鬼臼苦素对人结直肠癌HCT-15细胞增殖和周期的影响及其作用机制

doi:10.16098/j.issn.0529-1356.2020.02.014

摘要/Abstract

摘要：

目的探讨鬼臼苦素（PPP）对人结直肠癌细胞系HCT-15细胞增殖和周期的影响及其作用机制。方法不同浓度PPP处理HCT-15细胞，采用细胞计数试剂盒8（CCK-8）法检测HCT-15细胞的增殖活性；倒置显微镜下观察HCT-15细胞的形态变化；流式细胞术检测HCT-15细胞周期的改变；Western blotting检测增殖细胞核抗原（PCNA）及细胞周期蛋白D1（cyclinD1）的蛋白表达水平。结果 CCK-8法结果显示，PPP能显著抑制HCT-15细胞的增殖活性，并呈剂量-时间依懒性；倒置显微镜下观察发现，细胞失去原有形态回缩为圆形，折光性降低，活细胞数量明显减少；流式细胞术结果显示，G₂/M期细胞比例显著升高，G₀/G₁期和S期细胞比例明显降低；Western blotting结果显示，PCNA、细胞周期蛋白D1的表达水平明显降低。结论 PPP可显著抑制人结直肠癌HCT-15细胞的增殖，其作用机制可能是通过下调PCNA、cyclinD1的表达水平，进而将细胞阻滞于G₂/M期发挥抑制作用。

关键词: 鬼臼苦素, 结直肠癌HCT-15细胞, 流式细胞术, 免疫印迹法, 人

Abstract:

Objective To investigate the effect of picropodophyllin(PPP) on proliferation and cell cycle of human colorectal cancer cell line HCT-15 and to clarify the related mechanism. Methods Different concentrations of PPP were used to treat HCT-15 cells, the proliferative activity of HCT-15 cells was detected by cell courting kit-8(CCK-8). Morphological changes of HCT-15 cells were observed under inverted microscope. Flow cytometry was used to detect the changes of HCT-15 cell cycle and Western blotting was used to detect the expression of proliferating cell nuclear antigen (PCNA) and cell cyclinD1. Results The result of CCK-8 assay showed that PPP inhibited the proliferation of HCT-15 cells in a dose-time-dependent manner. Under the inverted microscope, it was found that the HCT-15 cells lost their original shape and became round, the refractive index decreased, and the number of living cells decreased significantly. Flow cytometry showed that the proportion of G₂/M cells increased significantly, and the proportion of G₀/G₁ phase and S phase cells decreased significantly. Western Blot result showed that the expression of PCNA, cyclinD1 protein was significantly decreased. Conclusion PPP significantly inhibited the proliferation of human colorectal cancer HCT-15 cells, suggesting that the mechanism might be to block the cells in G₂/M phase by down-regulating the expression of PCNA, cyclinD1 protein.

Key words: Picropodophyllin, Colorectal cancer cells HCT-15, Flow cytometry, Western blotting, Human

中图分类号:

R735.3

周丽丽王瑞婷赵杨薛晶刘赛璇申兴斌. 鬼臼苦素对人结直肠癌HCT-15细胞增殖和周期的影响及其作用机制[J]. 解剖学报, 2020, 51(2): 233-238.

ZHOU Li-li WANG Rui-ting ZHAO Yang XUE Jing LIU Sai-xuan SHEN Xing-bin. Effects of picropodophyllin on proliferation and cell cycle of human colorectal cancer HCT-15 cells and its mechanism#br#[J]. Acta Anatomica Sinica, 2020, 51(2): 233-238.

参考文献

［1］ Ferlay J, Soerjomataram Ⅰ, Dikshit R, et al. Cancer incidence and mortality worldwide: Sources, methods and major patterns in GLOBOCAN 2012［J］. Int J Cancer, 2015, 136(5): 359-386.
［2］ Binefa G, Rodriguez-Moranta F, Teule A, et al. Colorectal cancer: from prevention to personalized medicine［J］. World J Gastroenterol, 2014, 20(22):6786-6808.
［3］ Ling CQ, Yue XQ, Ling C. Three advantages of using traditional Chinese medicine to prevent and treat tumor［J］. J Integr Med, 2014, 12(4):331-335.
［4］ Tao WW, Luo X, Cui B, et al. Practice of traditional Chinese medicine for psycho-behavioral intervention improves quality of life in cancer patients: a systematic review and metaanalysis［J］. Oncotarget, 2015, 6(37):39725-39739.
［5］ Str ?mberg T, Feng X, Delforoush M, et al.Picropodophyllin inhibits proliferation and survival of diffuse large B-cell lymphoma cells［J］. Med Oncol, 2015, 32(7):188.
［6］ Tomizawa M, Shinozaki F, Motoyoshi Y, et al. Picropodophyllin and sorafenib synergistically suppress the proliferation and motility of hepatocellular carcinoma cells［J］. Oncol Lett, 2014, 8(5):2023-2026.
［7］ Lu XS, Wang LD, Xin JM, et al. Picropodophyllin inhibits epithelial ovarian cancer cells in vitro and in vivo［J］. Biochem biophy Res Commun, 2013, 435(3): 385-390.
［8］ Vasileanu D, Weng WH, Gimita A, et al. The insulin-like growth factor-Ⅰ receptor inhibitor PPP produces only very limited resistance in tumor cells exposed to long-term selection［J］. Oncogene, 2006, 25(22):3186-3195.
［9］ Gallagher EJ, LeRoith D. The proliferating role of insulin and insulin-like growth factors in cancer［J］. Trends Endocrinol Metab, 2010, 21(10): 610-618.
［10］ Hongo A, Kuramoto H, Nakamura Y,et al. Antitumor effects of a soluble insulin-like growth factor Ⅰ receptor in human ovarian cancer cells: adva -ntage of recombinant protein administration in vivo［J］. Cancer Res, 2003, 63(22):7834-7839.
［11］ Wu GB, Huang ChM.Targeted treatment of Insulin-like growth Factor Ⅰ receptor［J］. Journal of International Oncology, 2006，33(4):45-49.(in Chinase)
武广彬，黄昌明. 胰岛素样生长因子Ⅰ型受体肿瘤靶向治疗［J］. 国际肿瘤学杂志，2006，33(4)：45-49.
［12］ Wan J, Li XM, Shu SR, et al. Effects of lentivirus-mediated siRNA targeting IGF-IR on migration and invasion abilities of endometrial carcinoma cell［J］. Chin J Pathophysiol, 2012, 28(8):1352-1357.
［13］ Gualberto A, Pollak M. Emerging role of insulin-like growth factor receptor inhibitors in oncology: early clinical trial results and future directions［J］. Oncogene, 2009, 28(34):3009-3021.
［14］ Liu XQ, Sun LY, Liu ShSh, et al. Expression and clinical significance of IGF-Ⅰ, IGF-IR and GLUT-4 in colorectal carcinoma［J］. Journal of Chengde Medical College, 2018, 35(1):8-11.(in Chinese)
刘晓倩，孙立园，刘莎莎，等. 大肠癌组织IGF-Ⅰ、IGF-IR、GLUT-4的表达及意义［J］.承德医学院学报，2018，35(1)：8-11.
［15］ Wang I, Cao XX, Chen Q, et al. D1XDC1 targets p21 and CyclinD1 via PI3K pathway activatin to promote colon cancer cell proliferation［J］. Cancer Sci, 2009, 100(10):1801-1808.
［16］ Str?mberg T, Ekman S, Girnita L, et al. IGF-Ⅰ receptor tyrosine kinase inhibition by the cyclolignan PPP induces G2/M-phase accumulation and apoptosis in multiple myeloma cell ［J］. Blood, 2006, 107(2):669-678.
［17］ Lee MH, Cho Y, Jung BC, et al. Parkin induces G2/M cell cycle arrest in TNF-alpha-treated Hela cells［J］. Biochem Biophys Res Commun, 2015, 464(1):63-69.
［18］ Yan Y, Hein AL, Greer PM, et al. A novel function of HER2/Neu in the activation of G2/M checkpoint in response to gamma-irradiation［J］. Oncogene, 2015, 34(17):2215-2226.
［19］ Cao CY, Li JY, Li J, et al. Long non-coding RNA Uc.187 is upregulated in preeclampsia and modulates proliferation, apoptosis and invasion of HTR-8/SVneo trophoblast cells［J］. J Cell Biochem, 2017, 118(6):1462-1470.
［20］ Shi J, Liang YG. Effect of andrographolide on cell growth, apoptosis and expression of proliferating cell nuclear antigen proteinin the human skin carcinoma A431 cell line［J］. Acta Anatomica Sinica, 2013, 44(1):73-78.(in Chinese)
石静，梁勇刚. 穿心莲内酯对人皮肤基底细胞癌 A431 细胞生长、凋亡及增殖细胞核抗原蛋白表达的影响［J］. 解剖学报，2013，44(1)：73-78.
［21］ Martín MA, Goya L, Ramos S. Preventive effects of cocoa and cocoa antioxidants in colon cancer［J］. Diseases, 2016, 4(1):6.
［22］ Abdulkadir SN, Ali NR, Alchalabi NJ. Pathological study of oral squamous cell carcinoma by application of P53 and PCNA (Immunohisto chemical Approach)［J］. Int J Curr Microbiol App Sci, 2016, 5(4):91-100.
［23］ Ren J, Cheng ChL, Lu L, et al. Expression of serum markers PCNA and Bax in patients with lung cancer and their relationship with prognosis［J］. Chongqing Medicine, 2017, 46(10): 1399-1401.(in Chinese)
任洁，程春来，芦兰，等. 肺癌血清标志物PCNA及Bax的表达及与预后的关系［J］. 重庆医学，2017，46(10)：1399-1401.
［24］ Chen JC, Chen YJ, Lin CY, et al. Amphiregulin enhances alpha6betal integrin expression and cell motility in human chondrosarcoma cells through Ras/Raf/MEK/ERK/AP-1 pathway［J］. Oncotarget, 2017, 8(15):25830.

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