目的 探究大黄酸调控血管内皮细胞糖脂代谢功能的机制。方法 用棕榈酸干预人脐静脉内皮细胞（human umbilical vein endothelial cells，HUVECs），随后用2.5、5.0、10.0 μmol/L大黄酸处理24 h。qRT-PCR检测脂肪酸氧化（fatty acid oxidation，FAO）及糖酵解的关键酶[肉碱棕榈酰转移酶-1A（carnitine palmitoyltransferase-1A，CPT-1A）、葡萄糖转运蛋白1（glucose transporter protein 1，GLUT1）、6-磷酸果糖-2-激酶/果糖-2,6-双磷酸酶3（6-phosphofructose-2-kinase/fructose-2,6-bisphosphatase 3，PFKFB3）、己糖激酶2（hexokinase 2，HK2）]的表达情况；Seahorse XF96分析仪检测FAO和糖酵解水平；ELISA法检测一氧化氮（nitric oxide，NO）和血管内皮生长因子（vascular endothelial growth factor，VEGF）水平；Western blotting及免疫荧光观察一氧化氮合酶（endothelial nitric oxide synthase，eNOS）、VEGF、CPT-1A、PFKFB3表达情况；利用分子对接预测大黄酸的作用靶点。给予PPARα抑制剂norathyriol、FAO抑制剂etomoxir、糖酵解抑制剂3PO进行干预，观察大黄酸调节内皮细胞糖脂代谢功能的变化。结果 棕榈酸能显著上调HUVECs糖脂代谢酶CPT-1A、GLUT1、PFKFB3、HK2的表达（P＜0.05、0.01）；与模型组比较，10.0 μmol/L大黄酸显著上调CPT-1A的mRNA表达（P＜0.05），并显著下调GLUT1、PFKFB3、HK2的mRNA表达（P＜0.01）。代谢表型实验发现，大黄酸可以促进FAO并抑制糖酵解，加入PPARα抑制剂后上述现象发生逆转。细胞功能评价结果显示，大黄酸能促进血管舒张因子NO并抑制病理性血管生成因子VEGF水平（P＜0.05、0.01）。分子对接发现大黄酸与PPARα的亲和力较高。3PO联用大黄酸可抑制糖酵解、促进FAO，可以升高eNOS、NO水平，降低VEGF水平；大黄酸联用etomoxir能抑制糖酵解及FAO，并恢复eNOS、NO水平，降低VEGF水平。结论 大黄酸通过激活PPARα促进FAO并抑制糖酵解，进而调节血管内皮细胞NO、VEGF表达。

Objective To explore the mechanism of rhein in regulating function of glucose and lipid metabolism in vascular endothelial cells. Methods Human umbilical vein endothelial cells (HUVECs) was used for intervention with palmitic acid, followed by treatment with 2.5, 5.0, 10.0 μmol/L rhein for 24 h. qRT-PCR was used to detect the expressions of fatty acid oxidation (FAO) and key enzymes of glycolysis [carnitine palmitoyltransferase-1A (CPT-1A), glucose transporter protein 1 (GLUT1), 6-phosphofructose-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3) and hexokinase 2 (HK2)]; Seahorse XF96 analyzer was used to detect FAO and glycolysis levels; ELISA was used to detect the levels of nitric oxide (NO) and vascular endothelial growth factor (VEGF); Western blotting and immunofluorescence were used to observe the expressions of endothelial nitric oxide synthase (eNOS), VEGF, CPT1-A and PFKFB3; Molecular docking was used to predict the target of rhein. After intervened with PPARα inhibitor norathyriol, FAO inhibitor etomoxir and glycolysis inhibitor 3PO, the changes in regulation of endothelial cell glucose and lipid metabolism function by rhein was observed. Results Palmitic acid could significantly up-regulate the expressions of glucose and lipid metabolism enzymes (CPT-1A, GLUT1, PFKFB3, HK2) in HUVECs (P < 0.05, 0.01); Compared with model group, 10.0 μmol/L rhein significantly up-regulated the mRNA expression of CPT-1A (P < 0.05), and significantly down-regulated the mRNA expressions of GLUT1, PFKFB3 and HK2 (P < 0.01). Metabolic phenotype experiments found that rhein could promote FAO and inhibit glycolysis, and this phenomenon was reversed after the addition of PPARα inhibitors. The results of cellular functional evaluation showed that rhein could promote vasodilator factor NO and inhibit the level of pathological angiogenic factor VEGF (P < 0.05, 0.01). Molecular docking revealed a high affinity between rhein and PPARα. The combination of 3PO and rhein could inhibit glycolysis, promote FAO, increase eNOS and NO levels, and reduce VEGF level; The combination of rhein and etomoxir could inhibit glycolysis and FAO, restore eNOS and NO levels, and reduce VEGF level. Conclusion Rhein promotes FAO and inhibits glycolysis by activating PPARα, thereby regulating the expressions of NO and VEGF in vascular endothelial cells.

R285.5

国家自然科学基金面上项目（81873287）