Cadmium (Cd) is an established human lung carcinogen and causes severe injury to many other organs. Cd is widely used in industries and also occurs as an important environmental pollutant and an ingredient in cigarette. Thus, occupational exposure and cigarette smoking pose risks to human health. Although Cd binds to protein sulfhydryls (PSH) and DNA bases, most Cd short-term mutagenicity tests are negative. Thus, the molecular mechanisms and intracellular events leading to cell injury/cancer development elicited by Cd remain unclear. Recent findings of Cd-induced severe injury to microtubules (MT) in human bronchial epithelia cells (NHBEC) and rat lung RFL6 cells led to the following working hypothesis to explain the molecular mechanisms of the MT cytoskeletal toxicology relevant to Cd bronchogenic carcinogenesis. The binding of Cd to the sulfhydryls of MT proteins (e.g. tubulin), perturbs its dynamics in favor of MT disassembly, which occurs as a collective result of blocking essential tubulin PSH preventing GDP-bound tubulins from exchanging their GDP for GTP, stimulating GTP hydrolysis on MT, and interference with MT associated proteins (MAPs). Subsequently, the increased free tubulin triggers an autoregulatory feedback mechanism inducing tubulin mRNA instability and inhibition of tubulin synthesis, thus further worsening the MT integrity. In intact cells the MT integrity can be modulated by cellular glutathione (GSH) and metallothionein. Because of their vital roles in many cellular processes, the MT structural impairment could translate into functional deficiencies resulting in pathological consequences namely, cell cycle perturbation and chromosomal instability, two key events intrinsically involved in the mechanisms of Cd chemical pathology leading to lung carcinogenesis. The overall goal of the proposed research is to test this hypothesis at multiple levels focusing on molecular mechanisms and consequences of Cd-induced MT pathology in vitro, in cultured lung and in an animal model.
Specific Aims are: 1. In vitro studies with purified MT proteins to examine Cd interaction with """"""""assembly-critical"""""""" tubulin -SH groups and GTP hydrolysis on MT and the exchange of GDP for GTP on tubulin. 2. Studies with cultured lung cells to examine: Cd binding to tubulin and MAPs; expression of tubulin protein and mRNA; molecular changes in MAPS and their phosphorylation; and modulation of MT injury by GSH, metallothionein and PSH. 3. Studies with cultured lung cells to examine pathological consequences of Cd-induced MT injury focusing on cell cycle perturbation and chromosomal instability. 4A. To conduct a rat model study to link MT damage to chromosomal instability and lung carcinogenesis. and 4B. To create an in vivo model closer to human lung bronchogenic neoplasia by injecting nude mice with Cd- transformed HBEC. The outcome of the proposed research should enhance our understanding of the molecular basis of Cd lung carcinogenesis in humans.

National Institute of Health (NIH)
National Institute of Environmental Health Sciences (NIEHS)
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Alcohol and Toxicology Subcommittee 4 (ALTX)
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Tinkle, Sally S
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Boston University
Schools of Medicine
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Li, Wande; Zhao, Yinzhi; Gantz, Donald L et al. (2003) Nickel (Ni2+) enhancement of microtubule assembly in vitro is dependent on GTP function. Toxicol Appl Pharmacol 193:202-8