Fig 1: Quantitative proteomic and biochemical analyses identify the differentially regulated proteins by anemoside B4 upon TNBS-induced colitis colon tissue from rats. Rats were orally administrated with 80 mg/kg TNBS. Anemoside B4 was continuously injected i.p at the doses of 10 mg/kg twice a day for 8 days. Rats were sacrificed on day 8 after colitis induction. a The volcano plot for the MS identified 604 proteins in colon tissue. Each point showed the log2 (Fold change) in the x axis vs. their corresponding -log10 (P-value) in the y axis. The filter boundaries were set so that a fold change above 1.5, or a fold change below 0.67, were considered significant, and a P-value of 0.05 was used as the cutoff value. The horizontal dotted line indicates the cutoff P-value of 0.05 and all points above this line have P < 0.05. The black points indicate 591 quantified unaltered proteins. The green points and red points indicated the down-regulated and up-regulated proteins, respectively. The circle indicated the proteins used for the biochemical validation. b The biological processes of the differentially regulated proteins analyzed by the online STRING database. c Quantification of S100A9 mRNA expression. d Western blotting analysis of S100A9 protein level. c, d Data were shown as mean ± SD (n = 3). ##: P < 0.001 vs. vehicle group; ***: P < 0.001 vs. TNBS group. (E) Proteins in the downstream or upstream of S100A9 signaling pathway were detected by Western blotting in colon tissues (n = 3). f Analysis of S100A9 protein level. #: P < 0.05, ##: P < 0.01, ###: P < 0.001 vs. control group. **: P < 0.01 vs. TNBS group
Fig 2: Anemoside B4 attenuates the effect of S100A9 recombinant protein on the phosphorylation and activation of NF-κB and MAPKs in Caco-2 cells. Human Caco-2 cells were treated with S100A9 recombinant protein (1 µg/mL) for 1 h, and cell were treated with anemoside B4 for 6 h. a Phosphorylation of p65, ERK1/2, JNK1/2 and p38 MAPK and total proteins were analyzed by Western blotting. b Analysis of S100A9 and its downstream protein level. Data were shown as mean ± SD (n = 3). Each experiment was repeated for three times. #: P < 0.05 vs. control group. *: P < 0.05 vs. S100A9 group
Fig 3: Anemoside B4 attenuates the recruitment of inflammatory cells to colon. a Sections of colon tissue were immunostained with FITC-S100A9 (red) and FITC-CD11b (green) and the immunofluorescence was detected by confocal laser-scanning microscope. Shown were data from three rats in one experiment (n = 3 per group). (B) MPO in colon tissues (U/g) was determined by ELISA kits. Data were shown as mean ± SD (n = 3)
Fig 4: The expression of S100A9 and its relationship to tissue remodeling were investigated using tissues from patients with chronic rhinosinusitis (CRS), with a focus on the clinical implications. The ethmoid mucosa from the control and CRS without nasal polyp (CRSsNP) groups, as well as the nasal polyp tissue from the CRS with nasal polyp (CRSwNP) group, were analyzed. (A) Numerous inflammatory cells, including neutrophils and eosinophils, were observed in hematoxylin and eosin (H&E) stained specimen in patients with CRSwNP. (B) Immunohistochemistry shows a stronger expression of S100A9 in patients with CRSwNP than in control or those with CRSsNP. (C) Within the CRSwNP, tissues with squamous metaplasia showed more prominent expression of S100A9 than non‐squamous metaplasia tissues. (D) Multiplex immunofluorescence was performed on nasal polyps from CRSwNP and control ethmoid tissue. The expression of small proline‐rich protein 2A (SPRR2A) and matrix metalloproteinase 1 (MMP1) in the epithelial tissue remodeling lesions showed findings consistent with the expression of pan‐cytokeratin, an epithelial marker, and CD44, an epithelial–mesenchymal transition marker. In addition, the expression of S100A9 was also observed in squamous epithelium. (E) The relationship between myeloperoxidase (MPO), a neutrophil marker, and S100A9 in the epithelial and sub‐epithelial layers was analyzed in a high‐power field based on the basement membrane (dotted line). (F) In the squamous epithelium, more number of cells expressing only S100A9 were compared to those co‐expressing MPO. (G) The expression of S100A9 in the sub‐epithelial tissue was mostly consistent with MPO. (H) As S100A9 expression increased in the tissue, the clinical symptoms score (sino‐nasal outcome test; SNOT‐22) in patients with CRS became more severe. (I) Furthermore, the expression of S100A9 was found to be proportional to the Lund–Mackay computed tomography (CT) score, which indicates the severity of CRS. IHC, immunohistochemistry; SM, squamous metaplasia. * p < 0.05; ** p < 0.01; **** p < 0.0001.
Fig 5: Disease and biofunctions. (A) The top 20 diseases and biofunctions showed that the effect of S100A9 on human nasal epithelial cells was related to seven cellular movements and five dermatological diseases. It was also associated with three inflammatory diseases and two infectious diseases. (B) A graph showing 20 detailed items, p‐value, and the number of related molecules. The item about dermatological diseases was related to squamous metaplasia.
Supplier Page from Abcam for Recombinant Human S100A9 protein (His tag C-Terminus)