Ghrelin对同型半胱氨酸诱导大鼠心肌微血管内皮细胞损伤和炎性反应的保护作用

Effects of ghrelin on homocysteine-induced rat cardiac microvascular endothelial cell dysfunction and inflammatory response

目的探讨Ghrelin对同型半胱氨酸(Hcy)诱导大鼠心肌微血管内皮细胞(CMECs)损伤和炎性反应的保护作用及作用机制。方法取雄性SD大鼠心肌细胞培养鉴定后,分别以不同浓度的Hcy、不同浓度Ghrelin孵育24 h,采用MTT法检测细胞活力。另选细胞随机分为对照组、Hcy组(0.25 mmol/L)、Ghrelin组(100 ng/L)和混合组,Hoechst染色计算细胞凋亡率,NO试剂盒检测NO的分泌活性,ELISA检测白细胞介素6(IL-6)、细胞间黏附分子1(ICAM-1)的分泌,免疫印迹法检测NF-κB蛋白的表达。结果不同浓度Hcy细胞活性较不同浓度Ghrelin明显降低(P<0.05,P<0.01)。与对照组比较,Hcy组NO的分泌活性显著降低,细胞凋亡率显著上升,IL-6和ICAM-1的分泌显著增加,NF-κB的表达显著增加,差异有统计学意义(P<0.05,P<0.01)。与Hcy组比较,混合组可以明显抑制上述指标(P<0.05)。结论 Hcy可诱发大鼠CMECs的损伤和炎性反应,而Ghrelin预处理对Hcy诱导的损伤及炎性反应有明显的抑制作用,其保护作用机制可能与抑制NF-κB信号通路有关。

Objective To investigate the effects of ghrelin on homocysteine（Hcy）-induced rat cardiac microvascular endothelial cell（CMECs） dysfunction and inflammatory response and its possible mechanism. Methods CMECs were isolated from rat left ventricle and cultured with Hcy at the concentrations of 0.1,0.25,0.5,1 mmol/L respectively for 24 hours, or with Hcy 0.25 mmol/L＋ ghrelin at the concentrations of 0,10,100,1000 μg/L respectively for one hour. MTT was used to detect the cell viability. NO secretion was assessed by Griess reaction. The morphological assessment of apoptosis was studied by Hoechst staining. ELISA was used to detect the contents of the cell inflammatory factors. Western blot was used to examine the protein expression of the NF-κB. Results After treatment with Hcy,the survival rate and the activity of NO of CMECs reduced significantly,cell apoptosis rate rose significantly, and the secretion of inflammatory factors in- creased significantly. Moreover,compared with single Hcy-treated group, after pretreatment with ghrelin, the activity of CMECs was significantly higher （P 〈0.05）, the activity of NO significantly increased （P 〈 0.05）, and the expression of inflammatory cytokines obviously reduced （P 〈 0.05）. Western blot showed the expression of NF-κB elevated after treatment with Hcy,whereas reversed significantly after pretreatment with ghrelin. Conclusion It is suggested that ghrelin inhibits the homoeysteine-induced CMECs impairment and inflammatory response, which is likely mediated by inhibition of NF-κB signal pathway.