Gastroenterology

Gastroenterology

Volume 135, Issue 2, August 2008, Pages 529-538.e1
Gastroenterology

Basic—Alimentary Tract
Toll-Like Receptor Signaling in Small Intestinal Epithelium Promotes B-Cell Recruitment and IgA Production in Lamina Propria

https://doi.org/10.1053/j.gastro.2008.04.020Get rights and content

Background & Aims: Several lines of evidence support a role for Toll-like receptor (TLR) signaling to protect the intestine from pathogenic infection. We hypothesized that TLR signaling at the level of the intestinal epithelium is critical for mucosal immune responses. Methods: We generated transgenic mice that express a constitutively active form of TLR4 in the intestinal epithelium (V-TLR4 mice). Lamina propria cellularity was evaluated by immunostaining and flow cytometry. Immunoglobulin (Ig) A levels in the stool and serum were measured by enzyme-linked immunosorbent assay. Chemokine and cytokine expression were analyzed by quantitative polymerase chain reaction and enzyme-linked immunosorbent assay. Results: V-TLR4 transgenic mice reproduced normally and had a normal life span. Constitutive activity of TLR4 in the intestinal epithelium promoted recruitment of B cells and an increase in fecal IgA levels. Intestinal epithelial cells of V-TLR4 mice expressed higher levels of CCL20 and CCL28, chemokines known to be involved in B-cell recruitment, and of a proliferation-inducing ligand (APRIL), a cytokine that promotes T-cell-independent class switching of B cells to IgA. The changes in B-cell numbers and IgA levels were blocked by simultaneous expression in intestinal epithelial cells of M3, a herpes virus protein that binds and inhibits multiple chemokines. Conclusions: TLR signaling in the intestinal epithelial cells significantly elevated the production of IgA in the intestine. This effect was mediated by TLR-induced expression of a specific set of chemokines and cytokines that promoted both recruitment of B cells into the lamina propria and IgA class switching of B cells.

Section snippets

Mice

Plasmids containing the mouse villin promoter (pBS-Villin)21 and mCD4-hTLR4 fusion gene have been previously described.22 The mCD4-hTLR4 was polymerase chain reaction (PCR) amplified and cloned into pBS-Villin. The sequences were verified by sequencing. The transgene was gel purified, microinjected into fertilized eggs from [C57BL/6J × DBA/2]F2 mice, and the eggs transferred into oviducts of ICR foster mothers. Identification of the transgenic mice was done by PCR amplification using the

Generation of Transgenic Mice Expressing mCD4-hTLR4

To study the biologic effects of TLR signaling in the intestinal epithelium in vivo, we generated transgenic mice expressing a constitutively active form of TLR4 on IECs. The mice carry an mCD4-hTLR4 transgene that consists of the human TLR4 transmembrane and intracellular domains fused to the extracellular domain of murine CD4. This construct has been shown to signal similarly to endogenous TLR4 both in vitro and in vivo.22, 26 The transgene was driven by the villin promoter (Figure 1A), which

Discussion

Here, we report that increased activation of the TLR signaling in IECs induces a remarkable and unsuspected phenotype: the accumulation of large numbers of B cells and IgA+ cells in the intestine. This response reveals an important role for the IEC compartment in response to environmental signals and suggests a novel mechanism for maintenance of intestinal homeostasis. By mediating an increase in luminal IgA in response to TLR activation, IECs may prevent attachment and possibly invasion of

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    Supported by NIH grants P01 DK072201 (to S.A.L. and L.M.), AI052266 (to M.T.A.), and DK069594 (to M.T.A.); Career Development Award from CCFA (to M.F.); Uehara Memorial Foundation Research Fellowship (to M.F.); New York Crohn's Foundation and CCFA (to L.M.).

    Conflicts of interest: There are no conflicts of interest to disclose.

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