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Query: UNIPROT:P06889 (Mol)
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An air-liquid interface (biphasic) primary culture system in which guinea pig tracheal epithelial cells maintain morphologic characteristics of differentiated epithelium has been developed in this laboratory. In this report, we compared quantitatively cell populations of 8-day cultures to those of epithelial mucosa in intact trachea. In addition, high molecular weight glycoconjugates released by the cultured cells were isolated and characterized. Quantitative morphometric analysis revealed similar volume densities of ciliated, secretory, basal, and "other" cells in cultures and in intact tracheal surface epithelium, although the cultures tended to have smaller cells and contained fewer basal cells. High molecular weight glycoconjugates released apically by cell cultures and excluded from Sepharose CL-4B columns contained approximately 5% hyaluronic acid but undetectable amounts of other proteoglycans, such as chondroitin sulfate, heparan sulfate, and dermatan sulfate. The hyaluronidase-resistant glycoconjugates exhibited a peak buoyant density at 1.49 g/ml on cesium chloride density gradient centrifugation and were shown to contain mucin-type carbohydrate to peptide linkages (i.e., GalNAc to ser/thr) and an amino acid composition typical of respiratory mucins. The results indicate that this organotypic cell culture system mimics quite closely morphology of mucosal epithelium in intact airways and that the cells release high molecular weight glycoconjugates with biochemical properties of mucin-type glycoproteins. Thus, this in vitro system appears well-suited for studies of mucin secretion and other functions of respiratory epithelial cells.
Am J Respir Cell Mol Biol 1990 Feb
PMID:Characterization of guinea pig tracheal epithelial cells maintained in biphasic organotypic culture: cellular composition and biochemical analysis of released glycoconjugates. 230 71

Irradiation with a single dose of 30 Grey on the basal regions of the lungs of Sprague-Dawley rats induced a peribronchial and alveolar inflammation. Infiltration of mast cells in the edematous alveolar interstitial tissue and also in the peribronchial tissue were characteristic features of the lesion. The appearance of mast cells was already seen 4 wk after irradiation and by weeks 6 to 8 there was a heavy infiltration. The staining properties suggested that they were connective tissue-type mast cells. The infiltration of mast cells was paralleled by an accumulation of hyaluronan (hyaluronic acid) in the alveolar interstitial tissue 6 and 8 wk after irradiation. The recovery of hyaluronan (HA) during bronchoalveolar lavage (BAL) of the lungs also increased at this time. Treatment with a mast cell secretagogue, compound 48/80, induced a distinct reduction of granulated mast cells in the alveolar tissue. Regular treatment with compound 48/80 from the time of irradiation considerably reduced the HA recovery during BAL and the HA accumulation in the interstitial tissue but did not affect the interstitial infiltration of mononuclear cells and polymorphonuclear leukocytes. By contrast, an accumulation of HA in the alveolar interstitial space was induced when compound 48/80 was given not until mast cell infiltration of the lung had started. The effects of compound 48/80 indicate that the connective tissue response after lung irradiation is dependent on whether or not mast cell degranulation is induced before or after the mast cell infiltration of the alveolar tissue.(ABSTRACT TRUNCATED AT 250 WORDS)
Am J Respir Cell Mol Biol 1990 Feb
PMID:A mast cell secretagogue, compound 48/80, prevents the accumulation of hyaluronan in lung tissue injured by ionizing irradiation. 230 75

The rat renal papillary interstitum which contains abundant proteoglycans is a unique area important in renal function. These proteoglycans were studied ultrastructurally by ruthenium red fixation and staining and phosphate-buffered fixation before and after enzyme digestion. A tissue culture of rat renomedullary interstitial cells, the predominant cell of the renal papillary interstitum, was studied for its ability to synthesize proteoglycans and the proteoglycans were then analyzed. Tissue slices of whole rat renal inner medulla were also evaluated for their synthetic ability. In combination, these studies indicate that the dominant glycosaminoglycan is hyaluronic acid. The tissue culture of rat renal medullary interstitial cells synthesized glycosaminoglycans and on analysis, hyaluronic acid was found to be the chief glycosaminoglycan secreted by the renomedullary interstitial cells. Combined with the removal of the proteoglycans from tissue by leech hyaluronidase and testicular hyaluronidase, this suggests that the dominant glycosaminoglycan is hyaluronic acid. Hyaluronic acid is also synthesized by the intact papilla confirming the findings with the tissue culture. However, in addition, sulfated glycosaminoglycans were also synthesized by the intact papilla, presumably the product of the noninterstitial components of the papilla.
Exp Mol Pathol 1988 Dec
PMID:Glycosaminoglycans of the rat renomedullary interstitium: ultrastructural and biochemical observations. 246 72

The glycosaminoglycans (GAG) biosynthesized by a neoplastic human salivary duct cell line, HSGc, and by its nontumorigenic subclone, HSGc-E1, having a myoepithelial-like phenotype, were examined by incorporation of [3H]-acetate into GAG. The rate of GAG radiolabeling in HSGc-E1 was significantly greater than that in HSGc. The radiolabeled GAG recovered from HSGc-E1 showed a distribution of 22-32% in the cells and 68-78% secreted into the medium, while the amounts of GAG in the cells and medium of HSGc were equal. Two-dimensional electrophoresis of GAG extracted from the cells demonstrated that HSGc-E1 contained a much greater amount of heparan sulfate (HS, 53.5% of total), while HSGc synthesized hyaluronic acid (HA, 17.5%), HS 38.8%, chondroitin sulfate (Ch-S, 27.6%) and dermatan sulfate (DS, 16.1%). Moreover, treatment of HSGc with sodium butyrate or dibutyryl cyclic AMP (each is a potent inducer of differentiation to myoepithelial-like cells) strongly enhanced GAG synthesis, while dexamethasone (an inducer of differentiation to a more functional duct epithelium) did not stimulate GAG synthesis. These findings suggest that biosynthetic changes in the GAG content of neoplastic salivary cells are associated with their myoepithelial differentiation.
Virchows Arch B Cell Pathol Incl Mol Pathol 1989
PMID:Different contents of glycosaminoglycans in a human neoplastic salivary duct cell line and its subclone with a myoepithelial phenotype. 257 Apr 84

To study the roles played by cardiac valvular endothelium in normal and pathologic conditions, we have established and characterized a system of bovine valvular endothelial cells (VEC) in culture. Viable VEC from calf atrioventricular valves were obtained by a non-enzymatic procedure using 3 mM ethylenediamine-tetraacetic acid (EDTA) as dissociating agent. The cells grown in Dulbecco's modified Eagle's medium supplemented with non-essential amino acids, vitamins and 20% fetal calf serum, developed as monolayers of closely apposed polygonal cells which were subcultured for up to seven passages. VEC maintained in culture the general ultrastructure displayed in vivo, expressed von Willebrand factor, presented angiotensin converting enzyme activity and synthesized a rich extracellular matrix. VEC preserved the cell surface anionic sites (detected with cationized ferritin, pI 8.4) and cationic sites (visualized with haemeundecapeptide pI 4.85), and took up, especially by adsorptive endocytosis, albumin-gold conjugate. The cells were coupled by functional communicating (gap) junctions, as demonstrated by microinjection of 6-carboxyfluorescein. VEC in culture produced fibronectin, prostacyclin, hyaluronic acid and heparin-like glycosaminoglycans (identified by electrophoresis, enzyme digestion, and deaminative cleavage of molecules). These properties render cultured VEC a suitable model for investigating their functions and involvement in normal and pathologic heart valves.
J Mol Cell Cardiol 1988 Feb
PMID:Calf cardiac valvular endothelial cells in culture: production of glycosaminoglycans, prostacyclin and fibronectin. 284 May 11

Long-term synovial fibroblast cultures were exposed to interleukin 1 (IL-1) or prostaglandin E2 (PGE2). The normally spindle-shaped fibroblasts changed to stellate-shaped cells, resembling the HLA-DR-positive, collagenase-producing cells which are normally seen only in primary cultures from enzyme-digested rheumatoid synovial tissue. However, the IL-1- or PGE2-induced fibroblasts were not HLA-DR-positive. This suggests that these cell populations represent originally different cell lines or that the expression of HLA-DR antigens is not induced by the agents used. For further characterization of these stellate cells, the location of fibronectin and type I collagen was studied by specific antibodies and the pericellular coat around fibroblasts was visualized by the erythrocyte exclusion method. Both IL-1 and PGE2 treatments destroyed the intercellular fibronectin network. Type I collagen was detected as intracellular granules. The stellate fibroblasts were usually full of these granules in contrast to intact fibroblasts in which the number of collagen fluorescence granules varied greatly. The pericellular coat known to be formed mainly by hyaluronic acid was similar around spindle and stellate-shaped fibroblasts. Rheumatoid arthritis-derived fibroblasts did not differ from their non-rheumatoid counterparts in any of the experiments. The effect of IL-1 and PGE2 on fibroblasts simulates the interaction between mononuclear cells and fibroblasts in synovial stroma and also potentially the interactions between different cell types in synovial lining.
Virchows Arch B Cell Pathol Incl Mol Pathol 1986
PMID:Connective tissue components in synovial fibroblast cultures exposed to interleukin 1 and prostaglandin E2. 287 May 82

Analysis of mouse serum hyaluronidase by polyacrylamide gel electrophoresis, with the substrate high-molecular-weight hyaluronan (hyaluronic acid) included in the gel performed in mice of two different strains, BALB/cBy and C57BL/6By, reveals a pattern of multiple enzyme forms specific for each genotype. In BALB/c serum, seven different forms are present, only one of which is found in C57BL/6 serum. Segregation analysis of the enzyme polymorphism in backcross progeny and in recombinant inbred and bilineal congenic lines shows that the difference is due to a single locus, which we have designated as Hyal-1. Hyal-1 is linked to the histocompatibility locus H-7, on chromosome 9.
Somat Cell Mol Genet 1989 Jan
PMID:Hyal-1, a locus determining serum hyaluronidase polymorphism, on chromosome 9 in mice. 291 64

In contrast to glutaraldehyde-fixed vascular tissue with or without staining with cationic dye, the nonfibrous extracellular matrix of fast-frozen, freeze-dried rabbit aorta and renal artery contained a continuous reticulum of fine filaments, closely associated with collagen, elastin, and smooth muscle cells. Three morphologically distinct types of filament were distinguished; one type was selectively sensitive to chondroitinase ABC degradation, and therefore contains chondroitin and/or dermatan sulfate. The remaining filaments of the reticulum may represent the protein core of the proteoglycan monomer, and the hyaluronic acid backbone of the aggregate. Filaments associated with the surface of smooth muscle cells were usually linked to a continuous filament parallel to the cell surface, which was degraded by heparitinase and therefore contains heparan sulfate. The filaments linked directly to the cell surface were not degraded by either enzyme. The preservation of PG in fast-frozen material provides a significant improvement over that obtained by any presently available technique.
J Ultrastruct Mol Struct Res 1988 Feb
PMID:Proteoglycan in fast-frozen, freeze-dried, plastic-embedded rabbit arteries. 337 71

Heparin (5 U/ml) induced the release of LPL into the incubation medium of cardiac myocytes isolated from adult rat hearts. The secretion of LPL occurred in two phases: a rapid release (5-10 min of incubation with heparin) that was independent of protein synthesis followed by a slower rate of release that was inhibited by cycloheximide. The rapid release of LPL induced by heparin likely occurs from sites that are at or near the cell surface. LPL secretion could also be stimulated by heparan sulfate and dermatan sulfate, but not by hyaluronic acid, chondroitin sulfate or keratan sulfate. Heparin-releasable LPL activity measured in short-term incubations represented a large fraction (40-50%) of the initial LPL activity associated with myocytes, but the fall in cellular LPL activity following heparin was less than the amount of LPL activity secreted into the incubation medium. This discrepancy was not due to latency of LPL in the pre-heparin cell homogenates, but in part could be due to a three-fold greater affinity of the heparin-released enzyme for substrate as compared to LPL in post-heparin myocyte homogenates.
Mol Cell Biochem 1988 Jan
PMID:Secretion of lipoprotein lipase from myocardial cells isolated from adult rat hearts. 337 75

When human serum is diluted and pre-incubated at 37 degrees C in low ionic strength buffer (LIS, u = 0.07; made iso-osmotic with dextrose), a spontaneous activation of complement (C) is observed as determined by C4 and C3 electrophoretic conversion. In this paper it is postulated that most species of glycosaminoglycans (GAG) restricted non-specific fluid phase complement consumption induced by LIS, an effect which conserved complement and thereby enhanced the subsequent residual serum C mediated hemolytic activity. The capacity of glycosaminoglycans, to have modulated the hemolytic activity at low ionic strength, depended on the charge of the GAG species tested. In general, the GAG regulatory effects may have been due to GAG mediated restriction of spontaneous non-specific fluid phase C1 autoactivation, and/or restriction of activated C1 activity. Such effects would result in the subsequent reduction of the spontaneous fluid phase C4 and C3 consumption. Although the precise mechanisms responsible for the effects were not identified, it is speculated that the potentiation of C1 inhibitor function and direct effects on C1 might be involved. Overall, the relative specific activities of the glycosaminoglycans, on a weight basis, in mediating the fluid phase C regulatory effect were heparin greater than dermatan sulfate greater than chondroitin-6-sulfate greater than chondroitin-4-sulfate greater than hyaluronic acid and keratan sulfate. When much higher concns of heparin (greater than or equal 0.2 micrograms/ml) were used, complement mediated lysis of EA was inhibited, probably due to the direct inhibition of C1, even C1 which may have bound to the sensitized erythrocytes (EA). Results similar to that of heparin were obtained using greater than 1 mg/ml of dermatan sulfate or dextran sulfate. In contrast, pre-incubation of human serum in LIS with high concns (up to 10 mg/ml) of hyaluronic acid or chondroitin-4-sulfate, which are much less charged, continued to result only in the restriction of hemolytically non-specific (fluid phase) C consumption, resulting in a higher residual complement hemolytic activity. A theory is developed that the binding of polyionic GAG to C1 and to C1 INH may provide a charged local environment which simulates a relatively higher ionic strength. Chemical degradation of hyaluronic acid or chondroitin-4 or -6 sulfate resulted in lowering of this C modulating effect, indicative of the importance of the structural integrity of these charged glycosaminoglycans.(ABSTRACT TRUNCATED AT 400 WORDS)
Mol Immunol 1986 Aug
PMID:Glycosaminoglycans enhance complement hemolytic efficiency: theoretical considerations for GAG-complement-saliva interactions. 379 30


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