A slew of new studies presented at CROI revealed that a recently discovered subset of CD4 T cells – dubbed T-helper type 17 or Th17 cells for short – appear preferentially depleted in HIV and pathogenic SIV infections. Th17 cells were identified as a separate lineage of CD4 T cells in 2005; they are characterized by the production of the cytokines IL-17 and IL-22 and have been shown to be involved in both pro-inflammatory autoimmune diseases, and in protection against certain pathogens such as Klebsiella pneumonia and Salmonella typherium. The cytokines IL-17 and IL-22 also play a role in protecting mucosal sites from pathogens and maintaining the integrity of the mucosal barrier. Prior studies conducted outside of the HIV field indicate that Th17 cells make up less than 1% of circulating CD4 T cells in healthy people with slightly higher levels (~1-2%) found in the gut. The cytokine IL-23 appears involved in the development of Th17 CD4 T cells, which arise – like other Th subsets – from the activation of naïve CD4 T cells. The development of Th17 cells is also suppressed under certain conditions, particularly by the presence of type I & II interferons (e.g. alpha interferon and interferon gamma).
At CROI, Mirka Paiardini presented data showing that Th17 cell levels were similar or slightly increased in the blood of HIV-infected individuals compared to uninfected controls. But in the gut, the percentage of CD4 T cells producing IL-17 after in vitro stimulation (with the pan-T cell stimulating agents PMA and ionomycin or anti-CD3) was significantly lower in people with HIV and antiretroviral therapy did not fully restore the proportion of Th17 cells to those seen in uninfected individuals. Paiardini also showed that there was a significant inverse correlation between immune activation levels in both gut and peripheral blood (as assessed by the proportion of CD4 T cells expressing the proliferation marker Ki67) and the proportion of Th17 cells in the gut.
It has recently been reported that non-pathogenic SIV infection of sooty mangabeys is associated with gut CD4 T cell loss, so Paiardini measured Th17 levels in the gut of mangabeys to ascertain if they were also depleted, as they appear to be in HIV-infected humans. He reported that the proportion of gut Th17 cells remained stable in sooty mangabeys despite the overall reduction in the percentage of gut CD4 T cells. Paiardini hypothesized that preservation of gut Th17 cells in mangabeys preserves the integrity of the mucosal barrier and thus prevents translocation of microbial products and systemic immune activation, supporting this hypothesis with slides demonstrating the health of the epithelial barrier in SIV-infected animals. However it is perhaps worth noting that – as is so often the case with research results – cause and effect are not necessarily easy to tease apart. Immune activation in HIV infection is associated with elevated levels of type I interferons, so if Th17 cells are generated dynamically in the gut, immune activation would be expected to inhibit their differentiation and thus reduce the proportion of Th17 cells. In sooty mangabeys, where SIV infection does not result in persistent immune activation, Mark Feinberg has shown that type I interferons are not upregulated, suggesting that preservation of Th17 cells could be a consequence rather than a cause of the observed lack of activation.
Valentina Cecchinato from Genoveffa Franchini’s lab at the National Cancer Institute sounded a similar theme to Paiardini, showing that in a study of SIV-infected rhesus macaques, the percentage of Th17 cells was reduced in the gut while the proportion of Th1 (interferon gamma producing) CD4 T cells was unaffected. In contrast, both populations were depleted from the peripheral blood. Further analyses revealed an inverse correlation between SIV viral load and percentages of Th17 cells in the colon and rectum. But when Cecchinato looked for evidence of microbial translocation by measuring blood levels of lipopolysaccharides (LPS), there was no correlation between LPS levels and the proportion of Th17 cells in the gut (or anywhere else). This finding may be explained by the technical challenges associated with LPS measurement, but it could also be an indication that Th17 loss is an effect rather than a cause of virus-driven immune activation in these animals.
In the last of three talks on the topic, David Favre from UCSF showed that the proportion of Th17 cells also declines at mucosal sites in SIV-infected pigtailed macaques. Favre focused on the balance between T regulatory (Treg) cells (another CD4 T cell subset with the ability to suppress immune responses) and Th17 cells, noting that the balance is maintained in African green monkeys, a species that – like sooty mangabeys – does not develop immunodeficiency as a result of SIV infection. In contrast, the loss of Th17 cells dysregulated the Th17/Treg balance in pigtailed macaques.
At the close of CROI, conference chair Mario Stevenson highlighted these presentations as representing an important advance in the understanding of HIV pathogenesis. However, until the cause and effect relationship between immune activation and Th17 cell decreases is better understood, the data should probably be interpreted with caution.
Webcasts for the presentations are available on the CROI website (the fourth oral abstract session on Wednesday).
Study abstracts:
15th Conference on Retroviruses & Opportunistic Infections
Session 36 Oral Abstracts
New Insights into Mechanisms of Viral Pathogenecity
Abstract #115
Preferential Loss of Th17 CD4 T Cells in the Gastrointestinal Tract of HIV-infected Individuals but Not SIV-infected Sooty Mangabeys
Barbara Cervasi*1, J Brenchley2, M Paiardini1, S Gordon1, A Asher2, I Frank1, J Else3, D Douek2, and G Silvestri1
1Univ of Pennsylvania, Philadelphia, US; 2Vaccine Res Ctr, NIAID, NIH, Bethesda, MD, US; and 3Yerkes Natl Primate Res Ctr, Emory Univ, Atlanta, GA, US
Background: A new subset of CD4 T helper cells, designated Th17, has been recently identified, leading to a reappraisal of the classically proposed dichotomy in which memory CD4 T cells belong exclusively to either a Th1 or Th2 lineage. While Th17 cells may be involved in the pathogenesis of autoimmune diseases, emerging data suggest that they are important in mucosal immunity to extra-cellular bacteria infection. Here we examined the frequency, antigen specificity, functionality, phenotype, and infection frequency of Th17 cells in the peripheral blood and gastrointestinal tract of HIV-infected and uninfected humans and simian immunodeficiency virus (SIV) -infected and uninfected sooty mangabeys, a natural, non-progressing host species of SIV infection.
Methods: Peripheral blood- and gastrointestinal tract-derived cells were obtained from 30 HIV-infected individuals and 15 healthy controls; peripheral blood, gastrointestinal, and lymph node cells were also obtained from 12 naturally SIV-infected and 10 uninfected sooty mangabeys. Phenotypical and functional characterization of Th17 cells was performed by 10-color flow cytometry.
Results: Interleukin (IL) -17+, interferon (IFN) -g–, IL-4–, IL-2+, IL23R+ memory CD4+ Th17 cells can be identified in all the examined tissues of humans and sooty mangabeys, with the majority of Th17 cells being CCR5– in peripheral blood, but CCR5+ in the gastrointestinal tract. The main results of this study were: There is significant depletion of Th17 cells in the gastrointestinal tract of HIV-infected individuals than in uninfected individuals; SIV-infected sooty mangabeys maintain healthy frequencies of Th17 cells in the the gastrointestinal tract; Th17 cells are infected by HIV in vivo, but not preferentially so compared to Th1 cells; and human Th17 cells are specific for bacterial and fungal antigens, but not common viral antigens.
Conclusions: The preferential loss of Th17 in the the gastrointestinal tract of HIV-infected individuals represents a new mechanism of mucosal immune dysfunction during HIV infection. These data thus further elucidate the immunodeficiency of HIV disease and may provide a mechanistic basis for the enteropathy and associated microbial translocation that characterize HIV infection. Conversely, maintenance of gastrointestinal Th17 cells may account for the non-progressive nature of non-pathogenic SIV infection in sooty mangabeys.
Abstract #116
Preferential Loss of Th17 T Cells at Mucosal Sites Predicts AIDS Progression in Simian Immunodeficiency Virus-infected Macaques
Valentina Cecchinato*1, C Trindade1, J M Heraud1, A Laurence2, J Brenchley3, E Tryniszewska1,4, D Venzon1, D Douek3, J O'Shea2, and G Franchini1
1NCI, NIH, Bethesda, MD, US; 2Natl Inst of Arthritis and Musculoskeletal and Skin Diseases, NIH, Bethesda, MD, US; 3NIAID, NIH, Bethesda, MD, US; and 4Med Univ of Bialystok, Poland
Background: Depletion of CD4+ T cells in the gut is necessary, but not sufficient to cause AIDS in animal models, raising the possibility that differential depletion of CD4+ T cell subtypes may be important. Th17 is a recently identified lineage of effector CD4+ T-helper interleukin-17 (IL-17) -secreting cells, important in mucosal immunity to extracellular bacteria. Here we investigated whether the frequency of Th17 lymphocytes is modified during simian immunodeficiency virus (SIV) infection, particularly at mucosal sites.
Methods: We enrolled in the study 31 rhesus macaques, either naive or SIV-infected. Blood and lymphoid tissues were collected 2 weeks after infection or at the time of necropsy. The frequency of CD4+IL-17+ cells in tissues was determined by intracellular cytokines staining following stimulation with phorbol 12-myristate 13-acetate (PMA) and ionomycin. Flow cytometric analysis was performed to determine Th17 phenotype. Quantification of SIV Gag DNA in sorted memory CD4 T cells was performed by quantitative polymerase chain reaction (PCR). Human CD4+ T cells were infected in vitro with SIVmac251 and analyzed for IL-17 and p27 expression. Differences between groups were assessed by t-test. The Spearman rank correlation method was used to determine the correlation between the frequency of Th17 in tissues and viral load in plasma.
Results: Our data demonstrate that Th17 cells express CCR5 and can be infected in vitro and in vivo. We found that healthy macaques have a significantly higher number of Th17 T cells in the gut than in systemic tissues. Importantly, in the SIVmac251 model, Th17 cells are preferentially depleted within the first weeks of infection and their frequency is not restored to normal levels over time. Furthermore, we found that SIVmac251-infected macaques that could effectively control viral replication maintain normal levels of Th17 T cells in all tissues, and that there is a negative correlation between Th17 cell frequency at mucosal sites and plasma virus level, suggesting their importance in HIV/SIV pathogenesis.
Conclusions: Because Th17 cells play a central role in innate and adaptive immune response to extracellular bacteria, these data provide a rational explanation for the chronic enteropathy in HIV infection. Thus, therapeutic approaches to reconstitute Th17 numbers and/or function may be of benefit in HIV infection.
Abstract #117LB
Primary SIV Infection Causes Rapid Loss of the Balance between TH17 and T Regulatory Cell Populations in Pathogenic Infection of Non-human Primates
David Favre*1, S Lederer2, B Kanwar1, Z M Ma3, S Proll2, Z Kasakow1, C Miller3, M Katze2, and J McCune1
1Univ of California, San Francisco, US; 2Univ of Washington, Seattle, US; and 3Univ of California, Davis, US
Background: Progression to AIDS is observed after simian immunodeficiency virus (SIV) infection of some but not all non-human primates. To better define the basis for this dichotomy, we performed a comparative analysis of non-pathogenic infection of the African green monkey and pathogenic infection of the Asian pigtailed macaque. We hypothesized that a critical distinction between pathogenic and nonpathogenic SIV infections may lie in a shift in the equilibrium between pro- and anti-inflammatory host immune responses during acute infection.
Methods: To address this hypothesis, we investigated early virologic, immunologic, and gene expression events that occurred in the peripheral blood and in lymphoid organs, including the colonic mucosa, after acute infection of pigtailed macaques and African green monkeys with the same SIVagm.sab92018 isolate. In particular, we explored the absolute number and relative frequency of TH17 cells, which produce the pro-inflammatory cytokine interleukin (IL) -17, and FoxP3+ T regulatory cells (Treg), the function of which is instead immunosuppressive.
Results: After infection with SIVagm.sab92018, both African green monkeys and pigtailed macaques developed high peak and set point viral loads, but only pigtailed macaque showed generalized CD4+ T cell depletion and sustained immune activation and inflammation. In addition, the following novel features were observed. First, signs of chronic T cell activation and inflammation were detected in the colonic mucosa of both species, while systemic immune activation (e.g., in the blood and lymph node) was only sustained in pigtailed macaques. Second, an increase in the frequency of FoxP3+ Treg was detected early in the blood and in lymph nodes after infection of African green monkeys, but only at later time points in the colon of both African green monkeys and pigtailed macaques. Finally, and unexpectedly, the TH17 population was maintained in infected African green monkeys, but rapidly lost during the course of progressive SIV disease in pigtailed macaques.
Conclusions: These results suggest that balanced representation of both Treg and TH17 cells plays a unique and possibly determinative role in the setting of acute lentiviral infection. Because IL-17 acts to preserve the integrity of the mucosal barrier, thus enhancing host defenses against microbial agents, maintenance of robust TH17 function in mucosal tissue during SIV infection may decrease systemic inflammation and disease progression by preventing the immune activation that would otherwise occur after microbial translocation and spread.
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