Mechanical loading rescues mechanoresponsiveness in a human osteoarthritis explant model despite Wnt activation.

OBJECTIVES: Optimizing rehabilitation strategies for osteoarthritis necessitates a comprehensive understanding of chondrocytes' mechanoresponse in both health and disease, especially in the context of the interplay between loading and key pathways involved in osteoarthritis (OA) development, like canonical Wnt signaling. This study aims to elucidate the role of Wnt signaling in the mechanoresponsiveness of healthy and osteoarthritic human cartilage. METHODS: We used an ex-vivo model involving short-term physiological mechanical loading of human cartilage explants. First, the loading protocol for subsequent experiments was determined. Next, loading was applied to non-OA-explants with or without Wnt activation with CHIR99021. Molecular read-outs of anabolic, pericellular matrix and matrix remodeling markers were used to assess the effect of Wnt on cartilage mechanoresponse. Finally, the same set-up was used to study the effect of loading in cartilage from patients with established OA. RESULTS: Our results confirm that physiological loading maintains expression of anabolic genes in non-OA cartilage, and indicate a deleterious effect of Wnt activation in the chondrocyte mechanoresponsiveness. This suggests that loading-induced regulation of chondrocyte markers occurs downstream of canonical Wnt signaling. Interestingly, our study highlighted contrasting mechanoresponsiveness in the model of Wnt activation and the established OA samples, with established OA cartilage maintaining its mechanoresponsiveness, and mechanical loading rescuing the chondrogenic phenotype. CONCLUSION: This study provides insights into the mechanoresponsiveness of human cartilage in both non-OA and OA conditions. These findings hold the potential to contribute to the development of strategies that optimize the effect of dynamic compression by correcting OA pathological cell signaling.

Contribution of collagen degradation and proteoglycan depletion to cartilage degeneration in primary and secondary osteoarthritis: an in silico study.

OBJECTIVES: Current experimental approaches cannot elucidate the effect of maladaptive changes on the main cartilage constituents during the degeneration process in osteoarthritis (OA). In silico approaches, however, allow creating 'virtual knock-out' cases to elucidate these effects in a constituent-specific manner. We used such an approach to study the main mechanisms of cartilage degeneration in different mechanical loadings associated with the following OA etiologies: (1) physiological loading of degenerated cartilage, (2) injurious loading of healthy intact cartilage and (3) physiological loading of cartilage with a focal defect. METHODS: We used the recently developed Cartilage Adaptive REorientation Degeneration (CARED) framework to simulate cartilage degeneration associated with primary and secondary OA (OA cases (1)-(3)). CARED incorporates numerical description of tissue-level cartilage degeneration mechanisms in OA, namely, collagen degradation, collagen reorientation, fixed charged density loss and tissue hydration increase following mechanical loading. We created 'virtual knock-out' scenarios by deactivating these degenerative processes one at a time in each of the three OA cases. RESULTS: In the injurious loading of intact and physiological loading of degenerated cartilage, collagen degradation drives degenerative changes through fixed charge density loss and tissue hydration rise. In contrast, the two later mechanisms were more prominent in the focal defect cartilage model. CONCLUSION: The virtual knock-out models reveal that injurious loading to intact cartilage and physiological loading to degenerated cartilage induce initial degenerative changes in the collagen network, whereas, in the presence of a focal cartilage defect, mechanical loading initially causes proteoglycans (PG) depletion, before changes in the collagen fibril network occur.

Generation and characterization of human induced pluripotent stem cells (iPSCs) from hand osteoarthritis patient-derived fibroblasts.

Knowledge and research results about hand osteoarthritis (hOA) are limited due to the lack of samples and animal models of the disease. Here, we report the generation of two induced pluripotent stem cell (iPSC)-lines from patients with radiographic hOA. Furthermore, we wondered whether these iPSC-lines carried single nucleotide polymorphisms (SNPs) within genes that have been associated with hOA. Finally, we performed chondrogenic differentiation of the iPSCs in order to prove their usefulness as cellular models of the disease. We performed a non-integrative reprogramming of dermal fibroblasts obtained from two patients with radiographic rhizarthrosis and non-erosive hOA by introducing the transcriptional factors Oct4, Sox2, Klf4 and c-Myc using Sendai virus. After reprogramming, embryonic stem cell-like colonies emerged in culture, which fulfilled all the criteria to be considered iPSCs. Both iPSC-lines carried variants associated with hOA in the four studied genes and showed differences in their chondrogenic capacity when compared with a healthy control iPSC-line. To our knowledge this is the first time that the generation of iPSC-lines from patients with rhizarthrosis and non-erosive hOA is reported. The obtained iPSC-lines might enable us to model the disease in vitro, and to deeper study both the molecular and cellular mechanisms underlying hOA.

Generation of a human control iPS cell line (ESi080-A) from a donor with no rheumatic diseases.

Here, we report the establishment of the human iPS cell line N1-FiPS4F#7 generated from skin cells of a patient with no rheumatic diseases, thus obtaining an appropriate control iPS cell line for researchers working in the field of rheumatic diseases. The reprogramming factors Oct4, Sox2, Klf4 and c-Myc were introduced using a non-integrating reprogramming strategy involving Sendai Virus.

Usefulness of Mesenchymal Cell Lines for Bone and Cartilage Regeneration Research.

The unavailability of sufficient numbers of human primary cells is a major roadblock for in vitro repair of bone and/or cartilage, and for performing disease modelling experiments. Immortalized mesenchymal stromal cells (iMSCs) may be employed as a research tool for avoiding these problems. The purpose of this review was to revise the available literature on the characteristics of the iMSC lines, paying special attention to the maintenance of the phenotype of the primary cells from which they were derived, and whether they are effectively useful for in vitro disease modeling and cell therapy purposes. This review was performed by searching on Web of Science, Scopus, and PubMed databases from 1 January 2015 to 30 September 2019. The keywords used were ALL = (mesenchymal AND ("cell line" OR immortal*) AND (cartilage OR chondrogenesis OR bone OR osteogenesis) AND human). Only original research studies in which a human iMSC line was employed for osteogenesis or chondrogenesis experiments were included. After describing the success of the immortalization protocol, we focused on the iMSCs maintenance of the parental phenotype and multipotency. According to the literature revised, it seems that the maintenance of these characteristics is not guaranteed by immortalization, and that careful selection and validation of clones with particular characteristics is necessary for taking advantage of the full potential of iMSC to be employed in bone and cartilage-related research.

Induced pluripotent stem cells for cartilage repair: current status and future perspectives.

The establishment of cartilage regenerative medicine is an important clinical issue, but the search for cell sources able to restore cartilage integrity proves to be challenging. Human mesenchymal stromal cells (MSCs) are prone to form epiphyseal or hypertrophic cartilage and have an age-related limited proliferation. On the other hand, it is difficult to obtain functional chondrocytes from human embryonic stem cells (ESCs). Moreover, the ethical issues associated with human ESCs are an additional disadvantage of using such cells. Since their discovery in 2006, induced pluripotent stems cells (iPSCs) have opened many gateways to regenerative medicine research, especially in cartilage tissue engineering therapies. iPSCs have the capacity to overcome limitations associated with current cell sources since large numbers of autologous cells can be derived from small starting populations. Moreover, problems associated with epiphyseal or hypertrophic-cartilage formation can be overcome using iPSCs. iPSCs emerge as a promising cell source for treating cartilage defects and have the potential to be used in the clinical field. For this purpose, robust protocols to induce chondrogenesis, both in vitro an in vivo, are required. This review summarises the recent progress in iPSC technology and its applications for cartilage repair.