Abstract：
BACKGROUND: Vascular smooth muscle cell (VSMC) proliferation is involved in many types of arterial diseases, including neointima hyperplasia, in which Ca2+ has been recognized as a key player. However, the physiological role of Ca2+ release via inositol 1,4,5-trisphosphate receptors (IP3Rs) from endoplasmic reticulum in regulating VSMC proliferation has not been well determined.
RESULTS: Both in vitro cell culture models and in vivo mouse models were generated to investigate the role of IP3Rs in regulating VSMC proliferation. Expression of all 3 IP3R subtypes was increased in cultured VSMCs upon platelet-derived growth factor-BB and FBS stimulation as well as in the left carotid artery undergoing intimal thickening after vascular occlusion. Genetic ablation of all 3 IP3R subtypes abolished endoplasmic reticulum Ca2+ release in cultured VSMCs, significantly reduced cell proliferation induced by platelet-derived growth factor-BB and FBS stimulation, and also decreased cell migration of VSMCs. Furthermore, smooth muscle-specific deletion of all IP3R subtypes in adult mice dramatically attenuated neointima formation induced by left carotid artery ligation, accompanied by significant decreases in cell proliferation and matrix metalloproteinase-9 expression in injured vessels. Mechanistically, IP3R-mediated Ca2+ release may activate cAMP response element-binding protein, a key player in controlling VSMC proliferation, via Ca2+/calmodulin-dependent protein kinase II and Akt. Loss of IP3Rs suppressed cAMP response element-binding protein phosphorylation at Ser133 in both cultured VSMCs and injured vessels, whereas application of Ca2+ permeable ionophore, ionomycin, can reverse cAMP response element-binding protein phosphorylation in IP3R triple knockout VSMCs.
CONCLUSIONS: Our results demonstrated an essential role of IP3R-mediated Ca2+ release from endoplasmic reticulum in regulating cAMP response element-binding protein activation, VSMC proliferation, and neointima formation in mouse arteries.
RESULTS: Both in vitro cell culture models and in vivo mouse models were generated to investigate the role of IP3Rs in regulating VSMC proliferation. Expression of all 3 IP3R subtypes was increased in cultured VSMCs upon platelet-derived growth factor-BB and FBS stimulation as well as in the left carotid artery undergoing intimal thickening after vascular occlusion. Genetic ablation of all 3 IP3R subtypes abolished endoplasmic reticulum Ca2+ release in cultured VSMCs, significantly reduced cell proliferation induced by platelet-derived growth factor-BB and FBS stimulation, and also decreased cell migration of VSMCs. Furthermore, smooth muscle-specific deletion of all IP3R subtypes in adult mice dramatically attenuated neointima formation induced by left carotid artery ligation, accompanied by significant decreases in cell proliferation and matrix metalloproteinase-9 expression in injured vessels. Mechanistically, IP3R-mediated Ca2+ release may activate cAMP response element-binding protein, a key player in controlling VSMC proliferation, via Ca2+/calmodulin-dependent protein kinase II and Akt. Loss of IP3Rs suppressed cAMP response element-binding protein phosphorylation at Ser133 in both cultured VSMCs and injured vessels, whereas application of Ca2+ permeable ionophore, ionomycin, can reverse cAMP response element-binding protein phosphorylation in IP3R triple knockout VSMCs.
CONCLUSIONS: Our results demonstrated an essential role of IP3R-mediated Ca2+ release from endoplasmic reticulum in regulating cAMP response element-binding protein activation, VSMC proliferation, and neointima formation in mouse arteries.
摘要：
背景：血管平滑肌细胞（VSMC）增殖与许多类型的动脉疾病有关，包括新内膜增生,其中Ca2+被认为是关键角色。然而,内质网通过肌醇1,4,5-三磷酸受体（IP3Rs）释放Ca2在调节VSMC增殖中的生理作用尚未得到很好的确定。
结果：建立了体外细胞培养模型和体内小鼠模型，以研究IP3R在调节VSMC增殖中的作用。在血小板衍生的生长因子-BB和FBS刺激后，培养的VSMC以及血管闭塞后经历内膜增厚的左颈动脉中，所有3种IP3R亚型的表达均增加。所有3种IP3R亚型的遗传消融消除了培养的VSMCs中的内质网Ca2释放，血小板源性生长因子-BB和FBS刺激诱导的细胞增殖显着降低，并降低了VSMC的细胞迁移。此外，成年小鼠中所有IP3R亚型的平滑肌特异性缺失可显着减弱左颈动脉结扎诱导的新内膜形成，伴随着损伤血管中细胞增殖和基质金属蛋白酶-9表达的显着降低。机械上,IP3R介导的Ca2+释放可激活cAMP反应元件结合蛋白,控制VSMC扩散的关键角色，通过Ca2+/钙调蛋白依赖性蛋白激酶II和Akt。IP3R的丢失抑制了培养的VSMC和损伤血管中Ser133的cAMP反应元件结合蛋白磷酸化，而Ca2+可渗透的离子载体的应用，离子霉素,可以逆转IP3R三重敲除VSMC中cAMP反应元件结合蛋白的磷酸化。
结论：我们的结果证明了IP3R介导的内质网释放Ca2+在调节cAMP反应元件结合蛋白激活中的重要作用，VSMC增殖，和小鼠动脉中的新内膜形成。
结果：建立了体外细胞培养模型和体内小鼠模型，以研究IP3R在调节VSMC增殖中的作用。在血小板衍生的生长因子-BB和FBS刺激后，培养的VSMC以及血管闭塞后经历内膜增厚的左颈动脉中，所有3种IP3R亚型的表达均增加。所有3种IP3R亚型的遗传消融消除了培养的VSMCs中的内质网Ca2释放，血小板源性生长因子-BB和FBS刺激诱导的细胞增殖显着降低，并降低了VSMC的细胞迁移。此外，成年小鼠中所有IP3R亚型的平滑肌特异性缺失可显着减弱左颈动脉结扎诱导的新内膜形成，伴随着损伤血管中细胞增殖和基质金属蛋白酶-9表达的显着降低。机械上,IP3R介导的Ca2+释放可激活cAMP反应元件结合蛋白,控制VSMC扩散的关键角色，通过Ca2+/钙调蛋白依赖性蛋白激酶II和Akt。IP3R的丢失抑制了培养的VSMC和损伤血管中Ser133的cAMP反应元件结合蛋白磷酸化，而Ca2+可渗透的离子载体的应用，离子霉素,可以逆转IP3R三重敲除VSMC中cAMP反应元件结合蛋白的磷酸化。
结论：我们的结果证明了IP3R介导的内质网释放Ca2+在调节cAMP反应元件结合蛋白激活中的重要作用，VSMC增殖，和小鼠动脉中的新内膜形成。
