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BACKGROUND: Conventional natural killer (cNK) cells play an important role in the innate immune response by directly killing infected and malignant cells and by producing pro- and anti-inflammatory cytokines. Studies on their role in malaria and its complications have resulted in conflicting results. METHODS: Using the commonly used anti-NK1.1 depletion antibodies (PK136) in an in-house optimized experimental model for malaria-associated acute respiratory distress syndrome (MA-ARDS), the role of cNK cells was investigated. Moreover, flow cytometry was performed to characterize different NK cell populations. RESULTS: While cNK cells were found to be dispensable in the development of MA-ARDS, the appearance of a NK1.1+ cell population was observed in the lungs upon infection despite depletion with anti-NK1.1. Detailed characterization of the unknown population revealed that this population consisted of a mixture of monocytes and macrophages that bind the anti-NK1.1 antibody in an aspecific way. This aspecific binding may occur via Fcγ receptors, such as FcγR4. In contrast, in vivo depletion using anti-NK1.1 antibodies was proved to be specific for cNK cells. CONCLUSION: cNK cells are dispensable in the development of experimental MA-ARDS. Moreover, careful flow cytometric analysis, with a critical mindset in relation to potential aspecific binding despite the use of commercially available Fc blocking reagents, is critical to avoid misinterpretation of the results.
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Malaria , Síndrome de Dificultad Respiratoria , Ratones , Animales , Ratones Endogámicos C57BL , Síndrome de Dificultad Respiratoria/patología , Células Asesinas Naturales , Células Mieloides/patología , Malaria/complicacionesRESUMEN
Malaria is a hazardous disease caused by Plasmodium parasites and often results in lethal complications, including malaria-associated acute respiratory distress syndrome (MA-ARDS). Parasite sequestration in the microvasculature is often observed, but its role in malaria pathogenesis and complications is still incompletely understood. We used skeleton binding protein-1 (SBP-1) KO parasites to study the role of sequestration in experimental MA-ARDS. The sequestration-deficiency of these SBP-1 KO parasites was confirmed with bioluminescence imaging and by measuring parasite accumulation in the lungs with RT-qPCR. The SBP-1 KO parasites induced similar lung pathology in the early stage of experimental MA-ARDS compared to wildtype (WT) parasites. Strikingly, the lung pathology resolved subsequently in more than 60% of the SBP-1 KO infected mice, resulting in prolonged survival despite the continuous presence of the parasite. This spontaneous disease resolution was associated with decreased inflammatory cytokine expression measured by RT-qPCR and lower expression of cytotoxic markers in pathogenic CD8+ T cells in the lungs of SBP-1 KO infected mice. These data suggest that SBP-1-mediated parasite sequestration and subsequent high parasite load are not essential for the development of experimental MA-ARDS but inhibit the resolution of the disease.
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Pulmón/parasitología , Malaria/complicaciones , Proteínas de la Membrana/deficiencia , Plasmodium berghei/patogenicidad , Proteínas Protozoarias/metabolismo , Síndrome de Dificultad Respiratoria/prevención & control , Animales , Progresión de la Enfermedad , Femenino , Pulmón/metabolismo , Pulmón/patología , Malaria/parasitología , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas Protozoarias/genética , Síndrome de Dificultad Respiratoria/metabolismo , Síndrome de Dificultad Respiratoria/parasitología , Síndrome de Dificultad Respiratoria/patologíaRESUMEN
BACKGROUND: Malaria-associated acute respiratory distress syndrome (MA-ARDS) is an understudied complication of malaria and is characterized by pulmonary inflammation and disruption of the alveolar-capillary membrane. Its pathogenesis remains poorly understood. Since endothelial activation plays an important role in other malarial complications, the expression of two endothelial activation markers, von Willebrand factor (VWF) and angiopoietin-2 (ANG-2), was investigated in the lungs of patients with MA-ARDS. METHODS: Post-mortem lung sections of Plasmodium falciparum-infected patients without alveolar oedema (NA), P. falciparum-infected patients with alveolar oedema (MA-ARDS), and uninfected people who died accidentally with no pathological changes to the lungs (CON) were immunohistochemically stained for VWF and ANG-2, and were evaluated with semi-quantitative analysis. RESULTS: Alveolar oedematous VWF and ANG-2 and intravascular VWF staining were significantly increased in patients with MA-ARDS versus infected and uninfected control groups. The levels of VWF in the alveolar septa and endothelial lining of large blood vessels of patients with MA-ARDS was significantly decreased compared to controls. ANG-2 expression was increased in the alveolar septa of malaria patients without alveolar oedema versus control patients, while ANG-2+ leukocytes were increased in the alveoli in both infected patient groups. CONCLUSIONS: This study documents a high level of VWF and ANG-2, two endothelial activation markers in the oedematous alveoli of post-mortem lung sections of Thai patients with MA-ARDS. Decreased detection of VWF in the endothelial lining of blood vessels, in parallel with an increased presence of intravascular VWF staining suggests marked endothelial activation and Weibel-Palade body release in the lungs of patients with MA-ARDS.
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Angiopoyetina 2/metabolismo , Pulmón/fisiopatología , Malaria/complicaciones , Síndrome de Dificultad Respiratoria/diagnóstico , Factor de von Willebrand/metabolismo , Adulto , Femenino , Humanos , Pulmón/parasitología , Pulmón/patología , Masculino , Síndrome de Dificultad Respiratoria/parasitología , Síndrome de Dificultad Respiratoria/fisiopatología , Adulto JovenRESUMEN
BACKGROUND: Malaria-associated acute respiratory distress syndrome (MA-ARDS) is a complication of malaria with a lethality rate of up to 80% despite anti-malarial treatment. It is characterized by a vast infiltration of leukocytes, microhaemorrhages and vasogenic oedema in the lungs. Previously, a mouse model for MA-ARDS was developed by infection of C57BL/6 mice with the Edinburgh line NK65-E of Plasmodium berghei. RESULTS: Here, both host and parasite factors were demonstrated to play crucial roles in the development and severity of lung pathology. In particular, the genetic constitution of the host was an important determinant in the development of MA-ARDS. Both male and female C57BL/6, but not BALB/c, mice developed MA-ARDS when infected with P. berghei NK65-E. However, the New York line of P. berghei NK65 (NK65-NY) did not induce demonstrable MA-ARDS, despite its accumulation in the lungs and fat tissue to a similar or even higher extent as P. berghei NK65-E. These two commonly used lines of P. berghei differ in their red blood cell preference. P. berghei NK65-NY showed a stronger predilection for reticulocytes than P. berghei NK65-E and this appeared to be associated with a lower pathogenicity in the lungs. The pulmonary pathology in the C57BL/6/P. berghei NK65-E model was more pronounced than in the model with infection of DBA/2 mice with P. berghei strain ANKA. The transient lung pathology in DBA/2 mice infected with P. berghei ANKA coincided with the infection phase in which parasites mainly infected normocytes. This phase was followed by a less pathogenic phase in which P. berghei ANKA mainly infected reticulocytes. CONCLUSIONS: The propensity of mice to develop MA-ARDS during P. berghei infection depends on both host and parasite factors and appears to correlate with RBC preference. These data provide insights in induction of MA-ARDS and may guide the choice of different mouse-parasite combinations to study lung pathology.
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Modelos Animales de Enfermedad , Malaria/complicaciones , Plasmodium berghei/patogenicidad , Síndrome de Dificultad Respiratoria/patología , Animales , Femenino , Interacciones Huésped-Parásitos , Pulmón/patología , Malaria/parasitología , Masculino , Ratones Endogámicos BALB C , Ratones Endogámicos C57BLRESUMEN
A large proportion of HIV-coinfected visceral leishmaniasis (VL-HIV) patients exhibit chronic disease with frequent VL recurrence. However, knowledge on immunological determinants underlying the disease course is scarce. We longitudinally profiled the circulatory cellular immunity of an Ethiopian HIV cohort that included VL developers. We show that chronic VL-HIV patients exhibit high and persistent levels of TIGIT and PD-1 on CD8+/CD8- T cells, in addition to a lower frequency of IFN-γ+ TIGIT- CD8+/CD8- T cells, suggestive of impaired T cell functionality. At single T cell transcriptome and clonal resolution, the patients show CD4+ T cell anergy, characterised by a lack of T cell activation and lymphoproliferative response. These findings suggest that PD-1 and TIGIT play a pivotal role in VL-HIV chronicity, and may be further explored for patient risk stratification. Our findings provide a strong rationale for adjunctive immunotherapy for the treatment of chronic VL-HIV patients to break the recurrent disease cycle.
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Coinfección , Infecciones por VIH , Leishmaniasis Visceral , Humanos , Leishmaniasis Visceral/inmunología , Leishmaniasis Visceral/complicaciones , Leishmaniasis Visceral/parasitología , Infecciones por VIH/inmunología , Infecciones por VIH/complicaciones , Coinfección/inmunología , Masculino , Adulto , Femenino , Linfocitos T CD8-positivos/inmunología , Persona de Mediana Edad , Enfermedad Crónica , Linfocitos T CD4-Positivos/inmunología , EtiopíaRESUMEN
[This corrects the article DOI: 10.3389/fimmu.2020.628643.].
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Plasmodium parasites contain various virulence factors that modulate the host immune response. Malarial pigment, or hemozoin (Hz), is an undegradable crystalline product of the hemoglobin degradation pathway in the parasite and possesses immunomodulatory properties. An association has been found between Hz accumulation and severe malaria, suggesting that the effects of Hz on the host immune response may contribute to the development of malarial complications. Although the immunomodulatory roles of Hz have been widely investigated, many conflicting data exist, likely due to the variability between experimental set-ups and technical limitations of Hz generation and isolation methods. Here, we critically assess the potential immunomodulatory effects of Hz, its role in malarial complications, and its potential effects after parasite clearance.
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Hemoproteínas/inmunología , Interacciones Huésped-Parásitos/inmunología , Inmunomodulación , Malaria/inmunología , Humanos , Malaria/patología , Investigación/normas , Investigación/tendenciasRESUMEN
Malaria complications are often lethal, despite efficient killing of Plasmodium parasites with antimalarial drugs. This indicates the need to study the resolution and healing mechanisms involved in the recovery from these complications. Plasmodium berghei NK65-infected C57BL/6 mice develop malaria-associated acute respiratory distress syndrome (MA-ARDS) at 8 days post infection. Antimalarial treatment was started on this day and resulted in the recovery, as measured by the disappearance of the signs of pathology, in >80% of the mice. Therefore, this optimized model represents an asset in the study of mechanisms and leukocyte populations involved in the resolution of MA-ARDS. C-C chemokine receptor type 2 (CCR2) knock-out mice were used to investigate the role of monocytes and macrophages, since these cells are described to play an important role during the resolution of other inflammatory diseases. CCR2 deficiency was associated with significantly lower numbers of inflammatory monocytes in the lungs during infection and resolution and abolished the increase in non-classical monocytes during resolution. Surprisingly, CCR2 was dispensable for the development and the resolution of MA-ARDS, since no effect of the CCR2 knock-out was observed on any of the disease parameters. In contrast, the reappearance of eosinophils and interstitial macrophages during resolution was mitigated in the lungs of CCR2 knock-out mice. In conclusion, CCR2 is required for re-establishing the homeostasis of pulmonary leukocytes during recovery. Furthermore, the resolution of malaria-induced lung pathology is mediated by unknown CCR2-independent mechanisms.
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Homeostasis/inmunología , Leucocitos/inmunología , Malaria/inmunología , Plasmodium berghei/inmunología , Receptores CCR2/inmunología , Síndrome de Dificultad Respiratoria/inmunología , Animales , Homeostasis/genética , Leucocitos/patología , Malaria/genética , Malaria/patología , Ratones , Ratones Noqueados , Receptores CCR2/genética , Síndrome de Dificultad Respiratoria/genética , Síndrome de Dificultad Respiratoria/parasitología , Síndrome de Dificultad Respiratoria/patologíaRESUMEN
Neutrophil-depleting antibodies, such as anti-GR1 (RB6-8C5) and anti-Ly6G (1A8), are commonly used to study the in vivo function of neutrophils in murine disease models. Anti-Ly6G antibodies became the standard, because in contrast to anti-GR1, these do not bind Ly6C. The efficiency of the depletion needs to be carefully analysed as flow cytometry plots may be misinterpreted. For example, the staining intensity of GR1 on neutrophils (CD11b+ GR1hi) drops upon anti-Ly6G administration. We show that this drop is due to competition between anti-GR1 and anti-Ly6G antibodies. Neutrophil depletion with anti-Ly6G in naive mice was organ- and strain-specific. Furthermore, an incomplete anti-Ly6G-dependent neutrophil depletion was obtained in two immune-mediated mouse models, i.e. in malaria-infected C57BL/6 mice and in complete Freund's adjuvant (CFA)-challenged BALB/c mice. BrdU-incorporation studies show a slight increase in proliferating bone marrow neutrophils upon depletion in naive mice. Strikingly, depletion with anti-Ly6G in CFA-challenged BALB/c mice resulted in a significant increase in proliferating splenic neutrophils, causing a fast rebound of new immature neutrophils. In conclusion, our results emphasize the importance of careful panel design, gating strategies and duration of neutrophil depletion and highlight the context-dependent Ly6G depletion efficiency. It furthermore underlines the need for new tools to understand the in vivo role of neutrophils in immunological models.
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Anticuerpos Monoclonales/administración & dosificación , Antígenos Ly/inmunología , Tolerancia Inmunológica/efectos de los fármacos , Inflamación/inmunología , Neutrófilos/efectos de los fármacos , Animales , Anticuerpos Monoclonales/inmunología , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Humanos , Inflamación/inducido químicamente , Inyecciones Intraperitoneales , Ratones , Ratones Endogámicos C57BL , Neutrófilos/inmunología , Bazo/citología , Bazo/inmunologíaRESUMEN
BACKGROUND: Malaria-associated acute respiratory distress syndrome (MA-ARDS) is a lethal complication of severe malaria, characterized by marked pulmonary inflammation. Patient studies have suggested a link between von Willebrand factor (VWF) and malaria severity. OBJECTIVES: To investigate the role of VWF in the pathogenesis of experimental MA-ARDS. METHODS: Plasmodium berghei NK65-E (PbNK65) parasites were injected in Vwf+/+ and Vwf-/- mice. Pathological parameters were assessed following infection. RESULTS: In accordance with patients with severe malaria, plasma VWF levels were increased and ADAMTS13 activity levels were reduced in experimental MA-ARDS. ADAMTS13- and plasmin-independent reductions of high molecular weight VWF multimers were observed at the end stage of disease. Thrombocytopenia was VWF-independent because it was observed in both Vwf+/+ and Vwf-/- mice. Interestingly, Vwf-/- mice had a shorter survival time compared with Vwf+/+ controls following PbNK65 infection. Lung edema could not explain this shortened survival because alveolar protein levels in Vwf-/- mice were approximately two times lower than in Vwf+/+ controls. Parasite load, on the other hand, was significantly increased in Vwf-/- mice compared with Vwf+/+ mice in both peripheral blood and lung tissue. In addition, anemia was only observed in PbNK65-infected Vwf-/- mice. Of note, Vwf-/- mice presented with two times more reticulocytes, a preferential target of the parasites. CONCLUSIONS: This study suggests that parasite load together with malarial anemia, rather than alveolar leakage, might contribute to shortened survival in PbNK65-infected Vwf-/- mice. VWF deficiency is associated with early reticulocytosis following PbNK65 infection, which potentially explains the increase in parasite load.
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Malaria/sangre , Malaria/parasitología , Plasmodium berghei/patogenicidad , Síndrome de Dificultad Respiratoria/sangre , Reticulocitos/metabolismo , Enfermedades de von Willebrand/sangre , Factor de von Willebrand/metabolismo , Proteína ADAMTS13/sangre , Anemia/sangre , Anemia/parasitología , Animales , Modelos Animales de Enfermedad , Femenino , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Carga de Parásitos , Síndrome de Dificultad Respiratoria/parasitología , Reticulocitos/parasitología , Reticulocitosis , Trombocitopenia/sangre , Trombocitopenia/parasitología , Enfermedades de von Willebrand/genética , Factor de von Willebrand/genéticaRESUMEN
Malaria reduces host fitness and survival by pathogen-mediated damage and inflammation. Disease tolerance mechanisms counter these negative effects without decreasing pathogen load. Here, we demonstrate that in four different mouse models of malaria, adrenal hormones confer disease tolerance and protect against early death, independently of parasitemia. Surprisingly, adrenalectomy differentially affects malaria-induced inflammation by increasing circulating cytokines and inflammation in the brain but not in the liver or lung. Furthermore, without affecting the transcription of hepatic gluconeogenic enzymes, adrenalectomy causes exhaustion of hepatic glycogen and insulin-independent lethal hypoglycemia upon infection. This hypoglycemia is not prevented by glucose administration or TNF-α neutralization. In contrast, treatment with a synthetic glucocorticoid (dexamethasone) prevents the hypoglycemia, lowers cerebral cytokine expression and increases survival rates. Overall, we conclude that in malaria, adrenal hormones do not protect against lung and liver inflammation. Instead, they prevent excessive systemic and brain inflammation and severe hypoglycemia, thereby contributing to tolerance.
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Glándulas Suprarrenales/metabolismo , Encéfalo/inmunología , Citocinas/inmunología , Hormonas/inmunología , Hipoglucemia/inmunología , Hígado/inmunología , Pulmón/inmunología , Malaria/inmunología , Glándulas Suprarrenales/inmunología , Adrenalectomía , Animales , Glucemia/efectos de los fármacos , Encéfalo/efectos de los fármacos , Corticosterona/inmunología , Corticosterona/metabolismo , Citocinas/efectos de los fármacos , Dexametasona/farmacología , Modelos Animales de Enfermedad , Epinefrina/inmunología , Epinefrina/metabolismo , Glucocorticoides/inmunología , Glucocorticoides/farmacología , Glucógeno/metabolismo , Hidrocortisona/inmunología , Hidrocortisona/metabolismo , Inflamación , Hígado/efectos de los fármacos , Pulmón/efectos de los fármacos , Ratones , Mineralocorticoides/inmunología , Mineralocorticoides/metabolismo , Norepinefrina/inmunología , Norepinefrina/metabolismo , Plasmodium berghei , Plasmodium chabaudi , Tasa de SupervivenciaRESUMEN
Malaria is a severe disease and kills over 400,000 people each year. Malarial complications are the main cause of death and include cerebral malaria and malaria-associated acute respiratory distress syndrome (MA-ARDS). Despite antimalarial treatment, lethality rates of MA-ARDS are still between 20 and 80%. Patients develop pulmonary edema with hemorrhages and leukocyte extravasation in the lungs. The vascular endothelial growth factor-A (VEGF-A) and the placental growth factor (PlGF) are vascular permeability factors and may be involved in the disruption of the alveolar-capillary membrane, leading to alveolar edema. We demonstrated increased pulmonary VEGF-A and PlGF levels in lungs of mice with experimental MA-ARDS. Depletion of pathogenic CD8+ T cells blocked pulmonary edema and abolished the increase of VEGF-A and PlGF. However, neutralization of VEGF receptor-2 (VEGFR-2) with the monoclonal antibody clone DC101 did not decrease pulmonary pathology. The broader spectrum receptor tyrosine kinase inhibitor sunitinib even increased lung pathology. These data suggest that the increase in alveolar VEGF-A and PlGF is not a cause but rather a consequence of the pulmonary pathology in experimental MA-ARDS and that therapeutic inhibition of VEGF receptors is not effective and even contra-indicated.
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Linfocitos T CD8-positivos/inmunología , Malaria/complicaciones , Síndrome de Dificultad Respiratoria/etiología , Factor A de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Células Epiteliales Alveolares , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Neutralizantes , Linfocitos T CD8-positivos/fisiología , Citocinas/metabolismo , Modelos Animales de Enfermedad , Edema/etiología , Femenino , Regulación de la Expresión Génica , Inmunoglobulina G/sangre , Inmunohistoquímica , Pulmón/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Factor de Crecimiento Placentario/metabolismo , Plasmodium berghei , Síndrome de Dificultad Respiratoria/tratamiento farmacológico , Síndrome de Dificultad Respiratoria/patología , Factor A de Crecimiento Endotelial Vascular/genética , Receptor 1 de Factores de Crecimiento Endotelial Vascular/efectos de los fármacos , Receptor 1 de Factores de Crecimiento Endotelial Vascular/inmunología , Receptor 1 de Factores de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/efectos de los fármacos , Receptor 2 de Factores de Crecimiento Endotelial Vascular/inmunologíaRESUMEN
Coevolution of humans and malaria parasites has generated an intricate balance between the immune system of the host and virulence factors of the parasite, equilibrating maximal parasite transmission with limited host damage. Focusing on the blood stage of the disease, we discuss how the balance between anti-parasite immunity versus immunomodulatory and evasion mechanisms of the parasite may result in parasite clearance or chronic infection without major symptoms, whereas imbalances characterized by excessive parasite growth, exaggerated immune reactions or a combination of both cause severe pathology and death, which is detrimental for both parasite and host. A thorough understanding of the immunological balance of malaria and its relation to other physiological balances in the body is of crucial importance for developing effective interventions to reduce malaria-related morbidity and to diminish fatal outcomes due to severe complications. Therefore, we discuss in this review the detailed mechanisms of anti-malarial immunity, parasite virulence factors including immune evasion mechanisms and pathogenesis. Furthermore, we propose a comprehensive classification of malaria complications according to the different types of imbalances.
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Interacciones Huésped-Parásitos/inmunología , Malaria/inmunología , Malaria/parasitología , Plasmodium/inmunología , Animales , Humanos , Malaria/prevención & control , Factores de Virulencia/inmunologíaRESUMEN
Malaria is a global disease that clinically affects more than two hundred million people annually. Despite the availability of effective antimalarials, mortality rates associated with severe complications are high. Hepatopathy is frequently observed in patients with severe malarial disease and its pathogenesis is poorly understood. Previously, we observed high amounts of hemozoin or malaria pigment in livers from infected mice. In this study, we investigated whether hemozoin is associated with liver injury in different mouse malaria models. C57BL/6J mice infected with the rodent parasites Plasmodium berghei ANKA, P. berghei NK65 or P. chabaudi AS had elevated serum liver enzymes without severe histological changes in the liver, in line with the observations in most patients. Furthermore, liver enzymes were significantly higher in serum of P. chabaudi AS-infected mice compared to mice infected with the P. berghei parasite strains and a strong positive correlation was found between hepatic hemozoin levels, hepatocyte damage and inflammation in the liver with P. chabaudi AS. The observed liver injury was only marginally influenced by the genetic background of the host, since similar serum liver enzyme levels were measured in infected C57BL/6J and BALB/c mice. Intravenous injection of P. falciparum-derived hemozoin in malaria-free C57BL/6J mice induced inflammatory gene transcription in the liver, suggesting that hemozoin may be involved in the pathogenesis of malaria hepatopathy by inducing inflammation.