FHIT overexpression in HepG2 hepatoma cells affects growth and cyclin D1 expression in vitro

Ge,Jiayun; Shen,Simin; Zhang,Xiaowen; Wang,Kun; Liu,Bo; Sun,Deyun; Wang,Lin

doi:10.3892/etm.2013.1436

FHIT overexpression in HepG2 hepatoma cells affects growth and cyclin D1 expression in vitro

Authors:
- Jiayun Ge
- Simin Shen
- Xiaowen Zhang
- Kun Wang
- Bo Liu
- Deyun Sun
- Lin Wang
View Affiliations

Affiliations: Department of Hepatobiliary Surgery, The Second Affiliated Hospital of Kunming Medical University, Kunming, Yunnan 650101, P.R. China
Published online on: December 4, 2013 https://doi.org/10.3892/etm.2013.1436
Pages: 311-315

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

The aim of this study was to investigate the methylation status of fragile histidine triad (FHIT) and the effects of FHIT on cell growth and cyclin D1 expression in hepatoma cells. The total proteins from the human hepatoma cell lines HepG2, Hep3B and Huh7 were collected and the expression levels of FHIT were analyzed. The methylation status in the promoter region of FHIT in the hepatoma cells was measured using methylation‑specific polymerase chain reaction (PCR). The HepG2, Hep3B and Huh7 cells were subsequently treated with 5‑aza-2'-deoxycytidine (5‑azadc) and the restoration of FHIT expression was then examined. A p‑hemagglutinin (HA)‑FHIT plasmid was constructed and used to transfect the HepG2 cells, and the inhibitory effects of the transfection on cell growth were then assessed. In addition, HepG2 cells were cotransfected with the pHA‑FHIT plasmid and a cyclin D1 luciferase reporter plasmid, and the effects of FHIT on the activity of cyclin D1 transcription factor were analyzed using a luciferase assay. FHIT was observed to be expressed at a low level in Hep3B and HepG2 cells; however, it was expressed at a relatively high level in Huh7 cells. The promoter region of FHIT in the Hep3B and HepG2 cells was partially methylated, and 5‑azadc treatment induced an increased expression of FHIT. The increased expression of FHIT inhibited the growth of HepG2 cells. Cotransfection with the pHA‑FHIT plasmid significantly inhibited the transcriptional activity of the cyclin D1 promoter and decreased the expression of cyclin D1 in HepG2 cells. In conclusion, FHIT was partially methylated in the HepG2 and Hep3B hepatoma cells. The overexpression of FHIT inhibited cell growth and decreased the expression of cyclin D1 in HepG2 cells.

View Figures

View References

1	Plymoth A, Viviani S and Hainaut P: Control of hepatocellular carcinoma through hepatitis B vaccination in areas of high endemicity: perspectives for global liver cancer prevention. Cancer Lett. 286:15–21. 2009. View Article : Google Scholar : PubMed/NCBI
2	Li H, Zhang Y, Su T, et al: Hint1 is a haplo-insufficient tumor suppressor in mice. Oncogene. 25:713–721. 2006. View Article : Google Scholar : PubMed/NCBI
3	Wang L, Li H, Zhang Y, et al: HINT1 inhibits beta-catenin/TCF4, USF2 and NFkappaB activity in human hepatoma cells. Int J Cancer. 124:1526–1534. 2009. View Article : Google Scholar : PubMed/NCBI
4	Negrini M, Monaco C, Vorechovsky I, et al: The FHIT gene at 3p14.2 is abnormal in breast carcinomas. Cancer Res. 56:3173–3179. 1996.
5	Bugert P, Wilhelm M and Kovacs G: FHIT gene and the FRA3B region are not involved in the genetics of renal cell carcinomas. Genes Chromosomes Cancer. 20:9–15. 1997. View Article : Google Scholar : PubMed/NCBI
6	Sugimoto K, Yamada K, Miyagawa K, et al: Decreased or altered expression of the FHIT gene in human leukemias. Stem Cells. 15:223–228. 1997. View Article : Google Scholar : PubMed/NCBI
7	Tanaka H, Shimada Y, Harada H, et al: Methylation of the 5′ CpG island of the FHIT gene is closely associated with transcriptional inactivation in esophageal squamous cell carcinomas. Cancer Res. 58:3429–3434. 1998.
8	Croce CM, Sozzi G and Huebner K: Role of FHIT in human cancer. J Clin Oncol. 17:1618–1624. 1999.PubMed/NCBI
9	Simon B, Bartsch D, Barth P, et al: Frequent abnormalities of the putative tumor suppressor gene FHIT at 3p14.2 in pancreatic carcinoma cell lines. Cancer Res. 58:1583–1587. 1998.PubMed/NCBI
10	Calvisi DF, Conner EA, Ladu S, et al: Activation of the canonical Wnt/beta-catenin pathway confers growth advantages in c-Myc/E2F1 transgenic mouse model of liver cancer. J Hepatol. 42:842–849. 2005. View Article : Google Scholar : PubMed/NCBI
11	Behari J, Zeng G, Otruba W, et al: R-Etodolac decreases beta-catenin levels along with survival and proliferation of hepatoma cells. J Hepatol. 46:849–857. 2007. View Article : Google Scholar : PubMed/NCBI
12	Monga SP: Role of Wnt/beta-catenin signaling in liver metabolism and cancer. Int J Biochem Cell Biol. 43:1021–1029. 2011. View Article : Google Scholar : PubMed/NCBI
13	Lee HS, Park MH, Yang SJ, et al: Novel candidate targets of Wnt/beta-catenin signaling in hepatoma cells. Life Sci. 80:690–698. 2007. View Article : Google Scholar : PubMed/NCBI
14	Wang L, Zhang Y, Li H, et al: Hint1 inhibits growth and activator protein-1 activity in human colon cancer cells. Cancer Res. 67:4700–4708. 2007. View Article : Google Scholar : PubMed/NCBI
15	Zöchbauer-Müller S, Fong KM, Maitra A, et al: 5′ CpG island methylation of the FHIT gene is correlated with loss of gene expression in lung and breast cancer. Cancer Res. 61:3581–3585. 2001.
16	Ohta M, Inoue H, Cotticelli MG, et al: The FHIT gene, spanning the chromosome 3p14.2 fragile site and renal carcinoma-associated t(3;8) breakpoint, is abnormal in digestive tract cancers. Cell. 84:587–597. 1996. View Article : Google Scholar : PubMed/NCBI
17	Zheng S, Ma X, Zhang L, et al: Hypermethylation of the 5′ CpG island of the FHIT gene is associated with hyperdiploid and translocation-negative subtypes of pediatric leukemia. Cancer Res. 64:2000–2006. 2004.
18	Yang Q, Nakamura M, Nakamura Y, et al: Two-hit inactivation of FHIT by loss of heterozygosity and hypermethylation in breast cancer. Clin Cancer Res. 8:2890–2893. 2002.PubMed/NCBI
19	Leal MF, Lima EM, Silva PN, et al: Promoter hypermethylation of CDH1, FHIT, MTAP and PLAGL1 in gastric adenocarcinoma in individuals from Northern Brazil. World J Gastroenterol. 13:2568–2574. 2007. View Article : Google Scholar : PubMed/NCBI
20	Mirandola P, Gobbi G, Sponzilli I, et al: TRAIL-induced apoptosis of FHIT-negative lung cancer cells is inhibited by FHIT re-expression. J Cell Physiol. 220:492–498. 2009. View Article : Google Scholar : PubMed/NCBI
21	Werner NS, Siprashvili Z, Fong LY, et al: Differential susceptibility of renal carcinoma cell lines to tumor suppression by exogenous Fhit expression. Cancer Res. 60:2780–2785. 2000.PubMed/NCBI
22	Sevignani C, Calin GA, Cesari R, et al: Restoration of fragile histidine triad (FHIT) expression induces apoptosis and suppresses tumorigenicity in breast cancer cell lines. Cancer Res. 63:1183–1187. 2003.PubMed/NCBI
23	Zheng H, Tsuneyama K, Takahashi H, et al: Expression of PTEN and FHIT is involved in regulating the balance between apoptosis and proliferation in lung carcinomas. Anticancer Res. 27:575–581. 2007.PubMed/NCBI
24	Li H, Balajee AS, Su T, et al: The HINT1 tumor suppressor regulates both gamma-H2AX and ATM in response to DNA damage. J Cell Biol. 183:253–265. 2008. View Article : Google Scholar : PubMed/NCBI