Indian Journal of Biochemistry & Biophysics Vol. 43, June 2006, pp. 167-172 Augmented bone-matrix formation and osteogenesis under magnetic field stimulation in vivo XRD, TEM and SEM investigations Praveen Singh 1* , Rakesh C YashRoy 1 and M Hoque 2 1 Biophysics and Electron Microscopy Section, 2 Surgery Division, Indian Veterinary Research Institute, Izatnagar-243122, UP, India Received 02 June 2005; revised 25 January 2006 Bone is a composite biomaterial, which is formed, when proteins constituting collagen fibers attract calcium, phosphate and hydroxide ions in solution to nucleate atop the fibers. It grows into a hard structure of tiny crystallites of hydroxyapatite, aligned along the long axis of collagen fibers. The present work reports the stimulating effect of static magnetic field on microstructure and mineralization process of bone repair. A unilateral transverse fracture of mid-shaft of metacarpal was surgically created in healthy goats under thiopental sedation and xylocaine analgesia. Two bar magnets (~ 800 gauss/cm 2 field strength) were placed across the fracture line at opposite pole alignment immobilized in Plaster of Paris (POP) splint bandage for static magnetic field stimulation. Radiographs were taken at weekly intervals up to 45 days. Results show that formation of extra-cellular matrix and its microstructure can be influenced by non-invasive physical stimulus (magnetic field) for achieving an enhanced osteogenesis, leading to quicker regeneration of bone tissue in goats. X-ray diffraction (XRD) patterns of treated (magnetic field-exposed) and control samples revealed the presence and orientation of crystalline structures. Intensity of diffraction peaks corresponding to 310 and 222 planes were enhanced with respect to 211 families of reflections, indicating preferential alignment of the crystals. Also, the percent crystallinity and crystal size were increased in treated samples. The study provides a biophysical basis for augmented fracture healing under the influence of semi-aligned static magnetic field applied across the fracture line. Keywords: Fracture healing, Static magnetic field, Hydroxyapatite, Collagen fibers, XRD pattern, Electron microscopy There has been an increasing interest to investigate the effect of magnetic and electric fields on the biological systems 1-7 . Clinical studies have demonstrated the usefulness of magnetic and electromagnetic fields, in stimulating healing of chronically disjointed fractures in humans/animals 3-7 . The devices generating such fields have been approved for clinical use. The osteo-stimulating effect of pulse magnetic field (MF) has been reported 7,8 . Magnetic field has also been successfully used in treating delayed union 9 , non-union 10 and long- established cases of pseudo-arthroses 11 . Despite these studies, the biological processes underlying osteogenesis remain, only partially understood. In fact, experimental approach to the magnetic field effect on living system is complicated by various non-linearities (intensity, frequency and time windows of the fields) and peculiarities (cell type, age and treatment). However, considerable progress has been made to understand the cellular and molecular changes associated with application of biophysical stimuli such as electromagnetic field 11-15 and static magnetic field 16 . Each of these systems has shown substantial efficacy in controlled clinical trials 17 . In this paper, an attempt has been made to understand the effects of non-invasive stimulation by static magnetic field on morphology, ultra- microstructure of bone and on mineralization process of the fractured tissue. The results obtained are discussed in the light of existing models for coupling of magnetic field with bone tissue, bone cells and associated effects on extra-cellular matrix formation. The study throws light on modus operandi of augmented bone fracture healing process under the influence of static magnetic field. _____________ *Corresponding author Tel: 91-09359108482; Fax: 91-581-2303284, E-mail: psingh67@yahoo.com ;  Presently on deputation to Department of Electronics, Kyushu University, Fukuoka, Japan Ph 81-92-6427057, Fax 81-92-6423967 E-mail singh@belab.ed.kyushu-u.ac.jp Abbreviations MF, magnetic field; HAP, hydroxyapatite; XRD, X-ray diffraction; SEM, scanning electron microscopy; TEM, transmission electron microscopy.