Oh Namgil, Kim Sangho, Hosoya Kenji, Okumura Masahiro
Laboratory of Veterinary Surgery, Department of Veterinary Clinical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan.
J Vet Med Sci. 2014 May;76(5):629-36. doi: 10.1292/jvms.13-0482. Epub 2014 Jan 13.
The suppressive effects of nonsteroidal anti-inflammatory drugs (NSAIDs) on the bone healing process have remained controversial, since no clinical data have clearly shown the relationship between NSAIDs and bone healing. The aim of this study was to assess the compensatory response of canine bone marrow-derived mesenchymal stem cells (BMSCs) to several classes of NSAIDs, including carprofen, meloxicam, indomethacin and robenacoxib, on osteogenic differentiation. Each of the NSAIDs (10 µM) was administered during 20 days of the osteogenic process with human recombinant IL-1β (1 ng/ml) as an inflammatory stimulator. Gene expression of osteoblast differentiation markers (alkaline phosphatase and osteocalcin), receptors of PGE2 (EP2 and EP4) and enzymes for prostaglandin (PG) E2 synthesis (COX-1, COX-2, cPGES and mPGES-1) was measured by using quantitative reverse transcription-polymerase chain reaction. Protein production levels of alkaline phosphatase, osteocalcin and PGE2 were quantified using an alkaline phosphatase activity assay, osteocalcin immunoassay and PGE2 immunoassay, respectively. Histologic analysis was performed using alkaline phosphatase staining, von Kossa staining and alizarin red staining. Alkaline phosphatase and calcium deposition were suppressed by all NSAIDs. However, osteocalcin production showed no significant suppression by NSAIDs. Gene expression levels of PGE2-related receptors and enzymes were upregulated during continuous treatment with NSAIDs, while certain channels for PGE2 synthesis were utilized differently depending on the kind of NSAIDs. These data suggest that canine BMSCs have a compensatory mechanism to restore PGE2 synthesis, which would be an intrinsic regulator to maintain differentiation of osteoblasts under NSAID treatment.
非甾体抗炎药（NSAIDs）对骨愈合过程的抑制作用一直存在争议，因为尚无临床数据明确显示NSAIDs与骨愈合之间的关系。本研究的目的是评估犬骨髓间充质干细胞（BMSCs）对几类NSAIDs（包括卡洛芬、美洛昔康、吲哚美辛和罗贝考昔）在成骨分化方面的代偿反应。在成骨过程的20天中，将每种NSAIDs（10µM）与人重组IL-1β（1ng/ml）作为炎症刺激剂一起给药。通过定量逆转录-聚合酶链反应测量成骨细胞分化标志物（碱性磷酸酶和骨钙素）、PGE2受体（EP2和EP4）以及前列腺素（PG）E2合成酶（COX-1、COX-2、cPGES和mPGES-1）的基因表达。分别使用碱性磷酸酶活性测定、骨钙素免疫测定和PGE2免疫测定对碱性磷酸酶、骨钙素和PGE2的蛋白质产生水平进行定量。使用碱性磷酸酶染色、冯·科萨染色和茜素红染色进行组织学分析。所有NSAIDs均抑制碱性磷酸酶和钙沉积。然而，NSAIDs对骨钙素的产生没有显著抑制作用。在连续使用NSAIDs治疗期间，PGE2相关受体和酶的基因表达水平上调，而PGE2合成的某些途径根据NSAIDs的种类而有不同利用。这些数据表明犬BMSCs具有恢复PGE2合成的代偿机制，这将是在NSAIDs治疗下维持成骨细胞分化的内在调节因子。
