The AIDS-like disease of CD4C/human immunodeficiency virus transgenic mice is associated with accumulation of immature CD11bHi dendritic cells

Abstract

CD4C/human immunodeficiency virus (HIV) transgenic mice develop an AIDS-like disease. We used this model to study the effects of HIV-1 on dendritic cells (DC). We found a progressive decrease in total DC numbers in the lymph nodes, with a significant accumulation of CD11b(Hi) DC. In the thymus, the recovery of transgenic CD8alpha(+) DC had a tendency to be lower. Spleen DC were augmented in the marginal zone. Transgenic DC showed a decreased capacity to present antigen in vitro, consistent with their reduced major histocompatibility complex class II expression and impaired maturation profile. The accumulation of immature DC may contribute to disease and may reflect an adaptive advantage for the virus by favoring its replication and preventing the generation of fully functional antiviral responses.

FIG. 1.

Expression of the HIV-1 transgene in murine lymph node DC. Transgene expression in lymph node DC of CD4C/HIV^MutA transgenic mice was detected by in situ hybridization with an HIV-1-specific antisense riboprobe (a to c) and a sense probe as a control (d). The expression of MHC II (a and b) and CD11c (c) on the DC was determined by immunohistochemistry and appears in brown. Magnification, ×100; bar, 25 μm. Data are representative of at least nine experiments, each involving cells from pooled lymph nodes of sex- and age-matched transgenic (n = 3) and control nontransgenic (n = 3) mice.

FIG. 2.

Detection of DC in cryosections of lymph nodes and spleen of CD4C/HIV^MutA transgenic mice. The presence of DC was assessed on cryosections of peripheral lymph nodes (a) and spleen (b) of nontransgenic (non-Tg) and transgenic (Tg) mice at 3 months of age. IgM expression is in red (Texas Red [TR]) and CD11c expression is in green (fluorescein isothiocyanate [FITC]). Magnification, ×10; bar, 100 μm. Sections were analyzed by confocal microscopy. MZ, marginal zone; SC, subcapsular sinuses. Data are representative of at least three experiments, each involving the analysis of lymph node and spleen sections from sex- and age-matched transgenic (n = 2) and control nontransgenic (n = 2) mice.

FIG. 3.

Phenotypic characterization of lymph node DC in CD4C/HIV^MutA transgenic mice. DC were purified from peripheral lymph nodes of nontransgenic (non-Tg) and transgenic (Tg) mice. The data show FACS analysis on CD11c⁺ gated lymph node DC from nontransgenic and transgenic mice. Results are shown as mean fluorescence intensity. Data are shown for DC from animals 3 months of age and are representative of at least 12 experiments with mice ranging between 2 and 4 months of age, each experiment involving cells from pooled lymph nodes of sex- and age-matched transgenic (n = 3) and control nontransgenic (n = 3) mice.

FIG. 4.

DC functions are impaired in CD4C/HIV^MutA transgenic mice. DC were purified from peripheral lymph nodes of nontransgenic (non-Tg) and transgenic (Tg) mice at 6 to 8 and 16 to 18 weeks of age. DC were irradiated with 3,000 rads prior to culture. (a) For the allogeneic mixed leukocyte reaction, DC were cocultured at ratios of 1:10, 1:100, or 1:1,000 with purified resting CD4⁺ T cells from BALB/c mice in the presence or absence of IL-2 (100 U/ml). Proliferation was measured by [³H]thymidine incorporation for the last 18 h of a 48-h culture period. Experiments are shown separately for younger mice (6 to 8 weeks old) (left panel) and older (16 to 18 weeks old) mice (right panel). (b) To assess costimulatory capacity, DC isolated from nontransgenic (non-Tg) and transgenic (Tg) 16- to 18-week-old mice were cocultured on anti-CD3 (0.1 μg/ml)-coated plates with syngeneic resting CD4⁺ T cells purified from nontransgenic mice at ratios of 1:10, 1:100, and 1:1,000. T-cell proliferation was measured as in a. (c and d) The capacity to present pigeon cytochrome c (Pcc) or pigeon cytochrome c peptide to purified resting syngeneic pigeon cytochrome c-specific T-cell receptor CD4⁺ T cells from AD10 mice was assessed. Prior to culture, DC were incubated overnight with either medium alone, pigeon cytochrome c, or pigeon cytochrome c peptide at the indicated concentrations. DC were then cocultured with CD4⁺ T cells at the indicated DC-to-T-cell ratio. T-cell proliferative responses were measured as in a. Experiments are shown for older mice (16 to 18 weeks old). (a to d) Bars on graphs represent the mean ± standard error of the mean. For each experiment shown, the data are representative of at least four experiments, each involving DC from pooled lymph nodes of sex- and age-matched transgenic (n = 3) and control nontransgenic (n = 3) mice. Statistical analyses were performed according to Student's t test on data from all experiments. Differences between nontransgenic and transgenic DC that reached significance are shown in panels a (right panel), c, and d at P < 0.05 and in panel b at P < 0.001.

FIG. 5.

Bone marrow-derived DC from CD4C/HIV^MutA transgenic mice express the HIV transgene and are impaired in their maturation process. Bone marrow progenitors derived from the femur of nontransgenic (non-Tg) and transgenic (Tg) mice were allowed to mature for 6 days in the presence of 10% fetal bovine serum and GM-CSF. Cells were then recultured for an additional 24 h with (b) (right) or without (a and b) (left) lipopolysaccharide (10 μg/ml). The recovered cells showed dendritic morphology and expressed CD11c, as assessed by FACS. (a) These bone marrow-derived DC were analyzed for transgene expression by in situ hybridization with an HIV-1-specific antisense probe and a sense probe as a control. Counterstaining was done with hematoxylin and eosin. Magnification, ×40; bar, 50 μm. (b) Bone marrow-derived DC were analyzed by FACS for surface expression of MHC I, MHC II, CD40, and B7-2 (CD86) molecules. The data are representative of at least four different experiments, each involving cells from the bone marrow of sex- and age-matched transgenic (n = 3) and control nontransgenic (n = 3) mice.

FIG. 6.

Thymic DC from CD4C/HIV^MutA transgenic mice have reduced levels of MHC II and CD8α on their surface. DC were purified from the thymus of nontransgenic (non-Tg) and transgenic (Tg) mice 8 weeks of age and analyzed by FACS for surface expression of MHC II, CD8α, and CD11b. Data are representative of at least seven experiments with animals 6 to 16 weeks of age and are shown as a percentage and mean fluorescence intensity. Each experiment involved cells from pooled thymuses of sex- and age-matched transgenic (n = 3) and control nontransgenic (n = 3) mice.