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At least 1,297 records · Page 72Linked to original sources
PubMed1992

Carotenoids up-regulate connexin43 gene expression independent of their provitamin A or antioxidant properties.

Epidemiological evidence and studies in whole animals and cell culture have indicated that carotenoids have cancer chemopreventive action. In mouse C3H10T1/2 cells, this activity is highly correlated with the ability of carotenoids to up-regulate gap junctional intercellular communication. Here, we report that in mouse cells, carotenoids increase the expression of connexin43, a gene that encodes a major gap junction protein. This effect appears unrelated to their provitamin A or antioxidant properties, since carotenoids with and without provitamin A activity increased levels of connexin43 mRNA and protein, whereas the antioxidants methyl-bixin and alpha-tocopherol were inactive. Moreover, the active carotenoid canthaxanthin did not induce the vitamin A-inducible gene retinoic acid receptor-beta. Connexin43 is the first carotenoid-inducible gene described in mammals. By indicating an additional pathway through which carotenoids function, these data provide a mechanistic basis for cancer chemoprevention by carotenoids and may lead to a re-evaluation of carotenoid physiology.

Animals↗
PubMed1983

Membrane junctions in the subodontoblastic region. A freeze-fracture study of the human dental pulp.

The morphology of the subodontoblastic region in the human dental pulp was analysed using the freeze-fracturing technique. Multiple areas of focal intercellular junctions were found between subodontoblastic fibroblasts (or Höhl's cells). The connections were of the gap junction type. Similar junctions were observed between subodontoblastic cells and small-calibre (0.10-0.20 microns) fibres, presumably nerves. Tight junctions were not observed.

Adolescent↗
PubMed1990

Intercellular dye-coupling in intestinal smooth muscle. Are gap junctions required for intercellular coupling?

The dye Lucifer Yellow was injected into single smooth muscle cells in the guinea pig small intestine in order to study intercellular coupling. Dye-coupling was observed in both the circular and longitudinal muscle layers and was markedly reduced when the intercellular pH was lowered. These results suggest the presence of gap junctions among intestinal muscle cells, but are inconsistent with previous ultrastructural studies that failed to demonstrate such junctions in the longitudinal muscle.

Animals↗
PubMed1995

Location of focal silver staining at endothelial gaps in inflamed venules examined by scanning electron microscopy.

The century-old histological technique of silver nitrate staining has proven to be extremely useful for visualizing endothelial cell borders and localizing endothelial gaps, but the significance of the staining is still not fully understood. To gain some insight into what silver nitrate stains, we developed a method that enabled us to use scanning electron microscopy with backscattered and secondary electron imaging to examine silver staining at endothelial cell borders of venules of the rat tracheal mucosa. We found that in normal venules, silver lines followed the smooth contour of cell borders. However, 1 min after endothelial permeability was increased by substance P, cell borders were irregular and displaced from the silver lines by as much as 4.3 microns, and the lines were accompanied by three types of silver deposits. Most common (46% of total) were annulus-shaped silver deposits that surrounded endothelial gaps. These deposits averaged 1.5 microns in width, were positioned symmetrically across cell borders, and were located at a depth of 0.3 micron beneath the luminal surface. Many endothelial gaps were partitioned into multiple pores (mean, 2.4) by fingerlike processes of endothelial cells. Surprisingly, the gaps occupied only 5.4% of the total area of the silver deposits and constituted 0.15% of the luminal surface of the leaky postcapillary venules. A second type of silver deposit (19% of total) was positioned asymmetrically with respect to the cell border and marked sites where endothelial cell margins still overlapped but appeared to be vertically separated by obliquely oriented gaps. A third type marked gaps at three-cell junctions; these were no more abundant than deposits elsewhere around the cell perimeter, suggesting that three-cell junctions were not unusually leaky sites. We conclude that silver nitrate marks endothelial cell borders and outlines endothelial cell gaps by staining an element of intercellular junctions. The annular shape of many silver deposits around gaps suggests that junctional elements in the apposing cells are separated during gap formation but are still present at the gap perimeter.

Animals↗
PubMed1989

[Establishment of primary culture of cells derived from human uterine endometrium].

A culture of cells derived from the endometrium was established to study in vitro the effects of hormone on the endometrium. Endometrial tissue was cut into small pieces and suspended in trypsin and collagenase-containing medium. The primary cell culture was obtained by six passages. The following observations appeared to indicate that the cultured cells consisted of endometrial epithelium-derived adenocytes, endometrial stromal cells and fibroblasts: 1. The epithelium-derived adenocytes and the endometrial stromal cells were positive and the fibroblasts were negative for staining with keratin by the enzyme-labelled antibody method. 2. Transmission electron microscopic observation revealed microvilli on the surface of the adenocytes, abundant free ribosomes in the cytoplasm, and nucleoli in distinct nuclei whose margin was deeply stained. Junction complexes among the cells were also observed. There were no microvilli on the surface of the endometrial stromal cell. Filaments, mitochondria and secreting granules were distinctly seen in the cytoplasm. The adenocytes on the margin of the nucleus itself were more deeply stained, and the margin was notched. The fibroblasts were spindle-shaped, and there no basement membrane structure laterally along the surface of the fibroblast. There were no intercellular junction complexes but there were filaments in the intercellular borders and in the cytoplasm. Chromatin in the large oval nuclei was dispersed.

Adult↗
PubMed1999

Genetic and epigenetic changes of intercellular communication genes during multistage carcinogenesis.

During multistage carcinogenesis, the functions of several key genes involved in cell growth control must be damaged. Such genes include not only those involved in cell cycle control of individual cells, but also those involved in the coordination of cell growth throughout a given tissue through cell-cell communication. The most intimate form of intercellular communication is mediated by gap junctions. Gap junctional intercellular communication (GJIC) is known to transfer small water soluble molecules, including cAMP and IP3, from the cytoplasm of one cell to that of its neighbors; the growth of a given GJIC-associated cell is thus kept in check by other GJIC-connected cells. Most tumor cells have a reduced ability to communicate among themselves and/or with surrounding normal cells, confirming the importance of intact GJIC in growth control. When connexin (gap junction protein) genes are transfected into such cells, normal cell growth control is often recovered. Certain dominant-negative mutant connexin genes can reverse such tumor suppression. While these results suggest that connexin genes form a family of tumor suppressor genes, so far we have found no connexin gene mutations in human tumors; only two connexin gene mutations were found in chemically induced rat tumors. On the other hand, our recent studies suggest that connexin genes may be inactivated by hypermethylation of their promoter regions, suggesting that epigenetic inactivation of connexin genes may be a mechanism of GJIC disturbance in certain tumors. However, in many tumor cells connexins are normally expressed but aberrantly localized. The mechanisms of aberrant localization of connexins include lack of an appropriate cell-cell recognition apparatus and aberrant phosphorylation of connexins. These results suggest that GJIC disorders may occur not only because of aberrant expression of connexin genes themselves, but also as a result of disruption of various control mechanisms of the protein functions.

Animals↗
PubMed1997

Dissociation and reassembly of adherens junctions during experimental acute pancreatitis.

BACKGROUND & AIMS: The initial pathophysiological events that characterize acute pancreatitis include the formation of pancreatic edema. An interstitial accumulation of fluid, however, is incompatible with the presence of intact intercellular junctions between acinar cells. This study examined the major components of adherens junctions, E-cadherin, alpha-catenin, beta-catenin, and actin, during the initial phase of experimental pancreatitis. METHODS: Pancreatitis was induced in rats by 10 micrograms.kg-1.h-1 intravenous cerulein for up to 12 hours. Adherens junction proteins were localized by immunocytochemistry for fluorescence microscopy or electron microscopy, their expression was studied by slot blot analysis, and their association was investigated by immunoprecipitation and Western blot. RESULTS: During a rapid increase of E-cadherin-encoding RNA, E-cadherin protein declined only moderately and, unlike its cytoskeletal binding partner actin, was not proteolytically cleaved during pancreatitis. Morphologically, E-cadherin and beta-catenin were localized at the basolateral cell membrane from where they rapidly dissociated early in pancreatitis and to where they slowly relocalized during the subsequent course. E-cadherin/beta-catenin complexes disintegrated and reassembled completely in parallel on immunoprecipitation experiments. CONCLUSIONS: The dissociation of adherens junctions and the internalization, relocalization, and reassembly of their major components seem to represent the critical biochemical event at cell-cell contacts during edema formation and resolution in acute pancreatitis.

Actins↗
PubMed1976

Ultrastructure and surface topography of aggregates of human limb mesenchymal cells (HLM15) in vitro.

Tissue-like aggregates of human embryo fibroblasts can be created in vitro by limited aspiration of cells released from substrate during subcultivation. Aggregates increase in size, exhibit intercellular junctions, display a surface topography characteristic of cellular movement, elaborate an extracellular matrix and possess features of cellular death and phagocytosis. These cells, when introduced to a new culture environment, do not migrate away from one another as is common when a primary culture is started from tissue fragments. Instead, cells exhibit continued contact with each other, and develop complex junctional structures during that association. Cellular aggregates generated in this manner may provide a useful system for providing further information on cellular adhesion, intercellular communication, morphogenetic cell movements and the mechanisms of cell death.

Cell Adhesion↗
PubMed1975

Properties of surface and junctional membranes of embryonic cells isolated from blastula stages of Xenopus laevis.

1. Some membrane properties of endoderm and mesoderm cells isolated from late blastula stages of Xenopus laevis have been examined using electrophysiological techniques. 2. Cells were isolated by treatment of whole embryos with Ca-free EDTA containing media, or mechanically by micro-dissection, and cultured in Ca-containing Holtfreter solution (60 mM-NaCl) or Ringer solution (120 mM-NaCl). 3. Membrane potentials lay between -6 and -84 mV; specific membrane resistances ranged from 500 to 29,000 omega cm2; there was no difference between EDTA isolated and mechanically isolated cells. 4. Relative and absolute cation and anion conductances varied from cell to cell. Some cells were anion impermeable; the cation conductance ranged from 35 to 300 mumho/cm2. 5. The resting potential of some cells was largely determined by the concentration gradient and membrane permeability of K ions. In other cells the potential was maintained either by some other ion or by an electrogenic pump. [K]i came to approximately 130 mM in Ringer solution (the value pertaining in the intact embryo) and similar to 60 mM in Holtfreter solution. 6. In most pairs and small clumps of cells ionic current spread from one cell to the next; some single cells and groups of cells were uncoupled from their neighbours. 7. The junctional resistance lay between 10(5) and 10(8) omega; it behaved as a linear resistor in most cell pairs studied. In three pairs the intercellular junction showed rectifying properites. 8. By the late blastula stage of development presumptive endoderm and mesoderm cells form a heterogeneous population with widely varying passive membrane properties. The significance of these findings is discussed in relation to current hypotheses for the formation of spatial patterns during differentiation.

Animals↗
PubMed2001

Connexin43 null mutation increases infarct size after stroke.

Glial-neuronal interactions have been implicated in both normal information processing and neuroprotection. One pathway of cellular interactions involves gap junctional intercellular communication (GJIC). In astrocytes, gap junctions are composed primarily of the channel protein connexin43 (Cx43) and provide a substrate for formation of a functional syncytium implicated in the spatial buffering capacity of astrocytes. To study the function of gap junctions in the brain, we used heterozygous Cx43 null mice, which exhibit reduced Cx43 expression. Western blot analysis showed a reduction in the level of Cx43 protein and GJIC in astrocytes cultured from heterozygote mice. The level of Cx43 is reduced in the adult heterozygote cerebrum to 40% of that present in the wild-type. To assess the effect of reduced Cx43 and GJIC on neuroprotection, we examined brain infarct volume in wild-type and heterozygote mice after focal ischemia. In our model of focal stroke, the middle cerebral artery was occluded at two points, above and below the rhinal fissure. Four days after surgery, mice were killed, the brains were sectioned and analyzed. Cx43 heterozygous null mice exhibited a significantly larger infarct volume compared with wild-type (14.4 +/- 1.4 mm(3) vs. 7.7 +/- 0.82 mm(3), P < 0.002). These results suggest that augmentation of GJIC in astrocytes may contribute to neuroprotection after ischemic injury.

Animals↗
PubMed2003

Expression and localization of tight junction-associated proteins in human hair follicles.

Tight junctions (TJ) are barrier-forming intercellular junctions selectively sealing cells and controlling the paracellular pathway. They have been well-characterized in simple epithelia and endothelia but have only recently been described in stratified epithelia such as epidermis, oesophagus and oral mucosa. Various epithelial layers which are partly in morphogenic continuity with the epidermis and develop therefrom during early fetal life, build the human hair follicle. The barrier function of these epithelial layers seems to be important for the universal continuity of the barrier represented by the skin. We show the presence of the TJ proteins ZO-1, occludin, and various claudins in the hair follicle and demonstrate their impressive heterogeneous distribution pattern within a given stratum as well as within its different epithelial layers. Coexpression of the various TJ proteins, arguing for typical TJ structures, can be observed especially in cell layers facing the hair shaft and the stratum corneum, and in addition at the border between the outer and inner root sheaths. Usually they are found in close proximity to desmosomal and adherens junction proteins. The morphological and biological importance of these findings and the possible roles of TJ in hair follicles, e.g. in follicular penetration, are discussed.

Adherens Junctions↗
PubMed2003

The JAM family of junctional adhesion molecules.

Junctional adhesion molecules are a family of glycoproteins characterised by two immunoglobulin folds (VH- and C2-type) in the extracellular domain. Junctional adhesion molecule proteins localise to intercellular junctions of polarised endothelial and epithelial cells but can also be expressed on circulating leukocytes and platelets. In addition, they bind several ligands, in both a homophilic and heterophilic manner, and associate with several cytoplasmic partners. All these features represent the likely determinants for the role of junctional adhesion molecule proteins in processes as diverse as junction assembly, leukocyte transmigration and platelet activation.

Animals↗
PubMed2003

Oscillating fluid flow regulates gap junction communication in osteocytic MLO-Y4 cells by an ERK1/2 MAP kinase-dependent mechanism.

The present work was designed to investigate the effects of oscillating fluid flow on gap junctional intercellular communication (GJIC) and the gap junction protein connexin (Cx) 43 in osteocyte-like MLOY-4 cells. Cells were exposed for 1 h to oscillating fluid flow at a shear stress of +/-10 dyn/cm(2) and a frequency of 1 Hz in a parallel plate flow chamber. Control cells were incubated in the chamber but were not exposed to oscillating fluid flow. Functional analysis of GJIC indicated that MLOY-4 cells exposed to oscillating fluid flow established more gap junctions with an independent population of dye-labeled cells than did control cells. Phosphorylation of Cx43 was quantified by immunoprecipitation with an anti-Cx43 antibody followed by immunoblot analysis using an anti-phosphoserine antibody. Phosphoserine was normalized to Cx43 in each sample. Compared to control cells, phosphoserine content of Cx43 increased approximately twofold in cells exposed to oscillating fluid flow. The possible role of the extracellular signal regulated kinase (ERK1/2) in the flow-induced upregulation of GJIC was also investigated. The ERK1/2 inhibitor PD-98059 significantly attenuated the effects of oscillating fluid flow on MLOY-4 cells GJIC. These results indicate that oscillating fluid flow regulates GJIC in MLOY-4 cells via the ERK1/2 MAP kinase. In addition, increased serine phosphorylation of Cx43 correlates with the flow-induced increase in GJIC.

Animals↗
PubMed2003

The relationship between connexins, gap junctions, tissue architecture and tumour invasion, as studied in a novel in vitro model of HPV-16-associated cervical cancer progression.

Disruption of gap junctional intercellular communication (GJIC) and/or connexins (gap junction proteins) is frequently reported in malignant cell lines and tumours. Certain human papillomaviruses (HPV) associated with the development of cancers, especially of the cervix, have previously been reported to downregulate GJIC in vitro. There is also evidence for reduced gap junctions in cervical dysplasia. However, many squamous hyperproliferative conditions, including HPV-induced warts, often show extensive upregulation of certain connexins. The association between HPV and GJIC, and the mechanism and consequence of deregulated GJIC in cervical tumour progression, remains unclear. Therefore, using a variety of nonmalignant and malignant cell lines and an organotypic raft-culture system, we investigated the relationship between HPV, gap junctions and tumour progression. Established cervical tumour cell lines carrying HPV were unable to communicate via gap junctions (when assayed by dye-transfer techniques). This correlated with lack of connexin protein expression, while transfection with connexins 26 or 43 led to functional gap junction membrane plaques. On the other hand, immortal but nonmalignant cell lines that contained episomal or integrated HPV-16, but required feeder-layer and growth-factor support, were consistently well coupled, and expressed multiple connexins at membrane junctions. In vitro selection of feeder-layer and growth-factor-independent variants eventually lead to loss of GJIC, which correlated with loss of membrane and increased cytoplasmic connexin 43 localization. However, this was preceded by loss of differentiation and stromal invasion, as assayed on the organotypic raft-culture model. Using this model, a comparison between noncoupled, well-coupled and connexin-transfected cell lines revealed no firm correlation between GJIC and dysplasia, but GJIC appeared to favour increased stratification. These findings demonstrate that loss of GJIC is frequent, but appears to occur more as a consequence of, rather than being the cause of, epithelial dysplasia, and may be influenced by, but is not directly attributable to, HPV.

Animals↗
PubMed2000

Tetraspanins are localized at motility-related structures and involved in normal human keratinocyte wound healing migration.

We have described previously that beta1 integrins, which mediate keratinocyte cell adhesion and migration, are in ligand-occupied conformation at the basal surface but not at the lateral and apical surfaces of keratinocytes. This led us to study the cellular localization and function of tetraspanin molecules, which have been postulated to modulate integrin activity. We found that CD9 and CD81 are highly expressed by keratinocytes clearly delineating filopodia at lateral and apical surfaces. CD63 and CD151 are largely expressed in the intracellular compartment, although some membrane expression is observed. We found accumulation of CD9, CD81, and CD151 together with alpha3 and beta1 integrins at intercellular junctions. In low calcium medium, this intercellular space is crossed by a zipper of filopodia enriched in alpha3beta1 and tetraspanin proteins. Interestingly, the expression of CD9, CD81, and beta1 and alpha3 integrins was detected in the footprints and rippings of motile keratinocytes, suggesting their role in both adhesion to extracellular matrix and keratinocyte motility. beta1 integrins were only partially activated in the rips, whereas cytoskeleton-linking proteins such as talin were completely absent. On the other hand, antitetraspanin antibodies did not stain focal adhesions, which contain talin. The involvement of tetraspanins in keratinocyte motility was assessed in a wound healing migration assay. Inhibition of cell migration was observed with antibodies to CD9, CD81, beta1, and alpha3, and, to a lesser extent, to CD151. Together these results indicate that tetraspanin-integrin complexes might be involved in transient adhesion and integrin recycling during keratinocyte migration, as well as in intercellular recognition.

Antigens, CD↗
PubMed1996

Functional dissociation of paracellular permeability and transepithelial electrical resistance and disruption of the apical-basolateral intramembrane diffusion barrier by expression of a mutant tight junction membrane protein.

Tight junctions, the most apical of the intercellular junctions that connect individual cells in a epithelial sheet, are thought to form a seal that restricts paracellular and intramembrane diffusion. To analyze the functioning of tight junctions, we generated stable MDCK strain 2 cell lines expressing either full-length or COOH-terminally truncated chicken occludin, the only known transmembrane component of tight junctions. Confocal immunofluorescence and immunoelectron microscopy demonstrated that mutant occludin was incorporated into tight junctions but, in contrast to full-length chicken occludin, exhibited a discontinuous junctional staining pattern and also disrupted the continuous junctional ring formed by endogenous occludin. This rearrangement of occludin was not paralleled by apparent changes in the junctional morphology as seen by thin section electron microscopy nor apparent discontinuities of the junctional strands observed by freeze-fracture. Nevertheless, expression of both wild-type and mutant occludin induced increased transepithelial electrical resistance (TER). In contrast to TER, particularly the expression of COOH-terminally truncated occludin led to a severalfold increase in paracellular flux of small molecular weight tracers. Since the selectivity for size or different types of cations was unchanged, expression of wild-type and mutant occludin appears to have activated an existing mechanism that allows selective paracellular flux in the presence of electrically sealed tight junctions. Occludin is also involved in the formation of the apical/basolateral intramembrane diffusion barrier, since expression of the COOH-terminally truncated occludin was found to render MDCK cells incapable of maintaining a fluorescent lipid in a specifically labeled cell surface domain.

Amino Acid Sequence↗
PubMed1968

Ultrastructural studies on the permeability of the mesothelium to horseradish peroxidase.

Peritoneal mesothelium was exposed for 2-60 min to solutions of horseradish peroxidase by incubation in vitro, or after intraperitoneal injection in vivo. Peroxidase was localized, with the electron microscope in the intercellular clefts of the mesothelium, often along their entire lengths, in vesicles adjoining or contiguous with the clefts, and along the peritoneal and basal surfaces of the cell, and also in intracytoplasmic vacuoles. The intercellular junctions of peroxidase-treated mesothelium did not differ from those of controls: open and closed junctions were present in both groups. Intercellular localization was also obtained when the mesothelium was exposed to peroxidase during or after fixation. Although intracellular absorption of peroxidase and its incorporation into larger vacuoles were observed, there was no clearcut evidence of vesicular transport across the mesothelium in these experiments. These findings are consistent with physiologic data which postulate that mesothelial transport can be accounted for, at least in part, by passive diffusion through a system of pores, and they suggest that these pores are located in the intercellular clefts.

Animals↗
PubMedDate not supplied

Malignant fibrous histiocytoma of bone: an ultrastructural study.

An ultrastructural study of a case of malignant fibrous histiocytoma of bone in a 16-year-old skeletally mature female is presented. There were multiple metastatic bone lesions and a marked hypercalcemia. The cell population was similar to that found in soft tissue malignant fibrous histiocytoma lesions and comprised undifferentiated cells, fibroblastlike cells, histiocytes, and multinucleated giant cells. Occasional myofibroblasts and transitional cells with histiocytic and fibroblastic components were seen. The histiocytes were characterized by prominent Golgi bodies. Although there were many fibroblastlike cells, the rough endoplasmic reticulum was rarely as extensive or as well organized as in normal fibroblasts. The giant cells did not have the ultrastructural characteristics of osteoclasts, i.e., the clear zones and ruffled borders. Zonula adherens (belt desmosome) junctions were seen, but in general, intercellular junctions were poorly developed as to length and extent.

Adolescent↗