雌激素对骨折愈合过程中骨形态发生蛋白4 mRNA表达的影响

目的：探讨骨折愈合过程中雌激素对骨形态发生蛋白4(bone morphogenetic protein-4，BMP—4)基因表达的影响，了解BMP—4在骨折愈合过程中的作用。方法：180只雌性SD大鼠随机分成4组。OVXE1组为卵巢切除后1d开始雌激素替代治疗；OVXE2组为骨折后雌激素替代组；OVX组为雌激素缺乏组；SO组为正常对照组。所有动物在一侧胫骨中段造成骨折模型；各组动物分别在骨折后1、2、3、4、5、6、7、14、28d处死取材；标本制成石蜡切片，用地高辛标记的BMP-4 cDNA探针进行原位核酸分子杂交；实验结果经计算机图像处理系统分析并作统计学处理。结果：各组动物骨折后1—3d，骨折周围血肿及组织内BMP—4 cRMA检测为阳性，从第4d开始为阴性。OVX组的BMP—4 mRNA表达较任何一组都明显增多(P＜0．05)；SO组及OVXE1组最少。结论：BMP—4主要在骨折愈合早期起作用。雌激素对BMP-4基因的表达有抑制作用，将BMP—4基因的表达控制在一定程度内可能是雌激素调节骨折愈合的一种方式。     

雌激素对实验性骨质疏松症骨折愈合的影响

目的 研究不同剂量的雌激素对卵巢除后发生骨质疏检习股骨骨折愈合的影响。方法 成年ｂａｌｂ／ｃ小鼠卵巢除后３个月开始制作右股骨中段闭合骨折模型,同一天开始皮下注射高低两种剂量雌激素,小鼠于折骨后５、１０、１５、２０、３０天分批处死,采用Ｘ射线、骨痂称重、骨痂中钙盐沉积率测定、组织学检查、血清生化检测等方法研究雌激素对骨折愈合的影响,并和卵巢除组、假手术对照组进行比较。结果 卵巢除组骨痂较小,     

Spatial and temporal gene expression in chondrogenesis during fracture healing and the effects of basic fibroblast growth factor.

Chondrogenesis is an essential component of endochondral fracture healing, though the molecular and cellular events by which it is regulated have not been fully elucidated. In this study, we used a rat model of closed fracture healing to determine the spatial and temporal expression of genes for cartilage-specific collagens. Furthermore, to determine the effects of basic fibroblast growth factor (bFGF) on chondrogenesis in fracture healing, we injected 100 microg recombinant human bFGF into the fracture site immediately after fracture. In normal calluses, pro-alpha1(II) collagen mRNA (COL2A1) was detected in proliferative chondrocytes beginning on day 4 after the fracture, and pro-alpha1(X) collagen mRNA (COL10A1) in hypertrophic chondrocytes beginning on day 7. In FGF-injected calluses, the cartilage enlarged in size significantly. On day 14, both COL2A1- and COL10A1-expressing cells were more widely distributed, and the amounts of COL2A1 and COL10A1 mRNAs were both approximately 2-fold increased when compared with uninjected fractures. Temporal patterns of expression for these genes were, however, identical to those found in normal calluses. The number of proliferating cell nuclear antigen-positive cells was increased in the non-cartilaginous area in the bFGF-injected calluses by day 4. The present molecular analyses demonstrate that a single injection of bFGF enhances the proliferation of chondroprogenitor cells in fracture callus, and thus contributes to the formation of a larger cartilage. However, maturation of chondrocytes and replacement of the cartilage by osseous tissue are not enhanced by exogenous bFGF, and this results in the prolonged cartilaginous callus phase. We conclude that, in the healing of closed fractures of long bones, exogenous bFGF has a capacity to enlarge the cartilaginous calluses, but not to induce more rapid healing.     

Identification of Novel Gene Expression in Healing Fracture Callus Tissue by DNA Microarray

Fracture healing requires controlled expression of thousands of genes. Only a small fraction of these genes have been isolated and fewer yet have been shown to play a direct role in fracture healing. The purpose of this study was threefold: (1) to develop a reproducible open femur model of fracture healing that produces consistent fracture calluses for subsequent RNA extraction, (2) to use this model to determine temporal expression patterns of known and unknown genes using DNA microarray expression profiling, and (3) to identify and validate novel gene expression in fracture healing. In the initial arm of the study, a total of 56 wild-type C57BL/6 mice were used. An open, stabilized diaphyseal femur fracture was created. Animals were killed at 1, 5, 7, 10, 14, 21, and 35 days after surgery and the femurs were harvested for analysis. At each time point, fractures were radiographed and sectioned for histologic analyses. Tissue from fracture callus at all stages following fracture yielded reproducibly large amounts of mRNA. Expression profiling revealed that genes cluster by function in a manner similar to the histologic stages of fracture healing. Based on the expression profiling of fracture tissue, temporal expression patterns of several genes known to be involved in fracture healing were verified. Novel expression of multiple genes in fracture callous tissue was also revealed including leptin and leptin receptor. In order to test whether leptin signaling is required for fracture repair, mice deficient in leptin or its receptor were fractured using the same model. Fracture calluses of mice deficient in both leptin or leptin receptor are larger than wild-type mice fractures, likely due to a delay in mineralization, revealing a previously unrecognized role of leptin signaling in fracture healing. This novel model of murine fracture repair is useful in examining both global changes in gene expression as well as individual signaling pathways, which can be used to identify specific molecular mechanisms of fracture healing.     

Acidic fibroblast growth factor (aFGF) injection stimulates cartilage enlargement and inhibits cartilage gene expression in rat fracture healing

The effect of the administration of acidic fibroblast growth factor (aFGF) on normal fracture healing was examined in a rat fracture model. One microgram of aFGF was injected into the fracture site between the first and the ninth day after fracture either every other day or every day. aFGF-injected calluses were significantly larger than control calluses, although this does not imply an increased mechanical strength of the callus. Histology showed a marked increase in the size of the cartilaginous soft callus. Total DNA and collagen content in the cartilaginous portion of the aFGF-injected calluses were greater than those of controls, although the collagen content/DNA content ratio was not different between the aFGF-injected and control calluses. Fracture calluses injected with aFGF remained larger than controls until 4 weeks after fracture. The enlarged cartilaginous portion of the aFGF-injected calluses seen at 10 days after fracture was replaced by trabecular bone at 3 and 4 weeks. Northern blot analysis of total cellular RNA extracted separately from the cartilaginous soft callus and the bony hard callus showed decreased expression of type II procollagen and proteoglycan core protein mRNA in the aFGF-injected calluses when compared with controls. A slight decrease in types I and III procollagen mRNA expression was also observed. We concluded that aFGF injections induced cartilage enlargement and decreased mRNA expression for type II procollagen and proteoglycan core protein.     

Programmed removal of chondrocytes during endochondral fracture healing

This investigation tested the hypothesis that the removal of chondrocytes during endochondral fracture healing involves an ordered process of programmed cell death. To accomplish this, unilateral closed fractures were created in the femora of 36 Sprague-Dawley rats. The rats were killed in groups of four on days 1, 3, 7, 14, 21, 28, 42, 49, and 56 after fracture. The femora were embedded in paraffin and tested for expression of specific markers of fragmented DNA with use of a terminal deoxyuridyl transferase-mediated deoxyuridine triphosphatebiotin nick end labeling (TUNEL) technique. To determine the potential for trans–differentiation of chondrocytes to osteoblasts calluses were also hybridized to detect expression of osteocal in mRNA. Cell proliferation was assessed by an immunohistochemical detection method for proliferating cell nuclear antigen. A separate group of four rats was killed on day 28 to represent the later stage of the endochondral ossification, and the calluses were examined for cellular morphology with transmission electron microscopy. The results showed a coordination in both time and space of the activities of cellular proliferation and programmed cell death. Cell proliferation was most active in the earlier phases of fracture healing (days 1 through 14) although TUNEL expression was apparent in hypertrophic chondrocytes on day 14 after fracture and persisted until day 28. In the later stages of fracture healing (days 14 through 28), proliferating cell nuclear antigen was no longer synthesized in hard callus (intramembranous bone) and cell removal was the dominant activity in soft callus chondrocytes. Expression of osteocalcin mRNA was detected in osteoblasts but not in hypertrophic chondrocytes or in any other nonosteoblastic cell type. These findings support the hypothesis that the removal of chondrocytes during endochondral fracture healing is part of an ordered transition of tissue types in which the cellular mechanisms are genetically programmed to involve proliferation, maturation, and apoptotic cell death.     

Chemically modified tetracycline (CMT-8) and estrogen promote wound healing in ovariectomized rats: Effects on matrix metalloproteinase-2, membrane type 1 matrix metalloproteinase, and laminin-5 γ2-chain

Estrogen deficiency is associated with impaired cutaneous wound healing. Remodeling of the extracellular matrix in wound healing involves the action of matrix metalloproteinases on basement membrane zone components, especially laminin-5. We studied the effects of estrogen and a potent matrix metalloproteinase inhibitor, chemically modified non-antimicrobial tetracycline, CMT-8, on wound healing in ovariectomized rats. At the tissue level, laminin-5 gamma2-chain expression was decreased and the migration-inductive 80 kDa form of laminin-5 gamma2-chain was absent in ovariectomized rats when compared with sham and CMT-8- or estrogen-treated ovariectomized animals as detected by Western blotting. The highest levels of gelatinolytic activity (matrix metalloproteinase-2 and -9) were found in sham animals. Levels were reduced in ovariectomized rats and were lowest after treating ovariectomized rats with CMT-8 or estrogen as analyzed by functional activity assay and zymography. The total amount of membrane type 1-matrix metalloproteinase was unchanged in all groups. We conclude that CMT-8 and estrogen can promote wound healing in ovariectomized rats, not only by normalizing wound bed total collagen content and structure, but also by recovering the expression and processing of key molecules in wound healing, i.e., laminin-5 gamma2-chain. This study shows, for the first time, the role of estrogen and CMT-8 in laminin-5 gamma2-chain modulation in vivo.     

Gene Expression and Distribution of Key Bone Turnover Markers in the Callus of Estrogen-Deficient, Vitamin D-Depleted Rats

An experimental rat model was used to test the hypothesis that in osteoporosis (OP) the molecular composition of the extracellular matrix in the fracture callus is disturbed. OP was induced at 10 weeks of age by ovariectomy and a vitamin D₃-deficient diet, and sham-operated animals fed normal diet served as controls. Three months later a closed tibial fracture was made and stabilized with an intramedullary nail. After 3 and 6 weeks of healing, the animals were killed and the fracture calluses examined with global gene expression, in situ mRNA expression, and ultrastructural protein distribution of four bone turnover markers: osteopontin, bone sialoprotein, tartrate-resistant acid phosphatase, and cathepsin K. Global gene expression showed a relatively small number of differently regulated genes, mostly upregulated and at 3 weeks. The four chosen markers were not differently regulated, and only minor differences in the in situ mRNA expression and ultrastructural protein distribution were detected. Gene expression and composition of fracture calluses are not generally disturbed in experimental OP.     

血管内皮生长因子在骨折愈合中的自分泌作用

目的探讨血管内皮生长因子（VEGF）在骨折愈合中的自分泌作用。方法利用鼠股骨闭合性骨折模型，应用组织学和逆转录聚合酶链反应（RT-PCR）方法，观察骨折不同的愈合修复阶段中骨痂的组织学变化，检测各种相应骨痂组织中VEGF及其受体VEGFR1（Flt-1）和VEG-FR2（KDR／Flk-1）的mRNA的表达。结果骨折愈合是一个高度有序的组织学变化过程，骨痂内的膜内化骨，软骨形成，软骨内化骨等过程可同时或连续出现。vEGF（251bp）及受体Flt-1（272bp）和KDR／Flk-1（252bp）在骨折后第7、14天的软骨性骨痂（软骨组织）和骨性骨痂（小梁骨-膜内化骨和软骨内化骨）中均同时清晰表达。表达信号均匀，强度较大。结论骨折愈合过程中的软骨性骨痂和骨性骨痂中的软骨-骨细胞系统共表达VEGF及其两种受体，提示VEGF自分泌作用参与骨折愈合过程。     

Conditional deletion of IGF-I in osteocytes unexpectedly accelerates bony union of the fracture gap in mice

This study evaluated the effects of deficient IGF-I expression in osteocytes on fracture healing. Transgenic mice with conditional knockout (cKO) of Igf1 in osteocytes were generated by crossing Dmp1-Cre mice with Igf1 flox mice. Fractures were created on the mid-shaft of tibia of 12-week-old male cKO mice and wild-type (WT) littermates by three-point bending. At 21 and 28 days post-fracture healing, the increases in cortical bone mineral density, mineral content, bone area, and thickness, as well as sub-cortical bone mineral content at the fracture site were each greater in cKO calluses than in WT calluses. There were 85% decrease in the cartilage area and > 2-fold increase in the number of osteoclasts in cKO calluses at 14 days post-fracture, suggesting a more rapid remodeling of endochondral bone. The upregulation of mRNA levels of osteoblast marker genes (cbfa1, alp, Opn, and Ocn) was greater in cKO calluses than in WT calluses. μ-CT analysis suggested an accelerated bony union of the fracture gap in cKO mice. The Sost mRNA level was reduced by 50% and the Bmp2 mRNA level was increased 3-fold in cKO fractures at 14 days post-fracture, but the levels of these two mRNAs in WT fractures were unchanged, suggesting that the accelerated fracture repair may in part act through the Wnt and/or BMP signaling. In conclusion, conditional deletion of Igf1 in osteocytes not only did not impair, but unexpectedly enhanced, bony union of the fracture gap. The accelerated bony union was due in part to upregulation of the Wnt and BMP2 signaling in response to deficient osteocyte-derived IGF-I expression, which in turn favors intramembranous over endochondral bone repair.     

Differential inhibition of fracture healing by non-selective and cyclooxygenase-2 selective non-steroidal anti-inflammatory drugs.

Non-steroidal anti-inflammatory drugs (NSAIDs) specifically inhibit cyclooxygenase (COX) activity and are widely used as anti-arthritics, post-surgical analgesics, and for the relief of acute musculoskeletal pain. Recent studies suggest that non-specific NSAIDs, which inhibit both COX-1 and COX-2 isoforms, delay bone healing. The objectives of this study were 2-fold; first, to measure the relative changes in the normal expression of COX-1 and COX-2 mRNAs over a 42 day period of fracture healing and second, to compare the effects of a commonly used non-specific NSAID, ketorolac, with a COX-2 specific NSAID, Parecoxib (a pro-drug of valdecoxib), on this process. Simple, closed, transverse fractures were generated in femora of male Sprague-Dawley rats weighing approximately 450 g each. Total RNA was prepared from the calluses obtained prior to fracture and at 1, 3, 5, 7, 10, 14, 21, 35 and 42 days post-fracture and levels of COX-1 and COX-2 mRNA were measured using real time PCR. While the relative levels of COX-1 mRNA remained constant over a 21-day period, COX-2 mRNA levels showed peak expression during the first 14 days of healing and returned to basal levels by day 21. Mechanical properties of the calluses were then assessed at 21 and 35 days post-fracture in untreated animals and animals treated with either ketorolac or high or low dose parecoxib. At both 21 and 35 days after fracture, calluses in the group treated with the ketorolac showed a significant reduction in mechanical strength and stiffness when compared with controls (p<0.05). At the 21-day time point, calluses of the parecoxib treated animals showed a lower mean mechanical strength than controls, but the inhibition was not statistically significant. Based on physical analysis of the bones, 3 of 12 (25%) of the ketorolac-treated and 1 of 12 (8%) of the high dose parecoxib-treated animals showed failure to unite their fractures by 21 days, while all fractures in both groups showed union by 35 days. Histological analysis at 21 days showed that the calluses in the ketorolac-treated group contained substantial amounts of residual cartilage while neither the control nor the parecoxib-treated animals showed comparable amounts of cartilage at this stage. These results demonstrate that ketorolac and parecoxib delay fracture healing in this model, but in this study daily administration of ketorolac, a non-selective COX inhibitor had a greater affect on this process. They further demonstrate that a COX-2 selective NSAID, such as parecoxib (valdecoxib), has only a small effect on delaying fracture healing even at doses that are known to fully inhibit prostaglandin production.     

Delayed union of femoral fractures in older rats:decreased gene expression

Background

Fracture healing slows with age. While 6-week-old rats regain normal bone biomechanics at 4 weeks after fracture, one-year-old rats require more than 26 weeks. The possible role of altered mRNA gene expression in this delayed union was studied. Closed mid-shaft femoral fractures were induced followed by euthanasia at 0 time (unfractured) or at 1, 2, 4 or 6 weeks after fracture in 6-week-old and 12-15-month-old Sprague-Dawley female rats. mRNA levels were measured for osteocalcin, type I collagen α1, type II collagen, bone morphogenetic protein (BMP)-2, BMP-4 and the type IA BMP receptor.

Results

For all of the genes studied, the mRNA levels increased in both age groups to a peak at one to two weeks after fracture. All gene expression levels decreased to very low or undetectable levels at four and six weeks after fracture for both age groups. At four weeks after fracture, the younger rats were healed radiographically, but not the older rats.

Conclusions

(1) All genes studied were up-regulated by fracture in both age groups. Thus, the failure of the older rats to heal promptly was not due to the lack of expression of any of the studied genes. (2) The return of the mRNA gene expression to baseline values in the older rats prior to healing may contribute to their delayed union. (3) No genes were overly up-regulated in the older rats. The slower healing response of the older rats did not stimulate a negative-feedback increase in the mRNA expression of stimulatory cytokines.     

葛根素对去卵巢大鼠骨质疏松性骨折骨痂血管形成的影响

目的探讨葛根素对骨质疏松性骨折骨痂血管形成的影响及对骨折愈合的作用。方法60只雌性SD大鼠分为3组:假手术组、去势组、葛根素组,20只/组,去卵巢大鼠骨质疏松性骨折动物。观察评估骨折愈合情况,检测血清BMP-2和VEGF浓度,观察骨痂形态结构变化,检测骨痂BMP-2和VEGF表达。结果去势组较假手术组骨折愈合评分、血清BMP-2和VEGF浓度、骨痂BMP-2和VEGF表达、微血管数均显著降低(P<0.05),葛根素组较去势组上述指标均显著增高(P<0.05)。去势组骨痂组织可见少量骨小梁,生长稀疏,排列紊乱,大量纤维软骨细胞等纤维组织,成骨细胞及新生小血管少见;葛根素组骨痂可见较多骨小梁,生长较旺盛,排列有序,可见较多成熟的骨细胞,新生微小血管较多。结论葛根素通过介导去卵巢大鼠骨质疏松性骨折骨痂BMP-2和VEGF表达,促进骨痂血管形成,改善骨组织形态学,加快骨折愈合。     

甲状旁腺激素联合降钙素对大鼠骨质疏松性骨折骨生长因子的影响

目的探讨甲状旁腺素联合降钙素对大鼠骨质疏松性骨折骨相关生长因子及骨折愈合的影响。方法75只雌性SD大鼠随机分为:假手术组、去势组、甲状旁腺素组、降钙素组、联合用药组,15只/组,首先构建去卵巢大鼠骨质疏松性骨折动物模型。观察并评估骨折愈合,测定骨痂BMD和BMC,检测血清BMP-2、VEGF、TGF-β、IGF-1水平和骨痂蛋白表达,观察骨痂形态结构变化。结果去势组较假手术组骨折愈合评分、骨痂BMD和BMC、血清BMP-2、VEGF、TGF-β、IGF-1水平和骨痂蛋白表达均显著降低(P均<0.05),而联合用药组较去势组上述指标均显著升高(P均<0.05)。去势组骨痂骨小梁明显减少,生长稀疏,排列紊乱,大量间充质干细胞及纤维软骨细胞,成骨细胞及微血管少见;联合用药组骨痂大量骨小梁,生长旺盛,互相交错网状,排列致密有序,可见较多成熟的骨细胞及成骨细胞。结论甲状旁腺素联合降钙素通过介导提高相关骨生长因子表达,提高骨质疏松性骨折骨密度和矿物含量,改善骨组织形态学,加快骨折愈合。     

鲑鱼降钙素对骨质疏松大鼠骨折愈合的影响

目的　探讨鲑鱼降钙素 (密盖息 )对骨质疏松大鼠胫骨骨折愈合的影响。方法　Wis tar雌性大鼠卵巢摘除后 3个月开始制作左胫骨中段骨折模型 ,从术前 1周至术后 8周 ,连续每天皮下注射密盖息 2IU/kg。分别于术后 2 ,4,8周行X片检查和骨折处骨痂的组织学检查。 9周后检测腰椎和左侧胫骨中段的BMD、左胫骨扭转实验、腰椎体生物力学凹入实验。观察鲑鱼降钙对骨折愈合的影响 ,并和卵巢摘除组、雌激素注射组及假手术对照组进行比较。结果　密盖息组治疗 9周后BMD值明显升高 (P <0 0 5) ,骨折局部BMD明显高于治疗前 (P <0 0 5)。X片及组织学检查提示密盖息组比骨质疏松组的骨痂量多 ,愈合时间提前。密盖息组的凹入力和凹入应力明显高于骨质疏松组 (P <0 0 5) ,最大扭矩、剪切应力明显高于骨质疏松组 (P <0 0 5)。结论　鲑鱼降钙素能减少骨量丢失 ,促进矿化 ,加快骨痂的形成 ,促进骨折愈合 ,同时能提高骨生物力学特性和抗骨折能力     

补正续骨丸对去卵巢大鼠骨量及其相关骨折的作用机制研究

目的探索中药复方补正续骨丸对去卵巢大鼠骨量及相关骨折的影响。方法雌性大鼠卵巢切除法(OVX)造PMOP模型,分组给药6周后DXA行大鼠股骨BMD检测; HE染色测量股骨骨小梁密度; ELISA检测法检测大鼠血清中P1NP、β-CTX; Western blot法检测大鼠股骨中4EBP-1、p70S6K蛋白表达;同时比较不同处理组大鼠骨折愈合时间及相同时间骨痂密度,观察补正续骨丸对OVX大鼠骨折愈合时间及骨痂密度的影响。结果补正续骨丸可增加OVX大鼠股骨BMD、减少骨小梁断裂、降低去卵巢大鼠血清中β-CTX水平、抑制mTOR1信号通路中p70S6K、4EBP1的蛋白表达,补正续骨丸可促进OVX大鼠骨折处骨痂密度增加及减少骨折愈合时间。结论中药复方补正续骨丸通过抑制mTOR1信号通路中p70S6K、4EBP1蛋白的表达抑制骨吸收、促进OVX大鼠骨折的愈合。     

Diminished bone formation during diabetic fracture healing is related to the premature resorption of cartilage associated with increased osteoclast activity.

Histological and molecular analysis of fracture healing in normal and diabetic animals showed significantly enhanced removal of cartilage in diabetic animals. Increased cartilage turnover was associated with elevated osteoclast numbers, a higher expression of genes that promote osteoclastogenesis, and diminished primary bone formation. INTRODUCTION: Diminished bone formation, an increased incidence of nonunions, and delayed fracture healing have been observed in animal models and in patients with diabetes. Fracture healing is characterized by the formation of a stabilizing callus in which cartilage is formed and then resorbed and replaced by bone. To gain insight into how diabetes affects fracture healing, studies were carried out focusing on the impact of diabetes on the transition from cartilage to bone. MATERIALS AND METHODS: A low-dose treatment protocol of streptozotocin in CD-1 mice was used to induce a type 1 diabetic condition. After mice were hyperglycemic for 3 weeks, controlled closed simple transverse fractures of the tibia were induced and fixed by intramedullary pins. Histomorphometric analysis of the tibias obtained 12, 16, and 22 days after fracture was performed across the fracture callus at 0.5 mm proximal and distal increments using computer-assisted image analysis. Another group of 16-day samples were examined by microCT. RNA was isolated from a separate set of animals, and the expression of genes that reflect the formation and removal of cartilage and bone was measured by real-time PCR. RESULTS: Molecular analysis of collagen types II and X mRNA expression showed that cartilage formation was the same during the initial period of callus formation. Histomorphometric analysis of day 12 fracture calluses showed that callus size and cartilage area were also similar in normoglycemic and diabetic mice. In contrast, on day 16, callus size, cartilage tissue, and new bone area were 2.0-, 4.4-, and 1.5-fold larger, respectively, in the normoglycemic compared with the diabetic group (p < 0.05). Analysis of microCT images indicated that the bone volume in the normoglycemic animals was 38% larger than in diabetic animals. There were 78% more osteoclasts in the diabetic group compared with the normoglycemic group (p < 0.05) on day 16, consistent with the reduction in cartilage. Real-time PCR showed significantly elevated levels of mRNA expression for TNF-alpha, macrophage-colony stimulating factor, RANKL, and vascular endothelial growth factor-A in the diabetic group. Similarly, the mRNA encoding ADAMTS 4 and 5, major aggrecanases that degrade cartilage, was also elevated in diabetic animals. CONCLUSIONS: These results suggest that impaired fracture healing in diabetes is characterized by increased rates of cartilage resorption. This premature loss of cartilage leads to a reduction in callus size and contributes to decreased bone formation and mechanical strength frequently reported in diabetic fracture healing.