Arias E B, Cartee G D
Division of Kinesiology, University of Michigan, 401 Washtenaw Avenue, Ann Arbor, MI 48109, USA.
Acta Physiol Scand. 2005 Mar;183(3):281-9. doi: 10.1111/j.1365-201X.2004.01403.x.
Protein O-linked glycosylation is regulated in vivo by the concentration of hexosamine substrates. Calorie restriction (60% of ad libitum intake) for 20 days causes decreased UDP-N-acetylhexosamine levels and increased insulin-mediated glucose transport in rat skeletal muscle. Conversely, prolonged incubation (19 h) of muscle with O-(2-acetamido-2-deoxy-D-glucopyranosylidene)amino-N-phenyl-carbamate (PUGNAc; an inhibitor of N-acetyl-beta-D-glucosaminidase) is characterized by increased O-linked glycosylation and insulin resistance. We aimed to determine the calorie restriction effect on O-linked glycosylation and characterize the temporal relationship between PUGNAc-induced O-linked glycosylation and insulin resistance.
A calorie restriction protocol characterized by decreased muscle hexosamine levels will result in a global reduction in O-linked glycosylated proteins in muscle, and PUGNAc-induced insulin resistance will coincide with increased O-linked glycosylation.
Plantaris muscle and liver from rats (ad libitum or calorie restricted) were analysed for O-linked glycosylation using two antibodies against different O-linked N-acetylglucosamine epitopes. Also, rat epitrochlearis muscles were incubated for 8.5 h +/- 100 mum PUGNAc prior to measurement of [(3)H]-3-O-methylglucose transport and O-linked glycosylation.
Calorie restriction did not alter protein O-linked glycosylated levels in muscle or liver. Incubation with PUGNAc for 8.5 h resulted in increased in O-linked glycosylation but unaltered basal or insulin-stimulated glucose transport.
The delay between O-linked glycosylation and insulin resistance in muscle incubated with PUGNAc suggests an indirect, relatively slow mechanism for insulin resistance. The effect of calorie restriction on insulin action in muscle is unlikely to be the direct result of a global change in protein O-linked glycosylation.
蛋白质O-连接糖基化在体内受己糖胺底物浓度的调节。对大鼠进行20天的热量限制（自由摄入量的60%）会导致UDP-N-乙酰己糖胺水平降低，并使大鼠骨骼肌中胰岛素介导的葡萄糖转运增加。相反，用O-（2-乙酰氨基-2-脱氧-D-吡喃葡萄糖亚基）氨基-N-苯基氨基甲酸酯（PUGNAc；N-乙酰-β-D-氨基葡萄糖苷酶抑制剂）对肌肉进行长时间孵育（19小时）的特征是O-连接糖基化增加和胰岛素抵抗。我们旨在确定热量限制对O-连接糖基化的影响，并描述PUGNAc诱导的O-连接糖基化与胰岛素抵抗之间的时间关系。
以肌肉己糖胺水平降低为特征的热量限制方案将导致肌肉中O-连接糖基化蛋白的整体减少，且PUGNAc诱导的胰岛素抵抗将与O-连接糖基化增加同时出现。
使用两种针对不同O-连接N-乙酰葡糖胺表位的抗体，分析大鼠（自由摄食或热量限制）的比目鱼肌和肝脏的O-连接糖基化情况。此外，在测量[³H]-3-O-甲基葡萄糖转运和O-连接糖基化之前，将大鼠肱三头肌与100μM PUGNAc孵育8.5小时。
热量限制并未改变肌肉或肝脏中蛋白质O-连接糖基化水平。用PUGNAc孵育8.5小时导致O-连接糖基化增加，但基础或胰岛素刺激的葡萄糖转运未改变。
用PUGNAc孵育的肌肉中，O-连接糖基化与胰岛素抵抗之间的延迟表明胰岛素抵抗存在间接且相对缓慢的机制。热量限制对肌肉胰岛素作用的影响不太可能是蛋白质O-连接糖基化整体变化的直接结果。
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