lucifer-yellow and Liver-Neoplasms

One of the systems that regulate tissue homeostasis is gap junction intercellular communication (GJIC). It is accepted that the down-regulation of GJIC is linked to the tumor-promoting properties of carcinogens. In this study, the effect of some carcinogenic and non-carcinogenic polycyclic aromatic hydrocarbons (PAH) on GJIC was investigated. It was found that in hepatoma cell culture (Hep G2) carcinogenic PAH inhibited GJIC after 24 h exposure by 75-100% depending on the PAH structure. The inhibition effect on GJIC is reversible because removing the PAH by changing of culture medium restores the GJIC. The non-carcinogenic PAH do not significantly influence GJIC. alpha-Naphthoflavone, an inhibitor of PAH metabolism, has no effect on inhibition of GJIC by carcinogenic PAH. 2,3,7,8-Tetrachloro-p-dibenzodioxin, an aryl hydrocarbon (Ah) receptor ligand, inhibits GJIC by about 50% only after 48 h exposure. To clarify the role of formation of PAH metabolites and interaction with Ah receptor on inhibition of GJIC, we determined the effect of benzo/a/pyrene on hepatoma G27 cells in which neither mRNA of CYP1A1 nor Ah receptor was determined. As in Hep G2 cells, benzo/a/pyrene, unlike non-carcinogenic benzo/e/pyrene, inhibits GJIC. We conclude that in the studied hepatoma cells carcinogenic PAH inhibit GJIC directly (that is, not via their metabolites) and this effect is not associated with Ah receptor interaction.

Topics: Animals; Apoptosis; Carcinogens; Carcinoma, Hepatocellular; Cell Communication; Cell Proliferation; Fluorescent Dyes; Gap Junctions; Humans; Isoquinolines; Liver Neoplasms; Molecular Structure; Permeability; Polycyclic Aromatic Hydrocarbons; Rats; Receptors, Aryl Hydrocarbon; Tumor Cells, Cultured

To investigate gap junctional intercellular communication (GJIC) in hepatocellular carcinoma cell lines, and signal transduction mechanism of gap junction genes connexin32(cx32),connexin43(cx43) in human hepatocarcinogenesis.. Scarped loading and dye transfer (SLDT) was employed with Lucifer Yellow (LY) to detect GJIC function in hepatocellular carcinoma cell lines HHCC, SMMC-7721 and normal control liver cell line QZG. After Fluo-3AM loading, laser scanning confocal microscope (LSCM) was used to measure concentrations of intracellular calcium (Ca(2+))i in the cells. The phosphorylation on tyrosine of connexin proteins was examined by immunoblot.. SLDT showed that ability of GJIC function was higher in QZG cell than that in HHCC and SMMC-7721 cell lines. By laser scanning confocal microscopy, concentrations of intracellular free calcium (Ca(2+))i was much higher in QZG cell line (108.37 nmol/L) than those in HHCC (35.13 nmol/L) and SMMC-7721 (47.08 nmol/L) cells. Western blot suggested that only QZG cells had unphosphorylated tyrosine in Cx32 protein of 32 ku and Cx43 protein of 43 ku; SMMC-7721 cells showed phosphorylated tyrosine Cx43 protein.. The results indicated that carcinogenesis and development of human hepatocellular carcinoma related with the abnormal expression of cx genes and disorder of its signal transduction pathway, such as decrease of (Ca(2+))i, post-translation phosphorylation on tyrosine of Cx proteins which led to a dramatic disruption of GJIC.

Topics: Calcium Signaling; Carcinoma, Hepatocellular; Cell Communication; Cell Line; Connexin 43; Connexins; Fluorescent Dyes; Gap Junction beta-1 Protein; Gap Junctions; Gene Expression; Hepatocytes; Humans; Isoquinolines; Liver Neoplasms; Phosphorylation; Signal Transduction; Tumor Cells, Cultured; Tyrosine

Gap junction channels provide a pathway for exchange of ions and small molecules between coupled cells, and this exchange is believed to be critical for normal tissue growth and development. As a test for a role of gap junction-mediated intercellular communication in control of cell growth, we have compared growth rates of communication-deficient human tumor cells (SKHep1) with clones stably transfected with cDNA encoding the rat liver gap junction protein connexin 32. In culture, growth rates for parental and transfected clones were similar. However, when sizes of tumors were evaluated following injection of these clones into athymic nude mice, growth rates for two well-coupled clones were significantly lower than for communication-deficient or poorly coupled clones. This study demonstrates that growth rate of these tumor cells in situ is negatively correlated with strength of intercellular communication.

Topics: Animals; Carcinoma, Hepatocellular; Cell Division; Connexins; DNA; Fluorescent Antibody Technique; Fluorescent Dyes; Humans; Intercellular Junctions; Isoquinolines; Kinetics; Liver Neoplasms; Membrane Proteins; Mice; Mice, Nude; Neoplasm Transplantation; Plasmids; Transfection; Transplantation, Heterologous; Tumor Cells, Cultured