Aaron Schindeler

Bone remodeling during fracture repair : The cellular picture

Fracture healing is a complex event that involves the coordination of a variety of different processes. Repair is typically characterized by four overlapping stages: the initial inflammatory response, soft callus formation, hard callus formation, initial bony union and bone remodeling. However, repair can also be seen to represent a juxtaposition of two distinct forces: anabolism or tissue formation, and catabolism or remodeling. These anabolic/catabolic concepts are useful for understanding bone repair without giving the false impression of temporally distinct stages that operate independently. They are also relevant when considering intervention. In normal bone development, bone remodeling conventionally refers to the removal of calcified bone tissue by osteoclasts. However, in the context of bone repair there are two phases of tissue catabolism: the removal of the initial cartilaginous soft callus, followed by the eventual remodeling of the bony hard callus. In this review, we have attempted to examine catabolism/remodeling in fractures in a systematic fashion. The first section briefly summarizes the traditional four-stage view of fracture repair in a physiological manner. The second section highlights some of the limitations of using a temporal rather than process-driven model and summarizes the anabolic/catabolic paradigm of fracture repair. The third section examines the cellular participants in soft callus remodeling and in particular the role of the osteoclast in endochondral ossification. Finally, the fourth section examines the effects of delaying osteoclast-dependent hard callus remodeling and also poses questions regarding the crosstalk between anabolism and catabolism in the latter stages of fracture repair.

Ras-MAPK Signaling in Osteogenic Differentiation: Friend or Foe?†

THE ROLE OF Ras-mitogen–activated protein kinase (MAPK) signaling in osteogenic differentiation is currently a source of great controversy. On one side is robust evidence from in vitro studies, pharmacological therapy, and genetic disease indicating that increased Ras-MAPK signaling can antagonize bone formation. In this model, committed osteoprogenitors have a choice of either proliferation or further differentiation, with Ras-MAPK being a key regulator of that balance. In contrast, several conflicting in vitro studies have implied that Ras-MAPK is vital for supporting Runx2/Cbfa1 activity and subsequent osteogenic gene expression. As evidence mounts on both sides of the argument, it is important to step back and critically analyze the approaches taken thus far and the relative strength of their conclusions. This perspective attempts to provide a concise and unbiased summary of both lines of evidence to better understand the conflicting models. A particular focus has been placed on comparing differences in methodology. Whereas no single factor emerges as expedient to account for the divergent views, a clear lack of uniformity in experimental methods is evident. Perhaps the greatest variation was seen in the use of numerous cell lines and primary cell types at various stages of osteogenic commitment and differentiation. We speculate that uncommitted multipotent progenitors, committed osteoprogenitors, mature osteoblasts, and cells derived from other lineages may respond differently to identical stimuli, thus explaining some of the apparent conflict in the literature. As well as experimental consistency, we propose that systematic approaches for the independent examination of osteogenic commitment and osteogenic differentiation are of paramount importance for future progress.

The anabolic and catabolic responses in bone repair

The literature on fracture repair has been reviewed. The traditional concepts of delayed and nonunion have been examined in terms of the phased and balanced anabolic and catabolic responses in bone repair. The role of medical manipulation of these inter-related responses in the fracture healing have been considered.

Skeletal abnormalities in neurofibromatosis type 1: Approaches to therapeutic options

The skeleton is frequently affected in individuals with neurofibromatosis type 1, and some of these bone manifestations can result in significant morbidity. The natural history and pathogenesis of the skeletal abnormalities of this disorder are poorly understood and consequently therapeutic options for these manifestations are currently limited. The Childrens Tumor Foundation convened an International Neurofibromatosis Type 1 Bone Abnormalities Consortium to address future directions for clinical trials in skeletal abnormalities associated with this disorder. This report reviews the clinical skeletal manifestations and available preclinical mouse models and summarizes key issues that present barriers to optimal clinical management of skeletal abnormalities in neurofibromatosis type 1. These concepts should help advance optimal clinical management of the skeletal abnormalities in this disease and address major difficulties encountered for the design of clinical trials.

Decreased Bone Mineral Density in Neurofibromatosis Type 1: Results From a Pediatric Cohort

Neurofibromatosis type 1 (NF1) is a common genetic disorder affecting 1 in 3000 live births. It is well documented to be associated with bony deformities and other orthopaedic problems. Based on our observation that NF1 patients undergoing orthopaedic surgery often had osteopenic bone, we performed a study to assess the bone mineral density of a cohort of children with NF1 without orthopaedic defects. Twenty-three patients were recruited from the neurofibromatosis clinic. The bone mineral density of the total body, lumbar spine, and proximal femur was measured using dual-energy x-ray absorptiometry. Quantitative ultrasound was used to measure broadband ultrasonic attenuation at both heels. The groups mean dual-energy x-ray absorptiometry sex- and age-matched Z scores were below normal (−0.8 ± 1.1, −0.8 ± 1.2, −0.7 ± 0.8, −0.6 ± 1.1, −0.6 ± 0.9, −0.6 ± 1.1 for the total body, arms, legs, lumbar spine, and right and left femoral neck, respectively; all P < 0.01). Although some individuals had normal bone mass, 30% had total body Z scores below −1.5. The mean heel broadband ultrasonic attenuation Z score was also lower than normal (−0.8 ± 0.6; P < 0.001). Children with NF1 have a general tendency toward osteopenia, suggesting an abnormal underlying bone phenotype. This may be relevant when considering operative intervention and, if better understood, may partially explain poor bone healing associated with NF1.

A collagen-hydroxyapatite scaffold allows for binding and co-delivery of recombinant bone morphogenetic proteins and bisphosphonates.

An emerging paradigm in orthopedics is that a bone-healing outcome is the product of the anabolic (bone-forming) and catabolic (bone-resorbing) outcomes. Recently, surgical and tissue engineering strategies have emerged that combine recombinant human bone morphogenetic proteins (rhBMPs) and bisphosphonates (BPs) in order to maximize anabolism and minimize catabolism. Collagen-based scaffolds that are the current surgical standard can bind rhBMPs, but not BPs. We hypothesized that a biomimetic collagen-hydroxyapatite (CHA) scaffold would bind both agents and produce superior in vivo outcomes. Consistent with this concept, in vitro elution studies utilizing rhBMP-2 ELISA assays and scintillation counting of (14)C-radiolabeled zoledronic acid (ZA) confirmed delayed release of both agents from the CHA scaffold. Next, scaffolds were tested for their capacity to form ectopic bone after surgical implantation into the rat hind limb. Using CHA, a significant 6-fold increase in bone volume was seen in rhBMP-2/ZA groups compared to rhBMP-2 alone, confirming the ability of ZA to enhance rhBMP-2 bone formation. CHA scaffolds were found to be capable of generating mineralized tissue in the absence of rhBMP-2. This study has implications for future clinical treatments of critical bone defects. It demonstrates the relative advantages of co-delivering anabolic and anti-catabolic agents using a multicomponent scaffold system.

Recent insights into bone development, homeostasis, and repair in type 1 neurofibromatosis (NF1)

Neurofibromatosis type 1 (NF1) is one of the most common single gene syndromes and is typified by a range of characteristic but variably penetrant manifestations. The orthopaedic manifestations of congenital pseudarthrosis of the tibia (CPT) and scoliosis, along with other skeletal defects including sphenoid wing dysplasia, rib penciling, and gracile bones have been classically associated with NF1. Added to this, several recent studies have reported a high prevalence of osteoporosis or osteopenia in NF1 cohorts. Clues to the underlying molecular and cellular deficiencies that cause these bony defects can be gleaned from genetically modified mouse models of Nf1 gene deficiency. These studies suggest that a variety of different cell lineages may be adversely affected by Nf1 haploinsufficiency or by double inactivation of the Nf1 gene. Osteoblasts, osteoclasts, chondrocytes, fibroblasts, and vascular endothelial cells all express the Nf1 gene product, neurofibromin, and may be functionally compromised when levels are decreased or absent. This paper reviews the current literature on NF1 bone development, homeostatic regulation, and repair, and highlights some emerging themes that may have relevance for managing orthopaedic disorders that can arise in individuals with NF1.

The contribution of different cell lineages to bone repair: Exploring a role for muscle stem cells

An anabolic response driven by osteoblasts is critical for the process of bone healing. Current evidence suggests that these osteoblasts may arise from multiple tissue types and cell lineages. Stem cells present in the bone marrow, periosteum, local soft tissues, vasculature, and/or circulation have been shown to have osteogenic potential. Transplanted cells from these sources have also been shown to incorporate into induced ectopic bone or repaired bone. While these experiments demonstrate the latent capacity of different lineages to assume an osteoblastic phenotype under pro-osteogenic conditions, the actual contribution of the different lineages to various repair situations in vivo remains unclear. This review explores the data arising from different bone formation and repair models. We propose a model suggesting that cells arising from the local tissues, particularly muscle cells, may play an important role in fracture repair under situations where the periosteal and/or bone marrow progenitor populations are depleted.

Myogenic progenitors contribute to open but not closed fracture repair

BackgroundBone repair is dependent on the presence of osteocompetent progenitors that are able to differentiate and generate new bone. Muscle is found in close association with orthopaedic injury, however its capacity to make a cellular contribution to bone repair remains ambiguous. We hypothesized that myogenic cells of the MyoD-lineage are able to contribute to bone repair.MethodsWe employed a MyoD-Cre+:Z/AP+ conditional reporter mouse in which all cells of the MyoD-lineage are permanently labeled with a human alkaline phosphatase (hAP) reporter. We tracked the contribution of MyoD-lineage cells in mouse models of tibial bone healing.ResultsIn the absence of musculoskeletal trauma, MyoD-expressing cells are limited to skeletal muscle and the presence of reporter-positive cells in non-muscle tissues is negligible. In a closed tibial fracture model, there was no significant contribution of hAP+ cells to the healing callus. In contrast, open tibial fractures featuring periosteal stripping and muscle fenestration had up to 50% of hAP+ cells detected in the open fracture callus. At early stages of repair, many hAP+ cells exhibited a chondrocyte morphology, with lesser numbers of osteoblast-like hAP+ cells present at the later stages. Serial sections stained for hAP and type II and type I collagen showed that MyoD-lineage cells were surrounded by cartilaginous or bony matrix, suggestive of a functional role in the repair process. To exclude the prospect that osteoprogenitors spontaneously express MyoD during bone repair, we created a metaphyseal drill hole defect in the tibia. No hAP+ staining was observed in this model suggesting that the expression of MyoD is not a normal event for endogenous osteoprogenitors.ConclusionsThese data document for the first time that muscle cells can play a significant secondary role in bone repair and this knowledge may lead to important translational applications in orthopaedic surgery.Please see related article: http://www.biomedcentral.com/1741-7015/9/136

Models of Tibial Fracture Healing in Normal and Nf1 -Deficient Mice

Delayed union and nonunion are common complications associated with tibial fractures, particularly in the distal tibia. Existing mouse tibial fracture models are typically closed and middiaphyseal, and thus poorly recapitulate the prevailing conditions following surgery on a human open distal tibial fracture. This report describes our development of two open tibial fracture models in the mouse, where the bone is broken either in the tibial midshaft (mid‐diaphysis) or in the distal tibia. Fractures in the distal tibial model showed delayed repair compared to fractures in the tibial midshaft. These tibial fracture models were applied to both wild‐type and Nf1‐deficient (Nf1+/−) mice. Bone repair has been reported to be exceptionally problematic in human NF1 patients, and these patients can also spontaneously develop tibial nonunions (known as congenital pseudarthrosis of the tibia), which are recalcitrant to even vigorous intervention. pQCT analysis confirmed no fundamental differences in cortical or cancellous bone in Nf1‐deficient mouse tibiae compared to wild‐type mice. Although no difference in bone healing was seen in the tibial midshaft fracture model, the healing of distal tibial fractures was found to be impaired in Nf1+/− mice. The histological features associated with nonunited Nf1+/− fractures were variable, but included delayed cartilage removal, disproportionate fibrous invasion, insufficient new bone anabolism, and excessive catabolism. These findings imply that the pathology of tibial pseudarthrosis in human NF1 is complex and likely to be multifactorial.