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The fungus Candida albicans can cause mucosal infections including oropharyngeal candidiasis (OPC) in immunocompromised patients. In humans, an increased risk of fungal infections correlates with thrombocytopenia. However, our understanding of platelets and megakaryocytes (Mks) in mucosal fungal infections is almost entirely unknown. When megakaryocyte- and platelet-depleted mice were infected with OPC, the tongue showed higher fungal burden, due to decreased neutrophil accumulation. Protection depended on a distinct population of oral-resident Mks. Interleukin-17, important in antifungal immunity, was required since mice lacking the IL-17 receptor had decreased circulating platelets and their oral Mks did not expand during OPC. The secretion of the peptide toxin candidalysin activated human Mks to release platelets with antifungal capacity. Infection with a candidalysin-deficient strain resulted in decreased expansion of tongue Mks during OPC. This is the first time that a distinct megakaryocyte population was identified in the oral mucosa which is critical for immunity against fungal infection.
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Candidiasis Bucal , Enfermedades Transmisibles , Proteínas Fúngicas , Micosis , Humanos , Ratones , Animales , Candida albicans , Megacariocitos , Interleucina-17 , Antifúngicos , Candidiasis Bucal/microbiología , Mucosa BucalRESUMEN
Platelets are known for essential activities in hemostasis and for their important contribution to protection against infectious pathogens. Klebsiella pneumoniae is an opportunistic pathogen widely known to cause nosocomial infections. Recently, hypervirulent strains of K. pneumoniae have been emerging, which can cause severe infections in immunocompetent individuals. Combined with the increase in antibiotic resistance, it is important to understand how K. pneumoniae affects components of the immune system. We studied the interactions of human platelets with several K. pneumoniae strains (the wild type encapsulated strain, and a nonencapsulated mutant). Thrombin-stimulated whole human and mouse blood significantly inhibited bacterial growth compared to unstimulated whole blood. Furthermore, we investigated the effect of K. pneumoniae on platelet activation. Both strains induced significant increase in activation of both unstimulated and thrombin-stimulated human platelets. Additionally, only the nonencapsulated mutant increased aggregation of platelets in response to ADP. K. pneumoniae killing assays were then performed with washed platelets in the presence or absence of thrombin. Surprisingly, washed platelets failed to exhibit any effects on the growth of K. pneumoniae. We further explored the impact of platelets on monocyte-mediated killing of K. pneumoniae. Importantly, we found that activated platelets significantly enhanced monocyte-mediated killing of K. pneumoniae. This effect was likely due to the formation of platelet-monocyte aggregates in blood upon thrombin stimulation. Overall, this study highlights the role of platelets in mediating a protective response against K. pneumoniae and reinforces the importance of platelets in modulating leukocyte behavior.
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Plaquetas , Infecciones por Klebsiella , Animales , Ratones , Humanos , Klebsiella pneumoniae , Monocitos , Trombina/farmacología , Activación Plaquetaria , Infecciones por Klebsiella/microbiología , AntibacterianosRESUMEN
The paracrine and autocrine processes of the host response play an integral role in the success of scaffold-based tissue regeneration. Recently, the immunomodulatory scaffolds have received huge attention for modulating inflammation around the host tissue through releasing anti-inflammatory cytokine. However, controlling the inflammation and providing a sustained release of anti-inflammatory cytokine from the scaffold in the digestive inflammatory environment are predicated upon a comprehensive understanding of three fundamental questions. (1) How does the release rate of cytokine from the scaffold change in the digestive inflammatory environment? (2) Can we prevent the premature scaffold degradation and burst release of the loaded cytokine in the digestive inflammatory environment? (3) How does the scaffold degradation prevention technique affect the immunomodulatory capacity of the scaffold? This study investigated the impacts of the digestive inflammatory environment on scaffold degradation and how pre-mature degradation can be prevented using genipin crosslinking and how genipin crosslinking affects the interleukin-4 (IL-4) release from the scaffold and differentiation of naïve macrophages (M0). Our results demonstrated that the digestive inflammatory environment (DIE) attenuates protein retention within the scaffold. Over 14 days, the encapsulated protein released 46% more in DIE than in phosphate buffer saline (PBS), which was improved through genipin crosslinking. We have identified the 0.5 (w/v) genipin concentration as an optimal concentration for improved IL-4 released from the scaffold, cell viability, mechanical strength, and scaffold porosity, and immunomodulation studies. The IL-4 released from the injectable scaffold could differentiate naïve macrophages to an anti-inflammatory (M2) lineage; however, upon genipin crosslinking, the immunomodulatory capacity of the scaffold diminished significantly, and pro-inflammatory markers were expressed dominantly.
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Regeneración Tisular Dirigida/métodos , Inmunomodulación , Iridoides/farmacología , Macrófagos/efectos de los fármacos , Andamios del Tejido/química , Animales , Diferenciación Celular , Células Cultivadas , Colágeno , Inflamación/tratamiento farmacológico , Inflamación/prevención & control , Inyecciones , Interleucina-4/inmunología , Interleucina-4/metabolismo , Iridoides/uso terapéutico , Macrófagos/inmunología , Macrófagos/metabolismo , Macrófagos/patología , Ratones , Sistema Musculoesquelético/inmunología , PorosidadRESUMEN
Platelets have long been recognized for their role in maintaining the balance between hemostasis and thrombosis. While their contributions to blood clotting have been well established, it has been increasingly evident that their roles extend to both innate and adaptive immune functions during infection and inflammation. In this comprehensive review, we describe the various ways in which platelets interact with different microbes and elicit immune responses either directly, or through modulation of leukocyte behaviors.
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Platelet-leukocyte aggregates (PLAs) are associated with increased thrombosis risk. The influence of PLA formation is especially important for cancer patients, since thrombosis accounts for approximately 10% of cancer-associated deaths. Our objective was to characterize and quantify PLAs in whole blood samples from lung cancer patients compared to healthy volunteers with the intent to analyze PLA formation in the context of lung cancer-associated thrombosis. Consenting lung cancer patients (57) and healthy volunteers (56) were enrolled at the Dana Cancer Center at the University of Toledo Health Science Campus. Peripheral blood samples were analyzed by flow cytometry. Patient medical history was reviewed through electronic medical records. Most importantly, we found lung cancer patients to have higher percentages of platelet-T cell aggregates (PTCAs) than healthy volunteers among both CD4+ T lymphocyte and CD8+ T lymphocyte populations. Our findings demonstrate that characterization of PTCAs may have clinical utility in differentiating lung cancer patients from healthy volunteers and stratifying lung cancer patients by history of thrombosis.
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Plaquetas/patología , Neoplasias Pulmonares/sangre , Neoplasias Pulmonares/complicaciones , Linfocitos T/patología , Trombosis/sangre , Trombosis/etiología , Adulto , Anciano , Anciano de 80 o más Años , Linfocitos T CD4-Positivos/patología , Linfocitos T CD8-positivos/patología , Estudios de Casos y Controles , Agregación Celular , Femenino , Voluntarios Sanos , Humanos , Masculino , Persona de Mediana Edad , Factores de Riesgo , Adulto JovenRESUMEN
Candida albicans is a pervasive commensal fungus that is the most common pathogen responsible for invasive fungal infection (IFI). With incidence of IFI on the rise due to increasing susceptible populations, it is imperative that we investigate how Candida albicans interacts with blood components. When stimulating either human or mouse whole blood with thrombin, we saw a significant decrease in C. albicans survival. We then repeated Candida killing assays with thrombin-stimulated or unstimulated washed platelets and saw a similar decrease in CFU. To investigate whether killing was mediated through surface components or releasable products, platelets were pretreated with an inhibitor of actin polymerization (cytochalasin D [CytoD]). CytoD was able to abrogate C. albicans killing. Moreover, dilution of releasates from thrombin-stimulated platelets showed that the toxicity of the releasates on C. albicans is concentration dependent. We then investigated C. albicans actions on platelet activation, granule release, and aggregation. While C. albicans does not appear to affect alpha or dense granule release, C. albicans exerts a significant attenuation of platelet aggregation to multiple agonists. These results illustrate for the first time that platelets can directly kill C. albicans through release of their granular contents. Additionally, C. albicans can also exert inhibitory effects on platelet aggregation.
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Antifúngicos/metabolismo , Plaquetas/metabolismo , Plaquetas/microbiología , Candida albicans/inmunología , Factores Inmunológicos/metabolismo , Animales , Candida albicans/fisiología , Recuento de Colonia Microbiana , Humanos , Ratones , Viabilidad Microbiana/efectos de los fármacosRESUMEN
BACKGROUND: Platelets are widely recognized for their role in maintaining hemostasis. Recently, platelets have become appreciated for their varying roles in immunity, neuroprotection, and other physiological processes. While there are currently excellent methods to transiently deplete platelets and models of thrombocytopenia, studying the roles of platelets in chronic processes can be challenging. OBJECTIVE: Phenotypic characterization of the PF4-DTR mouse model of conditional platelet depletion compared to antibody depletion. METHODS: We describe the ability of the PF4-DTR mouse to maintain chronic platelet depletion, along with examining the bleeding phenotype compared to antibody-mediated platelet depletion. RESULTS: Systemic administration of diphtheria toxin resulted in >99% platelet depletion that can be maintained for >2 weeks. When compared to an antibody depletion model, PF4-DTR mice showed similar phenotypes when challenged with tail bleed and saphenous vein measurements of hemostasis. Mice depleted with diphtheria toxin were also able to undergo adoptive transfer of platelets. If the frequency and amount of diphtheria toxin is reduced, mice can be maintained at >40% depletion for >28 days, showing that this model is tunable. CONCLUSIONS: When compared to the gold standard of antibody-mediated depletion, PF4-DTR mice showed similar phenotypes and should be considered an important tool for examining the impact of thrombocytopenia over longer periods of time.
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This article was corrected to add a permissions credit to Fig 3 legend.
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In recent years, biomaterial- and scaffold-based immunomodulation strategies were implemented in tissue regeneration efforts for manipulating macrophage polarization (a.k.a. phenotype or lineage commitment, or differentiation). Yet, most of our understanding of macrophage phenotype commitment and phagocytic capacity is limited to how physical cues (extracellular matrix stiffness, roughness, and topography) and soluble chemical cues (cytokines and chemokines released from the scaffold) influence macrophage polarization. In the context of immune response-tissue interaction, the mechanical cues experienced by the residing cells within the tissue also play a critical role in macrophage polarization and inflammatory response. However, there is no compiled study discussing the effect of the dynamic mechanical environment around the tissues on macrophage polarization and the innate immune response. The aim of this comprehensive review paper is 2-fold; (a) to highlight the importance of mechanical cues on macrophage lineage commitment and function and (b) to summarize the important studies dedicated to understand how macrophage polarization changes with different mechanical loading modalities. For the first time, this review paper compiles and compartmentalizes the studies investigating the role of dynamic mechanical loading with various modalities, amplitude, and frequency on macrophage differentiation. A deeper understanding of macrophage phenotype in mechanically dominant tissues (i.e. musculoskeletal tissues, lung tissues, and cardiovascular tissues) provides mechanistic insights into the design of mechano-immunomodulatory tissue scaffold for tissue regeneration.
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Polaridad Celular , Inmunomodulación , Macrófagos/citología , Mecanotransducción Celular , Animales , Citocinas/inmunología , Humanos , Inmunidad Innata , Macrófagos/inmunología , Estrés Mecánico , Andamios del TejidoRESUMEN
Staphylococcus aureus is a major human pathogen that can cause mild to severe life-threatening infections in many tissues and organs. Platelets are known to participate in protection against S. aureus by direct killing and by enhancing the activities of neutrophils and macrophages in clearing S. aureus infection. Platelets have also been shown to induce monocyte differentiation into dendritic cells and to enhance activation of dendritic cells. Therefore, in the present study, we explored the role of platelets in enhancing bone marrow-derived dendritic cell (BMDC) function against S. aureus We observed a significant increase in dendritic cell phagocytosis and intracellular killing of a methicillin-resistant Staphylococcus aureus (MRSA) strain (USA300) by thrombin-activated platelets or their releasates. Enhancement of bacterial uptake and killing by DCs is mediated by platelet-derived CD40L. Coculture of USA300 and BMDCs in the presence of thrombin-activated platelet releasates invokes upregulation of the maturation marker CD80 on DCs and enhanced production of the proinflammatory cytokines tumor necrosis factor alpha (TNF-α), interleukin 12 (IL-12), and IL-6. Overall, these observations support our hypothesis that platelets play a critical role in the host defense against S. aureus infection. Platelets stimulate DCs, leading to direct killing of S. aureus and enhanced DC maturation, potentially leading to adaptive immune responses against S. aureus.
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Plaquetas/inmunología , Ligando de CD40/inmunología , Citotoxicidad Inmunológica/fisiología , Células Dendríticas/inmunología , Activación Plaquetaria/inmunología , Infecciones Estafilocócicas/inmunología , Staphylococcus aureus/inmunología , Antígeno B7-1/metabolismo , Citocinas/metabolismo , Humanos , Activación de Macrófagos/fisiología , Macrófagos/inmunología , Staphylococcus aureus Resistente a Meticilina/inmunología , Fagocitosis/inmunologíaRESUMEN
High blood pressure is a common cause of chronic kidney disease. Because CD40, a member of the tumor necrosis factor receptor family, has been linked to the progression of kidney disease in ischemic nephropathy, we studied the role of Cd40 in the development of hypertensive renal disease. The Cd40 gene was mutated in the Dahl S genetically hypertensive rat with renal disease by targeted-gene disruption using zinc-finger nuclease technology. These rats were then given low (0.3%) and high (2%) salt diets and compared. The resultant Cd40 mutants had significantly reduced levels of both urinary protein excretion (41.8 ± 3.1 mg/24 h vs. 103.7 ± 4.3 mg/24 h) and plasma creatinine (0.36 ± 0.05 mg/dl vs. 1.15 ± 0.19 mg/dl), with significantly higher creatinine clearance compared with the control S rats (3.04 ± 0.48 ml/min vs. 0.93 ± 0.15 ml/min), indicating renoprotection was conferred by mutation of the Cd40 locus. Furthermore, the Cd40 mutants had a significant attenuation in renal fibrosis, which persisted on the high salt diet. However, there was no difference in systolic blood pressure between the control and Cd40 mutant rats. Thus, these data serve as the first evidence for a direct link between Cd40 and hypertensive nephropathy. Hence, renal fibrosis is one of the underlying mechanisms by which Cd40 plays a crucial role in the development of hypertensive renal disease.
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Presión Sanguínea/genética , Antígenos CD40/genética , Hipertensión/genética , Enfermedades Renales/prevención & control , Riñón/metabolismo , Mutación , Proteinuria/prevención & control , Animales , Linfocitos B/metabolismo , Antígenos CD40/metabolismo , Movimiento Celular , Creatinina/sangre , Dieta Hiposódica , Modelos Animales de Enfermedad , Fibrosis , Predisposición Genética a la Enfermedad , Hipertensión/metabolismo , Hipertensión/fisiopatología , Riñón/patología , Riñón/fisiopatología , Enfermedades Renales/genética , Enfermedades Renales/metabolismo , Enfermedades Renales/fisiopatología , Activación de Linfocitos , Fenotipo , Fosforilación , Inhibidor 1 de Activador Plasminogénico/metabolismo , Proteinuria/genética , Proteinuria/metabolismo , Proteinuria/fisiopatología , Ratas Endogámicas Dahl , Ratas Mutantes , Eliminación Renal , Cloruro de Sodio Dietético , Linfocitos T/metabolismo , Factores de Tiempo , Familia-src Quinasas/metabolismoRESUMEN
Platelets are the chief effector cells in hemostasis. However, recent evidence suggests they have multiple roles in host defense against infection. Reports by us and others showed that platelets functionally contribute to protection against Staphylococcus aureus infection. In the current study, the capacity of mouse platelets to participate in host defense against S. aureus infection was determined by assessing two possibilities. First, we determined the ability of platelets to kill S. aureus directly; and, second, we tested the possibility that platelets enhance macrophage phagocytosis and intracellular killing of S. aureus In this study we report evidence in support of both mechanisms. Platelets effectively killed two different strains of S. aureus. A clinical isolate of methicillin-resistant S. aureus was killed by platelets (>40% killing in 2 h) in a thrombin-dependent manner whereas a methicillin-sensitive strain was killed to equal extent but did not require thrombin. Interestingly, thrombin-stimulated platelets also significantly enhanced peritoneal macrophage phagocytosis of both methicillin-resistant S. aureus and methicillin-sensitive S. aureus by >70%, and restricted intracellular growth by >40%. Enhancement of macrophage anti-S. aureus activities is independent of contact with platelets but is mediated through releasable products, namely IL-1ß. These data confirm our hypothesis that platelets participate in host defense against S. aureus both through direct killing of S. aureus and enhancing the antimicrobial function of macrophages in protection against S. aureus infection.
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Plaquetas/inmunología , Citotoxicidad Inmunológica/inmunología , Activación de Macrófagos/inmunología , Macrófagos/inmunología , Infecciones Estafilocócicas/inmunología , Animales , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Staphylococcus aureus/inmunologíaRESUMEN
Platelets are critical to hemostatic and immunological function, and are key players in cancer progression, metastasis, and cancer-related thrombosis. Platelets interact with immune cells to stimulate anti-tumor responses and can be activated by immune cells and tumor cells. Platelet activation can lead to complex interactions between platelets and tumor cells. Platelets facilitate cancer progression and metastasis by: (1) forming aggregates with tumor cells; (2) inducing tumor growth, epithelial-mesenchymal transition, and invasion; (3) shielding circulating tumor cells from immune surveillance and killing; (4) facilitating tethering and arrest of circulating tumor cells; and (5) promoting angiogenesis and tumor cell establishment at distant sites. Tumor cell-activated platelets also predispose cancer patients to thrombotic events. Tumor cells and tumor-derived microparticles lead to thrombosis by secreting procoagulant factors, resulting in platelet activation and clotting. Platelets play a critical role in cancer progression and thrombosis, and markers of platelet-tumor cell interaction are candidates as biomarkers for cancer progression and thrombosis risk.
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Monoamines, such as 5-HT and tyramine (TA), paralyze both free-living and parasitic nematodes when applied exogenously and serotonergic agonists have been used to clear Haemonchus contortus infections in vivo. Since nematode cell lines are not available and animal screening options are limited, we have developed a screening platform to identify monoamine receptor agonists. Key receptors were expressed heterologously in chimeric, genetically-engineered Caenorhabditis elegans, at sites likely to yield robust phenotypes upon agonist stimulation. This approach potentially preserves the unique pharmacologies of the receptors, while including nematode-specific accessory proteins and the nematode cuticle. Importantly, the sensitivity of monoamine-dependent paralysis could be increased dramatically by hypotonic incubation or the use of bus mutants with increased cuticular permeabilities. We have demonstrated that the monoamine-dependent inhibition of key interneurons, cholinergic motor neurons or body wall muscle inhibited locomotion and caused paralysis. Specifically, 5-HT paralyzed C. elegans 5-HT receptor null animals expressing either nematode, insect or human orthologues of a key Gαo-coupled 5-HT1-like receptor in the cholinergic motor neurons. Importantly, 8-OH-DPAT and PAPP, 5-HT receptor agonists, differentially paralyzed the transgenic animals, with 8-OH-DPAT paralyzing mutant animals expressing the human receptor at concentrations well below those affecting its C. elegans or insect orthologues. Similarly, 5-HT and TA paralyzed C. elegans 5-HT or TA receptor null animals, respectively, expressing either C. elegans or H. contortus 5-HT or TA-gated Cl- channels in either C. elegans cholinergic motor neurons or body wall muscles. Together, these data suggest that this heterologous, ectopic expression screening approach will be useful for the identification of agonists for key monoamine receptors from parasites and could have broad application for the identification of ligands for a host of potential anthelmintic targets.
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Animales Modificados Genéticamente/metabolismo , Antihelmínticos/farmacología , Conducta Animal/efectos de los fármacos , Caenorhabditis elegans/efectos de los fármacos , Agonistas de los Canales de Cloruro/farmacología , Descubrimiento de Drogas/métodos , Agonistas del Receptor de Serotonina 5-HT1/farmacología , Animales , Animales Modificados Genéticamente/genética , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/agonistas , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Drosophila/agonistas , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Subunidades alfa de la Proteína de Unión al GTP/química , Subunidades alfa de la Proteína de Unión al GTP/genética , Subunidades alfa de la Proteína de Unión al GTP/metabolismo , Haemonchus , Proteínas del Helminto/agonistas , Proteínas del Helminto/genética , Proteínas del Helminto/metabolismo , Humanos , Soluciones Hipotónicas/toxicidad , Interneuronas/efectos de los fármacos , Interneuronas/metabolismo , Actividad Motora/efectos de los fármacos , Proteínas del Tejido Nervioso/agonistas , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Receptores de Amina Biogénica/agonistas , Receptores de Amina Biogénica/genética , Receptores de Amina Biogénica/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismoRESUMEN
Platelets are anucleate cell fragments known for their central role in coagulation and vascular integrity. However, it is becoming increasingly clear that platelets contribute to diverse immunological processes extending beyond the traditional view of platelets as fragmentary mediators of hemostasis and thrombosis. There is recent evidence that platelets participate in: 1) intervention against microbial threats; 2) recruitment and promotion of innate effector cell functions; 3) modulating antigen presentation; and 4) enhancement of adaptive immune responses. In this way, platelets should be viewed as the underappreciated orchestrator of the immune system. This review will discuss recent and historical evidence regarding how platelets influence both innate and adaptive immune responses.