Efficient colonic drug delivery in domestic pigs employing a tablet formulation with dual control concept.

Efficient and reproducible colonic drug delivery remains elusive. The aim of this study was to demonstrate specific colonic delivery in vivo in domestic pigs with a novel tablet formulation based on a dual release control concept using 5-aminosalicylic acid (5-ASA) and caffeine as drug substances. The developed controlled colonic release (CCR) tablet formulation employs a pH-sensitive coating based on Eudragit® FS 30 D to prevent drug release in the upper gastrointestinal tract, and a xyloglucan-based matrix to inhibit drug release after coating removal in the small intestine and to allow microbiome-triggered drug release by enzymatic action in the colon. CCR tablets were administered to domestic pigs and plasma concentration data was analyzed by physiologically based pharmacokinetic modeling to estimate absorbed amounts from small and large intestine and in vivo drug release rates by model-dependent deconvolution using immediate release (IR) tablets and intravenous solutions as reference. Peak concentration times (tmax) and values (cmax) of CCR 5-ASA and caffeine tablets indicated strongly delayed drug absorption and the deduced absorbed amount as a function of time confirmed absorption overwhelmingly from the large intestine. The microbially cleaved marker molecule sulfasalazine administered alone or together with caffeine in CCR tablets reported, in combination with telemetry measurements, gastrointestinal transit times and site of absorption. Drug release from CCR tablets was inferred to take place predominantly at the site of absorption at a release rate of caffeine that was much larger in the colon than in the small intestine indicating enzymatically triggered release by the colonic microbiome. Xyloglucanase activity in rectal and cecal samples was consistent with release data and compound recovery in fecal droppings was consistent with 5-ASA bioavailability. The results provide evidence that the developed formulation can prevent premature drug release and provide targeted colonic drug delivery. Clinical relevance based on the comparability between pig and man is discussed.

A Bioinspired Orthopedic Biomaterial with Tunable Mechanical Properties Based on Sintered Titanium Fibers.

Inadequate mechanical compliance of orthopedic implants can result in excessive strain of the bone interface, and ultimately, aseptic loosening. It is hypothesized that a fiber-based biometal with adjustable anisotropic mechanical properties can reduce interface strain, facilitate continuous remodeling, and improve implant survival under complex loads. The biometal is based on strategically layered sintered titanium fibers. Six different topologies are manufactured. Specimens are tested under compression in three orthogonal axes under 3-point bending and torsion until failure. Biocompatibility testing involves murine osteoblasts. Osseointegration is investigated by micro-computed tomography and histomorphometry after implantation in a metaphyseal trepanation model in sheep. The material demonstrates compressive yield strengths of up to 50 MPa and anisotropy correlating closely with fiber layout. Samples with 75% porosity are both stronger and stiffer than those with 85% porosity. The highest bending modulus is found in samples with parallel fiber orientation, while the highest shear modulus is found in cross-ply layouts. Cell metabolism and morphology indicate uncompromised biocompatibility. Implants demonstrate robust circumferential osseointegration in vivo after 8 weeks. The biometal introduced in this study demonstrates anisotropic mechanical properties similar to bone, and excellent osteoconductivity and feasibility as an orthopedic implant material.

Oral Use of Therapeutic Carbon Monoxide for Anyone, Anywhere, and Anytime.

Carbon monoxide (CO) is a therapeutic gas with therapeutic potential in intestinal bowel disease. Therapeutic efficacy in the gastrointestinal tract (GIT) must be paired with safe and convenient use. Therefore, we designed an oral CO releasing system (OCORS) pairing tunable CO release into the GIT while preventing the release of any other molecule from within the device, causing safety concerns. The dimensions of the device, which is manufactured from 3D printed components, are within compendial limits. This is achieved by controlling CO decarbonylation from a molybdenum complex with a FeCl3 solution. OCORS' surrounding silicon membranes control release rates, as does the loading with carbonylated molybdenum complex and FeCl3 solution. Herein we describe the development of the system, the characterization of the CO releasing molecule (CORM), and the CO release kinetics of the overall system. Neither the CORM nor isocyanoacetate as a potential reaction byproduct were cytotoxic. Finally, we demonstrated by design validation in an in vivo porcine model that, except for the release of the therapeutic CO, OCORS isolates all components during transit through the stomach. We could show that OCORS generated and released CO locally into the stomach of the animals without systemic exposure, measured as the carboxyhemoglobin content in the blood of the pigs. In conclusion, OCORS derisks oral development by limiting patient exposure to (desirable) CO while preventing contact with any further (undesirable) chemical, by-, or degradation products. CO generating devices come in reach, which now can be used by anyone, anywhere, and anytime.

Histological Preparation and Evaluation of Cartilage Specimens.

In this chapter, an introduction is given into histological techniques to research related to hyaline cartilage and subchondral bone. Emphasis is placed on the importance to investigate cartilage and bone as a unit, which includes the transition zone of the calcified cartilage and tidemark. Reasons for the appropriate selection of histological methods are presented such as when to use (decalcified) specimens for routine paraffin embedding including immunohistology, cryosections of cartilage alone, or non-decalcified specimens for embedding in polymethylmethacrylate with or without additional biomaterials. Appropriate staining methods are also outlined. Apart from detailed laboratory protocols for different embedding and staining methods including open communication about difficulties related to the various techniques, also practical instructions for state-of-the-art evaluation methods and their strengths and weaknesses are given. Sample figures for scoring methods are included.

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.

In vitro Implementation of Photopolymerizable Hydrogels as a Potential Treatment of Intracranial Aneurysms.

Intracranial aneurysms are increasingly being treated with endovascular therapy, namely coil embolization. Despite being minimally invasive, partial occlusion and recurrence are more frequent compared to open surgical clipping. Therefore, an alternative treatment is needed, ideally combining minimal invasiveness and long-term efficiency. Herein, we propose such an alternative treatment based on an injectable, radiopaque and photopolymerizable polyethylene glycol dimethacrylate hydrogel. The rheological measurements demonstrated a viscosity of 4.86 ± 1.70 mPa.s, which was significantly lower than contrast agent currently used in endovascular treatment (p = 0.42), allowing the hydrogel to be injected through 430 µm inner diameter microcatheters. Photorheology revealed fast hydrogel solidification in 8 min due to the use of a new visible photoinitiator. The addition of an iodinated contrast agent in the precursor contributed to the visibility of the precursor injection under fluoroscopy. Using a customized light-conducting microcatheter and illumination module, the hydrogel was implanted in an in vitro silicone aneurysm model. Specifically, in situ fast and controllable injection and photopolymerization of the developed hydrogel is shown to be feasible in this work. Finally, the precursor and the polymerized hydrogel exhibit no toxicity for the endothelial cells. Photopolymerizable hydrogels are expected to be promising candidates for future intracranial aneurysm treatments.

Evaluation of D-dimer levels in aqueous humor of rabbit eyes with and without induced intraocular fibrin and fibrinolytic treatment.

OBJECTIVE: To analyze D-dimer concentrations in aqueous humor (AH) of rabbit eyes under physiological conditions, after induction of fibrin clots, and following fibrinolytic therapy. ANIMALS STUDIED: Prospective study measuring D-dimers in aqueous humor of rabbit eyes with induced fibrin clots (n = 44). PROCEDURES: Rabbits were purchased in two groups, which led to two temporally separated experimentation groups. Different treatment protocols were compared for their efficacy in fibrin reduction (slit-lamp examination, high-resolution ultrasound). AH was taken from left eyes before clot induction (baseline, day 1), 24 hours later after clot establishment/prior to drug administration (post-induction, day 2) and 48 hours after clot induction (post-treatment, day 3). An enzyme-linked immunosorbent assay (ELISA) was performed to measure intraocular D-dimer concentrations RESULTS: D-dimer concentrations were measurable in all samples. There were no differences in D-dimer levels across time points or treatments within the arrival groups. However, a significant difference in mean D-dimer levels was observed between the two arrival groups (group 1:3.1 µg/mL; group 2:6.1 µg/mL; P < .0001), which made a direct comparison of treatment groups impossible. Clinically, all eyes displayed fibrin clots in the anterior chamber and different treatment types led to significant differences in clot resolution (clot size reduction after intracameral treatment: 98%, topical treatment: 60%, no treatment: 40%). CONCLUSION: D-dimers were identified in all AH samples of rabbits with large variability between samples. D-dimer levels were neither predictive for differences in induced fibrin formation nor for drug efficacy.

Pulsatile Flow-Induced Fatigue-Resistant Photopolymerizable Hydrogels for the Treatment of Intracranial Aneurysms.

An alternative intracranial aneurysm embolic agent is emerging in the form of hydrogels due to their ability to be injected in liquid phase and solidify in situ. Hydrogels have the ability to fill an aneurysm sac more completely compared to solid implants such as those used in coil embolization. Recently, the feasibility to implement photopolymerizable poly(ethylene glycol) dimethacrylate (PEGDMA) hydrogels in vitro has been demonstrated for aneurysm application. Nonetheless, the physical and mechanical properties of such hydrogels require further characterization to evaluate their long-term integrity and stability to avoid implant compaction and aneurysm recurrence over time. To that end, molecular weight and polymer content of the hydrogels were tuned to match the elastic modulus and compliance of aneurysmal tissue while minimizing the swelling volume and pressure. The hydrogel precursor was injected and photopolymerized in an in vitro aneurysm model, designed by casting polydimethylsiloxane (PDMS) around 3D printed water-soluble sacrificial molds. The hydrogels were then exposed to a fatigue test under physiological pulsatile flow, inducing a combination of circumferential and shear stresses. The hydrogels withstood 5.5 million cycles and no significant weight loss of the implant was observed nor did the polymerized hydrogel protrude or migrate into the parent artery. Slight surface erosion defects of 2-10 µm in depth were observed after loading compared to 2 µm maximum for non-loaded hydrogels. These results show that our fine-tuned photopolymerized hydrogel is expected to withstand the physiological conditions of an in vivo implant study.

Sex-related differences in wall remodeling and intraluminal thrombus resolution in a rat saccular aneurysm model.

OBJECTIVE: Intracranial aneurysms (IAs) are more often diagnosed in women. Hormones and vessel geometry, which influences wall shear stress, may affect pathophysiological processes of the arterial wall. Here, the authors investigated sex-related differences in the remodeling of the aneurysm wall and in intraluminal thrombus resolution. METHODS: A well-characterized surgical side-wall aneurysm model was used in female, male, and ovariectomized rats. Decellularized grafts were used to model highly degenerated and decellularized IA walls and native grafts to model healthy IA walls. Aneurysm growth and thrombus composition were analyzed at 1, 7, 14, and 28 days. Sex-related differences in vessel wall remodeling were compared with human IA dome samples of men and pre- and postmenopausal women. RESULTS: At 28 days, more aneurysm growth was observed in ovariectomized rats than in males or non-ovariectomized female rats. The parent artery size was larger in male rats than in female or ovariectomized rats, as expected. Wall inflammation increased over time in all groups and was most severe in the decellularized female and ovariectomized groups at 28 days compared with the male group. Likewise, in these groups the most elastin fragmentation was seen at 28 days. In female rats, on days 1, 7, and 14, the intraluminal thrombus was mainly composed of red blood cells and fibrin. On days 14 and 28, macrophage and smooth muscle cell invasion inside the thrombus was shown, leading to the removal of red blood cells and deposition of collagen and elastin. On days 14 and 28, similar profiles of thrombus reorganization were observed in male and ovariectomized female rats. However, collagen content in thrombi and vessel wall macrophage content were higher in aneurysms of male rats at 28 days than in those of female rats. On day 28, thrombus coverage by endothelial cells was lower in ovariectomized than in female or male rats. Finally, analysis of human IA domes showed that endothelial cell coverage was lower in men and postmenopausal women than in younger women. CONCLUSIONS: Aneurysm growth and intraluminal thrombus resolution show sex-dependent differences. While certain processes (endothelial cell coverage and collagen deposition) point to a strong hormonal dependence, others (wall inflammation and aneurysm growth) seem to be influenced by both hormones and parent artery size.

Mechanisms of Suture Integration in Living Tissue: Biomechanical and Histological In Vivo Analysis in Sheep.

The potential of nonabsorbable suture material to augment tissue strength in the long-term is by far not exploited by most of the currently used sutures. The authors hypothesized that different sutures yield specific histological tissue reactions associated with specific mechanical shear resistance of the suture against the tissue. Four different suture types (Orthocord, Ethibond, FiberTape, and FiberWire) were implanted in 36 sheep shoulders (supraspinatus/greater tuberosity). One thread at each time point (6, 16, and 22 weeks) was used for histology, and 11 threads at each time point (0, 6, 16, and 22 weeks) were used for biomechanical longitudinal pullout testing. Histology included tissue maturity, activity of tissue reaction, and invasion of cells and tissue into the suture material. Fiber-Tape had the highest mean pullout strength at 6, 16, and 22 weeks of 4.4 N/cm (SD, 2.1 N/cm), 10.1 N/cm (SD, 5.1 N/cm), and 12.8 N/cm (SD, 6.0 N/cm), respectively. However, general pullout strength at 22 weeks was surprisingly low, particularly for Ethibond, Orthocord and FiberWire. The overall maturity of the surrounding tissue correlated (r=0.84, P=.001) with mechanical performance. Interestingly, in all 4 suture types, an intimate in- and on-growth of fibrous tissue to the filaments and into the space between suture fibers could be shown. However, for Ethibond, Orthocord, and FiberWire, the authors found an unexpected circumferential space around the sutures, often forming an inner and outer capsule, separating the sutures from the surrounding tissue with a shifting layer. [Orthopedics. 2019; 42(3):168-175.].

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.

Vascular supply of the femoral head in sheep-Implications for the ovine femoroacetabular impingement model.

Sheep hips have a natural non-spherical head similar to a cam-type deformity in human beings. By performing an intertrochanteric varus osteotomy, cam-type femoroacetabular impingement can be induced experimentally. In sheep, the aspherical portion is located superiorly-exactly matching the region where the superior retinacular vessels enter the femoral head-neck junction in human beings. In order to fully exploit the potential of this experimental FAI model, a safe osteochondroplasty of the superior asphericity would need to be done without the risk of avascular necrosis. The aim of this study was to describe the vascular anatomy of the femoral head in sheep from the aorta to the retinacular vessels in order to perform safe femoral osteochondroplasty of the superior femoral asphericity in sheep. Sixty-two ovine hips were analyzed using CT angiography (30 hips), post mortem intravascular latex injection (6 hips), vascular corrosion casting (6 hips), and analysis of the distribution of vascular foramina around the femoral head-neck junction in macerated ovine femora (20 hips). The ovine femoral head receives its blood supply from anterior retinacular arteries from the lateral femoral circumflex artery, and from posterior retinacular arteries from the medial femoral circumflex artery. The superior aspherical portion is free of vessels. Detailed knowledge about vascular anatomy of sheep hips is of clinical significance since it allows to perform osteochondroplasty of the superior aspherical portion in the experimental ovine FAI model safely without the risk of osteonecrosis. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2340-2348, 2018.

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.