Effects of Thrombopoietin (TPO) on Longitudinal Mouse Hind Limb Crush Injury Model

abstractApproximately 645 people suffer from blunt force trauma injury to the femur every day. The recovery time of such injury can last anywhere from 3-6 months. Thrombopoietin (TPO) was used as a growth factor to induce bone and muscle healing. In this study, nine separate mouse groups (10 mice per group) were used: Crush PBS, Crush TPO, Surgery PBS, and Surgery TPO at day 3 and day 17, and controls with no surgery/crush/treatment. Skeletal muscle was harvested from the following sites: experimental impact, experimental adjacent, and normal contralateral skeletal muscle as a control. The muscles were fixed, processed, sectioned, and stained with H&E and Massons Trichrome stains. The slides were reviewed for skeletal muscle injury, muscle necrosis, inflammation, muscle repair, and regeneration. In addition, F4/80, an immunostain for macrophages was performed. On microscopic examination at day 3 the most common histologic changes seen were sporadic muscle fiber vacuolation, focal necrosis of varying sizes, muscle contraction bands, and infiltration of macrophages. On day 17, the skeletal muscle injury was generally healed. The main histologic lesions seen were variable sizes of muscle fibers, early fibroplasia, fat infiltration, some macrophages, satellite cells, and neovascularization. Comparing the TPO treated mice versus the PBS control group, the lesions at both time points were less in the TPO treated mice

Improving Post-Operative Outcomes in Aged and Diabetic Obese Mice

Due to their small size, high metabolic rate, and large surface to volume ratio, mice are a challenge to work with surgically and peri-operatively. Working with mice that are more susceptible to anesthetic agents, aged, or obese (e.g., diabetic mice), provides even more challenges. In two separate studies, we found simple that supportive care measures during and after surgery improved post-operative outcomes

A Bibliometric Study of Authorship and Collaboration Trends Over the Past 30 Years in Four Major Musculoskeletal Science Journals

This study explored changes in bibliometric variables over the last 30 years for four major musculoskeletal science journals (BONE®), Calcified Tissue International® (CTI®), Journal of Bone and Mineral Research® (JBMR®), and Journal of Orthopaedic Research® (JOR®), with a specific focus on author gender. Bibliometric data were collected for all manuscripts in 1985 (BONE®, CTI®, JOR®), 1986 (JBMR®), 1995, 2005, and 2015; 2776 manuscripts met inclusion criteria. Manuscripts from Europe were more often published in BONE® or CTI®, while those from North America in JBMR® or JOR®. All journals demonstrated an increase over time in the number of authors (3.67–7.3), number of countries (1.1–1.4), number of institutions (1.4–3.1), and number of references (25.1–45.4). The number of manuscript pages increased (6.6–8.9) except for JOR® which showed a decline. CTI® had the lowest number of authors (4.9 vs. 5.6–6.8). There was a change in the corresponding author position from first to last for all journals; this change was highest for CTI® (35%) and lowest for BONE® (14.0%). All journals demonstrated an increase over time in female authors; however, CTI® was the highest amongst these four journals. The percentage of female first authors rose from 24.6 to 44.3% (CTI® 29.1–52.3%). The percentage of corresponding female authors rose from 17.5 to 33.6% (CTI® 22.9–40.0%). The proportion of female authors is increasing, likely reflecting the increasing number of women obtaining doctorates in science, medicine, and engineering

Historical Analysis of Bibliometric Trends in the Journal of Pediatric Orthopaedics With a Particular Focus on Sex

Background: Orthopaedics is the clinical discipline with the lowest percentage of female residents and faculty. Pediatric orthopaedics has a higher percentage of women than other orthopaedic subspecialties. It was the purpose of this study to examine bibliometric trends in the Journal of Pediatric Orthopaedics (JPO) with a specific focus on sex. Methods: A bibliometeric analysis for the years 2015, 2005, 1995, 1985, 1981 was performed. The names of first and corresponding authors; corresponding author position; country of origin; number of institutions, countries, authors, printed pages, and references was tabulated. Author sex was identified for the first and corresponding authors using the “Baby Name Guesser” (www.gpeters.com/names/baby-names.php). A P<0.05 was considered significant. Results: There were 746 publications; 68.7% were from North America. The average number of authors, corresponding author position, collaborating institutions, countries, and number of references increased, whereas the number of printed pages decreased. Asia had the greatest number of authors (4.4), with Australia/New Zealand the fewest (3.4). Sex was determined for 98.3% of the first authors and 98.5% of the corresponding authors. There was a significant increase in the number of female first authors over time (5.9% to 25.6%, P<10−6), especially in Europe and North America. There were significant increase in the number of female corresponding authors over time (5.8% to 17.6%, P=0.000009). There was a significant trend to have a greater percentage of both female first and corresponding authors over time (P=0.0005) with a reverse trend for both male first and corresponding authors (P<10−6). Conclusions: In this study, we noted that the number of female first and corresponding authors in Journal of Pediatric Orthopaedics has been steadily increasing. This should result in more female pediatric orthopaedic surgeons in academic faculty positions

Benchside to Bedside: The Launching of a Novel Bone Healing Agent

poster abstractThe ability to evolve a nascent idea into a successful entity requires navigation through a number of perils known to debilitate new ventures. Embryonic firms (or ideas) require sufficient development; from establishing an unambiguous approach, to attaining the necessary capital for evolution and growth, to fostering an environment and market for the idea or product. In the venture community, there are a number of advocates who possess the ability to contribute to new ventures (e.g. venture community support functions, venture capitalists, or informal contributors), and these individuals help navigate the startup or idea through inception to effectuation. Academic faculty, though, who often are not engrained into the local venture community, are frequently disadvantaged because their ideas or new firms come as an ancillary to their primary work. Already potentially impeded by the challenges presented by the legal constraints of providing sufficient equity for ideas to the university, developing a clear, effective path to market can be difficult for academic faculty. In addition to the systemic uncertainty, difficulty, and impediments faced by all entrepreneurs, academic faculty are constrained by limited time, funding, experience, and other resources – all related to their inclusion in the university or system. In order to alleviate these constraints and propel cutting-edge scientific breakthroughs and technological development, Innovation-to-Enterprise Central (ITEC) was initiated to assist academic faculties’ developments into the market – where, ultimately, these products will have the greatest utility to society. Osetofuse is an embryonic firm in the nascent stages of conceptualizing a revolutionary new product, which uses thrombopoietin as a novel bone healing agent. Through the ITEC program, Osteofuse has been able to facilitate the exploration of the potential value (clinically, economically, and societally) of its research and how the initial idea can be developed into a commercialized and monetized product. In the process, it has developed mechanisms to gauge the market’s acceptance of the product, the intellectual property and legal issue constraints facing the idea, potential commercialization streams and related valuations for marketization, and a quantitative analysis of projected revenue provisions. ITEC fosters continual compounding of knowledge capacity, as the trajectory of Osteofuse has not only inclined, but redirected because of specific uncovered data and insight from the program. As a result, Osteofuse has undergone dramatic transformation; in terms of both its formal identity and the potential approach to the market

ELEVATED LEVELS OF PLATELETS AND MDM2 EXPRESSION ARE CONTRIB-UTING FACTORS TO FACILITATING THE METASTASIS OF OSTEOSARCOMA

poster abstractOsteosarcoma (OS) is the most common form of primary bone cancer and the 6th leading cause of cancer in pediatric patients. A chart review of OS patients treated at this institution suggests that a high platelet count at di-agnosis is significantly (p=0.023) and inversely associated with the first year of survival. As the effects of platelet interaction with OS have been exten-sively researched and suggest that platelets may facilitate tumor metastasis, and the most important prognostic factor for OS patient survival is metasta-sis to the lungs, we hypothesized that platelets increase metastasis to the lungs and reduce survival. Therefore, we sought to determine whether in-creasing platelet numbers in a well characterized OS mouse model would de-crease survival and/or increase metastasis to the lungs. We found that thrombopoietin (TPO) treated mice, had increased platelet numbers, died earlier than placebo treated controls, and that lungs from TPO treated mice contained a small number of large tumor cells (most metastatic lesions were 2-4 cells), whereas lungs from placebo treated controls showed no signs of metastases. Next, an OS tissue microarray (TMA) was built from OS patients seen at our institution over the past 10 years. Mdm2, p53, TPO, and c-mpl expression were evaluated by immunohistochemical (IHC) staining followed by quantitation using the Aperio Imaging system and analysis software. C-mpl (TPO receptor) expression was higher in the metastatic than the primary tumors, suggesting that platelets may contribute to the metastasis of OS. Elevated levels of Mdm2 correlated with metastasis and lower levels of p53, as detected by IHC. In conclusion, both the mouse model and the human OS data were similar, suggesting that both platelets and Mdm2 promote metas-tases in OS

Aging-Related Reduced Expression of CXCR4 on Bone Marrow Mesenchymal Stromal Cells Contributes to Hematopoietic Stem and Progenitor Cell Defects

Aging impairs the regenerative potential of hematopoietic stem cells (HSC) and skews differentiation towards the myeloid lineage. The bone marrow (BM) microenvironment has recently been suggested to influence HSC aging, however the mechanisms whereby BM stromal cells mediate this effect is unknown. Here we show that aging-associated decreased expression of CXCR4 expression on BM mesenchymal stem cells (MSC) plays a crucial role in the development of the hematopoietic stem and progenitor cells (HSPC) aging phenotype. The BM MSC from old mice was sufficient to drive a premature aging phenotype of young HSPC when cultured together ex vivo. The impaired ability of old MSC to support HSPC function is associated with reduced expression of CXCR4 on BM MSC of old mice. Deletion of the CXCR4 gene in young MSC accelerates an aging phenotype in these cells characterized by increased production of reactive oxygen species (ROS), DNA damage, senescence, and reduced proliferation. Culture of HSPC from young mice with CXCR4 deficient MSC also from young mice led to a premature aging phenotype in the young HSPC, as evidenced by reduced hematopoietic regeneration and enhanced myeloid differentiation. Mechanistically, CXCR4 signaling prevents BM MSC dysfunction by suppressing oxidative stress, as treatment of old or CXCR4 deficient MSC with N-acetyl-L-cysteine (NAC), improved their niche supporting activity, and attenuated the HSPC aging phenotype. Our studies suggest that age-associated reduction in CXCR4 expression on BM MSC impairs hematopoietic niche activity with increased ROS production, driving an HSC aging phenotype. Thus, modulation of the SDF-1/CXCR4 axis in MSC may lead to novel interventions to alleviate the age-associated decline in immune/hematopoietic function

GATA-1 deficiency rescues trabecular but not cortical bone in OPG deficient mice

GATA-1(low/low) mice have an increase in megakaryocytes (MKs) and trabecular bone. The latter is thought to result from MKs directly stimulating osteoblastic bone formation while simultaneously inhibiting osteoclastogenesis. Osteoprotegerin (OPG) is known to inhibit osteoclastogenesis and OPG(-/-) mice have reduced trabecular and cortical bone due to increased osteoclastogenesis. Interestingly, GATA-1(low/low) mice have increased OPG levels. Here, we sought to determine whether GATA-1 knockdown in OPG(-/-) mice could rescue the observed osteoporotic bone phenotype. GATA-1(low/low) mice were bred with OPG(-/-) mice and bone phenotype assessed. GATA-1(low/low) × OPG(-/-) mice have increased cortical bone porosity, similar to OPG(-/-) mice. Both OPG(-/-) and GATA-1(low/low) × OPG(-/-) mice, were found to have increased osteoclasts localized to cortical bone, possibly producing the observed elevated porosity. Biomechanical assessment indicates that OPG(-/-) and GATA-1(low/low) × OPG(-/-) femurs are weaker and less stiff than C57BL/6 or GATA-1(low/low) femurs. Notably, GATA-1(low/low) × OPG(-/-) mice had trabecular bone parameters that were not different from C57BL/6 values, suggesting that GATA-1 deficiency can partially rescue the trabecular bone loss observed with OPG deficiency. The fact that GATA-1 deficiency appears to be able to partially rescue the trabecular, but not the cortical bone phenotype suggests that MKs can locally enhance trabecular bone volume, but that MK secreted factors cannot access cortical bone sufficiently to inhibit osteoclastogenesis or that OPG itself is required to inhibit osteoclastogenesis in cortical bone

Megakaryocytes Regulate Expression of Pyk2 Isoforms and Caspase-mediated Cleavage of Actin in Osteoblasts

The proliferation and differentiation of osteoblast (OB) precursors are essential for elaborating the bone-forming activity of mature OBs. However, the mechanisms regulating OB proliferation and function are largely unknown. We reported that OB proliferation is enhanced by megakaryocytes (MKs) via a process that is regulated in part by integrin signaling. The tyrosine kinase Pyk2 has been shown to regulate cell proliferation and survival in a variety of cells. Pyk2 is also activated by integrin signaling and regulates actin remodeling in bone-resorbing osteoclasts. In this study, we examined the role of Pyk2 and actin in the MK-mediated increase in OB proliferation. Calvarial OBs were cultured in the presence of MKs for various times, and Pyk2 signaling cascades in OBs were examined by Western blotting, subcellular fractionation, and microscopy. We found that MKs regulate the temporal expression of Pyk2 and its subcellular localization. We also found that MKs regulate the expression of two alternatively spliced isoforms of Pyk2 in OBs, which may regulate OB differentiation and proliferation. MKs also induced cytoskeletal reorganization in OBs, which was associated with the caspase-mediated cleavage of actin, an increase in focal adhesions, and the formation of apical membrane ruffles. Moreover, BrdU incorporation in MK-stimulated OBs was blocked by the actin-polymerizing agent, jasplakinolide. Collectively, our studies reveal that Pyk2 and actin play an important role in MK-regulated signaling cascades that control OB proliferation and may be important for therapeutic interventions aimed at increasing bone formation in metabolic diseases of the skeleton