A O Phillips, K Morrisey, R Steadman, J D Williams
{"title":"Decreased degradation of collagen and fibronectin following exposure of proximal cells to glucose.","authors":"A O Phillips, K Morrisey, R Steadman, J D Williams","doi":"10.1159/000020624","DOIUrl":null,"url":null,"abstract":"<p><strong>Background/aims: </strong>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.</p><p><strong>Methods: </strong>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.</p><p><strong>Results: </strong>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.</p><p><strong>Conclusions: </strong>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.</p>","PeriodicalId":12179,"journal":{"name":"Experimental nephrology","volume":"7 5-6","pages":"449-62"},"PeriodicalIF":0.0000,"publicationDate":"1999-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1159/000020624","citationCount":"22","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Experimental nephrology","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1159/000020624","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 22
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.
背景/目的:据报道,肾小管基底膜增厚和重复是糖尿病肾病的早期事件。本文概述的工作目的是研究葡萄糖对肾近端小管iv型胶原和纤维连接蛋白转换的调节作用和机制。方法:将人肾近端小管细胞(HPTC)原代培养物暴露于升高的d -葡萄糖浓度下,研究葡萄糖对iv型胶原和纤维连接蛋白生成的影响。随后,利用在多孔组织培养插入物上生长的猪近端管细胞系LLC-PK1,在极化系统中研究了纤维连接蛋白产生的调节机制。结果:融合生长阻滞HPTC与25 mM d -葡萄糖孵育导致iv型胶原和纤维连接蛋白的积累。这种增加并不依赖于两种蛋白的新基因转录。将HPTC暴露于25 mM d -葡萄糖中也会诱导组织金属蛋白酶抑制剂(TIMP-1和TIMP-2)和明胶酶a。然而,总体明胶溶解活性净下降。在组织培养插入物上培养的lc - pk1细胞的融合单层,在其顶端或基底外侧用25 mM d -葡萄糖孵育,导致仅在基底外侧室中可见纤维连接蛋白积累。在这些实验条件下,我们可以证明多元醇途径的激活,并且醛糖还原酶抑制剂山梨醇可以抑制葡萄糖反应中纤维连接蛋白浓度的增加。在细胞的顶端和底侧添加1mm山梨糖醇后也发现了纤维连接蛋白的积累,但在细胞的任何一侧添加25mm半乳糖后都没有。结论:这些数据表明,葡萄糖诱导的iv型胶原和纤维连接蛋白的积累与这些基质成分降解途径的改变有关。此外,就葡萄糖的应用而言,纤维连接蛋白的产生是非极性的,但就纤维连接蛋白的积累而言,却是极性的。葡萄糖诱导的纤维连接蛋白调节机制是通过多元醇途径激活介导的,更具体地说,与山梨醇对果糖的代谢有关。