Fig 1: Decreased circPan3 expression, ghrelin synthesis, and autophagy in the rat OA model. (A) Histological alterations of knee joint cartilage tissues in OA and CTL rats. The rat cartilage tissues were subjected to H&E staining. Rats that underwent the sham operation were used as the control group (CTL). (B) Safranin O/fast green staining of knee joint cartilage tissues from both groups. (C) The analysis results of cartilage thickness, chondrocyte density, and OARSI scores in knee joint tissues from both groups. (D) Plasma GHRL, IL-6, and IL-1β levels in both groups. The ELISA method was used to determine the levels of these factors. (E) Western blotting results of MMP13, ADAMTS5, beclin 1, and LC3 in the cartilage tissues of rats from both groups. (F) The expressions of circZfpm1, circSox6, and circPan3 in the knee joint cartilage tissues of rats in the OA model group. The relative expression levels of circRNAs were measured by RT–qPCR. (G) ROC curve analysis of circPan3, circSox6, and ghrelin as potential biomarkers of OA pathogenesis. (H) Bioinformatics predicted that circPan3 was formed from the back-splicing of Pan3 encoded pre-mRNA, which is located at chromosome 12. (I) Validation of the formation of circPan3 in rat chondrocytes. RT–PCR was carried out using divergent and opposite-directed primers and cDNA and gDNA as the templates. (J) Confirmation of the splice joint site of circPan3 in rat chondrocytes was assessed by DNA sanger sequencing. The joint site was indicated by an arrow. CTL, control; OA, osteoarthritis; OARSI, Osteoarthritis Research Society International; GHRL, ghrelin; IL-6, interleukin-6; IL-1β, interleukin-1β; MMP13, matrix metallopeptidase 13; ADAMTS5, ADAM metallopeptidase with thrombospondin type 1 motif 5; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; RT–qPCR, quantitative real-time polymerase chain reaction; RT–PCR, reverse transcription polymerase chain reaction; ELISA, enzyme linked immuneSorbent assay; ROC, receiver operating characteristic; cDNA, complementary DNA; gDNA, genomic DNA. *P < 0.05; **P < 0.01 versus CTL, n = 3.