Crook E D, Simmons S T, Daniels M, Singh L P
Department of Medicine, University of Mississippi Medical Center, Jackson 39216, USA.
J Investig Med. 2000 Nov;48(6):427-34.
The hexosamine biosynthesis pathway acts as a cellular glucose sensor and mediates many of the adverse effects of glucose. Increased flux through this pathway results in insulin resistance in rat fibroblasts and transgenic mice and upregulation of transforming growth factor beta (TGF-beta) transcriptional activity in rat kidney cells. The first and rate-limiting step in this pathway, which is responsible for the metabolism of glucose to glucosamine, is catalyzed by glutamine:fructose-6-phosphate amidotransferase (GFA).
Because of the known effects of hyperglycemia on mesangial cell (MC) function and growth factor regulation, we examined the regulation of GFA by glucose and TGF-beta in cultured SV40 rat MCs. GFA activity was assayed in cytosolic extracts of MCs using high-performance liquid chromatography.
Culturing in 10 and 25 mM of glucose for 24 hours resulted in 33.4% (P < 0.025) and 43.5% (P < 0.05) decreases in GFA activity when compared with cells cultured at 1 to 5 mM of glucose. The downregulation in GFA activity by high glucose (HG) required at least 6 hours in culture and persisted for several days. HG effects were not a result of osmolar changes or glucose-induced differences in glucose uptake. Like HG, treatment of MCs with TGF-beta (2 ng/mL) for 4 hours resulted in a 30% (P < 0.05) decrease in GFA activity in cells cultured at 1 mM glucose, but the effects of TGF-beta were not additive to those of HG. TGF-beta-mediated downregulation of GFA activity was inhibited by a TGF-beta-neutralizing antibody, but HG's effects were not. Insulin-like growth factor-1 (IGF-1) had similar effects as TGF-beta, but GFA activity was not regulated by angiotensin II.
GFA activity is downregulated by HG, TGF-beta, and IGF-1 in rat MCs. Downregulation of this cellular glucose sensor may be a protective mechanism against the harmful effects of excess glucose as seen in diabetes.
己糖胺生物合成途径作为一种细胞葡萄糖传感器，介导了葡萄糖的许多不良反应。该途径通量增加会导致大鼠成纤维细胞和转基因小鼠出现胰岛素抵抗，并使大鼠肾细胞中转化生长因子β（TGF-β）转录活性上调。此途径的第一步也是限速步骤，负责将葡萄糖代谢为葡糖胺，由谷氨酰胺：果糖-6-磷酸酰胺转移酶（GFA）催化。
鉴于高血糖对系膜细胞（MC）功能和生长因子调节的已知影响，我们研究了培养的SV40大鼠MC中葡萄糖和TGF-β对GFA的调节作用。使用高效液相色谱法测定MC胞质提取物中的GFA活性。
与在1至5 mM葡萄糖中培养的细胞相比，在10和25 mM葡萄糖中培养24小时导致GFA活性分别降低33.4%（P < 0.025）和43.5%（P < 0.05）。高糖（HG）对GFA活性的下调至少需要在培养6小时，且持续数天。HG的作用不是渗透压变化或葡萄糖诱导的葡萄糖摄取差异的结果。与HG一样，用TGF-β（2 ng/mL）处理MC 4小时会导致在1 mM葡萄糖中培养的细胞中GFA活性降低30%（P < 0.05），但TGF-β的作用与HG的作用无叠加性。TGF-β介导的GFA活性下调被TGF-β中和抗体抑制，但HG的作用未被抑制。胰岛素样生长因子-1（IGF-1）具有与TGF-β相似的作用，但GFA活性不受血管紧张素II调节。
在大鼠MC中，HG、TGF-β和IGF-1下调GFA活性。下调这种细胞葡萄糖传感器可能是一种针对糖尿病中过量葡萄糖有害影响的保护机制。
