Insulin-like growth factor 2 and its enterocyte receptor are not required for adaptation in response to massive small bowel resection

Abstract

Purpose: Enhanced structural features of resection-induced intestinal adaptation have been demonstrated following the administration of multiple different growth factors and peptides. Among these, the insulin-like growth factor (IGF) system has been considered to be significant. In this study, we employ mutant mouse strains to directly test the contribution of IGF2 and its enterocyte receptor (IGF1R) toward the adaptation response to massive small bowel resection (SBR).

Methods: IGF2-knockout (IGF2-KO) (n=8) and intestine specific IGF1R-knockout mice (IGF1R-IKO) (n=9) and their wild type (WT) littermates (n=5, n=7, respectively) underwent 50% proximal SBR. At post-operative day 7, structural adaptation was measured as crypt depth and villus height. Rates of enterocyte proliferation and apoptosis were also recorded.

Results: The successful deletion of IGF2 and IGF1R expression in the enterocytes was confirmed by RT-PCR and Western blot, respectively. Normal adaptation occurred in both IGF2-KO and IGF1R-IKO mice after 50% SBR. Post-operative rates of proliferation and apoptosis in both IGF2-KO and IGF1R-IKO mice were no different than their respective controls.

Conclusion: IGF2 and functional IGF1R signaling in enterocytes are both dispensable for resection-induced adaptation responses. The mechanism for IGF-stimulation of intestinal adaptation may involve other ligands or cellular compartments within the intestine.

Keywords: Insulin-like growth factor 1 receptor; Insulin-like growth factor-2; Intestinal adaptation; Short gut syndrome.

Figure 1

Western blotting confirming disrupted IGF1R expression in the crypt enterocytes. Villin Cre-ER (+); IGF1R (f/f) mice and their wild type littermates (Villin Cre-ER- (−)); IGF1R(f/f)were both injected with Tamoxifen for three days prior to resection. Lamin B1 was used as loading control. Successful deletion of IGF1R intestinal knockout was confirmed with all Villin Cre-ER(+); IGF1R (f/f) mice.

Figure 2

Percentage change in crypt depth and villus height for (A) IGF2-KO and (B) IGF1R-intestinal knockout (IKO) mice after small bowel resection. No differences in postoperative villus or crypt growth were observed between the mutant strains or corresponding wild-type (WT) mice in any parameter.

Figure 3

Post-operative rates of crypt cell proliferation in (A) IGF2-KO and (B) IGF1R- intestinal knockout (IKO) compared to their wild type (WT) littermates. BrdU was injected 90 minutes prior to harvest. Immunostaining was performed on tissue sections and BrdU- positive stained cells were counted in each crypt. A ratio was calculated as number of cells stained positive divided by total number of cells in the crypts. A minimum of 20 crypts were counted per mouse. No statistical differences were observed between either mouse strains versus their corresponding WT controls.

Figure 4

Post-operative crypt apoptosis index for (A) IGF2-KO and (B) IGF1R- intestinal knockout (IKO) mice compared to their wild type littermates. H&E stained sections of ileum were analyzed for apoptotic bodies. Apoptotic index was calculated as number of apoptotic bodies found per number of crypts. A minimum of 50 crypts were counted per mouse. There were no statistical differences observed between the mutant mice and their corresponding WT strains.