Changes in protein expression during multistage mouse skin carcinogenesis.

To directly compare the expression patterns of different proteins known to be altered during mouse skin carcinogenesis, serial sections of normal and hyperplastic skin and tumors from various stages of 7,12-dimethylbenz[a]anthracene-initiated, 12-O-tetradecanoylphorbol-13-acetate-promoted female SENCAR mice were examined by immunohistochemistry. In untreated, normal mouse skin, keratin 1 (K1) and transforming growth factor-beta1 (TGFbeta1) were strongly expressed in the suprabasal layers, whereas integrin alpha6beta4 was expressed only in basal cells and only moderate staining for transforming growth factor-alpha (TGFalpha) was seen. In hyperplastic skin, TGFalpha expression became stronger, whereas expression of another epidermal growth factor (EGF) receptor ligand, heparin-binding EGF-like growth factor (HB-EGF), was strongly induced in all epidermal layers from no expression in normal skin. Likewise, the gap-junctional protein connexin 26 (Cx26) became highly expressed in the differentiated granular layers of hyperplastic skin relative to undetectable expression in normal skin. Expression of cyclin D1 in the proliferative cell compartment was seen in all benign and malignant tumors but not in hyperplastic skin. Beginning with very early papillomas (after 10 wk of promotion), expression of alpha6beta4 in suprabasal cells and small, focal staining for keratin 13 (K13) were seen in some tumors. Later (after 20-30 wk), focal areas of gamma-glutamyl transpeptidase (GGT) activity appeared in a few papillomas, whereas TGFbeta1 expression began to decrease. Cx26 and TGFalpha staining became patchier in some late-stage papillomas (30-40 wk), whereas suprabasal alpha6beta4, K13, and GGT expression progressively increased and K1 expression decreased. Finally, in squamous cell carcinomas (SCCs), there was an almost complete loss of K1 and a further decline in TGFalpha, HB-EGF, TGFbeta1, and Cx26 expression. On the other hand, almost all SCCs showed suprabasal staining for alpha6beta4 and widespread cyclin D1 and K13 expression, whereas only about half showed positive focal staining for GGT activity.

Mechanism of phthalate-induced inhibition of hepatic mitochondrial beta-oxidation.

Studies show that peroxisome proliferators inhibit mitochondrial beta-oxidation of fatty acids. However, mechanism(s) of this inhibitory effect has not been identified. This study was undertaken to delineate such mechanism(s). Ketogenesis was significantly diminished in perfused livers from rats pre-treated with diethylhexyl phthalate (DEHP) compared with livers from control rats. Monethylhexyl phthalate (MEHP; 200 microM), a primary metabolite of DEHP and a known peroxisome proliferator, inhibited the oxidation of palmitic acid as well as its acyl-CoA and acylcarnitine derivatives in isolated mitochondria by about 50-60%. Similar concentrations of MEHP also inhibited mitochondrial respiration of succinate and malate plus glutamate. However, respiration of ascorbate was not influenced by MEHP. Considering the assembly of the mitochondrial respiratory chain, these data indicate that phthalates inhibit fatty acid metabolism as a result of inhibiting the respiratory chain at the level of the cytochrome c reductase. This effect may represent an early step in the mechanism by which phthalates cause hepatic peroxisome proliferation.

Skin carcinogenesis: characteristics, mechanisms, and prevention.

Skin carcinogenesis can be operationally and mechanistically divided into at least three major stages - initiation, promotion and progression. Variations among stocks and strains of mice to susceptibility to multistage skin carcinogenesis appear to be more related to alterations in tumor promotion than tumor initiation; however, the critical events have not been determined. In the mouse skin model the first stage is thought to involve the interaction of a tumor initiator with the genetic material of stem cells leading to an irreversible alteration in some aspect of growth control and/or differentiation, probably activating the Ha-ras oncogene. Some skin tumor promoters such as the phorbol esters, indole alkaloids, and polyacetates, appear to act through protein kinase C leading to specific phosphorylation of cellular proteins whereas others such as okadaic acid class of compounds appear to act through phosphatases also leading to an increase in phosphorylation. In addition, other types of tumor promoters such as peroxides, benzo(e)pyrene, and chrysarobin may act through a free radical mechanism. Regardless of the type, the major effect of the skin tumor promoters appears to be the specific expansion of the initiated stem cells in the skin. There is a very good correlation between the abilities of tumor promoters to induce a sustained hyperplasia and their tumor promoting activities. This appears to occur by both direct and indirect mechanisms involving the loss of glucocorticoid receptors, differentiation alterations, a direct growth stimulation of the initiated cells and/or selective cytotoxicity. A number of growth factors have recently been found to be increased during tumor promotion and may be responsible for the increase in cell proliferation. An inhibition of cell-cell communication and stimulation of differentiation of non-initiated cells appear to be important indirect mechanisms of further expanding the initiated cell population. The appearance of GGT and keratin 13 (K13) and the lack of expression of K1 and K10 were found to be good markers for skin tumor progression. These alterations occur at the time papillomas change from a diploid to aneuploid state which is mainly due to trisomies of chromosome 6 and 7. In order to evaluate a casual role for GGT in skin tumor progression, a functional GGT cDNA was transfected into two of our cell lines which normally produce papillomas when grafted into the skin of nude mice. The GGT positive cells and the vector transfected cells (controls) from one of the cell lines were cloned and injected into nude mice and placed into transplantation chambers.(ABSTRACT TRUNCATED AT 400 WORDS)