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标题:miR-93-3p alleviates lipopolysaccharide-induced inflammation and apoptosis in H9c2 cardiomyocytes by inhibiting toll-like receptor 4.
时间:2020-01-14 21:01:47
DOI:10.1016/j.prp.2018.08.024
PMID:30195636
作者:Bi;Tang;Ling
关键词:Apoptosis;Cardiomyocytes;LPS;TLR4;miR-93-3p
摘要:miR-93 is recently recognized to perform anti-inflammatory action in the pathological process of cardiomyocytes dysfunction. However, it remains unclear whether miR-93-3p involves in lipopolysaccharide (LPS)-induced inflammation and apoptosis in H9c2 cells. The present study aimed to investigate the functions of miR-93-3p and its target, toll-like receptor 4 (TLR4), in LPS-stimulated cardiomyocytes.Cell viability was analyzed by CCK-8 assay. AnnexinV-FITC/PI staining and lactate dehydrogenase (LDH) assay were used to evaluate the cell death. The mRNA and protein levels were assayed by RT-qPCR and western blotting, respectively. The targeted gene was predicted by a bioinformatics algorithm and confirmed by a dual luciferase reporter assay.LDH stimulation resulted in the suppression of cell viability and the increase in apoptosis rate, inflammatory cytokines and LDH levels, while inhibition of TLR4 with TAK-242 or overexpression of miR-93-3p dramatically blocked LPS-induced inflammation and apoptosis in cardiomyocytes. Intriguingly, bioinformatics analysis and experimental data suggested that TLR4 was a direct target of miR-93-3p, which could inhibit TLR4 expression by transfected with miR-93-3p mimics or elevate the expression of TLR4 by transfected with miR-93-3p inhibitors. Overexpression of TLR4 carried out an opposite effect to miR-93-3p and positively regulated LPS-induced inflammation and apoptosis in cardiomyocytes.miR-93-3p showed the protective effects against LPS-induced inflammation and apoptosis in cardiomyocytes by inhibiting TLR4 expression.
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目录:
  • miR-93-3p alleviates lipopolysaccharide-induced inflammation and apoptosis in H9c2 cardiomyocytes by inhibiting toll-like receptor 4
    • Introduction
    • Material and methods
      • Cell culture
      • LDH assay
      • Inflammatory cytokine
      • Apoptosis assay
      • Cell transfection and plasmid constructs
      • Luciferase reporter assay
      • Reverse transcription-quantitative polymerase chain reaction (RT-qPCR)
      • Western blotting
      • Statistical analysis
    • Results
      • LPS induces apoptosis and inflammation in H9c2 cells
      • TLR4 is activated in LPS-stimulated H9c2 cells
      • miR-93-3p is suppressed in LPS-stimulated H9c2 cells
      • TLR4 is a direct target of miR-93-3p
      • TLR4 possesses an antagonistic effect to miR-93-3p in LPS-induced inflammation and apoptosis in H9c2 cells
    • Discussion
    • Conclusion
    • Competing interests
    • Acknowledgements
    • References

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