Chronic complex musculoskeletal injuries that are slow to heal pose challenges to physicians and researchers alike. Orthobiologics is a relatively newer science that involves application of naturally found materials from biological sources (for example, cell-based therapies), and offers exciting new possibilities to promote and accelerate bone and soft tissue healing. Platelet-rich plasma (PRP) is an orthobiologic that has recently gained popularity as an adjuvant treatment for musculoskeletal injuries. It is a volume of fractionated plasma from the patient's own blood that contains platelet concentrate. The platelets contain alpha granules that are rich in several growth factors, such as platelet-derived growth factor, transforming growth factor-β, insulin-like growth factor, vascular endothelial growth factor and epidermal growth factor, which play key roles in tissue repair mechanisms. PRP has found application in diverse surgical fields to enhance bone and soft-tissue healing by placing supra-physiological concentrations of autologous platelets at the site of tissue damage. The relative ease of preparation, applicability in the clinical setting, favorable safety profile and possible beneficial outcome make PRP a promising therapeutic approach for future regenerative treatments. However, there is a large knowledge gap in our understanding of PRPs mechanism of action, which has raised skepticism regarding its potential efficacy and use. Thus, the aim of this review is to describe the various factors proposed to contribute to the biological activity of PRP, and the published pre-clinical and clinical evidence to support it. Additionally, we describe the current techniques and technology for PRP preparation, and review the present shortcomings of this therapy that will need to be overcome if it is to gain broad acceptance.

Recombinant parathyroid hormone (rPTH) therapy has been evaluated for skeletal repair in animal studies and clinical trials based on its known anabolic effects, but its effects on angiogenesis and fibrosis remain poorly understood. We examined the effects of rPTH therapy on blood vessel formation and osseous integration in a murine femoral allograft model, which caused a significant increase in small vessel numbers, and decreased large vessel formation (p < 0.05). Histology showed that rPTH also reduced fibrosis around the allografts to similar levels observed in live autografts, and decreased mast cells at the graft-host junction. Similar effects on vasculogenesis and fibrosis were observed in femoral allografts from Col1caPTHR transgenic mice. Gene expression profiling revealed rPTH induced angiopoietin-1 (8-fold), while decreasing angiopoietin-2 (70-fold) at day 7 of allograft healing. Finally, we demonstrate anti-angiopoietin-2 peptibody(L1-10) treatment mimics rPTH effects on angiogenesis and fibrosis. Collectively, these findings demonstrate that intermittent rPTH treatment enhances structural allograft healing by two processes: 1) anabolic effects on new bone formation via small vessel angiogenesis, and 2) inhibition of angiopoietin-2 mediated arteriogenesis. The latter effect may function as a vascular sieve to limit mast cell access to the site of tissue repair, which decreases fibrosis around and between the fractured ends of bone. Thus, rPTH therapy may be generalizable to all forms of tissue repair that suffer from limited biointegration and excessive fibrosis.

Management of various tumor metastases to bone has dramatically improved, but this is not so for renal cell carcinoma (RCC), which is a difficult surgical problem due to its great vascularity. Furthermore, the unique mechanisms that mediate RCC vasculogenesis in bone remain unknown. To understand this process we developed a xenograft model that recapitulates highly vascular RCC vs. less vascular tumors that metastasize to bone. Human tumor cell lines of RCC (786-O), prostate cancer (PC3), lung cancer (A549), breast cancer (MDA-MB231) and melanoma (A375) were transfected with firefly luciferase (Luc), injected into the tibiae of nude mice, and differences in growth, osteolysis and vascularity were assessed by longitudinal bioluminescent imaging (BLI), micro-CT for measurement of calcified tissues and vascularity and histology. The results showed that while RCC-Luc has reduced growth and osteolytic potential vs. the other tumor lines, it displayed a significant increase in vascular volume (p<0.05). This expansion was due to 3- and 5-fold increases in small and large vessel numbers respectively. In vitro gene expression profiling revealed that RCC-Luc expresses significantly (p<0.05) more vegf-a (10-fold) and 20-30-fold less ang-1 vs. the other lines. These data demonstrate the utility of this model to study the unique vasculogenic properties of RCC bone metastases.

Critically sized large bone defects commonly result from trauma, radical tumor resections or infections. Currently, massive allografting remain as the clinical standard to treat these critical defects. Unfortunately, allograft healing is limited by the lack of osteogenesis and bio-integration of the graft to the host bone. Based on its widely studied anabolic effects on the bone, we have proposed that teriparatide [recombinant parathyroid hormone (PTH1–34)] could be an effective adjuvant for massive allograft healing. In support of this theory, here we review studies that have demonstrated that intermittent PTH1–34 treatment enhances and accelerates the skeletal repair process via a number of mechanisms including: effects on mesenchymal stem cells (MSC), angiogenesis, chondrogenesis, bone formation and remodeling. We also review the current literature on the effects of PTH1–34 therapy on bone healing, and discuss this drug’s long term potential as an adjuvant for endogenous tissue engineering.

SUMMARY Preclinical studies on bone repair remain a high priority due to the unresolved clinical problems associated with treating critical segmental defects and complications of fracture healing. Over the last decade the murine femoral allograft model has gained popularity due to its standardized surgery and potential for examining a vast array of radiographic, biomechanical and histological outcome measures. Here, we describe these methods and a novel semi-automated histomorphometric approach to quantify the amount of bone, cartilage and undifferentiated mesenchymal tissue in demineralized paraffin sections of allografted murine femurs using the VisioPharm Image Analysis Software System.