Biomedical subjects

M J Fernstrom

Publications and source records attributed to M J Fernstrom.

6 recordsLinked to original sources

PubMed2000

Liver cell-specific transcriptional regulation of connexin32.

Gap junctional intercellular communication facilitates liver homeostasis and growth control in the liver. The major gap junction protein expressed by hepatocytes is connexin32 (Cx32) and non-parenchymal hepatic cells do not express this gene. We investigated the regulation of Cx32 transcription by trans-activating factors in liver cells. Transient transfection assays using deletions of the rat Cx32 promoter (nt -753 to -33) linked to the luciferase gene were performed in MH1C1 rat hepatoma cells that express endogenous Cx32 compared with WB-F344 rat liver epithelial cells that do not. The basal promoter element was located within nt -134 to -33 and was 1.4-fold more active in MH1C1 cells than WB-F344 cells whereas the entire promoter fragment (nt -754 to -33) was four-fold more active in MH1C1 cells. Specific nuclear protein-DNA complexes that bound to Sp1 consensus sites within the basal promoter were formed using nuclear extracts from both types of cells. Additional promoter sequences increased promoter activity more strongly in MH1C1 cells than WB-F344 cells and this was correlated with the binding of hepatocyte nuclear factor-1 (HNF-1) to two HNF-1 consensus sites centered at -187 and -736. Expression of HNF-1 and binding to these elements was only observed with MH1C1 cells. Other specific protein-DNA complexes were formed, however, that included YY-1- and NF-1-containing complexes, but these were not related to promoter activity. Dexamethasone increased Cx32 promoter activity and expression in MH1C1 cells, but had little effect in WB-F344 cells and did not alter protein-DNA complex formation. These data suggest that Sp1 is responsible for Cx32 promoter basal activity, that HNF-1 determines the cell-specific expression of Cx32, and that dexamethasone increases Cx32 expression through other mechanisms.

Animals↗

PubMed1999

Gap junctional intercellular communication and connexin43 expression in human ovarian surface epithelial cells and ovarian carcinomas in vivo and in vitro.

Gap junctional intercellular communication (GJIC) and the expression of gap junction proteins (connexins) are frequently decreased in neoplastic cells and have been increased by cAMP and retinoids. GJIC and connexin expression were investigated in early passage normal human ovarian surface epithelial (HOSE) cells, human ovarian adenocarcinoma cell lines (CaOV-3, NIH:OVCAR-3, SK-OV-3 and SW626) and surgical specimens of human serous cystadenocarcinomas. We hypothesized that GJIC and connexin expression would be decreased in neoplastic cells and would be increased by cAMP and retinoic acid. Cultured HOSE cells exhibited extensive fluorescent dye-coupling and connexin43 (Cx43) expression; other connexins were not detected. The ovarian adenocarcinoma cell lines had little dye-coupling or connexin expression. Deletions and rearrangements of the Cx43 gene were not detected by Southern blotting in the carcinoma lines. N6, 2'-O-dibutyryladenosine 3',5'-cyclic monophosphate and all-trans-retinoic acid inhibited cell proliferation, but did not enhance GJIC or Cx43 expression. Surface epithelial cells of benign ovaries expressed Cx43, but this expression was barely detectable in ovarian serous cystadenocarcinomas. Thus, normal HOSE cells had extensive GJIC and Cx43 expression whereas ovarian carcinoma cells had less and cAMP and retinoic acid did not change these, although both agents inhibited cell growth.

Adenocarcinoma↗

PubMed1998

Frequent reduction of gap junctional intercellular communication and connexin43 expression in human and mouse lung carcinoma cells.

The reduced gap junctional intercellular communication (GJIC) and gap junction protein (connexin) expression that have been noted in many neoplastic cell types may contribute to the neoplastic phenotype. We assessed GJIC (by fluorescent dye micro-injection) and connexin expression (by Northern blotting, Western blotting and immunohistochemistry) in five mouse and 17 human lung carcinoma cell lines; both measures were lower in neoplastic cells compared to non-transformed lung epithelial cells. Other connexins were not detected in these cells. Co-culture experiments indicated that carcinoma cell lines able to transfer dye among themselves (homologous GJIC) had little capacity for dye-coupling with non-transformed cells (heterologous GJIC). Southern blot analyses indicated that reductions in GJIC and connexin43 expression were not due to deletions or rearrangements of this gene, but were more likely accounted for by transcriptional down-regulation and/or post-transcriptional factors. No correlations between GJIC and known oncogene and tumor suppressor gene alterations in the human lung carcinoma cells were apparent, suggesting that other mechanisms down-regulate GJIC in these cells. Since the neoplastic cell lines exhibited low GJIC (either homologous or heterologous), this characteristic may be involved in expression of the neoplastic phenotype.

Adenocarcinoma↗

PubMed1997

Altered gap junctional intercellular communication in neoplastic rat esophageal epithelial cells.

Gap junctional intercellular communication (GJIC) is reduced in many neoplastic cells, but few data exist for esophageal neoplasms. GJIC was examined by fluorescent dye microinjection in two nontumorigenic and two highly tumorigenic rat esophageal epithelial cell lines. All lines expressed high levels of dye coupling in homologous cell culture. In cocultures of nontumorigenic and tumorigenic cells, however, only one of six cell combinations displayed significant heterologous GJIC. Northern, Western, and immunohistochemical analyses indicated that all four cell lines expressed comparable levels of connexin43 (Cx43), but not connexin32 or connexin26, and formed Cx43-containing gap junction plaques at cell-cell interfaces. Immunostaining of rat esophageal frozen sections demonstrated that esophageal epithelial cells expressed Cx43 in vivo. In normal epithelium, the highest expression was seen in the basal cells and little suprabasal staining was evident. In preneoplastic and neoplastic lesions of the esophageal epithelium which were induced by treating rats with N-nitrosomethylbenzylamine, Cx43 staining of the basal layer was also seen but appeared to be more diffuse compared to normal epithelium. In addition, suprabasal Cx43 staining was apparent in dysplastic and papillomatous lesions. These results indicate that Cx43 is expressed in normal and neoplastic rat esophageal cells and that the cells exhibit extensive homologous GJIC, but little heterologous GJIC. This lack of heterologous GJIC may be due to differences in cell adhesion proteins or other factors.

Animals↗

PubMed1995

Down-regulation by butylated hydroxytoluene of the number and function of gap junctions in epithelial cell lines derived from mouse lung and rat liver.

The mouse pneumotoxicant and lung and liver tumor promoter butylated hydroxytoluene (BHT) was examined for its effects on gap junctional intercellular communication (GJIC) in mouse lung epithelial (C10) and rat liver epithelial (WB-F344) cell lines. GJIC, as measured by fluorescent dye microinjection, was inhibited in both types of cells by BHT in dose- and time-dependent fashions. Inhibition was detected in WB-F344 cells at BHT concentrations > or = 62.5 microM and in C10 cells at concentrations > or = 150 microM after 4 h treatment. Inhibition occurred within 15-30 min and was reversed by removing BHT from the culture medium. The highly toxic BHT metabolite 6-t-butyl-2-(hydroxy-t-butyl)-4-methylphenol (BHTOH) and the non-toxic BHT metabolite, 2,6-di-t-butyl-4-hydroxymethylphenol (BHTBzOH) were also tested. In both cell lines BHTOH was a more potent inhibitor of GJIC than BHT, whereas BHTBzOH was ineffective. The mechanisms of inhibition of GJIC by BHT were also examined. The initial rapid inhibition detected within 15-30 min may have been due to gap junction channel closure or blockage, since no changes in gap junction number, connexin (Cx) 43 levels or Cx43 phosphorylation were observed. By 2-4 h, however, gap junctions were internalized into the cytoplasm, the number of immunodetectable plasma membrane gap junctions was reduced and phosphorylated Cx43-P2 was decreased. Treatment of the cells for 24 h with 12-O-tetradecanoylphorbol-13-acetate (TPA) prevented inhibition of GJIC by TPA, but not by BHT. Western blot analyses of TPA-treated WB-F344 or C10 cells revealed the presence of a hyperphosphorylated form of Cx43 (Cx43-P3) and no reduction in Cx43-P2, in contrast to BHT-treated cells. These data suggest that BHT and TPA inhibit lung and liver epithelial cell GJIC through distinct mechanisms.

Animals↗

PubMed1979

Oxytocin receptors: triggers for parturition and lactation?

Specific binding of tritiated oxytocin to uterine receptors of pregnant rats increases dramatically at term and is maximal during labor. In mammary glands the increase in binding is gradual, reaching a maximum during the lactation period. Concomitant changes in the sensitivity of the uterus and mammary gland to oxytocin indicate that the receptor concentration is of functional significance. Oxytocin receptors, therefore, may regulate the response of the target organs to circulating oxytocin and thereby control the onset of labor and lactation. Ovarian steroids participate in the regulation of oxytocin receptors in a manner as yet unclarified.

Animals↗