Department of Surgical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
Department of Gastroenterology and Rheumatology, The Fukushima Medical University School of Medicine, Fukushima, Japan.
Lab Invest. 2015 Feb;95(2):207-22. doi: 10.1038/labinvest.2014.143. Epub 2014 Dec 8.
Many commercially available cell lines have been in culture for ages, acquiring phenotypes that differ from the original cancers from which these cell lines were derived. Therefore, research on new cell lines could improve the success rates of translational research in cancer. We have developed methods for the isolation and culture of human pancreatic ductal adenocarcinoma (PDAC) cells from murine xenografts of human PDAC. We hypothesize that phenotypes of PDAC cells are modified by in vitro culture conditions over time and by in vivo implantation. Patient-derived xenografts were created in immunodeficient mice using surgically resected tumor specimens. These murine xenografts were then used to establish human PDAC cell lines in culture. Earlier (<5) passage and later (>20) passage cell lines were evaluated separately regarding proliferation, cell cycle, genetic mutations, invasiveness, chemosensitivity, tumorigenesis, epithelial-mesenchymal transition (EMT) status, and proteomics. Later passage cells accelerated their doubling time and colony formation, and were more concentrated in the G0/G1 phase and less in the G2/M checkpoint phase. Later passage cells were more sensitive to gemcitabine and 5-fluorouracil than earlier passage cells, but all four new cell lines were more chemo-resistant compared with commercial ATCC cell lines. EMT induction was observed when establishing and passaging cell lines in vitro and furthermore by growing them as subcutaneous tumors in vivo. This study demonstrates a novel approach to the establishment of PDAC cell lines and observes a process by which newly established cell lines undergo phenotypic changes during in vitro culture and in vivo tumorigenesis. This may help explain differences of treatment effects often observed between experiments conducted in vitro, in vivo, and in human clinical trials.
许多市售的细胞系已经培养了很长时间，获得了与这些细胞系来源的原始癌症不同的表型。因此，研究新的细胞系可以提高癌症转化研究的成功率。我们已经开发了从人胰腺导管腺癌（PDAC）的小鼠异种移植中分离和培养人 PDAC 细胞的方法。我们假设 PDAC 细胞的表型会随着时间的推移和体内植入而被体外培养条件所改变。使用手术切除的肿瘤标本在免疫缺陷小鼠中创建患者来源的异种移植物。然后，将这些小鼠异种移植物用于在培养中建立人 PDAC 细胞系。早期（<5 代）和晚期（>20 代）的细胞系分别在增殖、细胞周期、基因突变、侵袭性、化疗敏感性、致瘤性、上皮-间充质转化（EMT）状态和蛋白质组学方面进行了评估。晚期传代细胞的倍增时间和集落形成加快，G0/G1 期更多，G2/M 检查点期更少。晚期传代细胞对吉西他滨和 5-氟尿嘧啶的敏感性高于早期传代细胞，但与商业 ATCC 细胞系相比，所有四个新的细胞系的化疗耐药性更高。在体外建立和传代细胞系以及在体内作为皮下肿瘤生长时观察到 EMT 诱导。这项研究展示了一种建立 PDAC 细胞系的新方法，并观察到新建立的细胞系在体外培养和体内肿瘤发生过程中经历表型变化的过程。这可能有助于解释在体外、体内和人体临床试验中进行的实验中经常观察到的治疗效果差异。
