欣力康胶囊对肺癌A549细胞增殖抑制作用机制及有效成分分析

Mechanism of Xinlikang capsule inhibiting the proliferation of lung cancer A549 cells and the analysis of its effective components

目的 STAT1、STAT3和NF-κB信号通路是多种细胞因子和生长因子在细胞内传递信号的共同途径,其与肿瘤细胞增殖有密切关联。本研究观察欣力康胶囊提取物对A549细胞增殖的影响,同时研究欣力康胶囊对IFN-γ/STAT1、IL-6/STAT3和TNF-α/NF-κB信号通路的作用并分析其有效成分。方法利用MTT法检测欣力康提取物对A549细胞株增殖的影响;利用稳定转染了STAT1/STAT3-luciferase和NF-κB-luciferase报告基因质粒的细胞,分别检测欣力康提取物、欣力康10种药材提取物(丹参、当归、郁金、半枝莲、蛇莓、龙葵、黄芪、雪莲花、红参、轮环藤根)和10种药材对应单体成分(隐丹参酮、阿魏酸、姜黄素、野黄芩苷、齐墩果酸、澳洲茄碱、黄芪甲苷、秋水仙碱、人参皂苷、木兰花碱)对IFN-γ/STAT1、IL-6/STAT3和TNF-α/NF-κB信号通路的作用。结果欣力康提取物呈浓度依赖性的抑制A549细胞增殖,0.1、0.5、1、2.5和5mg/mL处理A549细胞,细胞存活率分别为(103.51±10.92)%、(78.14±6.03)%、(65.73±5.78)%、(40.41±9.20)%和(5.89±5.04)%,F=85.25,P<0.001;欣力康提取物对3条信号通路均有抑制作用,0.5、1和5mg/mL提取物对STAT1抑制率分别为(50.90±5.89)%、(70.24±2.39)%和(94.79±7.53)%(F=36.98,P<0.001),对STAT3抑制率分别为(30.39±3.86)%、(48.39±5.64)%和(97.40±5.34)%(F=41.23,P<0.001),对NF-κB抑制率分别为(-16.41±3.67)%、(1.01±3.85)%和(54.60±8.32)%(F=41.23,P<0.001);丹参、轮环藤根、半枝莲、蛇莓、雪莲花和龙葵提取物对3条信号通路均有不同程度抑制作用,10种单体成分对3条信号通路的抑制作用大部分都不同于其对应药材提取物作用。结论欣力康胶囊能够抑制A549细胞增殖,推断其作用机制可能与抑制IFN-γ/STAT1、IL-6/STAT3和TNF-α/NF-κB信号通路有关,其中丹参、轮环藤根、半枝莲、蛇莓、雪莲花和龙葵药材可能起主要作用,但其活性并非单一化合物,而是多种化合物成分共同作用的结果。

OBJECTIVE STAT1,STAT3 and NF-κB signaling pathways play an important role in the proliferation of cancer cells through regulating the expression of various cytokines and growth factors.This study aimed to observe the effect of Xinlikang Capsules on A549 proliferation and the downstream IFN-γ/STAT1,IL-6/STAT3 and TNF-α/NF-κB signal pathways and analyze its effective ingredients.METHODS A549 cell proliferation was tested by MTT assay.Cells stably transfected with STAT1/STAT3-luciferase and NF-κB-luciferase reporter plasmid were used to study the role of Xinlikang Capsule extract,Xinlikang 10 extracts(salvia,angelica,turmeric,scutellaria,duchesnea indica,solanum nigrum, astragalus,snowlotus,redginseng,ringrattan),and 10 herbs corresponding monomer components(cryptic tanshinone, ferulic acid,curcumin,wild baicalin,oleanolic acid,solanine,astragaloside,colchicine,ginsenoside,magnolia) in IFN-γ/ STAT1,IL-6/STAT3 and TNF-α/NF-κB signaling pathways.RESULTS Xinlikang extracts inhibited the proliferation of A549 cells in a concentration-dependent manner,the cell viability of A549 cells treated with extracts of 0.1,0.5,1,2.5 and 5 mg/ml Xinlikang was(103.51±10.92)%,(78.14±6.03)%,(65.73±5.78)%,(40.41±9.20)% and (5.89± 5.04)%,F=85.25,P<0.001.The STAT1 inhibition rate of the extracts of 0.5,1 and 5.0 mg/ml was (50.90± 5.89)%,(70.24±2.39)% and(94.79±7.53)%(F=36.98,P<0.001), respectively.The STAT3 inhibition rate was (30.39±3.86)%,(48.39±5.64)% and(97.40±5.34)%(F=41.23,P<0.001).The inhibition rates of NF-κB were (-16.41±3.67)%,(1.01±3.85)% and(54.60±8.32)%(F=41.23,P<0.001).Salvia miltiorrhiza extract,ring rat- tan extract,Scutellaria barbata extract,duchesnea indica extract,snow lotus extract and Solanum nigrum extract had var- ying inhibitory effects on the three signaling pathways.The inhibition of the three signal pathways by the 10 monomer components was mostly different from the corresponding herb extracts.CONCLUSIONS Xinlikang capsule can inhibit the proliferation of A549 cells,which may be related to the inhibition of IFN-γ/STAT1,IL-6/STAT3 and TNF-α/NF-κB sig- naling pathways.Salvia miltiorrhiza,ringrattan,scutellaria barbata,duchesnea indica,snow lotus and solanum nigrum may play a major role,but the biological activity of each medicinal material may be the result of a combination of multiple com-pound components,rather than the individual compound.