两种不同给药计算方法对体外药效实验结果的影响

Effect of two different dosage calculation methods on results of pharmacological experiment in vitro

目的:研究按浓度与按单个细胞药物含量两种不同计算方法对甲基阿魏酸(methyl ferulic acid,MFA)抑制TGF-β1刺激的体外培养人肝星状细胞(HSC-LX2)α-SMA表达的影响,评价两种给药计算方法在细胞实验中的应用效果。方法:体外培养HSC-LX2,MTT法测定MFA抑制HSC-LX2增殖的作用,由此确定MFA 1.25、2.5、5mg/L(其相应的单个细胞MFA含量为1.13×10-7、2.25×10-7和4.5×10-7 mg)3个浓度进行后续的实验;以两种不同的细胞数量接种到培养皿,分别以上述浓度和单个细胞MFA含量干预72h,采用RT-PCR检测细胞α-SMA mRNA的表达和Western blotting检测细胞内α-SMA蛋白含量。结果:在两种不同细胞数量情况下,以单个细胞MFA含量计算方法干预后,相同MFA含量组α-SMA mRNA和蛋白的表达趋势一致,且与MTT结果相吻合;而以浓度计算方法干预后,相同MFA浓度组α-SMA mRNA和蛋白表达差异较大。结论:以单个细胞药物含量计算方法在MFA对HSC-LX2细胞体外药效实验中的应用效果更稳定,更科学,实验可重复性较好。

Objective： To research the effect of two defferent dosage calculation methods （according to the concentration of drug and according to the drug content of single cell） on Methyl ferulic acid （MFA）which can inhibit the alpha SMA （a-SMA） expression in human hepatic stellate cells （HSC-LX2） stimulated by TGF-hetal in vitro. Methods： HSC-LX2 cells were cultured in vitro, and the inhibition role of cells proliferation by MFA was tested by MTT method, thus determining MFA 1.25, 2.5 and 5 mg/L （the corresponding single cell MFA contents are 1.13×10^-7,2.25×10^-7 and 4. 5 × 10^-7 mg, respectively） 3 concentrations for subsequent experiments by MTT results. The quantity of two different cells was inoculated to the petri dish, then respectively by the concentration and the content of single cell above mentioned for drug intervention for 72 hours, the mRNA expressions of crSMA were assayed by RT-PCR and the protein content of ccSMA was mesured by Western blotting. R^sults： Under the condition of two kinds of different cells, according to calculation method of single cell drug content for drug intervention, the trend of the a-SMA mRNA and protein expression was consistent in the same content of MFA group, which was conformity with the results of MTT; but according to calculation method of concentration for drug intervention, the mRNA and protein expressions of a-SMA were quite different in the same MFA concentration group. Condmion： The application effect of dosage calculation method according to the drug content of single cell in the pharmacological experiment of MFA acting on HSC-LX2 cells in Vitro is more stable and more scientific, and the experimental repeatability is good.