高分子量激肽原富含组氨酸区域抑制细胞伸展的机制分析

Inhibition of Cell Spreading by Recombinant Histidine-rich Domain of High Molecular Weight Kininogen and Its Mechanism

活化型高分子量激肽原 (activehighmolecularweightkininogen ,HKa)是组织培养板上体外连接蛋白 (vitronectin ,VN)促使细胞伸展的潜在抑制物 ,已证实轻链的富含组氨酸区域 (histidine richdomain ,HRD)是HKa抗细胞伸展的活性区域 .HK的重组HRD (r HRD)能够促使成纤维细胞伸展 .通过基于HRD序列的选择肽分析 ,定位了HRD的细胞伸展序列 .5个肽中的 3个能够使TIG 3细胞伸展 .P 1肽引起的细胞伸展能够被可溶性P 5肽或HKa所抑制 .P 2肽不能抑制P 1或P 5肽引起的细胞伸展 .r HRD以及 3种肽介导的细胞伸展能够被RGD合成肽以及抗αvβ3或α5β1整合素抗体所抑制 .结果提示 ,选择肽引起的细胞伸展是由整合素介导的 。

Active high molecular weight kininogen (HKa) is a potent inhibitor of cell spreading promoted by vitronectin on tissue culture plastic, and the effect requires interaction of the histidine-rich domain (HRD) with vitronectin substratum. Recombinant histidine-rich domain of HK (411-501) produced in bacteria (r-HRD), however, possessed cell spreading activity for human fibroblasts when adsorbed to ELISA plate. The cell adhesive sequence of HRD was localized by analysis of peptides based on the sequence of the HRD. Three of the five peptides (P-1, GKEQGHTRRHDWGHEKQRK, residues 402-420; P-2, HNLGHGHKHERDQGHGHQR, residues 421-439; and P-5, HKHGHGHGKHKNKGKKNGKH, residues 479-498) could promote the spreading of human fibroblasts. Cell spreading to P-1 was blocked by the soluble P-5 or HKa. P-2 did not inhibit cell spreading by P-1 or P-5. Cell spreading mediated by recombinant histidine-rich domain and all the three peptides was blocked by RGDS peptide and antibodies to the α vβ 3 or α 5β 1 integrins. These data suggest that spreading of cells owing to sequences in the histidine-rich domain is mediated by integrins even though this region does not have a RGD-containing sequence.