【摘要】 背景与目的:卵巢恶性肿瘤发病率、复发率、转移率均较高,有研究表明IL-12具有明显的抗肿瘤作用。本研究探讨白细胞介素-12(interleukin-12,IL-12)对人卵巢癌SKOV3细胞生长抑制作用机制及其对裸鼠皮下种植瘤成瘤能力的影响。方法:应用脂质体转染技术将含有小鼠IL-12全长基因的质粒转染到SKOV3细胞(SKOV3/IL-12组),同时以空白质粒载体转染SKOV3细胞(SKOV3/neo组)和未转染SKOV3细胞作为对照。ELISA法检测3组细胞上清中IL-12表达。MTT法检测3组SKOV3细胞的抑制率和细胞倍增时间。建立裸鼠皮下卵巢癌种植模型,分别接种SKOV3/IL-12、SKOV3/neo、SKOV3细胞,观察肿瘤生长情况。ELISA法检测裸鼠血清中IL-12及γ-干扰素(gamma interferon,IFN-γ)的表达,免疫组化法检测裸鼠种植瘤中血管内皮生长因子(vascular endothelial growth factor,VEGF)、微血管密度(microvessel density,MVD)及干扰素诱生蛋白-10(IFN-gamma-inducible protein10,IP-10)的表达。结果:转染48、60h后细胞上清中IL-12蛋白表达量,SKOV3/IL-12组[(473.0±38.0)pg/ml、(522.0±32.0)pg/ml]高于SKOV3/neo组[(16.0±1.3)pg/ml、(18.0±1.6)pg/ml]及SKOV3组[(16.0±1.2)pg/ml、(17.0±1.4)pg/ml](P<0.01)。SKOV3/IL-12组细胞增殖抑制率为63.7%,与对照组比较差异有统计学意义(P<0.01),SKVO3/IL-12组细胞倍增时间(45.8±2.7)h明显长于SKOV3/neo细胞组(27.6±2.2)h和SKOV3组(28.2±2.1)h(P<0.01)。裸鼠皮下卵巢癌种植模型中,SKOV3/IL-12组裸鼠成瘤率(5/8)低于对照组(8/8)(P<0.01);平均成瘤时间[(15.0±5.0)天]长于SKOV3/neo组[(7.9±3.2)天]和SKOV3组[(7.8±2.4)天](P<0.01);SKOV3/IL-12组种植瘤体积、重量和裸鼠体重低于SKOV3/neo组和SKOV3组(P<0.01),种植瘤生长速度缓慢,抑瘤率达61.3%。SKOV3/IL-12组裸鼠血清中IL-12、IFN-γ表达量高于SKOV3/neo组和SKOV3组(P<0.01)。SKOV3/IL-12组裸鼠种植瘤中IP-10阳性率(100%)高于对照组(P<0.01),VEGF阳性率(62.5%)及MVD低于对照组(P<0.01)。结论:转染IL-12能有效地抑制SKOV3细胞生长;转染IL-12的SKOV3/IL-12细胞在裸鼠皮下的成瘤能力降低;IL-12可通过诱导IFN-γ诱生IP-10抑制肿瘤血管形成而实现其抗卵巢癌的作用。更多还原

【Abstract】 BACKGROUND & OBJECTIVE: The incidence, recurrence rate, and metastasis rate of malignant ovarian tumors are high. Interleukin-12 (IL-12) has antitumor effect. This study was to investigate the effects of IL-12 on the proliferation of human ovarian carcinoma SKOV3 cells in vitro and in vivo. METHODS: The plasmid containing mouse IL-12 gene was transfected into SKOV3 cells (SKOV3/IL-12). Blank plasmid-transfected SKOV3 cells (SKOV3/neo) and untransfected SKOV3 cells were used as controls. The expression of IL-12 protein was detected by ELISA. The proliferation inhibition rate and population doubling time (PDT) of SKOV3 cells were evaluated by MTT assay. SKOV3/IL-12, SKOV3/neo and SKOV3 cells were inoculated in nude mice subcutaneously. Tumor growth was recorded. The serum levels of IL-12 and interferon-γ (IFN-γ) in mice was detected by ELISA. The expression of vascular endothelial growth factor (VEGF) and IFN-γ-inducible protein 10 (IP-10), and microvessel density (MVD) in tumor tissues was detected by immunohistochemistry. RESULTS: After transfection, the protein level of IL-12 was significantly higher in SKOV3/IL-12 cells than in SKOV3/neo and SKOV3 cells [(473.0±38.0) pg/ml vs. (16.0±1.3) pg/ml and (16.0±1.2) pg/ml at 48 h, (522.0±32.0) pg/ml vs. (18.0±1.6) pg/ml and (17.0±1.4) pg/ml at 60 h, P<0.01]. The proliferation inhibition rate of SKOV3/IL-12 cells was significantly higher than those of SKOV3/neo and SKOV3 cells (63.7% vs. 0% and 0%, P<0.01). The PDT was significantly longer in SKOV3/IL-12 cells than in SKOV3/neo and SKOV3 cells [(45.8±2.7) h vs. (27.6±2.2) h and (28.2±2.1) h, P<0.01]. In nude mice, the tumor formation rate of SKOV3/IL-12 cells was lower than those of control cells (5/8 vs. 8/8 and 8/8); the tumor formation time was significantly longer in SKOV3/IL-12 group than in SKOV3/neo group and SKOV3 group [(15.0±5.0) days vs. (7.9±3.2) days and (7.8±2.4) days, P<0.01]. The volume and weight of tumors and the body weight of mice were significantly lower in SKOV3/IL-12 group than in SKOV3/neo group and SKOV3 group (P<0.01). The serum levels of IL-12 and IFN-γ were significantly higher in SKOV3/IL-12 group than in SKOV3/neo group and SKOV3 group (P<0.01). The positive rate of VEGF and MVD were significantly lower and the positive rate of IP-10 was higher in SKOV3/IL-12 group than in SKOV3/neo group and SKOV3 group (P<0.01). CONCLUSIONS: Transfection of IL-12 can inhibit the proliferation and suppress the tumorigenic ability of SKOV3 cells. IL-12 can promote IFN-γ to induce IP-10 production, thereafter, inhibit neovascularization in tumors.更多还原