scholarly journals Characterization of a novel heparan sulfate proteoglycan found in the extracellular matrix of liver sinusoids and basement membranes.

1991 ◽  
Vol 113 (5) ◽  
pp. 1231-1241 ◽  
Author(s):  
C J Soroka ◽  
M G Farquhar
Keyword(s):  
Extracellular Matrix ◽  
Basement Membrane ◽  
Heparan Sulfate ◽  
Rat Liver ◽  
Core Protein ◽  
Heparan Sulfate Proteoglycan ◽  
Basement Membranes ◽  
Sulfate Proteoglycan ◽  
Sinusoidal Cell ◽  
Space Of Disse

A novel heparan sulfate proteoglycan (HSPG) present in the extracellular matrix of rat liver has been partially characterized. Proteoglycans were purified from a high salt extract of total microsomes from rat liver and found to consist predominantly (approximately 90%) of HSPG. A polyclonal antiserum raised against this fraction specifically recognized HSPG by immunoprecipitation and immunoblotting. The intact, fully glycosylated HSPG migrated as a broad smear (150-300 kD) by SDS-PAGE, but after deglycosylation with trifluoromethanesulfonic acid only a single approximately 40-kD band was seen. By immunocytochemistry this HSPG was localized in the perisinusoidal space of Disse associated with irregular clumps of basement membrane-like extracellular matrix material, some of which was closely associated with the hepatocyte sinusoidal cell surface. It was also localized in biosynthetic compartments (rough ER and Golgi cisternae) of hepatocytes, suggesting that this HSPG is synthesized and deposited in the space of Disse by the hepatocyte. The anti-liver HSPG IgG also stained basement membranes of hepatic blood vessels and bile ducts as well as those of kidney and several other organs (heart, pancreas, and intestine). An antibody that recognizes the basement membrane HSPG found in the rat glomerular basement membrane did not precipitate the 150-300-kD rat liver HSPG. We conclude that the liver sinusoidal space of Disse contains a novel population of HSPG that differs in its overall size, its distribution and in the size of its core protein from other HSPG (i.e., membrane-intercalated HSPG) previously described in rat liver. It also differs in its core protein size from HSPG purified from other extracellular matrix sources. This population of HSPG appears to be a member of the basement membrane HSPG family.

scholarly journals Matrix-associated heparan sulfate proteoglycan: core protein-specific monoclonal antibodies decorate the pericellular matrix of connective tissue cells and the stromal side of basement membranes.

1989 ◽  
Vol 109 (6) ◽  
pp. 3199-3211 ◽  
Author(s):  
A Heremans ◽  
B van der Schueren ◽  
B de Cock ◽  
M Paulsson ◽  
J J Cassiman ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Monoclonal Antibodies ◽  
Epithelial Cell ◽  
Heparan Sulfate ◽  
Core Protein ◽  
Structural Features ◽  
Heparan Sulfate Proteoglycan ◽  
Basement Membranes ◽  
Lung Fibroblasts ◽  
Sulfate Proteoglycan

Cultured human lung fibroblasts produce a large, nonhydrophobic heparan sulfate proteoglycan that accumulates in the extracellular matrix of the monolayer (Heremans, A., J. J. Cassiman, H. Van den Berghe, and G. David. 1988. J. Biol. Chem. 263: 4731-4739). A panel of four monoclonal antibodies, specific for four distinct epitopes on the 400-kD core protein of this extracellular matrix heparan sulfate proteoglycan, detects similar proteoglycans in human epithelial cell cultures. Immunohistochemistry of human tissues with the monoclonal antibodies reveals that these proteoglycans are concentrated at cell-matrix interfaces. Immunogold labeling of ultracryosections of human skin indicates that the proteoglycan epitopes are nonhomogeneously distributed over the width of the basement membrane. Immunochemical investigations and amino acid sequence analysis indicate that the proteoglycan from the fibroblast matrix shares several structural features with the large, low density heparan sulfate proteoglycan isolated from the Engelbreth-Holm-Swarm sarcoma. Thus, both epithelial cell sheets and individual mesenchymal cells accumulate a large heparan sulfate proteoglycan(s) at the interface with the interstitial matrix, where the proteoglycan may adopt a specific topological orientation with respect to this matrix.

scholarly journals Heparan sulfate proteoglycan is present in basement membrane as a double-tracked structure.

1989 ◽  
Vol 37 (5) ◽  
pp. 597-602 ◽  
Author(s):  
S Inoue ◽  
D Grant ◽  
C P Leblond
Keyword(s):  
Basement Membrane ◽  
Heparan Sulfate ◽  
Core Protein ◽  
Heparan Sulfate Proteoglycan ◽  
Basement Membranes ◽  
Sulfate Proteoglycan ◽  
Electron Microscopic ◽  
Gold Particles ◽  
Parallel Lines ◽  
Reichert's Membrane

Basement membranes contain 4.5-nm wide sets of two parallel lines, along which short prongs called "spikes" occur at regular intervals. The nature of this structure, referred to as "double tracks," was investigated in Lowicryl sections of mouse kidney and rat Reichert's membrane immunolabeled for basement membrane components using secondary antibodies conjugated to 5-nm gold particles. When the mouse glomerular basement membrane and rat Reichert's membrane were exposed to antibodies directed to the core protein of heparan sulfate proteoglycan, 95% or more of the gold particles were over double tracks, whereas after exposure of Reichert's membrane to antisera against laminin, collagen IV, or entactin, labeling of the double tracks remained at the random level. When heparan sulfate proteoglycan was incubated in Tris buffer, pH 7.4, at 35 degrees C for 1 hr, a precipitate resulted which, on electron microscopic examination, was found to consist of 5- to 6-nm wide sets of two parallel lines along which densities were observed. Immunolabeling confirmed the presence of the proteoglycan's core protein in the sets. Since double tracks were closely similar to this structure and were labeled with the same antibodies, they were likely to be also composed of heparan sulfate proteoglycan.

scholarly journals Heterogeneous distribution of a basement membrane heparan sulfate proteoglycan in rat tissues.

1987 ◽  
Vol 105 (4) ◽  
pp. 1901-1916 ◽  
Author(s):  
J R Couchman
Keyword(s):  
Basement Membrane ◽  
Heparan Sulfate ◽  
Core Protein ◽  
Buoyant Density ◽  
Heparan Sulfate Proteoglycan ◽  
Basement Membranes ◽  
Rat Tissues ◽  
Sulfate Proteoglycan ◽  
Heterogeneous Distribution

A heparan sulfate proteoglycan (HSPG) synthesized by murine parietal yolk sac (PYS-2) cells has been characterized and purified from culture supernatants. A monospecific polyclonal antiserum was raised against it which showed activity against the HSPG core protein and basement membrane specificity in immunohistochemical studies on frozen tissue sections from many rat organs. However, there was no reactivity with some basement membranes, notably those of several smooth muscle types and cardiac muscle. In addition, it was found that pancreatic acinar basement membranes also lacked the HSPG type recognized by this antiserum. Those basement membranes that lacked the HSPG strongly stained with antisera against laminin and type IV collagen. The striking distribution pattern is possibly indicative of multiple species of basement membrane HSPGs of which one type is recognized by this antiserum. Further evidence for multiple HSPGs was derived from the finding that skeletal neuromuscular junction and liver epithelia also did not contain this type of HSPG, though previous reports have indicated the presence of HSPGs at these sites. The PYS-2 HSPG was shown to be antigenically related to the large, low buoyant density HSPG from the murine Engelbreth-Holm swarm tumor. It was, however, confirmed that only a single population of antibodies was present in the serum. Despite the presence of similar epitopes on these two proteoglycans of different hydrodynamic properties, it was apparent that the PYS-2 HSPG represents a basement membrane proteoglycan of distinct properties reflected in its restricted distribution in vivo.

scholarly journals Association of malachite green-positive material with heparan sulfate proteoglycan double tracks in basement membrane of mouse kidney tubules.

1995 ◽  
Vol 43 (3) ◽  
pp. 293-297
Author(s):  
S Inoue
Keyword(s):  
Basement Membrane ◽  
Heparan Sulfate ◽  
Malachite Green ◽  
Heparan Sulfate Proteoglycan ◽  
Mouse Kidney ◽  
Basement Membranes ◽  
Kidney Cortex ◽  
Sulfate Proteoglycan ◽  
Kidney Tubules ◽  
Thin Sections

The presence of lipids in the basement membrane of the mouse kidney tubules was examined by histochemical staining with malachite green. Pieces of mouse kidney cortex were immersed in a fixative containing 3% glutaraldehyde and 0.1% malachite green in 0.067 M sodium cacodylate buffer, pH 6.8, for 18 hr at 4 degrees C. Control tissue was fixed in the same way except that no malachite green was added to the fixative. The tissue pieces were cryoprotected, frozen in Freon 22, and subjected to freeze-substitution in dry acetone containing 1% OsO4. Thin sections of Epon-embedded specimens were observed by electron microscopy at first without uranyl-lead counterstaining. The basement membrane of mouse kidney tubules was positively stained in a pattern composed of an irregular assembly of 5-8-nm wide strands. The nature of these malachite green-positive strands was further examined by counterstaining thin sections with uranyl-lead, and they were identified as 4.5-5-nm wide ribbon-like "double tracks" previously characterized as the form taken by heparan sulfate proteoglycan in basement membranes. It is concluded that lipids are present in the basement membrane of mouse kidney tubules in association with heparan sulfate proteoglycan.

scholarly journals Agrin Is a Major Heparan Sulfate Proteoglycan in the Human Glomerular Basement Membrane

1998 ◽  
Vol 46 (1) ◽  
pp. 19-27 ◽  
Author(s):  
Alexander J. Groffen ◽  
Markus A. Ruegg ◽  
Henri Dijkman ◽  
Thea J. van de Velden ◽  
Carin A. Buskens ◽  
...  
Keyword(s):  
Basement Membrane ◽  
Heparan Sulfate ◽  
Glomerular Basement Membrane ◽  
Heparan Sulfate Proteoglycan ◽  
Basement Membranes ◽  
Sulfate Proteoglycan ◽  
Linear Distribution ◽  
Cell Matrix ◽  
Sds Page ◽  
Strong Staining

Agrin is a heparan sulfate proteoglycan (HSPG) that is highly concentrated in the synaptic basal lamina at the neuromuscular junction (NMJ). Agrin-like immunoreactiv-ity is also detected outside the NMJ. Here we show that agrin is a major HSPG component of the human glomerular basement membrane (GBM). This is in addition to perlecan, a previously characterized HSPG of basement membranes. Antibodies against agrin and against an unidentified GBM HSPG produced a strong staining of the GBM and the NMJ, different from that observed with anti-perlecan antibodies. In addition, anti-agrin antisera recognized purified GBM HSPG and competed with an anti-GBM HSPG monoclonal antibody in ELISA. Furthermore, both antibodies recognized a molecule that migrated in SDS-PAGE as a smear and had a molecular mass of approximately 200–210 kD after deglycosylation. In immunoelectron microscopy, agrin showed a linear distribution along the GBM and was present throughout the width of the GBM. This was again different from perlecan, which was exclusively present on the endothelial side of the GBM and was distributed in a nonlinear manner. Quantitative ELISA showed that, compared with perlecan, the agrin-like GBM HSPG showed a sixfold higher molarity in crude glomerular extract. These results show that agrin is a major component of the GBM, indicating that it may play a role in renal ultrafiltration and cell matrix interaction.

scholarly journals Expression of Plasminogen Activator Pla ofYersinia pestis Enhances Bacterial Attachment to the Mammalian Extracellular Matrix

1998 ◽  
Vol 66 (12) ◽  
pp. 5755-5762 ◽  
Author(s):  
Kaarina Lähteenmäki ◽  
Ritva Virkola ◽  
Anne Sarén ◽  
Levente Emödy ◽  
Timo K. Korhonen
Keyword(s):  
Extracellular Matrix ◽  
Basement Membrane ◽  
Heparan Sulfate ◽  
Plasminogen Activator ◽  
Human Lung ◽  
Heparan Sulfate Proteoglycan ◽  
Bacterial Attachment ◽  
Extracellular Matrices ◽  
Type I ◽  
Sulfate Proteoglycan

ABSTRACT The effect of the plasminogen activator Pla of Yersinia pestis on the adhesiveness of bacteria to the mammalian extracellular matrix was determined. Y. pestis KIM D27 harbors the 9.5-kb plasmid pPCP1, encoding Pla and pesticin; the strain efficiently adhered to the reconstituted basement membrane preparation Matrigel, to the extracellular matrix prepared from human lung NCI-H292 epithelial cells, as well as to immobilized laminin. The isogenic strain Y. pestis KIM D34 lacking pPCP1 exhibited lower adhesiveness to both matrix preparations and to laminin. Both strains showed weak adherence to type I, IV, and V collagens as well as to human plasma and cellular fibronectin. The Pla-expressing recombinantEscherichia coli LE392(pC4006) exhibited specific adhesiveness to both extracellular matrix preparations as well as to laminin. The Pla-expressing strains showed a low-affinity adherence to another basement membrane component, heparan sulfate proteoglycan, but not to chondroitin sulfate proteoglycan. The degradation of radiolabeled laminin, heparan sulfate proteoglycan, or human lung extracellular matrix by the Pla-expressing recombinant E. coli required the presence of plasminogen, and degradation was inhibited by the plasmin inhibitors aprotinin and α2-antiplasmin. Our results indicate a function of Pla in enhancing bacterial adhesion to extracellular matrices. Y. pestis also exhibits a low level of Pla-independent adhesiveness to extracellular matrices.

scholarly journals Basement membrane-like matrix of teratocarcinoma-derived endodermal cells: presence of laminin and heparan sulfate in the matrix at points of attachment to cells.

1983 ◽  
Vol 31 (1) ◽  
pp. 35-45 ◽  
Author(s):  
I Leivo
Keyword(s):  
Cell Membrane ◽  
Basement Membrane ◽  
Heparan Sulfate ◽  
Type Iv Collagen ◽  
Heparan Sulfate Proteoglycan ◽  
Ruthenium Red ◽  
Basement Membranes ◽  
Sulfate Proteoglycan ◽  
Type Iv ◽  
The Matrix

Teratocarcinoma-derived endodermal PYS-2 cells are known to synthesize an extracellular matrix containing the basement membrane molecules laminin, type IV collagen, and heparan sulfate proteoglycan as major constituents (I. Leivo, K. Alitalo, L. Risteli, A. Vaheri, R. Timpl, J. Wartiovaara, Exp Cell Res 137:15-23, 1982). Immunoferritin techniques with specific antibodies were used in the present study to define the ultrastructural localization of the above constituents in the fibrillar network. Laminin was detected in matrix network adjacent to the basal cell membrane and in protruding matrix fibrils that connect the matrix to the cell membrane. Ruthenium red-stainable heparinase-sensitive 10- to 20-nm particles were often present at the junction of the attachment fibrils and the matrix network, or along the attachment fibrils. A corresponding distribution of ferritin label was observed for basement membrane heparan sulfate proteoglycan. Type IV collagen was found in the matrix network but not in the attachment fibrils. The results suggest that the PYS-2 cells are connected to their pericellular matrix by fibrils containing laminin associated with heparan sulfate-containing particles. These results may also have relevance for the attachment of epithelial cells to basement membranes.

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