Mucosa repair mechanisms of Tong-Xie-Yao-Fang mediated by CRH-R2 in murine, dextran sulfate sodium-induced colitis

Figure 1 Inhibition of CRH-R2 signaling aggravates symptoms of DSS-induced colitis in mice. A: Mice body weights measured for 16 d, and shown as percentage of weight change; B: Colon length; C: Representative photographs of colon lengths; D: DAI; E: Histological scores were evaluated on the 16^th day; F: Representative images of hematoxylin-eosin staining histology. Data are presented as mean ± standard deviation, n = 6-10 per group, scale bar = 200 μm. ^aP < 0.001 vs control group; ^gP < 0.05, ^bP < 0.001 vs DSS group; ^cP < 0.001 vs Ucn2 group; ^hP < 0.05 vs TXYF-L group; ^fP < 0.01, ^dP < 0.001 vs TXYF-M group; ⁱP < 0.05, ^eP < 0.001 vs TXYF-H group. CRH-R2: Corticotropin-releasing hormone-receptor 2; DAI: Disease activity index; DSS: Dextran sulfate sodium; TXYF: Tong-Xie-Yao-Fang.

Figure 2 Inhibition of CRH-R2 signaling increases secretion of inflammatory cytokines in colon tissues of DSS-induced ulcerative colitis mice. A-C: Real time-PCR-assessed mRNA level of TNF α (A), CXCL-1 (B) and IL 6 (C) in colon tissues. The mRNA level in each group was determined relative to the level in the control group (defined as 100%); E-F: Enzyme-linked immunosorbent assay-detected protein levels of TNF α (D), CXCL-1 (E) and IL 6 (F) in colon tissues. Data are presented as mean ± standard deviation, n = 6 per group. ^bP < 0.01, ^cP < 0.001 vs control group; ^dP < 0.05, ^eP < 0.01, ^fP < 0.001 vs DSS group; ^qP < 0.01, ^rP < 0.001 vs Ucn2 group; ^mP < 0.05, ⁿP < 0.01, ^oP < 0.001 vs TXYF-L group; ^gP < 0.05, ^hP < 0.01, ⁱP < 0.001 vs TXYF-M group; ^jP < 0.05, ^kP < 0.01, ^lP < 0.001 vs TXYF-H group. CRH-R2: Corticotropin-releasing hormone-receptor 2; DSS: Dextran sulfate sodium; TXYF: Tong-Xie-Yao-Fang.

Figure 3 Inhibition of CRH-R2 signaling promotes intestinal permeability in DSS-induced colitis. The FITC-dextran levels in serum were determined. Data are presented as mean ± standard deviation, n = 6 per group. ^aP < 0.001 vs control group; ^gP < 0.05, ^bP < 0.001 vs DSS group; ^eP < 0.001 vs Ucn2 group; ^fP < 0.001 vs TXYF-L group; ^dP < 0.001 vs TXYF-M group; ^cP < 0.001 vs TXYF-H group. CRH-R2: Corticotropin-releasing hormone-receptor 2; DSS: Dextran sulfate sodium; TXYF: Tong-Xie-Yao-Fang.

Figure 4 Inhibition of CRH-R2 signaling promotes colonic epithelial cell apoptosis in DSS-induced colitis. A-K: Representative images from TUNEL sections; L: Quantification of TUNEL data. Data are presented as mean ± standard deviation, n = 6 per group, scale bar = 50 μm. ^dP < 0.001 vs control group; ^aP < 0.05, ^bP < 0.01, ^cP < 0.001 vs DSS group; ^eP < 0.001 vs Ucn2 group; ^gP < 0.01 vs TXYF-L group; ^hP < 0.01 vs TXYF-M group; ^fP < 0.001 vs TXYF-H group. CRH-R2: Corticotropin-releasing hormone-receptor 2; DSS: Dextran sulfate sodium; TUNEL: Terminal deoxynucleotidyl transferase dUTP nick-end labeling; TXYF: Tong-Xie-Yao-Fang.

Figure 5 Inhibition of CRH-R2 signaling reduces epithelial cell proliferation in DSS-induced colitis. A-K: Representative images from Ki-67 immunoreactive sections; L: Quantification of Ki-67 immunohistochemistry data. Data presented as mean ± standard deviation, n = 6 per group, scale bar = 50 μm. ^cP < 0.05, ^dP < 0.01 vs control group; ^aP < 0.05, ^bP < 0.01 vs DSS group. ^fP < 0.01 vs Ucn2 group; ^eP < 0.05 vs TXYF-L group; ^gP < 0.01 vs TXYF-M group; ^hP < 0.01 vs TXYF-H group. CRH-R2: Corticotropin-releasing hormone-receptor 2; DSS: Dextran sulfate sodium; TXYF: Tong-Xie-Yao-Fang.