Faculty Bibliography

Background/Purpose: b-Tricalcium phosphate (b-TCP), the most common synthetic bone replacement product, is frequently used in craniofacial reconstruction. Although solid b-TCP can be absorbed over time, the slow degradation rate (1%-3%/year) predisposes this product to exposure, infection, and fracture, limiting its use in the growing face where implants are required to grow and remodel with the patient. Our tissue engineering laboratory has successfully leveraged 3D printers to manufacture 3D-printed bioactive ceramic (3DPBC) scaffolds composed of b-TCP in an architecture which optimizes the needs of rigidity with efficient vascular ingrowth, osteogenesis, and degradation kinetics. The latter qualities are further optimized when the osteogenic agent dipyridamole (DIPY) is used. This long-term animal study reports on the new degradation kinetics profile achievable through this novel manufacturing and tissue engineering protocol. Methods/Description: Twenty-two 1-month-old (immature) New Zealand white rabbits underwent creation of unilateral 10 mm calvarial defects with ipsilateral 3.5 +/- 3.5 mm alveolar defects. Each defect was repaired with b-TCP 3DPBC scaffolds coated with 1000 mM DIPY. Rabbits were killed at 8 weeks (n = 6), 6 months (n = 8), and 18 months (n = 8). Bone regeneration and scaffold degradation were calculated using micro-CT images and analyzed in Amira software. Cranial and maxillary suture patency and bone growth were qualitatively analyzed using histologic analysis.
Result(s): Results are reported as a percentage of volumetric space occupied by either scaffold or bone. When comparing time points 8 weeks, 6 months, and 18 months, scaffolds showed significant decreased defect occupancy in calvaria (23.6% +/- 3.6%, 15.2% +/- 1.7%, 5.1% +/- 3.4%; P < .001) and in alveoli (21.5% +/- 3.9%, 6.7% +/- 2.7%, 0.1% +/- 0.2%; P < .001), with annual degradation rates 55.9% and 94.2%, respectively. Between 8 weeks and 18 months, significantly more bone regenerated in calvarial defects (25.8% +/- 6.3% vs 55.7% +/- 10.3%, P < .001) and no difference was found in alveolar defects (28.4% +/- 6.8% vs 32.4% +/- 8.0%, P = .33). Histology showed vascularized, organized bone without suture fusion.
Conclusion(s): The degradation kinetics of b-TCP can be altered through 3D printing and addition of an osteogenic agent. Our study demonstrates an acceleration of b-TCP degradation from 1% to 3% a year to 55% to 95% a year. Absorbed b-TCP is replaced by vascularized bone and there is no damage noted to the growing suture. This additive manufacturing and tissue engineering protocol has implication to future reconstruction of the craniofacial skeleton

BACKGROUND:Vascular injury and inflammation during percutaneous coronary intervention (PCI) are associated with increased risk of post-PCI adverse outcomes. Colchicine decreases neutrophil recruitment to sites of vascular injury. The anti-inflammatory effects of acute colchicine administration before PCI on subsequent myocardial injury are unknown. METHODS:In a prospective, single-site trial, subjects referred for possible PCI (n=714) were randomized to acute preprocedural oral administration of colchicine 1.8 mg or placebo. RESULTS:=0.001). CONCLUSIONS:Acute preprocedural administration of colchicine attenuated the increase in interleukin-6 and high-sensitivity C-reactive protein concentrations after PCI when compared with placebo but did not lower the risk of PCI-related myocardial injury. Registration: URL: https://www.clinicaltrials.gov; Unique Identifiers: NCT02594111, NCT01709981.

Despite the introduction of numerous biologic agents for the treatment of rheumatoid arthritis (RA) and other forms of inflammatory arthritis, low-dose methotrexate therapy remains the gold standard in RA therapy. Methotrexate is generally the first-line drug for the treatment of RA, psoriatic arthritis and other forms of inflammatory arthritis, and it enhances the effect of most biologic agents in RA. Understanding the mechanism of action of methotrexate could be instructive in the appropriate use of the drug and in the design of new regimens for the treatment of RA. Although methotrexate is one of the first examples of intelligent drug design, multiple mechanisms potentially contribute to the anti-inflammatory actions of methotrexate, including the inhibition of purine and pyrimidine synthesis, transmethylation reactions, translocation of nuclear factor-κB (NF-κB) to the nucleus, signalling via the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway and nitric oxide production, as well as the promotion of adenosine release and expression of certain long non-coding RNAs.

In OA chondrocytes, there is diminished mitochondrial production of ATP and diminished extracellular adenosine resulting in diminished adenosine A2A receptor (A2AR) stimulation and altered chondrocyte homeostasis which contributes to the pathogenesis of OA. We tested the hypothesis that A2AR stimulation maintains or enhances mitochondrial function in chondrocytes. The effect of A2AR signaling on mitochondrial health and function was determined in primary murine chondrocytes, a human chondrocytic cell line (T/C-28a2), primary human chondrocytes, and a murine model of OA by transmission electron microscopy analysis, mitochondrial stress testing, confocal live imaging for mitochondrial inner membrane polarity, and immunohistochemistry. In primary murine chondrocytes from A2AR^-/- null mice, which develop spontaneous OA by 16 weeks, there is mitochondrial swelling, dysfunction, and reduced mitochondrial content with increased reactive oxygen species (ROS) burden and diminished mitophagy, as compared to chondrocytes from WT animals. IL-1-stimulated T/C-28a2 cells treated with an A2AR agonist had reduced ROS burden with increased mitochondrial dynamic stability and function, findings which were recapitulated in primary human chondrocytes. In an obesity-induced OA mouse model, there was a marked increase in mitochondrial oxidized material which was markedly improved after intraarticular injections of liposomal A2AR agonist. These results are consistent with the hypothesis that A2AR ligation is mitoprotective in OA.

BACKGROUND:Three-dimensionally-printed bioceramic scaffolds composed of β-tricalcium phosphate delivering the osteogenic agent dipyridamole can heal critically sized calvarial defects in skeletally mature translational models. However, this construct has yet to be applied to growing craniofacial models. In this study, the authors implanted three-dimensionally-printed bioceramic/dipyridamole scaffolds in a growing calvaria animal model and evaluated bone growth as a function of geometric scaffold design and dipyridamole concentration. Potential adverse effects on the growing suture were also evaluated. METHODS:Bilateral calvarial defects (10 mm) were created in 5-week-old (approximately 1.1 kg) New Zealand White rabbits (n = 16 analyzed). Three-dimensionally-printed bioceramic scaffolds were constructed in quadrant form composed of varying pore dimensions (220, 330, and 500 μm). Each scaffold was coated with collagen and soaked in varying concentrations of dipyridamole (100, 1000, and 10,000 μM). Controls consisted of empty defects. Animals were killed 8 weeks postoperatively. Calvariae were analyzed using micro-computed tomography, three-dimensional reconstruction, and nondecalcified histologic sectioning. RESULTS:Scaffold-induced bone growth was statistically greater than bone growth in empty defects (p = 0.02). Large scaffold pores, 500 μm, coated in 1000 μM dipyridamole yielded the most bone growth and lowest degree of scaffold presence within the defect. Histology showed vascularized woven and lamellar bone along with initial formation of vascular canals within the scaffold lattice. Micro-computed tomographic and histologic analysis revealed patent calvarial sutures without evidence of ectopic bone formation across all dipyridamole concentrations. CONCLUSION/CONCLUSIONS:The authors present an effective pediatric bone tissue-engineering scaffold design and dipyridamole concentration that is effective in augmentation of calvarial bone generation while preserving cranial suture patency.

OBJECTIVES/OBJECTIVE:To detail the greatest areas of unmet scientific and clinical needs in rheumatology. METHODS:The 21st annual international Advances in Targeted Therapies meeting brought together more than 100 leading basic scientists and clinical researchers in rheumatology, immunology, epidemiology, molecular biology and other specialties. During the meeting, breakout sessions were convened, consisting of 5 disease-specific groups with 20-30 experts assigned to each group based on expertise. Specific groups included: rheumatoid arthritis, psoriatic arthritis, axial spondyloarthritis, systemic lupus erythematosus and other systemic autoimmune rheumatic diseases. In each group, experts were asked to identify unmet clinical and translational research needs in general and then to prioritise and detail the most important specific needs within each disease area. RESULTS:Overarching themes across all disease states included the need to innovate clinical trial design with emphasis on studying patients with refractory disease, the development of trials that take into account disease endotypes and patients with overlapping inflammatory diseases, the need to better understand the prevalence and incidence of inflammatory diseases in developing regions of the world and ultimately to develop therapies that can cure inflammatory autoimmune diseases. CONCLUSIONS:Unmet needs for new therapies and trial designs, particularly for those with treatment refractory disease, remain a top priority in rheumatology.

Background/Purpose : Implant loosening due to loss of bone is the most common cause of total joint replacement revision surgeries. One of the main cause of osteolysis is the shed of wear particles from the implant as it causes an increased local inflammation and osteoclast number and activity. It is known that an A 2A adenosine receptor selective agonist (CGS21680, CGS) prevents bone loss in wear particle-induced osteolysis model in mice. But frequent administration requirements and its potential to cause side effects make it a less than optimal treatment. We therefore generated and tested a novel alendronate-CGS conjugate (MRS7216) that specifically localizes to bone targeting the agonist to the site of tissue injury and thereby diminishing the frequency of administration and curtailing systemic side effects. Methods : The conjugate was synthesized from CGS by sequential activation of the carboxylic acid moiety and reacting with the appropriate amino acid under basic conditions. A PEG 6 linker was incorporated to alendronic acid by direct coupling. Osteolysis in 6-8-week-old C57BL/6J (WT) or A 2A KO mice was induced by surgical implantation of 3mg of ultrahigh-molecular-weight-polyethylene particles over the calvaria. Mice received a weekly 10mg/kg intraperitoneal dose of MRS7216 conjugate, starting at the time of surgery. Other groups of mice were treated with equivalent weekly doses of alendronate-PEG 6 (AlenP) or saline respectively. An additional control group underwent sham surgery. New bone formation was studied by Calcein/Alizarin Red-labeling. After 2 weeks, animals were sacrificed and microCT and histology analyses were performed. The studies were approved by the Institutional Animal Care and Use Committee of NYU School of Medicine. Results : Receptor binding studies demonstrate that the Ki for CGS, 7216 conjugates and the control AlenP molecules were 21.5 nM, 69.2 nM and >10,000 nM respectively, indicating that MRS7216 efficiently binds the A2A adenosine receptor. MicroCT studies showed that WT mice treated with weekly doses of MRS7216 had a significant reduction in bone damage of 40% (p=0.04) compared to saline treated mice. In contrast, AlenP molecules did not prevent bone erosion. Similarly in A 2A KO mice MRS7216 treatment did not prevent bone damage. Histological analysis of TRAP stained samples showed a significant decrease of osteoclast number/high-power field (HPF) of 55% (p=0.03) in AlenP treated WT mice compared to the saline treated group. The osteoclast depletion was more dramatic in MRS7216 treated group with an 81% reduction of osteoclasts number/HPF (p=0.002). Additionally alkaline phosphatase staining in MRS7216 treated group, showed a significant increase in osteoblast number per HPF compared to saline (55%, p=0.01) and to AlenP group (45%, p=0.03). Furthermore Double bone labeling with calcein/alizarin red showed a significant increase on femurs bone formation of MRS7216 treated group compared to saline and to Alendronate group (p=0.0092 and p=0.0345). Conclusion : Alendronate-CGS conjugates represent a novel and specific therapeutic approach to inhibit osteolysis and stimulate new bone formation to prevent prosthetic failure in patients with prosthetic joints or other bone pathologies

Background/Purpose : Tenofovir is an anti-retroviral agent commonly used to treat human immunodeficiency virus (HIV)-infected patients as part of the drug regimen known as highly active anti-retroviral therapy (HAART). As many as 15% of patients taking tenofovir develop osteopenia resulting in pathological fractures. Recent studies in our lab indicate that tenofovir-induced osteopenia is due to reduction of extracellular adenosine resulting from tenofovir-mediated blockade of Pannexin-1, a transporter for ATP which is hydrolysed in the extracellular space to adenosine, and the effect of tenofovir on bone can be reversed by dipyridamole, an agent that blocks adenosine re-uptake. To further confirm this effect we studied the effect tenofovir on bone in Pannexin-1 knockout mice (PANX1KO) mice. Methods : PANX1KO animals were treated daily with 75mg/Kg of tenofovir, 25 mg/kg tenofovir or both during 4 weeks, after that bone mineral density (BMD) was measured. Additionally, primary osteoclasts were differentiated in the presence of different concentrations of Tenofovir and dipyridamole. The differentiation stage and extracellular ATP levels were studied. Results : Consistent with previous experiments, tenofovir treatment of extracellular ATP in cultures of wild type (WT) bone marrow-derived primary osteoclasts as well as an increase in osteoclast differentiation. However, the addition of dipyridamole inhibited osteoclast differentiation (p=0.0068). The dipyridamole-induced inhibition was reversed with Tenofovir in a dose dependent manner (0.0055). In contrast, osteoclasts from PANX1KO mice did not decrease ATP release when treated with Tenofovir (p= 0.3292). Additionally, PANX1KO mice osteoclast differentiation was also inhibited by dipyridamole (p=0.0005), which was not reversed by dipyridamole treatment (p= 0.9756). In WT mice, Tenofovir treatment reduced bone mineral density by 10% (p< 0.05, n=10) and this effect was reversed in the animals who received Dipyridamole in addition to Tenofovir. In contrast, tenofovir did not affect bone mineral density in PANX1KO mice (p >0.99, n=4). Conclusion : Tenofovir, a commonly used drug, induces osteopenia in many patients. The results presented here support the hypothesis that tenofovir reduces bone density by inhibiting ATP release with subsequent adenosinemediated inhibition of osteoclast differentiation. These effects can be reversed by treatment with dipyridamole