A scalable and highly immunogenic virus-like particle-based vaccine against SARS-CoV-2.

BACKGROUND: SARS-CoV-2 caused one of the most devastating pandemics in the recent history of mankind. Due to various countermeasures, including lock-downs, wearing masks, and increased hygiene, the virus has been controlled in some parts of the world. More recently, the availability of vaccines, based on RNA or adenoviruses, has greatly added to our ability to keep the virus at bay; again, however, in some parts of the world only. While available vaccines are effective, it would be desirable to also have more classical vaccines at hand for the future. Key feature of vaccines for long-term control of SARS-CoV-2 would be inexpensive production at large scale, ability to make multiple booster injections, and long-term stability at 4℃. METHODS: Here, we describe such a vaccine candidate, consisting of the SARS-CoV-2 receptor-binding motif (RBM) grafted genetically onto the surface of the immunologically optimized cucumber mosaic virus, called CuMVTT -RBM. RESULTS: Using bacterial fermentation and continuous flow centrifugation for purification, the yield of the production process is estimated to be >2.5 million doses per 1000-litre fermenter run. We demonstrate that the candidate vaccine is highly immunogenic in mice and rabbits and induces more high avidity antibodies compared to convalescent human sera. The induced antibodies are more cross-reactive to mutant RBDs of variants of concern (VoC). Furthermore, antibody responses are neutralizing and long-lived. In addition, the vaccine candidate was stable for at least 14 months at 4℃. CONCLUSION: Thus, the here presented VLP-based vaccine may be a good candidate for use as conventional vaccine in the long term.

Optical fibers for endoscopic high-power Er:YAG laserosteotomy.

SIGNIFICANCE: The highest absorption peaks of the main components of bone are in the mid-infrared region, making Er:YAG and CO2 lasers the most efficient lasers for cutting bone. Yet, studies of deep bone ablation in minimally invasive settings are very limited, as finding suitable materials for coupling high-power laser light with low attenuation beyond 2 µm is not trivial. AIM: The first aim of this study was to compare the performance of different optical fibers in terms of transmitting Er:YAG laser light with a 2.94-µm wavelength at high pulse energy close to 1 J. The second aim was to achieve deep bone ablation using the best-performing fiber, as determined by our experiments. APPROACH: In our study, various optical fibers with low attenuation (λ = 2.94 µm) were used to couple the Er:YAG laser. The fibers were made of germanium oxide, sapphire, zirconium fluoride, and hollow-core silica, respectively. We compared the fibers in terms of transmission efficiency, resistance to high Er:YAG laser energy, and bending flexibility. The best-performing fiber was used to achieve deep bone ablation in a minimally invasive setting. To do this, we adapted the optimal settings for free-space deep bone ablation with an Er:YAG laser found in a previous study. RESULTS: Three of the fibers endured energy per pulse as high as 820 mJ at a repetition rate of 10 Hz. The best-performing fiber, made of germanium oxide, provided higher transmission efficiency and greater bending flexibility than the other fibers. With an output energy of 370 mJ per pulse at 10 Hz repetition rate, we reached a cutting depth of 6.82 ± 0.99 mm in sheep bone. Histology image analysis was performed on the bone tissue adjacent to the laser ablation crater; the images did not show any structural damage. CONCLUSIONS: The findings suggest that our prototype could be used in future generations of endoscopic devices for minimally invasive laserosteotomy.

Industrial Development of Standardized Fetal Progenitor Cell Therapy for Tendon Regenerative Medicine: Preliminary Safety in Xenogeneic Transplantation.

Tendon defects require multimodal therapeutic management over extensive periods and incur high collateral burden with frequent functional losses. Specific cell therapies have recently been developed in parallel to surgical techniques for managing acute and degenerative tendon tissue affections, to optimally stimulate resurgence of structure and function. Cultured primary human fetal progenitor tenocytes (hFPT) have been preliminarily considered for allogeneic homologous cell therapies, and have been characterized as stable, consistent, and sustainable cell sources in vitro. Herein, optimized therapeutic cell sourcing from a single organ donation, industrial transposition of multi-tiered progenitor cell banking, and preliminary preclinical safety of an established hFPT cell source (i.e., FE002-Ten cell type) were investigated. Results underlined high robustness of FE002-Ten hFPTs and suitability for sustainable manufacturing upscaling within optimized biobanking workflows. Absence of toxicity or tumorigenicity of hFPTs was demonstrated in ovo and in vitro, respectively. Furthermore, a 6-week pilot good laboratory practice (GLP) safety study using a rabbit patellar tendon partial-thickness defect model preliminarily confirmed preclinical safety of hFPT-based standardized transplants, wherein no immune reactions, product rejection, or tumour formation were observed. Such results strengthen the rationale of the multimodal Swiss fetal progenitor cell transplantation program and prompt further investigation around such cell sources in preclinical and clinical settings for musculoskeletal regenerative medicine.

Development of Standardized Fetal Progenitor Cell Therapy for Cartilage Regenerative Medicine: Industrial Transposition and Preliminary Safety in Xenogeneic Transplantation.

Diverse cell therapy approaches constitute prime developmental prospects for managing acute or degenerative cartilaginous tissue affections, synergistically complementing specific surgical solutions. Bone marrow stimulation (i.e., microfracture) remains a standard technique for cartilage repair promotion, despite incurring the adverse generation of fibrocartilagenous scar tissue, while matrix-induced autologous chondrocyte implantation (MACI) and alternative autologous cell-based approaches may partly circumvent this effect. Autologous chondrocytes remain standard cell sources, yet arrays of alternative therapeutic biologicals present great potential for regenerative medicine. Cultured human epiphyseal chondro-progenitors (hECP) were proposed as sustainable, safe, and stable candidates for chaperoning cartilage repair or regeneration. This study describes the development and industrial transposition of hECP multi-tiered cell banking following a single organ donation, as well as preliminary preclinical hECP safety. Optimized cell banking workflows were proposed, potentially generating millions of safe and sustainable therapeutic products. Furthermore, clinical hECP doses were characterized as non-toxic in a standardized chorioallantoic membrane model. Lastly, a MACI-like protocol, including hECPs, was applied in a three-month GLP pilot safety evaluation in a caprine model of full-thickness articular cartilage defect. The safety of hECP transplantation was highlighted in xenogeneic settings, along with confirmed needs for optimal cell delivery vehicles and implantation techniques favoring effective cartilage repair or regeneration.

Do dGEMRIC and T2 Imaging Correlate With Histologic Cartilage Degeneration in an Experimental Ovine FAI Model?

BACKGROUND: Biochemical MRI of hip cartilage such as delayed gadolinium-enhanced MRI of cartilage (dGEMRIC) and T2 mapping is increasingly used to judge cartilage quality in the assessment of femoroacetabular impingement (FAI). The current evidence is sparse about which of these techniques yields a stronger correlation with histologic cartilage degeneration because of the difficulty in validating biochemical MRI techniques against histology in the clinical setting. Recently, an experimental ovine FAI model was established that induces chondrolabral damage and offers a validated platform to address these limitations. QUESTIONS/PURPOSES: In a sheep model, we asked: (1) Do dGEMRIC and/or T2 values of acetabular and femoral cartilage correlate with histologic cartilage degeneration as assessed with the Mankin score? (2) Do simultaneously measured dGEMRIC and T2 values correlate in an experimental ovine FAI model? METHODS: We performed an experimental pilot study on five female Swiss Alpine sheep (10 hips) that underwent postmortem MRI, including biochemical cartilage sequences, after a staged FAI correction had been performed on one side. No surgery was performed on the contralateral side, which served as a healthy control. In these sheep, an extraarticular intertrochanteric varus osteotomy was performed to rotate the naturally aspherical ovine femoral head into the acetabulum to induce cam-type FAI and chondrolabral damage comparable to human beings. After a 70-day ambulation period, femoral osteochondroplasty was performed and all sheep were euthanized after a total observation period of 210 days. Before they were euthanized, the sheep received a contrast agent and roamed and walked for at least 45 minutes. Hips were prepared to fit in a knee coil and MRI was performed at 3 T including a three-dimensional (3-D) dGEMRIC sequence, a two-dimensional (2-D) radial T2 mapping sequence, and a 2-D radial proton density-weighted sequence for morphologic cartilage assessment. Using specifically developed software, the 3-D dGEMRIC images and T2 maps were coregistered on the 2-D morphologic radial images. This enabled us to simultaneously measure dGEMRIC and T2 values using the identical regions of interest. dGEMRIC and T2 values of the acetabular and femoral cartilage were measured circumferentially using anatomic landmarks. After MRI, bone-cartilage samples were taken from the acetabulum and the femur and stained with toluidine blue for assessment of the histologic cartilage degeneration using the Mankin score, which was assessed in consensus by two observers. Spearman's rank correlation coefficient was used to (1) correlate dGEMRIC values and T2 values with the histologic Mankin score of femoroacetabular cartilage; and to (2) correlate dGEMRIC values and T2 values of femoroacetabular cartilage. RESULTS: A moderate to fair correlation between overall dGEMRIC values of the acetabular cartilage (R = -0.430; p = 0.003) and the femoral cartilage (R = -0.334; p = 0.003) versus the histologic Mankin score was found. A moderate correlation (R = -0.515; p = 0.010) was found among peripheral dGEMRIC values of the acetabulum, the superior femoral cartilage (R = -0.500; p = 0.034), and the histologic Mankin score, respectively. No correlation between overall and regional femoroacetabular T2 values and the histologic Mankin scores was found. No correlation between overall and regional femoroacetabular dGEMRIC values and T2 values was found. CONCLUSIONS: In this recently established sheep model, we found dGEMRIC values correlated well with histologic evidence of cartilage degeneration in the hip. This combination of a robust animal model and an accurate imaging technique appears to offer a noninvasive means to study the natural course of FAI and to compare the effectiveness of potential surgical options to treat it. CLINICAL RELEVANCE: This translational study supports the continuing use of dGEMRIC as a biomarker for prearthritic cartilage degeneration with the ultimate goal to identify patients who will benefit most from corrective FAI surgery. The value of T2 imaging of hip cartilage warrants further investigation.

Comparison of hock- and footpad-injection as a prostate adenocarcinoma model in rats.

BACKGROUND: Objective of this study is a feasibility-test comparing hock- and footpad-injection in rats with inoculated MatLyLu - adenocarcinoma tumor model. This study compares the development of an adenocarcinoma model (MatLyLu) in 12 Copenhagen rats. Two groups (n = 6) of animals were inoculated with 1 × 106 MatLyLu tumor cells solved in 0.1 ml NaCl either by footpad or hock injection. All animals were examined before tumor inoculation and before euthanasia using a 3.0 Tesla MRI. Histological evaluation of all organs was performed post mortem. RESULTS: Both types of injection were able to induce the adenocarcinoma model using MatLyLu tumor cells. The primary tumor could be visualized in MRI and confirmed histologically. Comparing the risk of reflux and the maximum injection volume during injection, the hock injection was superior to the footpad injection (less reflux, less anatomical restrictions for larger volumes). The hock injection induces a faster tumor growth compared to the footpad injection. As consequence the maximum level of long term discomfort after hock injection was reached earlier, even if it grew on a not weight bearing structure. Early lymph node tumor metastasis could not be observed macroscopically nor detected histologically. Therefore the reproducibility of the MatLyLu tumor model is questionable. CONCLUSION: Hock injection is a feasible alternative technique compared with footpad-injection in rats. It provides a save and easy injection method for various early-terminated applications with the potential to increase animal welfare during tumor models in rats.

Incorporating BMP-2 and skeletal muscle to a semitendinosus autograft in an oversized tunnel yields robust bone tunnel ossification in rabbits: Toward single-step revision of failed anterior cruciate ligament reconstruction.

BACKGROUND: Bone tunnel widening after anterior cruciate ligament (ACL) reconstruction is a known complication that can lead to graft failure. Subsequent revision surgery typically involves a two-stage procedure. The aim of this study was to test a novel autologous tendon graft retaining muscle tissue combined with Human Recombinant Bone Morphogenetic Protein-2 (rh-BMP-2) leading to rapid ossification of the muscle tissue, simultaneously replenishing bone stock and producing a mechanically stable bone-tendon insertion. METHODS: In 12 skeletally mature New Zealand rabbits, the ACL was resected and oversized bone tunnels were drilled to model tunnel widening. The ipsilateral semitendinosus muscle-tendon graft was harvested and folded twice. Muscle tissue was removed in the middle third but retained at both distal ends. One side was wrapped in a collagen sponge loaded with rh-BMP-2 while the other end was used as its own control. RESULTS: All animals were euthanized after six weeks. Micro-computed tomography (micro-CT) was used to analyze bone formation in 12 animals, with additional biomechanical testing to failure and histology performed for six animals each. Micro-CT showed that bone densities were higher by a factor of 2.4 in treated graft ends compared with their controls. Biomechanical testing showed a mean overall failure load of 37.5 N. Histology showed that the trabecular bone surrounding the implant was significantly (P = 0.0087) thicker on the treated (85.5 µm) compared with the control side (68.2 µm). CONCLUSIONS: We conclude that a semitendinosus graft retaining the muscle tissue stimulated by recombinant Bone Morphogenetic Protein-2 (BMP-2) allows robust osseointegration of the graft within an oversized bone tunnel in an animal model.

Comparative microstructural analysis of bone osteotomies after cutting by computer-assisted robot-guided laser osteotome and piezoelectric osteotome: an in vivo animal study.

Most industrial laser applications utilize computer and robot assistance, for guidance, safety, repeatability, and precision. In contrast, medical applications using laser systems are mostly conducted manually. The advantages can be effective only when the system is coupled to a robotic guidance, as operating by hand does not reach the required accuracy. We currently developed the first laser osteotome which offers preoperative planning based on CT data, robot guidance, and a precise execution of the laser cuts. In an animal trial, our system was used to create a grid pattern of the same depth on the inner layer of parietal bone in 12 adult sheep. The same bone cuts were done with piezoelectric osteotome on the contralateral side. The micro-CT and histological analysis showed more new mineralized bone in the laser group compared to the piezoelectric group. As well, a cutting pattern with especially a constant osteotomy depth in the laser group was demonstrated. The here presented autonomous osteotomy tool shows not only an advantage in early bone healing stage but additionally sharp bone cuts with a very high accuracy and freely selectable design cuts.

Neer Award 2016: reduced muscle degeneration and decreased fatty infiltration after rotator cuff tear in a poly(ADP-ribose) polymerase 1 (PARP-1) knock-out mouse model.

BACKGROUND: Disturbed muscular architecture, atrophy, and fatty infiltration remain irreversible in chronic rotator cuff tears even after repair. Poly (adenosine 5'-diphosphate-ribose) polymerase 1 (PARP-1) is a key regulator of inflammation, apoptosis, muscle atrophy, muscle regeneration, and adipocyte development. We hypothesized that the absence of PARP-1 would lead to a reduction in damage to the muscle subsequent to combined tenotomy and neurectomy in a PARP-1 knockout (KO) mouse model. METHODS: PARP-1 KO and wild-type C57BL/6 (WT group) mice were analyzed at 1, 6, and 12 weeks (total n = 84). In all mice, the supraspinatus and infraspinatus muscles of the left shoulder were detached and denervated. Macroscopic analysis, magnetic resonance imaging, gene expression analysis, immunohistochemistry, and histology were used to assess the differences in PARP-1 KO and WT mice. RESULTS: The muscles in the PARP-1 KO group had significantly less retraction, atrophy, and fatty infiltration after 12 weeks than in the WT group. Gene expression of inflammatory, apoptotic, adipogenic, and muscular atrophy genes was significantly decreased in PARP-1 KO mice in the first 6 weeks. DISCUSSION: Absence of PARP-1 leads to a reduction in muscular architectural damage, early inflammation, apoptosis, atrophy, and fatty infiltration after combined tenotomy and neurectomy of the rotator cuff muscle. Although the macroscopic reaction to injury is similar in the first 6 weeks, the ability of the muscles to regenerate was much greater in the PARP-1 KO group, leading to a near-normalization of the muscle after 12 weeks.

Human Bone Progenitor Cells for Clinical Application: What Kind of Immune Reaction Does Fetal Xenograft Tissue Trigger in Immunocompetent Rats?

The potential of human fetal bone cells for successful bone regeneration has been shown in vivo. In particular, it has been demonstrated that the seeding of these cells in porous poly-(l-lactic acid)/ß-tricalcium phosphate scaffolds improved the bone formation compared to cell-free scaffolds in skulls of rats. However, even if the outcome is an improvement of bone formation, a thorough analysis concerning any immune responses, due to the implantation of a xenograft tissue, is not known. As the immune response and skeletal system relationship may contribute to either the success or failure of an implant, we were interested in evaluating the presence of any immune cells and specific reactions of human fetal cells (also called human bone progenitor cells) once implanted in femoral condyles of rats. For this purpose, (1) cell-free scaffolds, (2) human bone progenitor cells, or (3) osteogenic human bone progenitor cells within scaffolds were implanted over 3, 7, 14 days, and 12 weeks. The key finding is that human bone progenitor cells and osteogenic human bone progenitor cells do not trigger any particular specific immune reactions in immunocompetent rats but are noted to delay some bone formation.

Head-Neck Osteoplasty has Minor Effect on the Strength of an Ovine Cam-FAI Model: In Vitro and Finite Element Analyses.

BACKGROUND: Osteochondroplasty of the head-neck region is performed on patients with cam femoroacetabular impingement (FAI) without fully understanding its repercussion on the integrity of the femur. Cam-type FAI can be surgically and reproducibly induced in the ovine femur, which makes it suitable for studying corrective surgery in a consistent way. Finite element models built on quantitative CT (QCT) are computer tools that can be used to predict femoral strength and evaluate the mechanical effect of surgical correction. QUESTIONS/PURPOSES: We asked: (1) What is the effect of a resection of the superolateral aspect of the ovine femoral head-neck junction on failure load? (2) How does the failure load after osteochondroplasty compare with reported forces from activities of daily living in sheep? (3) How do failure loads and failure locations from the computer simulations compare with the experiments? METHODS: Osteochondroplasties (3, 6, 9 mm) were performed on one side of 18 ovine femoral pairs with the contralateral intact side as a control. The 36 femurs were scanned via QCT from which specimen-specific computer models were built. Destructive compression tests then were conducted experimentally using a servohydraulic testing system and numerically via the computer models. Safety factors were calculated as the ratio of the maximal force measured in vivo by telemeterized hip implants during the sheep's walking and running activities to the failure load. The simulated failure loads and failure locations from the computer models were compared with the experimental results. RESULTS: Failure loads were reduced by 5% (95% CI, 2%-8%) for the 3-mm group (p = 0.0089), 10% (95% CI, 6%-14%) for the 6-mm group (p = 0.0015), and 19% (95% CI, 13%-26%) for the 9-mm group (p = 0.0097) compared with the controls. Yet, the weakest specimen still supported more than 2.4 times the peak load during running. Strong correspondence was found between the simulated and experimental failure loads (R2 = 0.83; p < 0.001) and failure locations. CONCLUSIONS: The resistance of ovine femurs to fracture decreased with deeper resections. However, under in vitro testing conditions, the effect on femoral strength remains small even after 9 mm correction, suggesting that femoral head-neck osteochondroplasty could be done safely on the ovine femur. QCT-based finite element models were able to predict weakening of the femur resulting from the osteochondroplasty. CLINICAL RELEVANCE: The ovine femur provides a seemingly safe platform for scientific evaluation of FAI. It also appears that computer models based on preoperative CT scans may have the potential to provide patient-specific guidelines for preventing overcorrection of cam FAI.

Dynamization at the near cortex in locking plate osteosynthesis by means of dynamic locking screws: an experimental study of transverse tibial osteotomies in sheep.

BACKGROUND: Locking plates are widely used in fracture fixation, mainly for meta-diaphyseal fractures, comminuted fractures, fractures with a critical-size bone defect, periprosthetic fractures, osteotomies, and fractures in osteoporotic bone. The aim of this animal study was to evaluate the effect on bone-healing of dynamization of locking plate constructs by means of new 5.0-mm dynamic locking screws (in the DLS group), which allow near-cortex micromotion, compared with a more rigid construct utilizing standard bicortical locking-head screws (in the LS group). Use of dynamic locking screws allows modulation of the stiffness of existing locking compression plate systems via parallel interfragmentary micromotion. METHODS: A standardized diaphyseal tibial osteotomy (90°, 3-mm fracture gap) was performed and stabilized with a six-hole large-fragment locking compression plate in twelve female sheep (six in each group). Radiographs were made postoperatively and then weekly from week three until sacrifice at nine weeks. Macroscopic, biomechanical, histologic, and radiographic assessments and microcomputed tomography were performed. RESULTS: The callus in the tested specimens in the DLS group had better biomechanical stability, with a significantly greater maximum failure moment (mean and standard deviation [SD] as a percentage of intact, 55.15 ± 20.65 compared with 26.80 ± 14.96 in the LS group; p = 0.021). The DLS group also had greater periosteal callus volume at the near cortex (mean volume and SD as a percentage of the tibial shaft volume, 36.21% ± 10.08% compared with 18.98% ± 8.61% in the LS group; p = 0.026) and in the intercortical region (mean volume and SD as a percentage of the bone volume of the tibial shaft, 3.56% ± 0.52% compared with 2.64% ± 0.98% in the LS group; p = 0.045), as shown by microcomputed tomography. The DLS group also had significantly greater torsional stiffness (mean and SD as a percentage of intact, 84.88 ± 13.51 compared with 58.89 ± 20.61 in the LS group; p = 0.027). CONCLUSIONS: Controlled micromotion and nearly homogeneous interfragmentary strain at the fracture site, together with the stable bicortical fixation achieved by the new dynamic locking screw, led to more uniform callus formation, significantly more callus formation at the near cortex, and biomechanically more competent bone-healing compared with use of rigid locking plate constructs with locking-head screws.

Remodeling of tissue-engineered bone structures in vivo.

Implant design for bone regeneration is expected to be optimized when implant structures resemble the anatomical situation of the defect site. We tested the validity of this hypothesis by exploring the feasibility of generating different in vitro engineered bone-like structures originating from porous silk fibroin scaffolds decorated with RGD sequences (SF-RGD), seeded with human mesenchymal stem cells (hMSC). Scaffolds with small (106-212 µm), medium (212-300 µm), and large pore diameter ranges (300-425 µm) were seeded with hMSC and subsequently differentiated in vitro into bone-like tissue resembling initial scaffold geometries and featuring bone-like structures. Eight weeks after implantation into calvarial defects in mice, the in vitro engineered bone-like tissues had remodeled into bone featuring different proportions of woven/lamellar bone bridging the defects. Regardless of pore diameter, all implants integrated well, vascularization was advanced, and bone marrow ingrowth had started. Ultimately, in this defect model, the geometry of the in vitro generated tissue-engineered bone structure, trabecular- or plate-like, had no significant impact on the healing of the defect, owing to an efficient remodeling of its structure after implantation.

Biocompatibility and osteoconduction of macroporous silk fibroin implants in cortical defects in sheep.

The goal of the presented study was to compare the biocompatibility and cellular responses to porous silk fibroin (SF) scaffolds produced in a water-based (UPW) or a solvent based process (HFIP) using two different SF sources. For that reason, four different SF scaffolds were implanted (n=6) into drill hole defects in the cancellous bone of the sheep tibia and humerus. The scaffolds were evaluated histologically for biocompatibility, cell-material interaction, and cellular ingrowth. New bone formation was observed macroscopically and histologically at 8 weeks after implantation. For semiquantitative evaluation, the investigated parameters were scored and statistically analyzed (factorial ANOVA). All implants showed good biocompatibility as evident by low infiltration of inflammatory cells and the absent encapsulation of the scaffolds in connective tissue. Multinuclear foreign body giant cells (MFGCs) and macrophages were present in all parts of the scaffold at the material surface and actively degrading the SF material. Cell ingrowth and vascularization were uniform across the scaffold. However, in HFIP scaffolds, local regions of void pores were present throughout the scaffold, probably due to the low pore interconnectivity in this scaffold type in contrast to UPW scaffolds. The amount of newly formed bone was very low in both scaffold types but was more abundant in the periphery than in the center of the scaffolds and for HFIP scaffolds mainly restricted to single pores.

The dynamisation of locking plate osteosynthesis by means of dynamic locking screws (DLS)-an experimental study in sheep.

In this in vivo study a new generation of locking screws was tested. The design of the dynamic locking screw (DLS) enables the dynamisation of the cortex underneath the plate (cis-cortex) and, therefore, allows almost parallel interfragmentary closure of the fracture gap. A 45° angle osteotomy was performed unilaterally on the tibia of 37 sheep. Groups of 12 sheep were formed and in each group a different osteotomy gap (0, 1 and 3mm) was fixed using a locking compression plate (LCP) in combination with the DLS. The healing process was monitored radiographically every 3 weeks for 6, respectively 12 weeks. After this time the sheep were sacrificed, the bones harvested and the implants removed. The isolated bones were evaluated in the micro-computed tomography unit, tested biomechanically and evaluated histologically. The best results of interfragmentary movement (IFM) were shown in the 0mm configuration. The bones of this group demonstrated histomorphometrically the most distinct callus formation on the cis-cortex and the highest torsional stiffness relative to the untreated limb at 12 weeks after surgery. This animal study showed that IFM stimulated the synthesis of new bone matrix, especially underneath the plate and thus, could solve a current limitation in normal human bone healing. The DLS will be a valuable addition to the locking screw technology and improve fracture healing.

Osseointegration and biocompatibility of different metal implants--a comparative experimental investigation in sheep.

BACKGROUND: In the present study, 4 different metallic implant materials, either partly coated or polished, were tested for their osseointegration and biocompatibility in a pelvic implantation model in sheep. METHODS: Materials to be evaluated were: Cobalt-Chrome (CC), Cobalt-Chrome/Titanium coating (CCTC), Cobalt-Chrome/Zirconium/Titanium coating (CCZTC), Pure Titanium Standard (PTST), Steel, TAN Standard (TANST) and TAN new finish (TANNEW). Surgery was performed on 7 sheep, with 18 implants per sheep, for a total of 63 implants. After 8 weeks, the specimens were harvested and evaluated macroscopically, radiologically, biomechanically (removal torque), histomorphometrically and histologically. RESULTS: Cobalt-Chrome screws showed significantly (p = 0.031) lower removal torque values than pure titanium screws and also a tendency towards lower values compared to the other materials, except for steel. Steel screws showed no significant differences, in comparison to cobalt-chrome and TANST, however also a trend towards lower torque values than the remaining materials. The results of the fluorescence sections agreed with those of the biomechanical test. Histomorphometrically, there were no significant differences of bone area between the groups. The BIC (bone-to-implant-contact), used for the assessment of the osseointegration, was significantly lower for cobalt-chrome, compared to steel (p = 0.001). Steel again showed a lower ratio (p = 0.0001) compared to the other materials. CONCLUSION: This study demonstrated that cobalt-chrome and steel show less osseointegration than the other metals and metal-alloys. However, osseointegration of cobalt-chrome was improved by zirconium and/or titanium based coatings (CCTC, TANST, TAN, TANNEW) being similar as pure titanium in their osseointegrative behavior.

Biocompatibility and Bone Formation of Flexible, Cotton Wool-like PLGA/Calcium Phosphate Nanocomposites in Sheep.

BACKGROUND: The purpose of this preliminary study was to assess the in vivo performance of synthetic, cotton wool-like nanocomposites consisting of a biodegradable poly(lactide-co-glycolide) fibrous matrix and containing either calcium phosphate nanoparticles (PLGA/CaP 60:40) or silver doped CaP nanoparticles (PLGA/Ag-CaP 60:40). Besides its extraordinary in vitro bioactivity the latter biomaterial (0.4 wt% total silver concentration) provides additional antimicrobial properties for treating bone defects exposed to microorganisms. MATERIALS AND METHODS: Both flexible artificial bone substitutes were implanted into totally 16 epiphyseal and metaphyseal drill hole defects of long bone in sheep and followed for 8 weeks. Histological and histomorphological analyses were conducted to evaluate the biocompatibility and bone formation applying a score system. The influence of silver on the in vivo performance was further investigated. RESULTS: Semi-quantitative evaluation of histology sections showed for both implant materials an excellent biocompatibility and bone healing with no resorption in the adjacent bone. No signs of inflammation were detectable, either macroscopically or microscopically, as was evident in 5 µm plastic sections by the minimal amount of inflammatory cells. The fibrous biomaterials enabled bone formation directly in the centre of the former defect. The area fraction of new bone formation as determined histomorphometrically after 8 weeks implantation was very similar with 20.5 ± 11.2 % and 22.5 ± 9.2 % for PLGA/CaP and PLGA/Ag-CaP, respectively. CONCLUSIONS: The cotton wool-like bone substitute material is easily applicable, biocompatible and might be beneficial in minimal invasive surgery for treating bone defects.

The use of BoneWelding® technology in spinal surgery: an experimental study in sheep.

The innovative BoneWelding(®) technology, where ultrasound energy bonds bioresorbable implants to bone, was tested for its feasibility in spine surgery and its local thermal effects. The three tested concepts consisted of implementation of a resorbable plating system, two converging polymer pins and suture anchors to the cervical vertebral bodies. Bioresorbable polylactide implants (PLDLLA 70/30) were inserted ventrally into the third and fourth vertebral body of seven sheep, of which six were sacrificed at 2 months and one sheep immediately after temperature measurements during implant insertion. Polymer screws were used as controls. Qualitative, semi-quantitative histological, and quantitative histomorphometrical evaluation showed excellent anchorage of the implants, new mineralized bone at the implant-bone interface, no inflammatory cell reaction or thermal damage to the adjacent bone in response to the novel insertion technology. The application of two converging pins, parallel inserted polymer pins, or fusion of the implant to the polymer plates did not affect the overall excellent tissue tolerance of the technology. Temperature increase during insertion was noticed but never exceeded 47°C for less than 1 s. The BoneWelding(®) technology was proven to be safe and easy to apply.

Comparison of Er:YAG laser and piezoelectric osteotomy: An animal study in sheep.

OBJECTIVES: It was the aim of this study to compare the feasibility of complete osteotomy of long bones in sheep using a newly designed variable square pulsed Er:YAG laser and piezoelectric surgery. In addition to uneventful bone healing after laser osteotomy, the goal was to assess the possibility to cut thick bony structures with both techniques in a surgically acceptable time frame of 2-3 minutes. MATERIAL AND METHODS: A tibia midshaft osteotomy was performed in 24 sheep using either an Er:YAG laser (n = 12) or piezoelectric device (n = 12). Laser and piezoelectric groups were divided in two subgroups (n = 6) with sheep sacrificed after 2 and 3 months, respectively. A complete radiological, histological and histomorphometric analysis was performed to compare the course of bone/fracture healing and remodelling. RESULTS: Laser and piezoelectric osteotomies of the sheep tibia up to a depth of 22 mm were possible without any thermal damage. Radiological and histological results after 2 months showed primary gap healing with distinct periosteal callus formation on the transcortex. After 3 months, radiological and histological analysis revealed less callus formation on the transcortex, with almost no visible osteotomy gap and a distinct formation of lamellar bone crossing the original osteotomy gap. CONCLUSION: Er:YAG laser osteotomy can successfully be used in long bones with a depth of up to 22 mm, thus challenging the dogma of adverse effects of laser osteotomy due to thermal or other damages.

Controlled release of tetracycline from biodegradable beta-tricalcium phosphate composites.

For use in the prevention of bone infections, a novel controlled release system composed of beta-tricalcium phosphate (TCP) granules with biodegradable coatings incorporating the antibiotic drug tetracycline (TC) was developed. Six formulations using poly(D,L-lactide) and poly(D,L-lactide-co-glycolide) as coating materials to incorporate TC were prepared and tested in vitro and in vivo. Release of TC from TCP composites was dependent on the biodegradability of the used polymers and on physical-chemical interactions of TC with the polymer materials. Three characteristic release profiles were obtained: slow release lasting up to 67 days, intermediate release with 60% of the total dose released up to day 20, and fast release with a high initial burst and 90% of TC released within 4 days. Even though TC decomposition products had formed during in vitro release, no cytotoxic effects on osteoblast-like cells were observed. The biological activity of TC after incorporation into PL(G)A films was confirmed using a TC-repressible promoter system in genetically engineered Chinese Hamster Ovary (CHO) cells. TC-loaded TCP composites implanted in ovine cancellous bone defects showed good biocompatibility and new bone formation in the histological evaluation. No differences in the cellular reactions were seen between antibiotic-loaded composites and the control group. These experimental results indicate the potential of coated TCP composites to be used as local carrier system for controlled TC delivery with different release kinetics and good in vitro and in vivo biocompatibility.