Decreased Degradation of Collagen and Fibronectin following Exposure of Proximal Cells to Glucose

Abstract

Background/Aims: Thickening and reduplication of the tubular basement membrane has been reported as an early event in diabetic nephropathy. The aim of the work outlined here was to examine the effects and mechanisms involved in the modulation of renal proximal tubular type-IV collagen and fibronectin turnover by glucose. Methods: The effect of glucose on type-IV collagen and fibronectin generation was studied by exposure of primary cultures of human renal proximal tubular cells (HPTC) to elevated D-glucose concentrations. Subsequently the mechanism of modulation of fibronectin generation was examined in a polarised system utilising the porcine proximal tubular cell line LLC-PK1 grown on porous tissue culture inserts. Results: Incubation of confluent growth-arrested HPTC with 25 mM D-glucose led to the accumulation of both type-IV collagen and fibronectin. This increase was not dependent on new gene transcription for either protein. Exposure of HPTC to 25 mM D-glucose also led to the induction of tissue inhibitor of metalloproteinases (TIMP-1 and TIMP-2) and also gelatinase A. There was, however, a net decrease in overall gelatinolytic activity. Incubation of confluent monolayers of LLC-PK1 cells grown on tissue culture inserts with 25 mM D-glucose on either their apical or basolateral aspect led to fibronectin accumulation seen only in the basolateral compartment. Under these experimental conditions, we can demonstrate polyol pathway activation, and furthermore the increase in fibronectin concentration in response to glucose was inhibited by the aldose reductase inhibitor sorbinil. Fibronectin accumulation was also demonstrated following both apical and basolateral addition of 1 mM sorbitol, but not following the addition of 25 mM galactose to either aspect of the cells. Conclusions: These data demonstrate that the glucose-induced accumulation of type-IV collagen and fibronectin was associated with alterations in the degradative pathway of these matrix components. In addition fibronectin generation in response to glucose was non-polar in terms of application of glucose, but polar in terms of fibronectin accumulation. The mechanisms of glucose-induced modulation of fibronectin were mediated by polyol pathway activation, and more specifically related to the metabolism of sorbitol to fructose.

GLUT2 protein at the rat proximal tubule brush border membrane correlates with protein kinase C (PKC)-βl and plasma glucose concentration

“…Tubulointerstitial fibrosis in diabetes is characterised by accumulation of extracellular matrix proteins, including fibronectin and collagen, leading eventually to tubulointerstitial scarring [20] and loss of tubular cell function. Unlike the glomerulus, exposure of human proximal tubular cells to elevated glucose levels does not affect collagen or fibronectin gene expression, but does result in a change in their degradation and rate of turnover [3,21]. The tubulointerstitium comprises much of the kidney substance, and tubular hypertrophy is responsible for most of the renal .001 compared with non-fasted control animals using Kruskal-Wallis ANOVA; # p<0.05 compared with nonfasted animals using unpaired t test enlargement seen in diabetes [4].…”

“…These studies have provided convincing evidence that overproduction of GLUT1 and protein kinase C (PKC)-βI are linked to glucose-induced mesangial cell damage and extracellular matrix deposition [1,2]. Diabetes also increases renal size, mainly due to proximal tubular cell hypertrophy and thickening of the tubular cell basement membrane [3], leading to eventual tubular cell loss and declining renal function [4]; however, the underlying mechanisms affecting proximal tubular cell function and their relationship to glucose transport have not been studied in any detail.…”

The proximal tubule in the pathophysiology of the diabetic kidney

“…Moreover, high glucose concentrations can induce collagen gene transcription and secretion in vitro in murine cortical tubular cells (182). Exposure of primary cultures of human renal proximal tubular cells to high glucose induces cell growth and increases the amount of type IV collagen and fibronectin (66,111) due to a net decrease in gelatinolytic activity (109).…”

The pathogenesis of diabetic retinopathy: old concepts and new questions

“…The imbalance of thromboxane and prostacylin enhances platelet hyperactivity. 120 Adhesion proteins 121 that are cofactors in the aggregation of human platelets and mediating the adenosine diphosphate (ADP)-induced response of these cells are also increased significantly.…”