Fukata Youko, Yu Xiao, Imachi Hitomi, Nishiuchi Takamasa, Lyu Jingya, Seo Kayoko, Takeuchi Akihiro, Iwama Hisakazu, Masugata Hisashi, Hoshikawa Hiroshi, Hosomi Naohisa, Iwasaki Yasumasa, Murao Koji
Department of Advanced Medicine, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe Miki-cho, Kita-gun, Kagawa, 761-0793, Japan.
Endocrine. 2014 Aug;46(3):644-50. doi: 10.1007/s12020-013-0134-5. Epub 2013 Dec 18.
High-density lipoprotein (HDL) mediates reverse cholesterol transport. In this process, the human homolog of the B class, type I scavenger receptor (SR-BI), CD36, and LIMPII analogous-1 (hSR-BI/CLA-1) facilitates the cellular uptake of cholesterol from HDL. In endothelial cells, HDL activates endothelial nitric oxide synthase (eNOS) via hSR-BI/CLA-1, and 17β-estradiol (E2) modulates nitric oxide (NO) synthesis. In this study, we elucidated the effect of E2 on hSR-BI/CLA-1 expression in human umbilical vein endothelial cells (HUVECs). HSR-BI/CLA-1 expression was examined by real-time PCR, western blot analysis and reporter gene assay in HUVECs incubated with E2. eNOS activity was assessed by detection of phosphorylation (Ser 1179) of eNOS. We investigated the effect of the constitutively active form or dominant negative form of protein kinase C on hSR-BI/CLA-1 promoter activity. Our results showed that E2 increased the endogenous expression of hSR-BI/CLA-1. E2 also enhanced the activity of the hSR-BI/CLA-1 promoter and the expression of its mRNA. However, bisindolylmaleimide I, an inhibitor of protein kinase C, blocked the stimulatory effect of E2 on hSR-BI/CLA-1 promoter activity. Moreover, constitutively active PKC increased the activity of the hSR-BI/CLA-1 promoter, and a dominant-negative mutant of PKC prevented E2 from stimulating promoter activity. In cells treated with E2, HDL stimulated the phosphorylation of serine 1179 of eNOS in HUVECs. These results suggested that E2 upregulates the expression of the endothelial hSR-BI/CLA-1 via the PKC pathway, which may be a novel mechanism of the anti-atherosclerotic potential of E2 in vascular endothelial cells.
高密度脂蛋白（HDL）介导逆向胆固醇转运。在此过程中，B类I型清道夫受体（SR-BI）的人类同源物、CD36和LIMPII类似物-1（hSR-BI/CLA-1）促进细胞从HDL摄取胆固醇。在内皮细胞中，HDL通过hSR-BI/CLA-1激活内皮型一氧化氮合酶（eNOS），而17β-雌二醇（E2）调节一氧化氮（NO）的合成。在本研究中，我们阐明了E2对人脐静脉内皮细胞（HUVECs）中hSR-BI/CLA-1表达的影响。在用E2孵育的HUVECs中，通过实时PCR、蛋白质印迹分析和报告基因测定法检测hSR-BI/CLA-1的表达。通过检测eNOS的磷酸化（Ser 1179）来评估eNOS活性。我们研究了蛋白激酶C的组成型活性形式或显性负性形式对hSR-BI/CLA-1启动子活性的影响。我们的结果表明，E2增加了hSR-BI/CLA-1的内源性表达。E2还增强了hSR-BI/CLA-1启动子的活性及其mRNA的表达。然而，蛋白激酶C抑制剂双吲哚基马来酰亚胺I阻断了E2对hSR-BI/CLA-1启动子活性的刺激作用。此外，组成型活性PKC增加了hSR-BI/CLA-1启动子的活性，并PKC的显性负性突变体阻止了E2刺激启动子活性。在用E2处理的细胞中，HDL刺激了HUVECs中eNOS丝氨酸1179的磷酸化。这些结果表明，E2通过PKC途径上调内皮hSR-BI/CLA-1的表达，这可能是E2在血管内皮细胞中抗动脉粥样硬化潜力的一种新机制。
