Glucosamine Modulates TNF-α-Induced ICAM-1 Expression and Function Through O-Linked and N-Linked Glycosylation in Human Retinal Pigment Epithelial Cells
Glucosamine Modulates TNF-α-Induced ICAM-1 Expression and Function Through O-Linked and N-Linked Glycosylation in Human Retinal Pigment Epithelial Cells
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DOI：
10.1167/iovs.11-9291
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发表时间：
2012-04-01
影响因子：
4.4
通讯作者：
Chen, Jiann-Torng
中科院分区：
文献类型：
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作者：
Chen, Ching-Long;Liang, Chang-Min;Chen, Jiann-Torng
目的。本研究旨在探讨葡萄糖胺（GlcN）对tnf α诱导的ARPE-19细胞间粘附分子1 （ICAM-1）表达的影响及其在ARPE-19细胞中的功能。我们用Western blotting定量了tnf - α诱导的ARPE-19细胞中ICAM-1的蛋白水平。在加入四氧嘧啶（O- linked n - acetylglucosamintransferase， OGT）抑制剂、O-（2-乙酰氨基-2-脱氧-d -glucopyranosylidene）氨基- n - phenyl氨基甲酸酯（PUGNAc） （N-acetylglucosaminidase, O- glcnacase [OGA]）抑制剂或OGT基因过表达后，比较GlcN对O-乙酰氨基葡萄糖转移酶（O- glcnacase）抑制剂后，对O-连接糖基化的影响，从而对ICAM-1表达的影响。Western blotting检测GlcN对ICAM-1 n -链糖基化的影响。采用NF-kappa B报告基因试验研究了o -链糖基化对核因子κ B （NF-kappa B）信号通路的影响。采用ICAM-1粘附实验检测GlcN对ICAM-1粘附活性的影响。在ARPE-19细胞中，GlcN、PUGNAc和OGT过表达抑制tnf - α诱导的ICAM-1表达和NF-kappa B活性。四氧嘧啶增加了tnf α诱导的ARPE-19细胞中ICAM-1的表达和NF-kappa B的活性。GlcN和tunicamycin降低了tnf - α诱导的ARPE-19细胞中ICAM-1的分子质量。蛋白酶体抑制剂MG-132抑制glcn诱导的tnf α诱导的ICAM-1分子质量的降低。GlcN还能减弱tnf - α诱导的icam的粘附活性。在ARPE-19细胞中，GlcN通过调节nf - κ B信号传导相关因子的o -连锁糖基化和减少tnf α诱导的ICAM-1的n -连锁糖基化来抑制ICAM-1的表达和功能。这些作用可能有助于gln介导的眼部抗炎作用。（中国眼科杂志，2012;53:2281-2291）DOI:10.1167/iovs.11-9291
PURPOSE. The purpose of this article was to investigate the effects of glucosamine (GlcN) on the TNF-alpha-induced expression of intercellular adhesion molecule 1 (ICAM-1) and the function of ICAM-1 in ARPE-19 cells in vitro.METHODS. We quantified protein levels of TNF-alpha-induced ICAM-1 in ARPE-19 cells with Western blotting. The effects of GlcN on O-linked glycosylation, and therefore on ICAM-1 expression, were compared after the addition of alloxan, an inhibitor of O-linked N-acetylglucosamine transferase (OGT), or O-(2-acetamido-2-deoxy-D-glucopyranosylidene) amino-N-phenylcarbamate (PUGNAc), an inhibitor of N-acetylglucosaminidase (O-GlcNAcase [OGA]), or after OGT gene overexpression. The effect of GlcN on the N-linked glycosylation of ICAM-1 was evaluated by the change in its molecular mass on Western blotting. The effect of O-linked glycosylation on the nuclear factor kappa B (NF-kappa B) signaling pathway was examined using an NF-kappa B reporter gene assay. The effect of GlcN on ICAM-1 adhesion activity was examined using an ICAM-1 adhesion assay.RESULTS. GlcN, PUGNAc, and OGT overexpression inhibited TNF-alpha-induced ICAM-1 expression and NF-kappa B activity in ARPE-19 cells. Alloxan increased ICAM-1 expression and NF-kappa B activity in TNF-alpha-induced ARPE-19 cells. GlcN and tunicamycin reduced the molecular mass of TNF-alpha-induced ICAM-1 in ARPE-19 cells. The proteasome inhibitor MG-132 suppressed the GlcN-induced reduction in the molecular mass of TNF-alpha-induced ICAM-1. GlcN also attenuated the adhesion activity of TNF-alpha-induced ICAM-1.CONCLUSIONS. GlcN inhibits ICAM-1 expression and functions by modulating the O-linked glycosylation of factors involved in NF-kappa B signaling and by reducing the N-linked glycosylation of TNF-alpha-induced ICAM-1 in ARPE-19 cells. These effects may contribute to the GlcN-mediated anti-inflammatory effects in the eye. (Invest Ophthalmol Vis Sci. 2012; 53: 2281-2291) DOI:10.1167/iovs.11-9291