Loss Of Functional Surface Area Research Articles

Epimorphin regulates the intestinal stem cell niche via effects on the stromal microenvironment.

Stem cell therapy is a potential therapeutic approach for disorders characterized by intestinal injury or loss of functional surface area. Stem cell function and proliferation are mediated by the stem cell niche. Stromal cells such as intestinal subepithelial myofibroblasts (ISEMFs) are important but poorly studied components of the stem cell niche. To examine the role of ISEMFs, we have previously generated mice with deletion of epimorphin ( Epim), an ISEMF protein and member of the syntaxin family of intracellular vesicle docking proteins that regulate cell secretion. Herein we explore the mechanisms for previous observations that Epim deletion increases gut crypt cell proliferation, crypt fission, and small bowel length in vivo. Stem cell-derived crypt culture techniques were used to explore the interaction between enteroids and myofibroblasts from Epim-/- and WT mice. Enteroids cocultured with ISEMFS had increased growth and crypt-like budding compared with enteroids cultured without stromal support. Epim deletion in ISEMFs resulted in increased enteroid budding and surface area compared with cocultures with wild-type (WT) ISEMFs. In primary crypt cultures, Epim-/- enteroids had significantly increased surface area and budding compared with WTs. However, stem cell assays comparing the number of Epim-/- vs. WT colony-forming units after first passage showed no differences in the absence of ISEMF support. Epim-/- vs. WT ISEMFs had increased Wnt4 expression, and addition of Wnt4 to WT cocultures enhanced budding. We conclude that ISEMFs play an important role in the stem cell niche. Epim regulates stem cell proliferation and differentiation via stromal contributions to the niche microenvironment. NEW & NOTEWORTHY The role of subepithelial intestinal myofibroblasts (ISEMFs) in the gut stem cell niche is controversial. We provide novel evidence supporting ISEMFs as important niche contributors. We show that the in vivo intestinal effects of deletion of myofibroblast Epim can be recapitulated in crypt stem cell cultures in vitro. ISEMFs support cocultured stem cell proliferation and enteroid growth, and these effects are augmented by deletion of Epim, a syntaxin that regulates myofibroblast cell secretion.

Increased apoptosis and accelerated epithelial migration following inhibition of hedgehog signaling in adaptive small bowel postresection

The intestinal epithelium undergoes a marked adaptive response following loss of functional small bowel surface area characterized by increased crypt cell proliferation and increased enterocyte migration from crypt to villus tip, resulting in villus hyperplasia and enhanced nutrient absorption. Hedgehog (Hh) signaling plays a critical role in regulating epithelial-mesenchymal interactions during morphogenesis of the embryonic intestine. Our previous studies showed that blocking Hh signaling in neonatal mice results in increased small intestinal epithelial crypt cell proliferation and altered enterocyte fat absorption and morphology. Hh family members are also expressed in the adult intestine, but their role in the mature small bowel is unclear. With the use of a model of intestinal adaptation following partial small bowel resection, the role of Hh signaling in the adult gut was examined by determining the effects of blocking Hh signaling on the regenerative response following loss of functional surface area. Hh-inactivating monoclonal antibodies or control antibodies were administered to mice that sustained a 50% intestinal resection. mRNA analyses of the preoperative ileum by quantitative real-time PCR revealed that Indian hedgehog was the most abundant Hh family member. The Hh receptor Patched was more abundant than Patched 2. Analyses of downstream targets of Hh signaling demonstrated that Gli3 was twofold more abundant than Gli1 and Gli2 and that bone morphogenetic protein (BMP)2 was most highly expressed compared with BMP1, -4, and -7. Following intestinal resection, the expression of Hh, Patched, Gli, and most BMP genes was markedly downregulated in the remnant ileum, and, in anti-Hh antibody-treated mice, expression of Patched 2 and Gli 1 was further suppressed. In Hh antibody-treated mice following resection, the enterocyte migration rate from crypt to villus tip was increased, and by 2 wk postoperation, apoptosis was increased in the adaptive gut. However, crypt cell proliferation, villus height, and crypt depth were not augmented. These data indicate that Hh signaling plays a role in adult gut epithelial homeostasis by regulating epithelial cell migration from crypt to villus tip and by enhancing apoptosis.

Regulation of PC4/TIS7 expression in adapting remnant intestine after resection.

The adaptive response of the small intestine to loss of functional surface area includes enhanced crypt cell proliferation and enterocyte differentiation. To better define the underlying molecular and cellular mechanisms, we have cloned rat genes that are specifically regulated in the adaptive gut after 70% small intestinal resection. One of these is the immediate early gene PC4/TIS7. Compared with sham-resected control ileum, PC4/TIS7 mRNA levels in the adaptive remnant ileum were markedly increased at 16 and 48 h but not 1 wk after resection. Greater augmentation of PC4/TIS7 mRNA levels occurred in the ileum compared with the duodenum and proximal jejunum. After resection, the changes in intestinal PC4/TIS7 mRNA levels also exceeded changes in extraintestinal levels. The demonstration by in situ hybridization that villus-associated, but not crypt, cells express PC4/TIS7 mRNA is consistent with a role in regulating cytodifferentiation. The pattern of expression in the Caco-2 cell line is also consistent with such a role. Although the precise function of PC4/TIS7 in adaptation remains unclear, the early and intestine-specific changes in mRNA levels after 70% resection suggest that it might augment the adaptive response by stimulating the production of differentiated enterocytes.