Figure 1. Formation of hyperproliferative crypt nodules in the small intestine of Hsp60^Δ/ΔIEC mice.

(a) Schematic illustration of the targeted genomic modifications to generate mice carrying a conditional knockout allele for Hsp60 (Flox allele). Expression of Cre recombinase and induction of Cre activity by tamoxifen generate the Hsp60 knockout specifically in intestinal epithelial cells (IEC) or intestinal stem cells (ISC). (b) Schedule for oral tamoxifen administration to induce HSP60 deficiency in IEC of adult mice. Agarose gels showing the presence of the knockout allele specifically in IEC isolates. (c) Representative H&E and corresponding HSP60 IHC stainings of Hsp60^flox/flox and Hsp60^Δ/ΔIEC mice along the intestinal tract. Images of HSP60 IHC in higher magnification show HSP60-deficient villus versus HSP60-positive crypt regions of the jejunum. HSP60-positive crypt nodules in Hsp60^Δ/ΔIEC mice were counted along the intestinal tract using HSP60 IHC stainings. The graph represents quantifications of Hsp60^Δ/ΔIEC mice (N=6) with >100 crypts counted per animal. Lines indicate mean numbers. One-way analysis of variance (ANOVA) followed by Dunn's test was used to test for significance. (d) Genomic DNA was isolated from villi and hyperproliferative crypt nodules of Hsp60^Δ/ΔIEC mice using laser-dissection microscopy (N=3). Presence and absence of the Hsp60 knockout allele and Cre transgene was determined via PCR. DNA control PCR were run to check for equal loading (e) Correlation (Pearson) of MT-UPR marker gene expression with Hsp60 mRNA levels in IEC isolated from jejunal fractions of villus tip and crypt bottom of Hsp60^Δ/ΔIEC mice (N=6). P values indicate one-sided significance. Asterisks indicate significant differences *P<0.05, **P<0.01, ***P<0.001; NS, not significant.