Fig 2: Inhibiting miR-204-5p expression regulates nesfatin-1-induced stimulation of IL-1β production in human OASFs. (A and B) miRNA target prediction software was used to identify miRNAs that potentially bind to the IL-1β 3’-UTR plasmid. (C) OASFs were incubated with nesfatin-1 and miR-204-5p levels were examined by qPCR. (D) Levels of miR-204-5p in normal and OA synovial tissues retrieved from the GEO dataset. (E and F) Cells were transfected with miR-204-5p mimic, then stimulated with nesfatin-1. IL-1β expression was examined by qPCR and ELISA. (G) Cells were treated with PI3K, Akt, AP-1 and NF-κB inhibitors, then stimulated with nesfatin-1 prior to qPCR analysis of miR-204-5p levels. *p < 0.05 compared with the control group; #p < 0.05 compared with the nesfatin-1-treated group.

Fig 3: ATG-125 reverses the impaired sucrose-induced insulin signaling in muscle atrophy in young adult mice. (A) Immunohistochemical analysis of p-IGF1R, p-IRS1, p-PI3K, p-AKT, p-mTOR, p-S6K and p-4EBP1 staining in muscle sections are shown. Red arrows highlight the positive staining. Scale bar, 100 µm. (B) Igf1r, Irs1, Pi3k, Akt, mtor, S6k and 4ebp1 mRNA expression levels were determined by reverse transcription-quantitative PCR. Data are presented as the mean ± the standard error of the mean of five independent repeats. *P<0.05 vs. Young adult mice; #P<0.05 vs. Sucrose-induced muscle atrophy mice. p-, phosphorylated; IGF1R, insulin-like growth factor 1 receptor; IRS1, insulin receptor substrate 1; PI3K, phospho-inositide 3-kinase; AKT, protein kinase B; mTOR, mechanistic target of rapamycin; S6K, ribosomal protein S6 kinase; 4EBP1, eukaryotic translation initiation factor 4E-binding protein 1.

Fig 4: The PI3K/Akt pathways mediate nesfatin-1-induced stimulation of IL-1β synthesis. (A–C) OASFs were treated with a PI3K inhibitor (Ly2942002) or Akt inhibitor (Akti), or transfected with p85 or Akt siRNAs, then stimulated with nesfatin-1. IL-1β expression was examined by qPCR and ELISA. (D) Cells were incubated with nesfatin-1 for the indicated time intervals; p85 and Akt phosphorylation was examined by Western blot. (E–F) Quantitative data for p-p85 and p-Akt expression. *p < 0.05 compared with the control group; #p < 0.05 compared with the nesfatin-1-treated group.