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The Journal of Immunology, 2006, 176: 2465-2469.
Copyright © 2006 by The American Association of Immunologists

Transepithelial Pathogen Uptake into the Small Intestinal Lamina Propria1

Alexandra Vallon-Eberhard2,*,{dagger}, Limor Landsman2,*, Nir Yogev*, Bernard Verrier{dagger} and Steffen Jung3,*

* Department of Immunology, The Weizmann Institute of Science, Rehovot, Israel; and {dagger} Formation de Recherche en Evolution 2736, Centre National de la Recherche Scientifique, BioMérieux, Institut Fédératif de Recherche 128 Biosciences Lyon Gerland, Lyon, France


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
The lamina propria that underlies and stabilizes the gut lining epithelium is densely populated with strategically located mononuclear phagocytes. Collectively, these lamina propria macrophages and dendritic cells (DC) are believed to be crucial for tissue homeostasis as well as the innate and adaptive host defense. Lamina propria DC were recently shown to gain direct access to the intestinal lumen by virtue of epithelium-penetrating dendrites. However, the role of these structures in pathogen uptake remains under debate. In this study, we report that entry of a noninvasive model pathogen (Aspergillus fumigatus conidia) into the murine small intestinal lamina propria persists in the absence of either transepithelial dendrites or lamina propria DC and macrophages. Our results suggest the existence of multiple pathogen entry pathways and point at the importance of villus M cells in the uptake of gut lumen Ags. Interestingly, transepithelial dendrites seem altogether absent from the small intestine of BALB/c mice suggesting that the function of lamina propria DC extensions resides in their potential selectivity for luminal Ags, rather than in general uptake or gut homeostasis.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
The surface of the gastrointestinal tract must protect the organism from harmful microorganisms while retaining permeability for nutrient absorption. The gut lumen-lining epithelium forms an efficient physical barrier. Exposure of the host to bacterial symbionts, commensals, and pathogens is further prevented through a number of nonspecific innate defense mechanisms. Local mucus production and a thick glycocalix reduce direct contact of the epithelium with the gut microflora and form a highly degradative microenvironment. Protection is also upheld through steady-state secretion of a broad spectrum of antimicrobial agents, such as {alpha}-defensins and secretory IgA Abs (1). Intestinal homeostasis is established and shaped by controlled host interaction with the commensal microflora through pathways allowing sensing and sampling of the gut content (2). The latter are likely to be also of crucial importance for immune protection against potential pathogens.

The luminal surface of the gut is lined with polarized, columnar epithelial cells. Joined by tight junctions dividing the apical and basolateral domains of neighboring cells, this single-cell layer restricts particle entry. Besides this barrier function, epithelial cells are actively involved in sensing of the gut microflora as indicated by prominent expression of membrane-bound and cytoplasmic "surveillance" proteins of the TLR and NOD/caspase recruitment domain superfamily (3). Triggering of these sensors potentially results in secretion of effector molecules and basolateral display of receptor ligands, such as CX3CL1, thereby transducing information to underlying lamina propria cells (4, 5). Surveillance of the gut content is further supported by a distinctive type of epithelial cell, the so-called M cell, that fenestrates the epithelial layer and is specialized in transepithelial uptake of particulate and soluble molecules. M cell formation was reported to depend on lymphoepithelial cross-talk (6) and M cells are believed to remain intimately associated with the mucosal-associated lymphoid tissue. Accordingly, they were reported to be restricted to follicle-associated epithelium that colocalize with unique small intestinal lymphoid structures, the Peyer’s patches. More recently, however, M cells were shown to be more widely distributed in the small intestine (7), suggesting extended exposure of the epithelium underlying lamina propria to the luminal content. A third mechanism that potentially enables immunosurveillance of the intestinal content is based on a cellular component of the immune system itself, the dendritic cells (DC).4 These cells constitute a major cellular component of the intestinal lamina propria and have been shown to penetrate epithelial tight junctions by virtue of extended dendrites (8). The epithelial barrier integrity is retained through expression of tight junction proteins by the lamina propria DC (lpDC) (8). Transgenic mice harboring green fluorescent lpDC (CX3CR1GFP mice) allowed us recently to demonstrate the existence of transepithelial dendrites in live intestinal tissue (4). Although these extensions enable lpDC to sense the content of the intestinal lumen, their role in pathogen uptake remains under debate.

In this study, we report lamina propria entry of a noninvasive particulate pathogen, i.e., conidia of the fungus Aspergillus fumigatus, in the absence of transepithelial dendrites. Furthermore, we show that pathogen uptake persists after conditional in situ depletion of all lamina propria mononuclear phagocytes, including CD11c+CX3CR1+ DCs (lpDC) and CD11c+CX3CR1– macrophages (lpM{phi}). Together with the observation that transepithelial lpDC extensions seem absent from the small intestine of BALB/c mice, these findings argue against a general role of transepithelial dendrites in the maintenance of intestinal homeostasis, but suggest the existence of multiple pathogen entry routes.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Disclosures
 References
 
Mice

This study involved the use of CD11c-diphtheria toxin receptor (DTR) transgenic mice (B6.FVB-Tg(Itgax-DTR/GFP)57Lan/J) (9) and CX3CR1GFP mice (10). C57BL/6 and BALB/c mice carrying the CX3CR1GFP allele were backcrossed >10 generations on the respective background. All mice were maintained under specific pathogen-free conditions and handled under protocols approved by The Weizmann Institute Animal Care Committee according to international guidelines.

Transformation of Aspergillus fumigatus and conidia isolation

A. fumigatus conidia (strain CBS 144.89) were provided by J.-P. Latgé (Aspergillus Unit, Pasteur Institute, Paris, France). Transformation was performed by electroporation using inflated spores. Briefly, conidia were incubated in yeast extract glucose 1% medium for 4 h at 37°C, washed, resuspended in yeast extract glucose 1% HEPES 20 mM (pH 8), and incubated for 1 h at 30°C. After centrifugation, spores were resuspended in electroporation buffer (10 mM Tris (pH 7.5), 270 mM sucrose, 1 mM lithium acetate). Plasmids (pAN7-1, pPgpd-DsRed) harboring a hygB resistance gene and DsRed gene, respectively (11) (provided by L. Mikkelsen, The Royal Veterinary and Agricultural University, Frederiksberg, Denmark) were added and electroporation was carried according the following conditions: 1 kV, 400 {omega}, 25 µF, in a 0.2-mm cuvette. Transformed spores were incubated for 15 min on ice in 1 ml of yeast extract glucose 1% medium, plated in minimum medium supplemented with 200 µg/ml hygromycin B (Invitrogen Life Technologies) and incubated at room temperature. For monosporic isolation, A. fumigatus spores were plated in sterilized water/Tween 20 0.1% at a final concentration of 10 spores/100 µl on malt agar 2% plates overlaid with 200 µg/ml hygromycin B. Single colonies were transferred to a new plate to allow production of conidia during 1wk at room temperature. The conidia stock was kept in sterile water at 4°C.

Isolation of small intestinal mononuclear phagocytes

Isolation of lamina propria cells was performed as previously described (4) with modifications. Briefly, following isolation from Rag–/–CX3CR1GFP/+ mice, small intestine was inverted on a polyethylene tube (diameter 2.08 mm), incubated with RPMI 1640 for 30 min at 37°C to loosen the epithelium, washed, and treated with 1 mM DTT (Sigma-Aldrich) for 10 min at room temperature. After a wash, tissue was incubated twice with EDTA 30 mM for 10 min at room temperature, washed, and digested with 36 U/ml collagenase IV (Sigma-Aldrich) and 2000 U/ml DNase I (Roche) in PBS for 1 h at 37°C. The digested tissue was then sheared by repeated pipetting to release the lamina propria cells. The resulting cell suspension was passed through a mesh (70-µm nylon) and analyzed.

Pathogen uptake assay

Mice were sacrificed and 2- to 3-cm sections spanning the small intestinal terminal ileum were cut out, ligated at both ends, and placed in a petri dish. Pathogens were injected with a syringe into the loop and incubated for 10 min at 37°C in RPMI 1640. The loop was subsequently opened at both ends and extensively washed with PBS. This study involved two pathogens: DsRed-transformed A. fumigatus conidia and Salmonella typhimurium strain CS093 (provided by A. Porgador, Ben Gurion University, Beer Sheva, Israel). Conidia were added at a final concentration of 107 conidia/ml and 2 x 108 S. typhimurium bacteria/ml. Bacteria were used in their late logarithmic phase of growth, by diluting a colony in Luria broth containing 0.3 M NaCl and incubating overnight at 37°C.

FACS analysis

Fluorochrome-labeled mAbs were purchased from BD Pharmingen or eBioscience and used according to the manufacturer’s protocols. CX3CR1 staining was performed as previously described using an fractalkine (FKN)-Fc fusion protein (provided by Millennium Biotherapeutics) (10). Cells were analyzed on a FACSCalibur cytometer (BD Biosciences) using CellQuest software (BD Biosciences).

Staining and microscopy of the small intestinal mucosa

After pathogen or mock challenge, ligated small intestinal loops were opened at their end, intensively flushed with PBS, opened by longitudinal incision, and rinsed again. For M cell staining, the cut tissue was fixed in 4% paraformaldehyde for 1 h on ice, incubated with biotin-conjugated anti-Ulex europaeus agglutinin (UEA-1; Vector Laboratories) at 20 µg/ml for 2 h on ice followed by staining with streptavidin-allophycocyanin. Living tissues were imaged with a Zeiss Axioskop II fluorescent microscope for three-color imaging. Image acquisition was conducted with Simple PCI software.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Disclosures
 References
 
The murine small intestinal lamina propria harbors two distinct mononuclear phagocyte populations

To characterize the mononuclear phagocyte content of the murine small intestinal lamina propria, we isolated tissue samples from the small bowel, prepared single-cell suspensions, and analyzed them by flow cytometry. Taking advantage of a mouse strain that harbors a targeted replacement of the gene encoding the chemokine receptor CX3CR1 by a reporter gene encoding GFP (10), we previously showed that the murine intestine harbors a distinct population of CD11c+CD11b+ DC that expresses CX3CR1 (4). Flow cytometric analysis of unfractioned small intestinal single-cell suspensions revealed the presence of additional CD11c+ mononuclear phagocytes that lack CX3CR1 expression (Fig. 1). The two CD11c+ populations differ with respect to maturation and activation markers, such as CD80, MHC class II, and the integrin CD11b, which show higher expression levels on the CX3CR1-expressing cells. CX3CR1+ lpDC are further characterized by expression of the DC marker 33D1 (12) and high level expression of the {alpha}E2 integrin CD103, recently reported for rat intestinal DC (13). The classical macrophage marker F4/80 (14) did not consistently discriminate between the two populations (data not shown). However, absence of CD14, the primary receptor for LPS, on the CX3CR1-negative CD11c+ cell population suggests that these cells correspond to a population of lpM{phi}, as described by Smythies et al. (15) for the human intestine. Interestingly, this mononuclear phagocyte dichotomy seems to be a characteristic general feature of the lamina propria because CD11c+ lung mononuclear phagocytes can also be discriminated on the basis of CD11b and CX3CR1 expression (16, 17) (L. Landsman, unpublished observation). Accordingly, we propose to refer to the CD11c+CD11b+CX3CR1+ population as lpDC and the CD11c+CD11b–CX3CR1– cells as lpM{phi}. Proximal and distal sections of the small intestine vary considerably in the microbial load. However, we did not observe significant differences between lamina propria cells isolated from the duodenum/jejunum and terminal ileum, for the markers analyzed (data not shown).


Figure 1
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  		FIGURE 1. Phenotypic dichotomy of lamina propria mononuclear phagocytes. Isolated small intestinal lamina propria cells of Rag–/– CX3CR1GFP/+ mice were stained with Abs for CD11c and one of the indicated surface markers. CX3CR1 was stained with FKN-Fc. Expression of surface markers of gated populations is shown in the right histograms. Gray-filled histograms: CD11c+ CX3CR1/GFP– cells (lpM{phi}) (Gate R1), open histograms: CD11c+ CX3CR1/GFP+ (lpDC) (Gate R2).

 
Neither transepithelial dendrites nor CD11c+ lamina propria cells are required for quantitative pathogen uptake

C57BL/6 CX3CR1GFP mice allow the in situ detection of transepithelial dendrites in intact intestinal tissue (4). The latter allow strategically located lpDC to sense the content of the intestinal lumen. Supporting their role in immunosurveillance, lpDC extensions are restricted to the terminal ileum (4), a region characterized by increased bacterial load, a unique microflora as well as activated epithelium and lamina propria, suggesting constant exposure to bacterial stimuli (4, 18, 19). Furthermore, formation of transepithelial dendrites can be induced by challenge of intestinal villi with commensal and enteropathogenic bacteria (4). However, a direct role of lpDC and trans-epithelial dendrites in pathogen sampling, as originally suggested by Rescigno et al. (20) remains controversial (21, 22). Challenging the general accessibility of the lamina propria, Macpherson and Uhr (23) failed to detect bacteria-harboring lpDC after gavage with GFP-expressing Escherichia coli. In contrast, we reported that orally administered invasive S. typhimurium entered the lamina propria even in absence of transepithelial dendrites, while entry of a noninvasive Salmonella mutant to the lamina propria was CX3CR1-dependent (4). To directly study dendrite/pathogen interactions at the intestinal surface, we investigated the potential of the noninvasive fungal pathogen, Aspergillus fumigatus, to induce transepithelial dendrites and enter the lamina propria. Both Aspergillus hyphae and conidia have been reported to interact with DC involving receptors of the IL-1R/TLR superfamily and C-type lectins (24, 25). To visualize the pathogen, we generated red fluorescent conidia by transforming A. fumigatus with an expression vector encoding DsRed protein (11). Using an ex vivo-ligated loop system that allows controlled pathogen exposure of intact tissue, we challenged intestinal villi of C57BL/6 CX3CR1GFP/+ mice with DsRed-expressing conidia. Aspergillus conidia readily induced the formation of transepithelial dendrites (Fig. 2B). Individual villi harbored extensions with previously reported globular endings (4) and extensions lacking the latter. Aspergillus conidia entered the CX3CR1GFP/+ lamina propria, where they seemed largely confined to GFP-expressing cells in the tip of the villi. Furthermore, we found transepithelial dendrites and globular structures to be associated with the conidia outside the epithelial cell layer (Fig. 2, C and D), suggesting their direct involvement in pathogen uptake. To further investigate this issue, we exposed conidia to villi of CX3CR1GFP/GFP mice that lack lpDC extensions (4). As shown in Fig. 2E, CX3CR1 deficiency did not impair pathogen entry as indicated by the abundance of noninvasive Aspergillus conidia in the lamina propria of CX3CR1GFP/GFP mice.


Figure 2
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  		FIGURE 2. Pathogen entry into small intestinal lamina propria. Fluorescent microscopic analysis of DsRed-transduced Aspergillus conidia challenged villi of C57BL/6:CX3CR1GFP mice (green, lpDC; red, conidia; diameter 2–3 µm); original magnification, x40). A, Unchallenged CX3CR1GFP/+ villus; B–D, Aspergillus conidia-challenged CX3CR1GFP/+ villi (note conidia in luminal globular structure (* in D); E, CX3CR1GFP/GFP C57BL/6 mouse; and F, DTx-treated, lpDC and lpM{phi}-depleted CD11c-DTR: CX3CR1GFP/+ C57BL/6 mouse. G, UEA-1+ M cells in the villus epithelium; H and H', colocalization of Aspergillus conidia with UEA-1+ M cells on villi of CX3CR1GFP/+ mice.

 
Only a fraction of the CD11c+ mononuclear phagocytes in the intestinal lamina propria expresses CX3CR1 (Fig. 1) and is hence visible in CX3CR1GFP mice. The above experiments thus do not exclude an involvement of CX3CR1-negative lamina propria CD11c+ cells (lpM{phi}) in pathogen uptake. Therefore, we investigated the uptake in a situation where all CD11c+ mononuclear phagocytes are missing from the lamina propria. To this end, we took advantage of an animal model that allows the diphtheria toxin (DTx)-induced ablation of CD11chigh cells in the intact organism (9). To visualize lpDC, the DTR transgene was crossed on the CX3CR1GFP background. Histological and flow cytometric analysis confirmed the rapid depletion of all CD11c+MHC II+ from the small intestine of toxin-treated CX3CR1GFP: CD11c-DTR tg mice (Fig. 3). We then prepared ligated intestinal loops from DTx-treated mice and challenged them with Aspergillus conidia. As seen in Fig. 2F, conidia uptake by the villi persisted in the absence of CD11c+ lamina propria cells.


Figure 3
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  		FIGURE 3. Conditional ablation of CD11c+ lamina propria mononuclear phagocytes. A, Histological analysis of terminal ileum of DTx-treated (left) and untreated (right) CX3CR1GFP/+:CD11c-DTR transgenic mice 24 h after i.p. toxin injection (original magnification, x10). B, Flow cytometric analysis of lamina propria cells of DTx-treated (left) and untreated (right) CD11c-DTR mice. Cells were stained with PE-conjugated anti-MHCII (I-Ab) and allophycocyanin-conjugated anti-CD11c Abs. Note quantitative ablation of all CD11chigh cells, including lpM{phi} and lpDC.

 
Taken together, there is unimpaired entry of our particulate model pathogen into the lamina propria of CX3CR1-deficient mice and mice depleted of both lpDC and lpM{phi}. This suggests that transepithelial dendrites do not significantly contribute to quantitative gut content sampling, which is likely to proceed via alternative routes. Kiyono and coworkers (7) recently reported the existence of intestinal villus M cells that like Peyer’s patch M cells can be identified on the basis of their display of (1-2)-fucose detected by UEA-1. Confirming these results, we could visualize numerous UEA-1+ cells on villi of the terminal ileum of C57BL/6 CX3CR1GFP mice (Fig. 2G). Furthermore, analysis of pathogen-challenged ligated loops revealed conidia clusters in the epidermal layer that coincided with the M cell staining (Fig. 2, H and H'). These results support the notion that intestinal villus M cells are a major entry route for particulate pathogens to the lamina propria.

Transepithelial dendrite formation of CX3CR1+ lpDC is mouse strain dependent

Absence of lpDC extensions in CX3CR1-deficient mice (CX3CR1GFP/GFP) (4) indicates a critical role of the CX3CR1 chemokine receptor and its membrane-tethered ligand CX3CL1 (FKN) in the penetration of the epithelial barrier. To test the general validity of this observation, we investigated the distribution of transepithelial extensions in another inbred mouse strain. Surprisingly, heterozygous mutant CX3CR1GFP/+ BALB/c mice lacked transepithelial dendrites in the terminal ileum both in steady state and after exposure to pathogens (Aspergillus conidia, Salmonella) (Fig. 4 and data not shown). GFP+ lpDC of CX3CR1GFP/+ BALB/c mice express surface CX3CR1 (data not shown). Furthermore, transepithelial dendrites are formed in F1 hybrid CX3CR1GFP/+ mice that inherited their wild-type CX3CR1 allele from the BALB/c and the mutant, CX3CR1GFP allele from the C57BL/6 parent (Fig. 4). The BALB/c allele thus encodes a functioning CX3CR1 receptor capable of promoting dendrite formation. Absence of transepithelial dendrites in BALB/c mice is thus a recessive phenotype not linked to the CX3CR1 locus but events upstream or downstream of CX3CR1 action.


Figure 4
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  		FIGURE 4. Absence of transepithelial dendrites in CX3CR1GFP/+ BALB/c mice. Fluorescent microscopic analysis of Salmonella-challenged small intestinal villi of CX3CR1GFP/GFP C57BL/6 mouse (A), CX3CR1GFP/+ BALB/c mouse (B), and unchallenged CX3CR1GFP/+ F1 (C57BL/6 x BALB/c) mouse (C).

 
The impact of the genetic background on the existence of transepithelial dendrites raised the question of whether their absence in C57BL/6 CX3CR1GFP/GFP mice is indeed due to lack of the CX3CR1 receptor, as we previously proposed (4). Alternatively, the phenotype of the C57BL/6 CX3CR1-deficient mice could result from homozygosity of the chromosomal region (chromosome no. 9) flanking the CX3CR1 GFP allele, which is derived from 129/sv mice (10). To this end, we tested whether transepithelial dendrites are present in mice homozygous for the respective chromosome or its subregion, i.e., F1 hybrid mice resulting from the following crosses: CX3CR1GFP/GFP B6 x 129/Sv and CX3CR1GFP/GFP B6 x CCR2 –/– B6 (the CCR2 locus is adjacent to the CX3CR1 locus and CCR2 mutant mice were generated using the same 129/sv ES cells as the CX3CR1 mutation (10, 26)). In both hybrid strains, we observed transepithelial dendrites upon bacteria challenge (data not shown) confirming our original interpretation that the absence of transepithelial dendrites in C57BL/6 CX3CR1GPF/GFP mice is due to their lack of the chemokine receptor CX3CR1. However, absence of lpDC extensions in CX3CR1GFP/+ BALB/c mice suggests control of their formation by multiple genetic factors.


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Disclosures
 References
 
Studies in the rat have established that lpDC sample enteric Ags and apoptotic epithelial cells in the lamina propria translocate to the mesenteric nodes and present their cargo to lymphocytes (27). The observation that lpDC can penetrate the epithelial tight junction barrier (8) thus led to speculations that besides Peyer’s patch-associated M cells, lpDC might be a major route of luminal Ag transport to lamina propria-draining lymph nodes. Such a scenario would be of critical importance for our understanding of immunosurveillance and pathogen invasion. In the present study, we focus on the most proximal step of this cascade of events, i.e., pathogen entry into the lamina propria. Using hosts that either lack small intestinal transepithelial dendrites (CX3CR1GFP/GFP mice; Ref.4) or were depleted of lpDC (DTx-treated CD11c-DTR transgenic mice (9)) we show that entry of fungal conidia by in large relies on an alternative uptake route. These results suggest that lpDC extensions are dispensable for quantitative sampling of luminal Ags, which are readily accessible to the lamina propria through intestinal villus M cells (7). The study of CX3CR1GFP/+ BALB/c mice revealed that lpDC of this inbred mouse strain seem unable to form transepithelial dendrites. BALB/c and C57BL/6 mice differ in the kinetics of postnatal formation of isolated lymphoid follicles and cryptopatches (27). In addition, Kennedy et al. (28) reported that C57BL/6 mice harbor a natural disruption of the secretory group II phospholipase A2 gene (sPLA2). Murine intestinal sPLA2 has bactericidal activity (29) and the observed high expression of sPLA2 in the intestine of BALB/c mice (28) might therefore affect the intestinal microflora. Furthermore, old BALB/c harbor more IgA+ B cells in their lamina propria than C57BL/6 mice and have elevated IgA titers in fecal extracts (30). However, it remains to be shown whether and how these facts are related to differences in transepithelial dendrite formation. Differential lpDC access to the intestinal lumen is likely to contribute to the well-established observation that inbred mouse strains vary significantly in their susceptibility to pathogen infection known to be under mono- or multigenic control (31).

Absence of transepithelial dendrites in CX3CR1GFP/GFP C57BL/6 and CX3CR1GFP/+ BALB/c mice impairs neither intestinal homeostasis, nor pathogen entry to the lamina propria. This study thus seems to argue against the importance of these structures in intestinal pathogen uptake. However, lpDC extensions are likely decorated with DC-restricted pathogen receptors (for a recent review, see Ref.32). Unlike the M cell route, the lpDC pathway might therefore allow for specific sensing and sampling of the intestinal lumen. Such a notion is also supported by our observation that some lpDC dendrites form upon epithelial penetration globular structures, i.e., an extended matrix for interaction with the gut content (4) (Fig. 2). The lpDC route could be an attractive target for the development of vaccination strategies. Furthermore, it could constitute a critical invasion path of pathogens targeting DC, such as HIV (33). However, a definitive answer on this topic will have to await experimental evidence showing lpDC-dependent entry of specific pathogens or Ag uptake by the transepithelial lpDC processes, migration of Ag-loaded lpDC to lymphoid organs, and activation of naive T cells.


   Acknowledgments
 
We thank Drs. J.-P. Latgé, A. Porgador, and L. Mikkelsen for kindly providing the fungus A. fumigatus, Salmonella, and plasmids, respectively.


   Disclosures
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
References
 
The authors have no financial conflict of interest.


   Footnotes
 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 This work was supported by a European Molecular Biology Organization short-term fellowship (to A.V.-E.) and the MINERVA foundation. S.J. is an Incumbent of the Pauline Recanati Career Development Chair and a Scholar of the Benoziyo Center for Molecular Medicine. Back

2 A.V.-E. and L.L. contributed equally to this work. Back

3 Address correspondence and reprint requests to Dr. Steffen Jung, Department of Immunology, The Weizmann Institute of Science, Rehovot 76100, Israel. E-mail address: s.jung{at}weizmann.ac.il Back

4 Abbreviations used in this paper: DC, dendritic cell; lpDC, lamina propria DC; lpM{phi}, lamina propria macrophage; FKN, fractalkine; UEA, Ulex europaeus agglutinin; DTR, diphtheria toxin receptor; DTx, diphtheria toxin; sPLA2, secretory group II phospholipase A2 gene. Back

Received for publication August 4, 2005. Accepted for publication November 30, 2005.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 

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