CAJ | 학술논문

目的：探讨天然小分子化合物青蒿琥酯（Akt）通过诱导人头颈部鳞癌细胞凋亡而抑制肿瘤生长的分子机制。方法培养人头颈部鳞癌细胞系UM-SCC-10A，利用四甲基偶氮唑蓝（MTT）法检测Akt半数抑制浓度（IC50）；在荧光显微镜下观察不同浓度（0、2.5、5、10、20、40μmol/L）Akt对细胞形态的影响；流式细胞仪检测细胞周期及凋亡情况；免疫印迹（Western blot）分析Akt诱导凋亡相关蛋白和细胞周期调节因子表达水平的变化。结果 Akt对UM-SCC-10A细胞的生长有明显抑制作用，且生长抑制率随药物浓度增加而增加；当Akt处理细胞48 h时，IC50为15.01μmol/L。荧光显微镜下，使用细胞核染料Hoechst33258观察到细胞核内出现凋亡小体；流式细胞仪分析显示Akt诱导细胞周期阻滞在G1期，细胞出现大量凋亡；Western blot结果显示P53、P21蛋白表达量上调、细胞周期蛋白D（Cy?cline D）下调；线粒体途径诱导的Bcl-2相关X蛋白（Bax）、细胞色素C（cytochrome C）及半胱氨酸天冬氨酸蛋白酶-3（caspase-3）表达上调，B细胞淋巴瘤/白血病-2（Bcl-2）、procaspase-3表达下调，线粒体膜电位降低。结论 Akt经由线粒体途径诱导UM-SCC-10A细胞凋亡，阻滞细胞周期于G1期，进而抑制肿瘤细胞增殖。
목적：탐토천연소분자화합물청호호지（Akt）통과유도인두경부린암세포조망이억제종류생장적분자궤제。방법배양인두경부린암세포계UM-SCC-10A，이용사갑기우담서람（MTT）법검측Akt반수억제농도（IC50）；재형광현미경하관찰불동농도（0、2.5、5、10、20、40μmol/L）Akt대세포형태적영향；류식세포의검측세포주기급조망정황；면역인적（Western blot）분석Akt유도조망상관단백화세포주기조절인자표체수평적변화。결과 Akt대UM-SCC-10A세포적생장유명현억제작용，차생장억제솔수약물농도증가이증가；당Akt처리세포48 h시，IC50위15.01μmol/L。형광현미경하，사용세포핵염료Hoechst33258관찰도세포핵내출현조망소체；류식세포의분석현시Akt유도세포주기조체재G1기，세포출현대량조망；Western blot결과현시P53、P21단백표체량상조、세포주기단백D（Cy?cline D）하조；선립체도경유도적Bcl-2상관X단백（Bax）、세포색소C（cytochrome C）급반광안산천동안산단백매-3（caspase-3）표체상조，B세포림파류/백혈병-2（Bcl-2）、procaspase-3표체하조，선립체막전위강저。결론 Akt경유선립체도경유도UM-SCC-10A세포조망，조체세포주기우G1기，진이억제종류세포증식。
Objective To study the influence of artesunate (Akt) on the proliferation and apoptosis of human head and neck squamous cell carcinoma (HNSCC), and to explore its molecular mechanism thereof. Methods The HNSCC cell line,UM-SCC-10A cells, was cultured in vitro. The Inhibitory concentration 50 (IC50) was examined by MTT assay. The cell mor?phological changes were observed under inverted light micro-scope after being interfered by 0, 2.5, 5, 10, 20 and 40μmol/L Akt. Cell cycle changes and apoptosis were measured by flow cytometry. And the expression of cell cycle regulators and apop?totic associated protein were detected by Western blot assay. Results MTT assay demonstrated that Akt significantly inhib?ited the proliferation of UM-SCC-10A cells in dose-dependent manner. After UM-SCC-10A cells were treated with Akt for 48 h, IC50 was 15.01μmol/L. Morphological changes of cell apoptosis such as karyopyknosis and conglomeration were ob?served by Hoechst 33258 staining. Flow cytometry showed that the apoptosis was associated with cell cycle arrest during the G1 phase. Western blot analysis showed that p53 and p21 protein was up-regulated and Cyclin D protein was down-regulat?ed. Furthermore, results revealed that Bcl-2 associated X protein induced by a mitochondrial pathway, cytochrome C and caspase-3 were up-regulated, and Bcl-2 and procaspase-3 were down-regulated. The mitochondrial membrane potential was reduced. Conclusion Artesunate can induce apoptosis of UM-SCC-10A cells via a mitochondrial pathway, which was associated with cell cycle arrest in the G1 phase. As a result, artesunate has an obvious inhibitory effects on proliferation of UM-SCC-10A cells.