis BCG.


RESULTS
M. leprae did not induce apoptosis in THP-1 cells, while BCG induced a significant loss of cell viability by 18 h post-infection at both (multiplicity of infection) MOI-10 and 20, with an increase by 48 h. BCG-induced cell death was accompanied by characteristic apoptotic DNA laddering in cells. Non-viable BCG had a limited effect on host cell death suggesting that BCG-induced apoptosis was a function of mycobacterial viability. M. leprae also activated lower levels of TNF-alpha secretion and TNF-alpha mRNA expression than BCG. Mycobacterium-induced activation of apoptotic gene expression was determined over a time course of infection. M. leprae reduced Bad and Bak mRNA expression by 18 h post-stimulation, with a further decrease at 48 h. Outcome of cell viability is determined by the ratio between pro- and anti-apoptotic proteins present in the cell. M. leprae infection resulted in downregulation of gene expression ratios, Bad/Bcl-2 mRNA by 39% and Bak/Bcl-2 mRNA by 23%. In contrast, live BCG increased Bad/Bcl-2 mRNA (29 %) but had a negligible effect on Bak/Bcl-2 mRNA. Heat killed BCG induced only a negligible (1-4 %) change in mRNA expression of either Bak/Bcl-2 or Bad/Bcl-2. Additionally, M. leprae upregulated the expression of anti-apoptotic gene Mcl-1 while, BCG downregulated Mcl-1 mRNA.


CONCLUSIONS
This study proposes an association between mycobacterium-induced apoptosis in THP-1 cells and the regulation of Bcl-2 family of proteins. M. leprae restricts apoptosis in THP-1 cells by downregulation of Bad and Bak and upregulation of Mcl-1 mRNA expression.
Hasan
Zahra
Z
Department of Pathology and Microbiology, The Aga Khan University, Karachi, Pakistan. zahra.hasan@aku.edu
Ashraf
Mussarat
M
Tayyebi
Ali
A
Hussain
Rabia
R
D017209:Apoptosis / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D002470:Cell Survival / Q000235:genetics; D015536:Down-Regulation / Q000235:genetics; D004587:Electrophoresis, Agar Gel / Q000379:methods; D005786:Gene Expression Regulation / Q000235:genetics; D006801:Humans; D008856:Microscopy, Fluorescence / Q000379:methods; D009163:Mycobacterium bovis / Q000472:pathogenicity / Q000502:physiology; D009166:Mycobacterium leprae / Q000472:pathogenicity / Q000502:physiology*; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009363:Neoplasm Proteins / Q000235:genetics*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics*; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000235:genetics; D014774:Virulence; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000235:genetics*; D051025:bcl-Associated Death Protein / Q000235:genetics*
C494819:BAD protein, human; C494772:BAK1 protein, human; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009363:Neoplasm Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D014409:Tumor Necrosis Factor-alpha; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051025:bcl-Associated Death Protein