Poria cocos Attenuated DSS-Induced Ulcerative Colitis via NF-κB Signaling Pathway and Regulating Gut Microbiota

Abstract

Ulcerative colitis (UC), as a chronic inflammatory disease, presents a global public health threat. However, the mechanism of Poria cocos (PC) in treating UC remains unclear. Here, LC-MS/MS was carried out to identify the components of PC. The protective effect of PC against UC was evaluated by disease activity index (DAI), colon length and histological analysis in dextran sulfate sodium (DSS)-induced UC mice. ELISA, qPCR, and Western blot tests were conducted to assess the inflammatory state. Western blotting and immunohistochemistry techniques were employed to evaluate the expression of tight junction proteins. The sequencing of 16S rRNA was utilized for the analysis of gut microbiota regulation. The results showed that a total of fifty-two nutrients and active components were identified in PC. After treatment, PC significantly alleviated UC-associated symptoms including body weight loss, shortened colon, an increase in DAI score, histopathologic lesions. PC also reduced the levels of inflammatory cytokines TNF-α, IL-6, and IL-1β, as evidenced by the suppressed NF-κB pathway, restored the tight junction proteins ZO-1 and Claudin-1 in the colon, and promoted the diversity and abundance of beneficial gut microbiota. Collectively, these findings suggest that PC ameliorates colitis symptoms through the reduction in NF-κB signaling activation to mitigate inflammatory damage, thus repairing the intestinal barrier, and regulating the gut microbiota.

Keywords: Poria cocos; gut microbiota; intestinal barrier function; ulcerative colitis.

Figure 1

PCE relieves DSS-induced colitis in mice. (A) Perianal condition in mice; (B) weight change during the experiment; (C) disease activity index (DAI) score; (D,E) representative images of colon length on d7; (F) spleen coefficients. PCE-L, low dose of PCE; PCE-M, medium dose of PCE; PCE-H, high dose of PCE. The values are expressed as mean ± SD (n = ten mice in each group) ^## p < 0.01, ^### p < 0.001 vs. control group; * p < 0.05, ** p < 0.01, *** p < 0.001 vs. model group.

Figure 2

PCE ameliorates DSS-induced intestinal structural damage in UC mice. (A) Representative H&E staining of colon tissue (200×); (B) histological score of H&E staining; (C) Alician staining of goblet cells (400×); (D) histological score of Alician staining. Red arrow indicates inflammatory damage before and after administration in the blank group and the model group. The values are expressed as mean ± SD (n = 3). ^## p < 0.01, ^### p < 0.001 vs. control group; * p < 0.05, ** p < 0.01, *** p < 0.001 vs. model group.

Figure 3

PCE decreased inflammatory cytokine levels in serum and colon tissue. (A,B) The expressions of TNF-α, IL-6, and IL-1β levels in serum and colonic tissue revealed by ELISA, respectively; (C) the mRNA expressions of TNF-α, IL-1β, and IL-6 were detected by performing qPCR. The values are expressed as the means ± SD (n = 3 of independent experiments in vitro), ^### p < 0.001 vs. control group; * p < 0.05, ** p < 0.01, and *** p < 0.001 vs. DSS group.

Figure 4

PCE inhibits ΙκΒα/ΝF-κΒ activity. (A) Immunoblots of ΙκΒα, P-ΙκΒα, and iNOS in the colon tissue of mice; (B,C) protein expression chart. The values are expressed as mean ± SD (n = 3 of independent experiments in vitro), ^### p < 0.001 vs. control group; * p < 0.05, ** p < 0.01, *** p < 0.001 vs. DSS group.

Figure 5

PCE attenuates DSS-induced damage to the TJs of intestinal epithelium in mice. (A–D) TJ proteins Claudin-1 and ZO-1 were observed by immunohistochemistry; (E–G) the expression of ZO-1, Claudin-1 proteins were determined by Western blotting. Red arrow indicates TJs damage before and after administration in the blank group and the model group. The values are expressed as mean ± SD (n = 3 of independent experiments in vitro), ^## p < 0.01, ^### p < 0.001 vs. control group; * p < 0.05, ** p < 0.01, *** p < 0.001 vs. DSS group.

Figure 6

Intestinal flora analyses in mice. (A) Alpha diversity; (B) abundance curve; (C) principal coordinates analysis; (D) non-metric multidimensional scaling. The values are expressed as mean ± SD. * p < 0.05, ** p < 0.01, and *** p < 0.001 vs. DSS group.

Figure 7

Intestinal microbiota species composition changes in mice. (A) Venn diagram; (B–H) relative abundance comparisons between Proteobacteria, Firmicutes, Pseudomonas, Oscillospira, Prevotella, Ruminococcus, and unidentified-Lachnospiraceae in three groups, respectively. ^# p < 0.05, ^## p < 0.01, and ^### p < 0.001 vs. control group; The values are expressed as mean ± SD, ** p < 0.01, *** p < 0.001 vs. DSS group.