Wang Yanqiang, Li Wei, Phay John E, Shen Rulong, Pellegata Natalia S, Saji Motoyasu, Ringel Matthew D, de la Chapelle Albert, He Huiling
1 Human Cancer Genetics Program and Department of Cancer Biology and Genetics, The Ohio State University Comprehensive Cancer Center, The Ohio State University , Columbus, Ohio.
2 Department of Surgery, The Ohio State University Comprehensive Cancer Center, The Ohio State University , Columbus, Ohio.
Thyroid. 2016 Aug;26(8):1131-40. doi: 10.1089/thy.2015.0518. Epub 2016 Jul 6.
Cell models are key instruments for in vitro studies of the thyroid. Permanent thyroid cell lines that are widely used in laboratory research typically originate from tumors. For many purposes, it is desirable to compare tumor cells with cells originating from normal tissue. However, such cultures grow slowly, have a highly limited life-span, and are known to lose their thyroid characteristics. The aim of the present study was to type coding and noncoding thyroid markers in different culture systems in an attempt to determine the optimal conditions for in vitro experimentation.
Human primary thyroid cells were isolated from histologically non-tumorous tissues. Two alternative media (6H and h7H) were used. The morphology and behavior of the ensuing monolayer (two-dimensional) cultures was monitored by microscopy. The expression of key thyroid-related genes (n = 9) was monitored by reverse transcription polymerase chain reaction on days 8, 21, and 43 after initiation. As a pilot study, the same markers were studied in a three-dimensional hanging-drop culture system.
In the cultures with 6H or h7H medium, the primary thyroid cells displayed growth in numbers and size. Most cells retained the main morphological characteristics of thyroid cells throughout the first two weeks of culture, and fibroblast-like cells appeared around day 19. By day 21, most thyroid gene markers were retained, but by day 43, several markers were no longer present. The lncRNA transcripts PTCSC2 (spliced) and PTCSC3 were the first to disappear. There were no fundamental differences between the two media in the early period of culture. In the three-dimensional system, most thyroid markers were retained by day 21.
Cultures of thyroid cells retain many thyroid characteristics up to day 21. Thereafter, fibroblast-like dedifferentiated cells begin to dominate.
细胞模型是甲状腺体外研究的关键工具。实验室研究中广泛使用的永久性甲状腺细胞系通常源自肿瘤。出于多种目的，将肿瘤细胞与源自正常组织的细胞进行比较是很有必要的。然而，此类培养物生长缓慢，寿命极为有限，且已知会丧失其甲状腺特征。本研究的目的是在不同培养系统中对编码和非编码甲状腺标志物进行分型，以试图确定体外实验的最佳条件。
从组织学上非肿瘤性组织中分离出人原代甲状腺细胞。使用了两种替代培养基（6H和h7H）。通过显微镜监测随后的单层（二维）培养物的形态和行为。在起始培养后的第8天、第21天和第43天，通过逆转录聚合酶链反应监测关键甲状腺相关基因（n = 9）的表达。作为一项初步研究，在三维悬滴培养系统中研究了相同的标志物。
在使用6H或h7H培养基的培养物中，原代甲状腺细胞数量和大小均有增长。在培养的前两周，大多数细胞保留了甲状腺细胞的主要形态特征，在第19天左右出现了成纤维细胞样细胞。到第21天，大多数甲状腺基因标志物得以保留，但到第43天，几种标志物不再存在。lncRNA转录本PTCSC2（剪接型）和PTCSC3是最先消失的。在培养早期，两种培养基之间没有根本差异。在三维系统中，到第21天大多数甲状腺标志物得以保留。
甲状腺细胞培养物在第21天之前保留许多甲状腺特征。此后，成纤维细胞样去分化细胞开始占主导地位。
