RESUMEN
There has been a consistent upward trend in ground-level ozone (O3) concentration in China. People living with HIV (PLWH) may be more vulnerable to the health impacts of O3 exposure due to their immunosuppressed state. This study aims to investigate the association between ambient O3 exposure and mortality among PLWH, as well as the potential exacerbating effects of a decreased CD4+ T cell level. Daily maximum 8-hour O3 concentrations were assigned to 7270 PLWH at a county level in Guangxi, China. Every 10-unit increase in ambient O3 concentration was associated with a significant rise in all-cause mortality ranging from 7.3 % to 28.7 % and a significant rise in AIDS-related mortality ranging from 8.4 % to 14.5 %. When PLWH had a higher CD4+ count (≥350 cells/µL), elevated O3 concentration was associated with increased blood CD4+ count at lag0 [percent change with 95 % confidence interval, 0.20(0.00, 0.40)], lag1 [0.26(0.06, 0.47)], and lag2 [0.23(0.03, 0.44)]; however, an opposite association was observed when CD4+ count was <350 cells/µL for half-year average [-2.45(-4.71, -0.14)] and yearly average [-3.42(-5.51, -1.29)] of O3 exposure. The association of O3 exposure with all-cause and AIDS-related mortality was more prominent among those with higher CD4+ count. Exploratory analysis revealed possible associations between O3 exposure and respiratory infections and clinical symptoms. These findings suggest potential synergistic effects between a compromised immune status and elevated O3 exposure levels on mortality risk among PLWH. Ambient O3 exposure should be considered as an emerging mortality risk factor for PLWH in the era of antiretroviral therapy, requiring further attention from researchers and healthcare professionals.
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Síndrome de Inmunodeficiencia Adquirida , Contaminantes Atmosféricos , Contaminación del Aire , Ozono , Humanos , Contaminación del Aire/análisis , Contaminantes Atmosféricos/análisis , Estudios Longitudinales , Linfocitos T , China/epidemiología , Ozono/efectos adversos , Ozono/análisis , Linfocitos T CD4-Positivos/química , Exposición a Riesgos Ambientales/análisis , Material Particulado/análisisRESUMEN
INTRODUCTION: Coronavirus disease 2019 (COVID-19) is a global pandemic that has affected millions of people worldwide. In this paper, we analyse the relationship between stress hyperglycaemia and disease severity in patients with COVID-19. MATERIAL AND METHODS: A total of 252 patients with COVID-19 were included in this study. The patients were divided into the following groups: COVID-19 with stress hyperglycaemia (SHG), COVID-19 with diabetes (DM), and COVID-19 with normal blood glucose (NG). The stress hyperglycaemia rate (SHR) was calculated using the fasting blood glucose (FBG)/glycated haemoglobin (HbA1c) ratio. To further compare the disease characteristics of different SHRs, we divided the SHR into low SHR and high SHR according to the SHR median. Correlations between the severity of the disease and other factors were analysed after adjusting for sex and age. Multivariate analysis was performed using logistic regression to analyse the risk factors predicting the severity of COVID-19. RESULTS: Compared with the NG group, the SHG group had higher disease severity (p < 0.001); the SHG group had higher HbA1c, FBG, SHR, blood urea nitrogen (BUN), interleukin 6 (IL-6), and neutrophil levels, while lymphocyte, CD3+ T cell, CD8+ T cell, CD4+ T cell, CD16+CD56 cell, and CD19+ cell counts were lower (p < 0.05). Compared with the NG group, the DM group had higher HbA1c, blood glucose, BUN, lactate dehydrogenase (LDH), erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), and neutrophils, while CD8+ T cell counts were lower (p < 0.05). Compared with the DM group, the SHG group had higher SHR and lower HbA1c, CD3+ T cell, CD4+ T cell, CD16+CD56 cell, and T cell ratio levels (p < 0.05). Compared to the low SHR group, the high SHR group had patients with more severe COVID-19 (p = 0.004). Also, the high SHR grouphad higher age, HbA1c, FBG, asparate aminotransferaze (AST), BUN, LDH, uric acid (UA), CRP, IL-6, and procalcitonin (PCT), while lymphocyte, CD3+ T cell, CD4+ T cell, CD8+ T cell, and CD19+ cell counts were lower (p < 0.05).Binary logistic regression analysis showed that SHR, gender, and lymphocyte count wererisk factorsfor the severity of COVID-19. CONCLUSION: Stress hyperglycaemia, as indicated by a higher SHR, is independently associated with the severity of COVID-19.
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COVID-19 , Hiperglucemia , Humanos , COVID-19/complicaciones , Interleucina-6 , Glucemia/análisis , Hemoglobina Glucada , Proteína C-Reactiva/análisis , Linfocitos T CD4-Positivos/química , Gravedad del Paciente , Estudios RetrospectivosRESUMEN
The current strategy to detect immunodominant T cell responses focuses on the antigen, employing large peptide pools to screen for functional cell activation. However, these approaches are labor and sample intensive and scale poorly with increasing size of the pathogen peptidome. T cell receptors (TCRs) recognizing the same epitope frequently have highly similar sequences, and thus, the presence of large sequence similarity clusters in the TCR repertoire likely identify the most public and immunodominant responses. Here, we perform a meta-analysis of large, publicly available single-cell and bulk TCR datasets from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected individuals to identify public CD4+ responses. We report more than 1,200 αßTCRs forming six prominent similarity clusters and validate histocompatibility leukocyte antigen (HLA) restriction and epitope specificity predictions for five clusters using transgenic T cell lines. Collectively, these data provide information on immunodominant CD4+ T cell responses to SARS-CoV-2 and demonstrate the utility of the reverse epitope discovery approach.
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COVID-19 , SARS-CoV-2 , Linfocitos T CD4-Positivos/química , Epítopos/análisis , Humanos , Receptores de Antígenos de Linfocitos T/genética , Especificidad del Receptor de Antígeno de Linfocitos TRESUMEN
BACKGROUND: Highly differentiated, senescent lymphocytes are pro-inflammatory and contribute to age-related systemic inflammation, called inflammageing. There are several reports of acute changes in senescent lymphocyte counts post exercise, which potentially have consequences for systemic inflammation. However, there is little consensus since the studies differ with respect to participants, exercise protocols, cellular markers assessed, and the time point of assessment post exercise. OBJECTIVE: We performed a systematic review and meta-analysis to assess the impact of exercise on senescent lymphocyte counts in blood immediately, 1 h and 2 h post exercise. METHODS: The search was performed in PubMed (MEDLINE), Web of Science, Embase, Scopus, and Cochrane, on January 11, 2021. The 13 studies selected tested aerobic exercise effects, mainly in young men. They assessed the counts of lymphocytes (CD4 T cells, CD8 T cells, and NK cells), with the following immune cell marker combinations: KLRG1+, CD57+ (only NK cells), EMRA T cells (CD45RA+CCR7-CD28-CD27-), CD28-CD27-, KLRG1+CD28-, and CD28-. Independent extraction of articles was done by 2 researchers. RESULTS: Standardized mean difference (SMD) and 95% confidence interval between baseline and post exercise showed significant increase (SMD >0.9, p < 0.003) in all types of senescent lymphocytes counts immediately post exercise. At 1 h post exercise, senescent CD4 T cells returned to baseline values (p = 0.74), CD8 T cells were reduced (-0.26 [-0.41; -0.11], p = 0.001), and senescent NK cells were raised (0.62 [0.14; 1.10], p = 0.01) above baseline. By 2 h post exercise, senescent CD4 T cells were reduced (-0.94 [-1.40; -0.48], p < 0.001), CD8 T cells remained below baseline (-0.53 [-1.04; -0.009], p = 0.04), and NK cells had returned to baseline values (-0.29 [-0.64; 0.07], p = 0.11). The main determinants of heterogeneity between studies were cytomegalovirus (CMV) serostatus and the characteristics of exercise protocols. CMV+ individuals had a higher immediate lymphocytosis and 1 h post lymphopenia than CMV- individuals. Exercise performed at higher intensities and shorter durations led to higher magnitude of change in senescent lymphocyte counts at all time-points. CONCLUSION: The differing effects of exercise on senescent NK cells and CD4 and CD8 T cells suggest differing susceptibility to factors modulating lymphocyte extravasation such as adrenaline and exercise intensity.
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Antígenos CD28 , Infecciones por Citomegalovirus , Antígenos CD28/análisis , Linfocitos T CD4-Positivos/química , Linfocitos T CD8-positivos/química , Ejercicio Físico , Citometría de Flujo , Humanos , Inflamación , Recuento de Linfocitos , MasculinoRESUMEN
OBJECTIVE: This study explored the role of IL-35 in CD4+ T lymphocyte and human skin fibroblast (HSF) activity and cytokine levels in systemic sclerosis. METHODS: Blood and skin biopsies were collected from 41 patients and 39 healthy controls to assess CD4+ T lymphocytes and IL-35-related factors. CD4+ T lymphocytes were co-cultured with HSFs, recombinant human IL-35 and IL-35 mAb to evaluate the cell viability, activation of CD4+ T lymphocytes and HSF cells. RESULTS: The proportion of blood Th1/Th2 was lower and Th17/Treg was higher in patients than in controls (P < 0.05). IL-35 and IL-17A levels were higher and IFN-γ, IL-10 and TGF-ß levels were lower in patients than in controls. IL-17A, forkhead box P3, TGF-ß1 and collagen type I (COL-1) mRNA and phospho (p)-signal transducer and activator of transcription (STAT) 1 and p-STAT4 were higher in skin tissues from patients than in those from controls (P < 0.05). IL-6 levels were higher, whereas IL-10 levels were lower in cell culture supernatants. α-Smooth muscle actin (α-SMA) and COL-1 proteins and Ki67 positivity were higher in CD4+ T + HSF cells from patients than in those from controls. Recombinant human IL-35 treatment inhibited proliferation (P < 0.001), but increased IL-10 and decreased IL-17A, α-SMA and COL-1 secretion into the conditioned medium of CD4+ T lymphocytes + HSFs from patients compared with those from controls. IL-35 mAb blocked the effects of IL-35 in CD4+ T + HSF cells (P < 0.05). CONCLUSIONS: IL-35 plays an inhibitory role in CD4+ T lymphocyte proliferation but induces Treg cell differentiation by STAT1 signalling activation, HSF proliferation and collagen expression in systemic sclerosis.
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Linfocitos T CD4-Positivos/química , Citocinas/sangre , Interleucinas/metabolismo , Esclerodermia Sistémica/metabolismo , Piel/metabolismo , Biopsia , Western Blotting , Estudios de Casos y Controles , Citocinas/análisis , Ensayo de Inmunoadsorción Enzimática , Femenino , Fibroblastos/metabolismo , Humanos , Interleucinas/análisis , Interleucinas/sangre , Masculino , Persona de Mediana Edad , Esclerodermia Sistémica/inmunología , Esclerodermia Sistémica/patología , Piel/química , Piel/patologíaRESUMEN
Detecting the entire repertoire of tumor-specific reactive tumor-infiltrating lymphocytes (TILs) is essential for investigating their immunological functions in the tumor microenvironment. Current in vitro assays identifying tumor-specific functional activation measure the upregulation of surface molecules, de novo production of antitumor cytokines, or mobilization of cytotoxic granules following recognition of tumor-antigens, yet there is no widely adopted standard method. Here we established an enhanced, yet simple, method for identifying simultaneously CD8+ and CD4+ tumor-specific reactive TILs in vitro, using a combination of widely known and available flow cytometry assays. By combining the detection of intracellular CD137 and de novo production of TNF and IFNγ after recognition of naturally-presented tumor antigens, we demonstrate that a larger fraction of tumor-specific and reactive CD8+ TILs can be detected in vitro compared to commonly used assays. This assay revealed multiple polyfunctionality-based clusters of both CD4+ and CD8+ tumor-specific reactive TILs. In situ, the combined detection of TNFRSF9, TNF, and IFNG identified most of the tumor-specific reactive TIL repertoire. In conclusion, we describe a straightforward method for efficient identification of the tumor-specific reactive TIL repertoire in vitro, which can be rapidly adopted in most cancer immunology laboratories.
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Antígenos de Neoplasias/inmunología , Linfocitos T CD4-Positivos/química , Linfocitos T CD8-positivos/química , Interferón gamma/análisis , Linfocitos Infiltrantes de Tumor/química , Proteínas de Neoplasias/análisis , Miembro 9 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/análisis , Factor de Necrosis Tumoral alfa/análisis , Antígenos CD/análisis , Apirasa/análisis , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Conjuntos de Datos como Asunto , Citometría de Flujo , Humanos , Cadenas alfa de Integrinas/análisis , Interferón gamma/biosíntesis , Interferón gamma/genética , Activación de Linfocitos/genética , Linfocitos Infiltrantes de Tumor/inmunología , Proteínas de Neoplasias/biosíntesis , Proteínas de Neoplasias/genética , Análisis de la Célula Individual , Transcriptoma , Microambiente Tumoral/inmunología , Factor de Necrosis Tumoral alfa/biosíntesis , Factor de Necrosis Tumoral alfa/genéticaRESUMEN
HLA-DQ8, a genetic risk factor in type I diabetes (T1D), presents hybrid insulin peptides (HIPs) to autoreactive CD4+ T cells. The abundance of spliced peptides binding to HLA-DQ8 and how they are subsequently recognised by the autoreactive T cell repertoire is unknown. Here we report, the HIP (GQVELGGGNAVEVLK), derived from splicing of insulin and islet amyloid polypeptides, generates a preferred peptide-binding motif for HLA-DQ8. HLA-DQ8-HIP tetramer+ T cells from the peripheral blood of a T1D patient are characterised by repeated TRBV5 usage, which matches the TCR bias of CD4+ T cells reactive to the HIP peptide isolated from the pancreatic islets of a patient with T1D. The crystal structure of three TRBV5+ TCR-HLA-DQ8-HIP complexes shows that the TRBV5-encoded TCR ß-chain forms a common landing pad on the HLA-DQ8 molecule. The N- and C-termini of the HIP is recognised predominantly by the TCR α-chain and TCR ß-chain, respectively, in all three TCR ternary complexes. Accordingly, TRBV5 + TCR recognition of HIP peptides might occur via a 'polarised' mechanism, whereby each chain within the αßTCR heterodimer recognises distinct origins of the spliced peptide presented by HLA-DQ8.
Asunto(s)
Diabetes Mellitus Tipo 1/metabolismo , Antígenos HLA-DQ/metabolismo , Insulina/metabolismo , Péptidos/metabolismo , Receptores de Antígenos de Linfocitos T/metabolismo , Secuencia de Aminoácidos , Linfocitos T CD4-Positivos/química , Linfocitos T CD4-Positivos/metabolismo , Diabetes Mellitus Tipo 1/genética , Antígenos HLA-DQ/química , Antígenos HLA-DQ/genética , Humanos , Insulina/química , Insulina/genética , Péptidos/química , Unión Proteica , Receptores de Antígenos de Linfocitos T/química , Receptores de Antígenos de Linfocitos T/genéticaRESUMEN
Mechanical forces acting on ligand-engaged T-cell receptors (TCRs) have previously been implicated in T-cell antigen recognition, yet their magnitude, spread, and temporal behavior are still poorly defined. We here report a FRET-based sensor equipped either with a TCR-reactive single chain antibody fragment or peptide-loaded MHC, the physiological TCR-ligand. The sensor was tethered to planar glass-supported lipid bilayers (SLBs) and informed most directly on the magnitude and kinetics of TCR-imposed forces at the single molecule level. When confronting T-cells with gel-phase SLBs we observed both prior and upon T-cell activation a single, well-resolvable force-peak of approximately 5 pN and force loading rates on the TCR of 1.5 pN per second. When facing fluid-phase SLBs instead, T-cells still exerted tensile forces yet of threefold reduced magnitude and only prior to but not upon activation.
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Transferencia Resonante de Energía de Fluorescencia/métodos , Antígenos de Histocompatibilidad/química , Receptores de Antígenos de Linfocitos T/química , Imagen Individual de Molécula/métodos , Anticuerpos de Cadena Única/química , Animales , Linfocitos T CD4-Positivos/química , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/química , Linfocitos T CD8-positivos/inmunología , Citocromos c/química , Transferencia Resonante de Energía de Fluorescencia/instrumentación , Antígenos de Histocompatibilidad/inmunología , Cinética , Ligandos , Membrana Dobles de Lípidos/química , Ratones , Péptidos/química , Receptores de Antígenos de Linfocitos T/inmunología , Imagen Individual de Molécula/instrumentación , Anticuerpos de Cadena Única/inmunología , Análisis Espacio-TemporalRESUMEN
PURPOSE: This pilot study aimed on generating insight on alterations in circulating immune cells during the use of FOLFIRINOX and gemcitabine/nab-paclitaxel in pancreatic ductal adenocarcinoma (PDAC). PATIENTS AND METHODS: Peripheral blood mononuclear cells were isolated before and 30 days after initiation of chemotherapy from 20 patients with advanced PDAC. Regulatory T cells (FoxP3+) and immune checkpoints (PD-1 and TIM-3) were analyzed by flow cytometry and immunological changes were correlated with clinical outcome. RESULTS: Heterogeneous changes during chemotherapy were observed in circulating T-cell subpopulations with a pronounced effect on PD-1+ CD4+/CD8+ T cells. An increase in FoxP3+ or PD-1+ T cells had no significant effect on survival. An increase in TIM3+/CD8+ (but not TIM3+/CD4+) T cells was associated with a significant inferior outcome: median progression-free survival in the subgroup with an increase of TIM-3+/CD8+ T cells was 6.0 compared to 14.0 months in patients with a decrease/no change (p = 0.026); corresponding median overall survival was 13.0 and 20.0 months (p = 0.011), respectively. CONCLUSIONS: Chemotherapy with FOLFIRNOX or gemcitabine/nab-paclitaxel induces variable changes in circulating T-cell populations that may provide prognostic information in PDAC.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Carcinoma Ductal Pancreático/tratamiento farmacológico , Proteínas de Punto de Control Inmunitario/efectos de los fármacos , Neoplasias Pancreáticas/tratamiento farmacológico , Linfocitos T Reguladores/efectos de los fármacos , Anciano , Albúminas/uso terapéutico , Linfocitos T CD4-Positivos/química , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD8-positivos/química , Linfocitos T CD8-positivos/efectos de los fármacos , Carcinoma Ductal Pancreático/inmunología , Desoxicitidina/análogos & derivados , Desoxicitidina/uso terapéutico , Femenino , Fluorouracilo/uso terapéutico , Factores de Transcripción Forkhead , Receptor 2 Celular del Virus de la Hepatitis A/análisis , Humanos , Proteínas de Punto de Control Inmunitario/análisis , Irinotecán/uso terapéutico , Leucovorina/uso terapéutico , Masculino , Persona de Mediana Edad , Oxaliplatino/uso terapéutico , Paclitaxel/uso terapéutico , Neoplasias Pancreáticas/inmunología , Proyectos Piloto , Receptor de Muerte Celular Programada 1/análisis , Receptor de Muerte Celular Programada 1/efectos de los fármacos , Supervivencia sin Progresión , Estudios Prospectivos , Linfocitos T Reguladores/química , GemcitabinaRESUMEN
Outcomes for triple negative breast cancer (TNBC) are poor and may be improved by increasing CD8+ tumor infiltrating lymphocytes (TIL) to augment antitumor immunity. Radiation (RT) can promote immunogenic cell death with increased antitumor T cell activity but also stimulates suppressive regulatory T cells (Tregs). Because metabolic alterations affect immune homeostasis and prior studies show caloric restriction (CR) combined with RT improves preclinical TNBC outcomes, we hypothesized that CR augments RT, in part, by altering intratumoral immunity. Using an in vivo model of TNBC, we treated mice with ad libitum (AL) diet, radiation, a CR diet, or CR + RT, and demonstrated an immune suppressive environment with a significant increase in CD4+ CD25+Foxp3+ Tregs after RT but not in CR-fed mice. CD8:Treg ratio in CR + RT TIL increased 4-fold compared with AL + RT mice. In vivo CD8 depletion was performed to assess the role of effector T cells in mitigating the effects of CR, and it was found that in mice undergoing CR, depletion of CD8 T cells resulted in increased tumor progression and decreased median survival compared with isotype control-treated mice. In addition, PD-1 expression on CD3+CD8+ T cells within the tumor microenvironment was significantly increased in CR + RT versus AL + RT treated mice as per immunofluorescence. Serum from breast cancer patients undergoing RT alone or CR and RT was collected pre- and postintervention, and a cytokine array demonstrated that patients treated with CR + RT had notable decreases in immunosuppressive cytokines such as IL-2Rγ, IL-10Rß, and TGF-ß2 and 3 compared with patients receiving RT alone. In conclusion, combining CR with RT decreases intratumoral Tregs, increases CD8:Treg, and increases PD-1 expression via a process dependent on CD8 T cells in a TNBC model. Breast cancer patients undergoing CR concurrently with RT also had significant reduction in immunosuppressive cytokine levels compared with those receiving RT alone.
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Restricción Calórica , Linfocitos Infiltrantes de Tumor/efectos de la radiación , Linfocitos T Reguladores/efectos de la radiación , Neoplasias de la Mama Triple Negativas/radioterapia , Microambiente Tumoral/efectos de la radiación , Adulto , Anciano , Animales , Linfocitos T CD4-Positivos/química , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/efectos de la radiación , Linfocitos T CD8-positivos/química , Linfocitos T CD8-positivos/metabolismo , Linfocitos T CD8-positivos/efectos de la radiación , Terapia Combinada/métodos , Progresión de la Enfermedad , Femenino , Citometría de Flujo , Factores de Transcripción Forkhead , Humanos , Subunidad gamma Común de Receptores de Interleucina/sangre , Subunidad beta del Receptor de Interleucina-10/sangre , Subunidad alfa del Receptor de Interleucina-2 , Depleción Linfocítica/métodos , Linfocitos Infiltrantes de Tumor/citología , Linfocitos Infiltrantes de Tumor/inmunología , Ratones , Ratones Endogámicos BALB C , Persona de Mediana Edad , Receptor de Muerte Celular Programada 1/metabolismo , Distribución Aleatoria , Linfocitos T Reguladores/citología , Linfocitos T Reguladores/inmunología , Factor de Crecimiento Transformador beta2/sangre , Factor de Crecimiento Transformador beta3/sangre , Neoplasias de la Mama Triple Negativas/sangre , Neoplasias de la Mama Triple Negativas/inmunología , Neoplasias de la Mama Triple Negativas/mortalidad , Microambiente Tumoral/inmunologíaRESUMEN
Peptide-major histocompatibility complex (pMHC) multimers enable the detection of antigen-specific T cells in studies ranging from vaccine efficacy to cancer immunotherapy. However, this technology is unreliable when applied to pMHC class II for the detection of CD4+ T cells. Here, using a combination of molecular biological and immunological techniques, we cloned sequences encoding human leukocyte antigen (HLA)-DP, HLA-DQ and HLA-DR molecules with enhanced CD4 binding affinity (with a Kd of 8.9 ± 1.1 µM between CD4 and affinity-matured HLA-DP4) and produced affinity-matured class II dimers that stain antigen-specific T cells better than conventional multimers in both in vitro and ex vivo analyses. Using a comprehensive library of dimers for HLA-DP4, which is the most frequent HLA allele in many ancestry groups, we mapped 103 HLA-DP4-restricted epitopes derived from diverse tumor-associated antigens and cloned the cognate T-cell antigen receptor (TCR) genes from in vitro-stimulated CD4+ T cells. The availability of affinity-matured class II dimers across HLA-DP, HLA-DQ and HLA-DR alleles will aid in the investigation of human CD4+ T-cell responses.
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Antígenos HLA , Antígenos de Histocompatibilidad Clase II , Coloración y Etiquetado/métodos , Antígenos CD4/química , Antígenos CD4/metabolismo , Linfocitos T CD4-Positivos/química , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/metabolismo , Células Cultivadas , Citometría de Flujo , Antígenos HLA/química , Antígenos HLA/metabolismo , Antígenos de Histocompatibilidad Clase II/química , Antígenos de Histocompatibilidad Clase II/metabolismo , Humanos , Unión ProteicaRESUMEN
A vaccine which is effective against the HIV virus is considered to be the best solution to the ongoing global HIV/AIDS epidemic. In the past thirty years, numerous attempts to develop an effective vaccine have been made with little or no success, due, in large part, to the high mutability of the virus. More recent studies showed that a vaccine able to elicit broadly neutralizing antibodies (bnAbs), that is, antibodies that can neutralize a high fraction of global virus variants, has promise to protect against HIV. Such a vaccine has been proposed to involve at least three separate stages: First, activate the appropriate precursor B cells; second, shepherd affinity maturation along pathways toward bnAbs; and, third, polish the Ab response to bind with high affinity to diverse HIV envelopes (Env). This final stage may require immunization with a mixture of Envs. In this paper, we set up a framework based on theory and modeling to design optimal panels of antigens to use in such a mixture. The designed antigens are characterized experimentally and are shown to be stable and to be recognized by known HIV antibodies.
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Vacunas contra el SIDA/biosíntesis , Antígenos Virales/química , Anticuerpos ampliamente neutralizantes/biosíntesis , Epítopos/química , Anticuerpos Anti-VIH/biosíntesis , Infecciones por VIH/prevención & control , VIH-1/inmunología , Vacunas contra el SIDA/química , Vacunas contra el SIDA/genética , Secuencia de Aminoácidos , Antígenos Virales/genética , Antígenos Virales/inmunología , Sitios de Unión , Anticuerpos ampliamente neutralizantes/química , Linfocitos T CD4-Positivos/química , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/virología , Cristalografía por Rayos X , Epítopos/genética , Epítopos/inmunología , Anticuerpos Anti-VIH/química , Proteína gp120 de Envoltorio del VIH/química , Proteína gp120 de Envoltorio del VIH/genética , Proteína gp120 de Envoltorio del VIH/metabolismo , Proteínas gp160 de Envoltorio del VIH/química , Proteínas gp160 de Envoltorio del VIH/genética , Proteínas gp160 de Envoltorio del VIH/metabolismo , Proteína gp41 de Envoltorio del VIH/química , Proteína gp41 de Envoltorio del VIH/genética , Proteína gp41 de Envoltorio del VIH/metabolismo , Infecciones por VIH/inmunología , Infecciones por VIH/virología , VIH-1/química , VIH-1/genética , Humanos , Modelos Moleculares , Mutación , Unión Proteica , Conformación Proteica en Hélice alfa , Conformación Proteica en Lámina beta , Ingeniería de Proteínas/métodos , Dominios y Motivos de Interacción de ProteínasRESUMEN
Inducible expression of neoantigens in mice would enable the study of endogenous antigen-specific naïve T cell responses in disease and infection, but has been difficult to generate because leaky antigen expression in the thymus results in central T cell tolerance. Here we develop inversion-induced joined neoantigen (NINJA), using RNA splicing, DNA recombination and three levels of regulation to prevent leakiness and allow tight control over neoantigen expression. We apply NINJA to create tumor cell lines with inducible neoantigen expression, which could be used to study antitumor immunity. We also show that the genetic regulation in NINJA mice bypasses central and peripheral tolerance mechanisms and allows for robust endogenous CD8 and CD4 T cell responses on neoantigen induction in peripheral tissues. NINJA will enable studies of how T cells respond to defined neoantigens in the context of peripheral tolerance, transplantation, autoimmune diseases and cancer.
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Antígenos de Neoplasias , Ingeniería Celular/métodos , Animales , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/metabolismo , Linfocitos T CD4-Positivos/química , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD8-positivos/química , Linfocitos T CD8-positivos/metabolismo , Femenino , Humanos , Ratones , Especificidad de Órganos/genética , Empalme del ARN/genética , Células Tumorales CultivadasRESUMEN
The interaction between T cell receptor (TCR) and peptide (p)-Human Leukocyte Antigen (HLA) complexes is the critical first step in determining T cell responses. X-ray crystallographic studies of pHLA in TCR-bound and free states provide a structural perspective that can help understand T cell activation. These structures represent a static "snapshot", yet the nature of pHLAs and their interactions with TCRs are highly dynamic. This has been demonstrated for HLA class I molecules with in silico techniques showing that some interactions, thought to stabilise pHLA-I, are only transient and prone to high flexibility. Here, we investigated the dynamics of HLA class II molecules by focusing on three allomorphs (HLA-DR1, -DR11 and -DR15) that are able to present the same epitope and activate CD4+ T cells. A single TCR (F24) has been shown to recognise all three HLA-DR molecules, albeit with different affinities. Using molecular dynamics and crystallographic ensemble refinement, we investigate the molecular basis of these different affinities and uncover hidden roles for HLA polymorphic residues. These polymorphisms were responsible for the widening of the antigen binding cleft and disruption of pHLA-TCR interactions, underpinning the hierarchy of F24 TCR binding affinity, and ultimately T cell activation. We expanded this approach to all available pHLA-DR structures and discovered that all HLA-DR molecules were inherently rigid. Together with in vitro protein stability and peptide affinity measurements, our results suggest that HLA-DR1 possesses inherently high protein stability, and low HLA-DM susceptibility.
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Antígenos/química , Antígenos HLA-DR/química , Receptores de Antígenos de Linfocitos T/química , Antígenos/inmunología , Linfocitos T CD4-Positivos/química , Linfocitos T CD4-Positivos/inmunología , Cristalografía por Rayos X , Células HEK293 , Antígenos HLA-DR/inmunología , Humanos , Receptores de Antígenos de Linfocitos T/inmunologíaRESUMEN
A low count of CD4+ and CD8+ lymphocytes is a hallmark laboratory finding in the coronavirus disease 2019 (COVID-19). Using flow cytometry, we observed significantly higher CD95 (Fas) and PD-1 expression on both CD4+ T and CD8+ T cells in 42 COVID-19 patients when compared to controls. Higher CD95 expression in CD4+ cells correlated with lower CD4+ counts. A higher expression of CD95 in CD4+ and CD8+ lymphocytes correlated with a lower percentage of naive events. Our results might suggest a shift to antigen-activated T cells, expressing molecules increasing their propensity to apoptosis and exhaustion during COVID-19 infection.
Asunto(s)
Linfocitos T CD4-Positivos/química , Linfocitos T CD8-positivos/química , COVID-19/inmunología , Subgrupos Linfocitarios/química , Linfopenia/etiología , Receptor de Muerte Celular Programada 1/sangre , Receptor fas/sangre , Adulto , Anciano , Anciano de 80 o más Años , Envejecimiento/sangre , Envejecimiento/inmunología , Apoptosis , COVID-19/sangre , COVID-19/complicaciones , Femenino , Humanos , Recuento de Linfocitos , Masculino , Persona de Mediana Edad , Pronóstico , SARS-CoV-2RESUMEN
Sterile α motif and histidine/aspartic acid domaincontaining protein 1 (SAMHD1) can inhibit reverse transcription of human immunodeficiency virus1 (HIV1) by hydrolyzing intracellular deoxyribonucleoside triphosphate. However, its role in HIV1 disease progression has not been extensively studied. To study the impacts of SAMHD1 on HIV1 disease progression, especially on DNA levels, we investigated SAMHD1 levels in the peripheral blood of HIV1 elite controllers (ECs), antiretroviral therapy (ART) naive viremic progressors (VPs) and patients with HIV1 receiving ART (HIVARTs) compared with healthy controls. In addition, the present study analyzed the relationship between SAMHD1 and interferonα, immune activation and HIV1 DNA levels. The results of the present study demonstrated elevated SAMHD1 expression in the peripheral blood mononuclear cells of all patients withHIV1, but higher SAMHD1 expression in the CD4+ T cells of only ECs compared with healthy controls. Immune activation was increased in the VPs and decreased in the ECs compared with healthy controls. Substantially lower HIV1 DNA levels were identified in ECs compared with those in VPs and HIVARTs. SAMHD1 expression was associated with low levels of immune activation. No significant correlation was observed between SAMHD1 and HIV1 DNA levels. Overall, the findings of the present study indicated that SAMHD1 was highly expressed in ECs, which may be associated with low immune activation levels, but was not directly related to HIV1 DNA levels.
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Linfocitos T CD4-Positivos/química , Linfocitos T CD4-Positivos/inmunología , ADN Viral/sangre , Infecciones por VIH/inmunología , VIH-1/química , Proteína 1 que Contiene Dominios SAM y HD/sangre , Proteína 1 que Contiene Dominios SAM y HD/inmunología , Adulto , Antirretrovirales/uso terapéutico , Femenino , Regulación Enzimológica de la Expresión Génica , Infecciones por VIH/sangre , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/metabolismo , VIH-1/genética , Humanos , Interferón-alfa/sangre , Leucocitos Mononucleares/química , Masculino , Persona de Mediana Edad , ARN Viral/sangre , Proteína 1 que Contiene Dominios SAM y HD/genética , Carga ViralRESUMEN
Dengue virus (DENV) is a mosquito-borne flavivirus that causes serious human disease. The current lack of an effective vaccine to simultaneously protect against the four serotypes of DENV in seronegative individuals is a major unmet medical need. Further, the immunological basis for protective immunity in the setting of DENV infection or vaccination is not fully understood. Our team has developed a live attenuated tetravalent dengue virus vaccine that provides complete protection in a human model of dengue virus challenge. The goal of this study was to define, in the context of protective human vaccination, the quality of vaccine-induced DENV-specific CD8+ and CD4+ T cells and the temporal dynamics associated with their formation and maintenance. Multifunctional, DENV-specific CD8+ and CD4+ T cells developed 8-14 days after vaccination and were maintained for at least 6 months. Virus-specific CD8 T+ cells were a mixture of effector memory T cells (TEM) and effector memory T cells re-expressing CD45RA (TEMRA), with TEM cells predominating until day 21 post-vaccination and TEMRA cells thereafter. The majority of virus-specific CD4+ T cells were TEM with a small fraction being TEMRA. The frequency of virus-specific CD8+ and CD4+ T cells were further skewed to the TEMRA phenotype following either a second dose of the tetravalent vaccine or challenge with a single serotype of DENV. Collectively, our study has defined the phenotypic profile of antiviral CD8+ and CD4+ T cells associated with protective immunity to DENV infection and the kinetics of their formation and maintenance.
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Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Vacunas contra el Dengue/inmunología , Virus del Dengue/inmunología , Antígenos Comunes de Leucocito/análisis , Subgrupos Linfocitarios/inmunología , Especificidad del Receptor de Antígeno de Linfocitos T , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Antígenos Virales/inmunología , Linfocitos T CD4-Positivos/química , Linfocitos T CD8-positivos/química , Ensayos Clínicos Fase I como Asunto , Citocinas/análisis , Virus del Dengue/genética , Epítopos/inmunología , Humanos , Inmunización Secundaria , Inmunogenicidad Vacunal , Memoria Inmunológica , Subgrupos Linfocitarios/química , Factores de Tiempo , Vacunación , Vacunas Atenuadas/inmunologíaRESUMEN
Single-cell RNA sequencing (scRNA-seq) allows the identification, characterization, and quantification of cell types in a tissue. When focused on B and T cells of the adaptive immune system, scRNA-seq carries the potential to track the clonal lineage of each analyzed cell through the unique rearranged sequence of its antigen receptor (BCR or TCR, respectively) and link it to the functional state inferred from transcriptome analysis. Here we introduce FB5P-seq, a FACS-based 5'-end scRNA-seq method for cost-effective, integrative analysis of transcriptome and paired BCR or TCR repertoire in phenotypically defined B and T cell subsets. We describe in detail the experimental workflow and provide a robust bioinformatics pipeline for computing gene count matrices and reconstructing repertoire sequences from FB5P-seq data. We further present two applications of FB5P-seq for the analysis of human tonsil B cell subsets and peripheral blood antigen-specific CD4 T cells. We believe that our novel integrative scRNA-seq method will be a valuable option to study rare adaptive immune cell subsets in immunology research.
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Subgrupos Linfocitarios/química , RNA-Seq/métodos , Receptores de Antígenos de Linfocitos B/genética , Receptores de Antígenos de Linfocitos T alfa-beta/genética , Análisis de la Célula Individual/métodos , Transcriptoma , Regiones no Traducidas 5' , Inmunidad Adaptativa , Adulto , Linfocitos B/química , Linfocitos T CD4-Positivos/química , Linaje de la Célula , Biología Computacional , Análisis Costo-Beneficio , Epítopos , Femenino , Citometría de Flujo , Humanos , Masculino , Tonsila Palatina/citología , RNA-Seq/economía , Análisis de la Célula Individual/economía , Flujo de TrabajoRESUMEN
Iron is a key factor at various stages of HIV life cycle and determines the progression of HIV infection. Data about cellular labile iron pool (LIP) in the settings of contemporary antiretroviral therapy (cART) are lacking. Yet LIP is directly related to the generation of reactive oxygen species, and may contribute to immune activation, dysfunction, and exhaustion. Using multiparameter flow cytometry, we evaluated LIP in CD4 and CD8 T cells from HIV+ patients with sustained viral suppression (SVS) as a result of continuous long-term cART. Based on the recovery of CD4/CD8 ratio, two patients' subgroups were defined: A (n = 26), CD4/CD8 > 0.9, and B (n = 37), CD4/CD8 < 0.9, with significantly differing CD4 absolute count (AC) (mean 752 vs. 571 cells/µL, p < .05). Although hemoglobin and serum iron had recovered in all patients, CD4 T cell LIP and CD8 T cell LIP were significantly higher than that of controls, both in the subgroup with complete (A) and with incomplete (B) immune recovery [mean CD4 mean fluorescence intensity (ΔMFI) 318.7 and 777.8 vs. 157.6; mean CD8 ΔMFI 359.5 and 628.7 vs. 179.2, analysis of variance p < .05 for both]. CD4 LIP correlated inversely with CD4 AC (R = -0.4, p < .01), and both CD4 LIP and CD8 LIP-with CD4/CD8 ratio (R = -0.4, p < .01). Thus, increased CD4 T cell LIP and CD8 T cell LIP in the settings of SVS and immune recovery are a sensitive marker of residual immune activation and may predict immune exhaustion in long-term cART-treated patients.
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Linfocitos T CD4-Positivos/química , Linfocitos T CD8-positivos/química , Infecciones por VIH/inmunología , Hierro/análisis , Carga Viral , Adulto , Terapia Antirretroviral Altamente Activa , Recuento de Linfocito CD4 , Relación CD4-CD8 , Progresión de la Enfermedad , Citometría de Flujo , Estudios de Seguimiento , Infecciones por VIH/fisiopatología , Humanos , Masculino , Persona de Mediana EdadRESUMEN
The highly homologous human leukocyte antigen (HLA)-DQ2 molecules, HLA-DQ2.5 and HLA-DQ2.2, are implicated in the pathogenesis of celiac disease (CeD) by presenting gluten peptides to CD4+ T cells. However, while HLA-DQ2.5 is strongly associated with disease, HLA-DQ2.2 is not, and the molecular basis underpinning this differential disease association is unresolved. We here provide structural evidence for how the single polymorphic residue (HLA-DQ2.5-Tyr22α and HLA-DQ2.2-Phe22α) accounts for HLA-DQ2.2 additionally requiring gluten epitopes possessing a serine at the P3 position of the peptide. In marked contrast to the biased T cell receptor (TCR) usage associated with HLA-DQ2.5-mediated CeD, we demonstrate with extensive single-cell sequencing that a diverse TCR repertoire enables recognition of the immunodominant HLA-DQ2.2-glut-L1 epitope. The crystal structure of two CeD patient-derived TCR in complex with HLA-DQ2.2 and DQ2.2-glut-L1 (PFSEQEQPV) revealed a docking strategy, and associated interatomic contacts, which was notably distinct from the structures of the TCR:HLA-DQ2.5:gliadin epitope complexes. Accordingly, while the molecular surfaces of the antigen-binding clefts of HLA-DQ2.5 and HLA-DQ2.2 are very similar, differences in the nature of the peptides presented translates to differences in responding T cell repertoires and the nature of engagement of the respective antigen-presenting molecules, which ultimately is associated with differing disease penetrance.