Cell differentiation regulates intestinal tight junctions

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The major functions of the small intestine include absorbing nutrients from food and acting as a filtration system, selectively allowing the passage of useful substances into the blood stream while preventing the loss of body fluids and the entry of harmful materials. The millions of intestinal cells are held together by junction proteins that work simultaneously as a filter and as a glue. Most intestinal cell are absorptive cells, and it is assumed that cell-cell connections made by this cell type determine the overall porosity of the intestine. However, there are several other types of mostly nonabsorptive cells whose filter properties are completely unknown but may contribute significantly to intestinal porosity, and this project will elucidate the function of those cell types. Tissues made of just one intestinal cell type will be synthesized and tested to determine the unique composition and porosity of filters made by only that cell type. The project will advance the fundamental understanding of how cell differentiation and composition affect the filtering capacity of the vertebrate intestine. The results will be relevant to human health because almost all allergic, inflammatory, and infectious diseases of the intestine allow nonselective movements of materials that can lead to illness and death, but there is no information on whether these diseases uniquely or equally affect the filters made by different cell types. Results will be disseminated via scientific publications, magazine articles, and blogs in public websites. Research, mentoring, and teaching opportunities will be provided to international collaborators, a postdoctoral researcher, graduate, undergraduate and high school students, and local teachers, leveraging programs at the institution that target students from underrepresented groups. Small-intestine stem cells continuously differentiate into absorptive enterocytes as well as secretory enteroendocrine, Paneth, goblet and other cell types that form tight junctions sealing paracellular spaces. The composition and barrier properties of junctions of stem-cell and secretory-cell types are virtually unknown, as permeability can presently only be determined in epithelial sheets composed mainly of enterocytes or of nonphysiological immortalized cells. This project employs innovative techniques that not only control in vitro differentiation of stem cells to produce primary cultures called enteroids comprised mainly of one cell type, but also force dedifferentiation of mature cells to reacquire stem-cell characteristics. This remarkable technical advance enables the planned experiments that test the hypothesis that differentiation of stem cells into absorptive and secretory cells alters the expression and location of tight-junction proteins, resulting in significant, cell-type-dependent variation in paracellular permeability of macromolecules. The role of the major junctional proteins occludin, tricellulin, and zonula occludens 1 in determining cell-type-dependent paracellular composition and permeability will be examined. High school, undergraduate, and graduate students and post-doctoral trainees will participate in this project which applies a modern approach to studies that increase knowledge of how cell differentiation and nonenterocyte junctions contribute to the barrier function of vertebrate epithelia.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.