Huangqi decoction ameliorated intestinal barrier dysfunction via regulating NF-κB signaling pathway in slow transit constipation model mice

Figure 4 Huangqi decoction treatment inhibited inflammation-related nuclear factor κB pathway and apoptosis in goblet cells. A: Differentially expressed genes (DEGs) of goblet cells between slow transit constipation (STC) and Huangqi decoction (HQD) group. Blue dot: DEGs in STC group; Red dot: DEGs in HQD group. DEGs were defined as those with a P value < 0.05 from the Wilcoxon rank sum test and a log2 fold change > 0.25; B: Gene ontology enrichment in goblet cells in STC and HQD groups. Odds ratio = gene ratio/background ratio; C: Scatterplot shows the correlation between inflammatory response and apoptosis pathway and nuclear factor κB (NF-κB) pathways in the goblet cells; D: Gene set enrichment analysis comparing transcriptomic profiles of goblet cells in STC and HQD group, with positive scores indicating enrichment in the HQD group, and negative scores indicating enrichment in the STC group; E: Spatial expression of the tumor necrosis factor alpha signaling via NF-κB pathway in epithelial cell regions of a control (CON) sample (left), an STC sample (middle), and an HQD sample (right); F: Violin plots showing the tumor necrosis factor alpha signaling via NF-κB pathway score in goblet cells regions from three spatial transcriptome sequencing samples; G: The heatmap shows the top transcription factors with the highest variability in scores for each group; H: Western blot analysis of p-IκB alpha, total IκB alpha, p-NF-κB P65, total NF-κB P65 in the LS174T cells with the treatment of HQD; I: Representative images of the immunohistochemical expression levels of NF-κB P65 in the colonic mucosa of mice from the CON, STC, and HQD groups were shown. The black scale bar represents 50 μm; J: Comparison of the average optical density of NF-κB P65 between the CON, STC, and HQD groups. ^aP < 0.05, STC vs CON; ^bP < 0.01, STC vs HQD; K: Immunocytochemistry imaging showed the nuclear translocation of NF-κB P65 subunit of NF-κB in LS174T cell. White scale bar represents 25 μm. CON: Control group; STC: Group of slow transit constipation model; HQD: Groups of treatment of Huangqi decoction; LPS: Lipopolysaccharide; TNFA: Tumor necrosis factor alpha; GAPDH: Glyceraldehyde-3-phosphate dehydrogenase; HQD-L: HQD low dose; HQD-M: HQD medium dose; HQD-H: HQD high dose.