The Journal of Immunology, 2006, 177: 2035-2036.
Copyright © 2006 by The American Association of Immunologists
Response to Comment on "CCR7 Is Critically Important for Migration of Dendritic Cells in Intestinal Lamina Propria to Mesenteric Lymph Nodes"
Myoung Ho Jang and
Masayuki Miyasaka
Laboratory of Immunodynamics, Department of Microbiology and Immunology, Osaka University Graduate School of Medicine, Osaka, Japan
In response to a letter by Dr. David Hume (1), we would like to point out that the following lines of evidence clearly indicate that the lamina propria dendritic cells (LP-DC) we reported (2) are indeed DC but not macrophages. As seen in Fig. 1A in our recent analysis in which we used a 17.5% Accudenz solution instead of the original 15.5% solution to enrich for light-density cells such as macrophages and DC from the small intestinal LP, CD11chighCD11blow (R1) and CD11chighCD11bhigh (R2) cell populations with dendritic morphology were abundantly found and were strongly positive for a DC-specific marker, DEC-205. With regard to F4/80 expression, R1 subset was clearly negative, but R2 was moderately positive. Functional analysis revealed that both R1 and R2 subsets can present alloantigen to naive T cells just as effectively as splenic DC and lymph node DC (Fig. 1B, left column) and can also present OVA efficiently to OVA-TCR expressing T cells (Fig. 1B, right column). Thus, these DEC-205+ LP-DC have the hallmark of DC, i.e., potent Ag-presenting abilities to T cells.
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FIGURE 1. CD11chigh subsets in the small intestinal LP are DC but not macrophages. A, Low-density lamina propria cells were isolated from the BALB/c small intestine using a 17.5% Accudenz gradient. Enriched cells were stained for CD11b, CD11c, DEC-205, and F4/80 and analyzed by flow cytometry. Several CD11c+ subsets could be recognized on the basis of their different CD11c/CD11b expression patterns: CD11chighCD11blow (R1), CD11chighCD11bhigh (R2), CD11cintCD11bhigh (R3), and CD11clowCD11bhigh (R4; consisting of mainly eosinophils). Histogram FACS profiles shown on the right were acquired after gating on the R1, R2, and R3 subsets. Shaded, isotype controls; open, stained for DEC-205 or F4/80. B, LP-DC were examined for their Ag-presenting abilities. FACS-sorted BALB/c LP-DC were either cocultured with CFSE-labeled C57BL/6 spleen cells for 3 days (left column) or pulsed with OVA protein first and then cocultured with CFSE-labeled OVA TCR transgenic CD4+ T cells (DO11.10; 1:20) for 3 days (right column). DC from other tissues were also treated in the same way. T cell proliferation was measured by CFSE dilution. Numbers within histograms indicate the percentage of proliferated cells.
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Then, are the intestinal F4/80-positive, MHC class II-positive phagocytic macrophages described by Dr. Hume’s group (3) also found in our analysis? The answer seems yes. They appear to represent CD11cintCD11bhigh cell population (R3) because they are strongly F4/80 positive, DEC-205 negative (Fig. 1A), and also strongly positive for MHC class II expression (data not shown). Although this cell population was hardly detectable in our original analysis (2), the use of 17.5% Accudenz solution instead of 15.5% solution in the present series of analysis allowed us to visualize such cells.
Collectively, by functional definition, LP-DC (2) are indeed DC with potent Ag-presenting abilities. Dr. Hume’s finding (3) is also correct that macrophages are present in substantial numbers in the small intestinal LP, but they appear clearly distinguishable from the LP-DC (2).
References
- Hume, D. A.. 2006. Comment on "CCR7 is critically important for migration of dendritic cells in intestinal lamina propria to mesenteric lymph nodes.". J. Immunol. 177: 2035[Free Full Text]
- Jang, M. H., N. Sougawa, T. Tanaka, T. Hirata, T. Hiroi, K. Tohya, Z. Guo, E. Umemoto, Y. Ebisuno, B.-G. Yang, et al 2006. CCR7 is critically important for migration of dendritic cells in intestinal lamina propria to mesenteric lymph nodes. J. Immunol. 176: 803-810. [Abstract/Free Full Text]
- Pavli, P., C. E. Woodhams, W. F. Doe, D. A. Hume. 1990. Isolation and characterization of antigen-presenting dendritic cells from the mouse intestinal lamina propria. Immunology 70: 40-47. [Medline]
