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PubMed1998

Effect of cytokines and nitric oxide on tight junctions in cultured rat retinal pigment epithelium.

PURPOSE: These experiments were designed to study the effect of cytokines and nitric oxide (NO) on rat retinal pigment epithelial (RPE) cell tight junctions in vitro. METHODS: Cultures of confluent RPE cells from retinas of PVG rats (a strain susceptible to development of experimental uveitis) were prepared on filters and incubated with various stimulants. The function of the tight junction was evaluated by measuring the transepithelial electrical resistance (TER) of the cell monolayer and the passive permeation of [3H]inulin across confluent RPE cells. The morphology of the intercellular junctions was visualized by immunolocalization of the tight junction-associated protein zonula occludens-1 (ZO-1) and F-actin. RESULTS: Seventy-two hours after plating, the RPE cell monolayer showed a mean TER level of 67.6+/-18.8 omega/cm2. A decrease in TER was observed after treatment with interferon-gamma (IFN-gamma) and lipopolysaccharide (LPS). The addition of tumor necrosis factor-alpha (TNF-alpha) accelerated the decrease of TER, whereas NG-monomethyl-L-arginine (L-NMMA) (an NO synthase [NOS] inhibitor) did not further modify the resistance decrease. In contrast, 3-morpholino-sydnonimine (SIN-1), a sydnonimine analog and NO donor, increased the TER. The variations of TER were correlated with the transepithelial fluxes of [3H]inulin and with tight junction morphologic changes of ZO-1 and F-actin immunostaining. CONCLUSIONS: Incubation with LPS associated with IFN-gamma and TNF-alpha induces alterations of RPE tight junctions, whereas NO is involved in the maintenance of their integrity. Cytokines and NO production could play a role in regulation of the blood-ocular barrier function and of the development of ocular inflammation.

Actins↗
PubMed1997

Effects of peroxisome proliferators and 12-O-tetradecanoyl phorbol-13-acetate on intercellular communication and connexin43 in two hamster fibroblast systems.

Effects of 12-O-tetradecanoyl phorbol 13-acetate (TPA) and the hepatic peroxisome proliferators (HPPs) clofibrate, methyl clofenapate (MCP), di(2-ethylhexyl)phthalate (DEHP) and mono(2-ethylhexyl)phthalate (MEHP) were studied in 2 gap junctional intercellular communication (GJIC) systems, metabolic cooperation in V79 cells and microinjection/dye transfer in Syrian hamster embryo (SHE) cells and V79 cells. TPA inhibited GJIC in both systems but was considerably more potent in V79 cells. SHE cells showed a rapid and transient inhibition of GJIC after exposure to HPPs, with maximal inhibition occurring at 5-15 min. The transient inhibition could be caused by metabolization of the compounds. Clofibrate and MEHP produced strong inhibition of metabolic cooperation in V79 cells at high concentrations, while the effect of MCP and DEHP was lower. However, DEHP, MEHP and clofibrate strongly inhibited dye transfer in V79 cells after a 30 min exposure. Clofibrate also showed a dose- and time-dependent effect on dye transfer in V79 cells. The phosphorylation status of the gap junction protein connexin43 (Cx43) changed minimally in SHE cells after exposure to TPA or HPPs. Cx43 from V79 cells was strongly affected by TPA, but not by HPPs. Immunofluorescence of Cx43 disappeared in both cell types when they were exposed to TPA and MEHP, but not to the other HPPs. Thus, there is no direct correlation between the inhibition of GJIC and changes in the phosphorylation status of Cx43 or the appearance of Cx43 in immunofluorescence experiments. The discrepancies may partly be explained by binding of accessory proteins to Cx43. We point out sequences that may be involved in such binding.

Animals↗
PubMed1983

Freeze-fracture studies of neonatal mouse incisors.

Differentiation of preodontoblasts to odontoblasts and preameloblasts to ameloblasts during development of the mouse mandibular incisor proceeds in a gradient from the area of the odontogenic organ, where undifferentiated ectomesenchymal and epithelial cells proliferate, toward the incisal tip where mature tooth tissues, dentin and enamel, are present. The freeze-fracture technique has been used in the work presented here to study cell membrane ultrastructure of preodontoblasts and preameloblasts at several stages of differentiation. At early stages of differentiation, cuboidal preameloblasts are joined together distally by numerous gap junctions. Relatively fewer junctions occur elsewhere on the lateral plasma membranes, but gap junctions frequently occur proximally between preameloblasts and stratum intermedium cells. As differentiation proceeds and the cells become columnar, distal and proximal junctions persist. Tight junctions, however, were not observed at any of the stages studied. Intramembrane particle concentration of the lateral preameloblast plasmalemma appears to increase as differentiation proceeds. Odontoblasts are also joined distally by numerous gap junctions which persists through later stages of differentiation. Although odontoblast cell processes were observed to project toward the preameloblast layer, no clear points of cell to cell contact or defined intercellular junctions between the two cell types were observed.

Ameloblasts↗
PubMed1982

Disruption of iridial blood-aqueous barrier in experimental diabetic rats.

The permeability pattern of iridial blood-aqueous barrier in streptozotocin-diabetic rats, maintained for 1-6 months, was studied with the light and electron microscopes, using horseradish peroxidase as a tracer. A reaction product of horseradish peroxidase was confined to the vascular lumina and to a small number of vesicles on the luminal portion of the endothelial cells in control rats and diabetic rats maintained for 1 month. In diabetic rats maintained for 2-6 months, endothelial cells contained a large number of horseradish peroxidase-labeled vesicles which opened into the basement membrane, and intercellular junctions of the endothelial cells were filled with horseradish peroxidase. Horseradish peroxidase was consistently found in the stroma around the vessels, and seemed to reach the anterior chamber through the intercellular spaces between the fibroblasts. However, the passage of horseradish peroxidase was blocked by the posterior epithelial cells.

Animals↗
PubMed1998

Specific localization of gap junction protein, connexin45, in the deep muscular plexus of dog and rat small intestine.

Cellular networks of pacemaker activity in intestinal movements are still a matter of debate. Because gap-junctional intercellular communication in the intestinal wall may provide important clues for understanding regulatory mechanisms of intestinal movements, we have attempted to clarify the distribution patterns of three types of gap junction proteins. Using antibodies for connexin40, connexin43, connexin45, smooth muscle actin, and vimentin, immunocytochemical observations were made with the confocal laser scanning microscope on cryosections of fresh-frozen small intestine and colon of the dog and rat. Connexin 45 was localized along the deep muscular plexus of the small intestine in both dog and rat. Double labeling studies revealed that connexin45 overlapped with vimentin -, but not actin-positive areas, indicating the fibroblast-like nature of the cells, rather than their being smooth muscle-like. Connexin43 immunoreactivity appeared along the smooth muscle cell surface in the outer circular layer of the small intestine of both animals. Connexin 40 immunoreactivity was not observed in the muscle layer other than in the wall of large blood vessels. It is suggested that connexin45-expressing cells along the deep muscular plexus of dog and rat small intestine are likely to act as a constituent of a pacemaker system, which may include a conductive system, by forming a cellular network operating via specific types of gap junctions.

Animals↗
PubMed2005

The spread of apoptosis through gap-junctional channels in BHK cells transfected with Cx32.

Programmed cell death (apoptosis) occurs both during normal development and as a result of various pathological conditions. An in vitro system was used to explore the transmission of death signals from apoptotic cells to cells with which they were coupled via gap junctions. Confluent cultures of baby hamster kidney (BHK) cells, stably transfected with the gap-junctional protein connexin32, were scrape loaded with cytochrome C (cyC), a mitochondria-derived apoptotic agent, to introduce the protein into cells injured by the cut. The cultures were subsequently analyzed for the presence of activated caspases, the distribution of TUNEL staining, and the binding of annexin V. Although cyC is too large to traverse the gap junctional channel, each of the assays revealed that apoptosis had spread from dying cells at the margin of the scrape to otherwise healthy neighboring cells to which they were coupled. This "bystander effect" was significantly reduced in the presence of agents that block gap junctional intercellular communication.

Animals↗
PubMedDate not supplied

Role of gap junctions in lung neoplasia.

Reduced gap junctional intercellular communication (GJIC) has been noted in many types of neoplastic cells and may contribute to the neoplastic phenotype. This study assessed GJIC (by fluorescent dye-coupling) and gap junction protein (connexin) expression in mouse and human lung carcinoma cell lines and investigated whether reduced GJIC was involved in their neoplastic phenotype. Dye-coupling and connexin43 (Cx43) expression were much lower in most of the carcinoma lines (16 of 22) compared to nontransformed lung epithelial cells. Other connexins were not detected. A poorly communicating mouse lung carcinoma cell line (E9) was transfected with Cx43 or transduced with Cx32 and several stable clones were isolated that had 2- to 4-fold increased dye coupling. When evaluated for growth in vitro, the population doubling times were increased and the saturation densities were decreased in the clones. When assessed for tumorigenicity, the parental E9 cells formed tumors with a 100% incidence (6/6 mice), whereas the clones varied in tumorigenic response (0-88% incidence). The best communicating clone (E9-2) was not tumorigenic. The highly communicating Cx32 clone, E9/32-9, gave a tumor incidence of 88%. These results suggest that restoration of GJIC by forced connexin expression can reduce the growth and tumorigenicity of lung carcinoma cells in a connexin-specific manner.

Adenocarcinoma↗
PubMed2006

Altered morphology and distribution of cellular junction proteins in non-lesional psoriatic epidermis: an insight into disease severity.

BACKGROUND: Psoriasis affects 2.7% of the world's population. Keratinocyte proliferation outside the basal layer suggests alterations in cell-cell interactions in affected epidermis. Anomalous expression of proteins forming intercellular junctions has been reported in lesional skin of psoriatic patients. In contrast, little is known about possible alterations in psoriatic non-lesional skin. METHODS: Ten clinically diagnosed psoriasis vulgaris patients and ten controls were studied. All patients were diagnosed with active but controlled psoriatic plates (PASI 3 to 5) and had not received any systemic treatment. The mean age was 43 years for patients and 43.5 years for controls. Four-mm2 skin samples were taken from lesional and non-lesional zones in patients and from abdomen in controls. Five-mum sections were examined for integrity and structural organization by fluorescent labeling of actin filaments and nuclei. Specific antibodies were utilized to localize occludin, E-cadherin, beta-catenin, and proliferation-specific keratins in sections and epidermal sheets. Samples were also processed for immunoblotting with occludin antibody. RESULTS: Lesional and non-lesional psoriatic epidermis from all patients showed keratinocyte hyperproliferation, lack of rete ridges and dermal papillae in the dermal-epidermal junction in some areas. Proteins forming tight and adherens junctions in non-lesional skin keratinocytes from two patients who during the course of the study evolved to uncontrolled disease, showed similar alterations to those observed in lesional skin of all the patients. However, the occludin isoforms expressed were apparently the same in all samples. CONCLUSIONS: Analysis of non-lesional skin in psoriatic patients diagnosed with controlled disease may provide clues about incipient structural abnormalities in the pathogenesis of psoriasis, providing an early diagnostic indicator for evolution to a generalized form of the disease.

Adherens Junctions↗
PubMed2003

The protein tyrosine phosphatase Pez is a major phosphatase of adherens junctions and dephosphorylates beta-catenin.

Cell-cell adhesion regulates processes important in embryonal development, normal physiology, and cancer progression. It is regulated by various mechanisms including tyrosine phosphorylation. We have previously shown that the protein tyrosine phosphatase Pez is concentrated at intercellular junctions in confluent, quiescent monolayers but is nuclear in cells lacking cell-cell contacts. We show here with an epithelial cell model that Pez localizes to the adherens junctions in confluent monolayers. A truncation mutant lacking the catalytic domain acts as a dominant negative mutant to upregulate tyrosine phosphorylation at adherens junctions. We identified beta-catenin, a component of adherens junctions, as a substrate of Pez by a "substrate trapping" approach and by in vitro dephosphorylation with recombinant Pez. Consistent with this, ectopic expression of the dominant negative mutant caused an increase in tyrosine phosphorylation of beta-catenin, demonstrating that Pez regulates the level of tyrosine phosphorylation of adherens junction proteins, including beta-catenin. Increased tyrosine phosphorylation of adherens junction proteins has been shown to decrease cell-cell adhesion, promoting cell migration as a result. Accordingly, the dominant negative Pez mutant enhanced cell motility in an in vitro "wound" assay. This suggests that Pez is also a regulator of cell motility, most likely through its action on cell-cell adhesion.

Adherens Junctions↗
PubMed1990

Effects on intercellular communication in human keratinocytes and liver-derived cells of polychlorinated biphenyl congeners with differing in vivo promotion activities.

Several purified polychlorinated biphenyl (PCB) congeners with differing toxicity/tumor promotional activities in rat liver in vivo were tested for their effects on gap-junctional intercellular communication (GJIC) in cell strains and lines derived from human liver and skin. This in vitro assay is being developed to detect various classes of tumor promoters. The 3-methylcholanthrene (MCA)-type cytochrome P450 inducer and hepatotoxic promoter 3,3',4,4'-tetrachlorobiphenyl was inactive in this assay for all of the cells tested, suggesting this promoter acts by other mechanisms. The phenobarbital-like enzyme inducer and less toxic promoter 2,2',4,4',5,5'-hexachlorobiphenyl inhibited GJIC in both liver and skin cells, whereas the 2,2',5,5'-tetrachlorobiphenyl congener, which does not act as a promoter in rat liver, inhibited GJIC only in the skin cell types and in one of the liver cell strains thought to be of bile duct origin. 2,3,4,4',5-Pentachlorobiphenyl, a mixed (phenobarbital plus MCA) inducer of cytochrome P450, inhibited GJIC in both liver and skin cells, suggesting that it may be a promoter in vivo. The results suggest that GJIC inhibition is a property of PCB congeners with phenobarbital-like enzyme induction capabilities, and that there exist some tissue/cell type differences in sensitivity to these congeners.

Carcinogens↗
PubMedDate not supplied

Role of connexin (gap junction) genes in cell growth control and carcinogenesis.

Gap junctional intercellular communication (GJIC) is considered to play a key role in the maintenance of tissue independence and homeostasis in multicellular organisms by controlling the growth of GJIC-connected cells. Gap junction channels are composed of connexin molecules and, so far, more than a dozen different connexin genes have been shown to be expressed in mammals. Reflecting the importance of GJIC in various physiological functions, deletion of different connexin genes from mice results in various disorders, including cancers, heart malformation or conduction abnormality, cataract, etc. The possible involvement of aberrant GJIC in abnormal cell growth and carcinogenesis has long been postulated and recent studies in our own and other laboratories have confirmed that expression and function of connexin genes play an important role in cell growth control. Thus, almost all malignant cells show altered homologous and/or heterologous GJIC and are often associated with aberrant expression or localization of connexins. Aberrant localization of connexins in some tumour cells is associated with lack of function of cell adhesion molecules, suggesting the importance of cell-cell recognition for GJIC. Transfection of connexin genes into tumorigenic cells restores normal cell growth, supporting the idea that connexins form a family of tumour-suppressor genes. Some studies also show that specific connexins may be necessary to control growth of specific cell types. We have produced various dominant-negative mutants of Cx26, Cx32 and Cx43 and showed that some of them prevent the growth control exerted by the corresponding wild-type genes. However, we have found that connexins 32, 37 and 43 genes are rarely mutated in tumours. In some of these studies, we noted that connexin expression per se, rather than GJIC level, is more closely related to growth control, suggesting that connexins may have a GJIC-independent function. We have recently created a transgenic mouse strain in which a mutant Cx32 is specifically overexpressed in the liver. Studies with such mice indicate that Cx32 plays a key role in liver regeneration after partial hepatectomy. A decade ago, we proposed a method to enhance killing of cancer cells by diffusion of therapeutic agents through GJIC. Recently, we and others have shown that GJIC is responsible for the bystander effect seen in HSV-tk/ganciclovir gene therapy. Thus, connexin genes can exert dual effects in tumour control: tumour suppression and a bystander effect for cancer therapy.

Animals↗
PubMed1998

Expression of gap junction proteins connexin 26 and connexin 43 in normal human breast and in breast tumours.

Gap junctional intercellular communication (GJIC) has been proposed as a cellular mechanism for tumour suppression and there is experimental evidence in support of this. If aberrant GJIC contributes to the formation of human breast tumours, one might expect that the connexins (gap junction proteins) expressed by epithelial cells in normal human breast would be down-regulated in tumour epithelial cells, or that tumour cells might show aberrant expression of other connexin family members. This study examines the immunocytochemical expression of connexins 26 (Cx26) and 43 (Cx43) in normal human breast, 11 benign breast lesions, two special-type carcinomas, and 27 invasive carcinomas of no special histological type (NST). Cx26 generally was not expressed at detectable level in normal human breast, but punctate Cx43 immunostaining of the myoepithelial cells was found. Cx43 staining of the myoepithelium was also a feature of the benign lesions and ductal carcinoma in situ (DCIS). In general, the epithelial cells of benign lesions failed to stain for either connexin. Similarly, a lobular carcinoma did not express Cx26 or Cx43, but there was punctate Cx43 in the epithelial cells of a mucoid carcinoma. Cx26 was up-regulated in the carcinoma cells of 15 of the 27 invasive NST carcinomas, although the staining was usually cytoplasmic and heterogeneous. Cx43 was expressed by stromal cells, possibly myofibroblasts, in all NST carcinomas. Furthermore, there was heterogeneous Cx43 expression in the carcinoma cells of 14 of the 27 NST carcinomas and the staining was often intercellular and punctate, characteristic of functional connexins. Up-regulated of Cx26 and/or Cx43 in the carcinoma cells of over two-thirds of invasive lesions of NST is not necessarily inconsistent with a tumour suppressor role for GJIC. However, the role of gap junctions in the formation and progression of solid human tumours is likely to be more complex than indicated from experimental systems.

Breast↗
PubMedDate not supplied

Connexin43 gene expression in the rabbit arterial wall: effects of hypercholesterolemia, balloon injury and their combination.

The specialized functions of endothelium require intercellular communication between endothelial cells within the monolayer, and between endothelium and other cells present in the vessel wall. This is accomplished by a combination of paracrine soluble mediators and direct gap-junctional intercellular communication (GJIC) mediated by a family of connexin proteins. A prominent connexin expressed by vascular cells in vivo and in vitro is connexin 43 (Cx43). We have investigated the in vivo gene regulation of Cx43 in the context of vascular pathology, as a result of mechanical injury, hypercholesterolemia or both. The aortoiliac bifurcation in the rabbit was examined following three types of insult: (1) diet-induced hypercholesterolemia resulting in macrophage-rich fatty streak lesions, (2) mechanical, stretch-denudation injury resulting in intimal smooth muscle cell (SMC) proliferation and (3) mechanical injury superimposed on hypercholesterolemia resulting in a complex vascular lesion having characteristics of both interventions. The normal rabbit iliac artery expressed approximately equal levels of Cx43 mRNA in the medial SMC layers and in the endothelium. In hypercholesterolemia-induced atherosclerosis, Cx43 expression was most prominent in macrophage foam cells even though normocholesterolemic precursor monocytes did not express Cx43 mRNA. Antibodies directed specifically to Cx43 protein confirmed the expression of macrophage gap junction protein in these cells. Medial SMC in hypercholesterolemia exhibited less Cx43 than their normal counterparts in control animals. Mechanical injury in the absence of hypercholesterolemia resulted in intimal thickening in which Cx43 expression in the intimal SMC was equivalent to that in the subjacent medial SMC, both being approximately equivalent to normal uninjured rabbit medial SMC expression. Cell-specific expression of Cx43 in combined mechanical injury/hypercholesterolemia was similar to that observed in hypercholesterolemia alone: Cx43 upregulation in macrophages, while medial SMC were downregulated. Normo- and hypercholesterolemic alveolar macrophages of the lung and Kupffer cells of the liver did not exhibit induction of Cx43 mRNA, nor did macrophages isolated from peritoneal or bronchial lavage fluid of the same animals. This work extends our previous finding of Cx43 upregulation in human atherectomy tissue and demonstrates that atherosclerotic lesions in situ, in a controlled animal model of atherosclerosis, exhibit cell-specific changes in Cx43 gene expression. Changes in medial SMC migration, proliferation and phenotype, as well as enhanced interactions between adherent/infiltrating monocytes and endothelium may be related to modified GJIC pathways in the vessel wall.

Animals↗
PubMed1999

Regulation of cell-cell communication in rat bone cells: the effect of phorbol esters.

The skin tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) is a potent inhibitor of gap junctional intercellular communication. In the present study, the inhibition of cell-cell communication by TPA has been investigated in primary bone cells from newborn rat calvaria, with an emphasis on the involvement of intracellular pH (pH(i)) and cytosolic calcium ([Ca(+2)](i)) in this process. The results show that TPA (5 x 10(-)(8) M) caused a complete inhibition of intercellular communication within 40-60 min. The intercellular communication was fully restored after overnight incubation in the presence of TPA. This effect was found to be associated with an elevation of pH(i). However, neither an increase of pH(i) alone nor exposure to TPA, under conditions preventing pH(i)-shift, were found to affect intercellular communication. It is suggested that the inhibition of intercellular communication, in the presence of TPA, depends on the pH(i)-shift itself rather than on the absolute value of pH(i). In addition, elevation of cytosolic calcium by ionomycin led to the termination of intercellular communication after 30 min. This inhibitory effect was abolished when the cells were incubated for overnight with TPA and then intracellular calcium was elevated by the addition of ionomycin. These results indicate that shift of pH(i) and the increase of intracellular calcium are involved in repression of intercellular communication by TPA.

Animals↗
PubMed1987

Selective lack of intercellular communication between transformed and nontransformed cells as a common property of chemical and oncogene transformation of BALB/c 3T3 cells.

BALB/c 3T3 cells can be transformed by transfection of an activated cellular oncogene as well as by chemicals. When the cells were transformed by pEJ-ras transfection, a marked increase in Mr 21,000 protein expression was found by Western blotting and immunohistochemical staining, whereas no such increase was detected in cells transformed by methylcholanthrene, suggesting two different molecular mechanisms. By directly microinjecting a fluorescent dye (Lucifer Yellow CH) into individual cells, we measured junctional intercellular communication among and between transformed and surrounding nontransformed cells. In both chemical and oncogene transformation studies, transformed cells and surrounding normal cells have similar capacities for gap-junctional communication, but there was complete lack of communication between transformed and nontransformed cells. When BALB/c 3T3 cells were transformed by methylcholanthrene initiation followed by phorbol ester promotion, again we saw no intercellular communication between transformed and nontransformed cells, suggesting that the observed selective communication block between transformed and nontransformed cells may be a general phenomenon in BALB/c 3T3 cells. These results indicate that selective lack of intercellular communication between transformed and surrounding normal cells may be an important phenomenon that separates transformed cells and nontransformed cells, permitting transformed cells to maintain autonomous growth.

Cell Communication↗
PubMed2003

Complex redundancy to build a simple epidermal permeability barrier.

To survive the transition from an aqueous in utero to a terrestrial ex utero environment, mice and humans must construct an epidermal permeability barrier in utero. Terminally differentiated epidermal cells, lipids and tight junctions are all essential to achieve this barrier. Recent analyses of mouse mutants with defects in structural components of the terminally differentiated epidermal cell, catalyzing enzymes, lipid processing, transcriptional regulators and the intercellular junctions have highlighted their essential function in establishing the epidermal permeability barrier. Particularly interesting examples include modulation of the expression of transglutaminase 1 enzyme, the transcription factor Klf4 and the claudin tight junction proteins. However, careful analysis of the various mutant phenotypes during embryonic development, as neonates and either as adults or transplanted skin, has revealed much more about the redundancy and compensatory mechanisms of the system. Molecular analysis of the various mouse mutants has demonstrated common pathways to compensate for loss of the epidermal barrier.

Animals↗
PubMed2004

Gap junctions are required for trophoblast proliferation in early human placental development.

Little is known about the role of gap junctional intercellular communication (GJIC) in human trophoblast differentiation, particularly during the formation of extravillous trophoblast (EVT) cell columns and their subsequent differentiation into invasive cells. We have identified transcripts for five connexin gap junction proteins in the early human placenta (Cx32, Cx37, Cx40, Cx43 and Cx45). Of these, Cx40 and Cx45 proteins immunolocalize to EVT in anchoring cell columns. Cx40 expression is prominent in the anchoring column throughout the first trimester of pregnancy (6-14 weeks gestation). We used first trimester placental villous explant cultures to determine the functional significance of the inhibition of GJIC in EVT cell proliferation and differentiation using two known GJIC inhibitors, carbenoxolone (CBX) and heptanol. The morphology of EVT outgrowths changed dramatically upon GJIC-blockade, from compact and organized outgrowths into a scattered group of rounded individual trophoblast cells, reminiscent of an early invasive phenotype. Furthermore, the inhibition of GJIC in placental explants by CBX or heptanol induced a switch away from the proliferative and towards an invasive EVT phenotype, as evident from (a) the loss of the proliferation marker Ki67 and (b) an increase in the invasive marker alpha1 integrin. We also utilized antisense oligonucleotides to inhibit Cx40 protein expression in placental explants. Cx40 antisense treatment also resulted in the abolishment of outgrowth EVT cell proliferation (as determined by Ki67 immunostaining). Together, these results suggest that gap junctions composed particularly of Cx40 channels are required for the proliferation of EVT cells in anchoring cell columns, and that a loss of GJIC contributes to differentiation to the invasive EVT phenotype.

Carbenoxolone↗
PubMed2000

Irsogladine upregulates expressions of connexin32 and connexin26 in the rat liver.

A gap junction is the channel for cell-to-cell communication and plays an important role in the maintenance of tissue homeostasis, control of cell growth and differentiation, and prevention of experimental hepatocarcino-genesis. Irsogladine, an antiulcer drug, augments gap junctional intercellular communication in gastric mucosa, but the effect of irsogladine on the liver remains uncertain. In this study the effects of irsogladine on the liver were investigated from the viewpoints of gap junctional protein connexin (Cx)32 and Cx26 in rats. Twelve rats were divided into a control group (n=6) and the irsogladine group (n=6) in which irsogladine (20 mg/kg per day) was administered orally for 3 days before sample collection, and the two groups were compared in regard to liver enzymes (serum aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactate dehydrogenase (LDH)), serum- and tissue-calcium concentrations, immunohistochemical expressions of Cx32 and Cx26, and RT-PCR analysis. In immunohistochemistry, analyzed using an image processor for analytical pathology (IPAP), the number of Cx32-positive spots was higher and the area of Cx26-positive spots were larger in the irsogladine group than those in the control group (P=0.036 and P=0.00032, respectively). In RT-PCR analysis, the mRNA of Cx32 or Cx26 in the irsogladine group showed a tendency to be higher than in the control group, but not significantly (Cx32, P=0.70; Cx26, P=0.07). Another 30 rats were used for measurements of cyclic-adenosine monophosphate (c-AMP) of the liver. c-AMP concentration was increased 1 h after the administration of irsogladine, which partially explained how the Cxs were upregulated. These findings may suggest that irsogladine upregulates Cx32 and Cx26 expressions in the liver of rats.

Journal Article↗