Overexpression of glyoxalase-I in bovine endothelial cells inhibits intracellular advanced glycation endproduct formation and prevents hyperglycemia-induced increases in macromolecular endocytosis
Overexpression of glyoxalase-I in bovine endothelial cells inhibits intracellular advanced glycation endproduct formation and prevents hyperglycemia-induced increases in macromolecular endocytosis
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DOI：
10.1172/jci119885
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发表时间：
1998-03-01
影响因子：
15.9
通讯作者：
Brownlee, M
中科院分区：
文献类型：
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作者：
Shinohara, M;Thornalley, PJ;Brownlee, M
丙酮醛（MG）是磷酸丙糖裂解产生的二羰基化合物，在体外可形成晚期糖基化终产物（AGEs）。谷胱甘肽酶-I催化MG转化为S-D-乳酰谷胱甘肽，后者又通过谷胱甘肽酶-II转化为D-乳酸。为了直接评价甘氨酰转移酶-I活性对细胞内AGE形成的影响，产生了稳定表达人甘氨酰转移酶-I的GM 7373内皮细胞，与新转染的对照细胞相比，这些细胞中的甘氨酰转移酶-I活性增加了28倍（21.80 +/- 0.1 vs.0.76 +/- 0.02 μ mol/min/mg蛋白，n = 3，P < 0.001）。在新转染的细胞中，30 mM葡萄糖孵育使MG和D-乳酸盐浓度在5 mM以上增加约2倍（分别为35.5 +/- 5.8 vs 19.6 +/- 1.6，P < 0.02，n = 3和21.0 +/- 1.3 vs 10.0 +/- 1.2 pmol/10（6）细胞，n = 3，P < 0.001）。相反，在glycoprotease-I转染的细胞中，30 mM葡萄糖孵育根本不增加MG浓度，而使酶产物D-乳酸增加> 10倍（分别为18.9 +/- 3.2对18.4 +/- 5.8，n = 3，P = NS，和107.1 +/- 9.0对9.4 +/- 0 pmol/10（6）细胞，n = 3，P < 0.001）。在暴露于30 mM葡萄糖后，neo细胞内AGE形成增加13.6倍（2.58 +/- 0.15对0.19 +/- 0.03总吸光度单位，n = 3，P < 0.001），伴随细胞内AGEs增加，这些细胞的大分子内吞作用增加了2.2倍。过表达的glycosidase-I完全防止高血糖诱导的AGE形成和增加的大分子内吞作用。
Methylglyoxal (MG), a dicarbonyl compound produced by the fragmentation of triose phosphates, forms advanced glycation endproducts (AGEs) in vitro. Glyoxalase-I catalyzes the conversion of MG to S-D-lactoylglutathione, which in turn is converted to D-lactate by glyoxalase-II. To evaluate directly the effect of glyoxalase-I activity on intracellular AGE formation, GM7373 endothelial cells that stably express human glyoxalase-I were generated, Glyoxalase-I activity in these cells was increased 28-fold compared to neo-transfected control cells (21.80 +/- 0.1 vs. 0.76 +/- 0.02 mu mol/min/mg protein, n = 3, P < 0.001). In neo-transfected cells, 30 mM glucose incubation increased MG and D-lactate concentration approximately twofold above 5 mM (35.5 +/- 5.8 vs, 19.6 +/- 1.6, P < 0.02, n = 3, and 21.0 +/- 1.3 vs. 10.0 +/- 1.2 pmol/10(6) cells, n = 3, P < 0.001, respectively). in contrast, in glyoxalase-I-transfected cells, 30 mM glucose incubation did not increase MG concentration at all, while increasing the enzymatic product D-lactate by > 10-fold (18.9 +/- 3.2 vs. 18.4 +/- 5.8, n = 3, P = NS, and 107.1 +/- 9.0 vs. 9.4 +/- 0 pmol/10(6) cells, n = 3, P < 0.001, respectively), After exposure to 30 mM glucose, intracellular AGE formation in neo cells was increased 13.6-fold (2.58 +/- 0.15 vs. 0.19 +/- 0.03 total absorbance units, n = 3, P < 0.001), Concomitant with increased intracellular AGEs, macromolecular endocytosis by these cells was increased 2.2-fold. Overexpression of glyoxalase-I completely prevented both hyperglycemia-induced AGE formation and increased macromolecular endocytosis.