Permeability measurement of macromolecules and assessment of mucosal antigen sampling using in vitro converted M cells.
BACKGROUND
M cells are located in the epithelial layer covering the gut-associated lymphoid tissue and are responsible for delivery of macromolecules and microorganisms to the underlying lymphoid cells. It has been shown that the human colonic cell line Caco-2 can be converted to M cells in vitro following coculture with isolated lymphocytes from murine Peyer's patches. Studies were undertaken to evaluate and characterize the transepithelial transport of select macromolecules across these in vitro derived M cells.


METHODS
Caco-2 cells were converted to M cells as reported previously. The morphology of Caco-2 cells and M cells was compared by transmission electron microscopy (TEM). The transport properties of macromolecules such as horseradish peroxidase, FITC-conjugated polystyrene beads, and radiolabeled dextrans were examined. The activation of murine antigen-specific T cells following transport of the antigen ovalbumin across the M-cell barrier was assessed by measuring cytokine production.


RESULTS
M cells were shown to be irregular in shape and have fewer and shorter microvilli compared to the Caco-2 cell progenitors. These cells were still able to form tight junctions and monolayers on polycarbonate membranes. Time-course studies demonstrated that the transport of polystyrene beads and large-molecular-weight dextrans at physiological temperature across M-cell-containing monolayers was size dependent and more rapid than across Caco-2 cell monolayers. The transport of dextrans was also shown to be temperature and concentration dependent. Befitting the role of the M cell in mucosal defense, protein antigen could be delivered by these cells in order to be processed and presented to antigen-specific CD4+ T lymphocytes.


CONCLUSIONS
The M-cell permeability model is a functional and practical system for evaluating the transport properties of macromolecules and assessing the potential for intestinal mucosal antigen sampling to elicit immunological responses.
Liang
E
E
Pharmaceutics Department, Boehringer Ingelheim Pharmaceuticals, Inc., PO Box 368, 900 Ridgebury Road, Ridgefield, CT 06877, USA.
Kabcenell
A K
AK
Coleman
J R
JR
Robson
J
J
Ruffles
R
R
Yazdanian
M
M
D000818:Animals; D000941:Antigens / Q000378:metabolism*; D018938:Caco-2 Cells; D004847:Epithelial Cells / Q000378:metabolism*; D005260:Female; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism* / Q000648:ultrastructure; D046911:Macromolecular Substances; D051379:Mice; D008807:Mice, Inbred BALB C; D010539:Permeability / Q000187:drug effects; D051381:Rats
D000941:Antigens; D046911:Macromolecular Substances