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. 2023 Oct 7;13:16920. doi: 10.1038/s41598-023-44158-8

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© This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Colon neoplasia is attenuated in βPix-deficient mice. (a) Treatment scheme for azoxymethane (AOM)/dextran sodium sulfate (DSS)-induced colon neoplasia in mice. Colon neoplasia was induced with four weekly intraperitoneal injections of 7.5 mg AOM/kg mouse body weight. One week after the last dose of AOM, 2.5% DSS was added to the animals’ drinking water for five days. In some experiments, bethanechol (400 µg) was added to the animals’ drinking water for 15 weeks after AOM/DSS treatment. Mice were euthanized 20 weeks after the first AOM injection and their colons were harvested for analysis. (b) Mouse body weights over the 20-week treatment period. Arhgef7 CKO and littermate control mouse weights were similar, including an approximately four-week period of weight loss associated with DSS treatment. Regardless of genotype, female mice weighed approximately 20% less than male mice. (c) Colon tumor numbers were reduced in Arhgef7 CKO mice. CKO mice (n = 30) had fewer colon tumors than control mice (n = 32) (p = 0.04). Each symbol represents the number of colon tumors detected in one mouse. Bars represent means ± SE. (d) Reduced numbers of adenocarcinomas in AOM/DSS-treated Arhgef7 CKO mice. Bar graph shows 24% fewer adenomas and 55% fewer adenocarcinomas in CKO compared to control mice. (p = 0.04). (e) Colon tumor numbers were reduced in Arhgef7 CKO mice treated with a MR agonist. After AOM/DSS treatment, bethanechol (400 µg/ml) was added to the animals’ drinking water for 15 weeks. CKO mice (n = 18) had substantially fewer colon tumors than littermate control mice (n = 20) (p = 0.03). Each symbol represents the number of colon tumors detected in one mouse. Bars represent means ± SE. (f) Expression of colon cancer-related genes was measured in adenocarcinomas harvested from 10 AOM/DSS-treated Arhgef7 CKO and 14 littermate control mice. Each symbol represents the mRNA level in an adenocarcinoma from one mouse relative to the mean values for that gene in control mice (arbitrarily set at 1). Bars represent means ± SE. (g) Representative immunoblots show paired expression of colon cancer-related proteins in normal colon and colon adenocarcinomas harvested from AOM/DSS-treated littermate control (n = 6) and Arhgef7 CKO (n = 6) mice. Original uncut immunoblots are shown in Supplemental Materials.