HIV-1 tissue pathogenesis after antiretroviral therapy Immune activation is now considered to be a driving force of various human pathologies, including HIV-1 disease. Currently, a combination of antiretroviral therapy (ART) has proven to be efficient in suppressing HIV-1 replication. However, lengthy suppression of HIV-1 replication by ART is associated with an increased risk of complications, including neurological and cardiovascular diseases. These diseases seem to be related to the residual immune activation in patients undergoing ART. Cytokines may play an important role in this residual immune activation. To investigate mechanisms of his phenomenon it is necessary to develop an ex vivo laboratory-controlled system reflecting what happens in vivo. We have developed such a system. As an experimental model we used ex vivo human lymphoid tissues where critical events in HIV-1 infection occur in vivo. In the present work we evaluated 33 cytokines released by donor matched human lymphoid tissues ex vivo productively infected with HIV-1 over 16 days of infection and treated or not with the antiviral, ritonavir. We evaluated soluble and EV-associated cytokines. We found that HIV-1 infection led to upregulation of several key cytokines. EV-associated cytokines were more likely to be elevated than soluble ones. Antivirals suppressed HIV in infected tissues. However, most of the cytokines upregulated upon HIV infection remained upregulated in the absence of HIV replication. EV-associated cytokines were more likely to remain elevated in spite of ART. In conclusion, the phenomenon of residual immune activation after successful ART is reproduced in a laboratory experimental system with human lymphoid tissue ex vivo opening a way to study this phenomenon under controlled laboratory conditions. Another complication in HIV-1 infected individuals undergoing ART is a failure to reconstitute the CD4+ T cells (immune non-responders, INR). 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-beta 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 coactivator 1- (PGC1) and transcription factor A for mitochondria. In vitro exposure to IL-15 allowed cells to complete division, restored the expression of PGC1, 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 INRs and enhance immune restoration in these subjects. In conclusion, our analysis showed that HIV-1 infected lymphoid tissues ex vivo upregulated production of many cytokines, both free and EV-associated, and the majority of these cytokines remained upregulated despite suppression of viral replication by ART. Also, in spite of ART a number of patients do not reconstitute their immune system. Mechanisms of both of these phenomena can now be investigated under controlled laboratory conditions resulting in the development of new therapeutic strategies. 2. Cytomegalovirus and EVs in atherosclerosis: Immune activation Acute cardiovascular syndrome (ACS) is associated with a general activation of the immune system that includes activation of many cells, in particular monocytes and platelets, leading to destabilization and rupture of coronary atherosclerotic plaques and to acute myocardial infarction (AMI). Recently, it was found that immune activation is associated with the release of EVs by activated cells that mediate cell-cell communication and play an important role in immune activation. One of the manifestations of the activation of monocytes and platelets is the formation of monocyte-platelet complexes (MPC). We analyzed MPC in vivo and in vitro and investigated the abilities of different monocyte subclasses to form MPC, characteristics of the cells and EVs involved in MPC formation, and MPC changes in AMI. We identified MPC by co-staining for platelet antigen CD41a and monocyte antigens CD14 and CD16. Platelet activation was evaluated by expression of phosphatidylserine (PS). Monocytes of different classes disproportionately formed MPC: although classical monocytes (CD14++CD16-) constituted the majority, MPC were preferentially formed by intermediate monocytes (CD14++CD16+). CD41-positive events in MPC exposed more PS than the circulating ones. AMI was associated with a 50% increase in circulating monocytes and with a threefold increase in MPC, in particular in those formed by classical monocytes. In AMI patients, MPC formed by intermediate monocytes contained more CD41a-positive events than MPC with other monocyte subsets, whereas in controls, MPC formed by classical monocytes carried more platelets than other MPC. The sizes of some of the CD41a+ events in MPC were smaller than those of regular platelets and may represent platelet-derived EV. Some of the aggregates seem to consist of monocytes and platelet-derived EVs. Binding of EVs to monocytes was confirmed in in vitro experiments when monocytes were co-incubated with platelet-generated EVs. There was association between the increase in MPC and their composition, and in-patient complications of AMI. Aggregation of EVs and platelets with monocytes in AMI patients is another manifestation of immune activation associated with atherosclerosis that plays an important role in this pathology and can be used as an AMI correlate. However, it remains to be understood what causes this immune activation. Our results as well as other data may indicate the presence of a specific antigen(s) towards which these lymphocytes are reactive. Herpesviruses, in particular CMV, are one of the main candidates causing persistent immunoactivation within plaques due to their ubiquity and ability to cycle between dormancy and replication, as well as by their ability to cause endothelial dysfunction. In our previous work, we showed that productive cytomegalovirus infection is more common in patients with acute coronary syndrome than in patients with chronic coronary artery disease and healthy volunteers. Moreover, in that work we also found a positive correlation between the cytomegaloviral DNA load and T-lymphocyte differentiation within the atherosclerotic plaques of patients with cardiovascular diseases Now, we analyzed the presence of cytomegaloviral DNA in plasma and endothelial function in 33 patients with ST-elevation myocardial infarction (STEMI) and 33 volunteers without cardiovascular diseases, using real-time polymerase chain reaction and a noninvasive test of flow-mediated dilation. We found that the presence of cytomegaloviral DNA in plasma of STEMI patients was significantly higher than in volunteers without cardiovascular diseases. Also, we found a significant prevalence in the number of copies of cytomegaloviral DNA in older hypertensive patients, indicating that one of the possible mechanisms of the development of hypertension in patients with cytomegalovirus infection could be the development of endothelial dysfunction triggered by cytomegalovirus. In summary, we found that cytomegalovirus infection is strongly associated with endothelial dysfunction in STEMI patients. Therefore, it is conceivable that preventing the reactivation of cytomegalovirus may enhance endothelial functions
| Fitzgerald, Wendy; Gomez-Lopez, Nardhy; Erez, Offer et al. (2018) Extracellular vesicles generated by placental tissues ex vivo: A transport system for immune mediators and growth factors. Am J Reprod Immunol 80:e12860 |
| Introini, Andrea; Fitzgerald, Wendy; Vanpouille, Christophe et al. (2018) Histoculture and Infection with HIV of Functional Human Lymphoid Tissue on Gelfoam®. Methods Mol Biol 1760:187-197 |
| Kaur, Sukhbir; Elkahloun, Abdel G; Arakelyan, Anush et al. (2018) CD63, MHC class 1, and CD47 identify subsets of extracellular vesicles containing distinct populations of noncoding RNAs. Sci Rep 8:2577 |
| Fitzgerald, Wendy; Freeman, Michael L; Lederman, Michael M et al. (2018) A System of Cytokines Encapsulated in ExtraCellular Vesicles. Sci Rep 8:8973 |
| Zicari, Sonia; Arakelyan, Anush; Palomino, Rogers Alberto Ñahui et al. (2018) Human cytomegalovirus-infected cells release extracellular vesicles that carry viral surface proteins. Virology 524:97-105 |
| Vagida, Murad; Arakelyan, Anush; Lebedeva, Anna et al. (2017) Flow analysis of individual blood extracellular vesicles in acute coronary syndrome. Platelets 28:165-173 |
| Tarca, Adi L; Fitzgerald, Wendy; Chaemsaithong, Piya et al. (2017) The cytokine network in women with an asymptomatic short cervix and the risk of preterm delivery. Am J Reprod Immunol 78: |
| Vanpouille, Christophe; Bernatchez, Jean A; Lisco, Andrea et al. (2017) A common anti-cytomegalovirus drug, ganciclovir, inhibits HIV-1 replication in human tissues ex vivo. AIDS 31:1519-1528 |
| Lebedeva, Anna; Vorobyeva, Daria; Vagida, Murad et al. (2017) Ex vivo culture of human atherosclerotic plaques: A model to study immune cells in atherogenesis. Atherosclerosis 267:90-98 |
| Christensen-Quick, Aaron; Vanpouille, Christophe; Lisco, Andrea et al. (2017) Cytomegalovirus and HIV Persistence: Pouring Gas on the Fire. AIDS Res Hum Retroviruses 33:S23-S30 |
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