人黑色素瘤细胞裸鼠肺转移模型的建立

目的建立整合素αvβ3高表达的黑色素瘤细胞肺转移模型。方法通过将不同数目的M21细胞经尾静脉接种裸鼠,适时处死后计数肺表面癌结节。通过实时定量PCR和明胶酶谱的方法比较肺转移灶细胞与亲代M21细胞差异。结果M21细胞1×10^6、2×10^6、5×10^6尾静脉接种裸鼠,均能形成肺转移灶,癌结节数目均值分别为：84±8、70±6、88±12,三组之间没有明显差异,阴性对照组未形成转移灶。M21肺转移灶细胞与亲代细胞相比,增殖增快,MMP-2活性增高,整合素αv和β3mRNA表达水平明显增高。结论M21细胞1×10^6经尾静脉接种裸鼠50d内即可100%成瘤。M21肺转移灶细胞具有更快的增殖能力,整合素αvβ3和MMP-2表达水平明显增高。本实验建立了稳定的肺转移模型,为黑色素瘤和整合素αvβ3的研究提供重要的动物模型。

Establishment of a Pulmonary Metastasis Model of Human Melanoma in Nude Mice

Objective To establish a pulmonary metastasis model of integrin αvβ3-positive human melanoma in nude mice,and identify the differences between the parent melanoma cells and its pulmonary metastatic cells.Methods 1×10^6,2×10^6,and 5×10^6 human melanoma M21 cells were injected into the tail vein of nude mice of different groups,respectively.The mice were sacrificed before spontaneous death,or on the 50th day after injection of M21 cells.The white metastatic foci on the surface of the lung were counted after fixed in Bouin＇s fixative solution.The lung and other organs were sent for pathological examination.Real-time PCR and gelatin zymography were carried out to identify the differences of integrin αvβ3 and MMP-2 between the melanoma cells and its pulmonary metastatic foci cells.Results Each mouse in the M21 cell injection groups had metastatic foci in the lung.The number of white metastatic foci were 84±8,70±6,88±12 after injection of 1×10^6,2×10^6,and 5×10^6 M21 cells,respectively.Differences between the three groups were not statistically significant.Organs of negative control group had no metastatic foci.The pulmonary metastatic cells had higher activity of MMP-2（2.76-fold）,and higher mRNA level of integrin αv and β3 subunit（1.8-and 3.0-fold,respectively）,compared with the parent M21 cells.The doubling time of the former was 23.4 h,while the latter was 34.7 h.Conclusion A pulmonary metastasis model of M21 in nude mice has been established successfully.This method is practical,reliable,easy to manipulate and only 1×10^6 melanoma cells and 50 days are needed.The establishment of the pulmonary metastasis model of the integrin αvβ3-positive human melanoma will benefit the researches relevant to both melanoma and integrin αvβ3.