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This Article Full Text Full Text (PDF) All Versions of this Article: 293/2/L393    most recent 00392.2006v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in ISI Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Lai, Y.-M. Articles by Antony, V. B. Search for Related Content PubMed PubMed Citation Articles by Lai, Y.-M. Articles by Antony, V. B.

Induction of cell cycle arrest and apoptosis by BCG infection in cultured human bronchial airway epithelial cells

Division of Pulmonary, Critical Care, and Sleep Medicine, Department of Medicine, College of Medicine, University of Florida, Gainesville, Florida

Submitted 3 October 2006 ; accepted in final form 21 May 2007

Bronchial airway epithelial cells (BAEpC) are among the first cells to encounter M. tuberculosis following airborne infection. However, the response of BAEpC to M. tuberculosis infection has been little studied. This study investigates the response of a human BAEpC cell line (BEAS-2B) to infection with Mycobacterium bovis Bacille Calmette Guerin (BCG). Cultured human BEAS-2B cells were experimentally infected with BCG. Uninfected BEAS-2B cultures were included as controls. Following infection, BEAS-2B cells were evaluated by various methods at various time points up to 3 days. Cell proliferation was evaluated by cellular bioreduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Distribution of cells along the cell cycle was evaluated by FACS analysis of cellular DNA. Apoptotic cells were identified by cell death ELISA and the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling method. Eighty-four apoptosis-relevant genes were screened by PCR gene microarray. Translation of Fas, Fas ligand (Fas-L), and Fas-associated death domain (FADD) were evaluated quantitatively by real-time PCR. Expression of Fas and FADD proteins was evaluated by immunofluorescence and Western blot. Activity of caspase-3 and caspase-8 was evaluated by colorimetric assay of their enzymatic activity. BCG infection of BEAS-2B cells inhibits proliferation, induces cell cycle arrest at the G₀/G₁ phase, causes apoptosis, modulates transcription of multiple apoptosis-relevant genes, promotes translation of Fas, Fas-L, and FADD, upregulates expression of Fas and FADD proteins, and increases activity of caspase-3 and caspase-8. Infection with BCG does not cause any significant change in the secretion of TGF-. The roles of Fas and FADD as mediators of BCG-induced apoptosis in BEAS-2B cells were tested by partial blockade of Fas and FADD expression with silencing RNA. Partial blockade of Fas or FADD expression results in a decreased apoptotic response to BCG infection. In conclusion, BCG induces cell cycle arrest and apoptosis in BEAS-2B cells. BCG induced apoptosis of BEAS-2B cells via the Fas death receptor pathway.

Bacille Calmette Guerin; Fas; Fas ligand; Fas-associated death domain protein; caspase-3; caspase-8