title: Intestinal subepithelial myofibroblasts support in vitro and in vivo growth of human small intestinal epithelium.
abstract: The intestinal crypt-niche interaction is thought to be essential to the function, maintenance, and proliferation of progenitor stem cells found at the bases of intestinal crypts. These stem cells are constantly renewing the intestinal epithelium by sending differentiated cells from the base of the crypts of Lieberkühn to the villus tips where they slough off into the intestinal lumen. The intestinal niche consists of various cell types, extracellular matrix, and growth factors and surrounds the intestinal progenitor cells. There have recently been advances in the understanding of the interactions that regulate the behavior of the intestinal epithelium and there is great interest in methods for isolating and expanding viable intestinal epithelium. However, there is no method to maintain primary human small intestinal epithelium in culture over a prolonged period of time. Similarly no method has been published that describes isolation and support of human intestinal epithelium in an in vivo model. We describe a technique to isolate and maintain human small intestinal epithelium in vitro from surgical specimens. We also describe a novel method to maintain human intestinal epithelium subcutaneously in a mouse model for a prolonged period of time. Our methods require various growth factors and the intimate interaction between intestinal sub-epithelial myofibroblasts (ISEMFs) and the intestinal epithelial cells to support the epithelial in vitro and in vivo growth. Absence of these myofibroblasts precluded successful maintenance of epithelial cell formation and proliferation beyond just a few days, even in the presence of supportive growth factors. We believe that the methods described here can be used to explore the molecular basis of human intestinal stem cell support, maintenance, and growth.
authors: Division of Gastroenterology and Nutrition, Department of Surgery, David Geffen School of Medicine, University of California, Los Angeles, California, United States of America.
authors: Nicholas
authors: N
authors: Lahar
authors: Nan Ye
authors: NY
authors: Lei
authors: Jiafang
authors: J
authors: Wang
authors: Ziyad
authors: Z
authors: Jabaji
authors: Stephaine C
authors: SC
authors: Tung
authors: Vaidehi
authors: V
authors: Joshi
authors: Michael
authors: M
authors: Lewis
authors: Matthias
authors: M
authors: Stelzner
authors: Martín G
authors: MG
authors: Martín
authors: James C Y
authors: JC
authors: Dunn
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D018920:Coculture Techniques; D020895:Enterocytes / Q000166:cytology* / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology* / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D007150:Immunohistochemistry; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology; D007413:Intestinal Mucosa / Q000166:cytology* / Q000254:growth & development / Q000378:metabolism; D007421:Intestine, Small / Q000166:cytology / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D016688:Mice, Inbred NOD; D018345:Mice, Knockout; D016513:Mice, SCID; D009130:Muscle, Smooth / Q000737:chemistry; D058628:Myofibroblasts / Q000166:cytology* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014746:Vimentin / Q000235:genetics / Q000378:metabolism
chemical_list: D000199:Actins; D036341:Intercellular Signaling Peptides and Proteins; D014746:Vimentin