Fig 2: Effect of EFBS on LPS-induced COX2, iNOS, and NF-?B p65 expression in lung tissue. The ICR mice were left untreated or challenged with LPS (5 mg/kg) for 6 h following no pretreatment or preadministration of 5 mg/kg DEX, 20 mg/kg EFBS, or 60 mg/kg EFBS. All mice were sacrificed 6 h after LPS administration, and lung tissue was collected for western blot analysis with specific antibodies. Data are expressed as mean ± SD (n = 3). *P < 0.05, **P < 0.01 compared with the vehicle treatment group. #P < 0.05, ##P < 0.01 compared with the DEX treatment group.

Fig 3: POG treatment suppresses inflammatory cytokine expression within LPS-treated RAW264.7 cells. (A) The activity of CCK8 cells was determined to affect RAW264.7 cells within the range of POG (12.5, 25, 50, 100,200 µmol/L). (B–D) qRT-PCR extraction of cellular mRNA for detecting IL-1ß, TNF-a, and IL-6 expression. (E–G) iNOS and COX-2 protein levels measured through WB assay. Data were means ± SD (n = 3). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Fig 4: Has_ circ_ 0001326 inhibition reversed the effect of luteolin on macrophages polarization. (a) The expression of has_circ_0001326 was detected using qRT-PCR in different concentration luteolin (10 nm, 100 nm, 1 µm, and 10 µm) treated THP-1 induced macrophages. (b) The expression of has_circ_0001326 was measured using qRT-PCR after siRNAs (si-NC, si-circ-1, si-circ-2, and si-circ-3) transfected into THP-1 induced macrophages. (c) qRT-PCR and (d) Western blot analysis was used to determine the expression of CD11B, INOS, ARG1, and FIZZ1 in CON (THP-1 induced macrophages), si-NC + luteolin (si-NC was transfected into THP-1 induced macrophages for 48 h followed with luteolin treatment for another 48 h), and si-circ + luteolin group (si-circ-3 was transfected into THP-1 induced macrophages for 48 h followed with luteolin treatment for another 48 h), GAPDH was as internal reference. HAS ELISA was used to test the secretion of TNF-a, IL-6, IL-10, and IL-RA in CON, si-NC + luteolin, and si-circ + luteolin group; * indicates the p value less than 0.05.

Fig 5: Validation of transcript expressions by quantitative real-time PCR and western blot among control, NOFD, IR, and IR-NOFD groups(A) The upregulated expression of HK2, ADM, VEGFA, and SLC2A1 in the IR-NOFD group without interfering FIH protein level, indicating the impeded enzymatic activity of FIH by NOFD. (B and C) The upregulated expressions of the 4 hub genes HIF1A, NOS2, EGFR, and CDKN1A (B), and the two lncRNAs, NONMMUT140549.1 and NONMMUT148249.1 (C), in IR-NOFD compared to IR groups by quantitative real-time PCR. (D) The absence of 18S and 28S bands in RNase-R-treated total RNA, indicating the successful degradation of linear RNAs. (E) The upregulated expressions of two circRNAs, circRNA_2909 and circRNA_0323, in IR-NOFD compared to IR groups by quantitative real-time PCR. (F) Validation for degrading linear RNA-HPRT in RNase-R-treated total RNA. (G–I) The upregulated expression of HIF1A, NOS2, EGFR, and CDKN1A in IR-NOFD group compared to IR, without interfering FIH protein level by western blot analysis. A housekeeping gene, HPRT, was used as control for normalization in quantitative real-time PCR. *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. Data are reported as mean ± SEM of three independent experiments.