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

血管平滑肌细胞从中膜向内膜的迁移在动脉粥样硬化的发生和发展中起着重要作用。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，据报道，其中一种蛋白质Calponin可以抑制平滑肌细胞的迁移，从而阻止实验性动脉粥样硬化的进展。Profilin是一种主要的肌动蛋白结合蛋白，被认为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，通过调控平滑肌细胞的迁移来防止动脉粥样硬化的进展。首先，我们从人骨骼肌细胞的cdna文库中克隆了一个人profilin基因。我们将该基因插入到表达载体中，并将其导入培养的血管平滑肌细胞。最后，Profilin基因的导入可明显加快细胞的迁移速度。但这项实验只在变态表情条件下进行。这一结果必须得到Profilin基因永久表达的细胞的证实。此外，我们开发了一种新的动脉粥样硬化模型，用于该项目的活体实验。该模型有一种独特的动脉粥样硬化斑块(一种不稳定的斑块)。斑块的病理与临床上有风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.