Templeton D M, Fan M Y
Department of Clinical Biochemistry, University of Toronto, Ontario, Canada.
Metabolism. 1996 Sep;45(9):1136-46. doi: 10.1016/s0026-0495(96)90014-1.
Hyperglycemic conditions are known to increase mRNA and protein levels of several extracellular matrix molecules in cultured mesangial cells, but accompanying increases in proteoglycan mRNA have not been found, and there are discrepant reports of normal or decreased proteoglycan synthesis with or without undersulfation in diabetic kidneys and hyperglycemic cultures. We examined the effects in proliferating cells of glucose on [35S]Sulfate incorporation into heparan and dermatan sulfates and on mRNA levels of decorin, biglycan, and basement membrane perlecan. In both mesangial cells and vascular smooth muscle cells, 30 mmol/L glucose caused a decrease of 15% to 25% in the amount of sulfate incorporated into each proteoglycan in cultures confluent for 1 to 4 days, compared with 10 mmol/L glucose. The effect showed no specificity for the class of proteoglycan and was not a consequence of changes in total protein synthesis, which increased, or cell proliferation, which was unaffected. No decrease in charge density of any of the proteoglycan fractions was observed by ion-exchange chromatography. Therefore, the decrease in labeling was due to a decrease in synthesis and not undersulfation. mRNA levels for biglycan and perlecan increased slightly and transiently, and these changes cannot account for the decreased synthesis. Decorin mRNA was detected only in smooth muscle cells, where it and biglycan were differentially affected by glucose, apparently at the transcriptional level; stabilities of the two messages were unaffected by glucose. Although transforming growth factor-beta 1 (TGF-beta 1) mRNA levels increased in response to glucose, the cytokine did not appear to regulate proteoglycan synthesis, because structural changes in proteoglycans elicited by addition of TGF-beta 1 to the culture medium did not occur in the hyperglycemic cultures. On the other hand, inhibition and downregulation of protein kinase C (PKC), while decreasing net sulfate incorporation into mesangial cell proteoglycans, prevented the effect of high glucose. We conclude that a high glucose concentration causes a general decrease in the synthesis of all classes of proteoglycans at a posttranscriptional level, and can do so without affecting the charge density of individual proteoglycan molecules.
已知高血糖状况会增加培养的系膜细胞中几种细胞外基质分子的mRNA和蛋白质水平，但未发现蛋白聚糖mRNA随之增加，并且关于糖尿病肾脏和高血糖培养物中蛋白聚糖合成正常或减少以及是否存在硫酸化不足存在相互矛盾的报道。我们研究了葡萄糖对增殖细胞中[35S]硫酸盐掺入硫酸乙酰肝素和硫酸皮肤素以及对核心蛋白聚糖、双糖链蛋白聚糖和基底膜多配体蛋白聚糖mRNA水平的影响。在系膜细胞和血管平滑肌细胞中，与10 mmol/L葡萄糖相比，30 mmol/L葡萄糖使培养1至4天的汇合细胞中掺入每种蛋白聚糖的硫酸盐量减少了15%至25%。该效应对蛋白聚糖类别无特异性，也不是总蛋白合成增加（总蛋白合成增加）或细胞增殖不受影响所导致的结果。通过离子交换色谱未观察到任何蛋白聚糖组分的电荷密度降低。因此，标记减少是由于合成减少而非硫酸化不足。双糖链蛋白聚糖和多配体蛋白聚糖的mRNA水平略有短暂升高，这些变化无法解释合成减少的现象。核心蛋白聚糖mRNA仅在平滑肌细胞中检测到，在平滑肌细胞中它和双糖链蛋白聚糖受葡萄糖的影响不同，显然是在转录水平；两种信使RNA的稳定性不受葡萄糖影响。尽管转化生长因子-β1（TGF-β1）的mRNA水平因葡萄糖而升高，但该细胞因子似乎并未调节蛋白聚糖的合成，因为在高血糖培养物中，向培养基中添加TGF-β1并未引起蛋白聚糖的结构变化。另一方面，蛋白激酶C（PKC）的抑制和下调虽然减少了系膜细胞蛋白聚糖中硫酸盐的净掺入量，但阻止了高葡萄糖的作用。我们得出结论，高葡萄糖浓度在转录后水平导致所有类别蛋白聚糖的合成普遍减少，并且可以在不影响单个蛋白聚糖分子电荷密度的情况下做到这一点。
