关键词: 透明质酸, 肺成纤维细胞, 基质金属蛋白酶, CD44, 整合素β₁

Abstract: Objective Low molecular weight HA was added into fibroblasts collagen gels to investigate the effect of HA on induction of matrix metalloproteinases(MMPs) and contraction and degradation of collagen gel, to explore the role of CD44 and integrin β₁ in the above effects. Methods 1) Lung fibroblasts were obtained by performing the primary human fetal lung tissue culture. 2) Experiments were divided into five groups: anti-CD44-Ab group(C), anti-integrin β₁-Ab group(B), IgG1 control group(I), HA group(H) and double distilled H₂O control group(M). 3) Fibroblasts were pretreated for 30 minutes with concentration of 25 mg/L of antibodies specific for CD44 receptor, integrin β₁ or nonimmune IgG1. 4) Mixing collagen, 2.5 times DMEM, HA and cell suspension, the mixture was cast into 24-well plates. Gelation occurred within 30 minutes, after which gels were released from the well and floated in culture medium. 5) The areas of collagen gels were measured every 24 hours, the supernatants of culture media were collected after first 24 hours and the concentration of MMPs is measured using an enzyme-linked immunosorbent assay(ELISA), collection of collagen gel after 72 hours, spectrophotometry was used to measure hydroxyproline. 6) Immunocytochemistry was used to analyze the expression of CD44. Results 1) HA increased the contraction of collagen gel of fibroblasts, while C group and B group attenuated the gel contraction significantly(P=0.000). 2) The content of hydroxyproline in fibroblasts collagen gel reduced significantly in B group, I group and H group(P＜0.05). 3) C group, B group and M group secreted less MMP-9 than other two groups from fibroblasts(P＜0.05); while only C group and M group secreted less MMP-1 than other three groups(P＜0.05). 4) Immunocytochemistry showed HA with different MW increasing the expression of CD44 receptor of fibroblasts, among which HA of 200 000 had the highest effect. Conclusion Hyaluronan induced MMPs secretion from human fibroblasts when cast into collagen gel, augmented the contraction of collagen gels and promoted 基金项目: 北京市优秀人才(20071D0300500083)资助项目, Supported by Excellent Person Project of Beijing(20071D0300500083) * Corresponding author, E-mail: cyh-birm@263.net degradation of collagen. These effects of HA could be completely inhibited by adding anti-CD44 antibody and partially attenuated by adding anti-integrin β₁ antibody.

Key words: hyaluronan, lung fibroblasts, matrix metalloproteinases, CD44, Integrin-β₁