Background: Many studies have reported that long noncoding RNAs (lncRNAs) could act as sponges for microRNAs (miRNAs) and play important roles in the regulation of osteoarthritis (OA). Yet, the underlying mechanisms of lncRNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) in OA are still unclear. Therefore, we aimed to explore the regulation mechanisms of MALAT1 in OA procession. Methods: IL-1β treatment in chondrocyte was used to mimic OA in vitro. MALAT1, miR-150-5p and AKT3 expression levels were detected via qRT-PCR. The protein levels of AKT3, MMP-13, ADAMTS-5, Bax, Bcl-2, cleaved-PARP, collagen II and aggracan were measured by western blot. MTT assay was performed to detect cell proliferation ability. The apoptosis of chondrocytes was determined using flow cytometry and western blot. Luciferase assay and RNA immunoprecipitation (RIP) assays were used to confirm the relationship among MALAT1, miR-150-5p and AKT3. Results: In our study, MALAT1 and AKT3 were upregulated while miR-150-5p was downregulated in OA in vitro and vivo. The level of miR-150-5p was negatively correlated with that of MALAT1 or AKT3. More importantly, overexpression of MALAT1 promoted the expression of AKT3 by negatively regulating miR-150-5p. MALAT1 knockdown inhibited cell proliferation, promoted apoptosis, increased MMP-13, ADAMTS-5 expression and decreased collagen II, aggracan expression in IL-1β treated chondrocytes. MALAT1 upregulation or AKT3 overexpression enhanced proliferation, inhibited apoptosis and extracellular matrix (ECM) degradation, which was undermined by overexpression of miR-150-5p. By contrast, miR-150-5p depletion rescued the effect of MALAT1 downregulation or loss of AKT3 on IL-1β-stimulated chondrocytes. Conclusion: MALAT1 was responsible for cell proliferation, apoptosis, and ECM degradation via miR-150-5p/AKT3 axis.

BackgroundCartilage tissue engineering is a promising approach for repairing and regenerating cartilage tissue. To date, attempts have been made to construct zonal cartilage that mimics the cartilaginous matrix in different zones. However, little attention has been paid to the chondrocyte density gradient within the articular cartilage. We hypothesized that the chondrocyte density gradient plays an important role in forming the zonal distribution of extracellular matrix (ECM).MethodsIn this study, collagen type II hydrogel/chondrocyte constructs were fabricated using a bioprinter. Three groups were created according to the total cell seeding density in collagen type II pre-gel: Group A, 2 × 107 cells/mL; Group B, 1 × 107 cells/mL; and Group C, 0.5 × 107 cells/mL. Each group included two types of construct: one with a biomimetic chondrocyte density gradient and the other with a single cell density. The constructs were cultured in vitro and harvested at 0, 1, 2, and 3 weeks for cell viability testing, reverse-transcription quantitative PCR (RT-qPCR), biochemical assays, and histological analysis.ResultsWe found that total ECM production was positively correlated with the total cell density in the early culture stage, that the cell density gradient distribution resulted in a gradient distribution of ECM, and that the chondrocytes’ biosynthetic ability was affected by both the total cell density and the cell distribution pattern.ConclusionsOur results suggested that zonal engineered cartilage could be fabricated by bioprinting collagen type II hydrogel constructs with a biomimetic cell density gradient. Both the total cell density and the cell distribution pattern should be optimized to achieve synergistic biological effects.

No abstract

As the connecting tissue between the hyaline articular cartilage and the subchondral bone, calcified cartilage zone (CCZ) plays a great role in the force transmission and materials diffusion. However, the questions that remain to be resolved are its mineral composition and organization. In this study, 40 healthy human knee specimens were harvested; first the CCZ was dissected and observed by Safranin O/fast green staining, then CCZ chemical characteristics were measured by using amino acid assay and X-ray diffraction. The percentage of dry weight of type II collagen as an organic compound of CCZ was 20.16% ± 0.96%, lower than that of the hyaline cartilage layer (61.39% ± 0.38%); the percentage of dry weight of hydroxyapatite as an inorganic compound was 65.09% ± 2.31%, less than that of subchondral bone (85.78% ± 3.42%). Our study provides the accurate data for the reconstruction of the CCZ in vitro and the elucidation of CCZ structure and function.