TNF-α is upregulated in T2DM patients with fracture and promotes the apoptosis of osteoblast cells in vitro in the presence of high glucose.

Fracture healing is regulated by proinflammatory mediators such as tumor necrosis factor-α (TNF-α), which poses influence on the balance between bone formation and remodeling. And the diabetes is thought to contribute to the delayed diabetic fracture healing. In the present study, we examined the promotion to proinflammatory cytokines and chemokines in type 2 diabetes mellitus (T2DM) patients with bone fractures, and then evaluated the promotion to TNF-α by the high glucose treatment in human osteoblast-like MG-63 cells and the regulatory role of the promoted TNF-α on the MG-63 cell apoptosis. It was demonstrated that there were significantly-upregulated high-sensitivity C-reactive protein (hsCRP) TNF-α, IL-1ß, IL-6, IFN-γ-inducible protein 10 (IP-10) and RANTES in T2DM patients with bone fracture. And the promotion to TNF-α and IL-1ß was confirmed in vitro in both mRNA and protein levels in high glucose-treated MG-63 cells. And either TNF-α or high glucose reduced the viability of MG-63 cells, promoted apoptosis and upregulated apoptosis-associated markers, such as released cytochrome c, cleaved caspase 3 and lyzed PARP. Moreover, there was a synergistic effect between TNF-α and high glucose. The viability reduction and the apoptosis induction of MG-63 cells were significantly higher in the group with both TNF-α and high glucose treatments, than in the group with singular TNF-α treatment. In conclusion, our study demonstrated that proinflammatory cytokines and chemokines were promoted in T2DM patients with bone fracture or in osteoblasts by the high glucose stimulation. TNF-α and high glucose synergistically reduced the viability and induced the apoptosis in the osteoblast-like MG-63 cells in vitro. It implies the significant regulatory role of TNF-α in the delayed fracture healing in T2DM.

Overexpression of hsa_circ_0094742 inhibits IL-1ß-induced decline in CHON-001 cell viability by targeting microRNA-127-5p.

Osteoarthritis (OA) is a public health problem that affects 240 million people globally; however, the current treatment options for OA are not effective. Therefore, there is still an urgent need to identify novel strategies to reduce the incidence and progression of OA. The circular RNA hsa_circ_0094742 was reported to be downregulated in patients with OA. However, the underlying mechanism remains unclear. The levels of hsa_circ_0094742 in CHON-001 were detected by reverse transcription quantitative polymerase chain reaction. Moreover, Cell Counting Kit-8 assay and Ki67 staining were used to determine the cell viability. The protein expression of biomarkers was detected by western blot analysis. In addition, the putative downstream target of hsa_circ_0094742 was predicted using the Circinteractome and TargetScan online databases. The putative targeting relationship was verified by dual luciferase reporter assay and fluorescence in situ hybridization. Next, cell apoptosis was determined by Annexin V/PI staining. hsa_circ_0094742 overexpression (OE) inhibited interleukin (IL)-1ß-induced decline in the viability of CHON-001 cells and primary human chondrocytes. Furthermore, IL-1ß-induced alterations in aggrecan, matrix metallopeptidase 13, X-linked inhibitor of apoptosis protein (XIAP), Bax and active caspase 3 were reversed by hsa_circ_0094742 OE. Luciferase reporter assay indicated that miR-127-5p was the downstream target of hsa_circ_0094742, and latexin was the target of miR-127-5p. hsa_circ_0094742 OE inhibited IL-1ß-induced decline in CHON-001 cell viability by targeting miRNA-127-5p. The findings of the present study revealed the biological rational of the use of hsa_circ_0094742 OE as an anti-IL-1ß effector in human chondrocytes. These findings may prompt further research on hsa_circ_0094742 as a potent circRNA target for the treatment of OA.

MiR-671 ameliorates the progression of osteoarthritis in vitro and in vivo.

OBJECTIVES: Expression of miR-671 was reported to be downregulated in articular cartilage of patients with OA compared to healthy individuals, indicating it may serve as potential biomarker for OA. However, the mechanism by which miR-671 regulates the progression of OA remains unclear. Here, we aimed to investigate the role of miR-671 in cartilage from patients with OA. METHODS: The expression of miR-671 and inflammation mediators in cartilage from patients with OA was analyzed by RT-PCR. In vitro, chondrocytes CHON-001 were stimulated with IL-1ß for 24 h for OA model establishment. Protein expression of MMP-13, aggrecan, and collagen II was measured by western blot. In vivo, the severity of OA in mice was determined by histological analysis. RESULTS: We found that the level of miR-671 was downregulated in OA tissues, plasma and IL-1ß treated CHON-001 cells, compared with control. MiR-671 mimics ameliorated IL-1ß-induced proliferation inhibition and apoptosis stimulation, as well as decreased protein levels of collagen II and aggrecan in CHON-001 cells. In vivo study showed miR-671 mimics alleviated the progression of OA in mice. CONCLUSION: These results indicated miR-671 play an important role during the pathogenesis of OA. Therefore, miR-671 may serve as a potential therapeutic target for the treatment of OA.