基于3D微流控芯片分析鸡血藤总鞣质对宫颈癌细胞HeLa的药理作用

目的:研究鸡血藤总鞣质的化学成分,并分析该有效部位对宫颈癌细胞HeLa的药效与作用机制,为鸡血藤抗宫颈癌提供实验依据。方法:运用UPLC-Q-TOF/MS对鸡血藤鞣质类化学成分进行定性分析。将不同质量浓度的鸡血藤总鞣质作用于宫颈癌细胞HeLa,通过3维(3D)微流控芯片筛选合适的给药浓度及时间。运用流式细胞仪测定鸡血藤总鞣质对宫颈癌细胞HeLa周期与凋亡的影响,分别应用Flow Jo v10. 0. 7及ModFit LT 3. 2软件进行分析。采用酶联免疫吸附测定(ELISA)测定经鸡血藤总鞣质作用后,宫颈癌细胞HeLa上清液中血管内皮生长因子-A(VEGF-A)与半胱天冬酶-3(Caspase-3)因子的含量变化。结果:鉴定并推断了鸡血藤总鞣质中15种成分。最终确定鸡血藤总鞣质低、中、高给药质量浓度分别为0. 5,1. 0,2. 0 g·L-1,36 h为最佳给药时间。经鸡血藤总鞣质作用后宫颈癌细胞HeLa早凋与晚凋比例明显增加,宫颈癌细胞HeLa DNA合成前期(G0/G1期)明显增加,DNA合成期(S期)与DNA合成后期(G2/M期)减小。经鸡血藤总鞣质作用后,宫颈癌细胞HeLa上清液中VEGF-A因子显著降低,Caspase-3因子显著增加。结论:鸡血藤中含有丰富的鞣质类成分,此类成分对宫颈癌细胞HeLa有明显的抑制其增殖并促进其凋亡的作用,为该有效部位的后续研究与开发提供了实验依据。

Analysis of Pharmacological Effect of Total Tannins from Spatholobi Caulis on Cervical Cancer HeLa Cells by 3D Microfluidic Chip

Objective:To study on the chemical constituents of total tannins from Spatholobi Caulis,and to analyze the pharmacodynamics and mechanism of total tannins from Spatholobi Caulis on cervical cancer HeLa cells. Method:UPLC-Q-TOF/MS was used to qualitatively analyze the composition of total tannins from Spatholobi Caulis.The appropriate concentration and time of administration were screened by 3 dimensional(3D) microfluidic chip.Flow cytometry was used to determine the effect of total tannins from Spatholobi Caulis on the cell cycle and apoptosis of cervical cancer HeLa cells and analyzed by FlowJo v10.0.7 and ModFit LT 3.2 software.Enzyme-linked immunosorbent assay(ELISA) was used to determine the changes of vascular endothelial growth factor(VEGF)-A and Caspase-3 factors in cervical cancer HeLa cells supernatant treated with total tannins from Spatholobi Caulis. Result:Total of 15 components in total tannins from Spatholobi Caulis were identified or inferred.The low,medium and high dosages of total tannins from Spatholobi Caulis were 0.5,1.0, 2.0 g·L^-1 and the best time of administration was 36 h.The proportions of early and late apoptosis of cervical cancer HeLa cells increased significantly in the apoptosis analysis after being treated by total tannins from Spatholobi Caulis.The DNA synthesis early phase(G₀/G₁ phase) of cervical cancer HeLa cells significantly increased,and the DNA synthesis phase(S phase) and the DNA synthesis late phase(G₂/M phase) reduced.After being treated with total tannins from Spatholobi Caulis,the expression of VEGF-A in cervical cancer HeLa cells supernatant was significantly decreased and the expression of Caspase-3 was significantly increased. Conclusion:Spatholobi Caulis is rich in tannins,which can significantly inhibit the proliferation of cervical cancer HeLa cells and promote its apoptosis.This paper can provide the basis for further research of total tannins from Spatholobi Caulis.