INHIBITION OF GAP JUNCTIONAL INTERCELLULAR COMMUNICATION IN HEPTACHLOR-TREATED AND HEPTACHLOR-EPOXIDE-TREATED NORMAL HUMAN BREAST EPITHELIAL-CELLS

Based on the concern of organochlorides in the environment and in huma n tissue, this study was designed to determine whether various noncyto toxic levels of heptachlor and heptachlor epoxide could inhibit, rever sibly, gap junctional intercellular communication in human breast epit helial cells (HBEC). Cytotoxicity and gap junctional intercellular com munication (GJIC) were evaluated by lactate dehydrogenase assay and fl uorescence redistribution after photobleaching analysis, respectively. Both heptachlor and heptachlor epoxide were noncytotoxic up to 10 mu g/ml. At this concentration, heptachlor and heptachlor epoxide inhibit ed GJIC of normal human breast epithelial cells after 1 h treatment. W ithin a 24 h treatment with heptachlor and heptachlor epoxide at 10 mu g/ml, recovery of GJIC had not returned. GJIC completely recovered af ter a 12 h treatment of 1 mu g/ml heptachlor epoxide, but it did not r ecover after a 24 h treatment of 1 mu g/ml heptachlor. RT-PCR and West ern blots were analyzed to determine whether the heptachlor or heptach lor epoxide might have altered the steady-state levels of gap junction mRNA and/or connexin protein levels or phosphorylation state. No sign ificant difference in the level of connexin 43 (Cx43) message between control and heptachlor-treated cells was observed. Western blot analys es showed hypophosphorylation patterns in cells treated with 10 mu g/m l heptachlor and heptachlor epoxide for 1 h with no recovery within 24 h. Immunostaining of Cx43 protein in normal HBEC indicated that hepta chlor and heptachlor epoxide caused a loss of Cx43 from the cell membr anes at noncytotoxic dose levels. Taken together, these results sugges t that heptachlor and heptachlor epoxide can alter GJIC at the post-tr anslational level, and that, under the conditions of exceeding a thres hold concentration in the breast tissue containing 'initiated' cells f or a long time and not being counteracted by anti-tumor-promoting chem icals, they could act as breast tumor promoters.