Modified House-Keeping Gene Expression in a Rat Tail Compression Loading-Induced Disc Degeneration Model

作者：Yurube Takashi; Takada Toru; Hirata Hiroaki; Kakutani Kenichiro; Maeno Koichiro; Zhang Zhongying; Yamamoto Junya; Doita Minoru; Kurosaka Masahiro; Nishida Kotaro^*

来源：Journal of Orthopaedic Research, 2011, 29(8): 1284-1290.

DOI：10.1002/jor.21406

• intervertebral disc degeneration
• animal model
• house-keeping gene
• real-time RT-PCR mRNA-quantification
• spine

House-keeping genes (HKGs) are generally used as endogenous controls for molecular normalization in quantitative PCR analysis. However, whether all the so-called HKGs are useful for intervertebral disc research is controversial. Our objective was, using a prevalidated rat tail static compression loading-induced disc degeneration model, to clarify the feasibility of common HKGs for gene-quantification in the nucleus pulposus cells. In real-time RT-PCR for five HKGs [beta-actin, beta-glucuronidase, beta-2 microglobulin, glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and lactate dehydrogenase A (LDHA)], static compression at 1.3 MPa for up to 56 days demonstrated messenger RNA (mRNA) expression levels of consistent beta-2 microglobulin and GAPDH, slightly up-regulated beta-glucuronidase, and fairly down-regulated beta-actin and LDHA. Especially, beta-actin had a drastic suppression to 0.15-fold in the loaded relative to unloaded discs at 7 days. In immunofluorescence, beta-actin showed a significant down-regulation to almost undetectable levels from 28 days, while GAPDH was constantly detected throughout. beta-Actin mRNA and protein-distribution are thought to be affected by the loading treatment, however, GAPDH mRNA and protein-distribution can retain relatively stable expressions. Under prolonged static compression, beta-actin and probably LDHA are inappropriate, and GAPDH is a feasible HKG as internal references in the disc cells.

• 出版日期2011-8