%0 Journal Article
%T Cell culture and passaging alters gene expression pattern and proliferation rate in rheumatoid arthritis synovial fibroblasts
%A Elena Neumann
%A Birgit Riepl
%A Anette Knedla
%A Stephanie Lef¨¨vre
%A Ingo H Tarner
%A Joachim Grifka
%A Jurgen Steinmeyer
%A Jurgen Sch£¿lmerich
%A Steffen Gay
%A Ulf M¨¹ller-Ladner
%J Arthritis Research & Therapy
%D 2010
%I BioMed Central
%R 10.1186/ar3010
%X RASF were passaged for up to 8 passages. RNA was isolated after each passage and cDNA arrays were performed to evaluate the RNA expression pattern during passaging. In addition, doubling time of the cells was also measured.From passages 2-4, mRNA expression did not change significantly. Gene expression in RASF started to change in passages 5-6 with 7-10% differentially expressed genes. After passages 7-8, more than 10% of the genes were differentially expressed. The doubling rate was constant for up to 5 passages and decreased after passages 6-8. After freezing, gene expression of the second passage is comparable to gene expression prior to freezing.The results of this study show, that experiments, which examine gene expression of RASF and shall reflect or imitate an in vivo situation, should be limited to early culture passages to avoid cell culture effects. It is not necessary to stop culturing SF after a few passages, but to keep the problems of cell culture in mind to avoid false positive results. Especially, when large-scale screening methods on mRNA level are used. Of note, freezing does not affect gene expression substantially.Predominant features of rheumatoid arthritis (RA) are synovial hyperplasia, synovial cell activation and articular inflammation associated with subsequent cartilage and bone destruction [1]. In this scenario, activated synovial fibroblasts (SF) are key players in joint destruction at the site of invasion into articular cartilage and bone [1-5]. They maintain their aggressive phenotype towards cartilage even when primarily cultured and thereafter co-implanted together with normal human cartilage into immunodeficient severe combined immunodeficient mice (SCID) mice for an extended period of time [5].To inhibit the progressive growth at the invasion zone followed by cartilage and bone degradation without interfering with physiologic matrix remodeling, identification of pathways operative specifically in RASF and not in SF of other origin (
%U http://arthritis-research.com/content/12/3/R83