Functional organization of the bovine rumen epithelium

The functional organization of the bovine rumen epithelium has been examined by electron and light microscopy combined with immunocytochemistry to define a transport model for this epithelium. Expression of connexin 43, an integral component of gap junctions, the tight-junction molecules claudin-1 and zonula occludens 1 (ZO-1), and the catalytic α-subunit of Na+-K+-ATPase was demonstrated by SDS-PAGE and Western blotting. From the lumen surface, four cell layers can be distinguished: the stratum corneum, the stratum granulosum, the stratum spinosum, and the stratum basale. Both claudin-1 and ZO-1 immunostaining showed plasma membrane staining, which was present at the stratum granulosum with decreasing intensity through the stratum spinosum to the stratum basale. The stratum corneum was negative for claudin-1 immunostaining. Transmission electron microscopy confirmed that occluding tight junctions were present at the stratum granulosum. Plasma membrane connexin 43 immunostaining was most intense at the stratum granulosum and decreased in intensity through stratum spinosum and stratum basale. There was intense immunostaining of the stratum basale for Na+-K+-ATPase, with weak staining of the stratum spinosum. Both the stratum granulosum and the stratum corneum were essentially negative. Stratum basale cells also displayed a high mitochondrial density relative to more apical cell layers. We conclude that epithelial barrier function may be attributed to the stratum granulosum and that cell-cell gap junctions allow diffusion to interconnect the barrier cell layer with the stratum basale where Na+-K+-ATPase is concentrated.

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Demonstration of gap junctions in frog skin epithelium

AJP Cell Physiology ◽

10.1152/ajpcell.1989.257.4.c658 ◽

1989 ◽

Vol 257 (4) ◽

pp. C658-C664 ◽

Cited By ~ 20

Author(s):

S. H. Shahin ◽

J. T. Blankemeyer

Keyword(s):

Gap Junctions ◽

Stratum Corneum ◽

Intercellular Junctions ◽

Lower Number ◽

Stratum Granulosum ◽

Intercellular Coupling ◽

Electron Microscopic ◽

Frog Skin Epithelium ◽

Stratum Spinosum ◽

Microscopic Techniques

The morphology and distribution of the intercellular junctions were investigated in isolated skin of Rana pipiens using various electron-microscopic techniques. Our evidence demonstrates the presence of gap junctions and suggests that the distribution of gap junctions is not homogeneous among the epithelial strata. Gap junctions were less frequent in the stratum corneum and stratum granulosum than in the stratum spinosum and stratum germinativum. These results support a model of widespread intercellular coupling, although the lower number of gap junctions in the stratum granulosum suggests a possible deficiency in intercellular coupling. Tight junctions were found only in two apical strata of the epithelium (stratum corneum and stratum granulosum). Desmosomes were located in all strata.

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The Ultrastructure of Monkey Gingival Epithelium

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010006009x ◽

1967 ◽

Vol 25 ◽

pp. 16-17

Author(s):

C.N. Sun

Keyword(s):

Epithelial Cells ◽

Macaca Nemestrina ◽

Stratum Granulosum ◽

Solution Ph ◽

Gingival Epithelial Cells ◽

Stratum Spinosum ◽

Cell Layers ◽

Gingival Epithelium ◽

The Individual ◽

Different Thickness

The present study demonstrates the ultrastructure of the gingival epithelium of the pig tail monkey (Macaca nemestrina). Specimens were taken from lingual and facial gingival surfaces and fixed in Dalton's chrome osmium solution (pH 7.6) for 1 hr, dehydrated, and then embedded in Epon 812.Tonofibrils are variable in number and structure according to the different region or location of the gingival epithelial cells, the main orientation of which is parallel to the long axis of the cells. The cytoplasm of the basal epithelial cells contains a great number of tonofilaments and numerous mitochondria. The basement membrane is 300 to 400 A thick. In the cells of stratum spinosum, the tonofibrils are densely packed and increased in number (fig. 1 and 3). They seem to take on a somewhat concentric arrangement around the nucleus. The filaments may occur scattered as thin fibrils in the cytoplasm or they may be arranged in bundles of different thickness. The filaments have a diameter about 50 A. In the stratum granulosum, the cells gradually become flatted, the tonofibrils are usually thin, and the individual tonofilaments are clearly distinguishable (fig. 2). The mitochondria and endoplasmic reticulum are seldom seen in these superficial cell layers.

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N-cadherin in adult rat cardiomyocytes in culture. II. Spatio-temporal appearance of proteins involved in cell-cell contact and communication. Formation of two distinct N-cadherin/catenin complexes

Journal of Cell Science ◽

10.1242/jcs.109.1.11 ◽

1996 ◽

Vol 109 (1) ◽

pp. 11-20 ◽

Cited By ~ 7

Author(s):

C.M. Hertig ◽

S. Butz ◽

S. Koch ◽

M. Eppenberger-Eberhardt ◽

R. Kemler ◽

...

Keyword(s):

Plasma Membrane ◽

Gap Junctions ◽

Connexin 43 ◽

Cell Contact ◽

Beta Catenin ◽

Rat Cardiomyocytes ◽

Cell Contacts ◽

Adult Rat ◽

Spatio Temporal ◽

Cell Cell

The spatio-temporal appearance and distribution of proteins forming the intercalated disc were investigated in adult rat cardiomyocytes (ARC). The ‘redifferentiation model’ of ARC involves extensive remodelling of the plasma membrane and of the myofibrillar apparatus. It represents a valuable system to elucidate the formation of cell-cell contact between cardiomyocytes and to assess the mechanisms by which different proteins involved in the cell-cell adhesion process are sorted in a precise manner to the sites of function. Appearance of N-cadherin, the catenins and connexin43 within newly formed adherens and gap junctions was studied. Here first evidence is provided for a formation of two distinct and separable N-cadherin/catenin complexes in cardiomyocytes. Both complexes are composed of N-cadherin and alpha-catenin which bind to either beta-catenin or plakoglobin in a mutually exclusive manner. The two N-cadherin/catenin complexes are assumed to be functionally involved in the formation of cell-cell contacts in ARC; however, the differential appearance and localization of the two types of complexes may also point to a specific role during ARC differentiation. The newly synthesized beta-catenin containing complex is more abundant during the first stages in culture after ARC isolation, while the newly synthesized plakoglobin containing complex progressively accumulates during the morphological changes of ARC. ARC formed a tissue-like pattern in culture whereby the new cell-cell contacts could be dissolved through Ca2+ depletion. Presence of cAMP and replenishment of Ca2+ content in the culture medium not only allowed reformation of cell-cell contacts but also affected the relative protein ratio between the two N-cadherin/catenin complexes, increasing the relative amount of newly synthesized beta-catenin over plakoglobin at a particular stage of ARC differentiation. The clustered N-cadherin/catenin complexes at the plasma membrane appear to be a prerequisite for the following gap junction formation; a temporal sequence of the appearance of adherens junction proteins and of gap junctions forming connexin-43 is suggested.

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Expression and localization of monocarboxylate transporters and sodium/proton exchangers in bovine rumen epithelium

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.00343.2006 ◽

2007 ◽

Vol 292 (2) ◽

pp. R997-R1007 ◽

Cited By ~ 72

Author(s):

C. Graham ◽

I. Gatherar ◽

I. Haslam ◽

M. Glanville ◽

N. L. Simmons

Keyword(s):

Family Members ◽

Short Chain ◽

Monocarboxylate Transporter ◽

Permeability Barrier ◽

Stratum Granulosum ◽

Rt Pcr ◽

Intense Staining ◽

Intracellular Location ◽

Bovine Rumen ◽

Rumen Epithelium

Monocarboxylate-H+ cotransporters, such as monocarboxylate transporter (MCT) SLC16A, have been suggested to mediate transruminal fluxes of short-chain fatty acids, ketone bodies, and lactate. Using an RT-PCR approach, we demonstrate expression of MCT1 (SLC16A1) and MCT2 (SLC16A7) mRNA in isolated bovine rumen epithelium. cDNA sequence from these PCR products combined with overlapping expressed sequence tag data allowed compilation of the complete open reading frames for MCT1 and MCT2. Immunohistochemical localization of MCT1 shows plasma membrane staining in cells of the stratum basale, with intense staining of the basal aspects of the cells. Immunostaining decreased in the cell layers toward the rumen lumen, with weak staining in the stratum spinsoum. Immunostaining in the stratum granulosum and stratum corneum was essentially negative. Since monocarboxylate transport will load the cytosol with acid, expression and location of Na+/H+ exchanger (NHE) family members within the rumen epithelium were determined. RT-PCR demonstrates expression of multiple NHE family members, including NHE1, NHE2, NHE3, and NHE8. In contrast to MCT1, immunostaining showed that NHE1 was predominantly localized to the stratum granulosum, with a progressive decrease toward the stratum basale. NHE2 immunostaining was observed mainly at an intracellular location in the stratum basale, stratum spinosum, and stratum granulosum. Given the anatomic localization of MCT1, NHE1, and NHE2, the mechanism of transruminal short-chain fatty acid, ketone body, and lactate transfer is discussed in relation to a functional model of the rumen epithelium comprising an apical permeability barrier at the stratum granulosum, with a cell syncitium linking the stratum granulosum to the blood-facing stratum basale.

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Connexin 43 K63-polyubiquitination on lysines 264 and 303 regulates gap junction internalization

10.1101/211607 ◽

2017 ◽

Cited By ~ 1

Author(s):

Rachael M. Kells-Andrews ◽

Rachel A. Margraf ◽

Charles G. Fisher ◽

Matthias M. Falk

Keyword(s):

Plasma Membrane ◽

Gap Junctions ◽

Gap Junction ◽

Connexin 43 ◽

Room Temperature ◽

Cell Communication ◽

Mapk Phosphorylation ◽

Annular Gap ◽

Summary Statement ◽

Triton X

ABSTRACTGap junctions (GJs) assembled from connexin (Cx) proteins play a pivotal role in cell-to-cell communication by forming channels that connect the cytosols of adjacent cells. Connexin 43, the best-studied Cx, is ubiquitously expressed in vertebrates. While phosphorylation is known to regulate multiple aspects of GJ function, much less is known about the role ubiquitination plays in these processes. Here we show by using ubiquitination-type specific antibodies and Cx43 lysine (K) to arginine (R) mutants that a portion of Cx43 in GJs can become K63-polyubiquitinated on K264 and K303. Relevant Cx43 K/R mutants assembled significantly larger GJ plaques, exhibited much longer protein half-lives and were internalization impaired. Interestingly, ubiquitin-deficient Cx43 mutants accumulated as hyper-phosphorylated polypeptides in the plasma membrane, suggesting that K63-polyubiquitination may be triggered by phosphorylation. Phospho-specific Cx43 antibodies revealed that upregulated phosphorylation affected serines 368, 279/282, and 255, well-known regulatory PKC and MAPK phosphorylation sites. Together, these novel findings suggest that upon internalization, some Cx43 in GJs becomes K63-polyubiquitinated, ubiquitination is critical for GJ internalization, and that K63-polyubiquitination may be induced by Cx phosphorylation.Summary StatementHere we show that connexin 43 in gap junctions becomes K63-poly ubiquitinated on lysines 264 and 303 and its requirement for gap junction endocytosis. These novel findings significantly contribute to our understanding of GJ turnover and patho-/physiology.Abbreviations usedAGJannular gap junctionAMSHassociated molecule with the SH3 domain of STAMCMEclathrin-mediated endocytosisCxConnexinCx43Connexin 43DUBdeubiquitinaseGJgap junctionMonoUbmonoubiquitinNedd4-1neural precursor cell expressed developmentally down-regulated protein 4-1PMplasma membranePolyUbpolyubiquitinTPA12-O-Tetradecanoylphorbol 13-AcetateTX-100Triton X-100RTroom temperatureUbubiquitin

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The permeability barrier in mammalian epidermis.

The Journal of Cell Biology ◽

10.1083/jcb.65.1.180 ◽

1975 ◽

Vol 65 (1) ◽

pp. 180-191 ◽

Cited By ~ 491

Author(s):

P M Elias ◽

D S Friend

Keyword(s):

Plasma Membrane ◽

Stratum Corneum ◽

Freeze Fracture ◽

Freeze Substitution ◽

Water Soluble ◽

Structural Basis ◽

Fracture Plane ◽

Permeability Barrier ◽

Stratum Granulosum ◽

Lamellar Bodies

The structural basis of the permeability barrier in mammalian epidermis was examined by tracer and freeze-fracture techniques. Water-soluble tracers (horesradish peroxidase, lanthanum, ferritin) were injected into neonatal mice or into isolated upper epidermal sheets obtained with staphylococcal exfoliatin. Tracers percolated through the intercellular spaces to the upper stratum granulosum, where further egress was impeded by extruded contents of lamellar bodies. The lamellar contents initially remain segregated in pockets, then fuse to form broad sheets which fill intercellular regions of the stratum corneum, obscuring the outer leaflet of the plasma membrane. These striated intercellular regions are interrupted by periodic bulbous dilatations. When adequately preserved, the interstices of the stratum corneum are wider, by a factor of 5-10 times that previously appreciated. Freeze-fracture replicas of granular cell membranes revealed desmosomes, sparse plasma membrane particles, and accumulating intercellular lamellae, but no tight junctions. Fractured stratum corneum displayed large, smooth, multilaminated fracture faces. By freeze-substitution, proof was obtained that the fracture plane had diverted from the usual intramembranous route in the stratum granulosum to the intercellular space in the stratum corneum. We conclude that: (a) the primary barrier to water loss is formed in the stratum granulosum and is subserved by intercellular deposition of lamellar bodies, rather than occluding zonules; (b) a novel, intercellular freeze-fracture plane occurs within the stratum corneum; (c) intercellular regions of the stratum corneum comprise an expanded, structurally complex, presumably lipid-rich region which may play an important role in percutaneous transport.

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Connexin 43 ubiquitination determines the fate of gap junctions: restrict to survive

Biochemical Society Transactions ◽

10.1042/bst20150036 ◽

2015 ◽

Vol 43 (3) ◽

pp. 471-475 ◽

Cited By ~ 7

Author(s):

Teresa M. Ribeiro-Rodrigues ◽

Steve Catarino ◽

Maria J. Pinho ◽

Paulo Pereira ◽

Henrique Girao

Keyword(s):

Plasma Membrane ◽

Gap Junctions ◽

Gap Junction ◽

Intercellular Communication ◽

Connexin 43 ◽

Transmembrane Proteins ◽

Post Translational Modifications ◽

Gap Junction Intercellular Communication

Connexins (Cxs) are transmembrane proteins that form channels which allow direct intercellular communication (IC) between neighbouring cells via gap junctions. Mechanisms that modulate the amount of channels at the plasma membrane have emerged as important regulators of IC and their de-regulation has been associated with various diseases. Although Cx-mediated IC can be modulated by different mechanisms, ubiquitination has been described as one of the major post-translational modifications involved in Cx regulation and consequently IC. In this review, we focus on the role of ubiquitin and its effect on gap junction intercellular communication.

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Histomorphological and Histochemical Studies on Esophagus in Gaddi Sheep (Ovis aries)

THE INDIAN JOURNAL OF VETERINARY SCIENCES AND BIOTECHNOLOGY ◽

10.21887/ijvsbt.14.2.5 ◽

2018 ◽

Vol 14 (2) ◽

Author(s):

Shabir Ahmad Malik ◽

Rajesh Rajput ◽

Mohd Rafiq ◽

Uiase Bin Farooq ◽

Harishbhai Gori

Keyword(s):

Connective Tissue ◽

Stratum Corneum ◽

Lamina Propria ◽

Periodic Acid ◽

Basal Layer ◽

Ovis Aries ◽

Stratum Granulosum ◽

Cell Nuclei ◽

Blood Capillaries ◽

Stratum Spinosum

The present work was conducted to study the histoarchitecture and histochemical characteristics of esophagus in six adult Gaddi sheep. Lamina epithelialis consisted of keratinized stratified squamous epithelium with four functional regions: stratum corneum, stratum granulosum, stratum spinosum and stratum basale. In the stratum spinosum layer, the cell nuclei appeared polygonal whereas in the stratum corneum the cell nuclei were flattened and condensed. The stratum spinosum was the thicker layer and stratum granulosum was a thin layer that contained basophilic keratohyalin granules. Pyknotic cells were observed towards the luminal side of stratum corneum. Blood capillaries and lymphoid aggregations in the form of dark stained cells were present in the connective tissue of lamina propria. Connective tissue of lamina propria layer was denser than the same of the submucosa. The tunica muscularis consisted of striated muscle cells throughout the length of esophagus. Stratum corneum of the stratified epithelium of esophagus showed strong periodic acid-Schiff reaction indicating accumulation of glycogen whereas the cells of the basal layer lacked glycogen. The intercellular spaces in the upper layers of stratum spinosum of the epithelium contained acidic mucopolysaccharides as indicated by their reactivity to alcian blue stain.

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