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2.
PLoS Pathog ; 20(4): e1012186, 2024 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-38648216

RESUMEN

In the bloodstream of mammalian hosts, African trypanosomes face the challenge of protecting their invariant surface receptors from immune detection. This crucial role is fulfilled by a dense, glycosylated protein layer composed of variant surface glycoproteins (VSGs), which undergo antigenic variation and provide a physical barrier that shields the underlying invariant surface glycoproteins (ISGs). The protective shield's limited permeability comes at the cost of restricted access to the extracellular host environment, raising questions regarding the specific function of the ISG repertoire. In this study, we employ an integrative structural biology approach to show that intrinsically disordered membrane-proximal regions are a common feature of members of the ISG super-family, conferring the ability to switch between compact and elongated conformers. While the folded, membrane-distal ectodomain is buried within the VSG layer for compact conformers, their elongated counterparts would enable the extension beyond it. This dynamic behavior enables ISGs to maintain a low immunogenic footprint while still allowing them to engage with the host environment when necessary. Our findings add further evidence to a dynamic molecular organization of trypanosome surface antigens wherein intrinsic disorder underpins the characteristics of a highly flexible ISG proteome to circumvent the constraints imposed by the VSG coat.


Asunto(s)
Tripanosomiasis Africana , Glicoproteínas Variantes de Superficie de Trypanosoma , Glicoproteínas Variantes de Superficie de Trypanosoma/metabolismo , Tripanosomiasis Africana/parasitología , Tripanosomiasis Africana/inmunología , Proteínas Protozoarias/metabolismo , Humanos , Glicoproteínas de Membrana/metabolismo , Animales
3.
Proc Natl Acad Sci U S A ; 120(4): e2208425120, 2023 Jan 24.
Artículo en Inglés | MEDLINE | ID: mdl-36669119

RESUMEN

Recurrent spillovers of α- and ß-coronaviruses (CoV) such as severe acute respiratory syndrome (SARS)-CoV, Middle East respiratory syndrome-CoV, SARS-CoV-2, and possibly human CoV have caused serious morbidity and mortality worldwide. In this study, six receptor-binding domains (RBDs) derived from α- and ß-CoV that are considered to have originated from animals and cross-infected humans were linked to a heterotrimeric scaffold, proliferating cell nuclear antigen (PCNA) subunits, PCNA1, PCNA2, and PCNA3. They assemble to create a stable mosaic multivalent nanoparticle, 6RBD-np, displaying a ring-shaped disk with six protruding antigens, like jewels in a crown. Prime-boost immunizations with 6RBD-np in mice induced significantly high Ab titers against RBD antigens derived from α- and ß-CoV and increased interferon (IFN-γ) production, with full protection against the SARS-CoV-2 wild type and Delta challenges. The mosaic 6RBD-np has the potential to induce intergenus cross-reactivity and to be developed as a pan-CoV vaccine against future CoV spillovers.


Asunto(s)
COVID-19 , Nanopartículas , Humanos , Animales , Ratones , SARS-CoV-2 , Anticuerpos Antivirales , COVID-19/prevención & control , Anticuerpos Neutralizantes , Glicoproteína de la Espiga del Coronavirus/genética
4.
FASEB J ; 38(16): e23893, 2024 Aug 31.
Artículo en Inglés | MEDLINE | ID: mdl-39177943

RESUMEN

Visceral leishmaniasis (VL) is characterized by an uncontrolled infection of internal organs such as the spleen, liver and bone marrow (BM) and can be lethal when left untreated. No effective vaccination is currently available for humans. The importance of B cells in infection and VL protective immunity has been controversial, with both detrimental and protective effects described. VL infection was found in this study to increase not only all analyzed B cell subsets in the spleen but also the B cell progenitors in the BM. The enhanced B lymphopoiesis aligns with the clinical manifestation of polyclonal hypergammaglobulinemia and the occurrence of autoantibodies. In line with earlier reports, flow cytometric and microscopic examination identified parasite attachment to B cells of the BM and spleen without internalization, and transformation of promastigotes into amastigote morphotypes. The interaction appears independent of IgM expression and is associated with an increased detection of activated lysosomes. Furthermore, the extracellularly attached amastigotes could be efficiently transferred to infect macrophages. The observed interaction underscores the potentially crucial role of B cells during VL infection. Additionally, using immunization against a fluorescent heterologous antigen, it was shown that the infection does not impair immune memory, which is reassuring for vaccination campaigns in VL endemic areas.


Asunto(s)
Linfocitos B , Médula Ósea , Memoria Inmunológica , Leishmania infantum , Leishmaniasis Visceral , Linfopoyesis , Bazo , Leishmaniasis Visceral/inmunología , Leishmaniasis Visceral/parasitología , Animales , Bazo/inmunología , Bazo/parasitología , Leishmania infantum/inmunología , Leishmania infantum/fisiología , Ratones , Médula Ósea/parasitología , Médula Ósea/inmunología , Linfocitos B/inmunología , Femenino , Ratones Endogámicos BALB C
5.
PLoS Pathog ; 17(11): e1010026, 2021 11.
Artículo en Inglés | MEDLINE | ID: mdl-34762705

RESUMEN

Salivarian trypanosomes are extracellular protozoan parasites causing infections in a wide range of mammalian hosts, with Trypanosoma evansi having the widest geographic distribution, reaching territories far outside Africa and occasionally even Europe. Besides causing the animal diseases, T. evansi can cause atypical Human Trypanosomosis. The success of this parasite is attributed to its capacity to evade and disable the mammalian defense response. To unravel the latter, we applied here for the first time a scRNA-seq analysis on splenocytes from trypanosome infected mice, at two time points during infection, i.e. just after control of the first parasitemia peak (day 14) and a late chronic time point during infection (day 42). This analysis was combined with flow cytometry and ELISA, revealing that T. evansi induces prompt activation of splenic IgM+CD1d+ Marginal Zone and IgMIntIgD+ Follicular B cells, coinciding with an increase in plasma IgG2c Ab levels. Despite the absence of follicles, a rapid accumulation of Aicda+ GC-like B cells followed first parasitemia peak clearance, accompanied by the occurrence of Xbp1+ expressing CD138+ plasma B cells and Tbx21+ atypical CD11c+ memory B cells. Ablation of immature CD93+ bone marrow and Vpreb3+Ly6d+Ighm+ expressing transitional spleen B cells prevented mature peripheral B cell replenishment. Interestingly, AID-/- mice that lack the capacity to mount anti-parasite IgG responses, exhibited a superior defense level against T. evansi infections. Here, elevated natural IgMs were able to exert in vivo and in vitro trypanocidal activity. Hence, we conclude that in immune competent mice, trypanosomosis associated B cell activation and switched IgG production is rapidly induced by T. evansi, facilitating an escape from the detrimental natural IgM killing activity, and resulting in increased host susceptibility. This unique role of IgM and its anti-trypanosome activity are discussed in the context of the dilemma this causes for the future development of anti-trypanosome vaccines.


Asunto(s)
Linfocitos B/inmunología , Citidina Desaminasa/fisiología , Cambio de Clase de Inmunoglobulina , Mutación , Análisis de la Célula Individual/métodos , Trypanosoma/genética , Tripanosomiasis/parasitología , Animales , Anticuerpos Antiprotozoarios/inmunología , Femenino , Isotipos de Inmunoglobulinas/inmunología , Activación de Linfocitos , Células B de Memoria/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Transcriptoma , Trypanosoma/inmunología , Tripanosomiasis/genética , Tripanosomiasis/inmunología
6.
Int J Mol Sci ; 24(6)2023 Mar 22.
Artículo en Inglés | MEDLINE | ID: mdl-36983063

RESUMEN

Nanobodies, also referred to as single domain-based VHHs, are antibody fragments derived from heavy-chain only IgG antibodies found in the Camelidae family. Due to their small size, simple structure, high antigen binding affinity, and remarkable stability in extreme conditions, nanobodies possess the potential to overcome several of the limitations of conventional monoclonal antibodies. For many years, nanobodies have been of great interest in a wide variety of research fields, particularly in the diagnosis and treatment of diseases. This culminated in the approval of the world's first nanobody based drug (Caplacizumab) in 2018 with others following soon thereafter. This review will provide an overview, with examples, of (i) the structure and advantages of nanobodies compared to conventional monoclonal antibodies, (ii) methods used to generate and produce antigen-specific nanobodies, (iii) applications for diagnostics, and (iv) ongoing clinical trials for nanobody therapeutics as well as promising candidates for clinical development.


Asunto(s)
Anticuerpos de Dominio Único , Anticuerpos Monoclonales/uso terapéutico , Fagocitosis , Inmunoglobulina G
7.
J Infect Dis ; 226(3): 528-540, 2022 08 26.
Artículo en Inglés | MEDLINE | ID: mdl-35363871

RESUMEN

BACKGROUND: Trypanosoma brucei brucei evades host immune responses by multiple means, including the disruption of B-cell homeostasis. This hampers anti-trypanosome vaccine development. Because the cellular mechanism underlying this pathology has never been addressed, our study focuses on the fate of memory B cells (MBCs) in vaccinated mice upon trypanosome challenge. METHODS: A trypanosome variant surface glycoprotein (VSG) and fluorescent phycoerythrin were used as immunization antigens. Functional and cellular characteristics of antigen-specific MBCs were studied after homologous and heterologous parasite challenge. RESULTS: Immunization with AnTat1.1 VSG triggers a specific antibody response and isotype-switched CD73+CD273+CD80+ MBCs, delivering 90% sterile protection against a homologous parasite challenge. As expected, AnTat1.1 VSG immunization does not protect against infection with heterologous VSG-switched parasites. After successful curative drug treatment, mice were shown to have completely lost their previously induced protective immunity against the homologous parasites, coinciding with the loss of vaccine-induced MBCs. A phycoerythrin immunization approach confirmed that trypanosome infections cause the general loss of antigen-specific splenic and bone marrow MBCs and a reduction in antigen-specific immunoglobulin G. CONCLUSIONS: Trypanosomosis induces general immunological memory loss. This benefits the parasites by reducing the stringency for antigenic variation requirements.


Asunto(s)
Trypanosoma brucei brucei , Tripanosomiasis Africana , Animales , Células B de Memoria , Ratones , Ficoeritrina , Glicoproteínas Variantes de Superficie de Trypanosoma
8.
Immunology ; 165(2): 219-233, 2022 02.
Artículo en Inglés | MEDLINE | ID: mdl-34775598

RESUMEN

Tuberculosis (TB) and malaria remain serious threats to global health. Bacillus Calmette-Guerin (BCG), the only licensed vaccine against TB protects against severe disseminated forms of TB in infants but shows poor efficacy against pulmonary TB in adults. Co-infections have been reported as one of the factors implicated in vaccine inefficacy. Given the geographical overlap of malaria and TB in areas where BCG vaccination is routinely administered, we hypothesized that virulence-dependent co-infection with Plasmodium species could alter the BCG-specific immune responses thus resulting in failure to protect against Mycobacterium tuberculosis. We compared virulent Plasmodium berghei and non-virulent Plasmodium chabaudi, their effects on B cells, effector and memory T cells, and the outcome on BCG-induced efficacy against M. tuberculosis infection. We demonstrate that malaria co-infection modulates both B- and T-cell immune responses but does not significantly alter the ability of the BCG vaccine to inhibit the growth of M. tuberculosis irrespective of parasite virulence. This malaria-driven immune regulation may have serious consequences in the early clinical trials of novel vaccines, which rely on vaccine-specific T-cell responses to screen novel vaccines for progression to the more costly vaccine efficacy trials.


Asunto(s)
Vacuna BCG/inmunología , Interacciones Huésped-Parásitos/inmunología , Interacciones Huésped-Patógeno/inmunología , Inmunogenicidad Vacunal , Mycobacterium tuberculosis/inmunología , Tuberculosis Pulmonar/prevención & control , Tuberculosis/prevención & control , Animales , Apoptosis , Recuento de Linfocito CD4 , Modelos Animales de Enfermedad , Femenino , Humanos , Malaria/inmunología , Malaria/parasitología , Células T de Memoria/inmunología , Células T de Memoria/metabolismo , Ratones , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Vacunas contra la Tuberculosis/inmunología , Vacunación
9.
PLoS Pathog ; 16(2): e1008170, 2020 02.
Artículo en Inglés | MEDLINE | ID: mdl-32012211

RESUMEN

Bovine African Trypanosomosis is an infectious parasitic disease affecting livestock productivity and thereby impairing the economic development of Sub-Saharan Africa. The most important trypanosome species implicated is T. congolense, causing anemia as most important pathological feature. Using murine models, it was shown that due to the parasite's efficient immune evasion mechanisms, including (i) antigenic variation of the variable surface glycoprotein (VSG) coat, (ii) induction of polyclonal B cell activation, (iii) loss of B cell memory and (iv) T cell mediated immunosuppression, disease prevention through vaccination has so far been impossible. In trypanotolerant models a strong, early pro-inflammatory immune response involving IFN-γ, TNF and NO, combined with a strong humoral anti-VSG response, ensures early parasitemia control. This potent protective inflammatory response is counterbalanced by the production of the anti-inflammatory cytokine IL-10, which in turn prevents early death of the host from uncontrolled hyper-inflammation-mediated immunopathologies. Though at this stage different hematopoietic cells, such as NK cells, T cells and B cells as well as myeloid cells (i.e. alternatively activated myeloid cells (M2) or Ly6c- monocytes), were found to produce IL-10, the contribution of non-hematopoietic cells as potential IL-10 source during experimental T. congolense infection has not been addressed. Here, we report for the first time that during the chronic stage of T. congolense infection non-hematopoietic cells constitute an important source of IL-10. Our data shows that hepatocyte-derived IL-10 is mandatory for host survival and is crucial for the control of trypanosomosis-induced inflammation and associated immunopathologies such as anemia, hepatosplenomegaly and excessive tissue injury.


Asunto(s)
Hepatocitos , Evasión Inmune , Interleucina-10/inmunología , Trypanosoma congolense , Tripanosomiasis Africana , Animales , Linfocitos B/inmunología , Linfocitos B/patología , Enfermedad Crónica , Modelos Animales de Enfermedad , Femenino , Hepatocitos/inmunología , Hepatocitos/parasitología , Hepatocitos/patología , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/patología , Activación de Linfocitos , Ratones , Monocitos/inmunología , Monocitos/patología , Linfocitos T/inmunología , Linfocitos T/patología , Trypanosoma congolense/inmunología , Trypanosoma congolense/patogenicidad , Tripanosomiasis Africana/inmunología , Tripanosomiasis Africana/patología
10.
Protein Expr Purif ; 185: 105906, 2021 09.
Artículo en Inglés | MEDLINE | ID: mdl-33991675

RESUMEN

Nanobodies are single-domain antibody constructs derived from the variable regions of heavy chain only (VHH) camelid IgGs. Their small size and single gene format make them amenable to various molecular biology applications that require a protein affinity-based approach. These features, in addition to their high solubility, allows their periplasmic expression, extraction and purification in E. coli systems with relative ease, using standardized protocols. However, some Nanobodies are recalcitrant to periplasmic expression, extraction and purification within E. coli systems. To improve their expression would require either a change in the expression host, vector or an increased scale of expression, all of which entail an increase in the complexity of their expression, and production cost. However, as shown here, specific changes in the existing standard E. coli culture protocol, aimed at reducing breakdown of selective antibiotic pressure, increasing the initial culture inoculum and improving transport to the periplasmic space, rescued the expression of several such refractory Nanobodies. The periplasmic extraction protocol was also changed to ensure efficient osmolysis, prevent both protein degradation and prevent downstream chelation of Ni2+ ions during IMAC purification. Adoption of this protocol will lead to an improvement of the expression of Nanobodies in general, and specifically, those that are recalcitrant.


Asunto(s)
Escherichia coli/metabolismo , Periplasma/metabolismo , Proteínas Recombinantes/aislamiento & purificación , Anticuerpos de Dominio Único/biosíntesis , Secuencia de Aminoácidos , Clonación Molecular , Medios de Cultivo/química , Medios de Cultivo/farmacología , Escherichia coli/genética , Expresión Génica , Vectores Genéticos/química , Vectores Genéticos/metabolismo , Presión Osmótica , Periplasma/genética , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/efectos de los fármacos , Proteínas Recombinantes/genética , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Anticuerpos de Dominio Único/genética , Anticuerpos de Dominio Único/aislamiento & purificación
11.
Parasite Immunol ; 41(10): e12664, 2019 10.
Artículo en Inglés | MEDLINE | ID: mdl-31325372

RESUMEN

Trypanosomosis is a chronic parasitic infection, affecting both humans and livestock. A common hallmark of experimental murine infections is the occurrence of inflammation and the associated remodelling of the spleen compartment. The latter involves the depletion of several lymphocyte populations, the induction of T-cell-mediated immune suppression, and the activation of monocyte/macrophage cell populations. Here, we show that in experimental T b brucei infections in mice, these changes are accompanied by the alteration of the spleen neutrophil compartment. Indeed, mature neutrophils are rapidly recruited to the spleen, and cell numbers remain elevated during the entire infection. Following the second peak of parasitemia, the neutrophil cell influx coincides with the rapid reduction of splenic marginal zone (MZ)B and follicular (Fo)B cells, as well as CD8+ T and NK1.1+ cells, the latter encompassing both natural killer (NK) and natural killer T (NKT) cells. This report is the first to show a comprehensive overview of all alterations in spleen cell populations, measured with short intervals throughout the entire course of an experimental T b brucei infection. These data provide new insights into the dynamic interlinked changes in spleen cell numbers associated with trypanosomosis-associated immunopathology.


Asunto(s)
Neutrófilos/inmunología , Trypanosoma brucei brucei/fisiología , Animales , Linfocitos B/inmunología , Linfocitos T CD8-positivos/inmunología , Humanos , Ratones , Ratones Endogámicos C57BL , Células T Asesinas Naturales/inmunología , Parasitemia/inmunología , Bazo/citología , Bazo/inmunología , Tripanosomiasis Africana/inmunología
12.
PLoS Pathog ; 12(7): e1005733, 2016 07.
Artículo en Inglés | MEDLINE | ID: mdl-27403737

RESUMEN

After infection with T. brucei AnTat 1.1, C57BL/6 mice lost splenic B2 B cells and lymphoid follicles, developed poor parasite-specific antibody responses, lost weight, became anemic and died with fulminating parasitemia within 35 days. In contrast, infected C57BL/6 mice lacking the cytotoxic granule pore-forming protein perforin (Prf1-/-) retained splenic B2 B cells and lymphoid follicles, developed high-titer antibody responses against many trypanosome polypeptides, rapidly suppressed parasitemia and did not develop anemia or lose weight for at least 60 days. Several lines of evidence show that T. brucei infection-induced splenic B cell depletion results from natural killer (NK) cell-mediated cytotoxicity: i) B2 B cells were depleted from the spleens of infected intact, T cell deficient (TCR-/-) and FcγRIIIa deficient (CD16-/-) C57BL/6 mice excluding a requirement for T cells, NKT cell, or antibody-dependent cell-mediated cytotoxicity; ii) administration of NK1.1 specific IgG2a (mAb PK136) but not irrelevant IgG2a (myeloma M9144) prevented infection-induced B cell depletion consistent with a requirement for NK cells; iii) splenic NK cells but not T cells or NKT cells degranulated in infected C57BL/6 mice co-incident with B cell depletion evidenced by increased surface expression of CD107a; iv) purified NK cells from naïve C57BL/6 mice killed purified splenic B cells from T. brucei infected but not uninfected mice in vitro indicating acquisition of an NK cell activating phenotype by the post-infection B cells; v) adoptively transferred C57BL/6 NK cells prevented infection-induced B cell population growth in infected Prf1-/- mice consistent with in vivo B cell killing; vi) degranulated NK cells in infected mice had altered gene and differentiation antigen expression and lost cytotoxic activity consistent with functional exhaustion, but increased in number as infection progressed indicating continued generation. We conclude that NK cells in T. brucei infected mice kill B cells, suppress humoral immunity and expedite early mortality.


Asunto(s)
Subgrupos de Linfocitos B/inmunología , Células Asesinas Naturales/inmunología , Activación de Linfocitos/inmunología , Bazo/inmunología , Tripanosomiasis Africana/inmunología , Animales , Western Blotting , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Trypanosoma brucei brucei/inmunología
13.
PLoS Pathog ; 12(9): e1005862, 2016 09.
Artículo en Inglés | MEDLINE | ID: mdl-27632207

RESUMEN

Animal African trypanosomosis is a major threat to the economic development and human health in sub-Saharan Africa. Trypanosoma congolense infections represent the major constraint in livestock production, with anemia as the major pathogenic lethal feature. The mechanisms underlying anemia development are ill defined, which hampers the development of an effective therapy. Here, the contribution of the erythropoietic and erythrophagocytic potential as well as of hemodilution to the development of T. congolense-induced anemia were addressed in a mouse model of low virulence relevant for bovine trypanosomosis. We show that in infected mice, splenic extramedullary erythropoiesis could compensate for the chronic low-grade type I inflammation-induced phagocytosis of senescent red blood cells (RBCs) in spleen and liver myeloid cells, as well as for the impaired maturation of RBCs occurring in the bone marrow and spleen. Rather, anemia resulted from hemodilution. Our data also suggest that the heme catabolism subsequent to sustained erythrophagocytosis resulted in iron accumulation in tissue and hyperbilirubinemia. Moreover, hypoalbuminemia, potentially resulting from hemodilution and liver injury in infected mice, impaired the elimination of toxic circulating molecules like bilirubin. Hemodilutional thrombocytopenia also coincided with impaired coagulation. Combined, these effects could elicit multiple organ failure and uncontrolled bleeding thus reduce the survival of infected mice. MIF (macrophage migrating inhibitory factor), a potential pathogenic molecule in African trypanosomosis, was found herein to promote erythrophagocytosis, to block extramedullary erythropoiesis and RBC maturation, and to trigger hemodilution. Hence, these data prompt considering MIF as a potential target for treatment of natural bovine trypanosomosis.


Asunto(s)
Anemia/metabolismo , Eritropoyesis , Hematopoyesis Extramedular , Oxidorreductasas Intramoleculares/metabolismo , Factores Inhibidores de la Migración de Macrófagos/metabolismo , Trypanosoma congolense/metabolismo , Tripanosomiasis Africana/metabolismo , Anemia/genética , Anemia/parasitología , Anemia/patología , Animales , Médula Ósea/metabolismo , Médula Ósea/parasitología , Médula Ósea/patología , Bovinos , Modelos Animales de Enfermedad , Eritrocitos/metabolismo , Eritrocitos/parasitología , Eritrocitos/patología , Hemodilución , Humanos , Oxidorreductasas Intramoleculares/genética , Factores Inhibidores de la Migración de Macrófagos/genética , Ratones , Ratones Noqueados , Bazo/metabolismo , Bazo/parasitología , Bazo/patología , Trombocitopenia/genética , Trombocitopenia/metabolismo , Trombocitopenia/parasitología , Trombocitopenia/patología , Tripanosomiasis Africana/genética , Tripanosomiasis Africana/patología
14.
PLoS Pathog ; 11(7): e1005065, 2015 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-26222157

RESUMEN

African trypanosomes are extracellular protozoan parasites causing a chronic debilitating disease associated with a persistent inflammatory response. Maintaining the balance of the inflammatory response via downregulation of activation of M1-type myeloid cells was previously shown to be crucial to allow prolonged survival. Here we demonstrate that infection with African trypanosomes of IL-27 receptor-deficient (IL-27R-/-) mice results in severe liver immunopathology and dramatically reduced survival as compared to wild-type mice. This coincides with the development of an exacerbated Th1-mediated immune response with overactivation of CD4+ T cells and strongly enhanced production of inflammatory cytokines including IFN-γ. What is important is that IL-10 production was not impaired in infected IL-27R-/- mice. Depletion of CD4+ T cells in infected IL-27R-/- mice resulted in a dramatically reduced production of IFN-γ, preventing the early mortality of infected IL-27R-/- mice. This was accompanied by a significantly reduced inflammatory response and a major amelioration of liver pathology. These results could be mimicked by treating IL-27R-/- mice with a neutralizing anti-IFN-γ antibody. Thus, our data identify IL-27 signaling as a novel pathway to prevent early mortality via inhibiting hyperactivation of CD4+ Th1 cells and their excessive secretion of IFN-γ during infection with African trypanosomes. These data are the first to demonstrate the essential role of IL-27 signaling in regulating immune responses to extracellular protozoan infections.


Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Interferón gamma/biosíntesis , Interleucinas/inmunología , Transducción de Señal/inmunología , Tripanosomiasis/inmunología , Animales , Muerte Celular , Interleucinas/genética , Interleucinas/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Trypanosoma/inmunología
15.
PLoS Pathog ; 11(6): e1004964, 2015 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-26070118

RESUMEN

African trypanosomes are the causative agents of Human African Trypanosomosis (HAT/Sleeping Sickness) and Animal African Trypanosomosis (AAT/Nagana). A common hallmark of African trypanosome infections is inflammation. In murine trypanosomosis, the onset of inflammation occurs rapidly after infection and is manifested by an influx of myeloid cells in both liver and spleen, accompanied by a burst of serum pro-inflammatory cytokines. Within 48 hours after reaching peak parasitemia, acute anemia develops and the percentage of red blood cells drops by 50%. Using a newly developed in vivo erythrophagocytosis assay, we recently demonstrated that activated cells of the myeloid phagocytic system display enhanced erythrophagocytosis causing acute anemia. Here, we aimed to elucidate the mechanism and immune pathway behind this phenomenon in a murine model for trypanosomosis. Results indicate that IFNγ plays a crucial role in the recruitment and activation of erythrophagocytic myeloid cells, as mice lacking the IFNγ receptor were partially protected against trypanosomosis-associated inflammation and acute anemia. NK and NKT cells were the earliest source of IFNγ during T. b. brucei infection. Later in infection, CD8+ and to a lesser extent CD4+ T cells become the main IFNγ producers. Cell depletion and transfer experiments indicated that during infection the absence of NK, NKT and CD8+ T cells, but not CD4+ T cells, resulted in a reduced anemic phenotype similar to trypanosome infected IFNγR-/- mice. Collectively, this study shows that NK, NKT and CD8+ T cell-derived IFNγ is a critical mediator in trypanosomosis-associated pathology, driving enhanced erythrophagocytosis by myeloid phagocytic cells and the induction of acute inflammation-associated anemia.


Asunto(s)
Anemia/inmunología , Eritrocitos/patología , Interferón gamma/inmunología , Células Mieloides/inmunología , Fagocitosis/inmunología , Tripanosomiasis Africana/inmunología , Traslado Adoptivo , Animales , Linfocitos T CD8-positivos/inmunología , Separación Celular , Modelos Animales de Enfermedad , Femenino , Citometría de Flujo , Células Asesinas Naturales/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células T Asesinas Naturales/inmunología , Tripanosomiasis Africana/complicaciones
16.
PLoS Pathog ; 11(6): e1004942, 2015 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-26110623

RESUMEN

African trypanosomiasis is a deadly neglected disease caused by the extracellular parasite Trypanosoma brucei. Current therapies are characterized by high drug toxicity and increasing drug resistance mainly associated with loss-of-function mutations in the transporters involved in drug import. The introduction of new antiparasitic drugs into therapeutic use is a slow and expensive process. In contrast, specific targeting of existing drugs could represent a more rapid and cost-effective approach for neglected disease treatment, impacting through reduced systemic toxicity and circumventing resistance acquired through impaired compound uptake. We have generated nanoparticles of chitosan loaded with the trypanocidal drug pentamidine and coated by a single domain nanobody that specifically targets the surface of African trypanosomes. Once loaded into this nanocarrier, pentamidine enters trypanosomes through endocytosis instead of via classical cell surface transporters. The curative dose of pentamidine-loaded nanobody-chitosan nanoparticles was 100-fold lower than pentamidine alone in a murine model of acute African trypanosomiasis. Crucially, this new formulation displayed undiminished in vitro and in vivo activity against a trypanosome cell line resistant to pentamidine as a result of mutations in the surface transporter aquaglyceroporin 2. We conclude that this new drug delivery system increases drug efficacy and has the ability to overcome resistance to some anti-protozoal drugs.


Asunto(s)
Resistencia a Medicamentos/efectos de los fármacos , Terapia Molecular Dirigida/métodos , Pentamidina/administración & dosificación , Tripanocidas/administración & dosificación , Tripanosomiasis Africana/tratamiento farmacológico , Animales , Anticuerpos Antiprotozoarios/administración & dosificación , Quitosano/administración & dosificación , Quitosano/farmacocinética , Modelos Animales de Enfermedad , Portadores de Fármacos/administración & dosificación , Portadores de Fármacos/farmacocinética , Ensayo de Cambio de Movilidad Electroforética , Femenino , Concentración 50 Inhibidora , Ratones , Ratones Endogámicos C57BL , Nanopartículas/uso terapéutico , Pentamidina/farmacocinética , Reacción en Cadena en Tiempo Real de la Polimerasa , Tripanocidas/farmacocinética
17.
Parasitology ; 142(3): 417-27, 2015 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-25479093

RESUMEN

African trypanosomes have been around for more than 100 million years, and have adapted to survival in a very wide host range. While various indigenous African mammalian host species display a tolerant phenotype towards this parasitic infection, and hence serve as perpetual reservoirs, many commercially important livestock species are highly disease susceptible. When considering humans, they too display a highly sensitive disease progression phenotype for infections with Trypanosoma brucei rhodesiense or Trypanosoma brucei gambiense, while being intrinsically resistant to infections with other trypanosome species. As extracellular trypanosomes proliferate and live freely in the bloodstream and lymphatics, they are constantly exposed to the immune system. Due to co-evolution, this environment however no longer poses a hostile threat, but has become the niche environment where trypanosomes thrive and obligatory await transmission through the bites of tsetse flies or other haematophagic vectors, ideally without causing severe side infection-associated pathology to their host. Hence, African trypanosomes have acquired various mechanisms to manipulate and control the host immune response, evading effective elimination. Despite the extensive research into trypanosomosis over the past 40 years, many aspects of the anti-parasite immune response remain to be solved and no vaccine is currently available. Here we review the recent work on the different escape mechanisms employed by African Trypanosomes to ensure infection chronicity and transmission potential.


Asunto(s)
Interacciones Huésped-Parásitos/inmunología , Trypanosoma brucei brucei/fisiología , Tripanosomiasis Africana , Animales , Variación Antigénica/inmunología , Humanos , Evasión Inmune/fisiología , Inmunidad Innata/fisiología , Región Variable de Inmunoglobulina/inmunología , Insectos Vectores/parasitología , Trypanosoma brucei brucei/inmunología , Tripanosomiasis Africana/inmunología , Tripanosomiasis Africana/parasitología , Tripanosomiasis Africana/transmisión , Moscas Tse-Tse/parasitología
18.
PLoS Pathog ; 8(3): e1002575, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22479178

RESUMEN

Brucella are facultative intracellular bacteria that chronically infect humans and animals causing brucellosis. Brucella are able to invade and replicate in a broad range of cell lines in vitro, however the cells supporting bacterial growth in vivo are largely unknown. In order to identify these, we used a Brucella melitensis strain stably expressing mCherry fluorescent protein to determine the phenotype of infected cells in spleen and liver, two major sites of B. melitensis growth in mice. In both tissues, the majority of primary infected cells expressed the F4/80 myeloid marker. The peak of infection correlated with granuloma development. These structures were mainly composed of CD11b⁺ F4/80⁺ MHC-II⁺ cells expressing iNOS/NOS2 enzyme. A fraction of these cells also expressed CD11c marker and appeared similar to inflammatory dendritic cells (DCs). Analysis of genetically deficient mice revealed that differentiation of iNOS⁺ inflammatory DC, granuloma formation and control of bacterial growth were deeply affected by the absence of MyD88, IL-12p35 and IFN-γ molecules. During chronic phase of infection in susceptible mice, we identified a particular subset of DC expressing both CD11c and CD205, serving as a reservoir for the bacteria. Taken together, our results describe the cellular nature of immune effectors involved during Brucella infection and reveal a previously unappreciated role for DC subsets, both as effectors and reservoir cells, in the pathogenesis of brucellosis.


Asunto(s)
Brucella/inmunología , Brucelosis/inmunología , Células Dendríticas/inmunología , Inmunidad Innata , Enfermedades Pulmonares/inmunología , Animales , Biomarcadores/metabolismo , Brucella/patogenicidad , Brucelosis/microbiología , Brucelosis/patología , Separación Celular , Células Dendríticas/microbiología , Células Dendríticas/patología , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Hígado/inmunología , Hígado/microbiología , Hígado/patología , Enfermedades Pulmonares/microbiología , Enfermedades Pulmonares/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Óxido Nítrico Sintasa de Tipo I/metabolismo , Óxido Nítrico Sintasa de Tipo II/metabolismo , Fenotipo , Bazo/inmunología , Bazo/microbiología , Bazo/patología
19.
Nitric Oxide ; 36: 36-43, 2014 Jan 30.
Artículo en Inglés | MEDLINE | ID: mdl-24269486

RESUMEN

Sepsis and septic shock result from an exacerbated systemic inflammatory reaction to infection. Their incidence is rising, and they have recently become the main cause of death in intensive care units. Septic shock is defined as sepsis accompanied by life-threatening refractory hypotension, for which excessive nitric oxide (NO), produced by inducible NO synthase iNOS, is thought responsible. LPS, a vital outer membrane component of Gram-negative bacteria, mimics most of the septic effects and is widely used as a model for septic shock. TLR4 is the signal-transducing receptor for LPS, evidenced by the resistance of TLR4-deficient C3H/HeJ and C57BL/10ScNJ mice. As expected, we found that TLR4 deficiency precludes LPS-induced cytokine production, independent of the purity of the LPS preparation. However, various conventional LPS preparations induced NO in TLR4-deficient mice to the same level as in control animals, while ultrapure LPS did not, indicating the presence of NO-producing contaminant(s). Nevertheless, despite identical iNOS induction pattern and systemic NO levels, the contaminant does not cause hypotension, hypothermia, or any other sign of morbidity. Using mice deficient for TLR2, TRL3, TLR4, TRL2x4, TLR9, MyD88 or TRIF, we found that the contaminant signals via TLR2 and MyD88. In conclusion, conventional LPS preparations generally used in endotoxic shock research contain TLR2 agonists that induce iNOS and high levels of systemic NO as such, and synergize with LPS towards the production of pro-inflammatory cytokines, morbidity and mortality. Surprisingly, the excessive iNOS-derived systemic NO production induced by impure LPS in TLR4⁻/⁻ is not accompanied by hypotension or morbidity.


Asunto(s)
Endotoxinas/metabolismo , Óxido Nítrico/metabolismo , Sepsis/metabolismo , Receptor Toll-Like 4/genética , Animales , Hipotensión/genética , Inflamación , Lipopolisacáridos/metabolismo , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Ratones Transgénicos , Óxido Nítrico Sintasa de Tipo II/metabolismo , Transducción de Señal , Receptor Toll-Like 2/genética
20.
Pathogens ; 13(3)2024 Feb 20.
Artículo en Inglés | MEDLINE | ID: mdl-38535532

RESUMEN

Trypanosomes are single-celled extracellular parasites that infect mammals, including humans and livestock, causing global public health concerns and economic losses. These parasites cycle between insect vectors, such as tsetse flies and vertebrate hosts, undergoing morphological, cellular, and biochemical changes. They have remarkable immune evasion mechanisms to escape the host's innate and adaptive immune responses, such as surface coat antigenic variation and the induction of the loss of specificity and memory of antibody responses, enabling the prolongation of infection. Since trypanosomes circulate through the host body in blood and lymph fluid and invade various organs, understanding the interaction between trypanosomes and tissue niches is essential. Here, we present an up-to-date overview of host-parasite interactions and survival strategies for trypanosomes by introducing and discussing the latest studies investigating the transcriptomics of parasites according to life cycle stages, as well as host cells in various tissues and organs, using single-cell and spatial sequencing applications. In recent years, this information has improved our understanding of trypanosomosis by deciphering the diverse populations of parasites in the developmental process, as well as the highly heterogeneous immune and tissue-resident cells involved in anti-trypanosome responses. Ultimately, the goal of these approaches is to gain an in-depth understanding of parasite biology and host immunity, potentially leading to new vaccination and therapeutic strategies against trypanosomosis.

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