Dept. of Medicine, Nephrology Division, Univ. of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390-8856, USA.
Am J Physiol Endocrinol Metab. 2010 Feb;298(2):E210-21. doi: 10.1152/ajpendo.00232.2009. Epub 2009 Nov 10.
Cells exposed to high glucose may undergo hypertrophy, proliferation, and apoptosis, but the role of hexosamine flux in mediating these effects has not been fully elucidated. Accordingly, we studied the effects of glucose and glucosamine on rat glomerular mesangial cells (MC) turnover. Compared with physiological glucose (5.6 mM), treatment with high glucose (25 mM) for 24 h stimulated MC proliferation, an effect that was mimicked by exposure to low concentrations of glucosamine (0.05 mM). The percentage of cells in G(0)/G(1) phase of the cell cycle was reduced with a concomitant increase of the number of cells in G(2)/M phase. Proliferating cell nuclear antigen, phosphorylated mammalian target of rapamycin [phospho-mTOR (Ser(2448))], and total regulatory-associated protein of mTOR were increased by high glucose and glucosamine treatment. Inhibition of glutamine:fructose-6-phosphate amidotransferase (GFAT), the rate-limiting enzyme for hexosamine flux, with 6-diazo-5-oxonorleucine (10 muM) and of mTOR with rapamycin both attenuated glucose-mediated MC proliferation. Higher glucosamine concentrations (0.25-10 mM) caused MC apoptosis after 48 h, and, in addition, GFAT overexpression also increased MC apoptosis (TdT-dUTP nick end-labeling-positive cells: 3.8 +/- 0.3 vs. 1.1 +/- 0.2% for empty vector; P < 0.05). Hence, hexosamine flux is an important determinant of MC proliferation and apoptosis. The proliferative response to high glucose and hexosamine flux is rapamycin-sensitive, suggesting that this effect is associated with signaling through rapamycin-sensitive mTOR complex 1 (mTORC1).
细胞暴露在高浓度葡萄糖中可能会经历肥大、增殖和凋亡，但己糖胺通量在介导这些效应中的作用尚未完全阐明。因此，我们研究了葡萄糖和氨基葡萄糖对大鼠肾小球系膜细胞（MC）更新的影响。与生理浓度葡萄糖（5.6 mM）相比，25 mM 高浓度葡萄糖处理 24 小时可刺激 MC 增殖，而暴露于低浓度氨基葡萄糖（0.05 mM）可模拟该作用。细胞周期 G0/G1 期的细胞比例减少，同时 G2/M 期的细胞数量增加。高浓度葡萄糖和氨基葡萄糖处理增加了增殖细胞核抗原、磷酸化哺乳动物雷帕霉素靶蛋白[磷酸化 mTOR（Ser2448）]和 mTOR 的调节相关蛋白的总量。用 6-二氮-5-氧正亮氨酸（10 μM）抑制己糖胺通量的限速酶谷氨酰胺：果糖-6-磷酸酰胺转移酶（GFAT）和用雷帕霉素抑制 mTOR，均可减弱葡萄糖介导的 MC 增殖。较高浓度的氨基葡萄糖（0.25-10 mM）在 48 小时后引起 MC 凋亡，并且 GFAT 过表达也增加了 MC 凋亡（TdT-dUTP 缺口末端标记阳性细胞：3.8 +/- 0.3 比空载体对照 1.1 +/- 0.2%；P < 0.05）。因此，己糖胺通量是 MC 增殖和凋亡的重要决定因素。对高浓度葡萄糖和己糖胺通量的增殖反应是雷帕霉素敏感的，这表明这种效应与通过雷帕霉素敏感的 mTOR 复合物 1（mTORC1）的信号转导有关。
