Abstract. Gap junctional intercellular communication (GJIC) of cultured mouse epidermal cells is mediated by a gap junction protein, connexin 43, and is dependent on the calcium concentration in the medium, with higher GJIC in a high-calcium (1.2 mM) medium . In several mouse epidermal cell lines, we found a good correlation between the level of GJIC and that of immunohistochemical staining of E-cadherin, a calciumdependent cell adhesion molecule, at cell-cell contact areas. The variant cell line P3/22 showed both low GJIC and E-cadherin protein expression in low-and high-Cal+ media. P3/22 cells showed very low E-cadherin mRNA expression. To test directly whether E-cadherin is involved in the Caz+-dependent regulation G

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1Excessive free radical generation is thought to contribute to tissue injury in a broad spectrum of diseases. A particular constraint in addressing this hypothesis has been the inability to assess free radical generation in vivo and the lack of information on drugs or vitamins which act as effective antioxidants in vivo. 2Traditional approaches have relied upon measures of substrate oxidizability or spin trapping of free radical adducts ex vivo. It is unknown how these measurements might relate, in a quantitative fashion, to the generation of reactive oxygen species in vivo. Isoeicosanoids are free radical catalyzed products of arachidonic acid. One of these compounds, 8‐epi prostaglandin F2α (8‐epi PGF2α) exhibits biological activity and may function as an autacoid. Specific analysis of this 8‐epi PGF2α isomer indicates that it is elevated in certain syndromes thought to be associated with oxidant stress. These include vascular reperfusion, paracetamol poisoning and liver cirrhosis. Apparently healthy individuals who smoke cigarettes or consume alcohol exhibit dose dependent increments in excretion of 8‐epi PGF2α. Excretion is depressed by antioxidant vitamins, although not by the nonspecific cyclooxygenase (COX) inhibitor, aspirin, even though 8‐epi PGF2α may be formed by either COX‐1 or COX‐2. 3Specific analysis of this and other isoeicosanoids may afford an opportunity to evaluate the effects of antioxidant interventions in human diseases characterized by excessive lipid peroxidation in vivo.

It is important for public health authorities to set a scientifically sound guideline value for the safe ingestion of copper in drinking water. To date, the principal health-based guideline values have been set by the US Environmental Protection Agency (1.3 mg Cu/L) and the World Health Organization (2.0 mg Cu/L). However, close examination of the data and assumptions used in the derivation of these values reveals a paucity of scientifically defensible information. Several international groups are now reviewing this issue, and others have begun epidemiologic studies that may provide useful copper exposure and toxicity data. Investigations of acute copper toxicity in human populations are most likely to affect future revisions of the guideline value for copper in drinking water.

We have studied the gap junctional intercellular communication (GJIC) of immortalized and tumourigenic human keratinocyte cell lines and of a spontaneously immortalized non-tumourigenic and a highly differentiating keratinocyte cell line (HaCaT) as the control. In homologous cultures, the GJIC capacity of five squamous cell carcinoma-derived cell lines was 1-27% that of the HaCaT cells. Ha-ras-transfected HaCaT cells with tumourigenic potential and an SV40 DNA-immortalized cell line had markedly reduced GJIC capacities. Northern analysis and immunohistochemistry showed that connexin (Cx) 43 is the major gap junction protein expressed in the communicating cells. They do not express Cx 26 or 32. The low or absent communication observed in certain cell lines was due in some to a lack of Cx 43 gene expression, but in others to aberrant localization of the gap junction protein. GJIC of these cell lines, as well as that of primary normal human epidermal keratinocytes, was susceptible to 12-O-tetradecanoylphorbol-13-acetate-mediated inhibition. Moreover, GJIC of HaCaT cells and their tumourigenic derivatives is Ca(2+)-dependent. These results, when compared with those previously obtained for mouse keratinocyte cell lines, reveal that GJIC of human keratinocytes was correlated to the degree of differentiation and is controlled in a similar way to that of murine keratinocytes. Aberrant GJIC seems to be a common feature of human and murine skin carcinogenesis.

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A rat liver gap junction (GJ) cDNA probe that detects mRNA encoding the 32 Kd GJ-protein (connexin 32) was employed to study GJ-protein gene expression in rat liver tumors induced by a single exposure to diethylnitrosamine (DEN) followed by exposure to 2-acetylaminofluorene (AAF)/CCl4/AAF or induced by systemic administration of N-ethyl-N-hydroxyethylnitrosamine (EHEN). All carcinomas generated by these carcinogens showed markedly reduced levels of GJ-protein mRNA. This may indicate that GJ-protein levels and gap-junctional intercellular communication (GJIC) capacity are also severely compromised. Moreover, all hyperplastic nodules also showed a reduced level of GJ-protein mRNA. Taken together with our earlier finding that the liver tumor promoter phenobarbital inhibits GJ-protein gene expression, these results suggest that deranged GJIC is a relatively early event in liver multistage carcinogenesis. A range of other cDNA probes was also used to characterize gene expression in the DEN-induced tumors. Induction of expression was seen for glutathione S-transferase (placental form) (GST-P), gamma-glutamyltranspeptidase (GGT), and c-raf but not for c-Ha-ras or c-myc.