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Objective Inter-observer agreement in the context of oral epithelial dysplasia (OED) grading has been notoriously unreliable and can impose barriers for developing new molecular markers and diagnostic technologies. This paper aimed to report the details of a 3-stage histopathology review and adjudication process with the goal of achieving a consensus histopathologic diagnosis of each biopsy. Study Design Two adjacent serial histological sections of oral lesions from 846 patients were independently scored by two different pathologists from a pool of four. In instances where the original two pathologists disagreed, a third, independent adjudicating pathologist conducted a review of both sections. If a majority agreement was not achieved, the third stage involved a face-to-face consensus review. Results Individual pathologist pair kappa values ranged from 0.251 – 0.706 (fair – good) before the 3-stage review process During the initial review phase, the two pathologists agreed on a diagnosis for 69.9% of the cases. After the adjudication review by a third pathologist, an additional 22.8% of cases were given a consensus diagnosis (agreement of 2 out of 3 pathologists). Following the face-to-face review, the remaining 7.3% of cases had a consensus diagnosis. Conclusion The use of the defined protocol resulted in a substantial increase (30%) in diagnostic agreement and has the potential to improve the level of agreement for establishing gold standards for studies based on histopathologic diagnosis.

Bone tissue healing is a dynamic process that is initiated by the recruitment of osteoprogenitor cells followed by their migration, proliferation, differentiation, and development of a mineralizing extracellular matrix. The work aims to manufacture a functionalized porous membrane that stimulates early events in bone healing for initiating a regenerative cascade. Layer‐by‐layer (LbL) assembly is proposed to modify the surface of osteoconductive electrospun meshes, based on poly(lactic‐co‐glycolic acid) and nanohydroxyapatite, by using poly(allylamine hydrochloride) and poly(sodium 4‐styrenesulfonate) as polyelectrolytes. Molecular cues are incorporated by grafting peptide fragments into the discrete nanolayers. KRSR (lysine‐arginine‐serine‐arginine) sequence is grafted to enhance cell adhesion and proliferation, NSPVNSKIPKACCVPTELSAI to guide bone marrow mesenchymal stem cells differentiation in osteoblasts, and FHRRIKA (phenylalanine‐histidine‐arginine‐arginine‐isoleucine‐lysine‐alanine) to improve mineralization matrix formation. Scanning electron microscopy, infrared spectroscopy, and X‐ray photoelectron spectroscopy demonstrate the successful surface functionalization. Furthermore, the peptide incorporation enhances cellular processes, with good viability and significant increase of alkaline phosphatase activity, osteopontin, and osteocalcin. The functionalized membrane induces a favorable in vivo response after implantation for four weeks in nonhealing rat calvarial defect model. It is concluded that the multilayer nanoencapsulation of biofunctional peptides using LbL approach has significant potential as innovative manufacturing technique to improve bone regeneration in orthopedic and craniofacial medical devices.

Bone graft procedures typically require surgeons to harvest bone from a second site on a given patient (Autograft) before repairing a bone defect. However, this results in increased surgical time, excessive blood loss and a significant increase in pain. In this context a synthetic bone graft with excellent histocompatibility, built in antibacterial efficacy and the ability to regenerate healthy tissue in place of diseased tissue would be a significant step forward relative to current state of the art philosophies. We developed a range of calcium-strontium-zinc-silicate glass based bone grafts and characterised their structure and physical properties, then evaluated their in vitro cytotoxicity and in vivo biocompatibility using standardised models from the literature. A graft (designated BT109) of composition 0.28SrO/0.32ZnO/0.40 SiO(2) (mol fraction) was the best performing formulation in vitro shown to induce extremely mild cytopathic effects (cell viability up to 95%) in comparison with the commercially available bone graft Novabone (cell viability of up to 72%). Supplementary to this, the grafts were examined using the standard rat femur healing model on healthy Wister rats. All grafts were shown to be equally well tolerated in bone tissue and new bone was seen in close apposition to implanted particles with no evidence of an inflammatory response within bone. Complimentary to this BT109 was implanted into the femurs of ovariectomized rats to monitor the response of osteoporotic tissue to the bone grafts. The results from this experiment indicate that the novel grafts perform equally well in osteoporotic tissue as in healthy tissue, which is encouraging given that bone response to implants is usually diminished in ovariectomized rats. In conclusion these materials exhibit significant potential as synthetic bone grafts to warrant further investigation and optimisation.

The regeneration of large bone defects remains clinically challenging. The aim of our study was to use a rat model to use nasal chondrocytes to engineer a hypertrophic cartilage tissue which could be remodelled into bone in vivo by endochondral ossification.Primary adult rat nasal chondrocytes were isolated from the nasal septum, the cell numbers expanded in monolayer culture and the cells cultured in vitro on polyglycolic acid scaffolds in chondrogenic medium for culture periods of 5-10 weeks. Hypertrophic differentiation was assessed by determining the temporal expression of key marker genes and proteins involved in hypertrophic cartilage formation. The temporal changes in the genes measured reflected the temporal changes observed in the growth plate. Collagen II gene expression increased 6fold by day 7 and was then significantly downregulated from day 14 onwards. Conversely, collagen X gene expression was detectable by day 14 and increased 100-fold by day 35. The temporal increase in collagen X expression was mirrored by increases in alkaline phosphatase gene expression which also was detectable by day 14 with a 30-fold increase in gene expression by day 35. Histological and immunohistochemical analysis of the engineered constructs showed increased chondrocyte cell volume (31-45 µm), deposition of collagen X in the extracellular matrix and expression of alkaline phosphatase activity. However, no cartilage mineralisation was observed in in vitro culture of up to 10 weeks. On subcutaneous implantation of the hypertrophic engineered constructs, the grafts became vascularised, cartilage mineralisation occurred and loss of the proteoglycan in the matrix was observed.Implantation of the hypertrophic engineered constructs into a rat cranial defect resulted in angiogenesis, mineralisation and remodelling of the cartilage tissue into bone. Micro-CT analysis indicated that defects which received the engineered hypertrophic constructs showed 38.48% in bone volume compared to 7.01% in the control defects. M A N U S C R I P T A C C E P T E D ACCEPTED MANUSCRIPT 4Development of tissue engineered hypertrophic cartilage to use as a bone graft substitute is exciting development in regenerative medicine. This is a proof of principal study demonstrating the potential of nasal chondrocytes to engineer hypertrophic cartilage which will remodel into bone on in vivo transplantation. This approach to making engineered hypertrophic cartilage grafts could form the basis of a new potential future clinical treatment for maxillofacial reconstruction.