Short Communication: Plasmacytoid Dendritic Cells from HIV-1 Elite Controllers Maintain a Gut-Homing Phenotype Associated with Immune Activation

Abstract

Lentivirus infections are characterized by a dramatic loss of mucosal CD4⁺ T cells, breakdown of the gut mucosa, and subsequent chronic immune activation. Residual immune activation persists even in patients controlling virus replication and remains a significant source of ongoing disease morbidities, but the causes are unclear. Plasmacytoid dendritic cells (pDCs), primary producers of interferon (IFN)-α, have been previously shown to be depleted from peripheral blood of HIV patients and simian immunodeficiency virus (SIV)-infected macaques, and most recently have been shown to accumulate in the gut mucosa. Although previous work has shown that pDC frequencies can be reduced in the circulation of HIV-1 Elite Controllers, it is unknown if gut-homing also occurs. In this new study we found that during progressive HIV-1 infection pDCs were depleted in peripheral blood compared to seronegative controls, and, correlating with plasma viremia, the remaining pDCs upregulated the gut-homing marker, α₄β₇. Even in HIV-1 Elite Controllers pDCs were significantly reduced in blood and α₄β₇ expression was still significantly upregulated compared to seronegative controls. Interestingly, pDC trafficking to the gut was associated with increased Ki67 and HLA-DR on circulating CD4⁺ and CD8⁺ T cells. Overall, these data suggest that gut trafficking of pDCs is independent of virus replication and could be mediated by alternative mechanisms, which in turn could contribute to residual immune activation in HIV-1 Elite Controllers.

Plasmacytoid dendritic cells (pDCs), also referred to as natural-interferon-producing cells, respond to microbial pathogens by rapidly producing interferon (IFN)-α, as well as other proinflammatory cytokines such as interleukin (IL)-12, tumor necrosis factor (TNF)-α, and macrophage inflammatory protein (MIP)-1β. This early cytokine production contributes to directly block viral replication and also initiates a cascade of other innate and adaptive immune responses. As first reported over a decade ago, multiple groups have verified that pDC numbers are severely reduced in blood during HIV infection, ^1

–4 and these findings have been verified in SIV-infected macaques models.^5
–7 Furthermore, loss of pDCs, which begins during primary infection, can also occur in HIV-1 Elite Controllers, and is only partially reversible by combination antiretroviral therapy (cART).⁸

Studies of acute pathogenic lentivirus infection in macaques have demonstrated a transient increase of pDCs in peripheral blood due to rapid egress from the bone marrow.^5,7 pDCs also redistribute to the lymph nodes early after infection where they undergo rapid apoptosis, but are excluded from lymph nodes during chronic infection due to ongoing fibrosis.^7,9,10 Interestingly, others and we have very recently refined this model to show that in fact pDCs are not necessarily depleted by HIV/SIV infection, but can accumulate in the gastrointestinal tract, depending on cell surface α₄β₇ expression.^11,12 pDCs have also been shown to accumulate in the spleen and lymph nodes in some human subjects,^13,14 suggesting pDCs in the circulation may not necessarily be depleted, but rather are redistributing to other tissues. Interestingly, these phenomena do not seem to occur in the nonpathogenic SIV natural hosts, sooty mangabeys and African green monkeys.^12,15

In this study we evaluated the frequency of pDCs and myeloid dendritic cells (mDCs) in the circulation of HIV-1 Elite Controllers, as well as α₄β₇ expression on both subsets as a surrogate marker for gut homing. Both the University of Alabama at Birmingham (UAB) and Harvard University IRB reviewed and gave approval for the studies herein. Written, informed consent was obtained from each of the study subjects.

HIV-1 Elite Controllers were defined by stable CD4⁺ T cell counts and sustained plasma viral loads below the threshold of detection as described previously.¹⁶ The median ages of persons included in this study were 45.4, 34.6, and 46.8 years in seronegative subjects, viremic HIV⁺ subjects, and Elite Controllers, respectively. Genders were 50%, 80%, and 60% male in the seronegative subjects, viremic HIV⁺ subjects, and Elite Controllers, respectively. Plasma HIV-1 RNA levels were determined using the Amplicor ultrasensitive HIV-1 monitor assay at UAB hospitals (version 1.5; Roche Diagnostics Systems) and absolute CD4⁺ T cell counts were determined in UAB diagnostic laboratories as described previously.¹⁷ All Elite Controllers had sustained viral loads of <50 copies/ml, plasma, while viremic HIV⁺ subjects had a median viral load of 50,800 copies/ml, plasma (range, 22,900 to 341,000 copies). Median CD4⁺ T cell counts were 781 and 639 in Elite Controllers and viremic HIV⁺ subjects, respectively. mDCs and pDCs were identified in peripheral blood, distinguishable by mutually exclusive expression of the cell-surface integrin, CD11c, and the IL-3Rα chain, CD123, respectively (Fig. 1A and B). pDCs were further confirmed by expression of blood dendritic cell antigen 2 (BDCA-2). Full phenotypes identified pDCs as CD3⁻CD14⁻CD20⁻HLA-DR⁺CD123⁺CD11c⁻BDCA-2⁺ and mDCs as CD3⁻CD14⁻CD20⁻HLA-DR⁺CD123⁻CD11c⁺ (the full antibody list is in Supplementary Table S1; Supplementary Data are available online at www.liebertpub.com/aid). In viremic HIV⁺ subjects and, surprisingly, in Elite Controllers, 3-fold and 2-fold losses in circulating pDCs frequencies were observed compared to seronegative controls (Fig. 1C). However, by comparison mDCs were not significantly different in any group, suggesting the loss of pDCs was cell specific. To assess if the recent phenomenon of pDC gut trafficking during SIV infection is recapitulated in infected humans we evaluated α₄β₇ expression on both DC subpopulations and found increased frequencies of α₄β₇ expression in both viremic HIV⁺ patients and Elite Controllers on pDCs but not mDCs (Fig. 1B and D). A similar phenomenon was observed for total surface α₄β₇ expression, as indicated by median fluorescence intensities (Fig. 1E).

FIG. 1.

Plasmacytoid dendritic cells (pDCs) but not myeloid dendritic cells (mDCs) are reduced in peripheral blood of HIV-1 viremic patients and Elite Controllers and have a gut-homing phenotype. (A) Representative gating strategy for human pDCs among live peripheral blood mononuclear cells. (B) Frequencies of pDCs (left panel) and mDCs (right panel) in HIV-naive patients (HIV⁻), viremic HIV patients (vHIV⁺), and Controllers. (C) Percentage of α₄β₇ ⁺ pDCs (left panel) and mDCs (right panel) in HIV-naive patients, viremic progressors, and controllers. (D) Median fluorescence intensity (MFI) of α₄β₇ expression on pDCs (left panel) and mDCs (right panel) in HIV-naive patients, viremic progressors, and controllers. (E) Total surface α₄β₇ expression. *p<0.05; **p<0.01; ***p<0.001; Mann–Whitney U test.

Since inflammatory cytokines such as IFN-α can induce apoptosis, generalized immune activation, and may be associated with breakdown of the gut mucosae and microbial translocation, we next evaluated whether measures of pDC gut homing were associated with systemic markers of chronic immune activation. Interestingly, α₄β₇ expression positively correlated, whereas declining frequencies of circulating pDCs (presumably homing to the gut) negatively correlated with HLA-DR and Ki67 expression in T cells as markers of immune activation (Table 1). No association, however, was found with CD4⁺ T cell counts or viral load. These data suggest that HIV disease induces gut trafficking of pDCs, which is in turn linked to immune activation and may occur independently of virus replication.

Table 1.

Correlation of Plasmacytoid Dendritic Cell Frequency and α ₄ β ₇ Expression with Markers of Immune Activation

	VL	CD4 count	HLA-DR⁺ CD4	HLA-DR⁺ CD8	Ki67⁺ CD4	Ki67⁺ CD8
% pDC in blood	R=−0.178	R=0.379	R=−0.509	R=−0.609	R=−0.278	R=−0.262
	p=0.454	p=0.454	p=0.004	p=0.001	p=0.136	p=0.162
% α₄β₇+pDCs	R=0.031	R=−0.277	R=0.435	R=0.504	R=0.417	R=0.367
	p=0.907	p=0.294	p=0.016	p=0.005	p=0.022	p=0.05

VL, viral load; pDC, plasmacytoid dendritic cell.

Despite its importance in trafficking and maintaining pDCs in the gastrointestinal tract, the mechanism of α₄β₇ imprinting remains unknown. Whether this occurs in the gastrointestinal tract (GI) or on de novo pDCs developing in the bone marrow is unclear. Recent reports indicate that α₄β₇ expression is well-correlated with an actual accumulation of the cells in the GI tract in both HIV-infected humans and macaques models,^11,12,18,19 and thus measuring α₄β₇ expression in blood pDCs could be a useful surrogate biomarker to predict gut homing and potentially ongoing immune activation.

Footnotes

Acknowledgments

This work was supported by an amFAR research grant (108547-53-RGRL) to R.K.R.

Author Disclosure Statement

No competing financial interests exist.

References

Donaghy

, Pozniak

, Gazzard

, et al.: Loss of blood CD11c(+) myeloid and CD11c(-) plasmacytoid dendritic cells in patients with HIV-1 infection correlates with HIV-1 RNA virus load. Blood, 2001; 98:2574–2576.

Feldman

, Stein

, Amrute

, et al.: Decreased interferon-alpha production in HIV-infected patients correlates with numerical and functional deficiencies in circulating type 2 dendritic cell precursors. Clin Immunol, 2001; 101:201–210.

Patterson

, Rae

, Hockey

, et al.: Plasmacytoid dendritic cells are highly susceptible to human immunodeficiency virus type 1 infection and release infectious virus. J Virol, 2001; 75:6710–6713.

Fitzgerald-Bocarsly

and Jacobs

: Plasmacytoid dendritic cells in HIV infection: Striking a delicate balance. J Leukoc Biol, 2010; 87:609–620.

Reeves

and Fultz

: Disparate effects of acute and chronic infection with SIVmac239 or SHIV-89.6P on macaque plasmacytoid dendritic cells. Virology, 2007; 365:356–368.

Brown

, Trichel

, and Barratt-Boyes

: Parallel loss of myeloid and plasmacytoid dendritic cells from blood and lymphoid tissue in simian AIDS. J Immunol, 2007; 178:6958–6967.

Brown

, Wijewardana

, Liu

, and Barratt-Boyes

: Rapid influx and death of plasmacytoid dendritic cells in lymph nodes mediate depletion in acute simian immunodeficiency virus infection. PLoS Pathog, 2009; 5:e1000413.

Chehimi

, Campbell

, Azzoni

, et al.: Persistent decreases in blood plasmacytoid dendritic cell number and function despite effective highly active antiretroviral therapy and increased blood myeloid dendritic cells in HIV-infected individuals. J Immunol, 2002; 168:4796–4801.

Biancotto

, Grivel

, Iglehart

, et al.: Abnormal activation and cytokine spectra in lymph nodes of people chronically infected with HIV-1. Blood, 2007; 109:4272–4279.

10.

Nies-Kraske

, Schacker

, Condoluci

, et al.: Evaluation of the pathogenesis of decreasing CD4(+) T cell counts in human immunodeficiency virus type 1-infected patients receiving successfully suppressive antiretroviral therapy. J Infect Dis, 2009; 199:1648–1656.

11.

Reeves

, Evans

, Gillis

, et al.: SIV infection induces accumulation of plasmacytoid dendritic cells in the gut mucosa. J Infect Dis, 2012; 206:1462–1468.

12.

Kwa

, Kannanganat

, Nigam

, et al.: Plasmacytoid dendritic cells are recruited to the colorectum and contribute to immune activation during pathogenic SIV infection in rhesus macaques. Blood, 2011; 118:2763–2773.

13.

Lehmann

, Lafferty

, Garzino-Demo

, et al.: Plasmacytoid dendritic cells accumulate and secrete interferon alpha in lymph nodes of HIV-1 patients. PLoS One, 2010; 5:e11110.

14.

Nascimbeni

, Perie

, Chorro

, et al.: Plasmacytoid dendritic cells accumulate in spleens from chronically HIV-infected patients but barely participate in interferon-alpha expression. Blood, 2009; 113:6112–6119.

15.

Diop

, Ploquin

, Mortara

, et al.: Plasmacytoid dendritic cell dynamics and interferon-alpha production during SIV infection with a non-pathogenic outcome. J Virol, 2008; 82:5145–5152.

16.

Yan

, Sabbaj

, Bansal

, et al.: HIV-specific CD8+ T cells from elite controllers are primed for survival. J Virol, 2013; 87:5170–5181.

17.

Bansal

, Jackson

, West

, et al.: Multifunctional T-cell characteristics induced by a polyvalent DNA prime/protein boost human immunodeficiency virus type 1 vaccine regimen given to healthy adults are dependent on the route and dose of administration. J Virol, 2008; 82:6458–6469.

18.

Lehmann

, Jung

, Forster

, et al.: Longitudinal analysis of distribution and function of plasmacytoid dendritic cells in peripheral blood and gut mucosa of HIV infected patients. J Infect Dis, 2014; 209:940–949.

19.

, Gillis

, Johnson

, and Reeves

: Multi-functional plasmacytoid dendritic cells redistribute to gut tissues during simian immunodeficiency virus infection. Immunology, 2013; 140:244–249.

Supplementary Material

Please find the following supplemental material available below.

For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.

For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.

0.02 MB

0.00 MB