晚期糖基化终产物修饰的β_2微球蛋白对人关节滑膜细胞的病理生物学作用

The pathobiological effect of β_2-microglobulin modified with advanced glycation end products on human type B synovial cells

目的 探讨晚期糖基化终产物（ＡＧＥ）修饰的β２微球蛋白（ＡＧＥ－β２ｍ）对人关节滑膜细胞的病理生物学作用。方法 分离、培养正常人关节Ｂ型滑膜细胞，与ＡＧＥ－β２ｍ共同孵育，用ＥＬＩＳＡ法检测培养上清中单核／巨噬细胞趋化蛋白１（ＭＣＰ－１）和ＲＡＮＴＥＳ水平，放射免疫法检测透明质酸（ＨＡ）、层黏蛋白和Ⅲ型前胶原水平，Ｎｏｒｔｈｅｒｎ ｂｌｏｔ分析培养细胞 ＭＣＰ－１和 ＲＡＮＴＥＳ的 ｍＲＮＡ表达，Ｗｅｓｔｅｒｎ ｂｌｏｔ分析培养上清中 ＨＡ的相对分子质量。结果 ＡＧＥ－β２ｍ以剂量依赖的方式刺激关节滑膜细胞分泌ＭＣＰ－１和ＲＡＮＴＥＳ，同时上调ＭＣＰ－１和ＲＡＮＴＥＳｍＲＮＡ的表达。抗入ＡＧＥ受体（ＲＡＧＥ）ＩｇＧ和Ｂ型滑膜细胞预孵育能明显抑制ＡＧＥ－β２ｍ的上述作用。ＡＧＥ－β２ｍ增加滑膜细胞ＨＡ的分泌量，但在ＡＧＥ－β２ｍ刺激下生成的 ＨＡ的相对分子质量（３３ ０００）明显小于正常 ＨＡ（３７ ０００）。ＡＧＥ－β２ｍ对滑膜细胞层黏蛋白和Ⅲ型胶原分泌无明显影响。结论ＡＧＥ－β２ｍ通过ＲＡＧＥ介导的作用，刺激人类关节Ｂ型滑膜细胞产生ＭＣＰ－１和ＲＡＮＴＥＳ，井促进滑膜细胞分泌炎症性小分子ＨＡ。

Objective To investigate the pathobiological effect of AGE-β2m on human type B synovial cells. Methods The normal human type B synovial cells(HSCs) were isolated and cultured in vitro with AGE-β2m. The levels of monocyte chemoattractant protein-1(MCP-1) and RANTES in the supernatants were measured by enzyme-linked immunosorbent assay(ELISA) . Hyaluronan(HA), laminin and procollagen type Ⅲ levels were determined by radio immunology assay. The molecular weight of HA was assessed by Western blot. The expression of mRNA was analyzed by Northern blots. Results AGE-β2m stimulated HSCs to produce MCP-1 and RANTES and enhanced the expression of MCP-1 and RANTES mRNA with a time-and dose-dependent manner. AGE-β2m induced effect was inhibited by preincubation of synovial cells with anti-AGE receptor(RAGE) IgG. The 3. 7 fold increased HA level was found in the supernatants of cells treated with AGE-β2m compared with those treated with medium alone. The molecular weight of HA was 33. 0 KD in AGE-β2m treated supernatants but 37. 0 KD in non-treated controls. There was no statistical significance in laminin and procollagen type Ⅲ levels between the supernatants of AGE-βm-treated and non-treated cells. Conclusions AGE-β2m stimulates HSCs to produce MCP-l and RANTES by a pathway mediated by RAGE. AGE-β2m induces production of HA with low molecular weight. These pathobiological effect of AGE-β2m might contribute to the local inflammatory response seen in DRA.