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Effects of high glucose on integrin activity and fibronectin matrix assembly by mesangial cells.

ABSTRACT: The filtration unit of the kidney is the glomerulus, a capillary network supported by mesangial cells and extracellular matrix (ECM). Glomerular function is compromised in diabetic nephropathy (DN) by uncontrolled buildup of ECM, especially type IV collagen, which progressively occludes the capillaries. Increased levels of the ECM protein fibronectin (FN) are also present; however, its role in DN is unknown. Mesangial cells cultured under high glucose conditions provide a model system for studying the effect of elevated glucose on deposition of FN and collagen IV. Imaging of mesangial cell cultures and analysis of detergent-insoluble matrix show that, under high glucose conditions, mesangial cells assembled significantly more FN matrix, independent of FN protein levels. High glucose conditions induced protein kinase C-dependent ?1 integrin activation, and FN assembly in normal glucose was increased by stimulation of integrin activity with Mn(2+). Collagen IV incorporation into the matrix was also increased under high glucose conditions and colocalized with FN fibrils. An inhibitor of FN matrix assembly prevented collagen IV deposition, demonstrating dependence of collagen IV on FN matrix. We conclude that high glucose induces FN assembly, which contributes to collagen IV accumulation. Enhanced assembly of FN might facilitate dysregulated ECM accumulation in DN.

SUBMITTER: Miller CG

PROVIDER: S-EPMC4142608 | biostudies-literature | 2014 Aug

REPOSITORIES: biostudies-literature

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Effects of high glucose on integrin activity and fibronectin matrix assembly by mesangial cells.

Miller Charles G CG Pozzi Ambra A Zent Roy R Schwarzbauer Jean E JE

Molecular biology of the cell 20140618 16

The filtration unit of the kidney is the glomerulus, a capillary network supported by mesangial cells and extracellular matrix (ECM). Glomerular function is compromised in diabetic nephropathy (DN) by uncontrolled buildup of ECM, especially type IV collagen, which progressively occludes the capillaries. Increased levels of the ECM protein fibronectin (FN) are also present; however, its role in DN is unknown. Mesangial cells cultured under high glucose conditions provide a model system for studyi ...[more]

PMID: 24943838

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Project description:Fibronectin matrix assembly is a cell-dependent process which is upregulated in tissues at various times during development and wound repair to support the functions of cell adhesion, migration, and differentiation. Previous studies have demonstrated that the alpha 5 beta 1 integrin and fibronectin's amino terminus and III-1 module are important in fibronectin polymerization. We have recently shown that fibronectin's III-1 module contains a conformationally sensitive binding site for fibronectin's amino terminus (Hocking, D.C., J. Sottile, and P.J. McKeown-Longo. 1994. J. Biol. Chem. 269: 19183-19191). The present study was undertaken to define the relationship between the alpha 5 beta 1 integrin and fibronectin polymerization. Solid phase binding assays using recombinant III-10 and III-1 modules of human plasma fibronectin indicated that the III-10 module contains a conformation-dependent binding site for the III-1 module of fibronectin. Unfolded III-10 could support the formation of a ternary complex containing both III-1 and the amino-terminal 70-kD fragment, suggesting that the III-1 module can support the simultaneous binding of III-10 and 70 kD. Both unfolded III-10 and unfolded III-1 could support fibronectin binding, but only III-10 could promote the formation of disulfide-bonded multimers of fibronectin in the absence of cells. III-10-dependent multimer formation was inhibited by both the anti-III-1 monoclonal antibody, 9D2, and amino-terminal fragments of fibronectin. A fragment of III-10, termed III-10/A, was able to block matrix assembly in fibroblast monolayers. Similar results were obtained using the III-10A/RGE fragment, in which the RGD site had been mutated to RGE, indicating that III-I0/A was blocking matrix assembly by a mechanism distinct from disruption of integrin binding. Texas red-conjugated recombinant III-1,2 localized to beta 1-containing sites of focal adhesions on cells plated on fibronectin or the III-9,10 modules of fibronectin. Monoclonal antibodies against the III-1 or the III-9,10 modules of fibronectin blocked binding of III-1,2 to cells without disrupting focal adhesions. These data suggest that a role of the alpha 5 beta 1 integrin in matrix assembly is to regulate a series of sequential self-interactions which result in the polymerization of fibronectin.

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Effects of high glucose on integrin activity and fibronectin matrix assembly by mesangial cells.

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Effects of high glucose on integrin activity and fibronectin matrix assembly by mesangial cells.

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