Endocrine Research Unit, College of Medicine, Mayo Clinic, Rochester, MN 55905, USA.
J Bone Miner Res. 2012 Apr;27(4):891-901. doi: 10.1002/jbmr.1502.
A complex network of transcription factors contributes to the establishment and maintenance of the osteoblastic phenotype. Although relatively few transcription factors, such as Runx2 and osterix, are essential to the process of osteoblastic differentiation, others serve the purpose of fine-tuning in response to various environmental and hormonal cues. The nuclear receptor (NR) superfamily of transcription factors are involved in numerous aspects of bone biology. In this study, we characterized the expression pattern of the entire NR superfamily in differentiating primary murine calvarial cells in order to identify novel NR regulatory patterns. Dynamic patterns of NR expression were observed throughout the differentiation process. Interestingly, retinoic acid receptor-related orphan receptor β (Rorβ) expression was markedly suppressed at later stages of differentiation. To gain further insight into the function of NRs in bone biology, the NR superfamily was also profiled in mouse bone marrow precursor cells isolated from either young (6-month) or aging, osteoporotic (18-22-month) mice. Of interest, Rorβ was potently overexpressed in the aged cohort. Collectively, these data provided evidence that Rorβ expression is inversely correlated with osteogenic potential, suggesting Rorβ may be an important and unexplored regulator of osteogenesis. To validate this hypothesis, a cell model stably expressing Rorβ in mouse osteoblastic MC3T3-E1 cells was produced (MC3T3-Rorβ). These cells displayed markedly suppressed bone nodule formation as well as reduced osteocalcin and osterix gene expression. Because these genes are Runx2 targets, we reasoned that Rorβ may interfere with Runx2 activity. Consistent with this, transient transfection analysis demonstrated that Rorβ inhibited Runx2-dependent activation of a Runx2-reporter construct. In summary, our data provide a comprehensive profile of NR expression during osteoblast differentiation and identify Rorβ as a novel regulator of osteogenesis and potentially of age-related bone loss through antagonism of Runx2 activity.
转录因子的复杂网络有助于成骨细胞表型的建立和维持。虽然相对较少的转录因子，如 Runx2 和 osterix，对于成骨细胞分化过程是必不可少的，但其他转录因子则用于精细调节以响应各种环境和激素信号。核受体（NR）转录因子超家族参与骨生物学的许多方面。在这项研究中，我们对分化中的原代鼠颅骨细胞中的整个 NR 超家族的表达模式进行了特征描述，以鉴定新的 NR 调节模式。在整个分化过程中观察到 NR 表达的动态模式。有趣的是，维甲酸受体相关孤儿受体β（Rorβ）的表达在分化的后期阶段明显受到抑制。为了更深入地了解 NR 在骨生物学中的功能，还在从小鼠骨髓前体细胞中鉴定的年轻（6 个月）或衰老、骨质疏松（18-22 个月）小鼠中对 NR 超家族进行了分析。有趣的是，Rorβ 在老年队列中被强烈过表达。总的来说，这些数据提供了证据表明 Rorβ 的表达与成骨潜能呈负相关，表明 Rorβ 可能是成骨作用的一个重要且尚未探索的调节因子。为了验证这一假设，在小鼠成骨细胞 MC3T3-E1 细胞中产生了稳定表达 Rorβ的细胞模型（MC3T3-Rorβ）。这些细胞的骨结节形成明显受到抑制，同时骨钙素和 osterix 基因的表达也降低。由于这些基因是 Runx2 的靶标，我们推测 Rorβ 可能干扰 Runx2 活性。一致，瞬时转染分析表明 Rorβ 抑制了 Runx2 依赖的 Runx2 报告基因构建体的激活。总之，我们的数据提供了成骨细胞分化过程中 NR 表达的综合图谱，并确定 Rorβ 是成骨作用的新调节因子，并且可能通过拮抗 Runx2 活性来调节与年龄相关的骨质流失。
