Immune nonresponder (INR) HIV-1–infected subjects are characterized by their inability to reconstitute the CD4+ T cell pool after antiretroviral therapy. This is linked to poor clinical outcome. Mechanisms underlying immune reconstitution failure are poorly understood, although, counterintuitively, INRs often have increased frequencies of circulating CD4+ T cells in the cell cycle. While cycling CD4+ T cells from healthy controls and HIV+ patients with restored CD4+ T cell numbers complete cell division in vitro, cycling CD4+ T cells from INRs do not. Here, we show that cells with the phenotype and transcriptional profile of Tregs were enriched among cycling cells in health and in HIV infection. Yet there were diminished frequencies and numbers of Tregs among cycling CD4+ T cells in INRs, and cycling CD4+ T cells from INR subjects displayed transcriptional profiles associated with the impaired development and maintenance of functional Tregs. Flow cytometric assessment of TGF-β activity confirmed the dysfunction of Tregs in INR subjects. Transcriptional profiling and flow cytometry revealed diminished mitochondrial fitness in Tregs among INRs, and cycling Tregs from INRs had low expression of the mitochondrial biogenesis regulators peroxisome proliferator–activated receptor γ coactivator 1-α (PGC1α) and transcription factor A for mitochondria (TFAM). In vitro exposure to IL-15 allowed cells to complete division, restored the expression of PGC1α and TFAM, and regenerated mitochondrial fitness in the cycling Tregs of INRs. Our data suggest that rescuing mitochondrial function could correct the immune dysfunction characteristic of Tregs in HIV-1–infected subjects who fail to restore CD4+ T cells during antiretroviral therapy.
Souheil-Antoine Younes, Aarthi Talla, Susan Pereira Ribeiro, Evgeniya V. Saidakova, Larisa B. Korolevskaya, Konstantin V. Shmagel, Carey L. Shive, Michael L. Freeman, Soumya Panigrahi, Sophia Zweig, Robert Balderas, Leonid Margolis, Daniel C. Douek, Donald. D. Anthony, Pushpa Pandiyan, Mark Cameron, Scott F. Sieg, Leonard H. Calabrese, Benigno Rodriguez, Michael M. Lederman
In the era of combined antiretroviral therapy (cART), lung diseases such as chronic bronchitis (CB) and COPD are common among persons living with HIV (PLWH), particularly smokers. Although smoking is highly prevalent among PLWH, HIV may be an independent risk factor for lung diseases; however, the role of HIV and cigarette smoke (CS) and their potential interaction in the development of chronic lung diseases among PLWH has not been delineated. To investigate this interaction, cynomolgus macaques were exposed to CS and/or simian-adapted human immunodeficiency virus (SHIV) and treated with cART. The development of CB and the lung functions were evaluated following CS±SHIV treatment. The results showed that in the lung, SHIV was a strong independent risk factor for goblet cell metaplasia/hyperplasia and mucus formation, MUC5AC synthesis, loss of tight junction proteins, and increased expression of Th2 cytokines/transcription factors. In addition, SHIV and CS synergistically reduced the lung function and increased the extrathoracic tracheal ring thickness. Interestingly, SHIV-infection generated significant numbers of HIV-gp120+ epithelial cells (HGECs) in small airways and alveoli and their numbers doubled in CS+SHIV-infected lungs. We conclude that even with cART, SHIV independently induces CB and pro-COPD changes in the lung and the effects are exacerbated by CS.
Hitendra S. Chand, Rodrigo Vazquez-Guillamet, Christopher M. Royer, Karin Rudolph, Neerad C. Mishra, Shashi P. Singh, Shah S. Hussain, Edward G. Barrett, Shannon Callen, Siddappa N. Byrareddy, Maria Cristina Vazquez Guillamet, Jawad Abukhalaf, Aryaz Sheybani, Vernat Exil, Veena Raizada, Hemant Agarwal, Madhavan Nair, Francois Villinger, Shilpa Buch, Mohan Sopori
BACKGROUND. Injectable depot medroxyprogesterone acetate (DMPA) is one of the most popular contraception methods in areas of high HIV seroprevalence. Evidence is accumulating that use of DMPA might be associated with an increased risk of HIV-1 acquisition by women; however, mechanisms of this association are not completely understood. The goal of this study was to gain insight into mechanisms underlying the possible link between use of DMPA and risk of HIV-1 acquisition, exploring transcription profiling of ectocervical tissues. METHODS. Healthy women received either DMPA (n = 31) or combined oral contraceptive (COC), which has not been linked to an increased risk of HIV acquisition (n = 32). We conducted a comparative microarray-based whole-genome transcriptome profiling of human ectocervical tissues before and after 6 weeks of hormonal contraception use. RESULTS. The analysis identified that expression of 235 and 76 genes was significantly altered after DMPA and COC use, respectively. The most striking effect of DMPA, but not COC, was significantly altered expression (mostly downregulation) of many genes strategically involved in the maintenance of mucosal barrier function; the alterations, as indicated by Ingenuity Pathway Analysis (IPA), were most likely due to the DMPA-induced estrogen deficiency. Furthermore, IPA predicted that transcriptome alterations related to ectocervical immune responses were in general compatible with an immunosuppressive effect of DMPA, but, in some women, also with an inflammatory-like response. CONCLUSION. Our results suggest that impairment of cervicovaginal mucosal integrity in response to DMPA administration is an important mechanism contributing to the potential increased risk of HIV-1 acquisition in DMPA users. TRIAL REGISTRATION. ClinicalTrials.gov NCT01421368. FUNDING. This study was supported by the United States Agency for International Development (USAID) under Cooperative Agreement GPO-A-00-08-00005-00.
Irina A. Zalenskaya, Neelima Chandra, Nazita Yousefieh, Xi Fang, Oluwatosin E. Adedipe, Suzanne S. Jackson, Sharon M. Anderson, Christine K. Mauck, Jill L. Schwartz, Andrea R. Thurman, Gustavo F. Doncel
Neutrophil extracellular traps (NETs) are involved in the pathogenesis of many infectious diseases, yet their dynamics and impact on HIV/SIV infection were not yet assessed. We hypothesized that SIV infection and the related microbial translocation trigger NET activation and release (NETosis), and investigated the interactions between NETs and immune cell populations and platelets. We compared and contrasted the levels of NETs between SIV-uninfected, SIV-infected, and SIV-infected antiretroviral-treated nonhuman primates. We also cocultured neutrophils from these animals with either peripheral blood mononuclear cells or platelets. Increased NET production was observed throughout SIV infection. In chronically infected animals, NETs were found in the gut, lung, liver, and in the blood vessels of kidney and heart. ART decreased NETosis, albeit above preinfection levels. NETs captured CD4+ and CD8+ T-cells, B-cells, and monocytes, irrespective of their infection status, potentially contributing to the indiscriminate generalized immune cell loss characteristic to HIV/SIV infection, and limiting the CD4+ T-cell recovery under ART. By capturing and facilitating aggregation of platelets, and through expression of increased tissue factor levels, NETs may also enhance HIV/SIV-related coagulopathy and promote cardiovascular comorbidities.
Ranjit Sivanandham, Egidio Brocca-Cofano, Noah Krampe, Elizabeth Falwell, Sindhuja Murali Kilapandal Venkatraman, Ruy M. Ribeiro, Cristian Apetrei, Ivona Pandrea
Long-lived HIV-1 reservoirs that persist despite antiretroviral therapy (ART) are a major impediment to a cure for HIV-1. We examined whether human liver macrophages (LMs), the largest tissue macrophage population, comprise an HIV-1 reservoir. We purified LMs from liver explants and included treatment with a T cell immunotoxin to reduce T cells to 1% or less. LMs were purified from 9 HIV-1–infected persons, 8 of whom were on ART (range 8–140 months). Purified LMs were stimulated ex vivo and supernatants from 6 of 8 LMs from persons on ART transmitted infection. However, HIV-1 propagation from LMs was not sustained except in LMs from 1 person taking ART for less than 1 year. Bulk liver sequences matched LM-derived HIV-1 in 5 individuals. Additional in vitro experiments undertaken to quantify the decay of HIV-1–infected LMs from 3 healthy controls showed evidence of infection and viral release for prolonged durations (>170 days). Released HIV-1 propagated robustly in target cells, demonstrating that viral outgrowth was observable using our methods. The t1/2 of HIV-1–infected LMs ranged from 3.8–55 days. These findings suggest that while HIV-1 persists in LMs during ART, it does so in forms that are inert, suggesting that they are defective or restricted with regard to propagation.
Abraham J. Kandathil, Sho Sugawara, Ashish Goyal, Christine M. Durand, Jeffrey Quinn, Jaiprasath Sachithanandham, Andrew M. Cameron, Justin R. Bailey, Alan S. Perelson, Ashwin Balagopal
Activation of HIV-1 reservoirs and induction of anti–HIV-1 T cells are critical to control HIV-1 rebound after combined antiretroviral therapy (cART). Here we evaluated in humanized mice (hu-mice) with persistent HIV-1 infection the therapeutic effect of TLR3 agonist and a CD40-targeting HIV-1 vaccine, which consists of a string of 5 highly conserved CD4+ and CD8+ T cell epitope-rich regions of HIV-1 Gag, Nef, and Pol fused to the C-terminus of a recombinant anti-human CD40 antibody (αCD40.HIV5pep). We show that αCD40.HIV5pep vaccination coadministered with poly(I:C) adjuvant induced HIV-1–specific human CD8+ and CD4+ T cell responses in hu-mice. Interestingly, poly(I:C) treatment also reactivated HIV-1 reservoirs. When administrated in therapeutic settings in HIV-1–infected hu-mice under effective cART, αCD40.HIV5pep with poly(I:C) vaccination induced HIV-1–specific CD8+ T cells and reduced the level of cell-associated HIV-1 DNA (or HIV-1 reservoirs) in lymphoid tissues. Most strikingly, the vaccination significantly delayed HIV-1 rebound after cART cessation. In summary, the αCD40.HIV5pep with poly(I:C) vaccination approach both activates replication of HIV-1 reservoirs and enhances the anti–HIV-1 T cell response, leading to a reduced level of cell-associated HIV-1 DNA or reservoirs. Our proof-of-concept study has significant implication for the development of CD40-targeting HIV-1 vaccine to enhance anti–HIV-1 immunity and reduce HIV-1 reservoirs in patients with suppressive cART.
Liang Cheng, Qi Wang, Guangming Li, Riddhima Banga, Jianping Ma, Haisheng Yu, Fumihiko Yasui, Zheng Zhang, Giuseppe Pantaleo, Matthieu Perreau, Sandra Zurawski, Gerard Zurawski, Yves Levy, Lishan Su
HIV post-treatment controllers (PTCs) represent a natural model of sustained HIV remission, but they are rare and little is known about their viral reservoir. We obtained 1450 proviral sequences after near-full-length amplification for 10 PTCs and 16 post-treatment non-controllers (NCs). Before treatment interruption, the median intact and total reservoir size in PTCs was 7-fold lower than in NCs, but the proportion of intact, defective and total clonally-expanded viral genomes was not significantly different between the two groups. Quantification of total, but not intact, proviral genome copies predicted sustained HIV remission as 81% of NCs, but none of the PTCs, had a total proviral genome >4 copies per million PBMCs. The results highlight the restricted intact and defective HIV reservoir in PTCs and suggest that total proviral genome burden could act as the first biomarker for identifying PTCs. Defective, but not intact, proviral copy numbers correlated with levels of cell-associated HIV RNA, activated NK cell percentages and both HIV-specific CD4+ and CD8+ responses. These results support the concept that defective HIV genomes lead to viral antigen production and interact with both the innate and adaptive immune systems.
Radwa Sharaf, Guinevere Q. Lee, Xiaoming Sun, Behzad Etemad, Layla M. Aboukhater, Zixin Hu, Zabrina L. Brumme, Evgenia Aga, Ronald J. Bosch, Ying Wen, Golnaz Namazi, Ce Gao, Edward P. Acosta, Rajesh T. Gandhi, Jeffrey M. Jacobson, Daniel Skiest, David M. Margolis, Ronald Mitsuyasu, Paul Volberding, Elizabeth Connick, Daniel R. Kuritzkes, Michael M. Lederman, Xu G. Yu, Mathias Lichterfeld, Jonathan Z. Li
HIV infection changes the lymph node (LN) tissue architecture, potentially impairing the immunologic response to antigenic challenge. The tissue-resident immune cell dynamics in virologically suppressed HIV+ patients on combination antiretroviral therapy (cART) are not clear. We obtained LN biopsies before and 10 to 14 days after trivalent seasonal influenza immunization from healthy controls (HCs) and HIV+ volunteers on cART to investigate CD4+ T follicular helper (Tfh) and B cell dynamics by flow cytometry and quantitative imaging analysis. Prior to vaccination, compared with those in HCs, HIV+ LNs exhibited an altered follicular architecture, but harbored higher numbers of Tfh cells and increased IgG+ follicular memory B cells. Moreover, Tfh cell numbers were dependent upon preservation of the follicular dendritic cell (FDC) network and were predictive of the magnitude of the vaccine-induced IgG responses. Interestingly, postvaccination LN samples in HIV+ participants had significantly (P = 0.0179) reduced Tfh cell numbers compared with prevaccination samples, without evidence for peripheral Tfh (pTfh) cell reduction. We conclude that influenza vaccination alters the cellularity of draining LNs of HIV+ persons in conjunction with development of antigen-specific humoral responses. The underlying mechanism of Tfh cell decline warrants further investigation, as it could bear implications for the rational design of HIV vaccines.
Eirini Moysi, Suresh Pallikkuth, Leslie R. De Armas, Louis E. Gonzalez, David Ambrozak, Varghese George, David Huddleston, Rajendra Pahwa, Richard A. Koup, Constantinos Petrovas, Savita Pahwa
BACKGROUND. The effect of a brief analytical treatment interruption (ATI) on the HIV-1 latent reservoir of individuals who initiate antiretroviral therapy (ART) during chronic infection is unknown. METHODS. We evaluated the impact of transient viremia on the latent reservoir in participants who underwent an ATI and at least 6 months of subsequent viral suppression in a clinical trial testing the effect of passive infusion of the broadly neutralizing Ab VRC01 during ATI. RESULTS. Measures of total HIV-1 DNA, cell-associated RNA, and infectious units per million cells (IUPM) (measured by quantitative viral outgrowth assay [QVOA]) were not statistically different before or after ATI. Phylogenetic analyses of HIV-1 env sequences from QVOA and proviral DNA demonstrated little change in the composition of the virus populations comprising the pre- and post-ATI reservoir. Expanded clones were common in both QVOA and proviral DNA sequences. The frequency of clonal populations differed significantly between QVOA viruses, proviral DNA sequences, and the viruses that reactivated in vivo. CONCLUSIONS. The results indicate that transient viremia from ATI does not substantially alter measures of the latent reservoir, that clonal expansion is prevalent within the latent reservoir, and that characterization of latent viruses that can reactivate in vivo remains challenging. TRIAL REGISTRATION. ClinicalTrials.gov NCT02463227 FUNDING. Funding was provided by the NIH.
D. Brenda Salantes, Yu Zheng, Felicity Mampe, Tuhina Srivastava, Subul Beg, Jun Lai, Jonathan Z. Li, Randall L. Tressler, Richard A. Koup, James Hoxie, Mohamed Abdel-Mohsen, Scott Sherrill-Mix, Kevin McCormick, E. Turner Overton, Frederic D. Bushman, Gerald H. Learn, Robert F. Siliciano, Janet M. Siliciano, Pablo Tebas, Katharine J. Bar
The human brain is an important site of HIV replication and persistence during antiretroviral therapy (ART). Direct evaluation of HIV infection in the brains of otherwise healthy individuals is not feasible; therefore, we performed a large-scale study of bone marrow/liver/thymus (BLT) humanized mice as an in vivo model to study HIV infection in the brain. Human immune cells, including CD4+ T cells and macrophages, were present throughout the BLT mouse brain. HIV DNA, HIV RNA, and/or p24+ cells were observed in the brains of HIV-infected animals, regardless of the HIV isolate used. HIV infection resulted in decreased numbers of CD4+ T cells, increased numbers of CD8+ T cells, and a decreased CD4+/CD8+ T cell ratio in the brain. Using humanized T cell–only mice (ToM), we demonstrated that T cells establish and maintain HIV infection of the brain in the complete absence of human myeloid cells. HIV infection of ToM resulted in CD4+ T cell depletion and a reduced CD4+/CD8+ T cell ratio. ART significantly reduced HIV levels in the BLT mouse brain, and the immune cell populations present were indistinguishable from those of uninfected controls, which demonstrated the effectiveness of ART in controlling HIV replication in the CNS and returning cellular homeostasis to a pre-HIV state.
Jenna B. Honeycutt, Baolin Liao, Christopher C. Nixon, Rachel A. Cleary, William O. Thayer, Shayla L. Birath, Michael D. Swanson, Patricia Sheridan, Oksana Zakharova, Francesca Prince, JoAnn Kuruc, Cynthia L. Gay, Chris Evans, Joseph J. Eron, Angela Wahl, J. Victor Garcia