A new organotypic model to study cell interactions in the intestinal mucosa.
BACKGROUND
Recently, we demonstrated that freshly elutriated monocytes differentiate into macrophages with a phenotype similar to that of intestinal macrophages in a three-dimensional model of intestinal epithelial cells. Here we describe a more organotypic model to study cell interactions in the intestinal mucosa.


METHODS
Primary intestinal fibroblasts and freshly elutriated blood monocytes (ratio 1:1) were embedded in collagen type I gels and cultured for 5 days. At day 5, intestinal epithelial cells (HT-29) were seeded on top of the collagen gels. After another 7 days collagen gels were harvested and fixed for immunohistochemical analysis. Cryosections of the aggregates were prepared and staining for monocyte/macrophage markers and basement membrane compounds was performed. Cell interactions inside the aggregates were examined by electron microscopy.


RESULTS
Intestinal fibroblasts contracted the collagen gels which formed stable three-dimensional aggregates within the first 5 days of culture. Intestinal epithelial cells formed a monolayer on top of the gels about 3 days after seeding. Intestinal fibroblasts were distributed randomly over the aggregate. Monocytes inside aggregates were localized in the vicinity to epithelial cells by positive staining for CD68. Typical monocyte/macrophage specific markers such as CD14, CD16, CD11b, CD11c and the co-stimulatory molecules CD80 and CD86 were down-regulated or not detectable on these cells after co-culture in three-dimensional aggregates. Omission of epithelial cells from the model was followed by impaired differentiation of intestinal macrophages.


CONCLUSIONS
In the three-dimensional organotypic cell culture model monocytes differentiate into intestinal-like macrophages when co-cultured with control intestinal fibroblasts and intestinal epithelial cells. Intestinal epithelial cells may be necessary for differentiation of intestinal macrophages.
Spottl
Tanja
T
Department of Internal Medicine I, University of Regensburg, Germany.
Hausmann
Martin
M
Gunckel
Manuela
M
Herfarth
Hans
H
Herlyn
Meenhard
M
Schoelmerich
Juergen
J
Rogler
Gerhard
G
D002450:Cell Communication / Q000502:physiology*; D018929:Cell Culture Techniques; D024042:Collagen Type I; D004847:Epithelial Cells / Q000502:physiology*; D005347:Fibroblasts / Q000502:physiology*; D019073:HT29 Cells; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology*; D008954:Models, Biological; D009000:Monocytes / Q000502:physiology*
D024042:Collagen Type I