双牻牛儿基转移酶-I在糖尿病加速性动脉粥样硬化中的作用及机制研究

Diabetes can accelerate the progression of atherosclerosis (AS), and becomes the main cause of death and disability in patients with diabetes. Geranylgeranyl transferase-I (GGTase-I) is the key enzyme mediating protein (e.g. Rac1) isoprenylation, and gradually highlights its role in vascular remodeling. However, the effect of GGTase-I on the diabetes-accelerated AS has not been reported, needing further explore. Our previous studies found that in diabetic mice, the AS was accelerated, the vascular smooth muscle cells (VSMCs) proliferated and migrated, and the expression of GGTase-I was upregulated. Meanwhile, in hypertensive rats, with the increase of VSMCs proliferation and GGTase-I expression, the reactive oxygen species (ROS) and NADPH oxidase (Nox) increased significantly. It is suggested that GGTase-I may be involved in the proliferation and migration of VSMCs induced by oxidative stress, while the latter is one of the key steps in the development of AS. Therefore, our team will build a diabetes-accelerated AS model using the conditional knockout mice (knockdown the GGTase-I gene in VSMCs), and the experiments will be carried out at the animal, cellular, and molecular levels, to explore the hypothesis “Does GGTase-I affect the proliferation and migration of VSMCs through Rac1-Nox-ROS mediated oxidative stress, and then participate in the diabetes-accelerated AS?” The results of this study hope to provide new ideas for the prevention and treatment of diabetic macrovascular disease.

糖尿病可加速动脉粥样硬化（AS），是患者致死致残的主因。双牻牛儿基转移酶-I（GGTase-I）是蛋白（包括Rac1）异戊二烯化修饰的关键酶，在血管重构中的作用逐步显现，但对糖尿病加速性AS的影响未见涉及。我们既往在糖尿病小鼠中发现AS加速、血管平滑肌细胞（VSMCs）增殖迁移伴GGTase-I表达亢进；另外在高血压大鼠中发现VSMCs增殖、GGTase-I表达上调的同时，活性氧（ROS）和NADPH氧化酶（Nox）显著增加。提示GGTase-I可能参与氧化应激诱导的VSMCs增殖迁移。而后者是AS进展的关键环节之一。因此，本项目拟利用条件性基因敲除小鼠（敲除VSMCs中GGTase-I）构建糖尿病加速性AS模型，从动物、细胞、分子水平探索“GGTase-I是否通过Rac1-Nox-ROS介导的氧化应激，影响VSMCs增殖迁移，从而参与糖尿病加速性AS？”为糖尿病大血管病变防治提供新思路。

氧化损伤诱导的血管平滑肌细胞（VSMCs）增殖是糖尿病加速性动脉粥样硬化的主要特征之一。 双牻牛儿基转移酶I(GGTase-I)是介导翻译后修饰的重要酶，尤其是小GTP酶（Rac1）的双牻牛儿基化。我们先前的研究发现，GGTase-I在糖尿病加速性动脉粥样硬化中起着重要作用。然而，其确切作用很大程度上尚不清楚。在本研究中，使用CRISPR/Cas9系统构建小鼠条件敲除VSMCs中GGTase-I(Pggt1bΔ/Δ)小鼠。并通过链脲佐菌素注射和动脉粥样硬化饮食诱导小鼠糖尿病加速性动脉粥样硬化模型。结果发现GGTase-I基因敲除可在减轻糖尿病加速性动脉粥样硬化进程，同时在体外实验中抑制高糖诱导的VSMC增殖。此外，在糖尿病病程16周后，Pggt1bΔ/Δ小鼠的α平滑肌肌动蛋白(α-SMA)和硝基酪氨酸水平、RAC1活性、p47phox和NOXO1表达、磷酸化ERK1/2和磷酸化JNK含量均低于野生型小鼠。同时，在高糖(22.2mM)体外培养Pggt1bΔ/Δ的VSMCs中也发现相似变化。总之，GGTase-I基因敲除有效地阻断了糖尿病加速性动脉粥样硬化，这种保护作用可能与抑制VSMC增殖有关。其潜在机制可能包括干扰Rac1 双牻牛儿基化，抑制NADPH氧化酶胞浆调节亚基的组装，减少氧化损伤，降低ERK1/2和JNK磷酸化。