Billiard J, Moran R A, Whitley M Z, Chatterjee-Kishore M, Gillis K, Brown E L, Komm B S, Bodine P V N
Women's Health Research Institute, Wyeth Research, Collegeville, Pennsylvania 19426, USA.
J Cell Biochem. 2003 May 15;89(2):389-400. doi: 10.1002/jcb.10514.
Osteoblast differentiation is a key aspect of bone formation and remodeling. To further our understanding of the differentiation process, we have developed a collection of conditionally immortalized adult human osteoblast cell lines representing discrete stages of differentiation. To evaluate changes in gene expression associated with differentiation, polyA((+)) RNA from pre-osteoblasts, early and late osteoblasts, and pre-osteocytes was subjected to gene chip analysis using the Affymetrix Hu6800 chip in conjunction with an Affymetrix custom chip enriched in bone and cartilage cDNAs. Overall, the expression of 47 genes was found to change threefold or more on both chips between the pre-osteoblastic and pre-osteocytic stages of differentiation. Many of the observed differences, including down-regulation of collagen type I and collagen-processing enzymes, reflect expected patterns and support the relevance of our results. Other changes have not been reported and offer new insight into the osteoblast differentiation process. Thus, we observed regulation of factors controlling cell cycle and proliferation, reflecting decreased proliferation, and increased apoptosis in pre-osteocytic cells. Elements maintaining the cytoskeleton, extracellular matrix, and cell-cell adhesion also changed with differentiation reflecting profound alterations in cell architecture associated with the differentiation process. We also saw dramatic down-regulation of several components of complement and other immune response factors that may be involved in recruitment and differentiation of osteoclasts. The decrease in this group of genes may provide a mechanism for controlling bone remodeling of newly formed bone. Our screen also identified several signaling proteins that may control osteoblast differentiation. These include an orphan nuclear receptor DAX1 and a small ras-related GTPase associated with diabetes, both of which increased with increasing differentiation, as well as a high mobility group-box transcription factor, SOX4, that was down-regulated during differentiation. In summary, our study provides a comprehensive transcriptional profile of human osteoblast differentiation and identifies several genes of potential importance in controlling differentiation of osteoblasts.
成骨细胞分化是骨形成和重塑的关键环节。为了进一步了解这一分化过程，我们构建了一组条件性永生化的成人人类成骨细胞系，它们代表了分化的不同阶段。为了评估与分化相关的基因表达变化，我们使用Affymetrix Hu6800芯片以及富含骨和软骨cDNA的Affymetrix定制芯片，对前成骨细胞、早期和晚期成骨细胞以及前骨细胞的polyA(+)RNA进行了基因芯片分析。总体而言，发现在分化的前成骨细胞阶段和前骨细胞阶段之间，有47个基因的表达在两种芯片上都发生了三倍或更多的变化。许多观察到的差异，包括I型胶原蛋白和胶原蛋白加工酶的下调，反映了预期的模式，并支持了我们结果的相关性。其他变化尚未见报道，为成骨细胞分化过程提供了新的见解。因此，我们观察到控制细胞周期和增殖的因子的调节，这反映了前骨细胞中增殖的减少和凋亡的增加。维持细胞骨架、细胞外基质和细胞间粘附的成分也随着分化而变化，反映了与分化过程相关的细胞结构的深刻改变。我们还看到补体的几个成分以及其他可能参与破骨细胞募集和分化的免疫反应因子的显著下调。这组基因的减少可能为控制新形成骨的骨重塑提供一种机制。我们的筛选还鉴定了几种可能控制成骨细胞分化的信号蛋白。其中包括一个孤儿核受体DAX1和一个与糖尿病相关的小的ras相关GTP酶，它们都随着分化程度的增加而增加，以及一个高迁移率族盒转录因子SOX4，它在分化过程中被下调。总之，我们的研究提供了人类成骨细胞分化的全面转录图谱，并鉴定了几个在控制成骨细胞分化中具有潜在重要性的基因。
