RESUMEN
BACKGROUND: The function of polymorphonuclear neutrophils (PMNs) decreases with age, which results in infectious and inflammatory complications in older individuals. The underlying causes are not fully understood. ATP release and autocrine stimulation of purinergic receptors help PMNs combat microbial invaders. Excessive extracellular ATP interferes with these mechanisms and promotes inflammatory PMN responses. Here, we studied whether dysregulated purinergic signaling in PMNs contributes to their dysfunction in older individuals. RESULTS: Bacterial infection of C57BL/6 mice resulted in exaggerated PMN activation that was significantly greater in old mice (64 weeks) than in young animals (10 weeks). In contrast to young animals, old mice were unable to prevent the systemic spread of bacteria, resulting in lethal sepsis and significantly greater mortality in old mice than in their younger counterparts. We found that the ATP levels in the plasma of mice increased with age and that, along with the extracellular accumulation of ATP, the PMNs of old mice became increasingly primed. Stimulation of the formyl peptide receptors of those primed PMNs triggered inflammatory responses that were significantly more pronounced in old mice than in young animals. However, bacterial phagocytosis and killing by PMNs of old mice were significantly lower than that of young mice. These age-dependent PMN dysfunctions correlated with a decrease in the enzymatic activity of plasma ATPases that convert extracellular ATP to adenosine. ATPases depend on divalent metal ions, including Ca2+, Mg2+, and Zn2+, and we found that depletion of these ions blocked the hydrolysis of ATP and the formation of adenosine in human blood, resulting in ATP accumulation and dysregulation of PMN functions equivalent to those observed in response to aging. CONCLUSIONS: Our findings suggest that impaired hydrolysis of plasma ATP dysregulates PMN function in older individuals. We conclude that strategies aimed at restoring plasma ATPase activity may offer novel therapeutic opportunities to reduce immune dysfunction, inflammation, and infectious complications in older patients.
RESUMEN
BACKGROUND: Adenosine triphosphate (ATP) enhances neutrophil responses, but little is known about the role of ATP in influenza infections. METHODS: We used a mouse influenza model to study if ATP release is associated with neutrophil activation and disease progression. RESULTS: Influenza infection increased pulmonary ATP levels 5-fold and plasma ATP levels 3-fold vs healthy mice. Adding ATP at those concentrations to blood from healthy mice primed neutrophils and enhanced CD11b and CD63 expression, CD62L shedding, and reactive oxygen species production in response to formyl peptide receptor stimulation. Influenza infection also primed neutrophils in vivo, resulting in formyl peptide receptor-induced CD11b expression and CD62L shedding up to 3 times higher than that of uninfected mice. In infected mice, large numbers of neutrophils entered the lungs. These cells were significantly more activated than the peripheral neutrophils of infected mice and pulmonary neutrophils of healthy mice. Plasma ATP levels of infected mice and influenza disease progression corresponded with the numbers and activation level of their pulmonary neutrophils. CONCLUSIONS: Findings suggest that ATP release from the lungs of infected mice promotes influenza disease progression by priming peripheral neutrophils, which become strongly activated and cause pulmonary tissue damage after their recruitment to the lungs.
Asunto(s)
Adenosina Trifosfato , Progresión de la Enfermedad , Pulmón , Activación Neutrófila , Neutrófilos , Infecciones por Orthomyxoviridae , Animales , Adenosina Trifosfato/metabolismo , Pulmón/patología , Pulmón/inmunología , Pulmón/metabolismo , Pulmón/virología , Neutrófilos/inmunología , Neutrófilos/metabolismo , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Orthomyxoviridae/virología , Infecciones por Orthomyxoviridae/metabolismo , Ratones , Modelos Animales de Enfermedad , Femenino , Ratones Endogámicos C57BL , Ratones Endogámicos BALB C , Especies Reactivas de Oxígeno/metabolismoRESUMEN
BACKGROUND: Sleep deficiencies, such as manifested in short sleep duration or insomnia symptoms, are known to increase the risk for multiple disease conditions involving immunopathology. Inflammation is hypothesized to be a mechanism through which deficient sleep acts as a risk factor for these conditions. Thus, one potential way to mitigate negative health consequences associated with deficient sleep is to target inflammation. Few interventional sleep studies investigated whether improving sleep affects inflammatory processes, but results suggest that complementary approaches may be necessary to target inflammation associated with sleep deficiencies. We investigated whether targeting inflammation through low-dose acetylsalicylic acid (ASA, i.e., aspirin) is able to blunt the inflammatory response to experimental sleep restriction. METHODS: 46 healthy participants (19F/27M, age range 19-63 years) were studied in a double-blind randomized placebo-controlled crossover trial with three protocols each consisting of a 14-day at-home monitoring phase followed by an 11-day (10-night) in-laboratory stay (sleep restriction/ASA, sleep restriction/placebo, control sleep/placebo). In the sleep restriction/ASA condition, participants took low-dose ASA (81 mg/day) daily in the evening (22:00) during the at-home phase and the subsequent in-laboratory stay. In the sleep restriction/placebo and control sleep/placebo conditions, participants took placebo daily. Each in-laboratory stay started with 2 nights with a sleep opportunity of 8 h/night (23:00-07:00) for adaptation and baseline measurements. Under the two sleep restriction conditions, participants were exposed to 5 nights of sleep restricted to a sleep opportunity of 4 h/night (03:00-07:00) followed by 3 nights of recovery sleep with a sleep opportunity of 8 h/night. Under the control sleep condition, participants had a sleep opportunity of 8 h/night throughout the in-laboratory stay. During each in-laboratory stay, participants had 3 days of intensive monitoring (at baseline, 5th day of sleep restriction/control sleep, and 2nd day of recovery sleep). Variables, including pro-inflammatory immune cell function, C-reactive protein (CRP), and actigraphy-estimated measures of sleep, were analyzed using generalized linear mixed models. RESULTS: Low-dose ASA administration reduced the interleukin (IL)-6 expression in LPS-stimulated monocytes (p<0.05 for condition*day) and reduced serum CRP levels (p<0.01 for condition) after 5 nights of sleep restriction compared to placebo administration in the sleep restriction condition. Low-dose ASA also reduced the amount of cyclooxygenase (COX)-1/COX-2 double positive cells among LPS-stimulated monocytes after 2 nights of recovery sleep following 5 nights of sleep restriction compared to placebo (p<0.05 for condition). Low-dose ASA further decreased wake after sleep onset (WASO) and increased sleep efficiency (SE) during the first 2 nights of recovery sleep (p<0.001 for condition and condition*day). Baseline comparisons revealed no differences between conditions for all of the investigated variables (p>0.05 for condition). CONCLUSION: This study shows that inflammatory responses to sleep restriction can be reduced by preemptive administration of low-dose ASA. This finding may open new therapeutic approaches to prevent or control inflammation and its consequences in those experiencing sleep deficiencies. TRIAL REGISTRATION: ClinicalTrials.gov NCT03377543.
RESUMEN
BACKGROUND: Sinonasal mucosal melanoma (SNMM) is a rare but aggressive tumor with a poor prognosis. The co-inhibitory receptors T cell immunoglobulin and mucinodomain containing-3 (TIM-3), lymphocyte activation gene-3 (LAG-3) and T cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain (TIGIT) are promising new targets in anti-cancer immunotherapy. The expression profiles of these immune checkpoint molecules (ICMs) and potential prognostic implications have not been characterized in SNMM yet. METHODS: Immunohistochemical staining for TIGIT, LAG-3 and TIM-3 was performed on tumor tissue samples from 27 patients with primary SNMM. Associations between ICM expression and demographic parameters, AJCC tumor stage, overall survival, and recurrence-free survival were retrospectively analyzed. RESULTS: SNMM patients with low numbers of TIGIT+ and TIM-3+ tumor infiltrating lymphocytes (TILs) in the primary tumor survived significantly longer than patients with a high degree of TIGIT+ and TIM-3+ TILs. High infiltration with TIM-3+ or TIGIT+ lymphocytes was associated with the higher T4 stage and decreased 5-year survival. CONCLUSION: We identified high densities of TIM-3+ and TIGIT+ TILs as strong negative prognostic biomarkers in SNMM. This suggests that TIM-3 and TIGIT contribute to immunosuppression in SNMM and provides a rationale for novel treatment strategies based on this next generation of immune checkpoint inhibitors. Prospective studies with larger case numbers are warranted to confirm our findings and their implications for immunotherapy.
Asunto(s)
Receptor 2 Celular del Virus de la Hepatitis A , Linfocitos Infiltrantes de Tumor , Melanoma , Receptores Inmunológicos , Humanos , Receptor 2 Celular del Virus de la Hepatitis A/metabolismo , Masculino , Receptores Inmunológicos/metabolismo , Receptores Inmunológicos/análisis , Linfocitos Infiltrantes de Tumor/inmunología , Linfocitos Infiltrantes de Tumor/metabolismo , Femenino , Persona de Mediana Edad , Melanoma/patología , Melanoma/inmunología , Melanoma/mortalidad , Melanoma/metabolismo , Anciano , Pronóstico , Adulto , Biomarcadores de Tumor/análisis , Biomarcadores de Tumor/metabolismo , Estudios Retrospectivos , Anciano de 80 o más Años , Neoplasias de los Senos Paranasales/patología , Neoplasias de los Senos Paranasales/inmunología , Neoplasias de los Senos Paranasales/metabolismo , Neoplasias de los Senos Paranasales/mortalidad , Mucosa Nasal/patología , Mucosa Nasal/inmunología , Mucosa Nasal/metabolismoRESUMEN
BACKGROUND: ATP enhances neutrophil responses, but little is known about the role of ATP in influenza infections. METHODS: We used a mouse influenza model to study if ATP release is associated with neutrophil activation and disease progression. RESULTS: Influenza infection increased pulmonary ATP levels 5-fold and plasma ATP levels 3-fold over the levels in healthy mice. Adding ATP at those concentrations to blood from healthy mice primed their neutrophils and enhanced CD11b and CD63 expression, CD62L shedding, and reactive oxygen species production in response to formyl peptide receptor (FPR) stimulation. Influenza infection also primed neutrophils in vivo, resulting in FPR-induced CD11b expression and CD62L shedding up to 3-times higher than that of uninfected mice. In infected mice, large numbers of neutrophils entered the lungs. These cells were significantly more activated than peripheral neutrophils of infected and pulmonary neutrophils of healthy mice. Plasma ATP levels of infected mice and influenza disease progression corresponded with the numbers and activation level of their pulmonary neutrophils. CONCLUSION: Our findings suggest that ATP release from the lungs of infected mice promotes influenza disease progression by priming peripheral neutrophils that become strongly activated and cause pulmonary tissue damage after their recruitment to the lungs.
RESUMEN
Bladder cancer is amongst the most common causes of cancer death worldwide. Muscle-invasive bladder cancer (MIBC) bears a particularly poor prognosis. Overexpression of purinergic P2X receptors (P2XRs) has been associated with worse outcome in several malignant tumors. Here, we investigated the role of P2XRs in bladder cancer cell proliferation in vitro and the prognostic value of P2XR expression in MIBC patients. Cell culture experiments with T24, RT4, and non-transformed TRT-HU-1 cells revealed a link between high ATP concentrations in the cell culture supernatants of bladder cell lines and a higher grade of malignancy. Furthermore, proliferation of highly malignant T24 bladder cancer cells depended on autocrine signaling through P2X receptors. P2X1R, P2X4R, and P2X7R expression was immunohistochemically analyzed in tumor specimens from 173 patients with MIBC. High P2X1R expression was associated with pathological parameters of disease progression and reduced survival time. High combined expression of P2X1R and P2X7R increased the risk of distant metastasis and was an independent negative predictor of overall and tumor-specific survival in multivariate analyses. Our results suggest that P2X1R/P2X7R expression scores are powerful negative prognostic markers in MIBC patients and that P2XR-mediated pathways are potential targets for novel therapeutic strategies in bladder cancer.
RESUMEN
Neutrophils (PMNs) require extracellular ATP and adenosine (ADO) to fight bacterial infections, which often have life-threatening consequences in pediatric patients. We wondered whether the ATP and ADO levels in the plasma of children change with age and if these changes influence the antimicrobial efficacy of the PMNs of these children. We measured plasma concentrations of ATP and ADO and the activities of the enzymes responsible for the breakdown of these mediators in plasma samples from healthy children and adolescents (n = 45) ranging in age from 0.2 to 15 years. In addition, using blood samples of these individuals, we compared how effective their PMNs were in the phagocytosis of bacteria. In an experimental sepsis model with young (10 days) and adolescent mice (10 weeks), we studied how age influenced the resilience of these animals to bacterial infections and whether addition of ATP could improve the antimicrobial capacity of their PMNs. We found that plasma ATP levels correlated with age and were significantly lower in infants (< 1 year) than in adolescents (12-15 years). In addition, we observed significantly higher plasma ATPase and adenosine deaminase activities in children (< 12 years) when compared to the adolescent population. The activities of these ATP and ADO breakdown processes correlated inversely with age and with the ability of PMNs to phagocytize bacteria. Similar to their human counterparts, young mice also had significantly lower plasma ATP levels when compared to adolescent animals. In addition, we found that mortality of young mice after bacterial infection was significantly higher than that of adolescent mice. Moreover, bacterial phagocytosis by PMNs of young mice was weaker when compared to that of older mice. Finally, we found that ATP supplementation could recover bacterial phagocytosis of young mice to levels similar to those of adolescent mice. Our findings suggest that rapid ATP hydrolysis in the plasma of young children lowers the antimicrobial functions of their PMNs and that this may contribute to the vulnerability of pediatric patients to bacterial infections.
Asunto(s)
Antiinfecciosos , Infecciones Bacterianas , Adolescente , Humanos , Ratones , Niño , Animales , Preescolar , Lactante , Neutrófilos/metabolismo , Adenosina/metabolismo , Adenosina Trifosfato/metabolismo , Infecciones Bacterianas/metabolismo , Antiinfecciosos/metabolismo , FagocitosisRESUMEN
BACKGROUND: Tumor-infiltrating lymphocytes (TILs) are important prognostic biomarkers in several types of cancers. The interplay between TIL subgroups and immune checkpoint molecules like programmed cell death ligand 1 (PD-L1) is a promising target for immunotherapy. However, the TIL landscape in sinonasal mucosal melanoma (SNMM) has not been sufficiently characterized yet and the prognostic value of TIL subgroups and PD-L1 expression remains uncertain. Here, we investigated subsets of TILs (CD3+, CD4+, CD8+, CD20+) and PD-L1 expression patterns in SNMM and assessed their prognostic value for recurrence-free and overall survival. METHODS: Immunohistochemical staining for CD3, CD4, CD8, CD20 and PD-L1 was performed on tumor tissue from 27 patients with primary SNMM. Patient history was obtained and associations between TIL subgroups or PD-L1 expression and AJCC tumor stage, overall survival, and recurrence-free survival were retrospectively analyzed. RESULTS: Patients with high CD3+ and CD8+ TILs in the primary tumor survived significantly longer than patients with SNMMs with a low number of CD3+ and CD8+ TILs. High CD3+ and high CD8+ TILs were associated with the lower T3 stage and increased 5-year survival. PD-L1 positivity in tumor cells was associated with advanced tumor stage. CONCLUSION: Our results indicate that high densities of CD3+ and CD8+ TILs are strong positive prognostic biomarkers for survival in SNMM. Prospective studies with larger case numbers are warranted to confirm our findings.
Asunto(s)
Melanoma , Neoplasias de los Senos Paranasales , Humanos , Estudios Prospectivos , Estudios Retrospectivos , Linfocitos Infiltrantes de Tumor , Antígeno B7-H1 , Pronóstico , Melanoma/patología , Neoplasias de los Senos Paranasales/patología , Biomarcadores/metabolismoRESUMEN
Polymorphonuclear neutrophils (PMNs) protect the host from invading microorganisms. However, excessively activated PMNs can also cause damage to host tissues under inflammatory conditions. Here we developed simple assays to determine the activation state of PMNs in human whole blood that contains soluble mediators known to influence PMN functions. Because mouse models are widely used to study the role of PMNs in infectious and inflammatory diseases, we adapted these assays for the rapid and reliable assessment of PMN functions in murine blood samples. Freshly collected whole blood samples were stimulated with agonists of the formyl peptide receptors (FPR) of PMNs and changes in reactive oxygen species (ROS) production and the expression of CD11b, CD62L (L-selectin), CD66b, and CD63 on the cell surface were analyzed with flow cytometry. We optimized these assays to minimize inadvertent interferences such as cell stress generated during sample handling and the loss of plasma mediators that regulate PMN functions. Human PMNs readily responded to the FPR agonist N-formyl-methionyl-leucyl-phenylalanine (fMLP). The most sensitive responses of human PMNs to fMLP were CD11b, CD62L, and CD66b expression with half maximal effective concentrations (EC50) of 5, 8, and 6 nM fMLP, respectively. CD63 expression and ROS production required markedly higher fMLP concentrations with EC50 values of 19 and 50 nM fMLP, respectively. Mouse PMNs did not respond well to fMLP and required significantly higher concentrations of the FPR agonist WKYMVm (W-peptide) to achieve equivalent cell activation. The most sensitive response of mouse PMNs was ROS production with an EC50 of 38 nM W-peptide. Because mice do not express CD66b, we only assessed the expression of CD62L, CD11b, and CD63 with EC50 values of 54, 119, and 355 nM W-peptide, respectively. Validation of our optimized assays showed that they sensitively detect the responses of human PMNs to priming with endotoxin in vitro as well as the corresponding responses of murine PMNs to bacterial infection in a sepsis model. We conclude that these optimized assays could be useful tools for the monitoring of patients with infections, sepsis, and other inflammatory conditions as well as for the design and interpretation of preclinical studies of these diseases in mouse models.
Asunto(s)
Activación Neutrófila , Sepsis , Humanos , Animales , Ratones , Especies Reactivas de Oxígeno/metabolismo , Activación Neutrófila/fisiología , Citometría de Flujo , Neutrófilos/metabolismo , Receptores de Formil Péptido/metabolismo , N-Formilmetionina Leucil-Fenilalanina/farmacologíaRESUMEN
RATIONALE: The increased mortality and morbidity seen in critically injured patients appears associated with systemic inflammatory response syndrome (SIRS) and immune dysfunction, which ultimately predisposes to infection. Mitochondria released by injury could generate danger molecules, for example, ATP, which in turn would be rapidly scavenged by ectonucleotidases, expressed on regulatory immune cells. OBJECTIVE: To determine the association between circulating mitochondria, purinergic signalling and immune dysfunction after trauma. METHODS: We tested the impact of hepatocyte-derived free mitochondria on blood-derived and lung-derived CD8 T cells in vitro and in experimental mouse models in vivo. In parallel, immune phenotypic analyses were conducted on blood-derived CD8 T cells obtained from trauma patients. RESULTS: Isolated intact mitochondria are functional and generate ATP ex vivo. Extracellular mitochondria perturb CD8+ T cells in co-culture, inducing select features of immune exhaustion in vitro. These effects are modulated by scavenging ATP, modelled by addition of apyrase in vitro. Injection of intact mitochondria into recipient mice markedly upregulates the ectonucleotidase CD39, and other immune checkpoint markers in circulating CD8+ T cells. We note that mice injected with mitochondria, prior to instilling bacteria into the lung, exhibit more severe lung injury, characterised by elevated neutrophil influx and by changes in CD8+ T cell cytotoxic capacity. Importantly, the development of SIRS in injured humans, is likewise associated with disordered purinergic signalling and CD8 T cell dysfunction. CONCLUSION: These studies in experimental models and in a cohort of trauma patients reveal important associations between extracellular mitochondria, aberrant purinergic signalling and immune dysfunction. These pathogenic factors with immune exhaustion are linked to SIRS and could be targeted therapeutically.
Asunto(s)
Antígenos CD , Linfocitos T CD8-positivos , Animales , Humanos , Ratones , Adenosina Trifosfato/metabolismo , Biomarcadores/metabolismo , Linfocitos T CD8-positivos/metabolismo , Mitocondrias , Síndrome de Respuesta Inflamatoria Sistémica/metabolismoRESUMEN
Tumor-infiltrating lymphocytes (TILs) are associated with improved survival in several types of cancers, including genitourinary cancers. However, multiple different scoring methods used to assess TILs complicate the comparison of different studies and are not always suitable for daily practice. In 2014, the International TILs Working Group (ITWG) proposed a simple and robust assessment method for a more standardized evaluation of TILs. Here, we validated this system in muscle-invasive urinary bladder cancer (MIBC). Patient history and histologic specimens from 203 patients with MIBC were retrospectively analyzed. The stromal TIL (sTIL) score was determined using the ITWG system and 3 groups were defined according to the degree of stromal lymphocytic infiltration: low (0-10%), intermediate (10-55%) and high (55-100%). Associations between sTIL score, clinicopathological variables, tumor-specific survival (TSS), overall survival (OS), and disease-free survival (DFS) were analyzed. High stromal lymphocytic infiltration was associated with significantly higher OS, TSS and DFS when compared to low grade sTILs. The survival benefit remained statistically significant in multivariate analyses, confirming that sTILs are a strong independent positive prognostic factor in patients with MIBC. In summary, the degree of sTILs as defined by the ITWG robustly predicts survival in MIBC patients. Prospective studies with larger case numbers are needed to determine whether sTILs should be included in staging guidelines and how they could aid in therapeutic decision making.
Asunto(s)
Carcinoma de Células Transicionales , Neoplasias de la Vejiga Urinaria , Humanos , Linfocitos Infiltrantes de Tumor/patología , Neoplasias de la Vejiga Urinaria/patología , Pronóstico , Estudios Retrospectivos , Estudios Prospectivos , Carcinoma de Células Transicionales/patologíaRESUMEN
ATP released into the bloodstream regulates immune responses and other physiological functions. Excessive accumulation of extracellular ATP interferes with these functions, and elevated plasma ATP levels could indicate infections and other pathological disorders. However, there is considerable disagreement about what constitutes normal plasma ATP levels. Therefore, we optimized a method to accurately assess ATP concentrations in blood. We found that rapid chilling of heparinized blood samples is essential to preserve in vivo ATP levels and that differential centrifugation minimizes inadvertent ATP release due to cell damage and mechanical stress. Plasma samples were stabilized with perchloric acid, etheno-derivatized, and delipidated for sensitive analysis of ATP and related compounds using high-performance liquid chromatography (HPLC) and fluorescence detection. We measured 33 ± 20 nM ATP, 90 ± 45 nM ADP, 100 ± 55 nM AMP, and 81 ± 51 nM adenosine in the blood of healthy human adults (n = 10). In critically ill patients, ATP levels were 6 times higher than in healthy subjects. The anticoagulant greatly affected results. ATP levels were nearly 8 times higher in EDTA plasma than in heparin plasma, while AMP levels were 3 times lower and adenosine was entirely absent in EDTA plasma. If EDTA blood was not immediately chilled, ATP, ADP, and AMP levels continued to rise, which indicates that EDTA interferes with the endogenous mechanisms that regulate plasma adenylate levels. Our optimized method eliminates artifacts that prevent accurate determination of plasma adenylates and will be useful for studying mechanisms that regulate adenylate levels and for monitoring of pathological processes in patients with infections and other diseases.
Asunto(s)
Adenosina Trifosfato , Adenosina , Adenosina Difosfato , Adenosina Monofosfato , Adenosina Trifosfato/análisis , Cromatografía Líquida de Alta Presión , Ácido Edético , HumanosRESUMEN
PURPOSE: Dysplasia and cancer of the upper aerodigestive tract are characterized by significant neoangiogenesis. This can be recognized by optical methods like the Storz Professional Image Enhancement System (SPIES). Up to now, there are no reports of using this novel technique for examining nasal diseases. The objective of this study was to evaluate the use of SPIES during sinus surgery to help differentiate various nasal pathologies and determine their extension. METHODS: Patients (n = 27) with different pathologies in the region of the paranasal sinuses were operated via functional endoscopic surgery using a 2D-HD-camera with white light and SPIES. In addition, 10 healthy individuals were examined. The system was evaluated using two different questionnaires. RESULTS: The handling and operation of SPIES was intuitive and easy. Use of SPIES did not prolong the procedure. There was no disturbing image distortion. SPIES seemed to improve the visualization, differentiation and evaluation of vascularization of paranasal pathologies and allowed for precise and accurate surgery. Compared to examination with the 2D-HD-camera and white light alone, SPIES appeared to facilitate the identification of mucosal pathologies. CONCLUSION: SPIES could be a promising adjunct tool to evaluate nasal pathologies intraoperatively. Especially in the case of vascularized tumors the enhanced image endoscopy seemed to be clearly superior to standard white light alone. In our study, the system facilitated the assessment of tumor extension and vascularization as well as the differentiation of healthy mucosa. Future randomized studies will be necessary to prove the potential of integrating this novel technique into the clinical routine for the differentiation of nasal pathologies and the improvement of resection margins during nasal tumor surgery.
Asunto(s)
Neoplasias Nasales , Enfermedades de los Senos Paranasales , Senos Paranasales , Endoscopía/métodos , Humanos , Aumento de la Imagen , Neoplasias Nasales/cirugía , Enfermedades de los Senos Paranasales/diagnóstico por imagen , Enfermedades de los Senos Paranasales/cirugía , Senos Paranasales/diagnóstico por imagen , Senos Paranasales/cirugíaRESUMEN
OBJECTIVES/HYPOTHESIS: Tumor-infiltrating lymphocytes (TILs) predict better outcome in several types of cancers. However, the prognostic value of TILs in sinonasal mucosal melanoma (SNMM) is uncertain. Here, we investigated whether TILs can be used as a prognostic indicator for survival in SNMM. STUDY DESIGN: Retrospective cohort study. METHODS: Patient history and histologic specimens from 27 patients with primary SNMM were retrospectively analyzed. TIL grade was determined and associations between TILs and AJCC tumor stage, overall survival, and recurrence-free survival were analyzed. RESULTS: Patients with TILs in the primary tumor classified as brisk or non-brisk survived significantly longer than patients with SNMMs lacking lymphocyte infiltrates. Brisk TILs were associated with the lower T3 stage and increased recurrence-free and 5-year survival. CONCLUSION: Our results indicate that TIL density is a strong prognostic factor for better survival in SNMM. Prospective studies with larger case numbers are warranted to determine whether TILs should be included in future AJCC staging guidelines. LEVEL OF EVIDENCE: 3 Laryngoscope, 132:1334-1339, 2022.
Asunto(s)
Melanoma , Neoplasias de los Senos Paranasales , Neoplasias Cutáneas , Humanos , Linfocitos Infiltrantes de Tumor/patología , Neoplasias de los Senos Paranasales/patología , Pronóstico , Estudios Prospectivos , Estudios Retrospectivos , Neoplasias Cutáneas/patologíaRESUMEN
Reported success rates of endoscopic choanal atresia (CA) surgery vary substantially due to a high heterogeneity in and between study groups. Comprehensive data on the unique patient cohort of newborns with bilateral CA are scarce. Our study aimed to close this gap by using narrow inclusion criteria and standardized surgical outcome parameters. A total of ten neonates who were diagnosed with bilateral complete CA and underwent endoscopic surgery at the Department of Otolaryngology, Head and Neck Surgery in the University Hospital of Munich between 2008 and 2017 were included. Preoperative findings, surgical procedures, outcome, and follow-up were analyzed. Standardized criteria were used to assess surgical outcome. Almost all patients (90%) required at least one revision procedure within the first 6 months after initial surgery because of symptomatic partial or complete restenosis. After that, all surviving patients remained asymptomatic until the end of the follow-up period.Conclusion: Endoscopic bilateral CA repair in neonates is a safe procedure with a high long-term success rate. However, compared to other patient groups with choanal obstruction, restenosis occurs frequently, and revision procedures are required in a large number of cases. This should be considered during preoperative planning and parent counseling. What is Known: ⢠Bilateral complete choanal atresia (CA) is a neonatal emergency that requires surgical intervention. ⢠Reported success rates of endoscopic choanal obstruction repair are highly variable and mostly derived from heterogenous study groups that do not reflect the situation in neonates adequately. What is New: ⢠This study focuses exclusively on newborns with complete bilateral CA who underwent endoscopic surgery within the first 28 days of life and uses standardized criteria to assess outcome. ⢠The long-term success rate of endoscopic bilateral CA repair in neonates is high; however, almost all patients require at least one revision procedure within the first 6 months.
Asunto(s)
Atresia de las Coanas , Atresia de las Coanas/cirugía , Endoscopía , Humanos , Recién Nacido , Nariz , Resultado del TratamientoRESUMEN
Extracellular diphosphate and triphosphate nucleotides are released from activated or injured cells to trigger vascular and immune P2 purinergic receptors, provoking inflammation and vascular thrombosis. These metabokines are scavenged by ectonucleoside triphosphate diphosphohydrolase-1 (E-NTPDase1 or CD39). Further degradation of the monophosphate nucleoside end products occurs by surface ecto-5'-nucleotidase (NMPase) or CD73. These ectoenzymatic processes work in tandem to promote adenosinergic responses, which are immunosuppressive and antithrombotic. These homeostatic ectoenzymatic mechanisms are lost in the setting of oxidative stress, which exacerbates inflammatory processes. We have engineered bifunctional enzymes made up from ectodomains (ECDs) of CD39 and CD73 within a single polypeptide. Human alkaline phosphatase-ectodomain (ALP-ECD) and human acid phosphatase-ectodomain (HAP-ECD) fusion proteins were also generated, characterized, and compared with these CD39-ECD, CD73-ECD, and bifunctional fusion proteins. Through the application of colorimetrical functional assays and high-performance liquid chromatography kinetic assays, we demonstrate that the bifunctional ectoenzymes express high levels of CD39-like NTPDase activity and CD73-like NMPase activity. Chimeric CD39-CD73-ECD proteins were superior in converting triphosphate and diphosphate nucleotides into nucleosides when compared with ALP-ECD and HAP-ECD. We also note a pH sensitivity difference between the bifunctional fusion proteins and parental fusions, as well as ectoenzymatic property distinctions. Intriguingly, these innovative reagents decreased platelet activation to exogenous agonists in vitro. We propose that these chimeric fusion proteins could serve as therapeutic agents in inflammatory diseases, acting to scavenge proinflammatory ATP and also generate anti-inflammatory adenosine.
Asunto(s)
5'-Nucleotidasa/farmacología , Antiinflamatorios/farmacología , Apirasa/farmacología , Inhibidores de Agregación Plaquetaria/farmacología , Agregación Plaquetaria/efectos de los fármacos , Ingeniería de Proteínas , 5'-Nucleotidasa/química , 5'-Nucleotidasa/genética , 5'-Nucleotidasa/metabolismo , Nucleótidos de Adenina/metabolismo , Antiinflamatorios/química , Antiinflamatorios/metabolismo , Apirasa/química , Apirasa/genética , Apirasa/metabolismo , Proteínas Ligadas a GPI/química , Proteínas Ligadas a GPI/genética , Proteínas Ligadas a GPI/metabolismo , Proteínas Ligadas a GPI/farmacología , Células HEK293 , Humanos , Concentración de Iones de Hidrógeno , Hidrólisis , Cinética , Inhibidores de Agregación Plaquetaria/química , Inhibidores de Agregación Plaquetaria/metabolismo , Conformación Proteica , Proteínas Recombinantes de Fusión/metabolismo , Transducción de Señal , Relación Estructura-Actividad , Especificidad por SustratoRESUMEN
T cells form an immune synapse (IS) with antigen-presenting cells (APCs) to detect antigens that match their TCR. Mitochondria, pannexin-1 (panx1) channels, and P2X4 receptors congregate at the IS where mitochondria produce the ATP that panx1 channels release in order to stimulate P2X4 receptors. P2X4 receptor stimulation causes cellular Ca2+ influx that up-regulates mitochondrial metabolism and localized ATP production at the IS. Here we show that P2Y11 receptors are essential players that sustain these T cell activation mechanisms. We found that P2Y11 receptors retract from the IS toward the back of cells where their stimulation by extracellular ATP induces cAMP/PKA signaling that redirects mitochondrial trafficking to the IS. P2Y11 receptors thus reinforce IS signaling by promoting the aggregation of mitochondria with panx1 ATP release channels and P2X4 receptors at the IS. This dual purinergic signaling mechanism involving P2X4 and P2Y11 receptors focuses mitochondrial metabolism to the IS where localized ATP production sustains synaptic activity in order to allow successful completion of T cell activation responses. Our findings have practical implications because rodents lack P2Y11 receptors, raising concerns as to the validity of rodent models to study treatment of infections and inflammatory conditions.
Asunto(s)
Sinapsis Inmunológicas/metabolismo , Activación de Linfocitos/inmunología , Mitocondrias/metabolismo , Receptores Purinérgicos P2/metabolismo , Linfocitos T/inmunología , Comunicación Autocrina , Linfocitos T CD4-Positivos/inmunología , Señalización del Calcio , AMP Cíclico/metabolismo , Humanos , Células Jurkat , Microtúbulos/metabolismo , Receptores Purinérgicos P2X4 , Transducción de Señal , Células U937RESUMEN
Intracellular ATP is the universal energy carrier that fuels many cellular processes. However, immune cells can also release a portion of their ATP into the extracellular space. There, ATP activates purinergic receptors that mediate autocrine and paracrine signaling events needed for the initiation, modulation, and termination of cell functions. Mitochondria contribute to these processes by producing ATP that is released. Here, we summarize the synergistic interplay between mitochondria and purinergic signaling that regulates T cell functions. Specifically, we discuss how mitochondria interact with P2X1, P2X4, and P2Y11 receptors to regulate T cell metabolism, cell migration, and antigen recognition. These mitochondrial and purinergic signaling mechanisms are indispensable for host immune defense. However, they also represent an Achilles heel that can render the host susceptible to infections and inflammatory disorders. Hypoxia and mitochondrial dysfunction deflate the purinergic signaling mechanisms that regulate T cells, while inflammation and tissue damage generate excessive systemic ATP levels that distort autocrine purinergic signaling and impair T cell function. An improved understanding of the metabolic and purinergic signaling mechanisms that regulate T cells may lead to novel strategies for the diagnosis and treatment of infectious and inflammatory diseases.
Asunto(s)
Inmunomodulación , Mitocondrias/metabolismo , Receptores Purinérgicos P2X/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Comunicación Celular/inmunología , Ciclo Celular/genética , Ciclo Celular/inmunología , Movimiento Celular/genética , Movimiento Celular/inmunología , Metabolismo Energético/genética , Metabolismo Energético/inmunología , Humanos , Sinapsis Inmunológicas/inmunología , Sinapsis Inmunológicas/metabolismo , Mitocondrias/genética , Transducción de SeñalRESUMEN
Microglia, the brain's resident macrophages, help to regulate brain function by removing dying neurons, pruning non-functional synapses, and producing ligands that support neuronal survival1. Here we show that microglia are also critical modulators of neuronal activity and associated behavioural responses in mice. Microglia respond to neuronal activation by suppressing neuronal activity, and ablation of microglia amplifies and synchronizes the activity of neurons, leading to seizures. Suppression of neuronal activation by microglia occurs in a highly region-specific fashion and depends on the ability of microglia to sense and catabolize extracellular ATP, which is released upon neuronal activation by neurons and astrocytes. ATP triggers the recruitment of microglial protrusions and is converted by the microglial ATP/ADP hydrolysing ectoenzyme CD39 into AMP; AMP is then converted into adenosine by CD73, which is expressed on microglia as well as other brain cells. Microglial sensing of ATP, the ensuing microglia-dependent production of adenosine, and the adenosine-mediated suppression of neuronal responses via the adenosine receptor A1R are essential for the regulation of neuronal activity and animal behaviour. Our findings suggest that this microglia-driven negative feedback mechanism operates similarly to inhibitory neurons and is essential for protecting the brain from excessive activation in health and disease.