|
|
|
| Gallbladder Cancer Cell Apoptosis and Up-regulated Expression of Caspase-3 Induced by Interference of Targeting MiRNA on Bmi-1 |
| WEI Dong1, ZOU Hao1, WANG Lin1, WANG Wen-ju2, Luo Zhi-ling3, ZHANG Xiao-wen1 |
1. Department of Hepatobiliary and Pancreatic Surgery, The Second Affiliated Hospital of Kunming Medical University, Kunming 650101, China;
2. Department of Research Laboratory Center, Yan'an Affiliated Hospital of Kunming Medical University, Kunming 650051, China;
3. The First Affiliated Hospital of Kunming Medical University, Kunming 650031, China |
|
|
|
Abstract Objective: Through targeted inhibition of oncogene Bmi-1 expression, to observe the induction of apoptosis and the effect of up-regulated Caspase-3 protein expression and to explore the mechanism of its formation in gallbladder cancer. Methods: The previously successfully constructed miRNA-Bmi-1 recombinant plasmid were transfected into GBC-SD cells and were divided into four groups of miRNABmi-1, miRNAScramble, Lipofectamine, GBC-SD. 48h after transfection, the mRNA and protein expression levels of Bmi-1 were measured by RT-PCR or Western blot, the cell growth was observed under inverted microscope; The apoptosis rate, cell cycle distribution and Caspase-3 protein levels were determined using flow cytometry. Results: RT-PCR and Western blot showed that Bmi-1 mRNA and protein expression of the gallbladder cancer cell in miRNA Bmi-1 group were significantly lower than those in the control groups (P<0.05). The inverted microscope showed a significantly higher cell death rate in the miRNA Bmi-1 group compared with the control groups; Cell cycle analysis showed that, in the miRNA Bmi-1 group, the cells were restrained at G0/G1 phase (72.20%±1.71), the cell number in G2/M and S phase decreased (18.30%±7.21, 9.50%±6.01) while apoptotic index increased (49.83%±5.19). Caspase-3 protein expression was increased (30.37%±8.10). Compared with the control groups, there was a significant difference (P<0.05). Conclusions: Targeting and silencing Bmi-1 can downregulate Bmi-1 mRNA and protein expression and induce the apoptosis in the gallbladder cancer cell, probably via the mechanism of restraining the early gallbladder cancer cell cycle at G0/G1 phase and up-regulating Caspase-3 protein expression. Bmi-1 may be involved in regulation of mitochondrial-dependent apoptotic pathway.
|
|
Received: 08 October 2013
Published: 25 December 2013
|
|
|
|
|
[1] Kayahara M, Nagakawa T. Recent trends of gallbladder cancer in Japan: an analysis of 4770 patients.Cancer, 2007, 110(3):572-580.
[2] Kumar J R, Tewari M, Rai A, et al. An objective assessment of demography of gallbladder cancer.J Surg Oncol, 2006, 93(8): 610-614.
[3] 邹声泉, 张林. 全国胆囊癌临床流行病学调查报告.中国实用外科杂志, 2000, 20(1):43-45. Zou S Q, Zhang L. Clinical epidemiologic characteristics of carcinoma of gallbladder in China. Chinese Journal of Practical Surgery, 2000, 20(1):43-45.
[4] 石景森.原发性胆囊癌的流行病学研究.肝胆胰外科杂志, 2003, 15(1):1-3. Shi J S. Epidemiologic study of primary carcinoma of the gallbladder. Journal of Hepatopancreatobiliary Surgery, 2003, 15(1):1-3.
[5] Zhu A X, Hong T S, Hezel A F, et al.Current management of gallbladder carcinoma. Oncologist, 2010, 15(2):168-181.
[6] Gourgiotis S, Kocher H M, Solaini L, et al.Gallbladder caneer.Am J Surg, 2008, 196(2):252-264.
[7] Chen Y S, Hsu Y H, Lee C F, et al.Metastatic gallbladder cancer presenting as a gingival tumor and deep neck infection. Kaohsiung J Med Sci, 2010, 26 (10):558-561.
[8] Park I K, Qian D, Kiel M, et al.Bmi-1 is required for maintenance of adult self-renewing haematopoietic stem cells.Nature, 2003, 423(6937):302-305.
[9] Min L, Dong Xiang S, Xiao Tong G, et a1.Clinicopathological and prognostic significance of Bmi-1 expression in human cervical cancer.Acta Obstet Gynecol Scand, 2011, 90(7):737-745.
[10] Yang G F, He W P, Cai M Y, et a1.Intensive expression of Bmi-1 is a new independent predictor of poor outcome in patients with ovarian carcinoma.BMC Cancer, 2010, 10:133.
[11] Xiao-Ting He, Xiu-Feng Cao, Lv Ji, et a1. Association between Bmi-1 and clinicopathological status of esophageal squamous cell carcinoma.World J Gastroenterol, 2009, 15(19):2389-2394.
[12] Isambert M, Leux C, Métairie S, et al. Incidentally discovered gallbladder cancer: When, why and which reoperation?. J Visc Surg, 2011, 148(2):e77-84.
[13] van Kemenade F J, Raaphorst F M, Blokzijl T, et al. Coexpression of BMI-1 and EZH2 polycomb-group proteins is associated with cycling cells and degree of malignancy in B-cell non-Hodgkin lymphoma. Blood, 2001, 97(12):3896-3901.
[14] Cui H, Hu B, Li T, et al.Bmi-1 is essential for the tumorigenicity of neuroblastoma cells.Am J Pathol, 2007, 170(4):1370-1378.
[15] Yip Y L, Tsang C M, Deng W, et al.Expression of Epstein-Barr virus-encoded LMP1 and hTERT extends the life span and immortalizes primary cultures of nasopharyngeal epithelial cells. J Med Virol, 2010, 82(10):1711-1723.
[16] Xiao J, Deng C. Knockdown of Bmi-1 impairs growth and invasiveness of human gastric carcinoma cells. Oncol Res, 2009, 17(11-12):613-620.
[17] Qin L, Zhang X, Zhang L, et al. Downregulation of BMI-1 enhances 5-fluorouracil-induced apoptosis in nasopharyngeal carcinoma cells. Biochem Biophys Res Commun, 2008, 371(3):531-535.
[18] Becker M, Korn C, Sienerth A R, et al.Polycomb group protein Bmi1 is required for growth of RAF driven non-small-cell lung cancer. PLoS One, 2009, 4(1): e4230.
[19] Yang M H, Hsu D S, Wang H W, et al. Bmi-1 is essential in Twist1-induced epithelial mesenchymal transition. Nat Cell Biol, 2010, 12(10):982-992.
[20] Zhang X Y, Dong Q G, Huang J S, et al. he expression of stem cell-related indicators as a prognostic factor in human lung adenocarcinoma. J Surg Oncol, 2010, 102(7):856-862.
[21] Hipfner D R, Cohen S M. Connecting proliferation and apoptosis in development and disease.Nat Rev Mol Cell Biol, 2004, 5(10):805-815.
[22] Alsuwaidi A R, Alsamri M T, Alfazari A S, et al.Lung tissue bioenergetics and caspase activity in rode.BMC Res Notes, 2013, 6:12.
[23] Porter A G, Jnicke R U.Emerging roles of caspase-3 in apoptosis.Cell Death Differ, 1999, 6(2):99-104.
[24] Jayaraman P, Sada-Ovalle I, Nishimura T, et al.IL-1β promotes antimicrobial immunity in macrophages by regulating TNFR signaling and caspase-3 activation. J Immunol, 2013, 190(8):4196-4204.
[25] D'Amelio M, Sheng M, Cecconi F.Caspase-3 in the central nervous system:beyond apoptosis.Trends Neurosci, 2012, 35(11):700-709. |
|
Viewed |
|
|
|
Full text
|
|
|
|
|
Abstract
|
|
|
|
|
Cited |
|
|
|
|
| |
Shared |
|
|
|
|
| |
Discussed |
|
|
|
|
