Divya Bhargav

Posterolateral intertransverse spinal fusion possible in osteoporotic rats with BMP-7 in a higher dose delivered on a composite carrier.

Study Design. Posterolateral intertransverse process spinal fusion (PLF) was performed in ovariectomized female rats using recombinant human BMP-7 (OP-1) delivered on a composite carrier. Objective. To investigate whether BMP-7 collagen on a composite carrier in a higher dose will enhance posterolateral spinal fusion in an estrogen deficiency rat model. Summary of Background Data. Osteoporosis is a systemic disease characterized by bone remodeling skewed in favor of excess bone resorption. This makes new bone formation and fixation of metallic implants difficult. Thus, treating osteoporotic patients who require posterior spinal fusion is challenging. Ovariectomized rats have been used as an osteoporotic model for posterolateral intertransverse process fusion. We have demonstrated in the past that endochondral bone formation in osteoporotic rats is delayed when compared with rats without osteoporosis. We have also shown that OP-1 Putty (BMP-7, collagen, and carboxy-methyl-cellulose) can overcome the effects of osteoporosis in a rat fracture model. However, it has not yet been demonstrated whether BMP-7 collagen composite carrier (Calstrux) can achieve a fusion in a process spinal fusion model in osteoporotic bone. Methods. A total of 42 ovariectomized Sprague-Dawley female rats were randomly assigned to 4 control and 2 experimental groups: (1) no Calstrux, no BMP; (2) 400 mg Calstrux alone; (3) 30 &mgr;g lactose + 400 mg Calstrux; (4) 90 &mgr;g lactose + 400 mg Calstrux; (5) 30 &mgr;g rhBMP-7 + 400 mg Calstrux; and (6) 90 &mgr;g rhBMP-7 + 400 mg Calstrux. Spinal fusion was evaluated by manual motion testing, microradiographs, computed tomographic scans, DEXA scans, and histology. Results. Ovariectomized rats receiving Calstrux alone or either dose of lactose and Calstrux did not show spinal fusion. Ovariectomized rats receiving 90 &mgr;g BMP-7 + 400 mg Calstrux showed significantly higher fusion rates than these control animals. (P < 0.0001). The rats receiving 30 &mgr;g BMP-7 + 400 mg Calstrux exhibited only partial fusion. Conclusion. BMP-7, delivered on a composite carrier, is able to overcome the detrimental effects of estrogen deficiency on posterolateral spinal fusion and generate a relatively robust fusion. The effect seems to be dose dependent.

Hyaluronan : its potential application in intervertebral disc regeneration

Hyaluronan (HA) is a ubiquitous component of extracellular matrix in human tissues with diverse functions in skeletal biology. The biophysical properties of HA, such as high viscosity, elasticity and highly negative charge, make it useful in various therapeutic procedures. Although intra-articular administration of HA has been extensively used in the management of osteoarthritis (OA), there is a paucity of data on the clinical application of HA in intervertebral disc repair. This review discusses the biology and signaling mechanisms of HA, the patho- physiology of disc degeneration and summarises current evidence relating to the role of HA in cell phenotype maintenance, differentiation of chondrocytes, intervertebral disc cells and bone marrow stromal cells, and its application in tissue engineering. Based on recent advances in the clinical outcomes of OA treatment, HA has demonstrated potential as a bio-polymer filler, therapeutic agent and cell carrier in the management of intervertebral disc degeneration.

Bone morphogenetic protein-7 accelerates fracture healing in osteoporotic rats

Background: Osteoporosis is characterized by low bone mass, bone fragility and increased susceptibility to fracture. Fracture healing in osteoporosis is delayed and rates of implant failure are high with few biological treatment options available. This study aimed to determine whether a single dose of bone morphogenetic protein-7 (BMP-7) in a collagen/carboxy-methyl cellulose (CMC) composite enhanced fracture healing in an osteoporotic rat model. Materials and Methods: An open femoral midshaft osteotomy was performed in female rats 3 months post-ovarectomy. Rats were randomized to receive either BMP-7 composite (n = 30) or composite alone (n = 30) at the fracture site during surgery. Thereafter calluses were collected on days 12, 20 and 31. Callus cross-sectional area, bone mineral density, biomechanical stiffness and maximum torque, radiographic bony union and histological callus maturity were evaluated at each time point. Results: There were statistically significant increases in bone mineral density and callus cross-section area at all time points in the BMP-7 group as compared to controls and biomechanical readings showed stronger bones at day 31 in the BMP-7 group. Histological and radiographic evaluation indicated significant acceleration of bony union in the BMP-7 group as compared to controls. Conclusion: This study demonstrated that BMP-7 accelerates fracture healing in an oestrogen-deficient environment in a rat femoral fracture healing model to scientific relevance level I. The use of BMP-7 composite could offer orthopedic surgeons an advantage over oestrogen therapy, enhancing osteoporotic fracture healing with a single, locally applied dose at the time of surgery, potentially overcoming delays in healing caused by the osteoporotic state.

Expression and functional roles of estrogen receptor GPR30 in human intervertebral disc.

Estrogen withdrawal, a characteristic of female aging, is associated with age-related intervertebral disc (IVD) degeneration. The function of estrogen is mediated by two classic nuclear receptors, estrogen receptor (ER)-α and -β, and a membrane bound G-protein-coupled receptor 30 (GPR30). To date, the expression and function of GPR30 in human spine is poorly understood. This study aimed to evaluate GPR30 expression in IVD, and its role in estrogen-related regulation of proliferation and apoptosis of disc nucleus pulposus (NP) cells. GPR30 expression was examined in 30 human adult NP and 9 fetal IVD. Results showed that GPR30 was expressed in NP cells at both mRNA and protein levels. In human fetal IVD, GPR30 protein was expressed in the NP at 12-14 weeks gestation, but was undetectable at 8-11 weeks. The effect of 17β-estradiol (E2) on GPR30-mediated proliferation and interleukin-1β (IL-1β)-induced apoptosis of NP cells was investigated. Cultured NP cells were treated with or without E2, GPR30 antagonist G36, and ER antagonist ICI 182,780. NP cell viability was tested by MTS assay. Apoptosis was determined by flow cytometry using fluorescence labeled annexin-V, TUNEL assay and immumnocytochemical staining of activated caspase-3. E2 enhanced cell proliferation and prevented IL-1β-induced cell death, but the effect was partially blocked by G36 and completely abrogated by a combination of ICI 182,780 and G36. This study demonstrates that GPR30 is expressed in human IVD to transmit signals triggering E2-induced NP cell proliferation and protecting against IL-1β-induced apoptosis. The effects of E2 on NP cells require both GPR30 and classic estrogen receptors.

Expression of growth differentiation factor 6 in the human developing fetal spine retreats from vertebral ossifying regions and is restricted to cartilaginous tissues.

During embryogenesis vertebral segmentation is initiated by sclerotomal cell migration and condensation around the notochord, forming anlagen of vertebral bodies and intervertebral discs. The factors that govern the segmentation are not clear. Previous research demonstrated that mutations in growth differentiation factor 6 resulted in congenital vertebral fusion, suggesting this factor plays a role in development of vertebral column. In this study, we detected expression and localization of growth differentiation factor 6 in human fetal spinal column, especially in the period of early ossification of vertebrae and the developing intervertebral discs. The extracellular matrix proteins were also examined. Results showed that high levels of growth differentiation factor 6 were expressed in the nucleus pulposus of intervertebral discs and the hypertrophic chondrocytes adjacent to the ossification centre in vertebral bodies, where strong expression of proteoglycan and collagens was also detected. As fetal age increased, the expression of growth differentiation factor 6 was decreased correspondingly with the progress of ossification in vertebral bodies and restricted to cartilaginous regions. This expression pattern and the genetic link to vertebral fusion suggest that growth differentiation factor 6 may play an important role in suppression of ossification to ensure proper vertebral segmentation during spinal development.

Basic Science of Adhesive Capsulitis

Primary idiopathic adhesive capsulitis is the subset of painful stiff shoulders in which no definitive cause for the “frozen shoulder” can be identified. The incidence of adhesive capsulitis is 2% to 5% in shoulder clinics and 10% to 20% in diabetics. Adhesive capsulitis has a left-sided predominance with an age distribution between 30 and 70 years and is more common in female individuals. The duration of symptoms ranges from 12 to 42 months. When untreated, 40% to 60% of patients with adhesive capsulitis have an objective residual loss of range of motion. Why the condition resolves and why it does not occur in younger and older patients are undetermined. Arthroscopic synovial biopsies in early cases of adhesive capsulitis show increase in synovial vascularity associated with small nerves, followed by subsynovial fibroblast proliferation and capsular fibrosis in patients with longer-standing symptoms. Fibrosis of the shoulder capsule with resultant loss of capsular volume and contracture of the ligaments is the major cause for restriction of shoulder range of movement. The sub cellular events and the cytokines and the sequence of triggered genes and proteins that lead to hyperemia, neoinnervation, fibroblast proliferation, fibrosis, and bone loss in adhesive capsulitis are yet to be elucidated.