The preservation of the chondrogenic phenotype and hypoxia-related physiological microenvironment are major challenges in the 2D culture of primary human chondrocytes. To address this problem, we develop a 3D culture system generating scaffold-free spheroids from human chondrocytes. Our results highlight the chondrogenic potential of cultured human articular chondrocytes in a 3D system combined with hypoxia independently of the cartilage source. After 14 days of culture, we developed spheroids with homogenous diameter and shape from hyaline cartilage donors. Spheroids generated in hypoxia showed a significantly increased glycosaminoglycans synthesis and up-regulated the expression of SOX9, ACAN, COL2A1, COMP, and SNAI1 compared to those obtained under normoxic conditions. Therefore, we conclude that spheroids developed under hypoxic conditions modulate the expression of chondrogenesis-related genes and native tissue features better than 2D cultures. Thus, this scaffold-free 3D culture system represents a novel in vitro model that can be used for cartilage biology research.

Giant cell tumor of bone (GCTB) is a primary bone tumor that affects skeletally mature people and whose main treatment is surgical. Because there are few pharmacological alternatives for the treatment of this tumor to find other molecules or compounds that could be potential therapeutic agents is desirable. Quercetin is a flavonoid with described antitumoral effect in different types of cancer cell lines that could be a possible option in GCTB treatment. However, there is no literature about the effect of quercetin on GCTB. In the present paper, we reported the ultrastructural changes in GCTB cells exposed to quercetin and also determined the expression of RIP1K, Caspase 3 and Caspase 8 on the exposed cells. For this purpose, GCTB sample was obtained from one patient and cultured. Quercetin affected all the histological components of the GCTB. The ultrastructural changes consisted mainly in necroptosis, autophagocytosis and secondary necrosis. This is the first report about quercetin effects on giant cell tumor of bone cultured cells. Further studies in other models could be done to support the use of quercetin as a complementary treatment in giant cell tumor of bone. Abbreviations: Giant cell tumor of bone (GCTB); transmission electron microscopy (TEM); reverse transcription – polymerase chain reaction (RT-PCR); receptor interacting protein kinase 1 (RIP1K); Dulbecco’s Modified Eagle’s Medium (DMEM).

Background and Objectives: Deposits of monosodium urate (MSU) crystals due to increased levels of uric acid (UA) have been associated with bone formation and erosion, mainly in patients with chronic gout. The synovial membrane (SM) comprises several types of cells, including mesenchymal stem cells (SM-MSCs); however, it is unknown whether UA and MSU induce osteogenesis through SM-MSCs. Materials and Methods: Cultures of SM were immunotyped with CD44, CD69, CD90, CD166, CD105, CD34, and CD45 to identify MSCs. CD90+ cells were isolated by immunomagnetic separation (MACS), colony-forming units (CFU) were identified, and the cells were exposed to UA (3, 6.8, and 9 mg/dL) and MSU crystals (1, 5, and 10 μg/mL) for 3 weeks, and cellular morphological changes were evaluated. IL-1β and IL-6 were determined by ELISA, mineralization was assessed by alizarin red, and the expression of Runx2 was assessed by Western blot. Results: Cells derived from SM and after immunomagnetic separation were positive for CD90 (53 ± 8%) and CD105 (52 ± 18%) antigens, with 53 ± 5 CFU identified. Long-term exposure to SM-MSCs by UA and MSU crystals did not cause morphological damage or affect cell viability, nor were indicators of inflammation detected. Mineralization was observed at doses of 6.8 mg/dL UA and 5 μg/mL MSU crystals; however, the differences were not significant with respect to the control. The highest dose of MSU crystals (10 μg/mL) induced significant Runx2 expression with respect to the control (1.4 times greater) and SM-MSCs cultured in the osteogenic medium. Conclusions: MSU crystals may modulate osteogenic differentiation of SM-MSCs through an increase in Runx2.