黑色素瘤模型的建立与评价
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摘要
为了快速有效地进行在体药物筛选,研究了以小鼠黑色素实体瘤组织中的原代细胞构建黑色素瘤模型的方法。从荷黑色素瘤小鼠的组织中提取原代肿瘤细胞,与体外培养的B16细胞通过皮下注射分别植入两组C57BL/6小鼠体内,考察两组小鼠肿瘤显现时间和生长速度。结果表明:当接种浓度为5.0×106个/mL、每只0.2mL时,小鼠黑色素瘤原代细胞和体外培养的B16细胞成瘤率分别为100%和80%,且小鼠右前肢肿瘤(大小约4mm3)出现时间分别约在第3天和第8天。紫杉醇对两种模型的初步评价显示,与体外培养的B16细胞模型相比,原代B16细胞模型是体内筛选和评价特定化合物抗肿瘤活性的一种可行而有效的方法,成瘤时间较短、成瘤率高,所得实验数据误差也较小。模型的建立为相关药物评价模型的开发研究提供了一条可借鉴的新途径。
In order to screen drug rapidly and effectively in vivo,the approach of the establishment for mouse melanoma model with primary cell from melanin solid tumor tissue of mouse is studied.The primary cell and the cultured B16 cells in vitro are implanted into two groups of C57 BL/6 mice by hypodermic,respectively.The appearing time and the growth rate of tumor in two groups of mouse are investigated.The results show that when the inoculation is 0.2 mL for each one(the cell concentration of 5.0×106/mL),the tumor formation rates of the primary tumor cells of mouse melanoma and in vitro cultured B16 cells are100% and 80%,respectively,and it takes about 3 and 8 days respectively that the tumor sizes become about 4 mm3.The preliminary evaluation of two models are carried out with taxol,showing that the primary B16 cell model is a feasible and effective method for screening and evaluating the antitumor activity of specific compounds in vivo compared with the B16 cell model cultured in vitro,and it has shorter tumor forming time,higher tumor forming rate and less error in the obtained experimental data,which provides a new way for the development and research of related drug evaluation models.
In order to screen drug rapidly and effectively in vivo,the approach of the establishment for mouse melanoma model with primary cell from melanin solid tumor tissue of mouse is studied.The primary cell and the cultured B16 cells in vitro are implanted into two groups of C57 BL/6 mice by hypodermic,respectively.The appearing time and the growth rate of tumor in two groups of mouse are investigated.The results show that when the inoculation is 0.2 mL for each one(the cell concentration of 5.0×106/mL),the tumor formation rates of the primary tumor cells of mouse melanoma and in vitro cultured B16 cells are100% and 80%,respectively,and it takes about 3 and 8 days respectively that the tumor sizes become about 4 mm3.The preliminary evaluation of two models are carried out with taxol,showing that the primary B16 cell model is a feasible and effective method for screening and evaluating the antitumor activity of specific compounds in vivo compared with the B16 cell model cultured in vitro,and it has shorter tumor forming time,higher tumor forming rate and less error in the obtained experimental data,which provides a new way for the development and research of related drug evaluation models.
引文
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[19]刘恺鸣,迟路湘,鲁向辉,等.大鼠脑微血管内皮细胞的原代培养[J].第三军医大学学报,2007,29(20):2011-2013.
[20]梁朝峰,郭英,石德金,等.大鼠脑皮质微血管内皮细胞的分离和培养[J].中国病理生理杂志,2008,24(5):1038-1040.
[21]王志强,梁锐,陈明清,等.血清胰蛋白酶消化法和组织块法原代培养人包皮成纤维细胞的比较[J].生命科学研究,2012,16(3):242-247.WANG Zhiqiang,LIANG Rui,CHEN Mingqing,et al.Comparing trypsin digestion and tissue piece method in primary culture of human foreskin fibroblasts[J].Life Science Research,2012,16(3):242-247.
[22]田林郁,李胜富,周东.大鼠脑微血管内皮细胞的分离、培养及不同纯化方法的比较[J].四川大学学报(医学版),2010,41(5):869-872.TIAN Linyu,LI Shengfu,ZHOU Dong.Separation culture and purification of rat brain micro-endothelial cell[J].Journal of Sichuan University(Medical Science Edition),2010,41(5):869-872.
[2]GREENLEE R T,HILLl-HARMON M B,MURRAY T,et al.Cancer statistics,2001[J].CA Cancer J Clin,2001,51(1):15-36.
[3]韩新峰.国际抗肿瘤药物市场趋势及制剂技术进展[J].河北工业科技,2010,27(3):207-209.HAN Xinfeng.Analysis of international anticancer drug market trends and preparation technology progress[J].Hebei Journal of Industrial Science and Technology,2010,27(3):207-209.
[4]GARBEA C,EIGENTLERA T K,KEILHOLZ U,et al.Systematic review of medical treatment in melanoma:Current status and future prospects[J].Oncologist,2011,16(1):5-24.
[5]CROWSON A N,MAGRO C M,MIHM M C.Prognosticators of melanoma,the melnoma report,and the sentinel lymph node[J].Mod Pathol,2006,19(sup2):71-87.
[6]BARNHILL R L,KATZEN J,SPATZPAT A,et al.The importance of mitotic rate as a prognostic factor for localized cutaneous melanoma[J].J Cutan Pathol,2005,32(4):268-273.
[7]FRANCKEN A B,SHAW H M,THOMPSON J F,et al.The prognostic importance of tumormitotic rate confirmed in 1317patients with primary cutaneous melanoma and long follow-up[J].Ann Surg Oncol,2004,11(4):426-433.
[8]王硕,苏杭,袁经权,等.实验小鼠在癌症研究中的应用及其进展[J].中国比较医学杂志,2011,21(9):63-67.WANG Shuo,SU Hang,YUAN Jingquan,et al.Application and recent advances of laboratory mouse in cancer research[J].Chinese Journal of Comparative Medicine,2011,21(9):63-67.
[9]孟星君,李孝东,刘俊,等.C57BL/6J小鼠黑色素瘤肺转移模型的构建[J].中国实验动物学报,2018,26(2):139-144.MENG Xingjun,LI Xiaodong,LIU Jun,et al.Establishment of a C57BL/6Jmouse model of metastatic melanoma in the lung[J].Chinese Journal of Experimental Animals,2018,26(2):139-144.
[10]王淑瑞,齐浩,刘建强,等.B16黑色素瘤移植模型的建立[J].西安文理学院学报(自然科学版),2006,9(1):18-20.WANG Shurui,QI Hao,LIU Jianqiang,et al.Modeling of B16melanoma cell in mice[J].Journal of Xi’an University of Arts and Sciences(Natural Science Edition),2006,9(1):18-20.
[11]刘小珍,郑智国,凌志强.肿瘤细胞原代培养与保存[J].中国肿瘤,2015,24(4):276-283.LIU Xiaozhen,ZHENG Zhiguo,LING Zhiqiang.Primary culture and preservation of tumor cells[J].Chinese Cancer,2015,24(4):276-283.
[12]熊玮.聚多巴胺表面修饰的紫杉醇纳米颗粒抗恶性黑色素瘤的作用研究[D].广州:南方医科大学,2016.XIONG Wei.Study on the Effect of the Polydopamine Surface Modified PTX-loaded Nanoparticles for Malignant Melanoma[D].Guangzhou:Southern Medical University,2016.
[13]张胜本,黄显凯,饶本强.SW-480裸鼠移植瘤模型的建立及其生物学特性[J].第三军医大学学报,2002,22(2):109-111.ZHANG Shengben,HUANG Xiankai,RAO Benqiang.Establishment of model of transplanted tumor of human highly differentiated colonic carcinoma cell line SW-480in nude mice[J].Journal of Third Military Medical University,2002,22(2):109-111.
[14]陈陵际.运用人癌裸小鼠移植瘤模型进行抗癌新药评价[J].上海实验动物科学,2001,21(4):247-250.CHEN Lingji.The criteria of new anticancer agents evaluation in human tumor nude mice model[J].Shanghai Laboratory Animal Science,2001,21(4):247-250.
[15]GIBSON D,AMBROSIO R E,SAMUEL M,et al.A method for isolating large Numbers of viable disaggregated cells from various human tissues for cell culture establishment[J].Vitro Cell Dev Biol,1986,22(9):529-534.
[16]LEE M Y,CHOU C Y,TANG M J,et al.Epithelial mesenchymal transition in cervical cancer:correlation with tumor progression,epidermal growth factor receptor over expression,and snail up-regulation[J].Clin Cancer Res,2008,14(15):4743-4750.
[17]RIDKY T W,CHOW J M,WONG D J,et al.Invasive three-dimensional organ otypic neoplasia from multiple normal human epithelia[J].Nat Med,2010,16(12):1450-1455.
[18]王义宝,刘云会,刘丽波,等.大鼠脑微血管内皮细胞的原代培养及其生物学行为初步探讨[J].神经解剖学杂志,2006,22(2):224-228.WANG Yibao,LIU Yunhui,LIU Libo,et al.Primary culture of brain m icrovascular endothelial cell and its biological behavior study[J].Chinese Journal of Neuroanatomy,2006,22(2):224-228.
[19]刘恺鸣,迟路湘,鲁向辉,等.大鼠脑微血管内皮细胞的原代培养[J].第三军医大学学报,2007,29(20):2011-2013.
[20]梁朝峰,郭英,石德金,等.大鼠脑皮质微血管内皮细胞的分离和培养[J].中国病理生理杂志,2008,24(5):1038-1040.
[21]王志强,梁锐,陈明清,等.血清胰蛋白酶消化法和组织块法原代培养人包皮成纤维细胞的比较[J].生命科学研究,2012,16(3):242-247.WANG Zhiqiang,LIANG Rui,CHEN Mingqing,et al.Comparing trypsin digestion and tissue piece method in primary culture of human foreskin fibroblasts[J].Life Science Research,2012,16(3):242-247.
[22]田林郁,李胜富,周东.大鼠脑微血管内皮细胞的分离、培养及不同纯化方法的比较[J].四川大学学报(医学版),2010,41(5):869-872.TIAN Linyu,LI Shengfu,ZHOU Dong.Separation culture and purification of rat brain micro-endothelial cell[J].Journal of Sichuan University(Medical Science Edition),2010,41(5):869-872.