Effects of VEGF Secreted by MCF-7 Breast Cancer Cells on Dendritic Cells Using siRNA

WANGHai-Yan;GEYin-Lin

Chinese Journal of Biochemistry and Molecular Biology ›› 2009, Vol. 25 ›› Issue (04) : 358-363.

PDF(351 KB)
PDF(351 KB)
Chinese Journal of Biochemistry and Molecular Biology ›› 2009, Vol. 25 ›› Issue (04) : 358-363.
Research Papers

Effects of VEGF Secreted by MCF-7 Breast Cancer Cells on Dendritic Cells Using siRNA

  • WANG Hai-Yan1), GE Yin-Lin2)*
Author information +
History +

Abstract

To investigate the effects of VEGF secreted by MCF-7 breast cancer cells on differentiation, maturation and function of dendritic cells (DCs) secreted by MCF-7 breast cancer cells, small interfering RNAs (siRNAs) directed against VEGF gene were designed and transfected into MCF-7 breast cancer cells at theoptimal concentration (100 nmol/L) using cationic liposome Lipofectamine 2000 TM, while the control group was transfected with Lipofectamine 2000TMonly. The culture supernatants were tested on normal DCs. The silening of VEGF gene expression in MCF-7 cell treated with siRNA was measured by Western blotting and ELISA assay to detect the VEGF protein expression and VEGF concentration, respectively Mononuclear cells were cultured with the culture supernatants from primary MCF-7 cells (control group) and from siRNAs treated MCF-7 cells (siRNA group). The phenotypes of DCs including CD1a,CD80,CD83,CD86 and HLA-DR were evaluated by flow cytometry. MTT assay was used to detect the cytotoxicity (inhibition rates) of DCsmediated tumor-specific cytotoxic T lymphocyte (CTL) against MCF-7 cells in twodifferent culture supernatants. The results showed that DCs in siRNA group had significantly higher expression of CD86,CD80,CD83 and HLA-DR than the control group while CD1a was much lower in siRNA group (P<0.01). The killing activities of CTLs mediated by DCs were significantly altered before and after siRNA transfection (P<0.01). In summary, our designed siRNAs specifically inhibited VEGFexpression in MCF-7 cells, the inhibition of DC maturation and function in culturesupernatants from siRNA transfected cells was significantly decreased. This suggested that VEGF might play an important role in tumor developmemt, progress and immunorepression.

Key words

dendritic cell / vascular endothelial growth factor(VEGF) / small interfering RNA (siRNA) / MCF-7 cells

Cite this article

Download Citations
WANGHai-Yan;GEYin-Lin. Effects of VEGF Secreted by MCF-7 Breast Cancer Cells on Dendritic Cells Using siRNA[J]. Chinese Journal of Biochemistry and Molecular Biology, 2009, 25(04): 358-363

References

[1] Tomoda M, Maehara Y, Kakeji Y, et al. Intratumoral neovascularization and growth pattern in early gastric carcinoma [J ].Cancer ,1999,85(11) :2340-2346
[2] 王辅林,韦立新, 陈乐真. 血管内皮生长因子、微血管密度与乳腺癌淋巴结转移及预后的关系[J]. 中华病理学杂志(Wang F L, Wei L X, Chen L Z.The relationship between vascular endothelial growth factor, microvascular density, lymph node metastasis and prognosis of breast carcinoma [J]. Chin J Pathol), 2000, 29(3):172-175
[3] Commeren D L, Van Soest P L, Karimi K,et al. Paradoxical effects of interleukin-10 on the maturation of murine myeloid dendritic cells[J]. Immunology,2003,110(2):188-196
[4] Ohm J E, Carbone D P. VEGF as a mediator of tumor-associated immunodeficiency[J]. Immunol Res, 2001,23(2-3):263-272
[5] Laxmanan S, Robertson S W, Wang E, et al. Vascular endothelial cell growth factor impairs the functional ability of dendritic cells through Id pathways [J].Biochem Biophys Res Commun, 2005, 334(1):193-198
[6] Almand B, Resser J R, Lindman B, et al. Clinical significance of defective dendritic cell differentiation in cancer [J]. Clin Cancer Res,2000, 6(5):1755-1766
[7] Orsini E, Guarini A, Chiaretti S, et al. The circulating dendritic cell compartment in patients with chronic lymphocytic leukemia is severely defective and unable to stimulate an effective T-cell response [J]. Cancer Res, 2003,63(15):4497-4506
[8] Dorsett Y, Tuschl T. siRNAs: applications in functional genomics and potential as therapeutics [J]. Nat Rev Drug Discov, 2004,3(4):318-329
[9] Takei Y, Kadomatsu K, Yuzawa Y, et al. A small interfering RNA targeting vascular endothelial growth factor as cancer therapeutics[J]. Cancer Res, 2004, 64(10):3365-3370
[10] 李霞,徐文华,曹明智,等. 下调VEGF基因表达对乳腺癌细胞MCF-7的增殖抑制作用[J]. 医学分子生物学(Li X, Xu W H, Cao M Z, et al. Inhibition of proliferation of breast cancer cells MCF-7 by down-regulating the expression of VEGF [J]. J Med Mol Biol), 2005, 2(4):262-265
[11] Restifo N P. Not so Fas: Re-evaluating the mechanisms of immune privige and tumor escape [J]. Nat med, 2000, 6(5): 493-495
[12]Thompson W D. The clinical manipulation of angiogenesis: pathology, side-effects , surprises and opportunities with novel human therapies[J]. J Pathol, 2000, 190 (3):330-337
[13] Chung J, Yoon S, Datta K, et al. Hypoxia-induced vascular endothelial growth factor transcription and protection from apoptosis are dependent on alpha6beta1 integrin in breast carcinoma cells [J]. Cancer Res, 2004, 64 (14): 4711-4716
[14] Folkman J. Clinical application of research on angiogenesis[J]. N Eng J Med, 1995, 333 (26): 1757-1763
[15] Schuler G, Schuler-Thumer B, Steinman R M. The use of dendritic cells in cancer immunotherapy[J]. Curr Opin Immuno1, 2003, 15(2):138-147
[16] Nishioka Y, Hua W, Nishmun N, et al. Genetic modification of dendritic cells and its application for cancer immunotherapy[J]. J Med Invest, 2002,49(1-2): 7-17
[17] Wright-Browni V, Mcclain K L,Talpaz M, et al. Physiology and pathophysiology of dendritic cells[J]. Hum pathol,1997, 28(5): 563-579
[18] Ardavin C, Iartinez del Hoyo G, Iartin P, et al. Origin an differentiation of dendritic cells[J]. Trends Immunol,2001, 22(12): 691-700
[19] 张淑华,葛银林,罗达亚,等. 小干扰RNA靶向VEGF基因在体内外抑制乳腺癌细胞的增殖[J].中国生物化学与分子生物学报(Zhang S Y, Ge Y L, Luo D Y, et al. Small interfering RNAs directed against VEGF gene inhibit proliferation of breast cancer cells in vitro and in vivo[J]. Chin J Biochem Mol Biol), 2007,23(1):68-73
[20]Boussiotis V A, Freeman G J, Gray G, et al. B7 but not intercelluar adhesion molecule-1 costimulation prevents the induction of human alloantigen-specific tolerance[J]. J Exp Med,1993,178(5):1753-1763
[21] Cuellar A, Fonseca A, Gomez A, et al. Effect of lipopolysaccharides on human dendritic cell cultures and its inhibition by polymyxin B[J]. Biomedica, 2004,24(4):413-422
[22] 席泓,古涛,朱一蓓. 凋亡肿瘤细胞负载的树突状细胞对抗原特异性CTL的激活[J]. 上海免疫学杂志(Xi H, Gu T, Zhu Y P. Study on tumor-specific cytotoxicity T lymphocytes activated by dendritic cells loaded with apoptotic tumor cells [J].Shanghai J Immunol), 2002, 22(5):297-300
[23] Banchereau J, Schucer-Thurner B, Palucka A K. Dendritic cell as vectors for therapy[J].Cell, 2001, 106(3):271-274




PDF(351 KB)

59

Accesses

0

Citation

Detail

Sections
Recommended

/