View Abstract

A7609 - Cigarette Smoke Exposure Alters the Airway Epithelial Cell Fate by Modulating Mitochondrial Morphology and Bcl-2 Levels
Author Block: H. S. Chand1, S. S. Hussain2, S. George2, S. P. Singh3, R. D. Jayant2, M. Nair2, M. L. Sopori3; 1Immunology, Florida International University, Miami, FL, United States, 2Immunology, Florida International University, MIAMI, FL, United States, 3Lovelace Respiratory Research Institute, Albuquerque, NM, United States.
Rationale: Airway mucous response is an important component of the lung innate immunity. However, persistent dysregulated mucous can lead to airway obstruction and impaired lung functions. Cigarette smoke (CS) is one of the primary risk factor associated with the chronic bronchitis (CB). However, apart from smoking cessation fewer treatment options are available to mitigate mucus exacerbations. We have previously shown that a mitochondrial outer membrane-associated protein, Bcl-2, is induced in hyperplastic mucous cells. Therefore, we investigated the underlying mechanisms by which CS modulates Bcl-2 and mitochondrial morphology to cause mucous hypersecretion. Methods: The role of CS exposure in modulating Bcl-2 and secretory mucin, MUC5AC, expression was investigated in primary human airway epithelial cells (HAECs). Bronchial biopsies from ex-smokers with CB and axial airways from mouse exposed to CS were analyzed for mitochondrial morphology and oxidative response, Bcl-2 and mucin expression by high-resolution microscopy and molecular techniques. Bcl-2 was suppressed by retroviral shRNA constructs (shBcl-2) and Bcl-2 activity was blocked using a small molecule BH3 mimetic compound, ABT-263 that disrupts the interaction of Bcl-2 with apoptotic proteins. Expression of secretory mucin genes, MUC5AC, MUC5B and MUC1, and the mucous-regulating transcription factors, Foxa3 and SPDEF, and the EGFR and ERK1/2 pathways were also probed. Results: Bronchial biopsies from ex-smokers with CMH showed significantly increased Bcl-2 and MUC5AC levels compared to control subjects with alteration in mitochondrial distribution and morphology. In a mouse model of CS exposure there was increased expression of Bcl-2 and Muc5ac in axial airways compared to controls. Similarly, cultured human airway epithelial cells (HAECs) exposed to CS displayed more fragmented and intermediate mitochondria. There was associated induction in the levels of Bcl-2 and MUC5AC via pathways involving EGFR and ERK1/2. CS treatment of differentiated HAECs caused a significant increase in the secretory mucin genes, MUC5AC, MUC5B and MUC16, and the mucous-regulating transcription factors, Foxa3 and SPDEF. Blocking Bcl-2 using shRNA approach or by using a small molecule inhibitor resulted in the downregulation of CS-induced mucus expression and oxidative stress via disrupting mitochondrial morphology and EGFR- and ERK-signaling resulting in induced apoptosis. This strategy significantly suppressed the number of CS-induced mucus cells in-vitro. Conclusions: CS exposure affects mucous response by modulating mitochondrial response and the levels of Bcl-2, the anti-apoptotic protein that promotes cell survival. Therefore, modulating the hyperplastic mucous cells by blocking Bcl-2 could be helpful in controlling the debilitating CS-induced mucous pathologies. Funded by: AI117560 and RG306208 (ALA)