通关藤苷H抑制低分化鼻咽癌细胞增殖及转移的作用机制研究

Study on tenacissoside H inhibiting proliferation and metastasis of poorly-differentiated human nasopharyngeal carcinoma cells

关键词:  通关藤苷H  鼻咽癌  侵袭转移

Key words:  Tenacissoside H  nasopharyngeal carcinoma  invasion and metastasis

摘要:

[目的] 研究通关藤苷H对低分化鼻咽癌（NPC）细胞增殖和转移抑制作用及其机制。[方法] 以人低分化NPC细胞株CNE-2为病理模型，采用细胞计数试剂CCK-8检测通关藤苷H对细胞存活率的影响。摸索适宜的药效浓度范围，流式细胞法检测细胞凋亡率，蛋白免疫印迹（Western blot）法检测磷酸化蛋白激酶B、磷酸化糖原合成酶激酶-3β、周期蛋白D1、上皮性钙黏附蛋白、神经性钙黏附蛋白和波形蛋白的含量，酶联免疫吸附（Elisa）法检测两种基质金属蛋白酶（MMP-2，MMP-9）细胞培养液中的浓度，Transwell检测CNE-2的转移侵袭能力。[结果] CCK-8结果显示，通关藤苷H可显著降低CNE-2细胞的存活率（P<0.05）；流式细胞法结果表明，通关藤苷H促进CNE-2细胞的凋亡（P<0.05）；Transwell检测结果显示CNE-2转移侵袭能力随通关藤苷H浓度增加而逐渐降低，Western blot结果显示随着通关藤苷H浓度的增加，Akt的磷酸化程度逐渐减少，GSK-3β的磷酸化修饰则逐渐增加，进而上调周期蛋白D1的表达；通关藤苷H可随浓度增加逐渐上调上皮性钙黏附蛋白和下调神经性钙黏附蛋白、波形蛋白的含量，并抑制MMP-2和MMP-9的分泌（P<0.05）。[结论] 通关藤苷H可通过影响磷酸化蛋白激酶B/磷酸化糖原合成酶激酶-3β/周期蛋白D1通路，并上调上皮性钙黏附蛋白和下调神经性钙黏附蛋白、波形蛋白的含量，减少MMP-2和MMP-9的分泌，抑制CNE-2细胞的增殖、侵袭和转移。

Abstract:

[Objective] To study the inhibitory effect of tenacissoside H on proliferation and metastasis of human poorly differentiated nasopharyngeal carcinoma cells and its mechanism.[Methods] The human nasopharyngeal carcinoma cell line CNE-2 was used as a pathological model. CCK-8 was used to detect the effect of tenacissoside H on cell viability to find the appropriate concentration range. Flow cytometry was used to detect apoptotic rate. Western blot was used to detect phosphorylated protein kinase B(p-Akt) and phosphorylated glycogen synthase kinase-3β(p-GSK-3β). The expressions of cyclin D1,E-cadhexin,N-cadherin and Vimentin,the concentration of two matrix metalloprotein(MMP-2,MMP-9) were detected by Elisa,and the metastasis and invasion ability of CNE-2 were detected by transwell.[Results] CCK-8 showed that tenacissoside H could significantly reduce the survival rate of CNE-2 cells (P<0.05);flow cytometry showed that tenacissoside H promoted the apoptosis of CNE-2 cells(P<0.05);transwell showed that CNE-2 metastasis and invasion ability decreased along with the gradual increase of tenacissoside H concentration Western blot showed that with the increase of tenacissoside H concentration,the phosphorylation of Akt decreased gradually,and the phosphorylation of GSK-3β was repaired. Tenacissoside H could gradually increase E-cadherin and N-cadhexin,Vimentin,MMP-2 and MMP-9(P<0.05).[Conclusion] Tenacissoside H can inhibit the proliferation of CNE-2 cells by affecting Akt/GSK-3β/Cyclin D1 pathway,and up-regulate E-cadherin and down-regulate the contents of N-cadhexin,Vimentin,also mitigated secretion of MMP-2 and MMP-9 to inhibit the invasion and metastasis of CNE-2 cells.