Analysis of fibrinolytic activation mechanism by staphylokinase in gene deficient mice

基因缺陷小鼠葡萄激酶纤溶激活机制分析

• 批准号: 09670061
• 负责人: OKADA Kiyotaka
• 金额: $ 1.98万
• 依托单位: Kinki University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 1998
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09670061/
• 关键词: staphylokinase; plasminogen; plasminogen activator; alpha2-antiplasmin; gene deficient mice; α_2ーアンチプラスミン

Staphylokinase (SAK) expresses plasminogen activator (PA) activity by forming a complex with plasminogen (Pig) or plasmin. Although this PA activity is inhibited by alpha2-antiplasmin (alpha2-AP), its activity is protected against the inhibition by alpha2-AP in the presnce of fibrin. The interaction between SAK and Pig were investigated using mutant SAKs, synthetic SAK peptides, purified pigs of several animals and gene deficient mice plasma.1). The interaction between mutant SAKs and pig.The deleted mutants of NH2-terminal amino acid region of SAK contracted by lacking the first 14, 18 and 25 amino acid region ( 14 SAK, 18 SAK and 25 SAK). The11-14 amino acid region of SAK was necessary to express PA activity after binding with Glu-plg.2). The interaction between synthetic SAK peptides and pig.The SAK peptides synthesized peptides consisted 10-30 amino acid in SAK amino acid sequence. The synthetic SAK peptides did not show PA activity. However, the synthetic nonadecapeptide (Glu22-Leu4O) of SAK bound to Glupig pig and enhanced pig activation by native SAK.3). The alpha2-AP and Pig gene deficient mice.The alpha2-AP gene deficient mice (alpha2-AP-/-) obtained by cloning and targeting of murine alpha2-AP gene. The pig gene deficient mice (Pig-/-)established breeding method and assay methods of pig gene in mice.4). The interaction between SAK and purified pigs in buffer and in alpha2-AP or Pig deficient mice plasma.The pigs purified from plasma of human, mice, rabbit or bovine. The interaction between SAK and purified pigs investigated in buffer and in alpha2-AP-/- or Pig-/- mice plasma. These findings indicate that (1) murine or bovine plg dose not react with SAK, (2) SAK-human piasmin complex activates murine or bovine plg, and (3) this murine plg activation is inhibited by murine alpha2-AP.

葡激酶(SAK)通过与纤溶酶原(Pig)或纤溶酶形成复合体来表达纤溶酶原激活物(PA)活性。虽然这种PA活性被α2-抗纤溶酶(α2-AP)抑制，但在纤维蛋白存在的情况下，它的活性被α2-AP所保护。用突变的SAK、合成的SAK多肽、几种动物的纯化猪和基因缺陷小鼠血浆研究了SAK与猪的相互作用。突变体SAKS与猪的相互作用：缺失SAK的前14、18和25个氨基酸区(14SAK、18SAK和25SAK)的缺失突变体。与Glu-plg2结合后，SAK的11-14个氨基酸区域是表达PA活性所必需的。合成的SAK多肽与猪的相互作用，合成的SAK多肽的氨基酸序列由10-30个氨基酸组成。合成的SAK多肽不具有PA活性。然而，人工合成的SAK非脂肽(Glu22-Leu4O)与Glupig猪结合，并增强天然SAK对猪的激活作用。α2-AP和猪基因缺陷小鼠。通过克隆和靶向小鼠α2-AP基因获得了α2-AP基因缺陷小鼠(α2-AP-/-)。猪基因缺陷小鼠(Pig-/-)建立了猪基因在小鼠中的育种方法和检测方法。SAK与纯化猪在缓冲液和α2-AP或猪缺陷小鼠血浆中的相互作用。在缓冲液和α2-AP-/-或Pig-/-小鼠血浆中研究了SAK与纯化猪的相互作用。这些结果表明：(1)小鼠或牛的PLG不与SAK反应，(2)SAK-人piasmin复合体激活小鼠或牛的PLG，(3)小鼠的这种激活被小鼠的α2-AP抑制。

项目成果

H.R.Lijnen, K.Okada, O.Matsuo, D.Collen, M.Dewerchin.: "alpha2-Antiplasmin gene-deficiency in mice is associated with enhanced fibrinolytic potential without overt bleeding." Blood. (in press). (1999)

H.R.Lijnen、K.Okada、O.Matsuo、D.Collen、M.Dewerchin.：“小鼠体内 alpha2-抗纤溶酶基因缺陷与纤维蛋白溶解潜力增强相关，但无明显出血。”