TAG HIV Basic Science, Vaccines, and Cure Project Blog

Trends in Immunology: Free Access Issue on Immune Senescence

The journal Trends in Immunology has published a special issue on immune senescence, and access to all articles is being offered free of charge (thanks to the sponsorship of the National Institutes of Health). The parallels between immune senescence in uninfected elderly individuals and people with HIV are increasingly well-recognized and include:

• Depletion of naive CD4 and CD8 T cells
• Involution of the thymus and decreased thymic output
• Skewing of the CD4:CD8 T cell ratio
• Accumulation of dysfunctional CD8 T cells lacking the CD28 co-stimulatory molecule
• Decreased responses to new immunizations
• Decreased recall (memory) T responses to common antigens

Research into immune senescence and its association with illness and mortality has gained considerably more attention recently, and may have the potential to offer important insights into the pathogenesis of HIV infection. Additional background on the topic, along with a link to a webcast presentation by immune senescence expert Rita Effros, can be found in a post from January of this year, "Accelerated Aging of the Immune System in HIV Infection."

Low Viral Loads in Elite Controllers

Several years ago, Bruce Walker and colleagues from Partners AIDS Research in Boston launched the largest ever effort to research immunological control of HIV infection, the international HIV controllers study. The project is now proceeding under the aegis of the new Ragon Institute of Massachusetts General Hospital, created recently thanks to the generous support of the Ragon family. The HIV controllers study is collecting blood samples from both elite controllers, defined based on having a viral load less than 50 copies in the absence of any treatment, and viremic controllers, defined based on a viral load of between 50 and 2000 copies in the absence of treatment. A number of papers have been published already describing results from the study, and the latest has just appeared online in the Journal of Infectious Diseases.

The purpose of this investigation was to quantify the level of viral load among elite controllers using an ultra-sensitive assay that can measure down to a single copy of HIV RNA per ml. Among 90 individuals studied, the median viral load level was just 2 copies/mL. A longitudinal analysis of a subset of 31 participants demonstrated that 2-5-fold fluctuations in viral load were common. In terms of the relationship with immune responses, the researchers document a significant correlation between the breadth and potency of neutralizing antibody responses against HIV and the level of viral load detected. The breadth and magnitude of HIV-specific CD8 T cell responses, as measured solely by interferon-gamma production, did not show a correlation with viral load at these low levels.  

An analysis of the relationship between viral load and the slope of CD4 decline was also conducted. The median time for which multiple CD4 measurements were available was 3.6 years (range 1 – 17.3), and the median number of measurements per person was seven. Although immune activation has previously been associated with CD4 T cell declines in elite controllers, in this analysis the median value of the slope per year was +11 cells/mm3.

Due the limited duration of follow up and number of measurements, the researchers note that in many cases the values were not significantly different from zero, indicating CD4 T cell counts were essentially stable over time. To try and get a better sense of which participants might be experiencing significant changes in CD4 counts, the researchers subsequently focused on individuals whose slopes were statistically different from zero. These analyses revealed that 5 out of 27 individuals (19%) with viral load levels less than 1 copy/mL experienced significant CD4 T cell count increases over time. The same was true for only 3 out of 50 individuals (3%) with HIV viral loads greater than 1 copy/mL. Conversely, 8 individuals experienced significant CD4 T cell declines, and all showed viral load levels over 1 copy/mL. An overall examination of the link between viral load level and CD4 T cell count slope showed a weak but significant correlation, with higher viral loads associated with CD4 T cell decline (r=-0.23; p=0.04).

Overall, the results suggest that the use of a viral load assay that is 250-fold more sensitive than those currently commercial available can reveal important information regarding elite control of HIV.

The Journal of Infectious Diseases 2009;200:000–000

DOI: 10.1086/605446

MAJOR ARTICLE

Persistent Low‐Level Viremia in HIV‐1 Elite Controllers and Relationship to Immunologic Parameters

Florencia Pereyra,1,2 Sarah Palmer,3,7 Toshiyuki Miura,1,6 Brian L. Block,1 Ann Wiegand,3 Alissa C. Rothchild,1 Brett Baker,1 Rachel Rosenberg,1 Emily Cutrell,1 Michael S. Seaman,4 John M. Coffin,5 and Bruce D. Walker1,6

1The Ragon Institute of Massachusetts General Hospital, Massachusetts Institute of Technology, and Harvard, 2Brigham and Women’s Hospital, Division of Infectious Diseases, 4Division of Viral Pathogenesis, Beth Israel Deaconess Medical Center, and 5Department of Molecular Biology and Microbiology, Tufts University, Boston Massachusetts; 3HIV Drug Resistance Program, National Cancer Institute, National Institutes of Health, Frederick, and 6Howard Hughes Medical Institute, Chevy Chase, Maryland; and 7Virology Department, Swedish Institute for Infectious Disease Control and Karolinska Institute, Solna, Sweden

Background. Human immunodeficiency virus type 1 (HIV‐1) elite controllers are able to control virus replication to levels below the limits of detection by commercial assays, but the actual level of viremia in these individuals is not well defined. Here, we quantify plasma HIV‐1 RNA in elite controllers and correlate this with specific immunologic parameters.

Methods. Plasma HIV‐1 RNA levels were quantified in 90 elite controllers with use of a real time reverse‐transcriptase polymerase chain reaction assay with a sensitivity of 0.2 copies/mL. HIV‐1–specific immune responses and longitudinal CD4+ T cell counts were examined.

Results. The median plasma HIV‐1 RNA level was 2 copies/mL (interquartile range, 0.2–14 copies/mL). A longitudinal analysis of 31 elite controllers demonstrated 2–5–fold fluctuations in viremia in the majority of individuals; 6 had persistent levels below 1 copy/mL. Viremia correlated directly with HIV‐1–specific neutralizing antibodies and Western blot reactivity but not with CD8+ T cell responses. Absolute CD4+ T cell decrease was more common among individuals with detectable viremia (p=.04).

Conclusions. Low‐level viremia is present in the majority of elite controllers and is associated with higher HIV‐1–specific antibody responses. Absolute CD4+ T cell loss is more common among viremic individuals, suggesting that even very low‐level viremia has negative consequences over time.

Received 29 January 2009; accepted 21 April 2009; electronically published 4 August 2009.

Reprints or correspondence: Florencia Pereyra, MD, Ragon Institute, Massachusetts General Hospital, 149 13th St, Charlestown, MA 02129

Potential conflicts of interest: none reported

Presented in part: 15th Conference on Retroviruses and Opportunistic Infections, Boston, Massachusetts, February 2008 (abstract 349).

Financial support: National Institutes of Health (AI28568 and AI30914 to B.D.W.), the Bill and Melinda Gates Foundation, the International AIDS Vaccine Initiative, and the Mark and Lisa Schwartz Foundation.

Immune Surveillance Below the Radar: Study Offers Explanation for Acyclovir’s Failure to Reduce HIV Risk

In a recent post on the Merck vaccine trial, I mentioned a new study from Larry Corey’s research group addressing the relationship between HSV-2 infection and enhanced susceptibility to HIV. The data were presented by Dr. Corey at the Keystone conference in March and were published online by Nature Medicine yesterday. The background to the work is that HSV-2 infection has been consistently associated with a 2 to 3-fold increased risk of acquiring HIV; a meta-analysis published in 2006 reported a relative risk of 3.1, 2.7 and 1.7 for women, heterosexual men and men who have sex with men, respectively. Recently analyzed data from the Merck vaccine trial are consistent with these findings in that HSV-2-infected participants were found to have approximately double the risk of acquiring HIV infection during the study. 

The mechanism by which HSV-2 infection increases HIV acquisition risk is not so clear, however, and has been the subject of debate. The general view is that local inflammation and damage to the integrity of the genital mucosa are plausible ways that HSV-2 infection may increase the chances of HIV transmission. This view led to the logical proposition that suppressing HSV-2 with chronic acyclovir treatment might be a means to also reduce the risk of HIV acquisition. Several large trials have now explored this hypothesis and, while acyclovir treatment was effective at reducing symptomatic HSV-2 reactivations, it did not reduce the incidence of HIV infection. 

The new study from Larry Corey’s laboratory set out to try and shed light on these trial results. The same group of researchers has previously shown that immune surveillance of HSV-2 is a far more active process than many had surmised. In a 2007 paper in the Journal of Experimental Medicine, they demonstrated that HSV-2-specific CD8+ T cells gather at sites of subclinical HSV-2 reactivation in the genital skin and persist for at least several months after HSV-2 DNA is no longer detectable. A year later in the Journal of Infectious Diseases, they published results of an intensive study in which participants took oral and anogenital swabs four times a day for 60 days. Analyzing the swabs for HSV-2 DNA, the researchers found that subclinical reactivations were frequent and typically lasted less than 12 hours, showing that there is an ongoing and dynamic effort on the part of the immune system to keep HSV-2 suppressed. 

These findings led the researchers to suspect that perhaps elevated levels of activated HSV-2-specific CD4 T cells would be present in the genital mucosa even during chronic acyclovir treatment, and this is exactly what they report in the Nature Medicine paper. The study initially took biopsies of genital skin during an acute, clinically symptomatic lesion and 2, 4 and 8 weeks after healing. Four participants subsequently initiated chronic suppressive acyclovir treatment at the start of their next symptomatic episode and had biopsies taken at 2, 4, 8, 16 & 20 weeks after healing. In all cases, control biopsies were obtained from an unaffected area of genital skin at each timepoint. 

During an acute HSV-2 lesion, the researchers found a “massive localized infiltration” of cells. Mean numbers of CD4 and CD8 T cells per mm² of skin were 655 and 618, respectively, compared to 68 and 55 per mm² of the unaffected skin sample. Although follow-up biopsies documented gradual clearance of HSV-2 and a decline in inflammation, elevated numbers of CD4 and CD8 T cells remained present locally for months. Eight weeks after healing, a median of 8-fold more CD4 T cells were present at the affected versus the unaffected site. Furthermore, even after 20 weeks of acyclovir treatment, the number of CD4 T cells remained significantly elevated. Additional analyses illustrated that the majority of these CD4 T cells expressed CCR5 and their presence was also associated with a significant increase in the numbers of DC-SIGN-expressing dendritic cells. The enrichment of CCR5-expressing CD4 T cells and DC-SIGN-expressing dendritic cells at the former lesion site was not significantly altered by chronic acyclovir treatment. 

The researchers acknowledge in the conclusion to the paper that the number of participants was small, but nevertheless state: “we feel that our central finding – that HSV reactivation leaves a residual inflammatory response not appreciated clinically – is typical of HSV-2 genital lesions.” They also note that “the wide anatomical distribution of HSV-2 in the male and female genital tract underscores the importance that these localized reservoirs of inflammatory cells are likely to have in HIV acquisition.” 

Nature Medicine

Published online: 2 August 2009 | doi:10.1038/nm.2006 

Persistence of HIV-1 receptor–positive cells after HSV-2 reactivation is a potential mechanism for increased HIV-1 acquisition 

Jia Zhu1,2, Florian Hladik1,3,4,6, Amanda Woodward1,3,6, Alexis Klock1,2, Tao Peng1,2, Christine Johnston1,3, Michael Remington2, Amalia Magaret1,2, David M Koelle1,2,3, Anna Wald1,2,3,5 & Lawrence Corey1,2,3,5 

Abstract 

To explore the mechanism by which herpes simplex virus (HSV)-2 infection is related to HIV-1 acquisition, we conducted in situ analysis of the cellular infiltrate from sequential biopsies of HSV-2 lesions from patients on and off antiviral therapy. CD4+ and CD8+ T cells and a mixed population of plasmacytoid and myeloid dendritic cells (DCs), including cells expressing the C-type lectin receptor DC-SIGN, persisted at sites of HSV-2 reactivation for months after healing, even with daily antiviral therapy. The CD4+ T cells that persisted reacted to HSV-2 antigen, were enriched for expression of the chemokine receptor CCR5, and were contiguous to DCs expressing the interleukin-3 receptor CD123 or DC-SIGN. Ex vivo infection with a CCR5-tropic strain of HIV-1 revealed greater concentrations of integrated HIV-1 DNA in cells derived from healed genital lesion biopsies than in cells from control skin biopsies. The persistence and enrichment of HIV receptor–positive inflammatory cells in the genitalia help explain the inability of anti–HSV-2 therapy to reduce HIV acquisition. 

1. Vaccine and Infectious Disease Institute, Fred Hutchinson Cancer Research Center, Seattle, Washington, USA.

2. Department of Laboratory Medicine, University of Washington, Seattle, Washington, USA.

3. Department of Medicine, University of Washington, Seattle, Washington, USA.

4. Department of Obstetrics and Gynecology, University of Washington, Seattle, Washington, USA.

5. Department of Epidemiology, University of Washington, Seattle, Washington, USA. 

6. These authors contributed equally to this work.