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[This corrects the article DOI: 10.3389/fimmu.2020.626431.].
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BACKGROUND AND AIMS: Hepatitis C virus (HCV) leads to a high rate of chronic infection and T cell dysfunction. Although it is well known that chronic antigenic stimulation is a driving force for impaired T cell functions, the precise mechanisms underlying immune activation-induced T cell dysfunctions during HCV infection remain elusive. APPROACH AND RESULTS: Here, we demonstrated that circulating CD4+ T cells from patients who are chronically HCV-infected exhibit an immune activation status, as evidenced by the overexpression of cell activation markers human leukocyte antigen-antigen D-related, glucose transporter 1, granzyme B, and the short-lived effector marker CD127- killer cell lectin-like receptor G1+ . In contrast, the expression of stem cell-like transcription factor T cell factor 1 and telomeric repeat-binding factor 2 (TRF2) are significantly reduced in CD4+ T cells from patients who are chronically HCV-infected compared with healthy participants (HP). Mechanistic studies revealed that CD4+ T cells from participants with HCV exhibit phosphoinositide 3-kinase/Akt/mammalian target of rapamycin signaling hyperactivation on T cell receptor stimulation, promoting proinflammatory effector cell differentiation, telomeric DNA damage, and cellular apoptosis. Inhibition of Akt signaling during T cell activation preserved the precursor memory cell population and prevented inflammatory effector cell expansion, DNA damage, and apoptotic death. Moreover, knockdown of TRF2 reduced HP T cell stemness and triggered telomeric DNA damage and cellular apoptosis, whereas overexpression of TRF2 in CD4 T cells prevented telomeric DNA damage. CONCLUSIONS: These results suggest that modulation of immune activation through inhibiting Akt signaling and protecting telomeres through enhancing TRF2 expression may open therapeutic strategies to fine tune the adaptive immune responses in the setting of persistent immune activation and inflammation during chronic HCV infection.
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Linfócitos T CD4-Positivos/imunologia , Diferenciação Celular/imunologia , Dano ao DNA/imunologia , Hepacivirus/genética , Hepatite C Crônica/genética , Hepatite C Crônica/imunologia , Telômero/genética , Adulto , Idoso , Apoptose/genética , Apoptose/imunologia , Células Cultivadas , Dano ao DNA/genética , Feminino , Técnicas de Silenciamento de Genes/métodos , Hepatite C Crônica/virologia , Humanos , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Infecção Persistente/genética , Infecção Persistente/imunologia , Infecção Persistente/virologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Viral/genética , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Serina-Treonina Quinases TOR/metabolismo , Proteína 2 de Ligação a Repetições Teloméricas/genética , Proteína 2 de Ligação a Repetições Teloméricas/metabolismo , Transdução Genética/métodos , Adulto JovemRESUMO
The hallmark of HIV/AIDS is a gradual depletion of CD4 T cells. Despite effective control by antiretroviral therapy (ART), a significant subgroup of people living with HIV (PLHIV) fails to achieve complete immune reconstitution, deemed as immune non-responders (INRs). The mechanisms underlying incomplete CD4 T cell recovery in PLHIV remain unclear. In this study, CD4 T cells from PLHIV were phenotyped and functionally characterized, focusing on their mitochondrial functions. The results show that while total CD4 T cells are diminished, cycling cells are expanded in PLHIV, especially in INRs. HIV-INR CD4 T cells are more activated, displaying exhausted and senescent phenotypes with compromised mitochondrial functions. Transcriptional profiling and flow cytometry analysis showed remarkable repression of mitochondrial transcription factor A (mtTFA) in CD4 T cells from PLHIV, leading to abnormal mitochondrial and T cell homeostasis. These results demonstrate a sequential cellular paradigm of T cell over-activation, proliferation, exhaustion, senescence, apoptosis, and depletion, which correlates with compromised mitochondrial functions. Therefore, reconstituting the mtTFA pathway may provide an adjunctive immunological approach to revitalizing CD4 T cells in ART-treated PLHIV, especially in INRs.
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Linfócitos T CD4-Positivos/metabolismo , Infecções por HIV/metabolismo , Infecções por HIV/virologia , HIV-1 , Mitocôndrias/metabolismo , Adulto , Idoso , Terapia Antirretroviral de Alta Atividade , Apoptose , Biomarcadores , Contagem de Linfócito CD4 , Linfócitos T CD4-Positivos/imunologia , Células Cultivadas , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Expressão Gênica , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , HIV-1/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Mitocôndrias/genética , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Estresse Fisiológico , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Carga Viral , Adulto JovemRESUMO
RUNX1 overlapping RNA (RUNXOR) is a long noncoding RNA and a key regulator of myeloid-derived suppressor cells (MDSCs) via targeting runt-related transcription factor 1 (RUNX1). We and others have previously reported MDSC expansion and inhibition of host immune responses during viral infections; however, the mechanisms regulating MDSC differentiation and suppressive functions, especially the role of RUNXOR-RUNX1 in the regulation of MDSCs in people living with HIV (PLHIV), remain unknown. In this study, we demonstrate that RUNXOR and RUNX1 expressions are upregulated in MDSCs that expand and accumulate in human PBMCs derived from PLHIV. We found that the upregulation of RUNXOR and RUNX1 is associated with the expressions of several key immunosuppressive molecules, including arginase 1, inducible NO synthase, STAT3, IL-6, and reactive oxygen species. RUNXOR and RUNX1 could positively regulate each other's expression and control the expressions of these suppressive mediators. Specifically, silencing RUNXOR or RUNX1 expression in MDSCs from PLHIV attenuated MDSC expansion and immunosuppressive mediator expressions, whereas overexpressing RUNXOR in CD33+ myeloid precursors from healthy subjects promoted their differentiation into MDSCs and enhanced the expression of these mediators. Moreover, loss of RUNXOR-RUNX1 function in MDSCs improved IFN-γ production from cocultured autologous CD4 T cells derived from PLHIV. These results suggest that the RUNXOR-RUNX1 axis promotes the differentiation and suppressive functions of MDSCs via regulating multiple immunosuppressive signaling molecules and may represent a potential target for immunotherapy in conjunction with antiviral therapy in PLHIV.
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Subunidade alfa 2 de Fator de Ligação ao Core/genética , Regulação da Expressão Gênica , Infecções por HIV/genética , Células Supressoras Mieloides/metabolismo , RNA Longo não Codificante/genética , Arginase/genética , Arginase/metabolismo , Diferenciação Celular/genética , Proliferação de Células/genética , Células Cultivadas , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Infecções por HIV/tratamento farmacológico , Infecções por HIV/metabolismo , Humanos , Células Supressoras Mieloides/citologia , Interferência de RNA , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/genética , Regulação para CimaRESUMO
The recent COVID-19 pandemic poses a serious threat to global public health, thus there is an urgent need to define the molecular mechanisms involved in SARS-CoV-2 spike (S) protein-mediated virus entry that is essential for preventing and/or treating this emerging infectious disease. In this study, we examined the blocking activity of human COVID-19 convalescent plasma by cell-cell fusion assays using SARS-CoV-2-S-transfected 293 T as effector cells and ACE2-expressing 293 T as target cells. We demonstrate that the SARS-CoV-2 S protein exhibits a very high capacity for membrane fusion and is efficient in mediating virus fusion and entry into target cells. Importantly, we find that COVID-19 convalescent plasma with high titers of IgG neutralizing antibodies can block cell-cell fusion and virus entry by interfering with the SARS-CoV-2-S/ACE2 or SARS-CoV-S/ACE2 interactions. These findings suggest that COVID-19 convalescent plasma may not only inhibit SARS-CoV-2-S but also cross-neutralize SARS-CoV-S-mediated membrane fusion and virus entry, supporting its potential as a preventive and/or therapeutic agent against SARS-CoV-2 as well as other SARS-CoV infections.
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COVID-19/imunologia , COVID-19/terapia , Glicoproteína da Espícula de Coronavírus/imunologia , Adulto , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , COVID-19/prevenção & controle , Fusão Celular/métodos , Feminino , Humanos , Imunização Passiva/métodos , Masculino , Fusão de Membrana/efeitos dos fármacos , Pessoa de Meia-Idade , Pandemias/prevenção & controle , Plasma/química , Receptores Virais/metabolismo , SARS-CoV-2/imunologia , SARS-CoV-2/patogenicidade , Glicoproteína da Espícula de Coronavírus/metabolismo , Internalização do Vírus/efeitos dos fármacos , Soroterapia para COVID-19RESUMO
HOXA transcript antisense RNA myeloid-specific 1 (HOTAIRM1) is a long non-coding RNA (lncRNA) that plays a pivotal role in regulating myeloid cell development via targeting HOXA1 gene expression. We and others have previously shown that myeloid-derived suppressor cells (MDSCs), a heterogeneous population of immature myeloid cells, expand during chronic viral (HCV, HIV) infections. However, the role of HOTAIRM1 in the development and suppression of MDSCs during viral infection remains unknown. In this study, we demonstrate that the expressions of HOTAIRM1 and its target HOXA1 are substantially upregulated to promote the expressions of immunosuppressive molecules, including arginase 1, inducible nitric oxide synthase, signal transducer and activator of transcription 3, and reactive oxygen species, in CD33+ myeloid cells derived from hepatitis C virus (HCV)-infected patients. We show that HCV-associated exosomes (HCV-Exo) can modulate HOTAIRM1, HOXA1, and miR124 expressions to regulate MDSC development. Importantly, overexpression of HOTAIRM1 or HOXA1 in healthy CD33+ myeloid cells promoted the MDSC differentiation and suppressive functions; conversely, silencing of HOTAIRM1 or HOXA1 expression in MDSCs from HCV patients significantly reduced the MDSC frequency and their suppressive functions. In essence, these results indicate that the HOTAIRM1-HOXA1-miR124 axis enhances the differentiation and suppressive functions of MDSCs and may be a potential target for immunomodulation in conjunction with antiviral therapy during chronic viral infection.
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Hepacivirus/fisiologia , Hepatite C Crônica/genética , Hepatite C Crônica/patologia , Proteínas de Homeodomínio/metabolismo , MicroRNAs/metabolismo , Células Supressoras Mieloides/metabolismo , Fatores de Transcrição/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Diferenciação Celular , Proliferação de Células , Inativação Gênica , Proteínas de Homeodomínio/genética , Humanos , Terapia de Imunossupressão , MicroRNAs/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores de Transcrição/genética , Regulação para Cima/genéticaRESUMO
Telomere erosion and mitochondrial dysfunction are prominent features of aging cells with progressive declines of cellular functions. Whether telomere injury induces mitochondrial dysfunction in human T lymphocytes, the major component of adaptive host immunity against infection and malignancy, remains unclear. We have recently shown that disruption of telomere integrity by KML001, a telomere-targeting drug, induces T cell senescence and apoptosis via the telomeric DNA damage response (DDR). In this study, we used KML001 to further investigate the role and mechanism of telomere injury in mitochondrial dysregulation in aging T cells. We demonstrate that targeting telomeres by KML001 induces mitochondrial dysfunction, as evidenced by increased mitochondrial swelling and decreased mitochondrial membrane potential, oxidative phosphorylation, mitochondrial DNA content, mitochondrial respiration, oxygen consumption, glycolysis, and ATP energy production. Mechanistically, we found that the KML001-induced telomeric DDR activated p53 signaling, which in turn repressed the expression of peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PGC-1α) and nuclear respiratory factor 1 (NRF-1), leading to T cell mitochondrial dysfunction. These results, forging a direct link between telomeric and mitochondrial biology, shed new light on the human T cell aging network, and demonstrate that the p53-PGC-1α-NRF-1 axis contributes to mitochondrial dysfunction in the setting of telomeric DDR. This study suggests that targeting this axis may offer an alternative, novel approach to prevent telomere damage-mediated mitochondrial and T cell dysfunctions to combat a wide range of immune aging-associated human diseases.
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Arsenitos/toxicidade , Linfócitos T CD4-Positivos/patologia , Mitocôndrias/patologia , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Transdução de Sinais , Compostos de Sódio/toxicidade , Telômero/patologia , Proteína Supressora de Tumor p53/metabolismo , Linfócitos T CD4-Positivos/efeitos dos fármacos , Dano ao DNA , Reparo do DNA/efeitos dos fármacos , Reparo do DNA/genética , Regulação para Baixo/genética , Redes Reguladoras de Genes/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Humanos , Mitocôndrias/efeitos dos fármacos , Biogênese de Organelas , Transdução de Sinais/efeitos dos fármacos , Telômero/efeitos dos fármacos , Regulação para Cima/genéticaRESUMO
RUNX1 overlapping RNA (RUNXOR) is a long non-coding RNA and plays a pivotal role in the differentiation of myeloid cells via targeting runt-related transcription factor 1 (RUNX1). We and others have previously reported that myeloid-derived suppressor cells (MDSCs) expand and inhibit host immune responses during chronic viral infections; however, the mechanisms responsible for MDSC differentiation and suppressive functions, in particular the role of RUNXOR-RUNX1, remain unclear. Here, we demonstrated that RUNXOR and RUNX1 expressions are significantly upregulated and associated with elevated levels of immunosuppressive molecules, such as arginase 1 (Arg1), inducible nitric oxide synthase (iNOS), signal transducer and activator of transcription 3 (STAT3), and reactive oxygen species (ROS) in MDSCs during chronic hepatitis C virus (HCV) infection. Mechanistically, we discovered that HCV-associated exosomes (HCV-Exo) can induce the expressions of RUNXOR and RUNX1, which in turn regulates miR-124 expression via STAT3 signaling, thereby promoting MDSC differentiation and suppressive functions. Importantly, overexpression of RUNXOR in healthy CD33+ myeloid cells promoted differentiation and suppressive functions of MDSCs. Conversely, silencing RUNXOR or RUNX1 expression in HCV-derived CD33+ myeloid cells significantly inhibited their differentiation and expressions of suppressive molecules and improved the function of co-cultured autologous CD4 T cells. Taken together, these results indicate that the RUNXOR-RUNX1-STAT3-miR124 axis enhances the differentiation and suppressive functions of MDSCs and could be a potential target for immunomodulation in conjunction with antiviral therapy during chronic HCV infection.
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Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Exossomos/virologia , Hepacivirus , MicroRNAs/metabolismo , Células Supressoras Mieloides/citologia , RNA Longo não Codificante , Fator de Transcrição STAT3/metabolismo , Adulto , Idoso , Arginase/metabolismo , Linfócitos T CD4-Positivos/citologia , Diferenciação Celular , Proliferação de Células , Feminino , Regulação Viral da Expressão Gênica , Genótipo , Hepatite C Crônica/metabolismo , Hepatite C Crônica/virologia , Humanos , Imunossupressores , Interferon gama/metabolismo , Masculino , Pessoa de Meia-Idade , Células Supressoras Mieloides/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico/metabolismo , Regulação para Cima , Carga ViralRESUMO
OBJECTIVE: Myeloid-derived suppressor cells (MDSCs) contribute to HIV progression by impairing antiviral immunity; however, the mechanisms responsible for MDSC development during HIV infection are incompletely understood. HOX antisense intergenic RNA myeloid 1 (HOTAIRM1) is a long noncoding RNA (lncRNA) that plays a pivotal role in regulating myeloid cell development via targeting HOXA1. The role of HOTAIRM1--HOXA1 in the differentiation and functions of MDSCs during HIV infection remains unclear. METHODS: In this study, we measured MDSC induction and suppressive functions by flow cytometry, RT-PCR, and co-culture experiments using CD33 myeloid cells derived from people living with HIV (PLHIV) on antiretroviral therapy (ART). We also manipulated the HOTAIRM1--HOXA1 axis in myeloid cells using knockdown and overexpression approaches. RESULTS: We demonstrate that HOTAIRM1 and HOXA1 expressions are reciprocally upregulated and are responsible for increased levels of immunosuppressive molecules, such as arginase 1 (Arg1), inducible nitric oxide synthase (iNOS), signal transducer and activator of transcription 3 (STAT3), and reactive oxygen species (ROS), in CD33 myeloid cells derived from PLHIV on ART. We found that overexpression of HOTAIRM1 or HOXA1 in CD33 cells isolated from healthy individuals promoted the differentiation and suppressive functions of MDSCs, whereas silencing of HOTAIRM1 or HOXA1 expression in MDSCs derived from PLHIV significantly inhibited the frequency of MDSCs and expressions of the immunosuppressive molecules and reduced their immunosuppressive effects on T cells. CONCLUSION: These results indicate that the HOTAIRM1--HOXA1 axis enhances differentiation and suppressive functions of MDSCs and could be a potential therapeutic target for immunomodulation during latent HIV infection.
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Infecções por HIV , Proteínas de Homeodomínio , MicroRNAs , Células Supressoras Mieloides , Fatores de Transcrição , Fármacos Anti-HIV/uso terapêutico , Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Infecções por HIV/tratamento farmacológico , Infecções por HIV/genética , Infecções por HIV/imunologia , Infecções por HIV/patologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Células Supressoras Mieloides/imunologia , Células Supressoras Mieloides/patologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Regulação para CimaRESUMO
CD4 T-cell depletion is a hallmark of HIV/AIDS, but the underlying mechanism is still unclear. We have recently shown that ataxia-telangiectasia-mutated (ATM) deficiency in CD4 T cells accelerates DNA damage, telomere erosion, and cell apoptosis in HIV-infected individuals on antiretroviral therapy (ART). Whether these alterations in ART-treated HIV subjects occur in vitro in HIV-infected CD4 T cells remains unknown. In this study, we employed a cellular model of HIV infection to characterize the mechanisms underlying CD4 T-cell destruction by analyzing the telomeric DNA damage response (DDR) and cellular apoptosis in highly permissive SupT1 cells, followed by the validation of our observations in primary CD4 T cells with active or drug-suppressed HIV infection. Specifically, we established an in vitro HIV T-cell culture system with viral replication and raltegravir (RAL; an integrase inhibitor) suppression, mimicking active and ART-controlled HIV infection in vivo We demonstrated that HIV-induced, telomeric DDR plays a pivotal role in triggering telomere erosion, premature T-cell aging, and CD4 T-cell apoptosis or depletion via dysregulation of the PI3K/ATM pathways. This in vitro model provides a new tool to investigate HIV pathogenesis, and our results shed new light on the molecular mechanisms of telomeric DDR and CD4 T-cell homeostasis during HIV infection.IMPORTANCE The hallmark of HIV infection is a gradual depletion of CD4 T cells, with a progressive decline of host immunity. How CD4 T cells are depleted in individuals with active and virus-suppressed HIV infection remains unclear. In this study, we employed a cellular model of HIV infection to characterize the mechanisms underlying CD4 T-cell destruction by analyzing the chromosome end (telomere) DNA damage response (DDR) and cellular apoptosis in a T-cell line (highly permissive SupT1 cells), as well as in primary CD4 T cells with active or drug-suppressed HIV infection. We demonstrated that HIV-induced telomeric DDR plays a critical role in inducing telomere loss, premature cell aging, and CD4 T-cell apoptosis or depletion via dysregulation of the PI3K/ATM pathways. This study sheds new light on the molecular mechanisms of telomeric DDR and its role in CD4 T-cell homeostasis during HIV infection.
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Ataxia Telangiectasia/genética , Linfócitos T CD4-Positivos/metabolismo , Infecções por HIV/imunologia , Telômero/metabolismo , Apoptose , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Linhagem Celular , Senescência Celular , Dano ao DNA , Células HEK293 , HIV-1/genética , Humanos , Replicação ViralRESUMO
Salicylic acid is widely studied for its role in biotic stress signaling in plants. Several SA-binding proteins, including SABP2 (salicylic acid-binding protein 2) has been identified and characterized for their role in plant disease resistance. SABP2 is a 29 kDA tobacco protein that binds to salicylic acid with high affinity. It is a methylesterase enzyme that catalyzes the conversion of methyl salicylate into salicylic acid required for inducing a robust systemic acquired resistance (SAR) in plants. Methyl salicylic acid is one of the several mobile SAR signals identified in plants. SABP2-interacting protein 428 (SIP428) was identified in a yeast two-hybrid screen using tobacco SABP2 as a bait. In silico analysis shows that SIP428 possesses the SIR2 (silent information regulatory 2)-like conserved motifs. SIR2 enzymes are orthologs of sirtuin proteins that catalyze the NAD+-dependent deacetylation of Nε lysine-acetylated proteins. The recombinant SIP428 expressed in E. coli exhibits SIR2-like deacetylase activity. SIP428 shows homology to Arabidopsis AtSRT2 (67% identity), which is implicated in SA-mediated basal defenses. Immunoblot analysis using anti-acetylated lysine antibodies showed that the recombinant SIP428 is lysine acetylated. The expression of SIP428 transcripts was moderately downregulated upon infection by TMV. In the presence of SIP428, the esterase activity of SABP2 increased modestly. The interaction of SIP428 with SABP2, it's regulation upon pathogen infection, and similarity with AtSRT2 suggests that SIP428 is likely to play a role in stress signaling in plants.
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T cells play a critical role in controlling viral infection; however, the mechanisms regulating their responses remain incompletely understood. Here, we investigated the role of topoisomerase IIA (Top2α, an enzyme that is essential in resolving entangled DNA strands during replication) in telomeric DNA damage and T cell dysfunction during viral infection. We demonstrated that T cells derived from patients with chronic viral (HBV, HCV, and HIV) infection had lower Top2α protein levels and enzymatic activity, along with an accumulation of the Top2α cleavage complex (Top2cc) in genomic DNA. In addition, T cells from virally infected subjects with lower Top2α levels were vulnerable to Top2α inhibitor-induced cell apoptosis, indicating an important role for Top2α in preventing DNA topological disruption and cell death. Using Top2α inhibitor (ICRF193 or Etoposide)-treated primary T cells as a model, we demonstrated that disrupting the DNA topology promoted DNA damage and T cell apoptosis via Top2cc accumulation that is associated with protein-DNA breaks (PDB) at genomic DNA. Disruption of the DNA topology was likely due to diminished expression of tyrosyl-DNA phosphodiesterase 2 (TDP2), which was inhibited in T cells in vitro by Top2α inhibitor and in vivo by chronic viral infection. These results suggest that immune-evasive viruses (HBV, HCV, and HIV) can disrupt T cell DNA topology as a mechanism of dysregulating host immunity and establishing chronic infection. Thus, restoring the DNA topologic machinery may serve as a novel strategy to protect T cells from unwanted DNA damage and to maintain immune competence.
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Linfócitos T CD4-Positivos/imunologia , Dano ao DNA , DNA Topoisomerases Tipo II/metabolismo , Viroses/genética , Viroses/imunologia , Adulto , Idoso , Apoptose/efeitos dos fármacos , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/enzimologia , Doença Crônica , Reparo do DNA , DNA Topoisomerases Tipo II/biossíntese , Proteínas de Ligação a DNA/antagonistas & inibidores , Proteínas de Ligação a DNA/metabolismo , Dicetopiperazinas , Etoposídeo/farmacologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Diester Fosfórico Hidrolases/metabolismo , Piperazinas/farmacologia , Poli(ADP-Ribose) Polimerase-1/antagonistas & inibidores , Poli(ADP-Ribose) Polimerase-1/metabolismo , Telomerase/antagonistas & inibidores , Telomerase/metabolismo , Telômero/efeitos dos fármacos , Telômero/enzimologia , Telômero/genética , Proteína 2 de Ligação a Repetições Teloméricas/metabolismo , Inibidores da Topoisomerase II/farmacologia , Viroses/enzimologia , Adulto JovemRESUMO
CD4 T cell death or survival following initial HIV infection is crucial for the development of viral reservoirs and latent infection, making its evaluation critical in devising strategies for HIV cure. Here we infected primary CD4 T cells with a wild-type HIV-1 and investigated the death and survival mechanisms in productively infected and bystander cells during early HIV infection. We found that HIV-infected cells exhibited increased programmed cell death, such as apoptosis, pyroptosis, and ferroptosis, than uninfected cells. However, productively infected (p24+) cells and bystander (p24-) cells displayed different patterns of cell death due to differential expression of pro-/anti-apoptotic proteins and signaling molecules. Cell death was triggered by an aberrant DNA damage response (DDR), as evidenced by increases in γH2AX levels, which inversely correlated with telomere length and telomerase levels during HIV infection. Mechanistically, HIV-infected cells exhibited a gradual shortening of telomeres following infection. Notably, p24+ cells had longer telomeres compared to p24- cells, and telomere length positively correlated with the telomerase, pAKT, and pATM expressions in HIV-infected CD4 T cells. Importantly, blockade of viral entry attenuated the HIV-induced inhibition of telomerase, pAKT, and pATM as well as the associated telomere erosion and cell death. Moreover, ATM inhibition promoted survival of HIV-infected CD4 T cells, especially p24+ cells, and rescued telomerase and AKT activities by inhibiting cell activation, HIV infection, and DDR. These results indicate that productively infected and bystander CD4 T cells employ different mechanisms for their survival and death, suggesting a possible pro-survival, pro-reservoir mechanism during early HIV infection.
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Efeito Espectador/imunologia , Linfócitos T CD4-Positivos/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Morte Celular Regulada/imunologia , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD4-Positivos/virologia , Sobrevivência Celular/imunologia , Dano ao DNA/imunologia , Feminino , Células HEK293 , Proteína do Núcleo p24 do HIV/imunologia , Infecções por HIV/patologia , Histonas/imunologia , Humanos , Masculino , Proteínas Proto-Oncogênicas c-akt/imunologia , Telômero/imunologiaRESUMO
HIV infection leads to a phenomenon of inflammaging, in which chronic inflammation induces an immune aged phenotype, even in individuals on combined antiretroviral therapy (cART) with undetectable viremia. In this study, we investigated T cell homeostasis and telomeric DNA damage and repair machineries in cART-controlled HIV patients at risk for inflammaging. We found a significant depletion of CD4 T cells, which was inversely correlated with the cell apoptosis in virus-suppressed HIV subjects compared to age-matched healthy subjects (HS). In addition, HIV CD4 T cells were prone to DNA damage that extended to chromosome ends-telomeres, leading to accelerated telomere erosion-a hallmark of cell senescence. Mechanistically, the DNA double-strand break (DSB) sensors MRE11, RAD50, and NBS1 (MRN complex) remained intact, but both expression and activity of the DNA damage checkpoint kinase ataxia-telangiectasia mutated (ATM) and its downstream checkpoint kinase 2 (CHK2) were significantly suppressed in HIV CD4 T cells. Consistently, ATM/CHK2 activation, DNA repair, and cellular functions were also impaired in healthy CD4 T cells following ATM knockdown or exposure to the ATM inhibitor KU60019 in vitro, recapitulating the biological effects observed in HIV-derived CD4 T cells in vivo. Importantly, ectopic expression of ATM was essential and sufficient to reduce the DNA damage, apoptosis, and cellular dysfunction in HIV-derived CD4 T cells. These results demonstrate that failure of DSB repair due to ATM deficiency leads to increased DNA damage and renders CD4 T cells prone to senescence and apoptotic death, contributing to CD4 T cell depletion or dysfunction in cART-controlled, latent HIV infection.