人胃癌双标细胞系及鼠原位模型的高效构建
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摘要
目的建立一种高效胃癌原位模型。方法采用慢病毒感染方法构建稳定表达荧光素酶及GFP的双标细胞系SGC-7901/FLUC/GFP。将细胞悬液注射入造模小鼠胃浆膜下层,建模后第3天通过生物发光成像观测建模情况。2周后处死小鼠,通过HE及冰冻切片观察移植瘤形态。结果 SGC-7901/FLUC/GFP双标细胞系构建成功,该稳定转染细胞系高表达荧光素酶及GFP,细胞数目与荧光及生物发光信号呈线性正相关(R2=0.9667,r2=0.9110),可用于肿瘤在体观测。手术后第3天通过生物发光实验证实模型构建成功,肿瘤位于胃区生长。2周后通过HE及冰冻切片可见异型肿瘤细胞生长,符合胃癌组织学特征。结论成功建立了简便、安全、高效的大规模胃癌原位模型,该方法便于普及,可为胃癌研究提供宝贵的研究平台和实验工具。
Objective To establish a novel orthotopic gastric cancer model with high efficiency. Methods The human gastric cancer cell line SGC-7901 was transfected to stable expression of luciferase and GFP by lentivirus transduction(SGC-7901/FLUC/GFP). The model was developed by orthotopic transplantation of SGC-7901/FLUC/GFP into the sub-serosal layer of the stomach of nude mice. Bioluminescence imaging was used to track and monitor the tumor growth on the 3 th day. Two weeks later,8 mice were sacrificed and tumors were excised for histological identification.Results SGC-7901/FLUC/GFP was high expression luciferase and GFP. Positive correlation between cell number and bioluminescence/fluorescence intensity was observed(R2= 0. 9667,r2= 0. 9110). The orthotopic model was successfully established. On the 3 th day,bioluminescence imaging showed the tumor growth at stomach region. The HE and frozen section also confirmed the tumor growth. This method showed high efficiency with easy procedure and less mice mortality. Conclusion This method is an easy,safe and high efficient way for the establishment of orthotopic gastric cancer model on the large scale,which would provide a valuable platform for gastric cancer research.
Objective To establish a novel orthotopic gastric cancer model with high efficiency. Methods The human gastric cancer cell line SGC-7901 was transfected to stable expression of luciferase and GFP by lentivirus transduction(SGC-7901/FLUC/GFP). The model was developed by orthotopic transplantation of SGC-7901/FLUC/GFP into the sub-serosal layer of the stomach of nude mice. Bioluminescence imaging was used to track and monitor the tumor growth on the 3 th day. Two weeks later,8 mice were sacrificed and tumors were excised for histological identification.Results SGC-7901/FLUC/GFP was high expression luciferase and GFP. Positive correlation between cell number and bioluminescence/fluorescence intensity was observed(R2= 0. 9667,r2= 0. 9110). The orthotopic model was successfully established. On the 3 th day,bioluminescence imaging showed the tumor growth at stomach region. The HE and frozen section also confirmed the tumor growth. This method showed high efficiency with easy procedure and less mice mortality. Conclusion This method is an easy,safe and high efficient way for the establishment of orthotopic gastric cancer model on the large scale,which would provide a valuable platform for gastric cancer research.
引文
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[13]YAMASHITA T.Manifestation of metastatic potential in human gastric cancer implanted into the stomach wall of nude mice[J].Jpn J Cancer Res,1988,79(8):945-951.
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[15]陈愉生,陈正伟,俞梅娥,等.肺腺癌脑转移动物模型的生物发光成像和SPECT/CT的比较[J].中国实验动物学报,2017,25(1):36-42.DOI:10.3969/j.issn.1005-4847.2017.01.007.CHEN Y S,CHEN Z W,YU M E,et al.Comparison between bioluminescence imaging and SPECT/CT of mouse models of brain metastasis from lung adenocarcinoma[J].Acta Laboratorium Animalis Scientia Sinica,2017,25(1):36-42.DOI:10.3969/j.issn.1005-4847.2017.01.007.
[16]苏乔,林颖,李雯雯,等.乳腺癌BALB/c小鼠生物发光活体成像和小动物~(18)F-FDG PET/CT成像的比较研究[J].中华普通外科学文献(电子版),2017,11(1):18-22.DOI:10.3877/cma.j.issn.1674-0793.2017.01.005.SU Q,LIN Y,LI W W,et al.Comparison between bioluminescent imaging and small-animal18F-FDG PET/CT in BALB/c mice with breast cancer[J].Chin Arch Gen Surg(Electronic Edition),2017,11(1):18-22.DOI:10.3877/cma.j.issn.1674-0793.2017.01.005
[17]张建文,陈道达,王国斌.人胃癌耐药细胞株裸鼠原位移植的实验研究[J].中华胃肠外科杂志,2001,4(3):189-191.ZHANG J W,CHEN D D,WANG G B,et al.Study of orthotopic implantation of multidrug-resistance gastric carcinoma cell in nude mice[J].Chin J Gastrointest Surg,2001,4(3):189-191.
[2]WU N,HE C,ZHU B,et al.3-phosphoinositide dependent protein kinase-1(PDK-1)promotes migration and invasion in gastric cancer cells through activating the NF-kappa B pathway[J].Oncol Res,2017,25(7):1153-1159.DOI:10.3727/096504017X14845839228545.
[3]KILLION J J,RADINSKY R,FIDLER I J.Orthotopic models are necessary to predict therapy of transplantable tumors in mice[J].Cancer Metastasis Rev,1998-1999,17(3):279-284.
[4]BIBBY M C.Orthotopic models of cancer for preclinical drug evaluation:advantages and disadvantages[J].Eur J Cancer,2004,40(6):852-857.DOI:10.1016/j.ejca.2003.11.021.
[5]RYGAARD J,POVLSEN C O.Heterotransplantation of a human malignant tumour to“Nude”mice[J].Acta Pathol Microbiol Scand,1969,77(4):758-760.DOI:10.1111/j.1699-0463.1969.tb04520.x.
[6]HU H,LIU J,YAO L,et al.Real-time bioluminescence and tomographic imaging of gastric cancer in a novel orthotopic mouse model[J].Oncol Rep,2012,27(6):1937-1943.DOI:10.3892/or.2012.1713.
[7]FURUKAWA T,FU X,KUBOTA T,et al.Nude mouse metastatic models of human stomach cancer constructed using orthotopic implantation of histologically intact tissue[J].Cancer Res,1993,53(5):1204-1208.
[8]曹钟元,赵振凯,相庚,等.人胃癌裸鼠原位移植瘤模型的建立及其活体荧光成像检测[J].中国肿瘤生物治疗杂志,2016,23(5):703-707.DOI:10.3872/j.issn.1007-385X.2016.05.020.CAO Z Y,ZHAO Z K,XIANG G J,et al.Establishment of an endogastric xenograft tumor modal in nude mice and its detection by in vivo fluorescence imaging system[J].Chin J Cancer Biother,2016,23(5):703-707.DOI:10.3872/j.issn.1007-385X.2016.05.020.
[9]张广顺,张光霁,申力,等.OB吻合胶粘贴法构建615小鼠胃癌原位移植瘤模型[J].浙江中医药大学学报,2013,37(5):587-590.DOI:10.3969/j.issn.1005-5509.2013.05.038.ZHANG G S,ZHANG G J,SHEN L,et al.The building of orthotopic transplanted model of gastric tumor in 615 mice with OB anastomotic glue pasted method[J].Journal of Zhejiang Chinese Medical University,2013,37(5):587-590.DOI:10.3969/j.issn.1005-5509.2013.05.038.
[10]孙培鸣,金润森,杜晓辉,等.工程化人胃癌裸鼠原位模型的建立及其活体荧光成像观察[J].南方医科大学学报,2012,32(12):1718-1721.DOI:10.3969/j.issn.1673-4254.2012.12.008.SUN P M,JIN R S,DU X H,et al.Establishment of a nude mouse model of orthotopic engineer gastric tumor and its in vivo fluorescence imaging[J].J South Med Univ,2012,32(12):1718-1721.DOI:10.3969/j.issn.1673-4254.2012.12.008.
[11]田树红,王日超,肖敏,等.裸小鼠胃癌原位移植模型的建立[J].中国比较医学杂志,2017,27(2):21-24.DOI:10.3969/j.issn.1671-7856.2017.02.004.TIAN S H,WANG R C,XIAO M,et al.Establishment of an orthotopic gastric cancer model in nude mice[J].Chinese Journal of Comparative Medicine,2017,27(2):21-24.DOI:10.3969/j.issn.1671-7856.2017.02.004.
[12]张杰,贾丽娟,梁冰,等.应用福爱乐医用胶粘贴法构建人胃癌组织块原位移植模型[J].江苏大学学报(医学版),2014,24(4):287-289.DOI:10.13312/j.issn.1671-7783.y140023.ZHANG J,JIA L J,LIANG B,et al.Application of FAL medical glue in establishing the human gastric cancer tissue mass orthotopic transplantation model[J].Journal of Jiangsu University(Medicine Edition),2014,24(4):287-289.DOI:10.13312/j.issn.1671-7783.y140023.
[13]YAMASHITA T.Manifestation of metastatic potential in human gastric cancer implanted into the stomach wall of nude mice[J].Jpn J Cancer Res,1988,79(8):945-951.
[14]WEISSLEDER R,PITTET M J.Imaging in the era of molecular oncology[J].Nature,2008,452(7187):580-589.DOI:10.1038/nature06917.
[15]陈愉生,陈正伟,俞梅娥,等.肺腺癌脑转移动物模型的生物发光成像和SPECT/CT的比较[J].中国实验动物学报,2017,25(1):36-42.DOI:10.3969/j.issn.1005-4847.2017.01.007.CHEN Y S,CHEN Z W,YU M E,et al.Comparison between bioluminescence imaging and SPECT/CT of mouse models of brain metastasis from lung adenocarcinoma[J].Acta Laboratorium Animalis Scientia Sinica,2017,25(1):36-42.DOI:10.3969/j.issn.1005-4847.2017.01.007.
[16]苏乔,林颖,李雯雯,等.乳腺癌BALB/c小鼠生物发光活体成像和小动物~(18)F-FDG PET/CT成像的比较研究[J].中华普通外科学文献(电子版),2017,11(1):18-22.DOI:10.3877/cma.j.issn.1674-0793.2017.01.005.SU Q,LIN Y,LI W W,et al.Comparison between bioluminescent imaging and small-animal18F-FDG PET/CT in BALB/c mice with breast cancer[J].Chin Arch Gen Surg(Electronic Edition),2017,11(1):18-22.DOI:10.3877/cma.j.issn.1674-0793.2017.01.005
[17]张建文,陈道达,王国斌.人胃癌耐药细胞株裸鼠原位移植的实验研究[J].中华胃肠外科杂志,2001,4(3):189-191.ZHANG J W,CHEN D D,WANG G B,et al.Study of orthotopic implantation of multidrug-resistance gastric carcinoma cell in nude mice[J].Chin J Gastrointest Surg,2001,4(3):189-191.