DC-SIGN/CD209 is a C-type lectin that is able to bind and transmit HIV-1, as discovered by Geijtenbeek et al. (4
), and it has been implicated in the capacity of DCs to stimulate resting T cells (1
). To assess the function of DC-SIGN in the noninflamed human lymph node, we have used a panel of anti-DC-SIGN mAbs, including new reagents obtained by immunization with recombinant vaccinia-DC-SIGN. Surprisingly, we found that DC-SIGN staining is primarily observed on macrophages throughout the medullary sinuses, as manifest by strong colabeling for two molecules that are abundant on macrophages, the CD68 lysosomal membrane protein and the mannose receptor, and a lack of DC markers such as CD205 and CD11c. In contrast, DCs in the T cell area of the cortex express high levels of CD205 and CD11c, but were not detectably stained with several new and previously available anti-CD209 mAb. Our approaches cannot determine whether DC-SIGN is not expressed at all, but only that the major depot of CD209 in the human lymph node are macrophages in the medulla and not DCs in the cortex. It will be important to learn how to isolate these cells from the lymphoid tissues, but at this time we have not succeeded in releasing significant numbers of DCs or macrophages with the very high levels of CD205 and CD209 that we observed in sections. Other reports have called attention to the detection of DC-SIGN on macrophages in lung and other sites (19
), but we report that in the noninflamed lymph node, DC-SIGN is abundant on most macrophages in the medulla, but not in most DCs in the T cell areas.
Our findings in sections are surprising relative to the previous literature. Geijtenbeek et al. (1
) identified some DC-SIGN+
cells in the T cell areas. Lore et al. (23
) reported DC-SIGN+
cells in the parafollicular T cell-rich areas of lymph nodes from patients with HIV and EBV infection, and that these cells were reduced in healthy controls. Soilleux et al. (19
) used a polyclonal anti-DC-SIGN serum to examine adult and fetal tissues and reported some cells with a dendritic morphology in the T cell areas and within sinuses in the cortex. Tailleux et al. (38
) studied DC-SIGN expression in lymph nodes from tuberculosis patients and reported DC-SIGN+
cells within the granulomas and subcapsular sinuses. Engering et al. (24
) reported DC-SIGN+
cells in the outer cortex in proximity to sinuses. While our study was under review, Krutzik et al. (37
) described how macrophages in leprosy lesions could express DC-SIGN. Our paper, for the first time, reports the abundance of this lectin in medullary macrophages in noninflamed lymph nodes and its paucity on many T cell area DCs.
Our tissue section results indicate that many DCs in the T cell areas lack the markers of monocyte-derived DCs, particularly high expression of DC-SIGN/CD209 and mannose receptor/CD206. It is possible that the equivalent of the cultured monocyte-derived DCs only develops under select circumstances, e.g., during an inflammatory response or in special tissue niches. We and others (19
) found that cytokines such as IL-4, IL-13, and IL-15 quickly up-regulated DC-SIGN expression on monocytes, so that cells with the phenotype of monocyte-derived DCs might accumulate in situations such as parasite infection and allergy. The reports of DC-SIGN+
cells in lymphatic sinuses, particularly in the subcapsular region of lymph nodes, may also reflect migratory monocyte-derived DCs responding to a stimulus in the periphery. Within the T cell areas, we did detect small foci in which there were CD11c- and DEC-205-positive DCs that coexpressed DC-SIGN. However, most DCs in the T cell area had features shared with the myeloid subset of DCs in blood, i.e., the cells expressed CD11c and DEC-205/CD205, but lacked DC-SIGN and mannose receptor. We propose that the finding of DC-SIGN-positive DCs in the T cell areas represent situations where the equivalent of monocyte-derived DCs are being generated in vivo, but that the major reservoir of DC-SIGN in normal lymph node is the medullary sinus macrophage rather than cortical DCs.
With the anti-DC-SIGN Abs we have generated and those already characterized, we also restudied the role of DC-SIGN as a factor that participates in two types of DC-T cell interaction. All the anti-DC-SIGN mAbs tested, in contrast to anti-HLA-DR, were unable to inhibit DC-induced proliferation of resting T cells in the MLR. These results differ from the initial conclusion that DC-SIGN supports primary immune responses that arose from the observation that anti-DC-SIGN mAbs could reduce MLR stimulation by ~60% (1
). However, we were unable to detect a block of the MLR with a panel of anti-DC-SIGN mAbs even at limiting doses of DCs and duration of MLR. We also looked at the contribution of DC-SIGN to transmission of HIV by monocyte-derived DCs and by blood DCs that had been induced to express DC-SIGN with IL-4. We found that the mAbs to DC-SIGN did not impose a major reduction on transmission of HIV by these DCs, whereas the same mAbs led to a major ~90% reduction of HIV transmission by DC-SIGN-transfected Raji cells. In both the MLR and HIV transmission assays, we additionally evaluated DCs in which DC-SIGN expression had been dampened with siRNA, and again, no blockade was noted, even though this approach markedly reduced HIV transmission from Raji DC-SIGN transfectants. These results are consistent with several other reports that molecules other than DC-SIGN can mediate virus transmission from DCs to T cells, using monocyte-derived DCs (6
). Likewise, there are types of DCs that lack DC-SIGN, i.e., Langerhans cells and both myeloid and plasmacytoid DCs in blood, that are able to transmit HIV to T cells in vitro. Clearly, DC-SIGN represents an exciting new mechanism by which pathogens are recognized by innate cells, but DCs use additional mechanisms to transmit HIV, and we suggest that macrophages within lymph nodes be considered in pursuing the functions of DC-SIGN in vivo.