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Gap junction intercellular communication capacity and connexin expression are reportedly decreased in human lung cancer. The mechanisms by which connexins, the gap junction proteins, act as tumor suppressors are unclear. In order to understand the involvement of connexins in tumorigenesis, we analyzed the effect of the heterologous deletion of Gja1 [the connexin43 (Cx43) gene] on the development of lung adenomas in mice. Heterozygous (Cx43(+/-)) and wild-type mice (Cx43(+/+)) were treated or not with single doses of urethane at 15 and 17 days after birth. Twenty-five weeks later, both the number and size of nodules were increased in Cx43(+/-) mice as compared with Cx43(+/+) mice. Moreover, the lesions were histologically more aggressive in the heterozygous mice. However, no increase in spontaneous lesions was observed in the lungs of untreated Cx43(+/-) mice. Heterozygous mice effectively presented lower expression of Cx43 genes and decreased amounts of Cx43. In conclusion, our results indicate that deletion of one allele of the Cx43 gene clearly favors the carcinogenic effect of urethane administration and results in a higher susceptibility to lung adenoma formation in mice.

Polychlorinated biphenyls (PCBs) are industrial chemicals which are highly persistent and widely distributed in the environment. We have previously shown that 3,4,5,3',4'-pentachlorobiphenyl (PCB 126) is a potent tumour promoter in two separate 20 week initiation-promotion studies. In the present study, rat livers from these two studies were further investigated for connexin expression. The results demonstrated that treatment with PCB 126 caused a decrease in the amount of the two major liver connexins, cx 26 and cx 32, in livers of treated animals. This reduction was also prominent after treatment at low doses, although gamma-glutamyl transpeptidase-positive foci had not developed in these livers. The quantity of cx 26 and cx 32 in immunostained liver sections was determined using a computerized fluorescence image analyzer. Western blot analysis of liver extracts confirmed these results. No changes in the RNA levels in the treated rats were seen, suggesting that the down-regulation of cx 26 and cx 32 is post-transcriptional.

The gap junctional communication capacity of six human carcinoma-derived tumorigenic cell lines (pancreatic, pharyngeal and cervical), two murine carcinoma-derived tumorigenic cell lines (bladder-derived and Ehrlich ascites) and one monkey non-tumorigenic cell line (kidney epithelium) have been compared by the dye-transfer technique. All the tumorigenic cell lines were communication defective, while the non-tumorigenic cell line was not. Moreover, six of the eight tumorigenic cell lines expressed a gap junction transcript coding for the connexin 43 (alpha 1). Finally, gap junction plaques were not detected between tumorigenic cells by immunofluorescence staining. Consequently, these data suggest that communication defects in tumorigenic cells may result from either abnormally low levels of translation of junction mRNA or alterations in the assembly of junction protein into cell surface plaques, and not from failure to produce junction transcripts. Furthermore, since induction of alpha 1 expression has been associated with dedifferentiation processes, the presence of alpha 1 mRNA might be a characteristic property of certain tumorigenic cells that originate from cells that do not normally express alpha 1 mRNA.

Transforming growth factor beta 1 (TGF-beta 1) enhances the yield of transformed foci of BALB/c 3T3 cells, but the continuous presence of TGF-beta 1 after foci formation inhibits the growth of transformed foci. The focus-forming ability of Ha-ras-, v-src- and PyMT-transformed cells growing on a monolayer of non-transformed cells was completely suppressed by TGF-beta 1, whereas growth of the transformed cells was little inhibited by TGF-beta 1 in the absence of their normal counterparts. The inhibition by TGF-beta 1 of focus formation by transformed BALB/c 3T3 cells on a normal cell monolayer remained when TGF-beta 1 was removed from the culture medium after 2 weeks. However, the transformed cells were not killed, since they grew in culture conditions under which only transformed cells are able to grow (soft agar). These results suggest that TGF-beta 1 suppresses growth of transformed cells in the presence of normal cells. Furthermore, when non-transformed cells were treated with TGF-beta 1 before co-culture with Ha-ras-transformed cells, formation of transformed foci was inhibited. When normal and transformed cells were cultured in the same dish but separated physically, focus formation was still inhibited. On the other hand, TGF-beta 1 enhanced the growth and changed the morphology of non-transformed cells only in the presence of transformed counterparts. The growth inhibitory effect of TGF-beta 1 on transformed cells and its growth stimulatory effect on non-transformed cells in co-culture conditions suggest the induction of reciprocal paracrine growth regulatory factors. As TGF-beta 1 inhibits the growth of transformed BALB/c 3T3 cells only in the presence of their normal counterparts, a paracrine negative growth control mechanism appears to be operating.

To investigate whether a selective intercellular communication exists between transformed epithelial cells and their counterparts, the homologous and heterologous communication capacities of four rat liver epithelial cell lines were compared with their expression of transformed phenotypes. All four cell lines showed reasonably good homologous junctional communication capacity, as measured by the dye-transfer assay. In heterologous co-cultures, the non-transformed cell line IAR 20 did not communicate with the transformed cell lines IAR 6-1 or IAR 27 F. These two cell lines showed a high degree of transformed phenotypes such as cell morphology, growth in soft agar and expression of gamma-glutamyltranspeptidase activity. Another cell line IAR 27 E, showed the least degree of transformation and it communicated with IAR 20 cells. Thus, it appears that there is an inverse correlation between the extent of expression of transformed phenotypes by rat liver epithelial cells and their ability to communicate with non-transformed counterparts. There was no heterologous intercellular communication between any combination of IAR 27 E, IAR 27 F and IAR 6-1 cell lines.