Safadi Fayez F, Xu Jie, Smock Steven L, Kanaan Reem A, Selim Abdul-Hafez, Odgren Paul R, Marks Sandy C, Owen Thomas A, Popoff Steven N
Department of Anatomy and Cell Biology, Temple University School of Medicine, Philadelphia, Pennsylvania 19140, USA.
J Cell Physiol. 2003 Jul;196(1):51-62. doi: 10.1002/jcp.10319.
Connective tissue growth factor (CTGF) is a secreted, extracellular matrix-associated signaling protein that regulates diverse cellular functions. In vivo, CTGF is expressed in many tissues with highest levels in the kidney and brain. The purpose of this study was twofold; first, to localize CTGF in normal bone in vivo during growth and repair, and second, to examine CTGF expression and function in primary osteoblast cultures in vitro and test its effect on bone formation in vivo. Northern and Western blot analyses confirmed that CTGF is expressed in normal long bones during the period of growth or modeling. In situ hybridization and immunohistochemical analysis demonstrated intense staining for CTGF mRNA and protein in osteoblasts lining metaphyseal trabeculae. Examination of CTGF expression in the fracture callus demonstrated that it was primarily localized in osteoblasts lining active, osteogenic surfaces. In primary osteoblast cultures, CTGF mRNA levels demonstrated a bimodal pattern of expression, being high during the peak of the proliferative period, abating as the cells became confluent, and increasing to peak levels and remaining high during mineralization. This pattern suggests that CTGF may play a role in osteoblast proliferation and differentiation as previously demonstrated for fibroblasts and chondrocytes. Treatment of primary osteoblast cultures with anti-CTGF neutralizing antibody caused a dose-dependent inhibition of nodule formation and mineralization. Treatment of primary osteoblast cultures with recombinant CTGF (rCTGF) caused an increase in cell proliferation, alkaline phosphatase activity, and calcium deposition, thereby establishing a functional connection between CTGF and osteoblast differentiation. In vivo delivery of rCTGF into the femoral marrow cavity induced osteogenesis that was associated with increased angiogenesis. This study clearly shows that CTGF is important for osteoblast development and function both in vitro and in vivo.
结缔组织生长因子（CTGF）是一种分泌型的、与细胞外基质相关的信号蛋白，可调节多种细胞功能。在体内，CTGF在许多组织中表达，在肾脏和大脑中水平最高。本研究有两个目的：第一，在生长和修复过程中对体内正常骨骼中的CTGF进行定位；第二，在体外原代成骨细胞培养物中检测CTGF的表达和功能，并测试其对体内骨形成的影响。Northern印迹和Western印迹分析证实，CTGF在生长或塑形期的正常长骨中表达。原位杂交和免疫组织化学分析表明，干骺端小梁表面的成骨细胞中CTGF mRNA和蛋白呈强染色。对骨折痂中CTGF表达的检测表明，它主要定位于活跃的、成骨表面的成骨细胞中。在原代成骨细胞培养物中，CTGF mRNA水平呈现双峰表达模式，在增殖期高峰时较高，随着细胞汇合而降低，在矿化过程中升高至峰值并保持较高水平。这种模式表明，CTGF可能在成骨细胞增殖和分化中发挥作用，正如之前在成纤维细胞和软骨细胞中所证明的那样。用抗CTGF中和抗体处理原代成骨细胞培养物会导致结节形成和矿化的剂量依赖性抑制。用重组CTGF（rCTGF）处理原代成骨细胞培养物会导致细胞增殖、碱性磷酸酶活性和钙沉积增加，从而建立了CTGF与成骨细胞分化之间的功能联系。将rCTGF体内递送至股骨髓腔可诱导成骨，这与血管生成增加有关。本研究清楚地表明，CTGF在体外和体内对成骨细胞的发育和功能都很重要。
