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 (iPSCs) represent an unlimited source of pluripotent cells capable of differentiating into any cell type of the body. Several studies have demonstrated the valuable use of iPSCs as a tool for studying the molecular and cellular mechanisms underlying disorders affecting bone, cartilage and muscle, as well as their potential for tissue repair. Musculoskeletal diseases are one of the major causes of disability worldwide and impose an important socio-economic burden. To date there is neither cure nor proven approach for effectively treating most of these conditions and therefore new strategies involving the use of cells have been increasingly investigated in the recent years. Nevertheless, some limitations related to the safety and differentiation protocols among others remain, which humpers the translational application of these strategies. Nonetheless, the potential is indisputable and iPSCs are likely to be a source of different types of cells useful in the musculoskeletal field, for either disease modeling or regenerative medicine. In this review, we aim to illustrate the great potential of iPSCs by summarizing and discussing the in vitro tissue regeneration preclinical studies that have been carried out in the musculoskeletal field by using iPSCs.

Human bone marrow-derived mesenchymal stromal cells (MSCs) obtained from aged patients are prone to senesce and diminish their differentiation potential, therefore limiting their usefulness for osteochondral regenerative medicine approaches or to study age-related diseases, such as osteoarthiritis (OA). MSCs can be transduced with immortalizing genes to overcome this limitation, but transduction of primary slow-dividing cells has proven to be challenging. Methods for enhancing transduction efficiency (such as spinoculation, chemical adjuvants, or transgene expression inductors) can be used, but several parameters must be adapted for each transduction system. In order to develop a transduction method suitable for the immortalization of MSCs from aged donors, we used a spinoculation method. Incubation parameters of packaging cells, speed and time of centrifugation, and valproic acid concentration to induce transgene expression have been adjusted. In this way, four immortalized MSC lines (iMSC#6, iMSC#8, iMSC#9, and iMSC#10) were generated. These immortalized MSCs (iMSCs) were capable of bypassing senescence and proliferating at a higher rate than primary MSCs. Characterization of iMSCs showed that these cells kept the expression of mesenchymal surface markers and were able to differentiate towards osteoblasts, adipocytes, and chondrocytes. Nevertheless, alterations in the CD105 expression and a switch of cell fate-commitment towards the osteogenic lineage have been noticed. In conclusion, the developed transduction method is suitable for the immortalization of MSCs derived from aged donors. The generated iMSC lines maintain essential mesenchymal features and are expected to be useful tools for the bone and cartilage regenerative medicine research.

Synovial fibrosis is a pathological process which contributes to joint pain and stiffness in several musculoskeletal disorders. Fucoidans, sulfated polysaccharides found in brown algae, have recently emerged as promising therapeutic agents. Despite the increasing amount of evidence suggesting the protective role of fucoidans in different experimental approaches of human fibrotic disorders, the effect of these sulfated polysaccharides on synovial fibrosis has not been investigated yet. By an in vitro experimental approach in fibroblast-like synoviocytes, we detected that fucoidans inhibit their differentiation into myofibroblasts with tumor cell-like characteristics and restore apoptosis. Composition and structure of fucoidan appear to be critical for the detected activity. Furthermore, protective effects of these sulfated polysaccharides are mediated by upregulation of nitric oxide production and modulation of TGF-β/smad pathway. Altogether, our results support the use of fucoidans as therapeutic compounds in the treatment of the fibrotic processes involved in rheumatic pathologies.

Objectives: This study investigates the role of retinol binding protein 4 (RBP4) in an articular context. RBP4, a vitamin A transporter, is linked to various metabolic diseases. Methods: Synovial fluid RBP4 levels were assessed in crystalline arthritis (CA) patients using ELISA. RBP4’s impact on articular cell types was analysed in vitro through RT-PCR and flow cytometry. Proteomic analysis was conducted on primary human osteoarthritis chondrocytes (hOACs). Results: Synovial fluid RBP4 concentrations in CA patients correlated positively with glucose levels and negatively with synovial leukocyte count and were elevated in hypertensive patients. In vitro, these RBP4 concentrations activated neutrophils, induced the expression of inflammatory factors in hOACs as well as synoviocytes, and triggered proteomic changes consistent with inflammation. Moreover, they increased catabolism and decreased anabolism, mitochondrial dysfunction, and glycolysis promotion. Both in silico and in vitro experiments suggested that RBP4 acts through TLR4. Conclusions: This study identifies relevant RBP4 concentrations in CA patients’ synovial fluids, linking them to hypertensive patients with a metabolic disruption. Evidence is provided that RBP4 acts as a DAMP at these concentrations, inducing robust inflammatory, catabolic, chemotactic, and metabolic responses in chondrocytes, synoviocytes, and neutrophils. These effects may explain RBP4-related metabolic diseases’ contribution to joint destruction in various rheumatic conditions like CA.

Different findings indicate that type 2 diabetes is an independent risk factor for osteoarthritis (OA). However, the mechanisms underlying the connection between both diseases remain unclear. Changes in the balance of hydrogen sulphide (H₂S) are thought to play an important role in the pathogenesis of diabetes and its complications, although its role is still controversial. In this study, we examined the modulation of H₂S levels in serum and chondrocytes from OA diabetic (DB) and non-diabetic (non-DB) patients and in cells under glucose stress, in order to elucidate whether impairment in H₂S-mediated signalling could participate in the onset of DB-related OA. Here, we identified a reduction in H₂S synthesis in the cartilage from OA-DB patients and in cells under glucose stress, which is associated with hyperglycaemia-mediated dysregulation of chondrocyte metabolism. In addition, our results indicate that H₂S is an inductor of the Nrf-2/HO-1 signalling pathway in cartilage, but is also a downstream target of Nrf-2 transcriptional activity. Thereby, impairment of the H₂S/Nrf-2 axis under glucose stress or DB triggers chondrocyte catabolic responses, favouring the disruption of cartilage homeostasis that characterizes OA pathology. Finally, our findings highlight the benefits of the use of exogeneous sources of H₂S in the treatment of DB-OA patients, and warrant future clinical studies.