Does early post-operative exercise influence bone healing kinetics? Preclinical evaluation of non-critical sized femur defect healing.

Physical exercise is a well-known modality for maintaining healthy locomotor mechanism. A detailed preclinical research on physical exercise effect on bone healing kinetics could help to improve the rehabilitation process after fracture treatment and bone remodeling. Our aim was to evaluate the effect of early post-operative exercise effect on bone microstructural changes in a rat model. Twenty Sprague Dawley male rats underwent bi-cortical 1.6 mm hole drilling in both femur diaphysis, after which (n = 10) underwent continuous treadmill training (TR) over two weeks, while the other group of rats (n = 10) was assigned to non-training (NT) control group. New bone formation labeling was performed by subcutaneous fluorochrome injections at day 5, 14 and 31. In vivo micro-computed tomography (µCT) scans were performed once a week during the 6-week post-operative period. Ten animals (five from each group) were euthanized at 3rd week while remaining animals were euthanized at 6th week. Femur samples were extracted and underwent ex vivo µCT and histological evaluation, while serum was used for evaluating alkaline phosphatase (ALP). µCT data demonstrated increased volume and surface of newly formed bone in defect area of TR group. Bone volume/Tissue volume (BV/TV) ratio and number of osteocytes showed an increase in TR group after 3-week period. Fluorochrome distances were increased between day 5 and 14 within the training group. Serum ALP level increased in both groups over 3- and 6-weeks. Post-operative exercise increases the bone healing kinetics and stimulates the new bone formation during and after the training protocol has ended.

The combined effect of zinc and calcium on the biodegradation of ultrahigh-purity magnesium implants.

Magnesium (Mg)-based implants are promising candidates for orthopedic interventions, because of their biocompatibility, good mechanical features, and ability to degrade completely in the body, eliminating the need for an additional removal surgery. In the present study, we synthesized and investigated two Mg-based materials, ultrahigh-purity ZX00 (Mg-Zn-Ca; <0.5 wt% Zn and <0.5 wt% Ca, in wt%; Fe-content <1 ppm) and ultrahigh-purity Mg (XHP-Mg, >99.999 wt% Mg; Fe-content <1 ppm), in vitro and in vivo in juvenile healthy rats to clarify the effect of the alloying elements Zn and Ca on mechanical properties, microstructure, cytocompatibility and degradation rate. Potential differences in bone formation and bone in-growth were also assessed and compared with state-of-the-art non-degradable titanium (Ti)-implanted, sham-operated, and control (non-intervention) groups, using micro-computed tomography, histology and scanning electron microscopy. At 6 and 24 weeks after implantation, serum alkaline phosphatase (ALP), calcium (Ca), and Mg level were measured and bone marrow stromal cells (BMSCs) were isolated for real-time PCR analysis. Results show that ZX00 implants have smaller grain size and superior mechanical properties than XHP-Mg, and that both reveal good biocompatibility in cytocompatibilty tests. ZX00 homogenously degraded with an increased gas accumulation 12 and 24 weeks after implantation, whereas XHP-Mg exhibited higher gas accumulation already at 2 weeks. Serum ALP, Ca, and Mg levels were comparable among all groups and both Mg-based implants led to similar relative expression levels of Alp, Runx2, and Bmp-2 genes at weeks 6 and 24. Histologically, Mg-based implants are superior for new bone tissue formation and bone in-growth compared to Ti implants. Furthermore, by tracking the sequence of multicolor fluorochrome labels, we observed higher mineral apposition rate at week 2 in both Mg-based implants compared to the control groups. Our findings suggest that (i) ZX00 and XHP-Mg support bone formation and remodeling, (ii) both Mg-based implants are superior to Ti implants in terms of new bone tissue formation and osseointegration, and (iii) ZX00 is more favorable due to its lower degradation rate and moderate gas accumulation.

Implant degradation of low-alloyed Mg-Zn-Ca in osteoporotic, old and juvenile rats.

Implant removal is unnecessary for biodegradable magnesium (Mg)-based implants and, therefore, the related risk for implant-induced fractures is limited. Aging, on the other hand, is associated with low bone-turnover and decreased bone mass and density, and thus increased fracture risk. Osteoporosis is accompanied by Mg deficiency, therefore, we hypothesized that Mg-based implants may support bone formation by Mg2+ ion release in an ovariectomy-induced osteoporotic rat model. Hence, we investigated osseointegration and implant degradation of a low-alloyed, degrading Mg-Zn-Ca implant (ZX00) in ovariectomy-induced osteoporotic (Osteo), old healthy (OH), and juvenile healthy (JH) groups of female Sprague Dawley rats via in vivo micro-computed tomography (µCT). For the Osteo rats, we demonstrate diminished trabecular bone already after 8 weeks upon ovariectomy and significantly enhanced implant volume loss, with correspondingly pronounced gas formation, compared to the OH and JH groups. Sclerotic rim development was observed in about half of the osteoporotic rats, suggesting a prevention from foreign-body and osteonecrosis development. Synchrotron radiation-based µCT confirmed lower bone volume fractions in the Osteo group compared to the OH and JH groups. Qualitative histological analysis additionally visualized the enhanced implant degradation in the Osteo group. To date, ZX00 provides an interesting implant material for young and older healthy patients, but it may not be of advantage in pharmacologically untreated osteoporotic conditions. STATEMENT OF SIGNIFICANCE: Magnesium-based implants are promising candidates for treatment of osteoporotic fractures because of their biodegradable, biomechanical, anti-bacterial and bone regenerative properties. Here we investigate magnesium‒zinc‒calcium implant materials in a rat model with ovariectomy-induced osteoporosis (Osteo group) and compare the related osseointegration and implant degradation with the results obtained for old healthy (OH) and juvenile healthy (JH) rats. The work applied an appropriate disease model for osteoporosis and focused in particular on long-term implant degradation for different bone conditions. Enhanced implant degradation and sclerotic rim formation was observed in osteoporotic rats, which illustrates that the setting of different bone models generates significantly modified clinical outcome. It further illustrated that these differences must be taken into account in future biodegradable implant development.

Anatomical similarity between the Sost-knockout mouse and sclerosteosis in humans.

Sclerosteosis, a rare autosomal recessive genetic disorder caused by a mutation of the Sost gene, manifests in the facial skeleton by gigantism, facial distortion, mandibular prognathism, cranial nerve palsy, and, in extreme cases, compression of the medulla oblongata. Mice lacking sclerostin reflect some symptoms of sclerosteosis, but this is the first report of the effect on the facial skeleton. We used geometric morphometrics (GMM) to analyze the deformations of the murine facial skeleton from the wild-type to the Sost gene knockout. Landmark coordinates were obtained by surface reconstructions from micro-computed tomography. Centroid size, principal component scores in shape space and form space, and asymmetry were computed by the standard GMM formulas, and dental and skeletal jaw lengths were examined as ratios. We show here that, compared to wild type controls, mice lacking Sost have larger centroid size (effect size, p-value: 4.59, <.001), higher mean asymmetry (1.14, .065), dental and skeletal mandibular prognathism (1.36, .010 and 5.92, <.001), a smaller foramen magnum (-1.71, .015), and calvaria that are more highly curved (form space p = 4.09, .002; shape space p = 12.82, .002). These features of mice lacking sclerostin largely correspond to the changes of the facial skeleton observed in sclerosteosis. This alignment further supports claims that the Sost gene plays a fundamental role in bony facial development in rodents and humans alike.

Impaired periodontium and temporomandibular joints in tumour necrosis factor-α transgenic mice.

AIM: Tumour necrosis factor (TNF)-α is a pathological factor causing the characteristic symptoms of periodontal disease and rheumatoid arthritis. In this study, we describe the phenotypes of human TNF-α transgenic mice (hTNFtg) with respect to their periodontium and the temporomandibular joint (TMJ). MATERIAL AND METHODS: Periodontal structures, TMJ and skull shape of hTNFtg mice and wild-type (WT) littermates were assessed by microcomputed tomography, automated segmentation, geometric morphometrics and histologic ground sections. RESULTS: We show that hTNFtg mice have an eroded lamina dura and reduced periodontal ligament space compared to (WT) littermates. Transgenic mice further exhibit severe destruction of the TMJ. Geometric morphometrics revealed that hTNFtg mice have a more laterally positioned TMJ with a concomitantly enlarged zygomatic process. Mandibular and maxillary teeth occluded properly. CONCLUSIONS: Our findings suggest that chronic inflammation in hTNFtg mice causes destructive changes of the periodontium and the TMJ.

Dimethyloxalylglycine lyophilized onto bone substitutes increase vessel area in rat calvarial defects.

AIM: Pharmacological inhibitors of prolyl hydroxylases, also termed hypoxia-mimetic agents (HMAs), when repeatedly injected can support angiogenesis and bone regeneration. However, the possible role of HMA loaded onto bone substitutes to support angiogenesis and bone regeneration under diabetic condition is unknown. The capacity of HMA loaded onto deproteinized bovine bone mineral (DBBM) to support angiogenesis and bone formation was examined in diabetic Wistar rats. METHODS: Diabetes was induced by intraperitoneal injection of streptozotocin. The HMA dimethyloxalylglycine (DMOG) and desferrioxamine (DFO) were lyophilized onto DBBM. Calvarial defects were created with a trephine drill and filled with the respective bone substitutes. After 4 weeks of healing, the animals were subjected to histological and histomorphometric analysis. RESULTS: In this report, we provide evidence that DMOG loaded onto DBBM can support angiogenesis in vivo. Specifically, we show that DMOG increased the vessel area in the defect site to 2.4% ± 1.3% compared with controls 1.1% ± 0.48% (P = 0.012). There was a trend toward an increased vessel number in the defect site with 38.6 ± 17.4 and 31.0 ± 10.3 in the DMOG and the control group (P = 0.231). The increase in angiogenesis, however, did not translate into enhanced bone formation in the defect area with 9.2% ± 7.1% and 8.4% ± 5.6% in DMOG and control group, respectively. No significant changes were caused by DFO. CONCLUSIONS: The results suggest that DMOG loaded onto DBBM can support angiogenesis, but bone formation does not increase accordingly in a type 1 diabetic rat calvarial defect model at the indicated time point.

Morphometric characteristics of cortical and trabecular bone in atrophic edentulous mandibles.

OBJECTIVES: Adaptations of the alveolar ridge after tooth loss have been well described. However, studies on the morphometric characteristics of cortical bone are rare; hence, this study of human atrophic edentulous mandibles was undertaken. MATERIAL AND METHODS: Total cortical area, porosity, and thickness, and the percentage of cortical area in the complete mandibular area as well as in an area (height, 10 mm) starting at the most caudal point of the trabecular compartment and extending in the coronal direction were determined in 185 thin ground sections of edentulous mandibles (incisor region, 49; premolar region, 76; molar region, 60; 95 from females and 90 from males; mean age, 78.2 years, SD ± 7.8 years; Caucasian donors; cause of death: cardiovascular disease). Further, mandibular height and width and degree of residual ridge resorption (RRR) were recorded. RESULTS: The percentage of cortical area in the complete mandibular area increased with increasing RRR. Yet, evaluation of the 10-mm caudal portion corresponding to the basal part of the mandibular body did not confirm these changes in cortical bone. Cortical porosity and thickness decreased from the mesial to the distal region. Cortical porosity was unaffected by RRR, while cortical thickness increased, mainly at lingual aspects. CONCLUSIONS: In conclusion, cortical bone remained stable in different degrees of RRR except for some modulations in the lingual aspects. Changes in the relative composition between cortical and trabecular bone are due to loss of height and total area, mainly at expense of trabecular bone area, but not to adaptations of the cortical bone.

Efficacy of fosfomycin compared to vancomycin in treatment of implant-associated chronic methicillin-resistant Staphylococcus aureus osteomyelitis in rats.

Fosfomycin monotherapy was compared to therapy with vancomycin for the treatment of implant-associated methicillin-resistant Staphylococcus aureus (MRSA) osteomyelitis in an experimental rat model. The proximal tibiae were inoculated with 15 µl of a suspension containing 1×10(8) to 5×10(8) CFU/ml of a clinical isolate of MRSA with simultaneous insertion of a titanium wire. Four weeks later, treatment was started for 28 days with either 50 mg/kg of body weight vancomycin intraperitoneally twice daily (n=11) or 75 mg/kg fosfomycin intraperitoneally once daily (n=10). Eleven animals were left untreated. After treatment, quantitative cultures from bone were found to be positive for MRSA in all animals in the untreated group (median, 3.29×10(6) CFU/g of bone) and the vancomycin group (median, 3.03×10(5) CFU/g of bone). In the fosfomycin group, MRSA was detectable in 2 out of 10 (20%) animals (3.42×10(2) and 1.51×10(3) CFU/g of bone). Vancomycin was superior to the no-drug control (P=0.002), and fosfomycin was superior to the no-drug control and vancomycin (P<0.001). The cultures from the wires were positive in all untreated animals (median, 2.5×10(3) CFU/implant), in 10 animals in the vancomycin group (median, 1.15×10(3) CFU/implant), and negative in all animals in the fosfomycin group. Based on the bacterial counts from the implants, vancomycin was not superior to the no-drug control (P=0.324), and fosfomycin was superior to the no-drug control and vancomycin (P<0.001). No emergence of resistance was observed. In conclusion, it was demonstrated that fosfomycin monotherapy is highly effective for the treatment of experimental implant-associated MRSA osteomyelitis.

Dental and periodontal phenotype in sclerostin knockout mice.

Sclerostin is a Wnt signalling antagonist that controls bone metabolism. Sclerostin is expressed by osteocytes and cementocytes; however, its role in the formation of dental structures remains unclear. Here, we analysed the mandibles of sclerostin knockout mice to determine the influence of sclerostin on dental structures and dimensions using histomorphometry and micro-computed tomography (µCT) imaging. µCT and histomorphometric analyses were performed on the first lower molar and its surrounding structures in mice lacking a functional sclerostin gene and in wild-type controls. µCT on six animals in each group revealed that the dimension of the basal bone as well as the coronal and apical part of alveolar part increased in the sclerostin knockout mice. No significant differences were observed for the tooth and pulp chamber volume. Descriptive histomorphometric analyses of four wild-type and three sclerostin knockout mice demonstrated an increased width of the cementum and a concomitant moderate decrease in the periodontal space width. Taken together, these results suggest that the lack of sclerostin mainly alters the bone and cementum phenotypes rather than producing abnormalities in tooth structures such as dentin.

Early bone apposition to hydrophilic and hydrophobic titanium implant surfaces: a histologic and histomorphometric study in minipigs.

OBJECTIVE: The first objective of this pilot study was to evaluate the impact of the hydrophilicity on the early phases of osseointegration. The second objective was to compare two hydrophilic implant surfaces with different geometries, surface roughness, and technologies achieving hydrophilicity. MATERIAL AND METHODS: Twelve weeks after extraction, all four quadrants of nine minipigs received three dental implants, alternating between hydrophilic microrough surfaces (INICELL and SLActive) and a conventional hydrophobic microrough surface. After 5, 10, and 15 days of submerged healing, ground sections were prepared and subjected to histologic and histomorphometric analysis. RESULTS: The histologic analysis revealed a similar healing pattern among the hydrophilic and hydrophobic implant surfaces, with extensive bone formation occurring between day 5 and day 10. With BIC values of greater than 50% after 10 days, all examined surfaces indicated favorable osseointegration at this very early point in healing. At day 15, the mean new bone-to-implant contact (newBIC) of one hydrophilic surface (INICELL; 55.8 ± 14.4%) was slightly greater than that of the hydrophobic microrough surface (40.6 ± 20.2%). At day 10 and day 15, an overall of 21% of the implants had to be excluded from analysis due to inflammations primarily caused by surgical complications. CONCLUSION: Substantial bone apposition occurs between day 5 and day 10. The data suggest that the hydrophilic surface can provoke a slight tendency toward increased bone apposition in minipigs after 15 days. A direct comparison of two hydrophilic surfaces with varying geometries is of limited relevance.