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, 7 (8), e1002170

Functional Cure of SIVagm Infection in Rhesus Macaques Results in Complete Recovery of CD4+ T Cells and Is Reverted by CD8+ Cell Depletion

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Functional Cure of SIVagm Infection in Rhesus Macaques Results in Complete Recovery of CD4+ T Cells and Is Reverted by CD8+ Cell Depletion

Ivona Pandrea et al. PLoS Pathog.

Abstract

Understanding the mechanism of infection control in elite controllers (EC) may shed light on the correlates of control of disease progression in HIV infection. However, limitations have prevented a clear understanding of the mechanisms of elite controlled infection, as these studies can only be performed at randomly selected late time points in infection, after control is achieved, and the access to tissues is limited. We report that SIVagm infection is elite-controlled in rhesus macaques (RMs) and therefore can be used as an animal model for EC HIV infection. A robust acute infection, with high levels of viral replication and dramatic mucosal CD4(+) T cell depletion, similar to pathogenic HIV-1/SIV infections of humans and RMs, was followed by complete and durable control of SIVagm replication, defined as: undetectable VLs in blood and tissues beginning 72 to 90 days postinoculation (pi) and continuing at least 4 years; seroreversion; progressive recovery of mucosal CD4(+) T cells, with complete recovery by 4 years pi; normal levels of T cell immune activation, proliferation, and apoptosis; and no disease progression. This "functional cure" of SIVagm infection in RMs could be reverted after 4 years of control of infection by depleting CD8 cells, which resulted in transient rebounds of VLs, thus suggesting that control may be at least in part immune mediated. Viral control was independent of MHC, partial APOBEC restriction was not involved in SIVagm control in RMs and Trim5 genotypes did not impact viral replication. This new animal model of EC lentiviral infection, in which complete control can be predicted in all cases, permits research on the early events of infection in blood and tissues, before the defining characteristics of EC are evident and when host factors are actively driving the infection towards the EC status.

Conflict of interest statement

The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Viral replication during acute and chronic SIVagm infection of RMs.
(a) Plasma viral load quantification showed high levels of viral replication during acute infection followed by complete control of viremia, with undetectable plasma viral loads, starting from days 72–98 p.i. on. (b) the overall pattern of SIVagm replication in the intestine generally paralleled that of plasma viral load, but control of viral replication was achieved at later time points (after day 132 p.i.). The detection limit of the assay was 102 copies/ml of plasma and 10 copies per 106 cells.
Figure 2
Figure 2. Control of SIVagm replication in RMs was confirmed by seroreversion of anti-SIVagm binding and neutralizing antibodies.
(a) Dynamics of anti-gp41 antibody titers in SIVagm-infected RMs by SIVagm.sab-specific ELISA. All animals seroconverted between days 14–21 post-infection. Antibody titers reached the highest levels by day 98 p.i., then continuously declined as VLs were controlled. (b) Neutralizing antibodies against SIVagm.sab92018 also vane with the control of viral replication.
Figure 3
Figure 3. Measurement of SIV RNA in serially sacrificed RMs infected with SIVagm.sab.
Six rhesus macaques were included in this study and were serially sacrificed at the peak of viral replication (day 9 pi-RMDJ52 and D10 pi-RMCV08), at the set-point (day 35 pi-RMDJ73 and D42 pi-RMEK15) and during chronic infection (D180pi), when infection is controlled (RMCM48 and RMCE26). (a) Quantification of plasma viral load showed that all these RMs replicated the virus at high levels during acute infection and that the virus load was controlled below detection limits in the RMs euthanized during chronic infection. (b) Quantitative RT–PCR analysis of SIVagm.sab RNA from over 30 tissues obtained at necropsy demonstrated that virus control is not due to a restriction of virus replication to a limited number of anatomical sites.
Figure 4
Figure 4. Changes in CD4+ T cells in SIVagm-infected RMs.
Acute viral replication resulted in a significant depletion of CD4+ T cells in blood (a), lymph nodes (b) and intestine (c), which was massive at mucosal sites. With the control of viral replication, complete recovery of CD4+ T cell was observed during the follow-up.
Figure 5
Figure 5. CD4+ T cell depletion pattern in SIVagm-infected macaques.
PBMCs were examined by polychromatic flow-cytometry for their correlated expression of cell surface CD3 vs CD4 vs CCR5 vs CD28 vs CD95 vs CXCR4 both before and 28 days after infection. Note the striking depletion of CD4+ memory T cells (CD95+), and within the memory population, the effector memory (CD28negCD95+) and the CCR5-expressing subsets. Meanwhile, CXCR4-expressing cells are well maintained during acute SIVagm.sab infection in RMs. This pattern is identical to the one reported by Picker et al., for CCR5 tropic viruses .
Figure 6
Figure 6. Dynamics of CD4+ T cell apoptosis during acute and controlled SIVagm.sab infection of rhesus macaques.
(a) Flow cytometric assessment of apoptotic and necrotic cells in the gut showed significant increases of both apoptotic and necrotic cells in association with the high levels of acute viral replication. Increases in apoptosis levels persisted longer than SIVagm viremia being probably responsible for the delays in CD4+ T cell restoration. (b) Immunohistochemistry for activated caspase-3. There is a significant increase in apoptosis during the acute infection in both intestine and LNs (middle panels) in SIVagm-infected RMs compared to the pre-inoculation samples from the same sites (upper panels). During late chronic infection the number of activated caspase-3 positive cells decrease in intestine and LNs (lower panels) to levels similar to the pre-infection ones, confirming the flow cytometry data. Note that, during acute SIVagm infection, both lamina propria and intestinal epithelia of RMs show an increased number of apoptotic cells (middle panel, left) thus explaining the transient bacterial translocation.
Figure 7
Figure 7. Longitudinal analysis of the microbial translocation in 5 SIVagm-infected RMs.
Only a transient increase in plasma levels of sCD14 was observed in SIVagm-infected RMs. Then, with the control of apoptosis, the sCD14 levels returned to baseline.
Figure 8
Figure 8. Dynamics of peripheral T cell activation, as assessed by changes in HLA-DR and Ki-67 expression on CD4+ and CD8+ T cells in SIVagm-infected RMs.
SIVagm infection of RMs induced transient levels of activation and proliferation of both CD3+ CD4+ and CD3+ CD8+ T cells. After the control of viral replication in tissues, both activation and proliferation markers returned to preinfection levels.
Figure 9
Figure 9. In vitro SIVagm replication on PBMCs from rhesus macaques and AGMs.
In vitro replication was tested on PBMCs originating from 3 RMs (DV11, DF93 and DD51). Each experiment was performed in triplicate. In vitro growth curves (±standard deviation) are shown.
Figure 10
Figure 10. Administration of cM-T807 depleting antibody resulted in a rebound of plasma VLs in all three SIVagm-infected RMs.
(a) cM-T807 successfully depleted anti-CD8 cells in periphery and resulted in a rebound of viral replication; (b) transient CD4+ T cell depletion in periphery and intestine was observed following the rebound of viral replication (c) increased in CD4+ T cell activation and proliferation (as assessed by the study of –DR and Ki-67) were observed, that lasted longer than the rebound in viral replication. CD8 depleted animals: BA38 (▴); V492 (▪); P373 (•).

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