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Fig. 3 | Journal of Neuroinflammation

Fig. 3

From: AMPK-autophagy-mediated inhibition of microRNA-30a-5p alleviates morphine tolerance via SOCS3-dependent neuroinflammation suppression

Fig. 3

AMPK activators upregulated the protein level of SOCS3 via the inhibition of microRNA-30a-5p. a BV-2 cells were treated with resveratrol (50 μM), AICAR (300 μM) or metformin (2.5 mM) for 12 h. Cell extracts were collected. The protein level of SOCS3 was evaluated by western blot (n = 3). b BV-2 cells were subjected to compound C (AMPK inhibitor, 20 μM) for 12 h, followed by exposure to metformin (2.5 mM) for 12 h. The protein level of SOCS3 and phosphorylation of AMPK were tested by western blot (n = 3). c BV-2 cells were treated with resveratrol (50 μM), AICAR (300 μM) or metformin (2.5 mM) for 12 h. The mRNA level of Socs3 was evaluated by real-time PCR (n = 3). d Screening based on bioinformatics (http://www.targetscan.org), five miRNAs were candidates that may target SOCS3. BV-2 cells were transfected with 100 pmol miRNA-30a-5p mimic, miRNA-19a-3p mimic, miRNA-203-5p mimic, miRNA-218-5p mimic, miRNA-455-5p mimic or negative control (NC) miRNA for 36 h. The protein level of SOCS3 was evaluated by western blot (n = 3). e BV-2 cells were transfected with 100 pmol miRNA-30a-5p mimic, miRNA-30a-5p inhibitor, negative control mimic or negative control inhibitor for 36 h. Western blot analysis showed the protein level of SOCS3 (n = 3). BV-2 cells were treated with resveratrol (50 μM), AICAR (300 μM) or metformin (2.5 mM) for 12 h. f The level of miRNA-30a-5p was evaluated by real-time PCR (n = 3). g The level of miRNA-30a (primary miRNA-30a-5p) was evaluated by real-time PCR (n = 3). Data were analyzed by one-way ANOVA. *p < .05, **p < .01, ***p < .001 vs. control group. ##p < .01, ###p < .001 vs. metformin-treated group

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