Semester

Spring

Date of Graduation

2007

Document Type

Dissertation

Degree Type

PhD

College

School of Medicine

Department

Physiology, Pharmacology & Neuroscience

Committee Chair

Yon Rojanasakul.

Abstract

Hexavalent chromium [Cr(VI)] compounds are redox cycling environmental carcinogens that induce apoptosis as the primary mode of cell death. Apoptosis is abnormally regulated in various disorders including cancer. Therefore, to understand the etiologies of these diseases, it is important to delineate the biochemical and molecular pathways involved in the regulation of apoptosis. The main objective of this study was to characterize the molecular mechanisms involved in Cr(VI)-induced apoptosis and malignant transformation. We found that both death receptor and mitochondrial pathways of apoptosis are involved in Cr(VI)-induced apoptosis, with the latter being more dominant. Consequently, overexpression of the mitochondrial anti-apoptotic protein Bcl-2 blocked Cr(VI)-induced apoptosis in human lung epithelial cells. We further observed that reactive oxygen species (ROS) play a critical role in Cr(VI)-induced apoptosis by acting through the mitochondrial death pathway. Superoxide anion (˙O2 -) was found to be the major ROS involved in Cr(VI)-induced apoptosis that exerted its effect by degrading Bcl-2 protein through the ubiquitin-proteasomal pathway. Furthermore, nitric oxide (NO) scavenged ˙O2 - to form peroxynitrite (ONOO-) and negatively regulated Cr(VI)-induced apoptosis. The mechanism by which NO exerted its anti-apoptotic effect involved upregulation of Bcl-2 via S-nitrosylation that prevented its ubiquitination and subsequent proteasomal degradation. Additionally, we established an in vitro model for studying Cr(VI)-induced malignant transformation by subjecting non-tumorigenic human lung epithelial Beas-2B cell line to long-term Cr(VI) exposure. Cr(VI) transformed cells exhibited clear signs of malignancy such as loss of contact inhibition and increased colony formation as compared to the passage-matched original cell-line. Cr(VI) transformed cells showed decreased apoptosis and ROS production and increased NO as well as Bcl-2 expression. These observations confirmed that NO mediated stabilization of Bcl-2 is an important event in Cr(VI) induced carcinogenesis. Taken together, our study reveals a novel regulatory mechanism that could be important in apoptosis resistance in response to Cr(VI) exposure. Additionally, this study demonstrated Cr(VI)-induced malignant transformation of a human lung epithelial cell line, establishing an important in vitro model for studying the molecular mechanisms involved in Cr(VI)-induced carcinogenesis. This study provides new mechanistic insights about environmental carcinogen induced human lung cancer. Since Cr(VI) is a paradigm of carcinogenic transition metals, the inferences from this study may be broadly applied to general metal carcinogenesis.

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