For colonoscopy, mice were anesthetized with isoflurane (Viatris, Tokyo, Japan), and feces were removed by injecting saline through a flexible feeding tube (Fuchigami, Kyoto, Japan). A rigid telescope (diameter, 1.9 mm; Smith and Nephew, London, UK) was rectally inserted into the mice for 4 cm using the Olympus CLH-250 Xenon Light Source (Olympus, Tokyo, Japan) as previously described [
19]. During endoscope withdrawal, a video of the colon was recorded using the Olympus Video System OTV-SC2 (Olympus) and TEAC UR-4MD Medical Video Recorder (TEAC, Tokyo, Japan). Then, colonoscopic findings were scored as follows: mucosal thickening (0 = transparent, 1 = moderate, 2 = marked, and 3 = non-transparent), vasculature (0 = normal, 1 = moderate, 2 = marked, and 3 = absent/bleeding), and granularity (0 = none, 1 = moderate, 2 = marked, and 3 = extreme) [
20]. For microscopic examination, colons were removed within 24 h of endoscopy after killing the mice under anesthesia. The colons were cut longitudinally, divided into three sections (proximal, middle, and distal), and fixed in 10% neutral-buffered formalin (FUJIFILM Wako Pure Chemical, Osaka, Japan). The tissues were embedded in paraffin wax and stained with hematoxylin and eosin. Tissue specimens for pathological evaluation were prepared by the Biopathology Institute (Kokuto, Japan). The three sections of colons were scored according to the histological criteria as follows: epithelial hyperplasia (0 = none, 1 = < 1.25-fold increase in epithelial length, 2 = 1.25–1.5-fold increase in epithelial length, 3 = 1.6–2-fold increase in epithelial length, and 4 = > 2-fold increase in epithelial length), goblet cell and crypt loss (0 = none, 1 = < 25%, 2 = 25–50%, 3 = 51–75%, and 4 = 76–100%), and inflammatory infiltration (0 = none, 1 = ≤ 25% increased presence in mucosa, 2 = > 25% increased presence in mucosa, 3 = < 25% infiltration in submucosa, 4 = < 75% infiltration in submucosa, and 5 = > 75% submucosa infiltration and sometimes transmural infiltration) [
21]. Colonoscopy and histology findings were scored by two investigators who were blinded to the treatment.