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Title: Enhanced Gadd45 expression and delayed G2/M progression are p53 dependent in zinc-supplemented human bronchial epithelial cells

item SHIH, RITA - University Of Maryland
item WONG, STEPHEN - University Of Maryland
item Schoene, Norberta
item ZHANG, JUN JUN - University Of Maryland
item LEI, KAI - University Of Maryland

Submitted to: Experimental Biology and Medicine
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/15/2010
Publication Date: 8/2/2010
Citation: Shih, R., Wong, S., Schoene, N.W., Zhang, J., Lei, K. 2010. Enhanced Gadd45 expression and delayed G2/M progression are p53 dependent in zinc-supplemented human bronchial epithelial cells. Experimental Biology and Medicine. 235(8):932-940.

Interpretive Summary: Zinc is an essential nutrient that is needed to regulate protein synthesis and function and is necessary for proper responses to stress for cell survival. Earlier we had shown that high amounts of zinc can be detrimental to the stress responses of proteins leading to conditions that can promote inflammation and cancer, especially in lung cells. To gain further information about mechanisms that produce such harmful effects with high zinc, we again looked at cultured normal cells from the lung grown in 4 levels of zinc (0.4, 4, 16, and 32 micromoles/liter). We confirmed that with the highest amount of zinc both messenger RNA and protein amounts for several stress-induced proteins were increased compared to the cells grown in 0.4, 4, and 16 micromoles/liter of zinc. Most importantly, we identified additional proteins that were altered by high zinc that provide new insights into mechanisms that explain the deleterious effects of high zinc. This is an important observation and is a significant advance in providing information to dietitians, nutritionists, and other researchers who advise consumers and conduct research on potential problems associated with over-supplementation with zinc.

Technical Abstract: Zinc is an essential nutrient for humans; however, this study demonstrated for the first time that an elevated zinc status, created by culturing cells at optimal plasma zinc concentration attainable by oral zinc supplementation, is cytotoxic for normal human bronchial epithelial (NHBE) cells. p53 plays a central role in the modulation of cell signal transduction in response to the stress from DNA damage, hypoxia, and oncogene activation. This study was designed to determine whether the previously reported increased Gadd45 expression and delayed G2/M cell cycle progression in zinc supplemented NHBE cells is p53-dependent, to decipher the mechanisms responsible for the regulation of Gadd45 expressions by p53, and to elucidate the Gadd45 functions in impaired cell growth and cell cycle progression in NHBE cells. Cells were cultured for one passage in different micromolar concentrations of zinc: <0.4uM (ZD)a severe zinc deficient culture medium; 4uM (ZN) as a normal zinc level in culture medium; 16uM (ZA) as normal human plasma zinc level; and 32uM (ZS) as the high end of plasma zinc attainable by oral supplementation. Inhibition of cell growth and up-regulation of p53 mRNA and protein expression were observed in ZS cells. Most importantly, ZS treatment also enhanced Gadd45 nuclear protein level and promoter activity, decreased CDK1-Cyclin B1 level, and delayed G2/M cell cycle progression. These changes were normalized to those observed in ZN by treating ZS cells with pifithrin, an inhibitor of p53 trans-activation activity. Thus, our findings support the p53 dependency of the Gadd45-CDK1/cyclin B1-G2/M cell cycle progression pathway in ZS NHBE cells.