摘要
[目的]研究苜蓿素对脂多糖(LPS)诱导的人脐静脉内皮细胞(HUVECs)炎症损伤的保护作用.[方法]以HUVECs为研究对象,用MTT法确定苜蓿素的处理浓度,用酶联免疫分析法确定LPS建模浓度以及检测HUVECs炎症相关因子(TNF-α、iNOS、COX-2、IL-1、IL-6)表达,用Western-blot法测定NF-κB和MAPK信号通路蛋白(β-Actin、P65、IKB、P-P65、P-IKB、P38、JNK、ERK1/2、P-P38、P-JNK、P-ERK1/2)表达.[结果]不同浓度的LPS均能促进TNF-α的表达(P<0.01),其中LPS浓度在2.5～7.5 μg/mL时TNF-α的表达量最高,选用2.5 μg/mL作为建模浓度,在该浓度下LPS促进炎症相关因子和炎症相关通路蛋白表达(P<0.01),而作为阳性对照辛伐他汀能抑制LPS诱导的炎症相关因子和炎症相关通路蛋白表达(P<0.01);苜蓿素浓度在3 μg/mL之内对HUVECs活性无显著差异,因而选用3、2、1 μg/mL作为苜蓿素高、中、低剂量组;苜蓿素高、中、低剂量组均能下调LPS诱导的炎症相关因子和炎症相关通路蛋白表达(P<0.01),而且苜蓿素对炎症相关因子的抑制效果弱于辛伐他汀,而对炎症相关通路蛋白的抑制效果强于辛伐他汀,各炎症相关因子表达随苜蓿素浓度升高而降低(P<0.01),炎症相关通路蛋白表达情况也有这个下降趋势.[结论]苜蓿素能够通过抑制NF-κB和MAPK信号通路消除LPS对HUVECs诱导的炎症损伤.
Abstract
[Objective]This study aimed to evaluate the protective effect of tricin on inflammatory in-jury in human umbilical vein endothelial cells(HUVECs)induced by lipopolysaccharide(LPS).[Method]HUVECs were used as the research subject.The MTT method was employed to determine the optimal con-centration of tricin for treatment.ELISA was utilized to determine the appropriate LPS concentration for modeling and to assess the expression of inflammation-related factors(TNF-α,iNOS,COX-2,IL-1,IL-6)in HUVECs.Western blot analysis was performed to detect the protein expression of NF-κB and MAPK signal pathways(β-Actin,P65,IKB,P-P65,P-IKB,P38,JNK,ERK1/2,P-P38,P-JNK,P-ERK1/2).[Result]Different concentrations of LPS were found to promote the expression of TNF-α(P<0.01),with the highest expression observed at LPS concentrations between 2.5 μg/mL and 7.5 μg/mL.Therefore,2.5 μg/mL was chosen as the modeling concentration.At this concentration,LPS induced the upregulation of inflammation-related factors and pathway proteins(P<0.01),while simvastatin,used as the positive con-trol,inhibited the expression of these factors and proteins induced by LPS(P<0.01).HUVECs exhibited no significant difference in activity within tricin concentrations up to 3 μg/ml;therefore,tricin high(3 μg/mL),medium(2 μg/mL),and low(1 μg/mL)dosage groups were selected.Tricin in these dosage groups effec-tively downregulated the expression of inflammation-related factors and pathway proteins induced by LPS(P<0.01).Tricin showed a weaker inhibitory effect on inflammation-related factors compared to simvas-tatin,while its inhibitory effect on inflammation-related pathway proteins was stronger than that of simvas-tatin.The expression of inflammation-related factors decreased with increasing tricin concentration(P<0.01),and a similar downward trend was observed for inflammation-related pathway proteins.[Conclusion]The findings indicated that tricin can attenuate LPS-induced inflammatory damage in HU-VECs by inhibiting the NF-κB and MAPK signaling pathways.
基金项目
西南山地生态循环农业国家级培育基地项目()
国家重点基础研究发展规划项项目(2014CB138701)
NETL
NSTL
万方数据