Abstract：Objective To establish a more rapid and convenient method for primary culture of renal tumor and normal renal tubular epithelial cells. Methods Fresh clear renal cell carcinoma tissue and renal cortical tissue from patients undergoing radical nephrectomy and partial nephrectomy in Beijing Chaoyang Hospital, Capital Medical University from January to March 2019 were selected. The tumor and normal primary cells were purified by mechanical segmentation, collagenase digestion and continuous cell screening and purification. The genetic background of the two primary cells was identified by transcriptomic sequencing (RNA-seq) technique, the expression of cell markers was identified by immunocytochemistry, and the ultrastructure of the primary cells was observed by transmission electron microscopy. Finally, the tumorigenesis characteristics of tumor cells were identified by the model of nude mice bearing tumors. Results A total of three human proximal tubular epithelial primary cells and eight renal clear cell carcinoma primary cells were isolated and purified. RNA-seq showed that the sequencing results were consistent with the genetic background of the two types of cells, and immunocytochemistry could identify the expression of marker proteins in the proximal renal tubular epithelial cells. Two kinds of cell specific ultrastructure could be observed by transmission electron microscope. The tumor bearing model of nude mice confirmed the tumorigenic properties of primary renal cell carcinoma cells. Conclusion A rapid and convenient processes and methods for primary culture of renal carcinoma cells and renal tubular epithelial cells is successfully established.
周晓光 王浩舟 李彦生 宋黎明. 一种基于RNA-seq验证的人肾癌细胞和正常细胞原代培养方法[J]. 中国医药导报, 2020, 17(26): 4-7,25.
ZHOU Xiaoguang WANG Haozhou LI Yansheng SONG Liming. A primary culture method of human renal carcinoma cells and normal cells based on RNA-seq validation. 中国医药导报, 2020, 17(26): 4-7,25.
[1] Atkins MB，Tannir NM. Current and emerging therapies for first-line treatment of metastatic clear cell renal cell carcinoma [J]. Cancer Treat Rev，2018，70：127-137.
[2] Brugarolas J. Molecular genetics of clear-cell renal cell carcinoma [J]. J Clin Oncol，2014，32（18）：1968-1976.
[3] Beroukhim R，Brunet JP，Di Napoli A，et al. Patterns of gene expression and copy-number alterations in von-hippel lindau disease-associated and sporadic clear cell carcinoma of the kidney [J]. Cancer Res，2009，69（11）：4674-4681.
[4] Lee J，Kotliarova S，Kotliarov Y，et al. Tumor stem cells derived from glioblastomas cultured in bFGF and EGF more closely mirror the phenotype and genotype of primary tumors than do serum-cultured cell lines [J]. Cancer Cell，2006，9（5）：391-403.
[5] van Staveren WC，Solís DY，Hébrant A，et al. Human cancer cell lines：Experimental models for cancer cells in situ? For cancer stem cells? [J]. Biochim Biophys Acta，2009，1795（2）：92-103.
[6] Pollard SM，Yoshikawa K，Clarke ID，et al. Glioma stem cell lines expanded in adherent culture have tumor-specific phenotypes and are suitable for chemical and genetic screens [J]. Cell Stem Cell，2009，4（6）：568-580.
[7] van de Wetering M，Francies HE，Francis JM，et al. Prospective derivation of a living organoid biobank of colorectal cancer patients [J]. Cell，2015，161（4）：933-945.
[8] Li XN，Wang ZJ，Ye CX，et al. Circular RNA circVAPA is up-regulated and exerts oncogenic properties by sponging miR-101 in colorectal cancer [J]. Biomed Pharmacother，2019（112）：108611.
[9] Wang M，Li X，Zhang J，et al. AHNAK2 is a Novel Prognostic Marker and Oncogenic Protein for Clear Cell Renal Cell Carcinoma [J]. Theranostics，2017，7（5）：1100-1113.
[10] Xu S，Zhang H，Chong Y，et al. YAP Promotes VEGFA Expression and Tumor Angiogenesis Though Gli2 in Human Renal Cell Carcinoma [J]. Arch Med Res，2019，50（4）：225-233.
[11] Tolkach Y，Ellinger J，Kremer A，et al. Apelin and apelin receptor expression in renal cell carcinoma [J]. Br J Cancer，2019，120（6）：633-639.
[12] 方开云，石明隽，肖瑛，等.原代肾小管上皮细胞的培养和鉴定[J].贵阳医学院学报，2008，33（2）：136-138.
[13] 余德芊，王富英，马元芬，等.微分离方法原代培养大鼠肾近曲小管上皮细胞[J].遵义医学院学报，2009，32（5）：448-450.
[14] 邢传平，钱震，李宁.肾细胞癌的组织病理和超微结构观察[J].电子显微学报，1997（5）：62-66.
[15] 许红民，李维华.肾细胞癌的超微结构分型[J].中国人民解放军军医进修学院学报，1990（3）：257-259，284.
[16] Abubakar MB，Gan SH. Molecular Targets in Advanced Therapeutics of Cancers：The Role of Pharmacogenetics [J]. Oncology，2016，91（1）：3-12.
[17] Okada S，Vaeteewoottacharn K，Kariya R. Application of Highly Immunocompromised Mice for the Establishment of Patient-Derived Xenograft （PDX） Models [J]. Cells，2019，8（8）：889.
[18] Meehan TF，Conte N，Goldstein T，et al. PDX-MI：Minimal Information for Patient-Derived Tumor Xenograft Models [J]. Cancer Res，2017，77（21）：e62-e66.
[19] 王梨名，陈佳，陈客宏，等.小鼠近端肾小管上皮细胞原代培养及鉴定[J].生理学报，2018，70（4）：406-412.
[20] 王裕，杨霞，高继萍，等.小鼠肾小管上皮细胞的培养和鉴定[J].山西医科大学学报，2014，45（11）：1106-1109， 1119.
[21] Vesey DA，Qi W，Chen X，et al. Isolation and primary culture of human proximal tubule cells [J]. Methods Mol Biol，2009，466：19-24.
[22] Qi W，Johnson DW，Vesey DA，et al. Isolation，propagation and characterization of primary tubule cell culture from human kidney [J]. Nephrology（Carlton），2007，12（2）：155-159.
[22] Valente MJ，Henrique R，Costa VL，et al. A rapid and simple procedure for the establishment of human normal and cancer renal primary cell cultures from surgical specimens [J]. PLoS One，2011，6（5）：e19337.
[24] Lobo NC，Gedye C，Apostoli AJ，et al. Efficient generation of patient-matched malignant and normal primary cell cultures from clear cell renal cell carcinoma patients：clinically relevant models for research and personalized medicine [J]. BMC Cancer，2016，16：485.
2020, Vol. 17 
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