目的:探讨驱虫斑鸠菊活性成分对人正常肝细胞(LO2)和人皮肤成纤维细胞(HFF)的细胞毒性,为将驱虫斑鸠菊药物临床用药提供实验依据。方法:以来源于人肝组织的正常肝细胞LO2和人正常皮肤组织成纤维细胞(HFF)为研究对象,通过药物细胞毒性实验测定LO2、HFF细胞在3个不同活性成分作用下的IC50值,采用CCK-8、总蛋白浓度测定、乳酸脱氢酶(LDH)释放3个不同的细胞毒性实验观察驱虫斑鸠菊活性成分对LO2和HFF细胞的细胞毒性。结果:体外毒性实验结果表明不同浓度的3个活性成分作用下LO2和HFF细胞存活率均显著降低。其中紫铆素、紫铆查尔酮、异鼠李素对LO2细胞的半数抑制浓度分别为67.537、157.383和214.043 μmol·L-1。与正常对照组相比较,紫铆素50、100 (P<0.05)、150、200、250、300、400(P<0.01)μmol·L-1实验组可显著减少LO2细胞的蛋白含量,同时增加细胞死亡率和LDH释放水平;异鼠李素浓度增加到200、250(P<0.05)、300、400(P<0.01)μmol·L-1时,可显著减少LO2细胞蛋白质含量,同时增加细胞死亡率和LDH释放水平,低浓度异鼠李素对LO2细胞存活率没有统计学意义;紫铆查尔酮组当浓度小于150 μmol·L-1时,对LO2细胞存活率没有影响,当浓度达到150、200(P<0.05)、250、300、400(P<0.01)μmol·L-1时,各剂量组均出现显著减少LO2细胞的蛋白含量,同时增加细胞死亡率和LDH释放水平的现象;紫铆素、紫铆查尔酮、异鼠李素对HFF细胞的半数抑制浓度分别为165.54、260.57和 178.04 μmol·L-1。与正常对照组相比较,紫铆素100、150、200(P<0.05)、250、300、400(P<0.01)μmol·L-1实验组,可显著减少HFF细胞的蛋白含量,同时增加细胞死亡率和LDH释放水平;异鼠李素浓度增加到150(P<0.05)、200、250、300、400(P<0.01) μmol·L-1时,可显著减少HFF细胞蛋白质含量,同时增加细胞死亡率和LDH释放水平,低浓度异鼠李素对HFF细胞存活率没有统计学意义;当紫铆查尔酮组浓度小于150 μmol·L-1时,对HFF细胞存活率没有影响,当浓度达到200、250、300(P<0.05)、400(P<0.01)μmol·L-1时,各剂量组出现均显著减少HFF细胞的蛋白含量,同时增加细胞死亡率和LDH释放水平的现象。结论:在本实验条件下,驱虫斑鸠菊活性成分紫铆素、异鼠李素、紫铆查尔酮对LO2和HFF有细胞毒性,提示临床用药时不能忽视驱虫斑鸠菊毒副作用。
Objective: To investigate the cytotoxic effect of the active components of Vernonia anthelmintica on human normal hepatocytes (LO2) and human normal skin tissue fibroblasts (HFF) cells, so as to provide experimental basis for its clinical use. Methods: Normal liver cells LO2 HFF were adopted.The IC50 values of LO2 and HFF cells under the action of three different active components were measured by drug cytotoxicity test. Three different cytotoxicity tests, including CCK-8, total protein concentration and LDH release were used to evaluate the cytotoxicity of the active components of Vernonia anthelmintica on to LO2 and HFF cells. Results: Considerably reduction in the survival rate at different active components of Vernonia anthelmintica concentrations were observed on LO2、HFF cell lines. The IC50 of butin, butein, isorhamnetin on the LO2 cell lines reached 67.537,157.383 and 214.043 μmol·L-1,respectively. Compared with the control group, 50, 100(P<0.05), 150, 200, 250, 300, 400(P<0.01) μmol·L-1 experimental group could significantly reduce the protein contents of LO2 cells, while increase cell mortality and LDH release levels;when the concentration of isorhamnetin was increased to 200, 250(P<0.05), 300,400(P<0.01) μmol·L-1, the protein contents of LO2 cells could be significantly reduced, while cell mortality and LDH release levels were increased. Isorhamnetin at lower concentrations had no statistical significance on the survival rate of LO2 cells; the concentration was less than 150 μmol·L-1 in the butein group, and there was no effect on the survival rate of LO2 cells. When the concentration reaches 150,200 (P<0.05), 250, 300, 400 (P<0.01) μmol·L-1, the protein contents of LO2 cells were significantly decreased,while the cell mortality and LDH release levels were increased.The IC50 of butin、 butein、isorhamnetin on the HFF cell lines reached to 165.54, 260.57 and 178.04 μmol·L-1,respectively. Compared with the control group, butin 100, 150, 200 (P<0.05), 250, 300, 400 (P<0.01) μmol·L-1 experimental group could significantly reduce the protein content of HFF cells. While the cell mortality and LDH release levels were increased;When the concentration of isorhamnetin was increased to 150 (P<0.05), 200, 250, 300, 400 (P<0.01) μmol·L-1, the protein content of HFF cells could be significantly reduced, while the cell mortality and LDH release levels were increased. Isorhamnetin at lower concentrations had no statistical significance on the survival rate of HFF cells; the concentration was less than 150 μmol·L-1 in the butein group, and there was no effect on the survival rate of HFF cells. When the concentration reached 200, 250, 300 (P<0.05), 400 (P<0.01) μmol·L-1, the protein contents of HFF cells were significantly decreased, while the cell mortality and LDH release were increased. Conclusion: The active components of Vernoniaanthelminticum, butin, butein, isorhamnetin are cytotoxic to LO2 and HFF, suggesting that the side effects of Vernoniaanthelminticus could not be ignored in clinical administration.
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