Gene therapy of atherosclerosis by regulation of actin binding protein gene.

血管平滑肌细胞从内侧向内膜的迁移在动脉粥样硬化的发生和发展中起重要作用。因此，我们有可能通过控制迁移来预防动脉粥样硬化的发生和发展。肌动蛋白结合蛋白与平滑肌细胞的迁移密切相关。例如，其中一种蛋白质，钙钙蛋白，据报道可以抑制平滑肌细胞的迁移，从而防止实验性动脉粥样硬化的进展。Profilin是主要的肌动蛋白结合蛋白之一，被怀疑在细胞运动中起重要作用。在这个项目中，我们评估了profilin是否可以作为基因治疗的靶点，通过调节平滑肌细胞的迁移来预防动脉粥样硬化的进展。首先，我们从人类骨骼肌细胞cDNA文库中克隆了一个人类profilin基因。我们将cDNA插入表达载体，转染到培养的血管平滑肌细胞中。最后，转染profilin基因后，迁移速度大大加快。但实验只在动态表达的条件下进行。该结果必须通过永久表达profilin基因的细胞来证实。此外，我们开发了一种新的动脉粥样硬化模型，用于本项目的体内实验。该模型具有独特的动脉粥样硬化斑块（不稳定斑块）。斑块的病理与临床上危险的人类动脉粥样硬化非常相似。该模型对于评估包括profilin基因治疗在内的各种干预措施的效果非常有用。

Migration of vascular smooth muscle cells from the medial to the intimal membrane is important in initiation and development of atherosclerosis. Therefore we can possibly prevent the initiation and development of atherosclerosis through a control of the migration. Actin-binding proteins are closely involved in migration of smooth muscle cells. For example one of the proteins, calponin, was reported to inhibit the migration of smooth muscle cells and consequently to prevent the progress of experimental atherosclerosis. Profilin is one of the major actin-binding proteins and is suspected to play an inportant role in cell movement. In this project, we assessed if profilin could be a target of gene therapy by which we can prevent the progress of atherosclerosis through regulation of the smooth muscle cell migration. First we cloned a human profilin gene from a cDNA library of human skeletal muscle cells. We inserted the cDNA into an expression vector and transfected it into cultured vascular smooth muscle cells. Finally migration was rather accelerated by the transfection of profilin gene. But the experiment was done only under a condition of trangent expression. The result must be confirmed by cells with permanent expression of profilin gene. Additionally, we developed a new model of atherosclerosis to use it in in vivo experiments of this project. The model has a unique atherosclerotic plaque (an unstable plaque). The pathology of the plaque is quite similar to that of clinically risky human atherosclerosis. The model is very useful to assess the effects of various interventions including the profilin gene therapy.