GDF-5 deficiency in mice delays Achilles tendon healing.

The aim of this study was to examine the role of one of the growth/differentiation factors, GDF-5, in the process of tendon healing. Specifically, we tested the hypothesis that GDF-5 deficiency in mice would result in delayed Achilles tendon repair. Using histologic, biochemical, and ultrastructural analyses, we demonstrate that Achilles tendons from 8-week-old male GDF-5 -/- mice exhibit a short-term delay of 1-2 weeks in the healing process compared to phenotypically normal control littermates. Mutant animals took longer to achieve peak cell density, glycosaminoglycan content, and collagen content in the repair tissue, and the time course of changes in collagen fibril size was also delayed. Revascularization was delayed in the mutant mice by 1 week. GDF-5 deficient Achilles tendons also contained significantly more fat within the repair tissue at all time points examined, and was significantly weaker than control tissue at 5 weeks after surgery, but strength differences were no longer detectable by 12-weeks. Together, these data support the hypothesis that GDF-5 may play an important role in modulating tendon repair, and are consistent with previously posited roles for GDF-5 in cell recruitment, migration/adhesion, differentiation, proliferation, and angiogenesis.     

Accelerated hypertrophic chondrocyte kinetics in GDF‐7 deficient murine tibial growth plates

The Growth/Differentiation Factors (GDFs) are a subgroup of the Bone Morphogenetic Proteins (BMPs) well known for their role in joint formation and chondrogenesis. Mice deficient in one of these signaling molecules, GDF‐5, have recently been shown to exhibit a decreased rate of endochondral bone growth in the proximal tibia due to a significantly longer hypertrophic phase duration. GDF‐7 is a related family member, which exhibits a high degree of sequence identity with GDF‐5. The purpose of the present study was to determine whether GDF‐7 deficiency also alters the endochondral bone growth rate in mice and, if so, how this is achieved. Stereologic and cell kinetic parameters in proximal tibial growth plates from 5‐week‐old female GDF‐7 ?/? mice and wild type control littermates were examined. GDF‐7 deficiency resulted in a statistically significant increase in growth rate (+26%; p = 0.0084) and rate of cell loss at the chondrosseous junction (+25%; p = 0.0217). Cells from GDF‐7 deficient mice also exhibited a significantly shorter hypertrophic phase duration compared to wild type controls (?27%; p = 0.0326). These data demonstrate that, in the absence of GDF‐7, the rate of endochondral bone growth is affected through the modulation of hypertrophic phase duration in growth plate chondrocytes. These findings further support a growing body of evidence implicating the GDFs in the formation, maturation, and maintenance of healthy cartilage. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:986–990, 2008     

Mechanical and geometric changes in the growing femora of BMP-5 deficient mice

We examined the growth-related changes in femoral geometry and torsional strength in BMP-5 deficient short-ear mice over a 22-week time interval (“long-term” changes). Four groups of female mice (n = 6 per group) were examined: short-ear animals and their heterozygous control littermates at 4 and 26 weeks of age. In agreement with findings previously observed in a mixed-gender group of adult mice (26 weeks), the femora of short-ear animals were significantly smaller in length and cross section at both ages. The magnitudes of the differences between genotypes were comparable at each age, indicating that the overall rates of appositional and endochondral growth were similar for both genotypes over the 22-week period. In the adult animals, short-ear femora were 27 ± 7% weaker in torsional strength due to their smaller cross-sectional geometry. However, bone strength in adult short-ear mice appeared to be adequate for animal size: No significant difference was detected in maximum femoral torque when normalized by body mass. In 4-week old animals, BMP-5 deficiency was associated with a 27 ± 6% lower body mass, but the torsional strength of the femur was not significantly different from that of controls. Cross-sectional geometry was smaller in 4-week old short-ear mice, but the apparent bone material ultimate shear stress was elevated by 33 ± 10%, thereby resulting in a whole bone torsional strength equivalent to that of the larger control mice. While the data suggest a higher material strength in the 4-week-old short-ear animals, no significant difference in the level of bone mineralization was detectable between genotypes at either age.     

Localization of Pigment Epithelium-Derived Factor in Growing Mouse Bone

Pigment epithelium-derived factor (PEDF) is a potent anti-angiogenic factor found in a wide range of fetal and adult tissues, where it is thought to play a role in the regulation of angiogenesis during development. The temporal expression of PEDF during endochondral bone formation has not previously been reported. In this study, we analysed the expression pattern of PEDF in growing mouse hindlimbs from newborn day one through to maturation at week 9, using immunohistochemistry and in situ hybridization. PEDF expression was demonstrated in chondrocytes within the resting, proliferative and upper hypertrophic zones of the epiphyseal growth plate. The pattern of expression was consistent throughout the developmental stages of the mouse. In addition, PEDF was expressed by osteoblasts lining the bone spicules in the ossification zone of metaphyseal bone, as well as by osteoblasts lining cortical periosteum. These novel results demonstrate that PEDF is developmentally expressed in both cartilage and bone cells during endochondral bone formation, and strongly suggest that it may play a regulatory role in the processes of chondrocyte and osteoblast differentiation, endochondral ossification, and bone remodelling during growth and development of long bones.     

Bone morphogenetic proteins and growth factors: emerging role in regenerative orthopaedic surgery*

Bone morphogenetic proteins (BMPs) were discovered by Urist and colleagues in 1965 and later defined as multifunctional cytokines involved in osteoinduction. BMPs are members of the transforming growth factor-β superfamily, with the exception of the BMP-1. Presently, at least 20 BMPs have been identified and studied, but only BMP 2, 4 and 7 have been able in vitro to stimulate the entire process of stem cell differentiation into osteoblastic mature cells. In preclinical and clinical studies, BMPs have demonstrated potential in osteoinduction and have been approved for clinical use in treating open fractures of the long bones and nonunions and in vertebral arthrodesis. Additional clinical uses of these molecules are under investigation and the possibility of using gene therapy in selected pathologies seems the most appealing. *Proceedings of the Consensus Conference 'TSS in hip and knee replacement" (Rapallo, 22–24 June 2006)     

生长分化因子-5诱导小鼠骨髓基质干细胞向软骨分化过程中对缝隙连接蛋白43表达的影响

目的探讨生长分化因子-5(GDF-5)在小鼠骨髓基质干细胞(BMSCs)向软骨分化过程中对缝隙连接蛋白43(Cx43)表达的影响。方法体外培养小鼠BMSCs,贴壁细胞传代,取第3代细胞,加入地塞米松、VitC、胰岛素和GDF-5诱导培养,72 h后免疫细胞化学和阿尔辛蓝染色分别检测软骨细胞特异性Ⅱ型胶原和蛋白多糖的表达；在诱导24、48和72 h后进行如下检测：MTT法测定GDF-5对小鼠BMSCs增殖的影响；RT-PCR、Western blotting和免疫细胞化学法检测GDF-5对Cx43蛋白表达的影响。结果MTT结果显示不同时间GDF-5对小鼠BMSCs的增殖无影响；RT-PCR、Western blotting和免疫细胞化学结果表明诱导后不同时间均有Cx43 mRNA和蛋白的表达；诱导72 h免疫细胞化学显示有Ⅱ型胶原蛋白的表达,阿尔辛蓝染色阳性,有蛋白多糖基质的分泌。结论GDF-5可以通过上调缝隙连接蛋白Cx43的表达来促进小鼠BM- SCs向软骨方向的分化。