Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, Rochester, New York, USA.
Stem Cells. 2011 Oct;29(10):1601-10. doi: 10.1002/stem.703.
Patients with chronic inflammatory disorders, such as rheumatoid arthritis, often have osteoporosis due to a combination of Tumor necrosis factor-induced increased bone resorption and reduced bone formation. To test if TNF inhibits bone formation by affecting the commitment and differentiation of mesenchymal stem cells (MSCs) into osteoblasts, we examined the osteogenic potential of MSCs from TNF transgenic (TNF-Tg) mice, a model of chronic inflammatory arthritis. MSC-enriched cells were isolated from bone marrow stromal cells using negative selection with anti-CD45 antibody coated magnetic beads. The expression profile of MSC surface markers the osteogenic, chondrogenic, and adipogenic properties of CD45(-) cells were confirmed by FACS and cell differentiation assays. MSC-enriched CD45(-) cells from TNF-Tg mice formed significantly decreased numbers of fibroblast and ALP(+) colonies and had a decreased expression of osteoblast marker genes. As TNF may upregulate ubiquitin ligases, which negatively regulate osteoblast differentiation, we examined the expression levels of several ubiquitin ligases and found that Wwp1 expression was significantly increased in MSC-enriched CD45(-) cells of TNF-Tg mice. Wwp1 knockdown rescued impaired osteoblast differentiation of TNF-Tg CD45(-) cells. Wwp1 promotes ubiquitination and degradation of JunB, an AP-1 transcription factor that positively regulates osteoblast differentiation. Injection of TNF into wild-type mice resulted in decreased osteoblast differentiation of MSCs and increased JunB ubiquitination, which was completely blocked in Wwp1(-/-) mice. Thus, Wwp1 targets JunB for ubiquitination and degradation in MSCs after chronic exposure to TNF, and inhibition of Wwp1 in MSCs could be a new mechanism to limit inflammation-mediated osteoporosis by promoting their differentiation into osteoblasts.
患有慢性炎症性疾病的患者，如类风湿关节炎，由于肿瘤坏死因子诱导的骨吸收增加和骨形成减少，常患有骨质疏松症。为了测试 TNF 是否通过影响间充质干细胞（MSCs）向成骨细胞的定向分化和分化来抑制骨形成，我们检查了 TNF 转基因（TNF-Tg）小鼠模型，一种慢性炎症性关节炎的模型中的 MSC 的成骨潜能。使用抗 CD45 抗体包被的磁珠进行阴性选择，从骨髓基质细胞中分离 MSC 富集细胞。通过 FACS 和细胞分化实验，确认 MSC 表面标志物的表达谱，MSC 富集的 CD45(-)细胞的成骨、成软骨和成脂特性。TNF-Tg 小鼠 MSC 富集的 CD45(-)细胞形成的成纤维细胞和 ALP(+)集落数量明显减少，成骨细胞标记基因的表达降低。由于 TNF 可能上调负向调节成骨细胞分化的泛素连接酶，我们检查了几种泛素连接酶的表达水平，发现 TNF-Tg 小鼠 MSC 富集的 CD45(-)细胞中 Wwp1 的表达显著增加。Wwp1 敲低可挽救 TNF-Tg CD45(-)细胞受损的成骨细胞分化。Wwp1 促进 JunB 的泛素化和降解，JunB 是一种正向调节成骨细胞分化的 AP-1 转录因子。TNF 注射到野生型小鼠中导致 MSC 中成骨细胞分化减少，JunB 泛素化增加，而在 Wwp1(-/-)小鼠中完全被阻断。因此，在慢性暴露于 TNF 后，Wwp1 在 MSC 中靶向 JunB 进行泛素化和降解，抑制 MSC 中的 Wwp1 可能是通过促进其分化为成骨细胞来限制炎症介导的骨质疏松症的新机制。
