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目的研究人IL-12对结肠癌干细胞(cancer stem cells,CSCs)生物学特性的影响及其机制。方法 IL-12/慢病毒重组质粒转染结肠CSCs构建IL-12过表达细胞模型,将IL-12过表达CSCs接种NOD/SCID小鼠,观察成瘤情况,Transwell检测结肠CSCs侵袭性,流式细胞仪检测细胞凋亡,免疫组化检测CK20表达,Western blot检测STAT6、p-STAT6、survivin、IL-12和IL-4蛋白表达。结果体外实验证明,IL-12使CSCs侵袭性降低40%,细胞成球能力降低,凋亡增加[(45.2±5.6)%vs(4.1±1.2)%]。转染IL-12的CSCs在小鼠体内肿瘤生长能力降低,抑瘤率63.53%;慢病毒/IL-12转染CSCs形成的肿瘤体积(189±21)mm3,质量(1 958.2±41.2)mg;而空载体转染CSCs形成的肿瘤体积(1 785±32)mm3,质量(5 369.8±41.3)mg。Western blot检测表明,IL-12过表达显著降低结肠CSCs上清IL-4和p-STAT6表达。结论 IL-12降低结肠CSCs自我更新能力及侵袭性,促进其分化和凋亡,可能和IL-4/STAT6通路有关。
Objective To investigate the effect and mechanism of human IL-12 on the biological characteristics of colon cancer stem cells (CSCs). Methods IL-12 overexpression cells were transfected with CS-CSCs by IL-12 / lentiviral recombinant plasmids. NOD / SCID mice were inoculated with IL-12 overexpression CSCs. The tumorigenesis was observed. Transwell assay was used to detect the invasiveness of colon CSCs. Cell apoptosis was detected by immunohistochemistry and protein expression of STAT6, p-STAT6, survivin, IL-12 and IL-4 were detected by Western blot. Results In vitro experiments demonstrated that IL-12 decreased the invasiveness of CSCs by 40%, decreased the ability of spheroid formation and increased apoptosis ([(45.2 ± 5.6)% vs (4.1 ± 1.2)%]. The CSCs transfected with IL-12 reduced the tumor growth rate in vivo, with a tumor inhibition rate of 63.53%. The volume of tumor formed by lentivirus / IL-12 transfected CSCs was (189 ± 21) mm3 and mass was (1 958.2 ± 41.2) mg (1785 ± 32) mm3 and mass (5 369.8 ± 41.3) mg, respectively. Western blot showed that IL-12 overexpression significantly decreased the expression of IL-4 and p-STAT6 in colonic CSCs supernatant. Conclusion IL-12 can decrease the self-renewal capacity and invasiveness of CSCs and promote their differentiation and apoptosis, which may be related to the IL-4 / STAT6 pathway.