Small hairpin RNA against Bcl-2 increases MTX-induced apoptosis in Raji cells
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Abstract
Objective
The aim of this study was to study the effect of Bcl-2 small hairpin RNA (shRNA) enhancing methotrexate (MTX)-induced apoptosis of Raji cells.
Methods
Expression plasmid with Bcl-2 shRNA was transfected into Raji cells by Lipofectmine 2000 and then treated with MTX. At 48 h of transfection, the expression level of Bcl-2 mRNA and protein was evaluated by RT-PCR and immunofluorescence. MTT assay was used to analyze cell proliferation at 24, 48 and 72 h. Apoptosis was detected by Giemsa staining and flow cytometric cell cycle analysis.
Results
After transfection with Bcl-2 shRNA, the expression levels of Bcl-2 mRNA and protein in Raji cells decreased (P < 0.05). Using Giemsa staining, cells transfected with Bcl-2 shRNA combined with MTX at 48 h displayed changes of apoptosis. MTX significantly inhibited the growth of cells after transfected with Bcl-2 shRNA (P < 0.05). Apoptotic rates of the Raji cells treated with Bcl-2 shRNA combined with MTX significantly increased (P < 0.05), compared with either control shRNA / MTX combination or MTX-treatment cells alone.
Conclusion
Our results suggest the shRNA against Bcl-2 mRNA could increase MTX-induced apoptosis of Raji cells.
Key word
Bcl-2 small hairpin RNA (shRNA) Raji cells apoptosis methotrexate (MTX)Preview
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References
- 1.Hammond SM, Boettcher S, Caudy AA, et al. Argonaute 2, a link between genetic and biochemical analyses of RNAi. Science, 2001, 293: 1146–1150.CrossRefPubMedGoogle Scholar
- 2.Gross A, McDonnell JM, Korsmeyer SJ. BCL-2 family members and the mitochondria in apoptosis. Genes Dev, 1999, 13: 1899–1911.CrossRefPubMedGoogle Scholar
- 3.Bettaieb A, Dubrez-Daloz L, Launay S, et al. Bcl-2 proteins: targets and tools for chemosensitisation of tumor cells. Curr Med Chem Anticancer Agents, 2003, 3: 307–318.CrossRefPubMedGoogle Scholar
- 4.Marzo I, Naval J. Bcl-2 family members as molecular targets in cancer therapy. Biochem Pharmacol, 2008, 76: 939–946.CrossRefPubMedGoogle Scholar
- 5.Pan D, Wei L, Yao M, et al. Down-regulation of CT120A by RNA interference suppresses lung cancer cells growth and sensitizes to ultraviolet-induced apoptosis. Cancer Lett, 2006, 235: 26–33.CrossRefPubMedGoogle Scholar
- 6.He DM, Zhang Y, Liu GX, et al. Construction of Bcl-2 shRNA expression plasmid and its inhibition on the growth of HL-60 cells. Chin J Modern Med (Chinese), 2005, 15: 3736–3742.Google Scholar
- 7.Hao JH, Gu QL, Liu BY, et al. Inhibition of the proliferation of human gastric cancer cells SGC-7901 in vitro and in vivo using Bcl-2 siRNA. Chin Med J, 2007, 120: 2105–2111.PubMedGoogle Scholar
- 8.Ocker M, Neureiter D, Lueders M, et al. Variants of bcl-2 specific siRNA for silencing antiapoptotic bcl-2 in pancreatic cancer. Gut, 2005, 54: 1298–1308.CrossRefPubMedGoogle Scholar
- 9.Nagamatsu K, Tsuchiya F, Oguma K, et al. The effect of small interfering RNA (siRNA) against the Bcl-2 gene on apoptosis and chemosensitivity in a canine mammary gland tumor cell line. Res Vet Sci, 2008, 84: 49–55.CrossRefPubMedGoogle Scholar
- 10.Lei XY, Zhong M, Feng LF, et al. siRNA-mediated Bcl-2 and Bcl-xl gene silencing sensitizes human hepatoblastoma cells to chemotherapeutic drugs. Clin Exp Pharmacol Physiol, 2007, 34: 450–456.CrossRefPubMedGoogle Scholar