In recent years, it has been suggested that oxidative stress could contribute to biological effects of polychlorinated biphenyl (PCB) exposures. During the current period of support we used in vitro cultures of human non-malignant breast (MCF10A) and prostate (RWPE-1) epithelial cells to determine the cellular effects of PCB 3, PCB 77, PCB 153, Aroclor 1254, and 2-(4-chlorophenyl)benzo-1,4-quinone (4-CI-BQ, a metabolite of PCB 3). 4-CI-BQ and PCB 153 (3 ?mu?M, 3-5 d) significantly increased mitochondria-generated reactive oxygen species (ROS) levels resulting in DNA damage, changes in MnSOD activity and cyclin D1 protein levels, inhibition in cell proliferation, and increased cytotoxicity. Antioxidant-treatment of cells prior to and following the PCB treatment ameliorated these effects. Interestingly, 1 micro molar of PCB 153 treatment decreased cyclin D1 protein turnover, while higher concentrations (3-20 micro molar) enhanced cyclin D1 degradation. This biphasic response of PCB 153 was associated with a small increase in cell number for the lower concentrations, and inhibition in cell proliferation for the higher concentrations. PCB 153-induced changes in cyclin D1 protein levels were also associated with changes in pyruvate kinase and hexokinase II protein levels. These results led us to hypothesize that redox signaling mediated by ROS (O{2}* and H2O2) disrupts coordinate regulation of SOD2 and cyclin D1 expression, which contributes to both stimulatory and cytostatic effects of PCBs on cellular proliferation.
Aim 1 : Determine if PCB-induced ROS-signaling perturbs transitions between quiescent (G{0}) and proliferative (G{1}, S, G{2}, and M) growth in human epithelial (breast and lung), and lung fibroblasts cultured in vitro.
Aim 2 : Determine the mechanisms regulating SOD2 activity in PCB-treated quiescent and proliferating epithelial and fibroblast cells cultured In vitro.
Aim 3 : Determine the mechanisms regulating cyclin D1 protein levels in PCB-treated epithelial and fibroblast cells cultured in vitro.
Aim 4 : Determine whether loss of SOD2 expression perturbs PCB-induced alterations in cyclin D1 expressions in vivo in breast, liver, and lung tissues of C57BL/6 Floxed SOD2 mice. Determine if application of small molecular weight antioxidants could suppress these effects.
The proposed research is innovative because of the hypothesis that inter-organelle communication via ROS-signaling is essential to prevent aberrant proliferation following PCB-induced oxidative injury. The application of antioxidants being tested is also innovative because it could lead to the development of viable redox-based countermeasures to mitigate PCB-induced abnormalities in cellular growth.
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