基于临床肿瘤标本的胃癌转移模型建立
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摘要
目的建立基于临床肿瘤标本的胃癌转移模型,为胃癌的转移研究提供个体化动物模型。方法将胃癌新鲜的手术标本移植到裸鼠皮下,建立胃癌患者异种移植(patient-derived xenograft,PDX)模型。进一步通过手术将皮下瘤组织原位移植到裸鼠胃部肌层,连续观察裸鼠的体征状态,通过近红外荧光活体成像技术检测肿瘤转移的发生。解剖荷瘤小鼠,将肺部转移灶进一步移植裸鼠皮下获得实体瘤。HE染色观察原发瘤与转移瘤的结构特征,(short tandem repeat) STR分析原发瘤和转移瘤的遗传特性。PCR-Array分析转移瘤和原发瘤中转移相关基因的表达。结果成功建立胃癌PDX模型,移植瘤组织结构与患者保持基本一致;通过胃部原位移植发现编号C19751的小鼠发生肺和肝的转移。其中肺转移灶皮下移植后获得了实体瘤,STR分析显示原发瘤保持了与肺转移瘤一致的遗传特征。PCR-Array结果显示,与原发瘤相比,转移瘤中CXCL12,IGF1和MMP2基因表达均显著上调。结论利用临床肿瘤标本成功建立胃癌转移模型,为胃癌转移研究提供了良好的个体化模型。
Objective To establish a mouse model of gastric cancer metastasis derived from clinical tumor specimens, and to provide individualized animal models for the metastasis of gastric cancer. Methods A patient-derived xenograft(PDX) model was established by subcutaneous transplantation of fresh clinical gastric cancer specimens into nude mice. Then, the tumor tissue from the PDX model was transplanted into the muscle layer of mouse stomach. The physical status of these nude mice was observed continuously. The occurrence of tumor metastasis was detected by near-infrared fluorescence(NIRF) optical imaging in vivo. After dissecting the tumor-bearing mice, lung metastatic lesions were further subcutaneously transplanted into nude mice to obtain tumors. The histopathological structures of the primary and metastatic tumors were observed by HE staining, and the genetic characteristics were analyzed using STR tests. Furthermore, the expression of metastasis-related genes was detected by PCR-Array analysis. Results A PDX model of gastric cancer is successfully established, and the histopathological characteristics of tumors in the nude mice were basically consistent with those of the patients. Metastatic lesions in the lung and liver were found in the mouse No.C19751 following orthotopic gastric transplantation. Apparent tumors were formed after subcutaneous transplantation of lung metastatic lesions, and STR analysis showed that the primary tumors well maintained the genetic characteristics of the lung metastatic tumors. The PCR-Array result demonstrated that the expressions of CXCL12, IGF1 and MMP2 in metastatic tumors were significantly upregulated compared with that in the primary tumors. Conclusions A PDX metastasis model of gastric cancer is successfully established in nude mice by xenografting clinical tumor specimens, providing an appropriate individualized metastasis model for research on gastric cancer.
Objective To establish a mouse model of gastric cancer metastasis derived from clinical tumor specimens, and to provide individualized animal models for the metastasis of gastric cancer. Methods A patient-derived xenograft(PDX) model was established by subcutaneous transplantation of fresh clinical gastric cancer specimens into nude mice. Then, the tumor tissue from the PDX model was transplanted into the muscle layer of mouse stomach. The physical status of these nude mice was observed continuously. The occurrence of tumor metastasis was detected by near-infrared fluorescence(NIRF) optical imaging in vivo. After dissecting the tumor-bearing mice, lung metastatic lesions were further subcutaneously transplanted into nude mice to obtain tumors. The histopathological structures of the primary and metastatic tumors were observed by HE staining, and the genetic characteristics were analyzed using STR tests. Furthermore, the expression of metastasis-related genes was detected by PCR-Array analysis. Results A PDX model of gastric cancer is successfully established, and the histopathological characteristics of tumors in the nude mice were basically consistent with those of the patients. Metastatic lesions in the lung and liver were found in the mouse No.C19751 following orthotopic gastric transplantation. Apparent tumors were formed after subcutaneous transplantation of lung metastatic lesions, and STR analysis showed that the primary tumors well maintained the genetic characteristics of the lung metastatic tumors. The PCR-Array result demonstrated that the expressions of CXCL12, IGF1 and MMP2 in metastatic tumors were significantly upregulated compared with that in the primary tumors. Conclusions A PDX metastasis model of gastric cancer is successfully established in nude mice by xenografting clinical tumor specimens, providing an appropriate individualized metastasis model for research on gastric cancer.
引文
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[7] 张贺,张彩勤,赵勇,等. 基于临床手术标本的胰腺癌原位移植模型建立及评价 [J]. 中国实验动物学报. 2018(03): 296-301.Zhang H, Zhang CQ, Zhao Y, et al. Establishment and characterization of a patient-derived orthotopic xenograft (PODX) model of pancreatic cancer [ J ]. Acta Lab Animalis Sci Sin, 2018, 26 (03): 296 - 301.
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[10] 张贺,陈薛,谭邓旭,等.基于肿瘤个体化治疗药物筛选的异种移植模型评价策略[J].中国实验动物学报. 2018(04): 523-527. Zhang H, Chen X, Tan DX, et al. Evaluation strategy of patient-derived xenograft models based on drug-screening of individualized treatment [J]. Acta Lab Anim Sci Sin, 2018, 26(04): 523-527.
[11] Monsky WL, Mouta Carreira C, Tsuzuki Y, et al. Role of host microenvironment in angiogenesis and microvascular functions in human breast cancer xenografts: Mammary fat pad versus cranial tumors1 [J]. Clin Cancer Res, 2002, 8(4):1008-1013.
[12] Sicklick JK, Leonard SY, Babicky ML, et al. Generation of orthotopic patient-derived xenografts from gastrointestinal stromal tumor [J]. J Transl Med. 2014, 12: 41.
[13] Murakami T, Murata T, Kawaguchi K, et al. Cervical cancer patient-derived orthotopic xenograft (PDOX) is sensitive to cisplatinum and resistant to Nab-paclitaxel [J]. Anticancer Res. 2017, 37(1): 61-66.
[14] Lang H, Béraud C, Bethry A, et al. Establishment of a large panel of patient-derived preclinical models of human renal cell carcinoma [J]. Oncotarget. 2016, 7(37): 59336.
[15] 王勤周,张成,李丽,等. 七甲川菁荧光染料在肝细胞癌移植模型活体成像中的应用 [J]. 实验动物与比较医学. 2018(04): 250-254.Wang QZ, Zhang C, Li L, et al. The Application of heptamethine cyanine dye in optical imaging of hepatocellular carcinoma transplantation model [J]. Lab Anim Comp Med, 2018, 38(4): 250-254.
[16] Shi C, Wu JB, Pan D. Review on near-infrared heptamethine cyanine dyes as theranostic agents for tumor imaging, targeting, and photodynamic therapy [J]. J Biomed Optics. 2016, 21(5): 50901.
[17] Balkwill F. The significance of cancer cell expression of the chemokine receptor.CXCR4 [J]. Semin Cancer Biol. 2004, 14(3): 171-179.
[18] 罗玉政,陈波,李红樱. 沉默IGF1R表达对胃癌细胞侵袭与迁移能力的影响及机制 [J]. 中国临床研究. 2018(02): 163-166.Luo YZ, Chen B, Li HY. Effect of silencing IGF1R expression on invasion and migration abilities of gastric cancer cells and its mechanism [J]. Chin J Clin Res 2018, 31(02): 163-166.
[19] 李倩玉,韩冬艳,蒋虹伟,等. p28、MMP2、β-catenin在胃癌及淋巴结转移癌中的表达 [J]. 诊断学理论与实践. 2016(02): 169-173.Li QY, Han DY, Jiang HW, et al. Expressions of p28, MMP2 and β-catenin proteins in human gastric carcinoma and metastatic lymph nodes [J]. J Diagn Concepts Practice, 2016, 15(02): 169-173.
[2] Cassidy JW, Caldas C, Bruna A. Maintaining tumor heterogeneity in patient-derived tumor xenografts [J]. Cancer Res. 2015, 75(15): 2963-2968.
[3] Hiroshima Y, Maawy A, Zhang Y, et al. Patient-derived mouse models of cancer need to be orthotopic in order to evaluate targeted anti-metastatic therapy[J]. Oncotarget. 2016, 7(44): 71696.
[4] Izumchenko E, Meir J, Bedi A, et al. Patient-derived xenografts as tools in pharmaceutical development [J].. Clin Pharmacol Ther.2016, 99(6): 612-621.
[5] Kawaguchi K, Igarashi K, Murakami T, et al. Tumor-targeting Salmonella typhimurium A1-R sensitizes melanoma with a BRAF-V600E mutation to vemurafenib in a patient-derived orthotopic xenograft (PDOX) nude mouse model [J]. J Cell Biochem. 2017, 118(8): 2314-2319.
[6] Shi C, Wu JB, Chu GC, et al. Heptamethine carbocyanine dye-mediated near-infrared imaging of canine and human cancers through the HIF-1alpha/OATPs signaling axis [J]. Oncotarget. 2014, 5(20): 10114-10126.
[7] 张贺,张彩勤,赵勇,等. 基于临床手术标本的胰腺癌原位移植模型建立及评价 [J]. 中国实验动物学报. 2018(03): 296-301.Zhang H, Zhang CQ, Zhao Y, et al. Establishment and characterization of a patient-derived orthotopic xenograft (PODX) model of pancreatic cancer [ J ]. Acta Lab Animalis Sci Sin, 2018, 26 (03): 296 - 301.
[8] Rapic S, Vangestel C, Verhaeghe J, et al. Characterization of an orthotopic colorectal cancer mouse model and its feasibility for accurate quantification in positron emission tomography [J]. Mol Imag Biol, 2017, 19(5): 762-771.
[9] Zhu W, Cai M, Tong Z, et al. Overexpression of EIF5A2 promotes colorectal carcinoma cell aggressiveness by upregulating MTA1 through C-myc to induce epithelial-mesenchymal transition [J]. Gut. 2012, 61(4): 562-575.
[10] 张贺,陈薛,谭邓旭,等.基于肿瘤个体化治疗药物筛选的异种移植模型评价策略[J].中国实验动物学报. 2018(04): 523-527. Zhang H, Chen X, Tan DX, et al. Evaluation strategy of patient-derived xenograft models based on drug-screening of individualized treatment [J]. Acta Lab Anim Sci Sin, 2018, 26(04): 523-527.
[11] Monsky WL, Mouta Carreira C, Tsuzuki Y, et al. Role of host microenvironment in angiogenesis and microvascular functions in human breast cancer xenografts: Mammary fat pad versus cranial tumors1 [J]. Clin Cancer Res, 2002, 8(4):1008-1013.
[12] Sicklick JK, Leonard SY, Babicky ML, et al. Generation of orthotopic patient-derived xenografts from gastrointestinal stromal tumor [J]. J Transl Med. 2014, 12: 41.
[13] Murakami T, Murata T, Kawaguchi K, et al. Cervical cancer patient-derived orthotopic xenograft (PDOX) is sensitive to cisplatinum and resistant to Nab-paclitaxel [J]. Anticancer Res. 2017, 37(1): 61-66.
[14] Lang H, Béraud C, Bethry A, et al. Establishment of a large panel of patient-derived preclinical models of human renal cell carcinoma [J]. Oncotarget. 2016, 7(37): 59336.
[15] 王勤周,张成,李丽,等. 七甲川菁荧光染料在肝细胞癌移植模型活体成像中的应用 [J]. 实验动物与比较医学. 2018(04): 250-254.Wang QZ, Zhang C, Li L, et al. The Application of heptamethine cyanine dye in optical imaging of hepatocellular carcinoma transplantation model [J]. Lab Anim Comp Med, 2018, 38(4): 250-254.
[16] Shi C, Wu JB, Pan D. Review on near-infrared heptamethine cyanine dyes as theranostic agents for tumor imaging, targeting, and photodynamic therapy [J]. J Biomed Optics. 2016, 21(5): 50901.
[17] Balkwill F. The significance of cancer cell expression of the chemokine receptor.CXCR4 [J]. Semin Cancer Biol. 2004, 14(3): 171-179.
[18] 罗玉政,陈波,李红樱. 沉默IGF1R表达对胃癌细胞侵袭与迁移能力的影响及机制 [J]. 中国临床研究. 2018(02): 163-166.Luo YZ, Chen B, Li HY. Effect of silencing IGF1R expression on invasion and migration abilities of gastric cancer cells and its mechanism [J]. Chin J Clin Res 2018, 31(02): 163-166.
[19] 李倩玉,韩冬艳,蒋虹伟,等. p28、MMP2、β-catenin在胃癌及淋巴结转移癌中的表达 [J]. 诊断学理论与实践. 2016(02): 169-173.Li QY, Han DY, Jiang HW, et al. Expressions of p28, MMP2 and β-catenin proteins in human gastric carcinoma and metastatic lymph nodes [J]. J Diagn Concepts Practice, 2016, 15(02): 169-173.