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MicroRNA-26b inhibits cell proliferation and cytokine secretion in human RASF cells via the Wnt/GSK-3β/β-catenin pathway.

Sun J, Yan P, Chen Y, Chen Y, Yang J, Xu G, Mao H, Qiu Y - Diagn Pathol (2015)

Bottom Line: Conversely, GSK-3β and CyclinD1 expression levels were markedly higher in the miR-26b inhibitor group compared to Mock and NC group (P < 0.05).Transfection of miR-26b mimics significantly reduced the cell proliferation of RAFLS, compared to the Mock and NC groups, and miR-26b inhibitors increased the proliferative capacity of RAFLS compared to Mock and NC groups (P < 0.05).MiR-26b regulates β-catenin and CyclinD1 levels by inhibiting GSK-3β expression, which in-turn alters the Wnt/GSK-3β/β-catenin pathway to lower RAFLS proliferation and elevate cell apoptosis and the secretion of TNF-α,IL-1β and IL-6 cytokines.

View Article: PubMed Central - PubMed

Affiliation: Nursing Office, Linyi People's Hospital, Linyi, 276000, P. R. China. sunjiling0515@163.com.

ABSTRACT

Background: Rheumatoid arthritis (RA) is a chronic systemic auto- immune disease characterized by joint synovitis. Recent evidence suggests that rheumatoid arthritis synovial fibroblasts (RASFs) promote joint destruction. In this study, we investigated the role of microRNA-26b (miR-26b) in cell proliferation and inflammatory cytokine secretion using patient-derived Rheumatoid arthritis fibroblast-like synoviocyte (RAFLS) to understand pathways influencing rheumatoid arthritis.

Methods: RAFLS were cultured in vitro and transfected with miR-26b mimics (experimental group) and negative sequence (control group). The protein levels of Wnt4, Wnt5ɑ, GSK-3β, CyclinD1, Ser9-GSK-3β and β-catenin were detected by western blot analysis. Tumor Necrosis Factor-ɑ (TNF-ɑ), IL- 1β, and IL-6 levels were quantified by Enzyme-linked Immunosorbent Assay (ELISA). RAFLS proliferation and apoptosis were measured by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide (MTT) assay and flow cytometry, respectively.

Results: GSK-3β and CyclinD1 expression levels were lower in miR-26b mimic group compared to Mock group and negative control (NC) group. Conversely, GSK-3β and CyclinD1 expression levels were markedly higher in the miR-26b inhibitor group compared to Mock and NC group (P < 0.05). Transfection of miR-26b mimics significantly increased the, levels of Ser9-GSK-3β and β-catenin in comparison to Mock and NC groups, while transfection of miR-26b inhibitors showed the opposite effect. In miR-26b mimic group, TNF-α, IL- 1β and IL-6 levels were lower than the Mock and NC groups, while in miR-26b inhibitor group, these cytokine levels were higher than the Mock and NC groups (P < 0.05). Transfection of miR-26b mimics significantly reduced the cell proliferation of RAFLS, compared to the Mock and NC groups, and miR-26b inhibitors increased the proliferative capacity of RAFLS compared to Mock and NC groups (P < 0.05). The miR-26b mimic group exhibited higher RAFLS apoptosis rate compared to Mock and NC group and miR-26b inhibitor group showed significantly lower RAFLS apoptosis rate compared to Mock and NC groups (P < 0.05).

Conclusions: MiR-26b regulates β-catenin and CyclinD1 levels by inhibiting GSK-3β expression, which in-turn alters the Wnt/GSK-3β/β-catenin pathway to lower RAFLS proliferation and elevate cell apoptosis and the secretion of TNF-α,IL-1β and IL-6 cytokines. Therefore, our results show that miR-26B plays a central role in inhibiting the inflammation associated with rheumatoid arthritis.

Virtual slides: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/9063056861547150.

No MeSH data available.


Related in: MedlinePlus

The ELISA results of TNF-ɑ,L- 1βand IL-6 levels, *, the comparison between Mock group and NC group, P < 0.05. NC, negative control
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Fig5: The ELISA results of TNF-ɑ,L- 1βand IL-6 levels, *, the comparison between Mock group and NC group, P < 0.05. NC, negative control

Mentions: TNF-α, IL- 1β and IL-6 levels were quantified by ELISA (Fig. 5). TNF-α, IL- 1β and IL-6 levels showed no statistical differences between the Mock and NC groups (all P > 0.05). However, in the miR-26b mimic group, these cytokine levels were significantly lower than the Mock and NC groups (all P < 0.05). On the other hand, in the miR-26b inhibitor group, the cytokines levels were higher than in the Mock and NC groups (all P < 0.05).Fig. 5


MicroRNA-26b inhibits cell proliferation and cytokine secretion in human RASF cells via the Wnt/GSK-3β/β-catenin pathway.

Sun J, Yan P, Chen Y, Chen Y, Yang J, Xu G, Mao H, Qiu Y - Diagn Pathol (2015)

The ELISA results of TNF-ɑ,L- 1βand IL-6 levels, *, the comparison between Mock group and NC group, P < 0.05. NC, negative control
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4472173&req=5

Fig5: The ELISA results of TNF-ɑ,L- 1βand IL-6 levels, *, the comparison between Mock group and NC group, P < 0.05. NC, negative control
Mentions: TNF-α, IL- 1β and IL-6 levels were quantified by ELISA (Fig. 5). TNF-α, IL- 1β and IL-6 levels showed no statistical differences between the Mock and NC groups (all P > 0.05). However, in the miR-26b mimic group, these cytokine levels were significantly lower than the Mock and NC groups (all P < 0.05). On the other hand, in the miR-26b inhibitor group, the cytokines levels were higher than in the Mock and NC groups (all P < 0.05).Fig. 5

Bottom Line: Conversely, GSK-3β and CyclinD1 expression levels were markedly higher in the miR-26b inhibitor group compared to Mock and NC group (P < 0.05).Transfection of miR-26b mimics significantly reduced the cell proliferation of RAFLS, compared to the Mock and NC groups, and miR-26b inhibitors increased the proliferative capacity of RAFLS compared to Mock and NC groups (P < 0.05).MiR-26b regulates β-catenin and CyclinD1 levels by inhibiting GSK-3β expression, which in-turn alters the Wnt/GSK-3β/β-catenin pathway to lower RAFLS proliferation and elevate cell apoptosis and the secretion of TNF-α,IL-1β and IL-6 cytokines.

View Article: PubMed Central - PubMed

Affiliation: Nursing Office, Linyi People's Hospital, Linyi, 276000, P. R. China. sunjiling0515@163.com.

ABSTRACT

Background: Rheumatoid arthritis (RA) is a chronic systemic auto- immune disease characterized by joint synovitis. Recent evidence suggests that rheumatoid arthritis synovial fibroblasts (RASFs) promote joint destruction. In this study, we investigated the role of microRNA-26b (miR-26b) in cell proliferation and inflammatory cytokine secretion using patient-derived Rheumatoid arthritis fibroblast-like synoviocyte (RAFLS) to understand pathways influencing rheumatoid arthritis.

Methods: RAFLS were cultured in vitro and transfected with miR-26b mimics (experimental group) and negative sequence (control group). The protein levels of Wnt4, Wnt5ɑ, GSK-3β, CyclinD1, Ser9-GSK-3β and β-catenin were detected by western blot analysis. Tumor Necrosis Factor-ɑ (TNF-ɑ), IL- 1β, and IL-6 levels were quantified by Enzyme-linked Immunosorbent Assay (ELISA). RAFLS proliferation and apoptosis were measured by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide (MTT) assay and flow cytometry, respectively.

Results: GSK-3β and CyclinD1 expression levels were lower in miR-26b mimic group compared to Mock group and negative control (NC) group. Conversely, GSK-3β and CyclinD1 expression levels were markedly higher in the miR-26b inhibitor group compared to Mock and NC group (P < 0.05). Transfection of miR-26b mimics significantly increased the, levels of Ser9-GSK-3β and β-catenin in comparison to Mock and NC groups, while transfection of miR-26b inhibitors showed the opposite effect. In miR-26b mimic group, TNF-α, IL- 1β and IL-6 levels were lower than the Mock and NC groups, while in miR-26b inhibitor group, these cytokine levels were higher than the Mock and NC groups (P < 0.05). Transfection of miR-26b mimics significantly reduced the cell proliferation of RAFLS, compared to the Mock and NC groups, and miR-26b inhibitors increased the proliferative capacity of RAFLS compared to Mock and NC groups (P < 0.05). The miR-26b mimic group exhibited higher RAFLS apoptosis rate compared to Mock and NC group and miR-26b inhibitor group showed significantly lower RAFLS apoptosis rate compared to Mock and NC groups (P < 0.05).

Conclusions: MiR-26b regulates β-catenin and CyclinD1 levels by inhibiting GSK-3β expression, which in-turn alters the Wnt/GSK-3β/β-catenin pathway to lower RAFLS proliferation and elevate cell apoptosis and the secretion of TNF-α,IL-1β and IL-6 cytokines. Therefore, our results show that miR-26B plays a central role in inhibiting the inflammation associated with rheumatoid arthritis.

Virtual slides: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/9063056861547150.

No MeSH data available.


Related in: MedlinePlus