Figure 4 from Human Colon Cancer–Derived <i>Clostridioides difficile</i> Strains Drive Colonic Tumorigenesis in Mice

Abstract

<p><i>C. difficile</i>–associated tumorigenesis is toxin B (TcdB)–dependent. <b>A,</b> GF <i>Apc<sup>Min/+</sup></i> mice were gavaged with WT or isogenic <i>tcdA<sup>−</sup></i>, <i>tcdB<sup>−</sup></i>, or <i>tcdA<sup>−</sup>tcdB<sup>−</sup></i> TargeTron mutant strains of M7404, an epidemic strain of ribotype 027, either simultaneously with the 3728T isolates or 1 week later. Tumor counts at 8 weeks p.i. were analyzed. Crosses represent mouse deaths. <i>n</i> = 2–7 mice per group, from two independent experiments. <b>B,</b> Summary graph showing colonic tumor induction of all TcdB<sup>−</sup> versus TcdB<sup>+</sup> M7404 strains in combination with the 3728T isolates in GF <i>Apc<sup>Min/+</sup></i> mice. <b>C</b> and <b>D,</b><i>C. difficile</i> stool CFUs and Vero toxin titers. Mann–Whitney <i>P</i> value is shown. <b>E–M,</b> Experimental design for 2-week experiments with M7404 strains: mice were gavaged with the 3728T isolates − Cd 1 week prior to <i>C. difficile</i> gavage with either the <i>tcdB<sup>−</sup></i> (<i>n</i> = 6 mice) or WT M7404 strain (<i>n</i> = 8 mice). <b>F</b> and <b>G,</b><i>C. difficile</i> stool CFUs and toxin titers. <b>H,</b> Representative crypts with BF invasion, stained in red for EUB338 and yellow for Cd198. <b>I,</b><i>C. difficile</i> tissue invasion. <b>J,</b> Representative H&E images and quantitation of histopathologic changes at 2 weeks p.i. <b>K,</b> Representative images of β-catenin, Ki-67, and PCNA immunofluorescent staining of distal colonic epithelium in M7404 TcdB<sup>−</sup> (top)– versus M7404 WT (bottom)–inoculated mice. Arrow denotes a dysplastic focus in the M7404 WT–inoculated mouse. <b>L,</b> Quantitation and representative images of crypt depth in the distal colon. <b>M,</b> Host gene expression in colon. Dashed line represents 3728T isolates − Cd-negative control. Statistics shown are Mann–Whitney <i>t</i> tests. ns, not significant.</p>

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