Evaluation of Osteoconductive Scaffolds in the Canine Femoral Multi-Defect Model

Treatment of large segmental bone defects remains an unsolved clinical challenge, despite a wide array of existing bone graft materials. This project was designed to rapidly assess and compare promising biodegradable osteoconductive scaffolds for use in the systematic development of new bone regeneration methodologies that combine scaffolds, sources of osteogenic cells, and bioactive scaffold modifications. Promising biomaterials and scaffold fabrication methods were identified in laboratories at Rutgers, MIT, Integra Life Sciences, and Mayo Clinic. Scaffolds were fabricated from various materials, including poly(L-lactide-co-glycolide) (PLGA), poly(L-lactide-co-ɛ-caprolactone) (PLCL), tyrosine-derived polycarbonate (TyrPC), and poly(propylene fumarate) (PPF). Highly porous three-dimensional (3D) scaffolds were fabricated by 3D printing, laser stereolithography, or solvent casting followed by porogen leaching. The canine femoral multi-defect model was used to systematically compare scaffold performance and enable selection of the most promising substrate(s) on which to add cell sourcing options and bioactive surface modifications. Mineralized cancellous allograft (MCA) was used to provide a comparative reference to the current clinical standard for osteoconductive scaffolds. Percent bone volume within the defect was assessed 4 weeks after implantation using both MicroCT and limited histomorphometry. Bone formed at the periphery of all scaffolds with varying levels of radial ingrowth. MCA produced a rapid and advanced stage of bone formation and remodeling throughout the defect in 4 weeks, greatly exceeding the performance of all polymer scaffolds. Two scaffold constructs, TyrPC[subscript PL]/TCP and PPF4[subscript SLA]/HA[subscript PLGA Dip], proved to be significantly better than alternative PLGA and PLCL scaffolds, justifying further development. MCA remains the current standard for osteoconductive scaffolds.United States. Army Medical Research and Materiel Command (Armed Forces Institute of Regenerative Medicine)United States. Office of Naval ResearchUnited States. Air Force. Office of the Surgeon GeneralUnited States. NavyNational Institutes of Health (U.S.)United States. Veterans AdministrationCleveland Clinic Foundatio

Severe Osteogenesis Imperfecta in Cyclophilin B–Deficient Mice

Osteogenesis Imperfecta (OI) is a human syndrome characterized by exquisitely fragile bones due to osteoporosis. The majority of autosomal dominant OI cases result from point or splice site mutations in the type I collagen genes, which are thought to lead to aberrant osteoid within developing bones. OI also occurs in humans with homozygous mutations in Prolyl-3-Hydroxylase-1 (LEPRE1). Although P3H1 is known to hydroxylate a single residue (pro-986) in type I collagen chains, it is unclear how this modification acts to facilitate collagen fibril formation. P3H1 exists in a complex with CRTAP and the peptidyl-prolyl isomerase cyclophilin B (CypB), encoded by the Ppib gene. Mutations in CRTAP cause OI in mice and humans, through an unknown mechanism, while the role of CypB in this complex has been a complete mystery. To study the role of mammalian CypB, we generated mice lacking this protein. Early in life, Ppib-/- mice developed kyphosis and severe osteoporosis. Collagen fibrils in Ppib-/- mice had abnormal morphology, further consistent with an OI phenotype. In vitro studies revealed that in CypB–deficient fibroblasts, procollagen did not localize properly to the golgi. We found that levels of P3H1 were substantially reduced in Ppib-/- cells, while CRTAP was unaffected by loss of CypB. Conversely, knockdown of either P3H1 or CRTAP did not affect cellular levels of CypB, but prevented its interaction with collagen in vitro. Furthermore, knockdown of CRTAP also caused depletion of cellular P3H1. Consistent with these changes, post translational prolyl-3-hydroxylation of type I collagen by P3H1 was essentially absent in CypB–deficient cells and tissues from CypB–knockout mice. These data provide significant new mechanistic insight into the pathophysiology of OI and reveal how the members of the P3H1/CRTAP/CypB complex interact to direct proper formation of collagen and bone

Germline TGF-β receptor mutations and skeletal fragility: A report on two patients with Loeys–Dietz syndrome

Loeys-Dietz syndrome (LDS, OMIM # 609192) caused by heterozygous mutations in TGFBR1 and TGFBR2 has recently been described as an important cause of familial aortic aneurysms. These patients have craniofacial and skeletal features that overlap with the Marfan syndrome (MFS), and more importantly, have significant vascular fragility as is seen in MFS and Ehlers-Danlos syndrome Type IV (EDS-IV). The skeletal phenotype with respect to low bone mineral density and skeletal fragility is not clear. We present two patients with LDS with significant skeletal fragility. The first is a 17-year-old male who had talipes equinovarus, diaphragmatic and inguinal and herniae, aortic root dilatation necessitating surgical repair, craniofacial and skeletal dysmorphism consistent with LDS, and a history of numerous fragility fractures leading to significant skeletal deformity. He was found to be heterozygous for a c.923T \u3e C transition in exon 4 of TGFBR2. The second is a 26-year-old male with submucous cleft palate, talipes equinovarus, pectus excavatum requiring surgery, inguinal hernia, and aneurysms in the ascending aorta, abdominal aorta, carotid, subclavian, vertebral and brachial arteries requiring surgical repairs. He also had craniofacial and skeletal dysmorphism consistent with LDS, multiple fractures in childhood, low bone mineral density, and was found to be heterozygous for a c.1561 T \u3e C transition in exon 7 of TGFBR2. These case studies highlight the importance of paying close attention to fractures and bone density in patients with LDS. Osteopenia or osteoporosis may become increasingly important issues as earlier detection and treatment of the vascular complications of LDS improves life expectancy in these patients

Enhanced Bone Morphogenetic Protein-2-Induced Ectopic and Orthotopic Bone Formation by Intermittent Parathyroid Hormone (1–34) Administration

Bone morphogenetic proteins (BMPs) play a central role in local bone regeneration strategies, whereas the anabolic features of parathyroid hormone (PTH) are particularly appealing for the systemic treatment of generalized bone loss. The aim of the current study was to investigate whether local BMP-2-induced bone regeneration could be enhanced by systemic administration of PTH (1–34). Empty or BMP-2-loaded poly(lactic-co glycolic acid)/poly(propylene fumarate)/gelatin composites were implanted subcutaneously and in femoral defects in rats (n = 9). For the orthotopic site, empty defects were also tested. Each of the conditions was investigated in combination with daily administered subcutaneous PTH (1–34) injections in the neck. After 8 weeks of implantation, bone mineral density (BMD) and bone volume were analyzed using microcomputed tomography and histology. Ectopic bone formation and almost complete healing of the femoral defect were only seen in rats that received BMP-2-loaded composites. Additional treatment of the rats with PTH (1–34) resulted in significantly (p < 0.05) enhanced BMD and bone volume in the BMP-2 composites at both implantation sites. Despite its effect on BMD in the humerus and vertebra, PTH (1–34) treatment had no significant effect on BMD and bone volume in the empty femoral defects and the ectopically or orthotopically implanted empty composites. Histological analysis showed that the newly formed bone had a normal woven and trabecular appearance. Overall, this study suggests that intermittent administration of a low PTH dose alone has limited potential to enhance local bone regeneration in a critical-sized defect in rats. However, when combined with local BMP-2-releasing scaffolds, PTH administration significantly enhanced osteogenesis in both ectopic and orthotopic sites