慢病毒介导NK4过表达对人肺腺癌移植瘤生长的影响

目的：研究瘤内注射慢病毒介导的NK4过表达对肺腺癌移植瘤生长、转移以及微血管生成的影响。方法：构建人肺腺癌A549裸鼠皮下移植瘤模型并随机分为3组。通过瘤内注射的方式分别向三组注射过表达慢病毒LV-NK4 (50 μl, 6×108 TU/ml)、空载慢病毒LV-con (50 μl, 6×108 TU/ml)和PBS(50 μl)。观察28天后将裸鼠处死，测量瘤体积及瘤重。HE染色观察瘤组织、肺组织情况。采用TUNEL法检测各组瘤体细胞凋亡情况。免疫组化染色检测c-Met蛋白的表达，微血管密度(MDV)检测评估血管生成。结果：小鼠荷瘤实验显示，LV-NK4组瘤体积(316.166±89.290 mm3)小于LV-con组(812.714±348.967mm3)和PBS组(838.598±390.780mm3)，差异具有统计学意义，P＜0.05。LV-NK4组瘤重(0.4344±0.1234g)小于LV-con组(0.7018±0.2227g)和PBS组(0.7829±0.1510g)，差异具有统计学意义，P＜0.05。肿瘤组织病理结果显示LV-NK4组凋亡、坏死明显增多；LV-NK4组微血管密度(MDV)为(11.2±2.78)，较LV-con组(18.90±3.479)和PBS组(22.00±5.40)明显减少，P＜0.01。LV-con组和PBS组肺部可见明显肿瘤转移灶，LV-NK4组未见明显转移灶，RT-PCR/WB检测证明移植瘤组织中NK4的表达增加。免疫组化染色显示LV-NK4组裸鼠瘤组织c-Met蛋白表达明显减少。结论：慢病毒介导的NK4过表达抑制肺腺癌移植瘤的生长及肺转移。

The effect of lentivirus-mediated NK4 overexpression on the growth of lung adenocarcinoma xenograft tumor in nude mice

Objective　To study the effect of NK4 overexpression mediated by intratumoral injection of lentivirus on the growth, invasion, metastasis and microangiogenesis of lung adenocarcinoma xenografts in nude mice. Methods　A549 cells were implanted into the right flank of BALB/c mice. After modeling, the mice were randomly divided into three groups: the PBS group, the LV-con group and the LV-NK4 group, 5 mice in each group. Mice in the three groups were received intratumoral injections of PBS (25 µL), LV-con (25 μL, 6×108 TU/mL) and LV-NK4 (25 μL, 6×108 TU/mL) respectively, twice at an interval of 3 days. The tumor volumes and weights of xenografts were measured, and the inhibitory rate of tumor was calculated at the end of the experiments. The tumor tissues and lung tissues in each group were observed by hematoxylin-eosin staining. The expressions of c-Met and CD31 were detected by immunohistochemistry, and microvessel density (MVD) was used to detect angiogenesis. The apoptosis of tumor cells in each group was detected by TUNEL. The expression levels of NK4 mRNA and protein in xenograft tumors were detected by real-time PCR and Western blot assay, respectively. Results　The xenograft tumor model of human lung adenocarcinoma was established successfully. The volume of tumor was smaller in the LV-NK4 group than that of the LV-con group and the PBS group (P＜0.05). There was no significant difference between the LV-con group and the PBS group. The tumor weight was significantly decreased in the LV-NK4 group compared with the LV-con group and the PBS group (P＜0.05). The tumour suppression rate was 44.51% in the LV-NK4 group and 10.35% in the LV-con group. The necrosis and apoptosis in tumor tissue of nude mice were significantly more in the LV-NK4 group than those in the LV-con group and the PBS group. The expression of c-Met in tumor tissue was significantly decreased in the LV-NK4 group, and the microangiogenesis was inhibited. There were no obvious lung metastases in the LV-NK4 group while there were obvious metastases in the LV-con group and the PBS group. The expression levels of NK4 mRNA and protein were significantly up-regulated in the LV-NK4 group than those of the other two groups. Conclusion　Lentivirus-mediated NK4 overexpression can inhibit the growth and lung metastasis of lung adenocarcinoma xenograft in nude mice by inhibiting the expression of c-Met protein and the formation of microvessels.