Low-intensity pulsed ultrasound affects RUNX2 immunopositive osteogenic cells in delayed clinical fracture healing

Introduction: Osteogenic cell proliferation and differentiation play an important role in adequate fracture healing, and is target for osteoinductive therapies in delayed fracture healing. The aim of this study was to investigate whether low-intensity pulsed ultrasound enhances fracture healing at the tissue level in patients with a delayed union of the osteotomized fibula through an effect on the presence of RUNX2 immunopositive osteogenic cells. The effect was studied in both atrophic and hypertrophic delayed unions. Materials and methods: Biopsies were obtained from 6 female and 1 male patient (age 43-63) with a delayed union of the osteotomized fibula after a high tibial osteotomy treated for 2-4 months with or without low-intensity pulsed ultrasound in a randomized prospective double-blind placebo-controlled trial. Immunolocalization of RUNX2 protein was performed to identify osteogenic cells. Histomorphometrical analysis was performed to determine the number of cells expressing RUNX2 located within and around the newly formed woven bone at the fracture end (area of new bone formation), and up to 3 mm distant from the fracture end. Results: Cells expressing RUNX2 were present in all histological sections of control and low-intensity pulsed ultrasound-treated bone evaluated. Within the area of new bone formation, RUNX2 immunopositive cells were found in the undifferentiated soft connective tissue, at the bone surface (presumably osteoblasts), and within the newly formed woven bone. Low-intensity pulsed ultrasound treatment of fibula delayed unions significantly reduced the number of RUNX2 immunopositive cells within the soft connective tissue at the fracture ends, whereas the number of RUNX2 immunopositive cells at the bone surface was not affected. The number of RUNX2 immunopositive cells was similar for the atrophic and hypertrophic delayed unions. Conclusions: Immunolocalization of RUNX2 positive cells in delayed unions of the fibula reveals that delayed clinical fracture healing does not result in impairment of osteogenic cell proliferation and/or differentiation at the tissue level, even if delayed unions are clinically regarded as atrophic. Reduced number of osteogenic RUNX2 immunopositive cells within the soft connective tissue, and unchanged number of RUNX2 immunopositive cells at the bone surface, implicate that low-intensity pulsed ultrasound does not increase osteogenic cell presence, but likely affects osteogenic cell differentiation

Low-intensity pulsed ultrasound affects human articular chondrocytes in vitro

We investigated whether low-intensity pulsed ultrasound (LIPUS) stimulates chondrocyte proliferation and matrix production in explants of human articular cartilage obtained from donors suffering from unicompartimental osteoarthritis of the knee, as well as in isolated human chondrocytes in vitro. Chondrocytes and explants were exposed to LIPUS (30 mW/cm2; 20 min/day, 6 days). Stimulation of [35S]-sulphate incorporation into proteoglycans by LIPUS was 1.3-fold higher in degenerative than in collateral monolayers as assessed biochemically and 1.9-fold higher in explants as assessed by autoradiography. LIPUS decreased the number of cell nests containing 1-3 chondrocytes by 1.5 fold in collateral and by 1.6 fold in degenerative explants. LIPUS increased the number of nests containing 4-6 chondrocytes by 4.8 fold in collateral and by 3.9 fold in degenerative explants. This suggests that LIPUS stimulates chondrocyte proliferation and matrix production in chondrocytes of human articular cartilage in vitro. LIPUS might provide a feasible tool for cartilage tissue repair in osteoarthritic patients, since it stimulates chondrocyte proliferation and matrix production