Experimental Induction of Physeal Injuries by Fracture, Drill, and Ablation Techniques: Analyses of Immunohistochemical Findings.

BACKGROUND: Although physeal fractures and physeal bars can result in significant clinical consequences to growth and development of the injured physis, little orthopaedic research has focused upon this topic. Our objective was to extend a previously developed rat model to examine the immunohistochemical features following surgical application of techniques disrupting the physis. METHODS: Physes were surgically disrupted using fracture (control), epiphyseal scrape (ES), or epiphyseal drill (ED). After 1, 3, 6, 10, or 21 days, animals were euthanized, sites processed for histology and immunohistochemical localization of vascular endothelial growth factor (VEGF), Factor VIII, Sox-9, PTHrP (parathyroid hormone-related protein) and PTHrP-R (parathyroid hormone-related protein receptor) in resting, proliferative, and hypertrophic physeal zones. Incidence of physeal bars, vertical septa and islands within the metaphysis was quantified. Semiquantitative analysis of immunohistochemistry was performed. RESULTS: Physeal bars, vertical septa, and displaced cartilage islands were present each of the surgical treatments. Fisher's exact test showed a statistically significant increase in the presence of physeal bars (P=0.002) and vertical septa (P=0.012) in the ED group at 10 and 21 days. Analysis of VEGF showed significant differences among the surgical treatments involving the resting zone, and the proliferative zone for days 1, 6, and 21 (P≤0.02) with greater mean scores present in the fracture (control) group, followed by the ED group; the lowest scores were present in the ES group. PTHrP-R immunolocalization showed significant differences among treatments in the hypertrophic zone at days 6 and 21 (P=0.022 and 0.044, respectively). CONCLUSIONS: On the basis of the type of surgical treatment, results show significant differences in the presence of VEGF (reflecting the vascular bed) in the resting and proliferating zones at days 1, 6, and 21. VEGF localization was less abundant in the ED group (which had more physeal bars), suggesting that lack of vascular ingrowth plays a role in physeal bar formation. CLINICAL RELEVANCE: Basic science data presented here provide insight into the importance of the various regions of the physis and its repair and continued growth after physeal fracture. We suggest that a better understanding of the cellular basis of physeal arrest following physeal fracture may have future relevance for the development of treatments to prevent or correct arrest.

The Role of the Basement Plate in Physeal Bar Formation.

BACKGROUND: Physeal fractures and resultant physeal bars can pose significant problems in skeletal development for the injured growing child. Although now well-recognized, only a small body of experimental literature covering this problem is available. The goal of this study was to help further develop an understanding of the different regions of the physis and the way in which each region responds to injury/fracture. METHODS: This Institutional Animal Care and Use Committee (IACUC)-approved study assessed bar formation using radiologic and histologic methods and measured leg lengths of skeletally immature rats. The right tibia was used as the control to measure leg length discrepancy (LLD), and the left tibia received either a fracture only (F), an epiphyseal scrape (ES), an epiphyseal drilling procedure (ED), or metaphyseal drilling (MD). Radiographs and LLD measurements were obtained at postoperative days 0, 21, and 56. RESULTS: A significant LLD was present at day 56 in the ED group (P=0.01). Radiographic identification of bars showed significant evidence of bar formation for the ES and ED groups at 21 days and the ED group at 56 days (P<0.05). Histologic examination showed a high incidence of histologic physeal bar formation in the ES, ED, and MD groups at 21 and 56 days. CONCLUSIONS: Findings showed that the physis was able to continue to grow following an injury to the physis' hypertrophic region. MD produced little effects with few physeal bars and little LLD. By postoperative day 56, ED animals showed greater LLD than ES animals. Penetration of the basement plate was more likely to lead to bar formation/growth retardation than was ablation of the epiphyseal region of the physis (including resting cells). CLINICAL RELEVANCE: Data presented here provides insight into the importance of different regions of the physis and its repair/continued growth after physeal fracture. We suggest that a better understanding of the physiological cause of physeal arrest after physeal fracture will be important for the development of treatments to prevent physeal arrest or to treat physeal arrest after it occurs.

Glucocorticoid Steroid and Alendronate Treatment Alleviates Dystrophic Phenotype with Enhanced Functional Glycosylation of α-Dystroglycan in Mouse Model of Limb-Girdle Muscular Dystrophy with FKRPP448L Mutation.

Fukutin-related protein-muscular dystrophy is characterized by defects in glycosylation of α-dystroglycan with variable clinical phenotypes, most commonly as limb-girdle muscular dystrophy 2I. There is no effective therapy available. Glucocorticoid steroids have become the standard treatment for Duchenne and other muscular dystrophies with serious adverse effects, including excessive weight gain, immune suppression, and bone loss. Bisphosphonates have been used to treat Duchenne muscular dystrophy for prevention of osteoporosis. Herein, we evaluated prednisolone and alendronate for their therapeutic potential in the FKRPP448L-mutant mouse representing moderate limb-girdle muscular dystrophy 2I. Mice were treated with prednisolone, alendronate, and both in combination for up to 6 months. Prednisolone improved muscle pathology with significant reduction in muscle degeneration, but had no effect on serum creatine kinase levels and muscle strength. Alendronate treatment did not ameliorate muscle degeneration, but demonstrated a limited enhancement on muscle function test. Combined treatment of prednisolone and alendronate provided best improvement in muscle pathology with normalized fiber size distribution and significantly reduced serum creatine kinase levels, but had limited effect on muscle force generation. The use of alendronate significantly mitigated the bone loss. Prednisolone alone and in combination with alendronate enhance functionally glycosylated α-dystroglycan. These results, for the first time, demonstrate the efficacy and feasibility of this alliance treatment of the two drugs for fukutin-related protein-muscular dystrophy.

Matrix metalloproteinase-12 immunolocalization in the degenerating human intervertebral disc and sand rat spine: Biologic implications.

Matrix metalloproteinase-12 (MMP-12; macrophage metalloelastase) degrades a number of extracellular matrix components which are present in the intervertebral disc, including type IV collagen, fibronectin, laminin, chondroitin sulfates, elastin and fibrinogen. MMP-12 has recently discovered relationships with cytokines and chemokines which also relate to disc cell biology. To date, no study has assessed immunolocalization of MMP-12 in degenerating human intervertebral disc tissue. Immunocytochemical localization was performed on 18 human disc specimens and on lumbar spines of the sand rat, a small animal model with well-recognized age-related disc degeneration. In the human disc, intracellular localization was present in both the annulus and nucleus portions of the disc. The sand rat degenerating disc also showed MMP-12 disc localization, with additional presence in chondrocytes of the vertebral endplate of older animals. This is the initial characterization of the presence of MMP-12 in the human and sand rat disc, and in chondrocytes of the vertebral endplate in older sand rats with degenerating discs. Findings are important because they document the presence of an additional MMP-12 in disc tissue, thus expanding our understanding of disc extracellular matrix remodeling, and because they provide novel information on the presence of MMP-12 in the cartilage endplate as it undergoes sclerosis during disc degeneration in the aging sand rat.