IL-23基因转染对荷瘤小鼠肿瘤组织血管生成的影响

Effect of IL-23 gene transfection on angiogenesis in tumor-bearing mice

  • 摘要: 目的 将小鼠IL-23(mIL-23)基因转染小鼠乳腺癌细胞(MA-891),检测转染后小鼠乳腺癌组织内VEGF的表达及微血管密度,了解VEGF的表达是否受IL-23基因转染的影响。方法 应用逆转录病毒载体(LXSN)将含IL-23的基因质粒经ψ2(eco-tropic)和PA317(amphotropic)两种细胞包装,G418筛选获得携带IL-23基因的病毒后,转染MA-891细胞,经G418筛选后获得IL-23/MA-891阳性克隆。用ELISA法检测IL-23/MA-891细胞分泌IL-23的能力;用MTT比色法检测细胞体外增殖能力;分别取接种IL-23/MA-891、LXSN/MA-891、MA-891细胞30天时的小鼠肿瘤组织,采用RT-PCR法以及免疫组化法检测三组小鼠肿瘤组织中VEGF的表达。结果 1)免疫组化法检测显示,接种IL-23/MA-891细胞组小鼠肿瘤组织中VEGF的表达水平明显低于接种LXSN/MA-891和MA-891细胞组小鼠的肿瘤组织(P<0.01);CD34阳性的微血管计数(MVD)为18.23±6.92,明显低于接种LXSN/MA-891(36.13±10.40)和MA-891细胞组(38.16±12.30)小鼠的肿瘤组织(P<0.01)。2)RT-PCR法检测显示,接种IL-23/MA-891细胞组小鼠肿瘤组织中VEGFmRNA表达水平明显低于接种LXSN/MA-891细胞和MA-891细胞组小鼠的瘤组织(P<0.01)。结论 转染IIL-23基因的小鼠乳腺癌细胞在荷瘤小鼠体内通过分泌IL-23,可能降低细胞VEGF的表达,抑制肿瘤微血管生成,从而发挥抗肿瘤作用。

     

    Abstract: Objective To study the expression of VEGF and the microvascular density in breast cancer tissue of mice transfected with IL-23(mIL-23)and to examine whether the expression of VEGF is affected by IL-23 gene transfection.MethodsIL-23 gene was transfected into two packing cell lines(ecotropic ψ2 and amphotropic PA317),respectively,with a retrovirus vector(LXSN).Positive cellular clones that can produce retroviruses carrying IL-23 gene were screened by G418.Retroviruses were used to transduce the mIL-23 gene into mouse mammary cancer cells(MA-891).After screened by G418,IL-23/MA-891 cells expressing IL-23 protein were obtained.Expression of IL-23 protien in IL-23/MA-891 cells was detected by ELISA.Proliferation of MA-891,LXSN/MA-891 and IL-23/MA-891 cells was detected by MTT in vitro.Three kinds of inoculated tumor were harvested on day 30 after s.c injection.Expression of mRNA and protein of VEGF was detected by RT-PCR and immunohistochemistry.Microvascular density(stained with CD34)in the grafted tumor tissues was detected by immunohistochemistry,observed under optical microscope and the value of MVD was calculated.ResultsIn the IL-23 gene-transfected tumor group,the expression of mRNA and protein of VEGF was lower than that in LXSN/MA-891 and MA-891 groups.Meanwhile,the microvascular density was lower in IL-23/MA-891 tumor cell group(18.23±6.92)than in LXSN/MA-891 group(36.13±10.40)and MA-891 group(38.16±12.30,P<0.01).ConclusionIL-23 secreted from IL-23/MA-891 cells exerts its anti-tumor effect by reducing the expression of VEGF and MVD,and by inhibiting the angiogenesis in tumors.

     

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