Reduced Simian Immunodeficiency Virus Replication in Macrophages of Sooty Mangabeys Is Associated with Increased Expression of Host Restriction Factors

Abstract

Macrophages are target cells of HIV/SIV infection that may play a role in AIDS pathogenesis and contribute to the long-lived reservoir of latently infected cells during antiretroviral therapy (ART). In previous work, we and others have shown that during pathogenic SIV infection of rhesus macaques (RMs), rapid disease progression is associated with high levels of in vivo macrophage infection. In contrast, during nonpathogenic SIV infection of sooty mangabeys (SMs), neither spontaneous nor experimental CD4(+) T cell depletion results in substantial levels of in vivo macrophage infection. To test the hypothesis that SM macrophages are intrinsically more resistant to SIV infection than RM macrophages, we undertook an in vitro comparative assessment of monocyte-derived macrophages (MDMs) from both nonhuman primate species. Using the primary isolate SIVM949, which replicates well in lymphocytes from both RMs and SMs, we found that infection of RM macrophages resulted in persistent SIV-RNA production while SIV-RNA levels in SM macrophage cultures decreased 10- to 100-fold over a similar temporal course of in vitro infection. To explore potential mechanisms responsible for the lower levels of SIV replication and/or production in macrophages from SMs we comparatively assessed, in the two studied species, the expression of the SIV coreceptor as well as the expression of a number of host restriction factors. While previous studies showed that SM monocytes express lower levels of CCR5 (but not CD4) than RM monocytes, the level of CCR5 expression in MDMs was similar in the two species. Interestingly, we found that SM macrophages exhibited a significantly greater increase in the expression of tetherin (P = 0.003) and TRIM22 (P = 0.0006) in response to alpha interferon stimulation and increased expression of multiple host restriction factors in response to lipopolysaccharide stimulation and exposure to SIV. Overall, these findings confirm, in an in vitro infection system, that SM macrophages are relatively more resistant to SIV infection compared to RM macrophages, and suggest that a combination of entry and postentry restriction mechanisms may protect these cells from productive SIV infection.

Importance: This manuscript represents the first in vivo comparative analysis of monocyte-derived macrophages (MDMs) between rhesus macaques, i.e., experimental SIV hosts in which the infection is pathogenic and macrophages can be infected, and sooty mangabeys, i.e., natural SIV hosts in which the infection is nonpathogenic and macrophages are virtually never infected in vivo. This study demonstrates that mangabey-derived MDMs are more resistant to SIV infection in vitro compared to macaque-derived MDMs, and provides a potential explanation for this observation by showing increased expression of specific retrovirus restriction factors in mangabey-derived macrophages. Overall, this study is important as it contributes to our understanding of why SIV infection is nonpathogenic in sooty mangabeys while it is pathogenic in macaques, and is consistent with a pathogenic role for in vivo macrophage infection during pathogenic lentiviral infection.

FIG 1

Nonspecific esterase activity and surface marker expression in SM and RM macrophages. (a and b) Nonspecific esterase staining in SM (a) and RM (b) monocyte-derived macrophages. (c) CD14, HLA-DR, and CD11b staining (red lines) relative to unstained controls (blue lines) in monocyte-derived macrophages from SM and RM.

FIG 2

SIV_M949 infection of RM and SM macrophages. (a and b) Fold change in SIV gag RNA levels in the supernatant of monocyte-derived macrophages from RMs (a) and SMs (b) after in vitro infection with SIV_M949. SIV gag RNA level at day 0 represents the amount of virus in the initial inoculation.

FIG 3

CCR5, CXCR6, and GPR15 expression in SM and RM macrophages. The relative CCR5, CXCR6, and GPR15 expression in SM and RM monocyte-derived macrophages was measured via real-time PCR and normalized to the housekeeping gene GAPDH.

FIG 4

IFN-α-induced restriction factors in SM and RM macrophages. MDMs from SMs and RMs were stimulated with IFN-α, and the fold change in mRNA levels was quantified via real-time PCR relative to unstimulated controls for APOBEC3G, APOBEC3H, TRIM5α, SAMHD1, tetherin, TRIM22, MX1, MX2, and OAS2. All genes were normalized to the housekeeping gene GAPDH.

FIG 5

LPS-induced restriction factors in SM and RM macrophages. MDMs from SMs and RMs were stimulated with LPS, and the fold change in mRNA levels was quantified via real-time PCR relative to unstimulated controls for APOBEC3G, APOBEC3H, TRIM5α, SAMHD1, tetherin, and TRIM22. The relative expression of the LPS receptors CD14, MD2, and TLR4 was assessed in unstimulated MDMs from SMs and RMs. All genes were normalized to the housekeeping gene GAPDH.

FIG 6

Relative expression of restriction factors in SM and RM macrophages after SIV infection. The expression of restriction factors and classical ISGs MX1, MX2, and OAS2 were measured relative to the housekeeping gene GAPDH at day 21 after in vitro infection with SIV_M949 in MDMs of SMs and RMs. In this part of the study, we examined a total of five RM-derived MDMs and six SM-derived MDMs that were exposed to different doses of virus (i.e., ranging between 400 and 3,200 pg p27 equivalents) for a total of 7 and 10 data points for RM and SM, respectively.