Innes Wise

Patient-matched scaffold guided bone regeneration (SGBR) has been poised to revolutionize the management of critical-sized bone defects due to trauma, tumors, infection, and congenital deformities. However, translation has been hampered by the focus on bioresorbable scaffolds where the rate of scaffold degradation needs to be precisely and predictably matched to the rate of new bone formation. Furthermore, resorbable ceramics and polymers require augmentation by metal plates to overcome their mechanical limitations under load. Consequently, there are no published studies using SGBR for segmental defects without metal plates, which cause stress shielding and X-ray perturbation. Even with augmentation, very few studies use animal models where the loads are comparable to humans. Segmental defects of the mandible represent one of the most challenging examples of critical-sized bone defects due to the high tensile and shear stresses encountered during mastication. The ovine mandible is an excellent model because the high and repetitive loads exceed those encountered in humans. Here, we describe the first successful long-term reconstruction of ovine segmental mandibulectomy defects using permanent, patient-matched, numerically optimized, 3D-printed, thermally toughened, plasma-treated, and selectively-laser-sintered polyetherketone gyroid scaffolds. The scaffolds house a resorbable ceramic lattice infused with a stem cell laden hydrogel and serve as an osteoinductive reservoir of calcium. Durable clinical performance and osseointegration was established in vivo and ex vivo, indicating for the first time that there is a reliable and translatable patient-matched SGBR alternative to traditional bone grafts and metal plate fixation.

The development of small-diameter vascular grafts requires testing in large animal models before advancing to clinical trials. Vascular graft interposition implantation in sheep carotid arteries (CAs) is the most widely used model, but ovine CAs are prone to severe spasm following surgical manipulation, potentially impairing graft performance assessment. There is paucity in the literature on reducing sheep CA spasm using effective vasodilator therapeutic protocols. In this study, four healthy Merino cross White Suffolk wethers (1–2 years, 52.1 ± 0.8 kg) underwent CT angiography and CA graft surgery. CT angiography using iodinated contrast agent was performed with innominate artery access through the CA or ascending aortic arch access through the femoral artery. Sheep then underwent right CA sham surgery or left CA vascular graft implantation. A variety of vasodilators, including papaverine, sodium nitroprusside, verapamil, and their combination, were tested for preventing or treating CA spasms intraoperatively. Blood flow was reassessed immediately after CA surgery using CT angiography. The results showed that innominate artery access through the CA for CT angiography in sheep induced presurgical CA spasm with reduced arterial flow. Conversely, ascending aortic arch access through the femoral artery for CT angiography did not cause CA spasm and maintained arterial flow. During CA graft surgery, surgical trauma induced CA spasm, which was prevented by localized intra-arterial administration of vasodilators papaverine hydrochloride and verapamil before significant surgical manipulation.

Considerable research is being undertaken to develop novel biomaterials-based approaches for surgical reconstruction of bone defects. This extends to three-dimensional (3D) printed materials that provide stable, structural, and functional support in vivo . However, few preclinical models can simulate in vivo human biological conditions for clinically relevant testing. In this study we describe a novel ovine model that allows evaluation of in vivo osteogenesis via contact with bone and/or periosteum interfaced with printed polymer bioreactors loaded with biomaterial bone substitutes. The infraspinous scapular region of 14 Dorset cross sheep was exposed. Vascularized periosteum was elevated either attached to the infraspinatus muscle or separately. In both cases, the periosteum was supplied by the periosteal branch of the circumflex scapular vessels. In eight sheep, a 3D printed 4-chambered polyetheretherketone bioreactor was wrapped circumferentially in vascularized periosteum. In 6 sheep, 12 double-sided 3D printed 2-chambered polyetherketone bioreactors were secured to the underlying bone allowing direct contact with the bone on one side and periosteum on the other. Our model enabled simultaneous testing of up to 24 (12 double-sided) 10 × 10 × 5 mm bioreactors per scapula in the flat contact approach or a single 40 × 10 mm four-chambered bioreactor per scapula using the periosteal wrap. De novo bone growth was evaluated using histological and radiological analysis. Of importance, the experimental model was well tolerated by the animals and provides a versatile approach for comparing the osteogenic potential of cambium on the bone surface and elevated with periosteum. Furthermore, the periosteal flaps were sufficiently large for encasing bioreactors containing biomaterial bone substitutes for applications such as segmental mandibular reconstruction.

Autologous bone replacement remains the preferred treatment for segmental defects of the mandible; however, it cannot replicate complex facial geometry and causes donor site morbidity. Bone tissue engineering has the potential to overcome these limitations. Various commercially available calcium phosphate-based bone substitutes (Novabone^®, BioOss^®, and Zengro^®) are commonly used in dentistry for small bone defects around teeth and implants. However, their role in ectopic bone formation, which can later be applied as vascularized graft in a bone defect, is yet to be explored. Here, we compare the above-mentioned bone substitutes with autologous bone with the aim of selecting one for future studies of segmental mandibular repair. Six female sheep, aged 7–8 years, were implanted with 40 mm long four-chambered polyether ether ketone (PEEK) bioreactors prepared using additive manufacturing followed by plasma immersion ion implantation (PIII) to improve hydrophilicity and bioactivity. Each bioreactor was wrapped with vascularized scapular periosteum and the chambers were filled with autologous bone graft, Novabone^®, BioOss^®, and Zengro^®, respectively. The bioreactors were implanted within a subscapular muscle pocket for either 8 weeks (two sheep), 10 weeks (two sheep), or 12 weeks (two sheep), after which they were removed and assessed by microCT and routine histology. Moderate bone formation was observed in autologous bone grafts, while low bone formation was observed in the BioOss^® and Zengro^® chambers. No bone formation was observed in the Novabone^® chambers. Although the BioOss^® and Zengro^® chambers contained relatively small amounts of bone, endochondral ossification and retained hydroxyapatite suggest their potential in new bone formation in an ectopic site if a consistent supply of progenitor cells and/or growth factors can be ensured over a longer duration.

Periosteum is a highly vascularized membrane lining the surface of bones. It plays essential roles in bone repair following injury and reconstruction following invasive surgeries. To broaden the use of periosteum, including for augmenting in vitro bone engineering and/or in vivo bone repair, we have developed an ex vivo perfusion bioreactor system to maintain the cellular viability and metabolism of surgically resected periosteal flaps. Each specimen was placed in a 3D printed bioreactor connected to a peristaltic pump designed for the optimal flow rates of tissue perfusate. Nutrients and oxygen were perfused via the periosteal arteries to mimic physiological conditions. Biochemical assays and histological staining indicate component cell viability after perfusion for almost 4 weeks. Our work provides the proof-of-concept of ex vivo periosteum perfusion for long-term tissue preservation, paving the way for innovative bone engineering approaches that use autotransplanted periosteum to enhance in vivo bone repair.

Ketamine-based total intravenous anaesthesia techniques are commonly used in equine practice for ponies requiring short procedures such as castration in field conditions. When a longer duration of recumbency than provided by the initial dose of anaesthetic agents is required, administration of supplementary ‘top-up’ doses of anaesthetic agents is required. Ideally, a single dose of anaesthetic agents would reliably achieve a longer duration of action whilst maintaining adequate anaesthetic, surgical and recovery qualities. This prospective, randomised and blinded study aimed to compare the UK-licensed induction dose of ketamine with an increased dose in ponies undergoing castration in field conditions. The hypothesis was that an increased dose would produce a longer duration of action without negatively affecting qualities of anaesthesia, surgical conditions and recovery. Ponies were randomly allocated to receive ketamine at either 2.2 mg kg^-1 (K2.2) or 3 mg kg^-1 (K3) combined with diazepam 20 μg kg^-1 following pre-anaesthetic medication with romifidine and butorphanol. Quality of anaesthesia, surgery and recovery were scored using simple descriptive scales (SDS) and timings of key events recorded. Top-up doses of ketamine 0.5 mg kg^-1 were administered if anaesthesia was inadequate during surgery. Time of top-up doses and total ketamine doses were recorded. Data were analysed using Student t-tests or the Mann-Whitney U test (p < 0.05). Thirty-six ponies completed the study. Six ponies enrolled were excluded due to cryptorchidism or surgical complications that required deviation from the anaesthetic protocol. There were no differences in timing of events recorded, number of ponies requiring top-up ketamine, timing and frequency of top-ups or total ketamine dose. Scores for anaesthetic and recovery qualities, and surgical conditions were similar between groups. Both induction doses of ketamine provided a similar duration of action and provided conditions suitable to anaesthetise ponies undergoing castration.