https://orcid.org/0000-0001-5525-6482
Figures & data
Figure 1. NEAT1 and inflammation levels are elevated in OME
(a) RT-qPCR showed NEAT1 expression in serum and middle ear effusion of OME. (b) ELISA showed the levels of IL-1β, IL-6, IL-8, and TNF-a in serum and middle ear effusion. **P< 0.01, ***P< 0.001.
Figure 2. NEAT1 regulates inflammation, cell proliferation, and apoptosis in LPS-treated HMEECs
(a) RT-qPCR showed NEAT1 expression in LPS-treated HMEEC compared to that in the control group. (b) ELISA showed the levels of IL-1β, IL-6, IL-8, and TNF-a in LPS-treated HMEEC. (c) RT-qPCR showed NEAT1 expression in LPS-treated HMEECs transfected with shNC or shNEAT1. (d) ELISA showed the levels of IL-1β, IL-6, IL-8, and TNF-a in LPS-treated HMEECs transfected with shNC or shNEAT1. (e and f) CCK-8 and flow cytometry assays showed cell proliferation and apoptosis in LPS-stimulated HMEECs transfected with shNC or shNEAT1. *P< 0.05, **P< 0.01, ***P< 0.001.
Figure 3. miR-495 is a target of NEAT1
(a) The binding site between NEAT1 and miR-495 was predicted by starBase website. (b) RT-qPCR analysis showed miR-495 expression in serum and middle ear effusion of OME. (c) RT-qPCR showed miR-495 expression in LPS-induced HMEECs. (d) Luciferase reporter assay was performed to testify the interaction between miR-495 and NEAT1 in HMEECs. (e) RIP assay was performed to determine the enrichment of miR-495 and NEAT1 in anti-IgG and anti-Ago2. (f) RT-qPCR analysis showed miR-495 expression in LPS-treated HMEECs transfected with shNC or shNEAT1. **P< 0.01, ***P< 0.001.
Figure 4. NEAT1 promotes OME progression by targeting miR-495 in HMEECs
(a) ELISA showed the levels of IL-1β, IL-6, IL-8, and TNF-a in LPS-treated HMEECs transfected with shNC, shNEAT1, shNEAT1+ NC inhibitor, and shNEAT1+ miR-495 inhibitor. (b and c) CCK-8 and flow cytometry assays showed cell proliferation and apoptosis in LPS-treated HMEECs transfected with shNC, shNEAT1, shNEAT1+ NC inhibitor, and shNEAT1+ miR-495 inhibitor. *P< 0.05, **P< 0.01, ***P< 0.001.
Figure 5. NEAT1 activates p38 MAPK signaling pathway by targeting miR-495 in LPS-treated HMEECs
(a) Western blot showed the expression of phosphorylated p38 MAPK in LPS-treated HMEECs transfected with pcDNA3.1, pcDNA3.1/NEAT1, pcDNA3.1/NEAT1+ miR-495 mimics. (b–d) ELISA, CCK-8, and flow cytometry assays showed inflammatory cytokine levels, cell proliferation, and apoptosis in LPS-treated HMEECs treated with pcDNA3.1, pcDNA3.1/NEAT1, pcDNA3.1/NEAT1+ SB203580. *P< 0.05, **P< 0.01.
Data availability statement
The datasets used and/or analyzed during the current study are available from the corresponding author upon reasonable request.