Identification of suitable reference genes in bone marrow stromal cells from osteoarthritic donors
| Autor: | Henriette Bretschneider, Maik Stiehler, Theresa Schildberg, Juliane Rauh |
|---|---|
| Rok vydání: | 2013 |
| Předmět: |
Stromal cell
Gene Expression Bone Marrow Cells Biology Real-Time Polymerase Chain Reaction Importin 8 Reference genes Osteoarthritis Gene expression medicine Humans RNA Messenger Gene Aged Medicine(all) Genetics Nuclear Proteins RNA-Binding Proteins Mesenchymal Stem Cells Cell Biology General Medicine Middle Aged TATA-Box Binding Protein beta Karyopherins Molecular biology Neoplasm Proteins Reverse transcription polymerase chain reaction Real-time polymerase chain reaction medicine.anatomical_structure Bone marrow Algorithms Developmental Biology |
| Zdroj: | Stem Cell Research. 11:1288-1298 |
| ISSN: | 1873-5061 |
| DOI: | 10.1016/j.scr.2013.08.015 |
| Popis: | Bone marrow stromal cells (BMSCs) are key cellular components for musculoskeletal tissue engineering strategies. Furthermore, recent data suggest that BMSCs are involved in the development of Osteoarthritis (OA) being a frequently occurring degenerative joint disease. Reliable reference genes for the molecular evaluation of BMSCs derived from donors exhibiting OA as a primary co-morbidity have not been reported on yet. Hence, the aim of the study was to identify reference genes suitable for comparative gene expression analyses using OA-BMSCs. Passage 1 bone marrow derived BMSCs were isolated from n=13 patients with advanced stage idiopathic hip osteoarthritis and n=15 age-matched healthy donors. The expression of 31 putative reference genes was analyzed by quantitative reverse transcription polymerase chain reaction (qRT-PCR) using a commercially available TaqMan(®) assay. Calculating the coefficient of variation (CV), mRNA expression stability was determined and afterwards validated using geNorm and NormFinder algorithms. Importin 8 (IPO8), TATA box binding protein (TBP), and cancer susceptibility candidate 3 (CASC3) were identified as the most stable reference genes. Notably, commonly used reference genes, e.g. beta-actin (ACTB) and beta-2-microglobulin (B2M) were among the most unstable genes. For normalization of gene expression data of OA-BMSCs the combined use of IPO8, TBP, and CASC3 gene is recommended. |
| Databáze: | OpenAIRE |
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