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Current regenerative cartilage therapies are associated with several drawbacks such as dedifferentiation of chondrocytes during expansion and the formation of fibrocartilage. Optimized chondrocyte expansion and tissue formation could lead to better clinical results of these therapies. In this study, a novel chondrocyte suspension expansion protocol that includes the addition of porcine notochordal cell-derived matrix was used to self-assemble human chondrocytes from osteoarthritic (OA) and nondegenerate (ND) origin into cartilage organoids containing collagen type II and proteoglycans. Proliferation rate and viability were similar for OA and ND chondrocytes and organoids formed had a similar histologic appearance and gene expression profile. Organoids were then encapsulated in viscoelastic alginate hydrogels to form larger tissues. Chondrocytes on the outer bounds of the organoids produced a proteoglycan-rich matrix to bridge the space between organoids. In hydrogels containing ND organoids some collagen type I was observed between the organoids. Surrounding the bulk of organoids in the center of the gels, in both OA and ND gels a continuous tissue containing cells, proteoglycans and collagen type II had been produced. No difference was observed in sulphated glycosaminoglycan and hydroxyproline content between gels containing organoids from OA or ND origin after 28 days. It was concluded that OA chondrocytes, which can be harvested from leftover surgery tissue, perform similar to ND chondrocytes in terms of human cartilage organoid formation and matrix production in alginate gels. This opens possibilities for their potential to serve as a platform for cartilage regeneration but also as an in vitro model to study pathways, pathology, or drug development., (© 2023 The Authors. Journal of Orthopaedic Research® published by Wiley Periodicals LLC on behalf of Orthopaedic Research Society.)

Integration of an implant with the surrounding tissue is a major challenge in cartilage regeneration. It is usually assessed with in vivo animal studies at the end-stage of implant development. To reduce animal experimentation and at the same time increase screening throughput and speed up implant development, this study examined whether integration of allogeneic cell-based implants with the surrounding native cartilage could be demonstrated in an ex vivo human osteochondral culture model. Chondrocytes were isolated from smooth cartilage tissue of fresh human tibial plateaus and condyles. They were expanded for 12 days either in three-dimensional spinner flask cultures to generate organoids, or in two-dimensional culture flasks for standard cell expansion. Three implant groups were created (fibrin+organoids, fibrin+cells, and fibrin only) and used to fill a Ø 6 mm full-depth chondral defect created in human osteochondral explants (Ø 10 mm, bone length cut to 4 mm) harvested from a second set of fresh human tibial plateaus. Explants were cultured for 1 or 28 days in a double-chamber culture platform. Histology showed that after 28 days the organoids on the interface of the defect remodeled and merged, and cells migrated through the fibrin glue bridging the space between the organoids and between the organoids and the native cartilage. For both conditions, newly formed tissue rich in proteoglycans and collagen type II was present mainly on the edges and in the corners of the defect. In these matrix-rich areas, cells resided in lacunae and the newly formed tissue integrated with the surrounding native cartilage. Biochemical analysis revealed a statistically significant effect of culture time on glycosaminoglycan (GAG) content, and showed a higher hydroxyproline (HYP) content for organoid-filled implants compared with cell-filled implants at both timepoints. This ex vivo human osteochondral culture system provides possibilities for exploration and identification of promising implant strategies based on evaluation of integration and matrix production under more controlled experimental conditions than possible in vivo .

To reduce animal experimentation and to overcome translational issues in cartilage tissue engineering, there is a need to develop an ex vivo human tissue-based approach. This study aims to demonstrate that a human osteochondral explant at different stages of osteoarthritis (OA) can be kept in long-term culture while preserving its viability and composition. Osteochondral explants with either a smooth or fibrillated cartilage surface, representing different OA stages, were harvested from fresh human tibial plateaus. Explants were cultured for 2 or 4 weeks in a double-chamber culture platform. The biochemical content of the cartilage of cultured explants did not significantly change over a period of 4 weeks and these findings were supported by histology. Chondrocytes mostly preserved their metabolic activity during culture and active bone and marrow were found in the periphery of the explants, while metabolic activity was decreased in the bone core in cultured explants compared to fresh explants. In fibrillated explants, chondrocyte viability decreased in the periphery of the sample in cultured groups compared to fresh explants (fresh, 94 ± 6%; cultured, 64% ± 17%, 2 weeks, and 69% ± 17%, 4 weeks; P < .05). Although biochemical and histological results did not show changes within the cartilage tissue, the viability of the explants should be carefully controlled for each specific use. This system provides an alternative to explore drug treatment and implant performance under more controlled experimental conditions than possible in vivo, in combination with clinically relevant human osteochondral tissue., (© 2020 The Authors. Journal of Orthopaedic Research® published by Wiley Periodicals LLC on behalf of Orthopaedic Research Society.)

Purpose: Animal models are frequently used to elucidate pathomechanism and pathophysiology of various disorders of the human intervertebral disc (IVD) and also to develop therapeutic approaches. Here we report morphological characteristics of the kangaroo lumbar IVDs and compare them with other animal models used in spine research., Methods: Twenty-five fresh-frozen cadaveric lumbar spines (T12-S1) derived from kangaroo carcases (Macropus giganteus) of undetermined age were first scanned in a C-Arm X-ray machine. A photograph of the axial section of the disc including a calibrated metric scale was also acquired. The digital radiographs and photographs were processed in ImageJ to determine the axial and sagittal plane dimensions for the whole disc (WD) and the nucleus pulposus (NP) and the mid-sagittal disc height for all the lumbar levels., Results: Our results suggest that the L6-S1 IVD in kangaroos is distinctly large compared with the upper lumbar IVDs. Based on previously published data, human lumbar IVDs are the largest of all the animal IVDs used in spine research, with camelid cervical IVDs being the closest relative in absolute dimensions (llamas: 78% in disc height, 40% in WD volume, and 38% in NP volume). Kangaroo L6-S1 IVD was approximately 51% in height, 20% in WD volume, and 20% in NP volume of the human lumbar IVD., Conclusions: We conclude that morphological similarities exist between a kangaroo and human lumbar IVD, especially with the lima bean shape in the axial plane, wedge shape in the sagittal plane, convexity at the cephalad endplates, and percentage volume occupied by the NP in the IVD. These slides can be retrieved under Electronic Supplementary Material.