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Exposure of LLC‐PK1 epithelial cell cultures to phorbol ester tumor promoters causes immediate translocation of protein kinase C‐α (PKC‐α) from cytosolic to membrane‐associated compartments. With a very similar time course, a dramatic and sustained increase in tight junctional (paracellular) permeability occurs. This increased permeability extends not only to salts and sugars but macromolecules as well. Fortyfold increases of transepithelial fluxes of biologically active EGF and insulin occur. Recovery of tight junction barrier function coincides with proteasomal downregulation of PKC‐α. The failure to downregulate activated membrane‐associated PKC‐α has correlated with the appearance of multilayered cell growth and persistent leakiness of tight junctions. Accelerated downregulation of PKC‐α results in only a partial and transient increase in tight junction permeability. Transfection of a dominant/ negative PKC‐α results in a slower increase in tight junction permeability in response to phorbol esters. In a separate study using rat colon, dimethylhydrazine (DMH)‐induced colon carcinogenesis has been preceded by linear increases in both the number of aberrant crypts and transepithelial permeability, as a function of weeks of DMH treatment. Adenocarcinomas of both rat and human colon have been found to have uniformly leaky tight junctions. Whereas most human colon hyperplastic and adenomatous polyps contain nonleaky tight junctions, adenomatous polyps with dysplastic changes did possess leaky tight junctions. Our overall hypothesis is that tight junctional leakiness is a late event in epithelial carcinogenesis but will allow for growth factors in luminal fluid compartments to enter the intercellular and interstitial fluid spaces for the first time, binding to receptors that are located on only the basal‐lateral cell surface, and causing changes in epithelial cell kinetics. Tight junctional leakiness is therefore a promotional event that would be unique to epithelial cancers.

1 The use of the cytoplasmic enzyme, alpha glutathione s-transferase (α-GST) as an early index of carbon tetrachloride (CCl4) toxicity in the rat was investigated and compared with a standard enzyme marker, aspartate aminotransferase (AST). The hepatotoxic effects of CCl 4 in the rat were determined in a time and dose-response study. 2 Following CCl 4 exposure, α-GST release was shown to be an earlier and more sensitive biomarker of hepatotoxicity than AST. 3 Significant increases in α-GST were detected 2 h after CCl4 exposure. Using the enzyme marker AST, this early hepatotoxic injury went undetected. At 6 and 16 h, α-GST was also a more sensitive indicator of hepatotoxicity than AST. 4 α-GST release was significantly increased at a dose of 5 μl/kg, the lowest concentration of CCl4 administered and clearly responded in a dose-dependent manner with increasing doses of CCl4. In contrast, release of AST did not reach statistical significance until a dose of 25 μl/kg. 5 Thus, these findings indicate that α-GST is a more sensitive and more accurate reflector of CCl4 induced hepatotoxicity than AST.

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