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BACKGROUND: Tumor PD-L1 expression is a predictive biomarker for patients with NSCLC receiving PD-(L)1 blockade agents. However, although increased tumor PD-L1 expression predicts responsiveness, clinical benefit has been observed regardless of tumor PD-L1 expression, suggesting the existence of other PD-L1 sources. The aim of our study was to analyze whether integrating systemic and tumor PD-L1 is more predictive of efficacy in patients with advanced NSCLC receiving PD-(L)1 blockade agents. MATERIAL AND METHODS: Twenty-nine healthy donors and 119 consecutive patients with advanced NSCLC treated with PD-(L)1 drug were prospectively included. Pretreatment blood samples were collected to evaluate PD-L1 levels on circulating immune cells, platelets (PLTs), platelet microparticles (PMPs), and the plasma soluble PD-L1 concentration (sPD-L1). Tumor PD-L1 status was assessed by immunohistochemistry. The percentages of circulating PD-L1 + leukocytes, sPD-L1 levels, and tumor PD-L1 were correlated with efficacy. RESULTS: No differences in the percentages of circulating PD-L1 + leukocytes were observed according to tumor PD-L1 expression. Significantly longer progression-free survival was observed in patients with higher percentages of PD-L1 + CD14 + , PD-L1 + neutrophils, PD-L1 + PLTs, and PD-L1 + PMPs and significantly longer overall survival was observed in patients with higher percentages of PD-L1 + CD14 + and high tumor PD-L1 expression. Integrating the PD-L1 data of circulating and tumor PD-L1 results significantly stratified patients according to the efficacy of PD-(L1) blockade agents. CONCLUSIONS: Our results suggest that integrating circulating PD-L1 + leukocytes, PLT, PMPs, and sPD-L1 and tumor PD-L1 expression may be helpful to decide on the best treatment strategy in patients with advanced NSCLC who are candidates for PD-(L)1 blockade agents.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Antígeno B7-H1 , Biomarcadores de Tumor/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/patología , Humanos , Inmunoterapia , Neoplasias Pulmonares/patologíaRESUMEN
OBJECTIVE: To study subclinical inflammation in intercritical gout patients and its relation to the estimated size of monosodium urate crystal deposition and cardiovascular risk factors. METHODS: We performed a secretome analysis and the quantification of cytokine and adipokine plasma levels [IL-1ß, IL-18, IL-6, soluble IL-6 receptor (sIL-6R), TNF-α, C-X-C motif chemokine 5, RANTES (Regulated upon Activation, Normal T Cell Expressed and Presumably Secreted), leptin, resistin and adiponectin] to analyse subclinical inflammation in intercritical gout patients. Since it is currently not feasible to determinate the whole body deposit of monosodium urate crystals, we created an indirect clinical classification to estimate it. Then we compared cytokine levels in controls and gout patients and in patients with different crystal deposition sizes. We also studied the association between cytokine-levels and the number of cardiovascular risk factors. RESULTS: Ninety consecutive patients attending a crystal arthritis unit were studied. IL-18, sIL-6R, RANTES, leptin and adiponectin were higher in intercritical gout patients than in controls. An association was observed between IL-18, sIL6-R and RANTES levels and the size of crystal deposition. IL-18, sIL6-R, RANTES and leptin were higher in patients with no cardiovascular risk factors compared with controls with no risk factors. CONCLUSION: Our results showed that the levels of some pro-inflammatory cytokines and metabolic proteins are elevated in intercritical gout patients. The levels of certain cytokines were related to the estimated size of the monosodium urate crystal deposition and to the number of cardiovascular risk factors. These cytokine changes may help to explain the increase in cardiovascular events in gout patients.
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Gota/metabolismo , Ácido Úrico , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Casos y Controles , Cristalización , Citocinas/sangre , Femenino , Humanos , Masculino , Persona de Mediana Edad , SecretomaRESUMEN
BACKGROUND: Anti-PD-(L)1 blocking agents can induce immune-related adverse events (irAEs), which can compromise treatment continuation. Since circulating leukocyte-platelet (PLT) complexes contribute to inflammatory and autoimmune diseases, we aimed to analyze the role of these complexes as predictors of irAEs in non-small cell lung cancer (NSCLC) patients receiving anti-PD-(L)1. MATERIALS AND METHODS: Twenty-six healthy donors (HD) and 87 consecutive advanced NSCLC patients treated with anti-PD-(L)1 were prospectively included. Percentages of circulating leukocyte-PLT complexes were analyzed by flow cytometry and compared between HD and NSCLC patients. The association of leukocyte-PLT complexes with the presence and severity of irAEs was analyzed. RESULTS: NSCLC patients had higher percentages of circulating leukocyte-PLT complexes. Higher percentages of monocytes with bound PLT (CD14 + PLT +) were observed in patients who received prior therapies while CD4 + T lymphocytes with bound PLT (CD4 + PLT +) correlated with platelets counts. The CD4 + PLT + high percentage group presented a higher rate of dermatological irAEs while the CD4 + PLT + low percentage group showed a higher rate of non-dermatological irAEs (p < 0.001). A lower frequency of grade ≥ 2 irAEs was observed in the CD4 + PLT + high percentage group (p < 0.05). Patients with CD4 + PLT + low and CD14 + PLT + high percentages presented a higher rate of grade ≥ 3 irAEs and predominantly developed non-dermatological irAEs (p < 0.01). CONCLUSIONS: Our results suggest that circulating leukocyte-PLT complexes and the combination of CD4 + PLT + and CD14 + PLT + percentages can be used as a predictive biomarker of the development and severity of irAEs in advanced NSCLC patients receiving anti-PD-(L)1 agents.
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Antígeno B7-H1/metabolismo , Plaquetas/patología , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos/diagnóstico , Inhibidores de Puntos de Control Inmunológico/efectos adversos , Leucocitos/patología , Neoplasias Pulmonares/tratamiento farmacológico , Adenocarcinoma del Pulmón/tratamiento farmacológico , Adenocarcinoma del Pulmón/inmunología , Adenocarcinoma del Pulmón/metabolismo , Adenocarcinoma del Pulmón/patología , Anciano , Biomarcadores de Tumor/análisis , Carcinoma de Pulmón de Células no Pequeñas/inmunología , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/patología , Carcinoma de Células Escamosas/tratamiento farmacológico , Carcinoma de Células Escamosas/inmunología , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patología , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos/etiología , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos/metabolismo , Femenino , Estudios de Seguimiento , Humanos , Neoplasias Pulmonares/inmunología , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Masculino , Persona de Mediana Edad , Pronóstico , Estudios Prospectivos , Tasa de SupervivenciaRESUMEN
BACKGROUND AND AIMS: Crohn's disease [CD] is a chronic, systemic inflammatory disease characterised by periods of remission and flare-ups. It has been associated with a disturbed gastrointestinal barrier function, an increase in the transport of luminal contents into the tissue, and lower immune tolerance. METHODS: Peripheral blood samples were collected from healthy controls and 33 adult active flare-up CD patients. We classified patients as onset or relapse flare-up subjects, according to the days of disease evolution. Plasma levels of lipopolysaccharide-binding protein [LBP], fatty acid-binding proteins [FABP], and antibodies against bacterial lysates, interferons [IFN] and interleukin-6 [IL6] were measured by enzyme-linked immunosorbent assay [ELISA] in each group of patients. RESULTS: Onset CD patients had higher plasma levels of LBP [57.32 ± 38.86 vs 30.22 ± 9.80 µg/ml] and IFNα [1.25 ± 0.23 vs 0.95 ± 0.36 log10pg/ml] and lower levels of immunoglobulins G and A [IgG and IgA] antibodies against bacterial lysates than relapse CD patients. We also observed a subgroup of onset patients with the highest levels of LBP. In this subgroup, LBP correlated negatively with C-reactive protein [CRP]. Onset and relapse CD patients had similar levels of FABP6 and FABP2, though LBP and FABP6 correlated positively only in relapse patients. In relapse patients, anti-E coli IgG antibodies correlated positively with systemic IL6 and IFNα levels. CONCLUSIONS: Our findings suggest that onset and relapse flare-ups in adult CD patients are related to different systemic immune-related bacterial events. Characterising these differences may provide insights into the aetiology of Crohn's disease, and would help in the selection of appropriate therapies.
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Anticuerpos/sangre , Enfermedad de Crohn/inmunología , Enfermedad de Crohn/microbiología , Escherichia coli/inmunología , Proteínas de Fase Aguda , Adolescente , Adulto , Anticuerpos Antibacterianos/sangre , Proteínas Portadoras/sangre , Estudios de Casos y Controles , Enfermedad de Crohn/sangre , Proteínas de Unión a Ácidos Grasos/sangre , Femenino , Hormonas Gastrointestinales/sangre , Humanos , Inmunoglobulina A/sangre , Inmunoglobulina G/sangre , Inmunoglobulina M/sangre , Interferón-alfa/sangre , Interleucina-6/sangre , Masculino , Glicoproteínas de Membrana/sangre , Persona de Mediana Edad , Recurrencia , Brote de los Síntomas , Adulto JovenRESUMEN
Soluble factors released from platelets can modulate the immune response of leukocytes. We and others have recently found that T lymphocytes with bound platelets have reduced proliferation and IFN-γ and IL-17 production. Thus, we speculate that if we induce the binding of platelets to lymphocytes, we will be able to regulate the inflammatory response. When we cocultured platelets with lymphocytes at different ratios, we were able to increase the percentage of lymphocytes with bound platelets. The coculture of platelets with lymphocytes in the presence of stimulation decreased the production of IFN-γ and TNF-α, T cell proliferation, and the expression of CD25, PD-L1, and SLAM. However, this coculture increased CD39 expression. All of these effects were dependent on the dose of platelets and operated indistinctly with platelets from different healthy donors. When platelets were cocultured in the same compartment with lymphocytes, we observed less IFN-γ and TNF-α production and T lymphocyte proliferation than in cultures with platelets separated from lymphocytes by a 0.4-µm pore size filter. The binding of platelets to lymphocytes was blocked with anti-P-selectin Abs, and when this occurred we observed higher IFN-γ and TNF-α production than in nonblocked conditions. The cocultures of platelets with synovial fluid cells from rheumatoid arthritis patients reduced inflammatory cytokine production and increased IL-10 production. These results suggest that platelet binding to lymphocytes effectively regulates T lymphocyte function. This mechanism could be easily applied to reduce inflammatory responses.
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Artritis Reumatoide/inmunología , Plaquetas/inmunología , Citocinas/biosíntesis , Activación de Linfocitos/inmunología , Linfocitos T/inmunología , Técnicas de Cocultivo , Femenino , Citometría de Flujo , Humanos , Masculino , Persona de Mediana EdadRESUMEN
INTRODUCTION: Adalimumab is a fully human anti-tumor necrosis factor α (anti-TNFα) monoclonal antibody that specifically blocks the interaction of TNFα with its receptors. It binds both soluble and transmembrane TNFα. We hypothesized that blocking these TNFα signals regulates the altered TNFα production in rheumatoid arthritis (RA) patients. METHODS: We compared, by flow cytometry, Toll-like receptor induction levels of membrane and intracellular TNFα in monocytes (iTNFα + CD14+ cells) from 12 patients before and after adalimumab treatment with those from 5 healthy donors. RESULTS: Before starting the treatment, the percentage of iTNFα+ CD14+ cells in the RA patients was significantly lower than that in healthy donors (mean ± SEM = 33.16 ± 4.82% vs 66.51 ± 2.4%, P < 0.001). When we added in vitro TNFα to healthy donor culture cells, levels of iTNFα+ CD14+ cells decreased, suggesting that the TNFα signal was responsible for the iTNFα+ CD14+ cell downregulation observed in the RA patients. After 2, 6 and 12 adalimumab injections, we observed significant blocking of membrane and soluble TNFα and a progressive increase in iTNFα+ CD14+ cells in ten patients with a good to moderate response as defined by the European League Against Rheumatism (EULAR) criteria. Levels of iTNFα+ CD14+ cells after 12 injections in these 10 patients were comparable to levels in healthy donors. In two patients, iTNFα+ CD14+ cell upregulation was not observed, and their EULAR-defined responses had not improved. The first patient developed antiadalimumab antibodies, explaining why adalimumab was not able to block membrane and soluble TNFα. In the second patient, adalimumab was discontinued because of adverse effects, which led to a decrease in iTNFα+ CD14+ cells to levels measured before treatment. CONCLUSIONS: Our findings suggest that adalimumab treatment in RA patients can return iTNFα levels to those of healthy donors. This effect was not observed in the presence of neutralizing antiadalimumab antibodies.
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Anticuerpos Monoclonales Humanizados/uso terapéutico , Antirreumáticos/uso terapéutico , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/inmunología , Factor de Necrosis Tumoral alfa/biosíntesis , Adalimumab , Artritis Reumatoide/metabolismo , Membrana Celular/metabolismo , Citoplasma/metabolismo , Femenino , Citometría de Flujo , Humanos , Espacio Intracelular/metabolismo , Masculino , Persona de Mediana Edad , Monocitos/metabolismoRESUMEN
Expression of the scavenger receptor CD36 on lymphocytes is intriguing. We observed that a minor subpopulation of lymphocytes expressed CD36 on the cell surface. We investigated the source of CD36 and also the proliferation and cytokine production of these CD36(+) CD4(+) lymphocytes. Flow cytometry analysis and immunofluorescence microscopy showed that CD36(+) platelets were responsible for CD36 detection on lymphocytes. CD36 was then used as a tool to characterize lymphocytes with bound platelets. Activation-induced proliferation was lower in CD4(+) lymphocytes with bound platelets than lymphocytes without bound platelets. IL-17 and IFN-γ production was also reduced in lymphocytes with bound platelets. We then studied the presence of CD36(+) CD4(+) lymphocytes in RA patients. We observed that the percentage of CD4(+) lymphocytes with bound platelets was higher on RA patients than in healthy donors. RA patients with higher titers of anti-CCP, RF levels, and cardiovascular risk index presented a lower percentage of CD4(+) lymphocytes with bound platelets. These patients also had higher IL-17 and IFN-γ production. These results suggest that platelet-binding modifies lymphocyte function. This binding could be a regulatory mechanism in RA that confers a less severe phenotype.
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Artritis Reumatoide/inmunología , Plaquetas/fisiología , Antígenos CD36/fisiología , Linfocitos T CD4-Positivos/fisiología , Complejo CD3/análisis , Antígenos CD36/análisis , Citocinas/biosíntesis , Humanos , Memoria Inmunológica , Inmunofenotipificación , Activación de Linfocitos , Receptores de Antígenos de Linfocitos T/fisiologíaRESUMEN
Chemokine receptors are distinctively exposed on cells to characterize their migration pattern. However, little is known about factors that may regulate their expression. To determine the optimal conditions for an accurate analysis of chemokine receptors, we compared the expression of CCR2, CCR4, CCR5, CCR6, CXCR3 and CXCR4 on different leukocyte subsets using whole blood (WB) plus erythrocyte lysis and density gradient isolation (Ficoll). Most WB monocytes were CCR2+ (93.5 ± 2.9%) whereas 32.8 ± 6.0% of monocytes from Ficoll-PBMC expressed CCR2 (p<0.001). Significant reductions of CCR6 and CXCR3 on monocytes were also observed after Ficoll isolation (WB: 46.4 ± 7.5% and 57.1 ± 5.5%; Ficoll: 29.5 ± 2.2% and 5.4 ± 4.3% respectively) (p<0.01). Although comparable percentages of WB and Ficoll-PBMC monocytes expressed CCR4, CCR5 and CXCR4, Ficoll isolation significantly reduced the levels of CXCR4 (WB: MFI 5 ± 0.4 and Ficoll: MFI 3.3 ± 0.1) (p<0.05). Similarly to monocytes, CCR2, CXCR3 and CXCR4 were also reduced on lymphocytes. In addition, Ficoll isolation significantly reduced the percentage of CCR4 positive lymphocytes (WB: 90.2 ± 4.5% and Ficoll: 55 ± 4.1%) (p<0.01). The loss of expression of chemokine receptors after isolation of monocytes was not dependent on either the anticoagulant or the density gradient method. It was irreversible and could not be restored by LPS activation or in vitro macrophage differentiation. Experiments tagged with anti-CCR2 antibodies prior to density gradient isolation demonstrated that Ficoll internalized chemokine receptors. The method for cell isolation may alter not only the expression of certain chemokine receptors but also the respective functional migration assay. The final choice to analyze their expression should therefore depend on the receptor to be measured.
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Separación Celular/métodos , Regulación de la Expresión Génica , Leucocitos/citología , Leucocitos/metabolismo , Receptores de Quimiocina/metabolismo , Animales , Anticoagulantes/farmacología , Movimiento Celular/efectos de los fármacos , Células Cultivadas , Regulación hacia Abajo/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Leucocitos/efectos de los fármacos , Lipopolisacáridos/farmacología , Monocitos/citología , Monocitos/efectos de los fármacos , Monocitos/metabolismo , Coloración y Etiquetado , Factores de TiempoRESUMEN
Genetic polymorphisms of the renin-angiotensin system (RAS) have been implicated in the pathogenesis of nephropathy and end-stage renal disease (ESRD). The association between angiotensin-converting enzyme (ACE) gene polymorphism and nephropathy evolution was studied. A random sample of 161 subjects from the Nephrology Department (of Hospital de Sant Pau) were divided into two groups: (i) 117 with end-stage renal disease; (ii) 44 with established nephropathy; and (iii) control groups of 129 subjects. The ACE gene polymorphism was performed by using polymerase chain reaction. High DD genotype presentation was observed in the two groups of subjects with nephropathy (46.12 and 61.37%, respectively vs 35.66% in controls; P < 0.0482), and also, a decrease was observed in the II genotype (6.4 and 4.54%, respectively vs 13.17% in controls, P < 0.0404). Glomerular filtration rate (GFR) was evaluated after 44 months of follow up. An important decrease of GFR was observed in patients with DD polymorphism versus other genotypes (initial, 32.3 +/- 7.9 and at 44 months, 18.35 +/- 3.3 mL/min vs 31.4 +/- 11.9 and 11.7 +/- 3.2 mL/min; P < 0.039). In a non-longitudinal study of patients in ESRD, patients with an ACE-DD genotype had a lower period of time between diagnosis of nephropathy and ESRD than patients with other genotypes (10.45 +/- 9.32 vs 19.5 +/- 8.4 years; P < 0.034). In conclusion, the ACE gene that controls RAS response may influence the development and progression of nephropathy to ESRD. Patients who develop several types of nephropathy have a higher risk of severe evolution if they have a profile of ACE-DD genotype.