目的 探讨鼠尾草酚对人脐静脉血管内皮细胞(HUVEC)黏附淋巴细胞能力的影响及其机制.方法 将体外培养的人脐静脉血管内皮细胞(HUVEC)分为鼠尾草酚高剂量组、鼠尾草酚低剂量组、TNF-α组和对照组.鼠尾草酚高剂量组、鼠尾草酚低剂量组分别加入10、5μmol/L鼠尾草酚培养18 h,然后加入10 ng/mL TNF-α培养6 h;TNF-α组加入10 ng/mL TNF-α培养6h;对照组只加EGM-2培养液培养24 h.用淋巴细胞黏附试验观察各组HUVEC黏附淋巴细胞的能力(以相对荧光强度表示),用酶联免疫吸附法(ELISA)法检测各组HUVEC细胞培养液上清液中的趋化因子白细胞介素8(IL-8)和单核细胞趋化蛋白-1(MCP-1),用Western blotting法检测各组HUVEC细胞中的黏附分子血管细胞黏附分子1(VCAM-1)、细胞间黏附分子1(ICAM-1).结果 鼠尾草酚高剂量组、鼠尾草酚低剂量组、TNF-α组和对照组相对荧光强度分别为1.74±0.33、2.08±0.28、2.61±0.29、1.00±0.01,TNF-α组荧光强度高于对照组(P<0.05),鼠尾草酚低、高剂量组荧光强度低于TNF-α组(P均<0.05),且鼠尾草酚高剂量组低于鼠尾草酚低剂量组(P<0.05).四组HUVEC细胞培养液上清液中IL-8、MCP-1水平分别为1 910.00±156.33、3 766.00±336.88、5 629.00±324.61、961.00±48.13和952.00±57.81、1 289.00±223.46、2 285.00±209.61、211.00±6.36.四组HUVEC细胞中VCAM-1、ICAM-1相对表达量分别为0.38±0.03、0.51±0.05、0.99±0.01、0.08±0.02和0.33±0.06、0.42±0.05、0.98±0.02、0.06±0.01.与对照组相比,TNF-α组HUVEC细胞培养液上清液中IL-8、MCP-1蛋白水平及HUVEC细胞中VCAM-1、ICAM-1相对表达量升高(P均<0.05);与TNF-α组比较,鼠尾草酚低、高剂量组HUVEC细胞培养液上清液中IL-8、MCP-1蛋白水平及HUVEC细胞中VCAM-1、ICAM-1相对表达量下降(P均<0.05),且鼠尾草酚高剂量组低于鼠尾草酚低剂量组(P<0.05).结论 鼠尾草酚能下调HUVEC黏附淋巴细胞的能力.其机制可能是鼠尾草酚能抑制TNF-α诱导的HUVEC趋化因子IL-8、MCP-1及细胞黏附分子VCAM-1、ICAM-1表达上调.%Objective To investigate the effects of carnosol on adhesion of human umbilical vein endothelial cells (HUVEC) to lymphocytes and the mechanism.Methods HUVEC cultured in vitro were divided into four groups: the high-dose, low-dose carnosol groups were pretreated with 10, 5 μmol/L carnosol for 18 h and then were induced by 10 ng/mL TNF-α for 6 h;the TNF-α group was stimulated by 10 ng/mL TNF-α for 6 h;the control group was cultured with EGM-2 medium for 24 h.The ability of lymphocyte adhesion in HUVEC was analyzed by lymphocyte adhesion test (measured by relative fluorescence intensity).We examined interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) expression in cell culture supernates of HUVEC by ELISA.Western blotting was used to detect the expression of vascular cell adhesion molecule-1 (VCAM-1) and intracellular adhesion molecule-1 (ICAM-1).Results The relative fluorescence intensities were 1.74±0.33, 2.08±0.28, 2.61±0.29, and 1.00±0.01 in the high-dose carnosol group, low-dose carnosol group, TNF-α and control groups, and the TNF-α group had significantly higher relative fluorescence intensity than that in the control group (P<0.05), while the high-dose and low-dose groups had significantly lower relative fluorescence intensities than that in the TNF-α group (both P<0.05), and the high-dose carnosol group had significantly lower relative fluorescence intensity than that in the low-dose carnosol group (P<0.05).The expression levels of IL-8 and MCP-1 in the four groups were 1 910.00±156.33, 3 766.00±336.88, 5629.00±324.61, 961.00±48.13, and 952.00±57.81, 1 289.00±223.46, 2 285.00±209.61, 211.00±6.36;VCAM-1 and ICAM-1 expression levels were 0.38±0.03, 0.51±0.05, 0.99±0.01, 0.08±0.02, and 0.33±0.06, 0.42±0.05, 0.98±0.02, 0.06±0.01.We demonstrated that the TNF-α group had significantly higher IL-8 and MCP-1 levels as well as VCAM-1 and ICAM-1 expression levels as compared with those of the control group (all P<0.05), and the low-dose and high-dose carnosol groups had significantly lower IL-8 and MCP-1 as well as VCAM-1 and ICAM-1 expression levels as compared with those of the TNF-α group (all P<0.05), while the high-dose carnosol group had significantly lower IL-8 and MCP-1 as well as VCAM-1 and ICAM-1 expression levels than those in the low-dose carnosol group (all P<0.05).Conclusion Carnosol may inhibit lymphocyte adhesion of HUVEC, and its underlying mechanism may be associated with the inhibitory effect of carnosol on the expression of chemokines IL-8, MCP-1, and CAMs VCAM-1 and ICAM-1.
展开▼
