Continuous occurrence of both insufficient neovascularization and elevated vascular permeability in rabbit proximal femur during inadequate repair of steroid‐associated osteonecrotic lesions

Abstract

Objective. To examine the features of the intraosseous vasculature, the size of the marrow stem cell pool (MSCP), and expression of vascular endothelial growth factor A (VEGF) during inadequate repair of steroid-associated osteonecrotic lesions in rabbits.Methods. Steroid-associated osteonecrosis was induced in male rabbits. At 0, 1, 2, 4, and 6 weeks postinduction, vascularization and permeability indices were quantified by dynamic magnetic resonance imaging (MRI). In addition, the size of the MSCP in the hematopoietic and mesenchymal compartments was determined, and marrow mononuclear cells expressing specific surface markers for endothelial progenitor cells or periendothelial mural precursor cells were counted. At various time points after the rabbits were killed, the proximal femora were dissected to examine the intraosseous vasculature by angiography, histomorphometry, and ultramorphology. In addition, osteonecrotic lesion repair and marrow VEGF expression were evaluated.Results. Lesion formation without repair was observed at 2 weeks after induction of steroid-associated osteonecrosis. Rabbits displaying destructive repair (DR؉) and those displaying reparative osteogenesis (DR؊) from 4 weeks to 6 weeks postinduction were identified. From week 2 to week 6, the vascularization index was significantly lower in DR؉ rabbits compared with DR؊ rabbits, whereas the permeability index was significantly higher in DR؉ rabbits compared with DR؊ rabbits. The features of the intraosseous vasculature determined by angiography, histomorphometry, and ultramorphology were consistent with those determined by dynamic MRI. The MSCP size and number of marrow mononuclear cells expressing specific surface markers were all significantly lower in DR؉ rabbits than in DR؊ rabbits from week 1 to week 6. The increased VEGF expression at 2 weeks was maintained through week 6 in DR؉ rabbits, whereas VEGF expression decreased in DR؊ rabbits from week 2 to week 6.Conclusion. Continuous occurrence of both insufficient neovascularization and elevated vascular permeability is accompanied by a continuously lowlevel MSCP and uncontrolled VEGF expression during inadequate repair of steroid-associated osteonecrotic lesions.Pulsed steroids are frequently prescribed as lifesaving agents for serious infectious diseases such as severe acute respiratory syndrome and acquired immunodeficiency syndrome, or as disease-modifying agents for chronic autoimmune diseases such as systemic lupus erythematosus and rheumatoid arthritis. However, pulsed administration of steroids often results in osteonecrosis (also known as avascular necrosis). Orthopedic surgeons face the challenge of managing subchondral

Low-intensity Pulsed Ultrasound Enhances Bone Repair in a Rabbit Model of Steroid-associated Osteonecrosis

“…Osteogenesis and neovascularization play important roles during bone repair. In addition to the impaired activity of MSCs in the steroid-associated osteonecrosis rabbit model, it has been reported that there are effects on hematopoietic stem cells in this model [40]. Neovascularization is thought to be the result of hematopoietic stem cells and endothelial progenitor cell activity.…”

“…This might be because there were more, and smaller, new vessels in LIPUS-treated limbs. Because sufficient blood flow is a fundamental prerequisite for bone repair [40], LIPUS may be a potential approach for improving treatment of early-stage steroid-associated osteonecrosis.…”

Blockage of Src by Specific siRNA as a Novel Therapeutic Strategy to Prevent Destructive Repair in Steroid-Associated Osteonecrosis in Rabbits

“…A histogram was generated to display the thickness of the angiographic vessels. (30,34) Histological evaluation After decalcification by 10% EDTA, the proximal femurs were embedded in paraffin and cut into 5-mm-thick sections along the coronal plane, except for those used for fluorescence labeling. Sections were stained with H&E for routine histology and with tartrate-resistant acid phosphatase (TRAP; Sigma Diagnostics, St. Louis, MO, USA) for identification of osteoclasts.…”

“…(43) Appositional bone formation with osteoblast-like cells around the osteonecrotic lesion was classified as reparative osteogenesis, whereas fibrotic tissue creep linked to necrotic bone resorption was classified as destructive repair. (30) Evaluation of the repair pattern was done as follows: the entire area of each section, including epiphysis and metaphysis, was examined based on the characteristic histopathological features blindly by two pathologists. We quantified the destructive repair region and the reparative osteogenesis region on the entire section of each specimen.…”

“…(32) We aim to determine the role of VEGF in destructive repair by supplementing and neutralizing VEGF, respectively, and define the role of Src through blocking Src by means of in vivo gene knockdown technology-RNAi using siRNA (33) with and without supplement of VEGF in a rabbit SAON model. (30,34) …”