Macrophage are the principal reservoir and sustain high virus loads in rhesus macaques after the depletion of CD4+ T cells by a highly pathogenic simian immunodeficiency virus/HIV type 1 chimera (SHIV): Implications for HIV-1 infections of humans

Abstract

The highly pathogenic simian immunodeficiency virus/HIV type 1 (SHIV) chimeric virus SHIV(DH12R) induces a systemic depletion of CD4(+) T lymphocytes in rhesus monkeys during the initial 3-4 weeks of infection. Nonetheless, high levels of viral RNA production continue unabated for an additional 2-5 months. In situ hybridization and immunohistochemical analyses revealed that tissue macrophage in the lymph nodes, spleen, gastrointestinal tract, liver, and kidney sustain high plasma virus loads in the absence of CD4(+) T cells. Quantitative confocal immunofluorescence analysis indicated that greater than 95% of the virus-producing cells in these tissues are macrophage and less than 2% are T lymphocytes. Interestingly, the administration of a potent reverse transcriptase inhibitor blocked virus production during the early T cell phase but not during the later macrophage phase of the SHIV(DH12R) infection. When interpreted in the context of HIV-1 infections, these results implicate tissue macrophage as an important reservoir of virus in vivo. They become infected during the acute infection, gradually increase in number over time, and can be a major contributor to total body virus burden during the symptomatic phase of the human infection.

Figure 1

SHIV and CD4⁺ T lymphocyte levels in infected rhesus monkeys. (a) Viral RNA loads (red) and CD4⁺ T cell levels (black) in rhesus macaques inoculated intravenously with 4.1 × 10⁵ (Rh H27 and Rh H358), 16,400 (Rh 5980), and 656 (Rh 5981) TCID₅₀ of the SHIV_DH12R stock virus are shown. The black crosses indicate the times of animal killing because of their deteriorating clinical condition. (b and c) Plasma viral RNA (red) and CD4⁺ T cell levels (black) in SHIV_DH12R-infected rhesus macaques administered PMPA (30 mg per kg) for 28 days beginning 5 days (b) or 21 days (c) postinfection (PI) (blue rectangles). The cross indicates the time when monkey AG28 was killed.

Figure 2

Virus replication and CD4⁺ T cell depletion in lymphoid tissues during primary SHIV_DH12R infections of rhesus monkeys. Animals inoculated with 1.0 × 10⁵ TCID₅₀ of SHIV_DH12R were killed on day 10 [Rh AH5E (a–c)] or day 14 [Rh AE56 (d–f)]. Mesenteric lymph nodes (a, b, d, and e), spleen (c), and thymus (f) were examined by ISH (b, c, e, and f; visualized as dark blue) and immunohistochemistry for CD4 staining (a and d; visualized as brown). (Original magnifications: a, ×5; b, ×10; c, ×10; d, ×4; e, ×10; and f, ×10.)

Figure 3

Virus replication during the “macrophage phase” of SHIV_DH12R infection. Rhesus macaque Rh H27 was infected with SHIV_DH12R (4.1 × 10⁵ TCID₅₀) and killed at week 12.6. Specimens from mesenteric lymph node (a), ileum (b), spleen (c), liver (d), and kidney (e) were evaluated for virus replication by ISH. (Original magnifications: a, ×10; Inset, ×30; b, ×10; c, ×20; d, ×20; and e, ×10.)

Figure 4

Identification of virus-producing lymph node cells during the “macrophage phase” of SHIV_DH12R infection. Two individual mesenteric lymph node sections (a–c or d–f) were subjected to combination ISH using a riboprobe recognizing SHIV sequences (a and d; visualized in green) and immunostaining specific for macrophage (b; visualized in red) or CD3⁺ T lymphocytes (e; visualized in red). (c) Superimposed image of a with b. (f) Superimposed image of d and e.