Dendritic cells route human immunodeficiency virus to lymph nodes after vaginal or intravenous administration to mice

Abstract

We have developed a murine model to study the involvement of dendritic cells (DC) in human immunodeficiency virus (HIV) routing from an inoculation site to the lymph nodes (LN). Murine bone marrow-derived DC migrate to the draining LN within 24 h after subcutaneous injection. After incubation of these cells with heat-inactivated (Hi) HIV type 1 (HIV-1), HIV RNA sequences were detected in the draining LN only. Upon injection of DC pulsed with infectious HIV, the virus recovered in the draining LN was still able to productively infect human T cells. After a vaginal challenge with Hi HIV-1, the virus could be detected in the iliac and sacral draining LN at 24 h after injection. After an intravenous challenge, the virus could be detected in peripheral LN as soon as 30 min after injection. The specific depletion of a myeloid-related LN DC population, previously shown to take up blood macromolecules and to translocate them into the LN, prevented HIV transport to LN. Together, our data demonstrate the critical role of DC for HIV routing to LN after either a vaginal or an intravenous challenge, which does not require their infection. Therefore, despite the fact that the mouse is not infectable by HIV, this small animal model might be useful to test preventive strategies against HIV.

FIG. 1

Murine BM-derived DC transmit a productive HIV infection to human T-lymphoblastoid cells and to human T lymphocytes. The kinetics of HIV replication, measured by ELISA for p24 production, in cocultures of BM-derived DC with either HUT-78 cells (A) or human T lymphocytes (B) was determined. (A) HUT-78 cells were cocultured with Hi or infectious HIV-1_LAI-pulsed DC or with unpulsed DC as a control. (B) T lymphocytes were cocultured with HIV-1_LAI-pulsed DC or T lymphocytes in the presence of SEA. The control was unpulsed DC with T lymphocytes in the presence of SEA.

FIG. 2

Murine BM-derived DC route HIV to the draining LN after s.c. injection. (A) Section of a draining popliteal LN observed under a fluorescence microscope 24 h after injection of murine BM-derived, PKH2-stained DC into the footpads of DBA/2 mice. Note the presence of numerous fluorescent cells. Magnification, ×200. (B) Detection of HIV (left) and hCD4 (right) RNAs by RT-PCR in LN of nontransgenic mice, 24 h after injection of 25 μl containing 5 × 10⁵ Hi HIV-1_LAI-pulsed DC derived from hCD4-transgenic mice into the footpad. Pop, popliteal LN; Ing, inguinal LN.

FIG. 3

HIV remains infectious after its routing to the draining LN by DC. Twenty-five microliters of HIV-1_LAI supernatant (squares) or 25 μl containing 5 × 10⁵ HIV-1_LAI-pulsed DC (triangles) was injected into the footpads of normal mice. Twenty-four hours later, LN were harvested and the cells were cocultured with HUT-78 cells. The kinetics of HIV replication was measured by ELISA for p24 production.

FIG. 4

HIV is routed to draining LN after vaginal application. (A) Fluorescence-activated cell sorter analysis of LN cells 24 h after FITC application on the mouse vaginal mucosa. FITC-fluorescent cells could be detected almost exclusively within a subpopulation of LN DC (l-DC) characterized by expression of CD11c and MHC class II molecules, as previously described (46) (also see Fig. 5). Data are presented as FITC fluorescence histograms for l-DC from an iliac LN of a noninjected mouse (left panel) and from iliac and inguinal LN of an FITC-inoculated mouse (middle and right panels, respectively). Results are from the analysis of 2 × 10⁵ to 3 × 10⁵ LN cells. FITC staining is detected in about 30% of the l-DC-draining iliac LN. Results from one representative experiment of four independent experiments with two or three mice per group are shown. (B) Detection of HIV RNA by RT-PCR in the vaginal mucosae and inguinal, iliac, and sacral LN of mice, 24 h after inoculation of 25 μl of Hi HIV-1_LAI supernatant into the vaginal vault.

FIG. 5

Specific abrogation of s-DC in transgenic mice. Fluorescence-activated cell sorter analysis of inguinal LN of control and DC-depleted mice is shown. s-DC depletion was obtained by a 7-day GCV treatment of transgenic mice expressing the suicide gene for HSV-1 TK preferentially in DC (47, 48). Analysis of s-DC and l-DC subpopulations based on expression of the CD11c marker and MHC class II molecules was performed as previously described (46).