Figures & data
Figure 1. Effect of mepivacaine on miR-183-5p and PDCD4 expression in SH-SY5Y cells. (a) Mepivacaine could decline miR-183-5p level in a concentration-dependent manner. (b) Mepivacaine could augment PDCD4 mRNA expression level in a concentration-dependent manner. (c) We detected miR-183-5p expression in cells to verify the transfection efficiency. (d) PDCD4 mRNA level was measured after cell transfection and mepivacaine (10 mM) treatment. (e) PDCD4 protein level was measured after cell transfection and mepivacaine (10 mM) treatment.*P< 0.05, **P< 0.01 vs. sham group; #P< 0.05, ##P< 0.01, ###P< 0.001 vs. mepivacaine group
Figure 2. MiR-183-5p targeted PDCD4. (a) The forecasted target sequence of PDCD4 on the 3ʹ-UTR of miR-183-5p. Schematic diagrams showed the mutation in its binding site on the UTR. (b) Dual-luciferase reporter assay was conducted to verify the targeting relationship between miR-183-5p and PDCD4 in SH-SY5Y cells. Luciferase activity was decreased by miR-183-5p mimics. WT: wild-type; Mut: mutant. *P< 0.05, **P< 0.01 (* miR-183-5p vs. miR-NC)
Figure 3. Influence of miR-183-5p on apoptosis induced by mepivacaine. (a) Apoptosis was quantified by flow cytometry using propidium iodide exclusion. (b) Hoechst33342/PI double staining to monitor the apoptosis rate. (c) Western blot to further determine C-caspase-3, Bax, and Bcl-2 expression levels. MiR-183-5p mimics and miR-183-5p inhibitor changed the expression level of these proteins.*P< 0.05, **P< 0.01 vs. sham group; #P< 0.05, ##P< 0.01 vs. mepivacaine group
Figure 4. MiR-183-5p attenuated ROS and oxidative damage in SH-SY5Y cells. (a~b) ELISA kit to assay the content of malondialdehyde (MDA) and superoxide dismutase(SOD). (c) Qualitative detection of the steady-state levels of intracellular reactive oxygen species (ROS) by fluorescence microscopy.**P< 0.01, ***P< 0.001 vs. sham group; #P< 0.05, ##P< 0.01 vs. mepivacaine group
Figure 5. Impact of miR-183-5p on the release of inflammatory factors from SH-SY5Y cells. (a~d) Impact of miR-183-5p on the secretion of IL-6, TNF-α, IL-1β, and IL-8 by SH-SY5Y cells. (e) The protein expression of COX-2 and iNOS in the groups. β-actin was used as an invariant control for calculating protein fold changes. ***P< 0.001 vs. Sham group; #P< 0.05, ##P< 0.01, ###P< 0.001 vs. mepivacaine group
Data availability statement
The data used to support the findings in this study are available from the corresponding author upon request.