Lentivirus-delivered short hairpin RNA targeting SNAIL inhibits HepG2 cell growth

Liu,Jing; Jiang,Gang; Liu,Shihai; Liu,Zimin; Pan,Huazheng; Yao,Ruyong; Liang,Jun

doi:10.3892/or.2013.2552

Lentivirus-delivered short hairpin RNA targeting SNAIL inhibits HepG2 cell growth

Authors:
- Jing Liu
- Gang Jiang
- Shihai Liu
- Zimin Liu
- Huazheng Pan
- Ruyong Yao
- Jun Liang
View Affiliations

Affiliations: Department of Oncology, the Affiliated Hospital of the Medical College, Qingdao University, Qingdao 266003, P.R. China, Department of Radiology, the Affiliated Hospital of the Medical College, Qingdao University, Qingdao 266003, P.R. China, Department of Central Laboratory, the Affiliated Hospital of the Medical College, Qingdao University, Qingdao 266003, P.R. China, Department of Clinical Laboratory, the Affiliated Hospital of the Medical College, Qingdao University, Qingdao 266003, P.R. China
Published online on: June 19, 2013 https://doi.org/10.3892/or.2013.2552
Pages: 1483-1487

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related mortality worldwide, and the highest incidence rates are reported in East Asia. We previously showed that SNAIL is upregulated in HCC tissues. In the present study, we aimed to investigate RNA interference-mediated targeting of SNAIL on the growth of HepG2 cells. We constructed three RNA interference plasmids targeting the SNAIL gene and selected the most efficient shRNA expression cassette. After the lentivirus (LV)-SNAIL small interfering (si)RNA vector was transfected into the HepG2 cell line, cell proliferation was measured using the MTT assay. E-cadherin mRNA and protein expression levels were examined by quantitative PCR and western blotting, respectively. We successfully constructed an LV-SNAIL siRNA lentiviral vector and demonstrated that it suppressed the expression of the SNAIL gene in HepG2 cells. RNA interference of SNAIL by the LV-SNAIL siRNA construct significantly inhibited the growth of HepG2 cells, in addition to significantly increasing E-cadherin mRNA and protein expression. Our findings strongly suggest that SNAIL and E-cadherin play a significant role in HCC progression, and exhibit a negative correlation. Furthermore, the expression of E-cadherin may be responsible for the reduced proliferation and survival of HepG2 cells. Thus, the SNAIL signaling pathway may provide a novel therapeutic target for the treatment of HCC.

View Figures

View References

1	Parkin DM, Bray F, Ferlay J and Pisani P: Estimating the world cancer burden: Globocan 2000. Int J Cancer. 94:153–156. 2001. View Article : Google Scholar : PubMed/NCBI
2	Comijn J, Berx G, Vermassen P, et al: The two-handed E box binding zinc finger protein SIP1 downregulates E-cadherin and induces invasion. Mol Cell. 7:1267–1278. 2001. View Article : Google Scholar : PubMed/NCBI
3	Osada T, Sakamoto M, Ino Y, Iwamatsu A, Matsuno Y, Muto T and Hirohashi S: E-cadherin is involved in the intrahepatic metastasis of hepatocellular carcinoma. Hepatology. 24:1460–1467. 1996. View Article : Google Scholar : PubMed/NCBI
4	Tung-Ping Poon R, Fan ST and Wong J: Risk factors, prevention, and management of postoperative recurrence after resection of hepatocellular carcinoma. Ann Surg. 232:10–24. 2000.PubMed/NCBI
5	Clark HP, Carson WF, Kavanagh PV, Ho CP, Shen P and Zaqoria RJ: Staging and current treatment of hepatocellular carcinoma. Radiographics. 25(Suppl 1): S3–S23. 2005. View Article : Google Scholar : PubMed/NCBI
6	Hoshida Y, Moeini A, Alsinet C, Kojima K and Villanueva A: Gene signatures in the management of hepatocellular carcinoma. Semin Oncol. 39:473–485. 2012. View Article : Google Scholar : PubMed/NCBI
7	Hugo H, Ackland ML, Blick T, Lawrence MG, Clements JA, Williams ED and Thompson EW: Epithelial-mesenchymal and mesenchymal-epithelial transitions in carcinoma progression. J Cell Physiol. 213:374–383. 2007. View Article : Google Scholar : PubMed/NCBI
8	Neal CL, McKeithen D and Odero-Marah VA: Snail negatively regulates cell adhesion to extracellular matrix and integrin expression via the MAPK pathway in prostate cancer cells. Cell Adh Migr. 5:249–257. 2011. View Article : Google Scholar : PubMed/NCBI
9	Miyoshi A, Kitajima Y, Sumi K, Sato K, Haqiwara A, Koqa Y and Mivazaki K: Snail and SIP1 increase cancer invasion by upregulating MMP family in hepatocellular carcinoma cells. Br J Cancer. 90:1265–1273. 2004. View Article : Google Scholar : PubMed/NCBI
10	Cano A, Perez-Moreno MA, Rodriqo I, et al: The transcription factor Snail controls epithelial-mesenchymal transitions by repressing E-cadherin expression. Nat Cell Biol. 2:76–83. 2000. View Article : Google Scholar : PubMed/NCBI
11	Eger A, Aiqner K, Sondereqqer, et al: DeltaEF1 is a transcriptional repressor of E-cadherin and regulates epithelial plasticity in breast cancer cells. Oncogene. 24:2375–2385. 2005. View Article : Google Scholar : PubMed/NCBI
12	Yang J, Mani SA, Donaher JL, et al: Twist, a master regulator of morphogenesis, plays an essential role in tumor metastasis. Cell. 117:927–939. 2004. View Article : Google Scholar : PubMed/NCBI
13	Bolos V, Peinado H, Perez-Moreno MA, Fraga MF, Esteller M and Cana A: The transcription factor Slug represses E-cadherin expression and induces epithelial to mesenchymal transitions: a comparison with Snail and E47 repressors. J Cell Sci. 116:499–511. 2003. View Article : Google Scholar : PubMed/NCBI
14	Haraguchi M: The role of the transcriptional regulator snail in cell detachment, reattachment and migration. Cell Adh Migr. 3:259–263. 2009. View Article : Google Scholar : PubMed/NCBI
15	Peinado H, Olmeda D and Cano A: Snail, Zeb and bHLH factors in tumour progression: an alliance against the epithelial phenotype? Nat Rev Cancer. 7:415–428. 2007. View Article : Google Scholar : PubMed/NCBI
16	Palmer MB, Majumder P, Green MR, Wade PA and Boss JM: A 3′ enhancer controls Snail expression in melanoma cells. Cancer Res. 67:6113–6120. 2007.
17	Brummelkamp TR, Bernards R and Agami R: A system for stable expression of short interfering RNAs in mammalian cells. Science. 296:550–553. 2002. View Article : Google Scholar : PubMed/NCBI
18	Nishitsuji H, Ikeda T, Miyoshi H, Ohashi T, Kannaqi M and Masuda T: Expression of small hairpin RNA by lentivirus-based vector confers efficient and stable gene-suppression of HIV-1 on human cells including primary non-dividing cells. Microbes Infect. 6:76–85. 2004. View Article : Google Scholar : PubMed/NCBI
19	Chen D, Zheng X, Jiao X, Gao Y, Zhang K and Liang J: Transcriptional repressor snail and metastasis in hepatocellular carcinoma. Hepatogastroenterology. 59:1359–1365. 2012.PubMed/NCBI
20	Park JH, Sung IJ, Lee SW, Kim KW, Kim YS and Yoo MA: The zinc-finger transcription factor Snail downregulates proliferating cell nuclear antigen expression in colorectal carcinoma cells. Int J Oncol. 26:1541–1547. 2005.
21	Fan XJ, Wan XB, Yang ZL, et al: Snail promotes lymph node metastasis and Twist enhances tumor deposit formation through epithelial-mesenchymal transition in colorectal cancer. Hum Pathol. 44:173–180. 2012. View Article : Google Scholar
22	Sun M, Guo X, Qian X, et al: Activation of the ATM-Snail pathway promotes breast cancer metastasis. J Mol Cell Biol. 4:304–315. 2012. View Article : Google Scholar : PubMed/NCBI
23	Kim DH, Behlke MA, Rose SD, Chang MS, Choi S and Rossi JJ: Synthetic dsRNA Dicer substrates enhance RNAi potency and efficacy. Nat Biotechnol. 23:222–226. 2005. View Article : Google Scholar : PubMed/NCBI
24	Emadi Baygi M, Soheili ZS, Schmitz I, Sameie S and Schulz WA: Snail regulates cell survival and inhibits cellular senescence in human metastatic prostate cancer cell lines. Cell Biol Toxicol. 26:553–567. 2010.PubMed/NCBI
25	Becker KF, Rosivatz E, Blechschmidt K, Kremmer E, Sarbia M and Hofler H: Analysis of the E-cadherin repressor Snail in primary human cancers. Cells Tissues Organs. 185:204–212. 2007. View Article : Google Scholar : PubMed/NCBI
26	Merikallio H, Turpeenniemi-Hujanen T, Paakko P, et al: Snail promotes an invasive phenotype in lung carcinoma. Respir Res. 13:1042012. View Article : Google Scholar : PubMed/NCBI
27	Boutet A, Esteban M, Maxwell P and Nieto M: Reactivation of Snail genes in renal fibrosis and carcinomas: a process of reversed embryogenesis? Cell Cycle. 6:638–642. 2007. View Article : Google Scholar : PubMed/NCBI