2011 - ISBTS 2011 Symposium


Plenary Session II: Physiology & Mucosal Biology + Oral Communications 8

10.206 - Isolation and characterization of human primary enterocytes from small intestine

Presenter: Priti, Chougule, Gothenburg, Sweden
Authors: Priti Chougule1, Gustaf Herlenius1, Suchitra Sumitran-Holgersson1


206
Isolation and characterization of human primary enterocytes from small intestine

Priti Chougule, Gustaf Herlenius, Suchitra Sumitran-Holgersson

University of Gothenburg, Gothenburg, Sweden

Background: Cultures of human enterocytes represent valuable tools to study physiologic and pathophysiology of several inflammatory bowel diseases. Here, our aim was to develop a simple and reproducible culture model for the isolation and characterization of human enterocytes.

Methods: Enterocytes were isolated from samples of small intestine obtained from cadaveric donors using either enzymatic or mechanical procedure, followed by separation with immunomagnetic beads coated with anti-EpCAM antibodies. Light and electron microscopy, flow cytometry and immunocytochemistry techniques were used to characterize the isolated enterocytes. Paraffin embedded small intestine tissue from same donors were also stained for various markers.

Results: Enzymatic and mechanical methods yielded both elongated columnar and spherical cells. Both methods produced a high yield of intestinal epithelial cells with about 88-90 % viability. Magnetic separation of cells using anti-EpCAM antibodies resulted in highly reproducible yields of purified enterocytes. Immunohistochemical staining of normal small bowel biopsies confirmed that the cell cultures maintained an in vivo phenotype as reflected in cytokeratin expression and expression of markers restricted to intestinal enterocytes. Accordingly, cells expressed several markers of enterocyte brush border enzymes (sucrase-isomaltase and maltase glucoamylase). The mechanical method consistently resulted in a good yield of enterocytes with no adverse effect on phenotype composition of the recovered enterocytes or on cell viability.

Conclusions: The cell culture isolation technique and culture model described in the present study provides a unique in vitro system to study the biology of enterocytes in normal conditions as well as to study inflammatory processes in various small bowel disorders.


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