RESUMO
Antigen-specific multifunctional T cells that secrete interferon-γ, interleukin-2 and tumour necrosis factor-α simultaneously after activation are important for the control of many infections. It is unclear if these CD8(+) T cells are at an early or late stage of differentiation and whether telomere erosion restricts their replicative capacity. We developed a multi-parameter flow cytometric method for investigating the relationship between differentiation (CD45RA and CD27 surface phenotype), function (cytokine production) and replicative capacity (telomere length) in individual cytomegalovirus (CMV) antigen-specific CD8(+) T cells. This involves surface and intracellular cell staining coupled to fluorescence in situ hybridization to detect telomeres (flow-FISH). The end-stage/senescent CD8(+) CD45RA(+) CD27(-) T-cell subset increases significantly during ageing and this is exaggerated in CMV immune-responsive subjects. However, these end-stage cells do not have the shortest telomeres, implicating additional non-telomere-related mechanisms in inducing their senescence. The telomere lengths in total and CMV (NLV)-specific CD8(+) T cells in all four subsets defined by CD45RA and CD27 expression were significantly shorter in old compared with young individuals in both a Caucasian and an Asian cohort. Following stimulation by anti-CD3 or NLV peptide, similar proportions of triple-cytokine-producing cells are found in CD8(+) T cells at all stages of differentiation in both age groups. Furthermore, these multi-functional cells had intermediate telomere lengths compared with cells producing only one or two cytokines after activation. Therefore, global and CMV (NLV)-specific CD8(+) T cells that secrete interferon-γ, interleukin-2 and tumour necrosis factor-α are at an intermediate stage of differentiation and are not restricted by excessive telomere erosion.
Assuntos
Envelhecimento/imunologia , Linfócitos T CD8-Positivos/imunologia , Senescência Celular , Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Ativação Linfocitária , Encurtamento do Telômero , Telômero/imunologia , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Envelhecimento/etnologia , Envelhecimento/genética , Povo Asiático/genética , Biomarcadores/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/virologia , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Citocinas/imunologia , Citocinas/metabolismo , Citomegalovirus/patogenicidade , Infecções por Citomegalovirus/genética , Infecções por Citomegalovirus/metabolismo , Infecções por Citomegalovirus/virologia , Citometria de Fluxo , Humanos , Imunofenotipagem/métodos , Antígenos Comuns de Leucócito/imunologia , Antígenos Comuns de Leucócito/metabolismo , Londres , Fenótipo , Singapura , Telômero/genética , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/imunologia , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo , População Branca/genética , Adulto JovemRESUMO
T cell senescence is thought to contribute to immune function decline, but the pathways that mediate senescence in these cells are not clear. Here, we evaluated T cell populations from healthy volunteers and determined that human CD8+ effector memory T cells that reexpress the naive T cell marker CD45RA have many characteristics of cellular senescence, including decreased proliferation, defective mitochondrial function, and elevated levels of both ROS and p38 MAPK. Despite their apparent senescent state, we determined that these cells secreted high levels of both TNF-α and IFN-γ and showed potent cytotoxic activity. We found that the senescent CD45RA-expressing population engaged anaerobic glycolysis to generate energy for effector functions. Furthermore, inhibition of p38 MAPK signaling in senescent CD8+ T cells increased their proliferation, telomerase activity, mitochondrial biogenesis, and fitness; however, the extra energy required for these processes did not arise from increased glucose uptake or oxidative phosphorylation. Instead, p38 MAPK blockade in these senescent cells induced an increase in autophagy through enhanced interactions between p38 interacting protein (p38IP) and autophagy protein 9 (ATG9) in an mTOR-independent manner. Together, our findings describe fundamental metabolic requirements of senescent primary human CD8+ T cells and demonstrate that p38 MAPK blockade reverses senescence via an mTOR-independent pathway.
Assuntos
Autofagia/fisiologia , Linfócitos T CD8-Positivos/fisiologia , Sistema de Sinalização das MAP Quinases/fisiologia , Complexos Multiproteicos/fisiologia , Serina-Treonina Quinases TOR/fisiologia , Proteínas Quinases p38 Ativadas por Mitógeno/fisiologia , Adulto , Proteínas Relacionadas à Autofagia , Senescência Celular , Dano ao DNA , Glicólise , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina , Potencial da Membrana Mitocondrial , Proteínas de Membrana/fisiologia , Mitocôndrias/fisiologia , Fosforilação Oxidativa , Proteínas de Transporte Vesicular/fisiologiaRESUMO
The mechanisms regulating memory CD8(+) T cell function and homeostasis during aging are unclear. CD8(+) effector memory T cells that re-express CD45RA increase considerably in older humans and both aging and persistent CMV infection are independent factors in this process. We used MHC class I tetrameric complexes that were mutated in the CD8 binding domain to identify CMV-specific CD8(+) T cells with high Ag-binding avidity. In individuals who were HLA-A*0201, CD8(+) T cells that expressed CD45RA and were specific for the pp65 protein (NLVPMVATV epitope) had lower avidity than those that expressed CD45RO and demonstrated decreased cytokine secretion and cytolytic potential after specific activation. Furthermore, low avidity NLVPMVATV-specific CD8(+) T cells were significantly increased in older individuals. The stimulation of blood leukocytes with CMV lysate induced high levels of IFN-α that in turn induced IL-15 production. Moreover, the addition of IL-15 to CD45RA(-)CD45RO(+) CMV-specific CD8(+) T cells induced CD45RA expression while Ag activated cells remained CD45RO(+). This raises the possibility that non-specific cytokine-driven accumulation of CMV-specific CD8(+)CD45RA(+) T cells with lower Ag-binding avidity may exacerbate the effects of viral reactivation on skewing the T cell repertoire in CMV-infected individuals during aging.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Antígenos Comuns de Leucócito/metabolismo , Fatores Etários , Afinidade de Anticorpos/imunologia , Antígenos/imunologia , Antígenos/metabolismo , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Antígeno HLA-A2/imunologia , Antígeno HLA-A2/metabolismo , Humanos , Imunofenotipagem , Interferon-alfa/biossíntese , Interleucina-15/imunologia , Interleucina-15/metabolismo , Ativação Linfocitária/imunologia , Monócitos/imunologia , Monócitos/metabolismo , Fosfoproteínas/imunologia , Proteínas da Matriz Viral/imunologiaRESUMO
The dysregulated immune response to CMV constitutes a major force driving T cell immunosenescence and growing evidence suggests that it is not a benign virus in old age. We show here that the PD-1/L pathway defines a reversible defect in CMV specific CD8(+) T cell proliferative responses in both young and old individuals. More specifically, highly differentiated CD45RA(+)CD27(-) CMV-specific CD8(+) T cells exhibit a proliferative deficit compared their central and effector memory counterparts, which is reversed following PD-L blockade. However, we also report that HLA-B(∗)07/TPR specific CD8(+) T cells express higher levels of PD-1 than HLA-A(∗)02/NLV specific cells and HLA-A(∗)02 individuals show a higher proliferative response to PD-L blockade, than HLA-B(∗)07 individuals, which we postulate may be due to the differing functional avidities for these two CMV-specific CD8(+) T cells populations. Nevertheless data presented here demonstrate that CMV-specific CD8(+) T cells can be functionally enhanced by perturbation of the PD-1/L signalling pathway, whose manipulation may provide a therapeutic modality to combat age-associated immune decline.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Citomegalovirus/imunologia , Antígenos HLA/imunologia , Receptor de Morte Celular Programada 1/imunologia , Transdução de Sinais/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígeno B7-H1/imunologia , Antígeno B7-H1/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/virologia , Diferenciação Celular/imunologia , Proliferação de Células , Células Cultivadas , Citomegalovirus/fisiologia , Citometria de Fluxo , Antígenos HLA/metabolismo , Antígenos HLA-A/imunologia , Antígenos HLA-A/metabolismo , Antígenos HLA-B/imunologia , Antígenos HLA-B/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Humanos , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/virologia , Pessoa de Meia-Idade , Receptor de Morte Celular Programada 1/metabolismo , Adulto JovemRESUMO
Contributions of the emissions from a U.K. regulated fossil-fuel power station to regional air pollution and deposition are estimated using four air quality modeling systems for the year 2003. The modeling systems vary in complexity and emphasis in the way they treat atmospheric and chemical processes, and include the Community Multiscale Air Quality (CMAQ) modeling system in its versions 4.6 and 4.7, a nested modeling system that combines long- and short-range impacts (referred to as TRACK-ADMS [Trajectory Model with Atmospheric Chemical Kinetics-Atmospheric Dispersion Modelling System]), and the Fine Resolution Atmospheric Multi-pollutant Exchange (FRAME) model. An evaluation of the baseline calculations against U.K. monitoring network data is performed. The CMAQ modeling system version 4.6 data set is selected as the reference data set for the model footprint comparison. The annual mean air concentration and total deposition footprints are summarized for each modeling system. The footprints of the power station emissions can account for a significant fraction of the local impacts for some species (e.g., more than 50% for SO2 air concentration and non-sea-salt sulfur deposition close to the source) for 2003. The spatial correlation and the coefficient of variation of the root mean square error (CVRMSE) are calculated between each model footprint and that calculated by the CMAQ modeling system version 4.6. The correlation coefficient quantifies model agreement in terms of spatial patterns, and the CVRMSE measures the magnitude of the difference between model footprints. Possible reasons for the differences between model results are discussed. Finally, implications and recommendations for the regulatory assessment of the impact of major industrial sources using regional air quality modeling systems are discussed in the light of results from this case study.
Assuntos
Poluição do Ar/análise , Monitoramento Ambiental/métodos , Modelos Teóricos , Centrais Elétricas , Reino UnidoRESUMO
The Second International Workshop on CMV & Immunosenescence was held in Cambridge, UK, 2-4th December, 2010. The presentations covered four separate sessions: cytomegalovirus and T cell phenotypes; T cell memory frequency, inflation and immunosenescence; cytomegalovirus in aging, mortality and disease states; and the immunobiology of cytomegalovirus-specific T cells and effects of the virus on vaccination. This commentary summarizes the major findings of these presentations and references subsequently published work from the presenter laboratory where appropriate and draws together major themes that were subsequently discussed along with new areas of interest that were highlighted by this discussion.
RESUMO
Short telomeres of circulating leukocytes are a risk factor for age-related diseases, such as atherosclerosis, but the exact mechanisms generating variations in telomere length are unknown. We hypothesized that induction of differentiated T cells during chronic CMV infection would affect T cell telomere length. To test this, we measured the amount of differentiated T cells and telomere length of lymphocytes during primary CMV infection as well as CMV-seropositive and -seronegative healthy individuals. After primary CMV infection, we observed an increase in highly differentiated cells that coincided with a steep drop in telomere length. Moreover, we found in a cohort of 159 healthy individuals that telomere shortening was more rapid in CMV-seropositive individuals and correlated with the amount of differentiated T cells in both CD4(+) T cells and CD8(+) T cells. Finally, we found that telomere length measured in blood leukocytes is correlated with lymphocyte telomere length. Thus, CMV infection induces a strong decrease in T cell telomere length, which can be explained by changes in the composition of the circulating lymphocyte pool.
Assuntos
Infecções por Citomegalovirus/imunologia , Subpopulações de Linfócitos T/patologia , Linfócitos T/patologia , Telômero/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antivirais/uso terapêutico , Diferenciação Celular/imunologia , Separação Celular , Infecções por Citomegalovirus/genética , Infecções por Citomegalovirus/patologia , DNA Viral/sangue , Citometria de Fluxo , Imunofluorescência , Ganciclovir/uso terapêutico , Humanos , Hibridização in Situ Fluorescente , Transplante de Rim/efeitos adversos , Transplante de Rim/imunologia , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Subpopulações de Linfócitos T/imunologia , Linfócitos T/imunologia , Telômero/virologia , Viremia/tratamento farmacológico , Adulto JovemRESUMO
Highly differentiated CD8+CD28-CD27- T cells have short telomeres, defective telomerase activity, and reduced capacity for proliferation, indicating that they are close to replicative senescence. In addition, these cells express increased levels of the senescence-associated inhibitory receptor KLRG1 and have poor capacity for IL-2 synthesis and defective Akt (ser(473)) phosphorylation after activation. It is not known whether signaling via KLRG1 contributes to any of the attenuated differentiation-related functional changes in CD8+ T cells. To address this, we blocked KLRG1 signaling during T-cell receptor activation using antibodies against its major ligand, E-cadherin. This resulted in a significant enhancement of Akt (ser(473)) phosphorylation and T-cell receptor-induced proliferative activity of CD8+CD28-CD27- T cells. Furthermore, the increase of proliferation was directly linked to the Akt-mediated induction of cyclin D and E and reduction in the cyclin inhibitor p27 expression. In contrast, the reduced telomerase activity in highly differentiated CD8+CD28(-)CD27- T cells was not altered by KLRG1 blockade, indicating the involvement of other mechanisms. This is the first demonstration of a functional role for KLRG1 in primary human CD8+ T cells and highlights that certain functional defects that arise during progressive T-cell differentiation toward replicative senescence are maintained actively by inhibitory receptor signaling.