Annexin A1 is a biomarker of T-tubular repair in skeletal muscle of nonmyopathic patients undergoing statin therapy.

Skeletal muscle complaints are a common consequence of cholesterol-lowering therapy. Transverse tubular (T-tubular) vacuolations occur in patients having statin-associated myopathy and, to a lesser extent, in statin-treated patients without myopathy. We have investigated quantitative changes in T-tubular morphology and looked for early indicators of T-tubular membrane repair in skeletal muscle biopsy samples from patients receiving cholesterol-lowering therapy who do not have myopathic side effects. Gene expression and protein levels of incipient membrane repair proteins were monitored in patients who tolerated statin treatment without myopathy and in statin-naive subjects. In addition, morphometry of the T-tubular system was performed. Only the gene expression for annexin A1 was up-regulated, whereas the expression of other repair genes remained unchanged. However, annexin A1 and dysferlin protein levels were significantly increased. In statin-treated patients, the volume fraction of the T-tubular system was significantly increased, but the volume fraction of the sarcoplasmic reticulum remained unchanged. A complex surface structure in combination with high mechanical loads makes skeletal muscle plasma membranes susceptible to injury. Ca(2+)-dependent membrane repair proteins such as dysferlin and annexin A1 are deployed at T-tubular sites. The up-regulation of annexin A1 gene expression and protein points to this protein as a biomarker for T-tubular repair.

PMMA-Cement-PLIF Is Safe and Effective as a Single-Stage Posterior Procedure in Treating Pyogenic Erosive Lumbar Spondylodiscitis-A Single-Center Retrospective Study of 73 Cases.

BACKGROUND: Surgical treatment for erosive pyogenic spondylodiscitis of the lumbar spine is challenging as, following debridement of the intervertebral and bony abscess, a large and irregular defect is created. Sufficient defect reconstruction with conventional implants using a posterior approach is often impossible. Therefore, we developed the "Cement-PLIF", a single-stage posterior lumbar procedure, combining posterior lumbar interbody fusion (PLIF) with defect-filling using antibiotic-loaded polymethylmethacrylate (PMMA). This study first describes and evaluates the procedure's efficacy, safety, and infection eradication rate. Radiological implant stability, bone-regeneration, sagittal profile reconstruction, procedure-related complications, and pre-existing comorbidities were further analyzed. METHODS: A retrospective cohort study analyzing 73 consecutive patients with a minimum of a one-year follow-up from 2000-2017. Patient-reported pain levels and improvement in infectious serological parameters evaluated the clinical outcome. Sagittal profile reconstruction, anterior bone-regeneration, and posterior fusion were analyzed in a.p. and lateral radiographs. A Kaplan-Meier analysis was used to determine the impact of pre-existing comorbidities on mortality. Pre-existing comorbidities were quantified using the Charlson-Comorbidity Index (CCI). RESULTS: Mean follow-up was 3.3 (range: 1-16; ±3.2) years. There was no evidence of infection persistence in all patients at the one-year follow-up. One patient underwent revision surgery for early local infection recurrence (1.4%). Five (6.9%) patients required an early secondary intervention at the same level due to minor complications. Radiological follow-up revealed implant stability in 70/73 (95.9%) cases. Successful sagittal reconstruction was demonstrated in all patients (p < 0.001). There was a significant correlation between Kaplan-Meier survival and the number of pre-existing comorbidities (24-months-survival: CCI ≤ 3: 100%; CCI ≥ 3: 84.6%; p = 0.005). CONCLUSIONS: The Cement-PLIF procedure for pyogenic erosive spondylodiscitis is an effective and safe treatment as evaluated by infection elimination, clinical outcome, restoration, and maintenance of stability and sagittal alignment.

L51P - A BMP2 variant with osteoinductive activity via inhibition of Noggin.

Bone morphogenetic proteins (BMP) have to be applied at high concentrations to stimulate bone healing. The limited therapeutic efficacy may be due to the local presence of BMP antagonists such as Noggin. Thus, inhibiting BMP antagonists is an attractive therapeutic option. We hypothesized that the engineered BMP2 variant L51P stimulates osteoinduction by antagonizing Noggin-mediated inhibition of BMP2. Primary murine osteoblasts (OB) were treated with L51P, BMP2, and Noggin. OB proliferation and differentiation were quantified with XTT and alkaline phosphatase (ALP) assays. BMP receptor dependent intracellular signaling in OB was evaluated with Smad and p38 MAPK phosphorylation assays. BMP2, Noggin, BMP receptor Ia/Ib/II, osteocalcin, and ALP mRNA expressions were analyzed with real-time PCR. L51P stimulated OB differentiation by blocking Noggin mediated inhibition of BMP2. L51P did not induce OB differentiation directly and did not activate BMP receptor dependent intracellular signaling via the Smad pathway. Treatment of OB cultures with BMP2 but not with L51P resulted in an increased expression of ALP, BMP2, and Noggin mRNA. By inhibiting the BMP antagonist Noggin, L51P enhances BMP2 activity and stimulates osteoinduction without exhibiting direct osteoinductive function. Indirect osteoinduction with L51P seems to be advantageous to osteoinduction with BMP2 as BMP2 stimulates the expression of Noggin thereby self-limiting its own osteoinductive activity. Treatment with L51P is the first protein-based approach available to augment BMP2 induced bone regeneration through inhibition of BMP antagonists. The described strategy may help to decrease the amounts of exogenous BMPs currently required to stimulate bone healing.

Inhibition of endogenous antagonists with an engineered BMP-2 variant increases BMP-2 efficacy in rat femoral defect healing.

Bone morphogenetic proteins (BMP) have been used successfully by orthopedic clinicians to augment bone healing. However, these osteoinductive proteins must be applied at high concentrations to induce bone formation. The limited therapeutic efficacy may be due to the local expression of BMP antagonists such as Noggin that neutralize exogenous and endogenous BMPs. If so, inhibiting BMP antagonists may provide an attractive option to augment BMP induced bone formation. The engineered BMP-2 variant L51P is deficient in BMP receptor type I binding, but maintains its affinity for BMP receptor type II and BMP antagonists including Noggin, Chordin and Gremlin. This modification makes L51P a BMP receptor-inactive inhibitor of BMP antagonists. We implanted ß-tricalcium phosphate ceramics loaded with BMP-2 and/or L51P into a critical size defect model in the rat femur to investigate whether the inhibition of BMP antagonist with L51P enhances the therapeutic efficacy of exogenous BMP-2. Our study reveals that L51P reduces the demand of exogenous BMP-2 to induce bone healing markedly, without promoting bone formation directly when applied alone.

Long-term cell-mediated protein release from calcium phosphate ceramics.

Efficient delivery of growth factors from carrier biomaterials depends critically on the release kinetics of the proteins that constitute the carrier. Immobilizing growth factors to calcium phosphate ceramics has been attempted by direct adsorption and usually resulted in a rapid and passive release of the superficially adherent proteins. The insufficient retention of growth factors limited their bioavailability and their efficacy in the treatment of bone regeneration. In this study, a coprecipitation technique of proteins and calcium phosphate was employed to modify the delivery of proteins from biphasic calcium phosphate (BCP) ceramics. To this end, tritium-labeled bovine serum albumin ([(3)H]BSA) was utilized as a model protein to analyze the coprecipitation efficacy and the release kinetics of the protein from the carrier material. Conventional adsorption of [(3)H]BSA resulted in a rapid and passive release of the protein from BCP ceramics, whereas the coprecipitation technique effectively prevented the burst release of [(3)H]BSA. Further analysis of the in vitro kinetics demonstrated a sustained, cell-mediated release of coprecipitated [(3)H]BSA from BCP ceramics induced by resorbing osteoclasts. The coprecipitation technique described herein, achieved a physiologic-like protein release, by incorporating [(3)H]BSA into its respective carriers, rendering it a promising tool in growth factor delivery for bone healing.