| Formaldehyde is a well-known environmental pollutant found in automobile emissions and tobacco smoke. However, formaldehyde is also a ubiquitous compound found in all biological systems liberated by the metabolism of a variety of exogenous and endogenous precursors. Various N-, O- and S-methyl compounds are oxidatively demethylated by demethylases resulting in formaldeyhyde formation. The International Agency for Research on Cancer (IARC) classified formaldehyde as a probable human carcinogen, and several cohort-control and case-control studies link formaldehyde exposure to nasopharyngeal carcinoma (IARC,2004). It is reported that the mechanism of cytotoxicity induced by formaldehyde has some thing to do with the gene regulation, and there is evidence that formaldehyde could induce DNA-protein crosslinks (DPC). But it is still controversial whether inhaled formaldehyde has a site distant toxicity. IARC concluded that there was sufficient epidemiological evidence for a causal association between leukemia and occupational exposure to formaldehyde, but experimental evidence was little, and stated that more research is needed in this area. To explore whether formaldehyde has a site distant toxicity, KCl-SDS assay was applied to determine the amount of DNA-protein crosslinks in the brain and testicle cells of formaldehyde inhaled Wistar male rats.Paxillin is a focal adhesion-associated, phosphotyrosine-containing protein that may play a role in several signaling pathways. Paxillin contains a number of motifs that mediate protein protein interactions, including LD motifs, LIM domains, an SH3 domain-binding site and SH2 domain-binding sites. These motifs serve as docking sites for cytoskeletal proteins, tyrosine kinases, serine/threonine kinases, GTPase activating proteins and other adaptor proteins, which play an important role in cell signaling. Recent study found that formaldehyde could activate MAP kinases and induce phosphorylation of Paxillin in HT-29 cells, and therefore induced serial signaling events, which provides us new ways to study the physiological and toxic mechanisms of formaldehyde. In the present study, we treated Hela cells with formaldehyde, and then investigated the expression and phosphorylation level of Paxillin in Hela cells, which aimed to provide evidence for research on formaldehyde induced cancer and formaldehyde as a signaling molecule.1. Study on DNA-protein crosslinks in Brain and Testicle Cells Induced by Formaldehyde in Male RatsTo explore the DNA-protein crosslinks (DPC) effect induced by gaseous formaldehyde (FA) in the brain and testicle cells of Wistar male rats. In the present study, Wistar male rats were randomly divided into four groups,6 in each and exposed to gaseous FA at doses of 0 (control group),0.5, 1.0 and 3.0 mg/m3 for 72 consecutive hours. The brain and testicle cells were used as experimental materials. The KCl-SDS assay was applied to determine the amount of DNA-protein crosslinks. Results showed that compared with the control group, no significant change in DPC was observed at the low concentrations exposure group (0.5mg/m3, P>0.05), but as formaldehyde concentration increased, the coefficient of DPC also increased gradually. Higher concentrations exposure (1.0,3.0mg/m3) resulted in significant elevation of DPC amount compared with the control group (P<0.01).2. The Regulation Mechanism of Formaldehyde on Paxillin Function in Hela CellsFormaldehyde is a gas and a free radical which is now recognized to have very important physiological roles. However, the mechanism by which formaldehyde functions is still poorly characterized. In the present study, we investigated the effect of formaldehyde on the focal adhesion protein Paxillin in Hela cells. We found that non-lethal formaldehyde concentrations had significant influences on both the levels of gene expression and the protein phosphorylation of Paxillin. Western blot analysis showed that when treated with formaldehyde at the concentrations of 0.5 mM,1.0 mM and 2.0 mM for 1 hour, or at the concentration of 1.0 mM for 0.5,1 and 2 hours, Paxillin expression in Hela cells was down-regulated and dose-dependent. On the other hand, the phosphorylation level of Paxillin tyrosine was detected by immunofluorescence assay and was significantly increased when Hela cells were incubated with 2.0 mM formaldehyde for 2 hours. Our data suggest that formaldehyde may play its physiological role via modification of the Paxillin signaling pathway, i.e. via the down regulated gene expression level and the up regulated tyrosine phosphorylation level of Paxillin.
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