Integrin overexpression induced by high glucose and by human diabetes: potential pathway to cell dysfunction in diabetic microangiopathy.

Abstract

The nature of the process leading to the acellular nonperfused capillaries of diabetic microangiopathy remains unknown. Because these capillaries manifest thickened basement membranes, we asked whether the process causing deposition of excess extracellular matrix in diabetes modifies cell-matrix interactions in a direction that would compromise cell renewal. In 44 individual isolates of human umbilical vein endothelial cells we observed that high glucose concentrations (30 mM) induce coordinate increases in the levels of mRNAs encoding fibronectin and the fibronectinspecific integrin receptor as5fl as well as in the cognate proteins. Expression of the integrin subunit a3, component of the a3f81 polyspecific receptor for fibronectin, laminin, and collagen, was also up-regulated by high glucose. Overexpression of integrins correlated with increased cell attachment to exogenous fibronectin and laminin as well as to complex matrix. Moreover, cells exhibited firmer steady-state adhesion to their own matrix. To correlate these in vitro observations with events in human diabetic retinopathy we measured integrin levels in retinal trypsin digests prepared from 10 patients with 8.2 ± 1.6 (mean ± SE) years of diabetes and 10 age-and sex-matched nondiabetic controls. Microvessels of diabetic patients showed increased immunostaining for B13 integrin (P = 0.025) when compared with control microvessels. These data show that high glucose and diabetes increase integrin expression and thus alter the interaction of vascular endothelial cells with their basement membranes in the direction of firmer cell-matrix adhesion. This could compromise the migration and replication critical to the reendothelialization process and contribute to microvascular occlusion.The most characteristic chronic sequelae of diabetes mellitus (retinopathy leading to blindness, nephropathy leading to renal failure) are local manifestations of a widespread occlusive microangiopathy (1-3). Histologically the nonperfused capillaries are acellular and surrounded by thickened basement membranes, which represent the identifying feature of diabetic microangiopathy (4). The nature of the process leading to accelerated cellular demise and whether it relates to the accumulation of basement membrane material have remained outstanding issues (5).Among the modulatory influences exerted by the extracellular matrix on endothelial cells, some are specifically related to the amount of matrix. It has been shown that increasing concentrations of fibronectin and, to a lesser extent, of laminin and collagen IV reduce endothelial cell migration and replication (6). Because these functions are critical to the reendothelialization process (6), it could be hypothesized that diabetic vessels become occluded because excess matrixThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact. compromises prompt and efficient reendothe...

Cytoskeletal changes induced by excess extracellular matrix impair endothelial cell replication

“…*p < 0.05 vs control cells plated on plastic and not receiving cytochalasin treatment models. Endothelial cells cultured in the presence of high glucose concentrations upregulate the synthesis of several basement membrane components and relevant integrins, adhere more firmly to their matrix and show decreased replication [12,13]. In this study the ECM produced under high glucose conditions proved sufficient to reduce the replication of endothelial cells, in the absence of high ambient glucose.…”

“…Integrin function as measured in an attachment assay was increased in cells grown on both 1 and 10 m g/ml fibronectin (144 ± 30 and 147 ± 37 % of control cells grown on plastic, respectively, p < 0.01). There was also a modest upregulation of the expression of the integrin subunits [13].…”

“…For the attachment assay, cells were released from the dishes by trypsinization and plated in 96-well microtitre plates coated overnight with fibronectin at 2.5 m g/ml and postcoated with bovine serum albumin (BSA) to block nonspecific binding. Attachment was tested after 30 min at 37°C as previously described [13]. Cultures companion to those providing cells for the attachment assay were simultaneously harvested for RNA extraction.…”

A study of capillary pericyte viability on extracellular matrix produced by endothelial cells in high glucose

“…Impaired replication of endothelium on ECM obtained from other endothelial cells previously grown in high glucose concentrations was described previously [7,24] and a role for excess exogenous fibronectin, altering cell-ECM interactions through increased adhesion and cytoskeletal rearrangements, was hypothesized [5]. High glucose was shown to increase type IV collagen and fibronectin mRNA synthesis [25] and to upregulate the trascription of their genes [7] in cultured human endothelial cells.…”

The effects of glucose-induced oxidative stress on growth and extracellular matrix gene expression of vascular smooth muscle cells

“…There are numerous reports of increased extracellular matrix proteins (both mRNA expression and protein concentrations) at variable time points and glucose concentrations in differing cell types. For example, in human endothelial cells, mRNA expression of both fibronectin and collagen IV were increased (by approximately 150 %) after 17 days exposure to 30 mmol/l glucose [52], whereas others have shown similar changes in endothelial cells after only 5 days in 30 mmol/l glucose [53]. In human mesangial cells cultured in 30 mmol/l D-glucose, increased mRNA expression of fibronectin, laminin and collagen IV were not apparent at 7 days but were increased at 14 days [54].…”