Troubles and answers of identity authentication of human tumor cell lines established in China

doi:10.16098/j.issn.0529-1356.2019.02.010

Abstract

Abstract:

Objective To authenticate the identity of human tumor cell lines established in China and make sure the authenticated cell lines will be used in science and technology research in China. Methods We collected and extracted genomic DNA of those tumor cells, amplify short tandem repeat (STR) loci by PCR and performed electrophoretic analysis to get STR profiles (as the DNA finger print of the cell line). Comparing their STR profiles against international STR database and those of their donor tissues, crosscontaminated cell lines will be excluded from the service list. PCR detection of human papilloma virus type 18（HPV-18）fragment were performed to double check HeLa crosscontaminated cell lines. Suspected cells contaminated by T leukemia cells were checked by detection characteristics of T cells, such as expression of some surface markers. Results Altogether 46 samples of 37 human tumor cell lines were collected from different labs. All 6 cell lines established in our center showed identical STR profile with their original tissue or primary culture. Among the tumor cell lines established by others 8 cell lines showed unique STR profile, indicating authentic. The other 23 (62.2%) of them showed identical STR with HeLa or with high evaluation value (EV) value, HPV fragment were found in their genomic DNA indicating replacement by or hybrid with HeLa. Some tumor cell lines established in China were cross-contaminated by human colorectal cancer cell line HCT-8, cervical squamous cancer line SiHa, T leukemia cell line CCRF-CEM and a rat cell line. Conclusion A large proportion of human tumor cell lines established by Chinese scholars were cross-contaminated. So scientists must first do authenticate these cell lines before experiments or obtain cell lines from China Infra-structure of Cell Line Resource. Furthermore, when trying to establish cell lines, preserve original tissue/DNA for future authentication.

Key words: Short tandem repeat DNA typing, Cross-contamination, Characterization, PCR, Tumor cell, Cell culture, Human

BIAN Xiao-cui LIU Yu-qin YANG Zhen-li FENG Hai-liang. Troubles and answers of identity authentication of human tumor cell lines established in China[J]. Acta Anatomica Sinica, 2019, 50(2): 201-210.

References

［1］ Bian X, Yang Z, Feng H, et al. A combination of species identification and STR profiling identifies cross-contaminated cells from 482 human tumor cell lines［J］. Sci Rep, 2017,7(1): 9774.

［2］ Ye F, Chen C, Qin J, et al. Genetic profiling reveals an alarming rate of cross-contamination among human cell lines used in China［J］. FASEB J, 2015,29(10): 4268-4272.

［3］ Huang Y, Liu Y, Zheng C, et al. Investigation of cross-contamination and misidentification of 278 widely used tumor cell lines［J］. PLoS One, 2017,12(1): e0170384.

［4］ Feng H, Zhang Y, Liu K, et al. Intrinsic gene changes determine the successful establishment of stable renal cancer cell lines from tumor tissue［J］. Int J Cancer, 2017,140(11): 2526-2534.

［5］ Yang ZhL, Xu YL, Bian XC, et al. Faciliated primary culture and amplification of breast cancer cells and their biological properties［J］. Basic and Clinical Medicine, 2017,37(2): 224-229. (in Chinese)

杨振丽, 徐雅丽, 卞晓翠, 等. 乳腺癌组织细胞的高效原代培养及其特性［J］. 基础医学与临床, 2017,37(2): 224-229.

［6］ Wang R, Bian XC, Liu YQ, et al. Establishment and characterization of a laryngeal squamous cell carcinoma cell line［J］. Chinese Journal of Otorhinolaryngology Head and Neck Surgery, 2017,52(1): 44-48. (in Chinese)

王茹, 卞晓翠, 刘玉琴, 等. 喉鳞状细胞癌细胞系的建立及其生物学特性研究［J］. 中华耳鼻咽喉头颈外科杂志, 2017,52(1): 44-48.

［7］ Group of Cell Biology, Institute of Cancer Prevention and Treatment Chinese Acdemy of Medical Sciences. Establishment of human esophageal cancer epithelial cell line ［J］. National Medical Journal of China, 1976,56(7): 412-415. (in Chinese)

中国医学科学院肿瘤防治研究所细胞生物组. 人体食管癌上皮细胞株的建立［J］. 中华医学杂志, 1976,56(7): 412-415.

［8］ Han YL, Wei F, Xu X, et al. Establishment and comparative genomic hybridization analysis of human esophageal carcinomas cell line EC9706 ［J］. Chinese Journal of Medical Genetics, 2002,19(6): 455-457. (in Chinese)

韩亚玲, 魏芳, 徐昕, 等. 人食管鳞癌细胞系EC9706的建立及其比较基因组杂交分析［J］. 中华医学遗传学杂志, 2002,19(6): 455-457.

［9］ Zhao XB, Zhang YD, Su YK, et al. Establishment and characterization of a Xeno grafted human cervical carcinoma in nude mice and homologous cell line in vitro［J］. Journal of Shandong University(Health Sciences), 1998,(2): 155-160. (in Chinese)

赵兴波, 张义读, 苏应宽. 人子宫颈癌细胞系HCC-94的建立及其生物学特性［J］. 山东大学学报(医学版), 1998,(2): 155-160.

［10］Li MZh, Zhao LJ, Li XP, et al. Establishment of a new cell line of primary human endometrial carcinoma by improved explants culture and enzyme-cloning digestion［J］. Chinese Journal of Clinical Obstetrics and Gynecology, 2014,(2): 141-144. (in Chinese)

李明珠, 赵丽君, 李小平, 等. 应用微小组织块法培养和胰酶滤纸片克隆消化法建立人子宫内膜癌细胞系的研究［J］. 中国妇产科临床杂志, 2014,(2): 141-144.

［11］Wang ShL, Li F. Establishment and characterization of human brain malignant glioblastoma cell lines TJ899 and TJ905 in vitro ［J］. Tianjin Medical Journal, 1996,(7): 416-418. (in Chinese)

王淑兰, 李峰. 人脑恶性胶质母细胞瘤体外细胞系TJ899及TJ905的建立及其特征［J］. 天津医药, 1996,(7): 416-418.

［12］Du ZW, Xu GD, Wang Y, et al. Establishment and characterization of human brain malignant glioma cell line SHG-44 in vitro［J］.Journal of Medical Research, 1984,6(4): 241-243. (in Chinese)

杜子威, 徐庚达, 王尧, 等.人脑恶性胶质瘤体外细胞系SHG-44的建立及其特征［J］. 医学研究杂志, 1984,6(4): 241-243.

［13］Zhang ZX, Qian LJ, Zhang YY, et al. Establishment of human cecum undifferentiated adenocarcinoma cell line Hce-8693 and its biological characteristics［J］. Journal of Chinese Oncology, 1989,(3): 1-4. (in Chinese)

张宗显, 钱丽娟, 张奕荫, 等. 人盲肠未分化腺癌细胞系Hce 8693的建立及其生物学特性［J］. 肿瘤学杂志, 1989,(3): 1-4.

［14］Wang KH. Establishment of human gastric poorly differentiated mucinous adenocarcinoma cell line MGc80-3 and preliminary observation of its biological characteristics［J］. Acta Biologiae Experimentalis Sinica, 1983,(3): 20-30. (in Chinese)

王凯华. 人体胃低分化黏液腺癌细胞系MGc80-3的建立及其生物学特性的初步观察［J］. 实验生物学报, 1983,(3): 20-30.

［15］Cai YH, Gu YZh, Zhang JQ, et al. Establishment of human gastric cancer cell line BGC-823［J］. Journal of Peking University(Health Sciences), 1986,18(2): 136-139. (in Chinese)蔡玉晖, 谷钰之, 张嘉庆, 等. 人体胃癌细胞系BGC-823的建立［J］. 北京大学学报(医学版), 1986,18(2): 136-139.

［16］Lin ChH, Fu ZhM, Liu YL, et al. The establishment of human gastric carcinoma cell line(SGC-7901) ［J］. Tumor, 1981,1(1): 1-4. (in Chinese)

林超鸿, 富志民, 刘亚伦, 等. 人体胃腺癌细胞株SGC-7901的建立［J］. 肿瘤, 1981,1(1): 1-4.

［17］Chen RM, Zhu DH, Ye XZh, et al. Establishment and characterization of human hepatocellular carcinoma cell line (BEL-7402) in vitro ［J］. Chinese Science Bulletin, 1975,20(9): 434-436. (in Chinese)

陈瑞铭, 朱德厚, 叶秀珍, 等. 人体肝癌体外细胞株(BEL-7402)的建立及其特征［J］. 科学通报, 1975,20(9): 434-436.

［18］Liu B, Wang ZhM, Li ZhT, et al. Establishment and characterization of cholangiocarcinoma cell line HCCC-9810［C］.21 Century Surgical Symposium, 2002. (in Chinese)

刘博, 王占民, 李兆亭, 等. 胆管细胞癌细胞系HCCC-9810的建立及特性研究［C］. 21世纪外科研讨会, 2002.

［19］Wu ZhY, Huang JSh, Huang XB. Brief report on human liver cell culture ［J］. Jiangsu Medical Journal, 1978,(9): 8-11. (in Chinese)

吴忠一, 黄建生, 黄锡兵. 人体肝脏细胞培养简报［J］. 江苏医药, 1978,(9): 8-11.

［20］Dong RCH, Zhou RH, Lü FD, et al. Establishment of human hepatocellular carcinoma cell line SMMC-7721 and preliminary observation of its biological characteristics ［J］. Academic Journal of Second Military Medical University, 1980,(1): 5-9. (in Chinese)

董荣春, 周荣华, 吕发度, 等. SMMC-7721人体肝癌细胞株的建立及其生物学特性的初步观察［J］. 第二军医大学学报, 1980,(1): 5-9.

［21］Mou HZh, Xu ShH, Zhang YY, et al. The establishment of human ovarian carcinoma cell line HO-8901 and its characteristics［J］. Chinese Journal of Obstetrics and Gynecology, 1994,(3): 162-164. (in Chinese)

牟瀚舟, 许沈华, 张奕荫, 等. 人卵巢癌细胞系HO-8910的建立及其生物学特性［J］. 中华妇产科杂志, 1994,(3): 162-164.

［22］Xu ShH, Qian LJ. Establishment of a highly metastatic human ovarian cancer cell line HO-8910PM and its biological characteristics［J］. Chinese Journal of Pathology, 1998,27(6): 451-452. (in Chinese)

许沈华, 钱丽娟. 高转移人卵巢癌细胞系HO-8910PM的建立及其生物学特性［J］. 中华病理学杂志, 1998,27(6): 451-452.

［23］Xiong ShG. Establishment of human ovarian cancer cell line COC1 COC2 and effects of autocrine TGF-beta on growth and tumor immunity［D］. Peking Union Medical College, 1993.(in Chinese)

熊世钢. 人卵巢癌细胞株COC1 COC2的建立及自分泌TGF-β促生长和对肿瘤免疫的影响［D］. 中国协和医科大学, 1993.

［24］Chen HZh, Cai HB. Establishment of human ovarian cancer cell line COC1/DDP and its biological characteristics ［J］. Medical Scientific Research in Wuhan, 1997,(2):9.(in Chinese)

陈惠祯, 蔡红兵. 人卵巢癌耐药细胞亚株COC1/DDP的建立及其生物学特性的研究［J］. 武汉市医学科研, 1997,(2):9.

［25］He RG, Xu XQ, Zhou XJ, et al. The establishment and some biological characteristics of a squamous cell cancer cell line of human tongue (Tca 8113) ［J］. Tumor, 1983,3(3): 97-100. (in Chinese)

何荣根, 徐秀祺, 周晓健, 等. 人舌鳞状细胞癌Tca8113细胞系的建立及其生物学特性［J］. 肿瘤, 1983,3(3): 97-100.

［26］Cancer Research on Prevention and Treatment. Establishment of human nasopharyngeal carcinoma epithelioid cell lines and spindle cell lines［J］. 1978,6(3): 18-23. (in Chinese)中国医学科学院肿瘤研究所病毒室. 人体鼻咽癌上皮样细胞株和梭形细胞株的建立［J］. 肿瘤防治研究, 1978,6(3): 18-23.

［27］Gu ShY, Zhao WP, Zeng Y, et al. An epithelsal cell line established from pooly differentiated nasopharyngeal carcinoma［J］. Cancer, 1983,2(2): 70-72. (in Chinese)

谷淑燕, 赵文平, 曾毅, 等. 从低分化鼻咽癌患者建立鼻咽癌上皮细胞株［J］. 癌症, 1983,2(2): 70-72.

［28］Wang H, Lai BT, Zhan XP, et al. Establishment of five lung cancer cell lines and comparison of their biological characteristics［J］. National Medical Journal of China, 1985,65(4): 208-211. (in Chinese)

汪蕙, 赖百塘, 占秀萍, 等. 5个人肺癌细胞系的建立及其生物学特性的比较［J］. 中华医学杂志, 1985,65(4): 208-211.

［29］Liang MD, Jia W, Hu MY, et al. Establishment and biological characteristics of Gejiu lung adenocarcinoma cell line (GLC-82) ［J］. Journal of Kunming Medical University, 1984,5(1): 1-6. (in Chinese)

梁明达, 贾伟, 胡美英, 等. 个旧肺腺癌细胞系(GLC-82)的建立及其生物学特性［J］. 昆明医科大学学报, 1984,5(1): 1-6.

［30］Wu ShF, Su JZh, Wang EZh, et al. Establishment of human lung adenocarcinoma cell line SPC-A-1 and its biological characteristics［J］. Science in China (Series B), 1982, 12(10):913-921. (in Chinese)

吴善芳，苏建中，王恩忠，等. 人体肺腺癌细胞系——SPC-A-1的建立及其生物学特性［J］. 中国科学（B辑）, 1982, 12(10):913-921.

［31］Yu M, Selvaraj SK, Liang-Chu MM, et al. A resource for cell line authentication, annotation and quality control［J］. Nature, 2015,520(7547): 307-311.

［32］Yang ZhL, Bian XC, Feng HL, et al. Detection of six main species of viruses in human cell lines［J］. Chinese Journal of Experimental and Clinical Virology, 2017,31(5): 457-461. (in Chinese)

杨振丽, 卞晓翠, 冯海凉, 等. 人源细胞系中6种主要病毒的检测［J］. 中华实验和临床病毒学杂志, 2017,31(5): 457-461.

［33］Strong MJ, Baddoo M, Nanbo A, et al. Comprehensive high-throughput RNA sequencing analysis reveals contamination of multiple nasopharyngeal carcinoma cell lines with HeLa cell genomes［J］. J Virol, 2014,88(18): 10696-10704.

［34］MacLeod RA, Dirks WG, Matsuo Y, et al. Widespread intraspecies cross-contamination of human tumor cell lines arising at source［J］. Int J Cancer, 1999,83(4): 555-563.

［35］Drexler HG, Dirks WG, Matsuo Y, et al. False leukemia-lymphoma cell lines: an update on over 500 cell lines［J］. Leukemia, 2003,17(2): 416-426.

［36］Capes-Davis A, Theodosopoulos G, Atkin Ⅰ, et al. Check your cultures! A list of cross-contaminated or misidentified cell lines［J］. Int J Cancer, 2010,127(1): 1-8.

［37］Identity crisis［J］. Nature, 2009,457(7232): 935-936.

［38］Feng HL, Wang ChJ, Gu B, et al. Establishment of a simian virus 40 T antigen transfected human umbilical vein endothelial cell line PUMC-HUVEC-T1［J］.Acta Anatomica Sinica, 2013, 44(2): 199-203. (in Chinese)

冯海凉, 王春景, 顾蓓, 等. 猿猴病毒40 T抗原转化的人脐静脉内皮细胞系PUMC-HUVEC-T1的建立［J］. 解剖学报, 2013, 44(2): 199-203.

［39］Americal Type Culture Collection Standards Development Organization Workgroup ASN-0002. Cell line misidentification: the beginning of the end［J］. Nat Rev Cancer, 2010,10(6): 441-448.

［40］Dirks WG, Drexler HG. Online verification of human cell line identity by STR DNA typing［J］. Methods Mol Biol, 2011,731: 45-55.

［41］Liu YQ. Shirt tandem repeat can be used as a gold standard for cell identity authentication. ［J］. Chinese Journal of Pathology, 2013,42(9): 647-648. (in Chinese)

刘玉琴. 短串联重复序列可作为细胞身份认证的金标准［J］. 中华病理学杂志, 2013,42(9): 647-648.

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