申请人在人血管平滑肌细胞（VSMC）中偶然新发现并克隆了小GTP结合蛋白Rac1的新亚型Rac1b。Rac1b是Rac1的多了一个外显子3B的剪接变异体，与乳腺结肠及肺肿瘤的发生与转移关系密切。我们已成功自制了优质特异抗体。预实验发现Rac1b在在体大鼠主动脉中无表达、在培养的人主动脉及冠状动脉VSMC中有表达、在损伤后第4周及第12周的大鼠颈动脉中高表达、免疫组化显示高表达的Rac1b局限在新生内膜中、蛋白质下拉实验显示新生内膜中的Rac1b具有GTP非依赖性的永久活性，提示其表达与VSMC增生和/或迁移密切相关。下一步，本课题要完成其在新生内膜中表达增高的1-16周时间曲线；然后在最佳时间点抑制其表达后观察新生内膜的变化；解明其是调节VSMC增生还是迁移；寻找是什么生长/细胞因子使其表达上调；并探索其促进VSMC增生和/或迁移的信号通路。从而试图找出能阻止血管损伤后管腔再狭窄的新药靶。

We first discovered and cloned Rac1b, a new subtype of Rac1, in human aortic smooth muscle cells (VSMC) cDNA library. Rac1b is a splicing varient of Rac1 with an extra exon 3b and plays an important role in the pathogenesis, invasion, and metastases of the breast, colon and lung cancer. In our pilot studies, we expressed and purified recombinant Rac1b from Ecoli and successfully made its specific antibodies. As shown by Western blotting, Rac1b is expressed in cultured human aortic (Ao184) and human coronary artery (Co396) VSMC. Surprisingly, it is highly expressed in rat left carotid artery at 4 and 12 weeks after balloon injury, but it is not detectable in the right carotid artery and in the aorta. Immunohistochemistry shows the expressed Rac1b is limited to the balloon injury-induced neointima. In protein pull-down experiment, not like Rac1, Rac1b from the rat neointima lysate binds to its downstream PAK-1 (protein Cdc42/Rac1-activated kinase 1) in a GTP-independent manner, indicating Rac1b is constitutively active in vivo. These results strongly indicate Rac1b's up-regulation is closely related to VSMC proliferation and/or migration. Next, we plan to: 1) establish a 1-16 week full time course of Rac1b's up-regulation in rat neointima; 2) test whether blocking Rac1b's expression prior to or at the peak of the up-regulation could retard the neointima formation in vivo; 3) study whether Rac1b's mainly regulate VSMC's proliferation and/or migration; 4) search for the pathological stimulants for Rac1b's up-regulation in cultured VSMC; and 5) determine the signal transduction pathway of Rac1b in VSMC. Our study aims to investigate whether Rac1b is a new drug-discovery target for effectively blocking neointima formation and restenosis after coronary artery intervention injury.