Surgery (St Vincent's) - Research Publications
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ItemIn situ handheld three-dimensional bioprinting for cartilage regeneration(WILEY, 2018-03)Articular cartilage injuries experienced at an early age can lead to the development of osteoarthritis later in life. In situ three-dimensional (3D) printing is an exciting and innovative biofabrication technology that enables the surgeon to deliver tissue-engineering techniques at the time and location of need. We have created a hand-held 3D printing device (biopen) that allows the simultaneous coaxial extrusion of bioscaffold and cultured cells directly into the cartilage defect in vivo in a single-session surgery. This pilot study assessed the ability of the biopen to repair a full-thickness chondral defect and the early outcomes in cartilage regeneration, and compared these results with other treatments in a large animal model. A standardized critical-sized full-thickness chondral defect was created in the weight-bearing surface of the lateral and medial condyles of both femurs of six sheep. Each defect was treated with one of the following treatments: (i) hand-held in situ 3D printed bioscaffold using the biopen (HH group), (ii) preconstructed bench-based printed bioscaffolds (BB group), (iii) microfractures (MF group) or (iv) untreated (control, C group). At 8 weeks after surgery, macroscopic, microscopic and biomechanical tests were performed. Surgical 3D bioprinting was performed in all animals without any intra- or postoperative complication. The HH biopen allowed early cartilage regeneration. The results of this study show that real-time, in vivo bioprinting with cells and scaffold is a feasible means of delivering a regenerative medicine strategy in a large animal model to regenerate articular cartilage.
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ItemProtocols for Culturing and Imaging a Human Ex Vivo Osteochondral Model for Cartilage Biomanufacturing Applications(MDPI, 2019-02-02)Cartilage defects and diseases remain major clinical issues in orthopaedics. Biomanufacturing is now a tangible option for the delivery of bioscaffolds capable of regenerating the deficient cartilage tissue. However, several limitations of in vitro and experimental animal models pose serious challenges to the translation of preclinical findings into clinical practice. Ex vivo models are of great value for translating in vitro tissue engineered approaches into clinically relevant conditions. Our aim is to obtain a viable human osteochondral (OC) model to test hydrogel-based materials for cartilage repair. Here we describe a detailed step-by-step framework for the generation of human OC plugs, their culture in a perfusion device and the processing procedures for histological and advanced microscopy imaging. Our ex vivo OC model fulfils the following requirements: the model is metabolically stable for a relevant culture period of 4 weeks in a perfusion bioreactor, the processing procedures allowed for the analysis of 3 different tissues or materials (cartilage, bone and hydrogel) without compromising their integrity. We determined a protocol and the settings for a non-linear microscopy technique on label free sections. Furthermore, we established a clearing protocol to perform light sheet-based observations on the cartilage layer without the need for tedious and destructive histological procedures. Finally, we showed that our OC system is a clinically relevant in terms of cartilage regeneration potential. In conclusion, this OC model represents a valuable preclinical ex vivo tool for studying cartilage therapies, such as hydrogel-based bioscaffolds, and we envision it will reduce the number of animals needed for in vivo testing.
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Item3D bioprinting of cartilage for orthopedic surgeons: reading between the lines(FRONTIERS MEDIA SA, 2015)Chondral and osteochondral lesions represent one of the most challenging and frustrating scenarios for the orthopedic surgeon and for the patient. The lack of therapeutic strategies capable to reconstitute the function and structure of hyaline cartilage and to halt the progression toward osteoarthritis has brought clinicians and scientists together, to investigate the potential role of tissue engineering as a viable alternative to current treatment modalities. In particular, the role of bioprinting is emerging as an innovative technology that allows for the creation of organized 3D tissue constructs via a "layer-by-layer" deposition process. This process also has the capability to combine cells and biomaterials in an ordered and predetermined way. Here, we review the recent advances in cartilage bioprinting and we identify the current challenges and the directions for future developments in cartilage regeneration.
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ItemHandheld Co-Axial Bioprinting: Application to in situ surgical cartilage repair(NATURE PORTFOLIO, 2017-07-19)Three-dimensional (3D) bioprinting is driving major innovations in the area of cartilage tissue engineering. Extrusion-based 3D bioprinting necessitates a phase change from a liquid bioink to a semi-solid crosslinked network achieved by a photo-initiated free radical polymerization reaction that is known to be cytotoxic. Therefore, the choice of the photocuring conditions has to be carefully addressed to generate a structure stiff enough to withstand the forces phisiologically applied on articular cartilage, while ensuring adequate cell survival for functional chondral repair. We recently developed a handheld 3D printer called "Biopen". To progress towards translating this freeform biofabrication tool into clinical practice, we aimed to define the ideal bioprinting conditions that would deliver a scaffold with high cell viability and structural stiffness relevant for chondral repair. To fulfill those criteria, free radical cytotoxicity was confined by a co-axial Core/Shell separation. This system allowed the generation of Core/Shell GelMa/HAMa bioscaffolds with stiffness of 200KPa, achieved after only 10 seconds of exposure to 700 mW/cm2 of 365 nm UV-A, containing >90% viable stem cells that retained proliferative capacity. Overall, the Core/Shell handheld 3D bioprinting strategy enabled rapid generation of high modulus bioscaffolds with high cell viability, with potential for in situ surgical cartilage engineering.
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ItemFunctional reconstruction of sarcoma defects utilising innervated free flaps.(Hindawi Limited, 2012)Soft-tissue reconstruction following preoperative radiotherapy and wide resection of soft tissue sarcoma remains a challenge. Pedicled and free tissue transfers are an essential part of limb sparing surgery. We report 22 cases of sarcoma treated with radiotherapy and wide excision followed by one-stage innervated free or pedicled musculocutaneous flap transfers. The resection involved the upper limb in 3 cases, the lower limb in 17, and the abdominal wall in 2. The flaps used for the reconstruction were mainly latissimus dorsi and gracilis. The range of motion was restored fully in 14 patients. The muscle strength of the compartment reconstructed was of grades 4 and 5 in all patients except one. The overall function was excellent in all the cases with functional scores of 71.2% in the upper limb and 84% in the lower limb. The only 2 major complications were flap necrosis, both revised with another flap, one of which was innervated with restoration of function. Innervated flaps are valuable alternatives for reconstruction after sarcoma resection in the extremity and in the abdominal wall. The excellent functional results are encouraging, and we believe that innervated muscle reconstruction should be encouraged in the treatment of sarcoma after radiotherapy and wide resection.
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ItemAdipose-Derived Mesenchymal Stem Cells in the Use of Cartilage Tissue Engineering: The Need for a Rapid Isolation Procedure(HINDAWI LTD, 2018)Mesenchymal stem cells (MSCs) have shown much promise with respect to their use in cartilage tissue engineering. MSCs can be obtained from many different tissue sources. Among these, adipose tissue can provide an abundant source of adipose-derived mesenchymal stem cells (ADMSCs). The infrapatellar fat pad (IFP) is a promising source of ADMSCs with respect to producing a cartilage lineage. Cell isolation protocols to date are time-consuming and follow conservative approaches that rely on a long incubation period of 24-48 hours. The different types of ADMSC isolation techniques used for cartilage repair will be reviewed and compared with the view of developing a rapid one-step isolation protocol that can be applied in the context of a surgical procedure.
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ItemCartilage Tissue Engineering Using Stem Cells and Bioprinting Technology-Barriers to Clinical Translation(FRONTIERS MEDIA SA, 2018-11-27)There is no long-term treatment strategy for young and active patients with cartilage defects. Early and effective joint preserving treatments in these patients are crucial in preventing the development of osteoarthritis. Tissue engineering over the past few decades has presented hope in overcoming the issues involved with current treatment strategies. Novel advances in 3D bioprinting technology have promoted more focus on efficient delivery of engineered tissue constructs. There have been promising in-vitro studies and several animal studies looking at 3D bioprinting of engineered cartilage tissue. However, to date there are still no human clinical trials using 3D printed engineered cartilage tissue. This review begins with discussion surrounding the difficulties with articular cartilage repair and the limitations of current clinical management options which have led to research in cartilage tissue engineering. Next, the major barriers in each of the 4 components of cartilage tissue engineering; cells, scaffolds, chemical, and physical stimulation will be reviewed. Strategies that may overcome these barriers will be discussed. Finally, we will discuss the barriers surrounding intraoperative delivery of engineered tissue constructs and possible solutions.
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ItemIn vivo biocompatibility of porous and non-porous polypyrrole based trilayered actuators(SPRINGER, 2017-11)Trilayered polypyrrole (PPy) actuators have high stress density, low modulus and have wide potential biological applications including use in artificial muscles and in limb prosthesis after limb amputation. This article examines the in vivo biocompatibility of actuators in muscle using rabbit models. The actuators were specially designed with pores to encourage tissue in growth; this study also assessed the effect of such pores on the stability of the actuators in vivo. Trilayered PPy actuators were either laser cut with 150 µm pores or left pore-less and implanted into rabbit muscle for 3 days, 2 weeks, 4 weeks and 8 weeks and retrieved subsequently for histological analysis. In a second set of experiments, the cut edges of pores in porous actuator strips were further sealed by PPy after laser cutting to further improve its stability in vivo. Porous actuators with and without PPy sealing of pore edges were implanted intramuscularly for 4 and 8 weeks and assessed with histology. Pore-less actuators incited a mild inflammatory response, becoming progressively walled off by a thin layer of fibrous tissue. Porous actuators showed increased PPy fragmentation and delamination with associated greater foreign body response compared to pore-less actuators. The PPy fragmentation was minimized when the pore edges were sealed off by PPy after laser cutting showing less PPy debris. Laser cutting of the actuators with pores destabilizes the PPy. This can be overcome by sealing the cut edges of the pores with PPy after laser. The findings in this article have implications in future design and manufacturing of PPy actuator for use in vivo.