Histone H3K36M mutation and trimethylation patterns in chondroblastoma.

AIMS: Histones are essential components of chromatin, and mutations in histones lead to alterations in methylation and acetylation, which play an important role in tumorigenesis. Most of the chondroblastomas harbour the H3K36M mutation. With the availability of a mutation-specific antibody, we sought to assess the sensitivity of this antibody and the alterations of histone methylation in a series of chondroblastoma cases. METHODS AND RESULTS: Immunohistochemical staining with antibodies against H3K36M, trimethylated histones (H3K27me3 and H3K36me3) and an osteoblastic marker (SATB2) was performed on 27 chondroblastomas from 27 patients. The clinical and radiological characteristics of each patient were reviewed. All 27 tumours showed typical radiological and histological features of chondroblastoma, with a subset of cases showing secondary aneurysmal bone cyst changes (11/27), giant-cell-rich foci (4/27), and matrix-rich areas mimicking chondromyxoid fibroma (1/27). All except one case (26/27, 96%) showed positive H3K36M immunostaining (nuclear). In the majority of cases, there was a diffuse staining pattern. Immunohistochemical staining for H3K27me3 and H3K36me3 showed a heterogeneous staining pattern in all cases, regardless of mutation status. None of the cases showed loss of positivity or diffuse positivity. Focal or diffuse SATB2 expression was seen in 21 of 26 tumours (81%). CONCLUSION: Our results demonstrate that the vast majority of chondroblastomas are positive for H3K36M by immunohistochemical analysis, confirming its diagnostic value. H3K27me3 expression and H3K36me3 expression are heterogeneous in these tumours.

Sarcomas of the mandible.

INTRODUCTION: Sarcomas of the mandible are extremely rare tumors, with osteosarcoma being the most common, followed by Ewing's sarcoma MATERIALS AND METHODS: A retrospective review of the clinical records, imaging studies, and pathology slides of patients with sarcoma of the mandible at a Tertiary Care Cancer Center from 1998 to 2014 was undertaken. The impact of neoadjuvant chemotherapy and postoperative radiotherapy with or without chemotherapy was studied, and factors impacting upon local control and disease-specific survival were analyzed. RESULTS: Twenty-two patients were treated over the study period, comprising of 15 males and seven females. External swelling, intraoral growth, or facial numbness were the presenting symptoms. Eighteen patients had osteosarcoma and four had the Ewing's sarcoma. Nine patients received neoadjuvant chemotherapy. All but one patient underwent surgery. Eleven had negative margins, with 90% recurrence-free survival at 3 years, compared to 10 with positive or close margins, leading to 67% recurrence-free survival. None of the patients receiving neoadjuvant chemotherapy developed recurrence and all were alive at 3 years. The impact of postoperative radiotherapy or adjuvant chemotherapy was not statistically significant. CONCLUSIONS: Wide surgical resection with negative margins remains the hallmark of surgical treatment.

Reproducibility of Serum Potassium Values in Serum From Blood Samples Stored for Increasing Times Prior to Centrifugation and Analysis.

GOALS: The goal of this work was to determine if immediate versus postponed centrifugation of samples affects the levels of serum potassium. METHODS: Twenty participants donated normal venous blood that was collected in four serum separator tubes per donor, each of which was analyzed at 0, 1, 2, or 4 hr on the Siemens Advia 1800 autoanalyzer. RESULTS: Coefficients of variation (CVs) for potassium levels ranged from 0% to 7.6% with a mean of 3 ± 2%. ANOVA testing of the means for all 20 samples showed a P-value of 0.72 (>0.05) indicating that there was no statistically significant difference between the means of the samples at the four time points. Sixteen samples were found to have CVs that were ≤5%. Two samples showed increases of potassium from the reference range to levels higher than the upper reference limit, one of which had a 4-hr value that was within the reference or normal range (3.5-5 mEq/l). Overall, most samples were found to have reproducible levels of serum potassium. CONCLUSIONS: Serum potassium levels from stored whole blood collected in serum separator tubes are, for the most part, stable at room temperature for at least 4 hr prior to analysis. However, some samples can exhibit significant fluctuations of values.

Rosai-Dorfman Disease of pancreas: rare aetiology mimicking malignancy.

Rosai-Dorfman disease (RDD) is a non-Langerhans cell histiocytosis which usually presents as painless lymphadenopathy. Extranodal involvement is known to occur in various organs, and less than ten cases with primary pancreatic involvement have been reported previously. This case report details the clinical course of an elderly female, presenting with upper abdominal discomfort and imaging suggestive of malignancy. Multiple non-diagnostic fine-needle aspirations were followed by surgical intervention. Histopathological evaluation revealed a pancreatic mass with characteristic features of RDD. The large hallmark RDD histiocytes showed pale, watery-clear cytoplasm, central round nucleus, and prominent nucleolus, with and without lymphocyte emperipolesis. The RDD histiocytes showed positive immunostaining for CD68, CD163, S100 (nuclear and cytoplasmic), OCT-2, Cyclin D1 and are negative for CD1a, Factor XIIIa, fascin and langerin. This case underscores the importance of considering RDD in the differential diagnosis of pancreatic masses alongwith comprehensive evaluation, multidisciplinary approach and pancreatic core needle biopsy evaluation.

DNA Mutational and Copy Number Variation Profiling of Primary Craniofacial Osteosarcomas by Next-Generation Sequencing.

BACKGROUND: Craniofacial osteosarcomas (CFOS) are uncommon malignant neoplasms of the head and neck with different clinical presentation, biological behavior and prognosis from conventional osteosarcomas of long bones. Very limited genetic data have been published on CFOS. METHODS: In the current study, we performed comprehensive genomic studies in 15 cases of high-grade CFOS by SNP array and targeted next generation sequencing. RESULT: Our study shows high-grade CFOS demonstrate highly complex and heterogenous genomic alterations and harbor frequently mutated tumor suppressor genes TP53, CDKN2A/B, and PTEN, similar to conventional osteosarcomas. Potentially actionable gene amplifications involving CCNE1, AKT2, MET, NTRK1, PDGFRA, KDR, KIT, MAP3K14, FGFR1, and AURKA were seen in 43% of cases. GNAS hotspot activating mutations were also identified in a subset of CFOS cases, with one case representing malignant transformation from fibrous dysplasia, suggesting a role for GNAS mutation in the development of CFOS. CONCLUSION: High-grade CFOS demonstrate highly complex and heterogenous genomic alterations, with amplification involving receptor tyrosine kinase genes, and frequent mutations involving tumor suppressor genes.

Musculoskeletal tissue engineering by endogenous stem/progenitor cells.

From its inception, tissue engineering has had three tenets: cells, biomaterial scaffolds and signaling molecules. Among the triad, cells are the center piece, because cells are the building blocks of tissues. For decades, cell therapies have focused on the procurement, manipulation and delivery of healthy cells for the treatment of diseases or trauma. Given the complexity and potential high cost of cell delivery, there is recent and surging interest to orchestrate endogenous cells for tissue regeneration. Biomaterial scaffolds are vital for many but not all, tissue-engineering applications and serve to accommodate or promote multiple cellular functions. Signaling molecules can be produced by transplanted cells or endogenous cells, or delivered specifically to regulate cell functions. This review highlights recent work in tissue engineering and cell therapies, with a focus on harnessing the capacity of endogenous cells as an alternative or adjunctive approach for tissue regeneration.

Anti-inflammatory treatment increases angiogenesis during early fracture healing.

OBJECTIVES: Both inflammation and angiogenesis are crucial for normal fracture healing. The goal of this work was to determine how anti-inflammatory treatment affects angiogenesis during early stages of fracture repair. METHODS: Tibia fractures were created in adult mice and animals were treated with indomethacin (2 mg/kg/day), a non-steroidal anti-inflammatory drug, or PBS once a day beginning from 1 day before fracture and continuing to 6 days after fracture. Animals were killed at 7, 14, and 28 days after injury for histomorphometric analysis of fracture healing. A second group of animals were killed at 3 and 7 days after injury to measure tissue levels of VEGF and interleukin-1 beta (IL-1ß). A third group of animals were killed at 3 and 7 days after injury for stereology analysis of macrophage and neutrophil infiltration and tissue vascularization. RESULTS: Indomethacin significantly decreased bone and cartilage formation at 7 days after fracture compared to controls. Indomethacin decreased the tissue levels of IL-1ß at 3 days after fracture but did not affect the recruitment of macrophages or neutrophils to injured limbs. Indomethacin-treated fractures had similar length density and surface density of vasculature as the controls at 3 days after injury. At 7 days after fracture, vasculature in indomethacin-treated fractures exhibited higher length density and surface density than that in controls. By 28 days after injury, indomethacin-treated fractures still exhibited defects in fracture repair. CONCLUSIONS: Anti-inflammatory treatments using indomethacin impair bone and cartilage formation and increase tissue vascularization in the callus during early fracture healing.

Tibial fracture decreases oxygen levels at the site of injury.

OBJECTIVES: Oxygen is an essential component for many aspects of tissue repair. However, the effect of oxygen levels on differentiation of stem cells into osteoblasts and chondrocytes during fracture healing is unknown, in part because of the difficulty in measuring oxygen during fracture healing. In this study we tested the feasibility of using electron paramagnetic resonance (EPR) oximetry to assess tissue oxygen partial pressure (pO(2)) after tibial fractures in mice. METHODS: Transverse tibia fractures were created by three-point bending in adult mice. Paramagnetic material, lithium phthalocyanine (LiPc), was implanted into the fracture site or adjacent to the periosteum in the contralateral leg immediately after fracture. Tissue pO(2) was assessed by EPR 90-110 minutes after implantation of the crystals. in a second experiment, LiPc was implanted into the fracture site and fracture repair and the bio-compatibility of LiPc were assessed at 14 and 28 days after injury. RESULTS: At the very early stage after fracture, injury significantly decreased tissue oxygenation at the fracture site. When animals were breathing 21% oxygen, pO(2) at the fracture site ((30.6 +/- 12.7 mmHg, n=7) was lower than that in contralateral legs (45.5 +/- 15.3 mmHg, n=7, p<0.01). breathing 100% inspired oxygen increased the pO(2) in both the fractured (72.8 +/- 28.2 mmHg; n=7) and contralateral legs (148.4 +/- 59.2 mmHg; n=7, p<0.01). in addition, LiPc crystals implanted into fracture sites did not interfere with normal fracture healing at 10 and 28 days post-injury. CONCLUSIONS: EPR oximetry is a valuable tool for monitoring oxygen levels during fracture repair in mice.

Solitary fibrous tumor with neuroendocrine and squamous dedifferentiation: a potential diagnostic pitfall.

Solitary fibrous tumor (SFT) is a ubiquitous mesenchymal neoplasm of deep soft tissue with variable and often unpredictable biological behavior. The lineage is presumed to be fibroblastic, and histological features range from benign to overtly malignant with rare tumors showing "dedifferentiation" or transformation to undifferentiated pleomorphic sarcoma. Dedifferentiation in mesenchymal neoplasms is a phenomenon of histologic progression of a well-differentiated neoplasm to a high-grade sarcoma, which can differentiate along divergent lines. It is extremely uncommon to encounter "transdifferentiation" to non-mesenchymal lineage and still maintaining the driver genetic event of the primary tumor. Herein, we report two diagnostically challenging SFTs with transformation to neuroendocrine and squamous phenotypes. The index case is a pelvic malignant SFT, which metastasized to the liver as a high-grade neuroendocrine carcinoma. The second case is a recurrent brain tumor initially presenting as a typical SFT and evolving into a dedifferentiated SFT with foci of squamous differentiation. Positive immunohistochemical stains for CD34 and STAT6 and the detection of NAB2-STAT6 fusion supported the diagnosis of dedifferentiated SFT in both cases. In the first case, molecular study also demonstrated that both the pelvic primary and liver metastasis harbored the same NAB2-STAT6 fusion. Dedifferentiation to a non-mesenchymal lineage/lineage infidelity can be a potential diagnostic pitfall in these tumors.

Shoulder Lesion in a 69 Year Old Woman.

Milwaukee Shoulder Syndrome (MSS) is a painful progressive arthropathy in which hydroxyapatite crystal deposition in synovial tissue induces lysosomal release of collagenase and neutral proteases. These enzymes are destructive to periarticular tissue, including the synovium, articular cartilage, rotator cuff muscles, and the intrasynovial cortical bone. MSS predominantly occurring in women (90%) over the age of 70 years of age with a clinical history marked by recurrent joint effusions and pain, which classically worsens at night. Our patient is a 69-year-old woman who presented with progressive shoulder pain, most prominent at night, with limited range of motion and swelling; intermittent discharge; and intermittent neck pain that radiated to her right upper extremity. Her medical history was notable for invasive carcinoma of the right breast treated with mastectomy and radiation. She was also treated with radiation therapy for right shoulder pain and a lucent right shoulder lesion presumed to be metastatic breast cancer. The remainder of her medical history consists of hypertension, diabetes mellitus, hyperlipidemia, and uneventful bilateral total knee arthroplasties. At presentation, she denied constitutional symptoms. Based on the patient's history and physical exam the differential diagnosis included primary and metastatic malignancy, radiation induced sarcoma and necrosis, infection, Charcot disease, and crystal arthropathies. Physical exam, laboratory findings, and imaging studies led us to the diagnosis of MSS.

Ischemia leads to delayed union during fracture healing: a mouse model.

Vascular damage accompanying skeletal injury leads to an ischemic environment, and in clinical settings the extent of vascular damage is directly correlated with failure of skeletal repair. However, the exact mechanism(s) underlying ischemia-related defects in bone healing are not well understood. To better understand the mechanism and to facilitate development of novel interventions to treat ischemic fractures, a mouse model of long bone fracture healing in an ischemic environment was created. Ischemia was induced by femoral artery resection prior to tibia fracture. Fractures were left unstabilized or were stabilized with custom-designed external fixators. Animals with intact femoral vessels served as controls. Tissues from non-stabilized fractures were analyzed at various times from 3 to 28 days after injury (n = 5/time point). Femoral artery resection severely impaired blood supply to the fractured limbs, and perfusion to the fracture sites did not recover until 14 days post-injury. Ischemia significantly decreased the callus size (p < 0.05), and decreased bone (p < 0.05) and cartilage (p < 0.05) matrix production during healing of non-stabilized fracture. The decreased formation of skeletal tissues in ischemic limbs was accompanied by decreased cell proliferation and increased apoptosis at early time points, and increased fibrous and fatty tissues adjacent to the fracture site during the third and fourth week after injury. These alterations led to a delayed-union. Complete fracture healing was not achieved in the majority (day 21 = 4/5; day 28 = 5/5) of ischemic animals, while all control mice (n = 5/5) had evidence of bony bridging by day 21. The ratio of cartilage to bone was similar in ischemic and control limbs at days 7 and 10 in non-stabilized fractures. In stabilized fractures, which healed through direct bone formation in the nonischemic controls, ischemia decreased the amount of bone formation at days 10 and 14 (n = 5/time point) but did not induce cartilage formation. These data reveal that an ischemic insult in the hind limb prior to fracture leads to a delayed union or a nonunion, but does not favor formation of cartilage over bone. This model will be useful for testing novel therapeutic regimens to stimulate fracture healing.

Assessing angiogenesis during fracture healing.

Angiogenesis, the sprouting of new capillaries from existing blood vessels, is crucial for normal fracture healing. Angiogenesis is a complex process involving a variety of growth factors and several cell types. The mechanism regulating angiogenesis during fracture repair is not well understood, and the relationships between angiogenesis, chondrogenesis, and osteogenesis are also undefined. In vivo animal models have been useful for determining angiogenic mechanisms. In particular, a murine model has been developed that offers the advantages of easy animal handling, low cost, reliable healing, and the availability of molecular and genetic techniques for research. However, the small size of mice provides challenges, including the inability to assess vascularization using techniques that have been employed in larger animals. Therefore, we developed and optimized techniques specifically for studying angiogenesis during mouse fracture repair. These techniques include blood vessel casting, micro-computed tomography (micro-CT), immunohistochemistry, in situ hybridization, and genetic labeling of endothelial cells. Blood vessel casting and micro-CT are useful for visualization of small blood vessels. Immunohistochemistry using anti-PECAM (platelet endothelial cell adhesion molecule) or CD34 antibodies and genetic approaches using Tie2-cre transgenic mice can be used to label endothelial cells, visualize blood vessels including capillaries, and provide structural information about the vascularization of the fracture callous. Lastly, expression patterns of important growth factors regulating angiogenesis could be assessed by molecular approaches such as in situ hybridization.

A Rare Case of Renal Cell Carcinoma With Leiomyomatous Stroma and Concomitant Ruptured Adrenal Aneurysm.

Here we report a rare case of coexisting renal cell carcinoma (RCC) with leiomyomatous stroma and a ruptured adrenal aneurysm. The patient was a 75-year-old woman with acute abdominal pain. Imaging studies showed a left peri-renal hematoma and a mass in the left kidney. Left nephrectomy and adrenalectomy were performed. Pathological examination showed a ruptured aneurysm in the left adrenal gland. The renal mass was composed of tubules and acini of epithelial cells and a prominent leiomyomatous stroma. The tumor cells were positive for carbonic anhydrase IX, cytokeratin 7, and negative for AMACR, consistent with clear cell (tubulo) papillary RCC.

Cellular basis for age-related changes in fracture repair.

The goal of this work was to define cellular and molecular changes that occur during fracture healing as animals age. We compared the molecular, cellular, and histological progression of skeletal repair in juvenile (4 weeks old), middle-aged (6 months old), and elderly (18 months old) mice at 3, 5, 7, 10, 14, 21, 28, and 35 days post-fracture, using a non-stabilized tibia fracture model. Our histological and molecular analyses demonstrated that there was a sharp decline in fracture healing potential between juvenile and middle-aged animals, while a more subtle decrease in healing potential was apparent between middle-aged and elderly mice. By three days after fracture, chondrocytes expressing Collagen type II, and osteoblasts expressing osteocalcin, were present in calluses of juvenile, but not middle-aged or elderly, mice. At day 5 immature chondrocytes and osteoblasts were observed in calluses of middle-aged and elderly mice. While at this time, chondrocytes in juvenile mice were expressing Collagen type X (ColX) indicating that chondrocyte maturation was already underway. At day 7, chondrocytes expressing ColX were abundant in middle-aged mice while a small domain of ColX-positive chondrocytes were observed in elderly mice. Further, in juvenile and middle-aged mice, but not elderly mice, vascular invasion of the cartilage was underway by day 7. Juvenile mice had replaced nearly all of the cartilage by day 14, while cartilage was still present in the callus of middle-aged mice at day 21 and in elderly mice at day 28. In addition to these delays, histomorphometry revealed that elderly and middle-aged mice formed less bone than juveniles (p<0.001), while cartilage production was unaffected (p>0.22). Collectively, these data suggest that enhancing cell differentiation, improving osteoblast function, and accelerating endochondral ossification may significantly benefit the elderly.

Protein-releasing polymeric scaffolds induce fibrochondrocytic differentiation of endogenous cells for knee meniscus regeneration in sheep.

Regeneration of complex tissues, such as kidney, liver, and cartilage, continues to be a scientific and translational challenge. Survival of ex vivo cultured, transplanted cells in tissue grafts is among one of the key barriers. Meniscus is a complex tissue consisting of collagen fibers and proteoglycans with gradient phenotypes of fibrocartilage and functions to provide congruence of the knee joint, without which the patient is likely to develop arthritis. Endogenous stem/progenitor cells regenerated the knee meniscus upon spatially released human connective tissue growth factor (CTGF) and transforming growth factor-ß3 (TGFß3) from a three-dimensional (3D)-printed biomaterial, enabling functional knee recovery. Sequentially applied CTGF and TGFß3 were necessary and sufficient to propel mesenchymal stem/progenitor cells, as a heterogeneous population or as single-cell progenies, into fibrochondrocytes that concurrently synthesized procollagens I and IIα. When released from microchannels of 3D-printed, human meniscus scaffolds, CTGF and TGFß3 induced endogenous stem/progenitor cells to differentiate and synthesize zone-specific type I and II collagens. We then replaced sheep meniscus with anatomically correct, 3D-printed scaffolds that incorporated spatially delivered CTGF and TGFß3. Endogenous cells regenerated the meniscus with zone-specific matrix phenotypes: primarily type I collagen in the outer zone, and type II collagen in the inner zone, reminiscent of the native meniscus. Spatiotemporally delivered CTGF and TGFß3 also restored inhomogeneous mechanical properties in the regenerated sheep meniscus. Survival and directed differentiation of endogenous cells in a tissue defect may have implications in the regeneration of complex (heterogeneous) tissues and organs.

MMP9 regulates the cellular response to inflammation after skeletal injury.

Like other tissue injuries, bone fracture triggers an inflammatory response, which plays an important role in skeletal repair. Inflammation is believed to have both positive and negative effects on bone repair, but the underlying cellular mechanisms are not well understood. To assess the role of inflammation on skeletal cell differentiation, we used mouse models of fracture repair that stimulate either intramembranous or endochondral ossification. In the first model, fractures are rigidly stabilized leading to direct bone formation, while in the second model, fracture instability causes cartilage and bone formation. We compared the inflammatory response in these two mechanical environments and found changes in the expression patterns of inflammatory genes and in the recruitment of inflammatory cells and osteoclasts. These results suggested that the inflammatory response could influence skeletal cell differentiation after fracture. We then exploited matrix metalloproteinase 9 (MMP9) that is expressed in inflammatory cells and osteoclasts, and which we previously showed is a potential regulator of cell fate decisions during fracture repair. Mmp9(-/-) mice heal stabilized fractures via endochondral ossification, while wild type mice heal via intramembranous ossification. In parallel, we observed increases in macrophages and T cells in the callus of Mmp9(-/-) compared to wild type mice. To assess the link between the profile of inflammatory cells and skeletal cell fate functionally, we transplanted Mmp9(-/-) mice with wild type bone marrow, to reconstitute a wild type hematopoietic lineage in interaction with the Mmp9(-/-) stroma and periosteum. Following transplantation, Mmp9(-/-) mice healed stabilized fractures via intramembranous ossification and exhibited a normal profile of inflammatory cells. Moreover, Mmp9(-/-) periosteal grafts healed via intramembranous ossification in wild type hosts, but healed via endochondral ossification in Mmp9(-/-) hosts. We observed that macrophages accumulated at the periosteal surface in Mmp9(-/-) mice, suggesting that cell differentiation in the periosteum is influenced by factors such as BMP2 that are produced locally by inflammatory cells. Taken together, these results show that MMP9 mediates indirect effects on skeletal cell differentiation by regulating the inflammatory response and the distribution of inflammatory cells, leading to the local regulation of periosteal cell differentiation.

The role of oxygen during fracture healing.

Oxygen affects the activity of multiple skeletogenic cells and is involved in many processes that are important for fracture healing. However, the role of oxygen in fracture healing has not been fully studied. Here we systematically examine the effects of oxygen tension on fracture healing and test the ability of hyperoxia to rescue healing defects in a mouse model of ischemic fracture healing. Mice with tibia fracture were housed in custom-built gas chambers and groups breathed a constant atmosphere of 13% oxygen (hypoxia), 21% oxygen (normoxia), or 50% oxygen (hyperoxia). The influx of inflammatory cells to the fracture site, stem cell differentiation, tissue vascularization, and fracture healing were analyzed. In addition, the efficacy of hyperoxia (50% oxygen) as a treatment regimen for fracture nonunion was tested. Hypoxic animals had decreased tissue vascularity, decreased bone formation, and delayed callus remodeling. Hyperoxia increased tissue vascularization, altered fracture healing in un-complicated fractures, and improved bone repair in ischemia-induced delayed fracture union. However, neither hypoxia nor hyperoxia significantly altered chondrogenesis or osteogenesis during early stages of fracture healing, and infiltration of macrophages and neutrophils was not affected by environmental oxygen after bone injury. In conclusion, our results indicate that environmental oxygen levels affect tissue vascularization and fracture healing, and that providing oxygen when fractures are accompanied by ischemia may be beneficial.

Vascular endothelial growth factor improves bone repair in a murine nonunion model.

OBJECTIVES: Vascular endothelial growth factor (VEGF) is a potent angiogenic factor that plays an important role during skeletal development and fracture healing. Previous experimental studies have shown that VEGF applied immediately after injury can stimulate bone repair in animal fracture nonunion models. However, the effectiveness of VEGF on an established fracture non-union has not been determined. the goal of this work was to test the ability of VEGF applied at a later stage on the healing of fracture nonunions. METHODS: In this study, a murine non-union model was induced by rapid distraction of a tibia osteotomy. this model exhibits radiological and histological evidence of impaired fracture healing at 7 days after the completion of distraction. VEGF (10 µg in 20 µl Pbs/day, n=10) or control (20 µl Pbs/day, n=10) was injected directly into the distraction gap through the posterior musculature on three consecutive days (7, 8, and 9 days after completing distraction). A third group of animals (n=10) with rapid distraction, but no injections, served as non-treated controls. Fracture healing was analyzed by x-ray, histology, and histomorphometry at 27 days after the last round of distraction. RESULTS: radiographs showed that half of the VEGF treated animals (5/10) achieved bony healing whereas the majority of Pbs treated (7/10) and non-treated controls (8/10) did not exhibit bone bridging. Histological and histomorphometric analyses demonstrated that VEGF increased, but not significantly, the amount of bone formed in the distraction gap (1.35 ± 0.35 mm(3)), compared to the saline treated (0.77 ± 0.25 mm(3), p=0.19) and non-treated animals (0.79 ± 0.23mm(3), p=0.12). CONCLUSIONS: Results from this study demonstrate that VEGF potentially promotes bone repair, warranting further research in this direction.