RESUMEN
Subcutaneous dirofilariasis, caused by the parasitic nematode Dirofilaria repens, is a growing concern in Europe, affecting both dogs and humans. This study focused on D. repens Dr20/22, a protein encoded by an alt (abundant larval transcript) gene family. While well-documented in L3 larvae of other filariae species, this gene family had not been explored in dirofilariasis. The research involved cloning Dr20/22 cDNA, molecular characterization, and evaluating its potential application in the diagnosis of dirofilariasis. Although Real-Time analysis revealed mRNA expression in both adult worms and microfilariae, the native protein remained undetected in lysates from both developmental stages. This suggests the protein's specificity for L3 larvae and may be related to a process called SLTS (spliced leader trans-splicing), contributing to stage-specific gene expression. The specificity of the antigen for invasive larvae positions it as a promising early marker for dirofilariasis. However, ELISA tests using sera from infected and uninfected dogs indicated limited diagnostic utility. While further research is required, our findings contribute to a deeper understanding of the molecular and immunological aspects of host-parasite interactions and could offer insights into the parasite's strategies for evading the immune system.
Asunto(s)
Dirofilaria repens , Dirofilariasis , Enfermedades de los Perros , Animales , Perros , Dirofilariasis/inmunología , Dirofilariasis/parasitología , Dirofilaria repens/genética , Dirofilaria repens/inmunología , Enfermedades de los Perros/parasitología , Enfermedades de los Perros/inmunología , Anticuerpos Antihelmínticos/inmunología , Anticuerpos Antihelmínticos/sangre , Proteínas del Helminto/genética , Proteínas del Helminto/inmunología , Proteínas del Helminto/metabolismo , Antígenos Helmínticos/inmunología , Antígenos Helmínticos/genética , Larva/inmunología , Formación de Anticuerpos/inmunologíaRESUMEN
BACKGROUND: Dirofilariasis is a vector-borne disease caused by parasitic nematodes of the genus Dirofilaria spp., considered an emerging concern in both veterinary and human medicine. Climate changes and human activities, such as pet travel, contribute to the spread of diseases to new non-endemic regions. Poland is dominated by subcutaneous dirofilariasis caused by D. repens infections. Cardiopulmonary dirofilariasis, also known as a heartworm disease is much more rare with only single autochthonous cases reported so far. Also, imported infections are observed sporadically in dogs traveling to endemic countries. In this study, we report the first case of a dog in Poland, never having traveled abroad, co-infected with Dirofilaria repens and Dirofilaria immitis. CASE PRESENTATION: A 14-year-old mixed breed, an intact male dog with fever, lightly pale mucosal membranes, moderate abdominal pain, and a mild cough was presented in a veterinary clinic in Warsaw, Poland. The examination of the blood sample collected for complete morphology and biochemistry revealed the presence of live microfilariae. Presence of the DNA of both microfilariae species was detected using Real-Time PCR with species-specific primers. CONCLUSIONS: Since the remaining diagnostic methods like Knott's test, antigen test or echocardiography did not reveal the presence of D. immitis, we discussed the impact of microfilariae periodicity and low worm burden infections on the limited efficiency of these techniques. We strongly recommend using a mixed diagnostic approach for the most sensitive and specific diagnosis since the ideal diagnostic method does not exist, and several factors may contribute to misdiagnosis. Furthermore, we considered factors that contribute to the uncontrolled spread of dirofilariasis such as climate changes, introduction of new species of mosquitoes competent for the transmission of the disease, and wildlife animals as an important reservoir of this parasitosis. Given that Poland shares borders with countries classified as endemic and pre-endemic for D. immitis, such as Slovakia and Ukraine, it is reasonable to anticipate a rise in autochthonous heartworm infections and shifts in the epidemiological pattern of dirofilariasis in the coming years.
Asunto(s)
Dirofilaria immitis , Dirofilaria repens , Dirofilariasis , Enfermedades de los Perros , Humanos , Animales , Perros , Masculino , Dirofilariasis/diagnóstico , Dirofilaria repens/genética , Polonia , Enfermedades de los Perros/epidemiología , Mosquitos Vectores , MicrofilariasRESUMEN
BACKGROUND: Dirofilaria immitis, also known as heartworm, is one of the most important parasitic nematodes of domestic dogs, causing a potentially serious disease, cardiopulmonary dirofilariosis, which can be lethal. This species seems to be less 'expansive' than its sister species Dirofilaria repens, and it is believed that climate change facilitates the spread of this parasite to new non-endemic regions. METHODS: In total, 122 heartworm isolates were analysed from nine endemic countries in Europe (Portugal, Spain, Italy, Greece, Hungary, Romania, Slovakia, and Ukraine) and a single isolate from Bangladesh by amplification and sequencing of two mitochondrial (mt) DNA markers: cytochrome c oxidase subunit 1 (COI) and dehydrogenase subunit 1 (NADH). The main aim of the current study was to determine the genetic diversity of D. immitis and compare it with D. repens haplotype diversity and distribution. DNA was extracted from adult heartworms or microfilariae in blood. Most isolates originated from dogs (Canis lupus familiaris) while 10 isolates originated from wildlife species from Romania, including eight isolates from golden jackals (Canis aureus), one isolate from a Eurasian otter (Lutra lutra) and one isolate from a red fox (Vulpes vulpes). RESULTS: Median spanning network analysis was based on the combined sequence (1721 bp) obtained from two mt markers and successfully delineated nine haplotypes (Di1-Di9). Haplotype Di1 was the dominant haplotype encompassing 91 out of the 122 sequences (75%) from all nine countries and four host species. Haplotype Di2 was the second most common haplotype, formed solely by 13 isolates from Italy. The remaining sequences were assigned to Di3-Di9 haplotypes, differing by 1-4 SNPs from the dominant Di1 haplotype. There was evidence for geographical segregation of haplotypes, with three unique haplotypes associated with Italy and four others associated with certain countries (Di4 and Di7 with Slovakia; Di8 with Greece; Di6 with Hungary). CONCLUSION: Diversity in D. immitis mt haplotypes was lower by half than in D. repens (9 vs. 18 haplotypes in D. immitis and D. repens, respectively), which may be associated with the slower expansion of heartworm in Central and NE Europe. NADH gene appears to be conserved in Dirofilaria sp. by showing lower genetic diversity than the analysed COI gene.
Asunto(s)
Canidae , Dirofilaria immitis , Nutrias , Perros , Animales , Dirofilaria immitis/genética , Haplotipos , NAD , Europa (Continente)/epidemiologíaRESUMEN
During chronic schistosome infections, a complex regulatory network is induced to regulate the host immune system, in which IL-10-producing regulatory B (Breg) cells play a significant role. Schistosoma mansoni soluble egg antigens (SEA) are bound and internalized by B cells and induce both human and mouse IL-10 producing Breg cells. To identify Breg-inducing proteins in SEA, we fractionated SEA by size exclusion chromatography and found 6 fractions able to induce IL-10 production by B cells (out of 18) in the high, medium and low molecular weight (MW) range. The high MW fractions were rich in heavily glycosylated molecules, including multi-fucosylated proteins. Using SEA glycoproteins purified by affinity chromatography and synthetic glycans coupled to gold nanoparticles, we investigated the role of these glycan structures in inducing IL-10 production by B cells. Then, we performed proteomics analysis on active low MW fractions and identified a number of proteins with putative immunomodulatory properties, notably thioredoxin (SmTrx1) and the fatty acid binding protein Sm14. Subsequent splenic murine B cell stimulations and hock immunizations with recombinant SmTrx1 and Sm14 showed their ability to dose-dependently induce IL-10 production by B cells both in vitro and in vivo. Identification of unique Breg cells-inducing molecules may pave the way to innovative therapeutic strategies for inflammatory and auto-immune diseases.
Asunto(s)
Linfocitos B Reguladores , Nanopartículas del Metal , Esquistosomiasis mansoni , Humanos , Animales , Ratones , Schistosoma mansoni , Esquistosomiasis mansoni/prevención & control , Interleucina-10/genética , Oro , Factores Inmunológicos , Tiorredoxinas/genética , Antígenos HelmínticosRESUMEN
Obesity-associated metabolic inflammation drives the development of insulin resistance and type 2 diabetes, notably through modulating innate and adaptive immune cells in metabolic organs. The nutrient sensor liver kinase B1 (LKB1) has recently been shown to control cellular metabolism and T cell priming functions of DCs. Here, we report that hepatic DCs from high-fat diet-fed (HFD-fed) obese mice display increased LKB1 phosphorylation and that LKB1 deficiency in DCs (CD11cΔLKB1) worsened HFD-driven hepatic steatosis and impaired glucose homeostasis. Loss of LKB1 in DCs was associated with increased expression of Th17-polarizing cytokines and accumulation of hepatic IL-17A+ Th cells in HFD-fed mice. Importantly, IL-17A neutralization rescued metabolic perturbations in HFD-fed CD11cΔLKB1 mice. Mechanistically, deficiency of the canonical LKB1 target AMPK in HFD-fed CD11cΔAMPKα1 mice recapitulated neither the hepatic Th17 phenotype nor the disrupted metabolic homeostasis, suggesting the involvement of other and/or additional LKB1 downstream effectors. We indeed provide evidence that the control of Th17 responses by DCs via LKB1 is actually dependent on both AMPKα1 salt-inducible kinase signaling. Altogether, our data reveal a key role for LKB1 signaling in DCs in protection against obesity-induced metabolic dysfunctions by limiting hepatic Th17 responses.
Asunto(s)
Proteínas Quinasas Activadas por AMP , Diabetes Mellitus Tipo 2 , Ratones , Animales , Proteínas Quinasas Activadas por AMP/metabolismo , Interleucina-17/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Obesidad/metabolismo , Hígado/metabolismo , Homeostasis , Células Dendríticas/metabolismoRESUMEN
Background: The parasitic trematode Fasciola hepatica evades host immune defenses through secretion of various immunomodulatory molecules. Fatty Acid Binding Proteins (fhFABPs) are among the main excreted/secreted proteins and have been shown to display anti-inflammatory properties. However, little is currently known regarding their impact on dendritic cells (DCs) and their subsequent capacity to prime specific CD4+ T cell subsets. Methodology/Principal Findings: The immunomodulatory effects of both native F. hepatica extracts and recombinant fhFABPs were assessed on monocyte-derived human DCs (moDCs) and the underlying mechanism was next investigated using various approaches, including DC-allogenic T cell co-culture and DC phenotyping through transcriptomic, proteomic and FACS analyses. We mainly showed that fhFABP1 induced a tolerogenic-like phenotype in LPS-stimulated moDCs characterized by a dose-dependent increase in the cell-surface tolerogenic marker CD103 and IL-10 secretion, while DC co-stimulatory markers were not affected. A significant decrease in secretion of the pro-inflammatory cytokines IL-12p70 and IL-6 was also observed. In addition, these effects were associated with an increase in both Th2-on-Th1 ratio and IL-10 secretion by CD4+ T cells following DC-T cell co-culture. RNA sequencing and targeted proteomic analyses identified thrombospondin-1 (TSP-1) as a non-canonical factor highly expressed and secreted by fhFABP1-primed moDCs. The effect of fhFABP1 on T cell skewing was abolished when using a TSP-1 blocking antibody during DC-T cell co-culture. Immunomodulation by helminth molecules has been linked to improved metabolic homeostasis during obesity. Although fhFABP1 injection in high-fat diet-fed obese mice induced a potent Th2 immune response in adipose tissue, it did not improved insulin sensitivity or glucose homeostasis. Conclusions/Significance: We show that fhFABP1 modulates T cell polarization, notably by promoting DC TSP-1 secretion in vitro, without affecting metabolic homeostasis in a mouse model of type 2 diabetes.
Asunto(s)
Diabetes Mellitus Tipo 2 , Fasciola hepatica , Animales , Células Dendríticas , Diabetes Mellitus Tipo 2/metabolismo , Proteínas de Unión a Ácidos Grasos/genética , Proteínas de Unión a Ácidos Grasos/metabolismo , Homeostasis , Interleucina-10/metabolismo , Ratones , Ratones Obesos , Proteómica , Trombospondina 1/metabolismoRESUMEN
Parasites may significantly affect the functioning of the host organism including immune response and gut-brain-axis ultimately leading to alteration of the host behavior. The impact of intestinal worms on the host central nervous system (CNS) remains unexplored. The aim of this study was to evaluate the effect of intestinal infection by the tapeworm Hymenolepis diminuta on behavior and functions of the CNS in rats. The 3 months old animals were infected, and the effects on anxiety, exploration, sensorimotor skills and learning processes were assessed at 18 months in Open Field (OF), Novel Object Recognition (NOR) and the Water Maze (WM) tests. After completing the behavioral studies, both infected and non-infected rats were sacrificed, and the collected tissues were subjected to biochemical analysis. The levels of neurotransmitters, their metabolites and amino acids in selected structures of the CNS were determined by HPLC. In addition, the gene expression profile of the pro- and anti-inflammatory cytokines (TNF-α, IL-1ß, IL-6 and IL-10) was evaluated by Real-Time PCR to determine the immune response within the CNS to the tapeworm infection. The parasites caused significant changes in exploratory behavior, most notably, a reduction of velocity and total distance moved in the OF test; the infected rats exhibited decreased frequency in the central zone, which may indicate a higher level of anxiety. Additionally, parasite infestation improved spatial memory, assessed in the WM test, and recognition of new objects. These changes are related to the identified reduction in noradrenaline level in the CNS structures and less pronounced changes in striatal serotonergic neurotransmission. H. diminuta infestation was also found to cause a significant reduction of hippocampal expression of IL-6. Our results provide new data for further research on brain function during parasitic infections especially in relation to helminths and diseases in which noradrenergic system may play an important role.
Asunto(s)
Helmintos , Himenolepiasis , Hymenolepis diminuta , Animales , Cognición , Conducta Exploratoria , Helmintiasis , Himenolepiasis/parasitología , Hymenolepis diminuta/fisiología , Interleucina-6 , Parasitosis Intestinales , Neurotransmisores , RatasRESUMEN
Dirofilaria repens is a parasitic nematode causing vector-borne disease (dirofilariasis), considered an emerging problem in veterinary and human medicine. Although main hosts are carnivores, particularly dogs, D. repens shows high zoonotic potential. The disease spreads uncontrollably, affecting new areas. Since there is no vaccine against dirofilariasis, the only way to limit disease transmission is an early diagnosis. Currently, diagnosis depends on the detection of microfilariae in the host bloodstream using modified Knott's test or multiplex PCR. However, the efficacy of tests relying on microfilariae detection is limited by microfilariae periodic occurrence. Therefore, a new reliable diagnostic test is required. Our study aimed to select new diagnostic markers for dirofilariasis with potential application in diagnostics. We focused on single epitopes to ensure high specificity of diagnosis and avoid cross-reactivity with the other parasite infections common in dogs. Using phage display technology and 12-mer peptides library, we selected epitopes highly reactive with IgG from sera of infected dogs. Additionally, our study presents the possibility of detecting D. repens specific cell-free DNA in dogs with no microfilaria but high IgG and IgM antibody levels against parasite somatic antigen.
Asunto(s)
Técnicas de Visualización de Superficie Celular/métodos , ADN de Helmintos/sangre , Dirofilaria repens/genética , Dirofilaria repens/aislamiento & purificación , Dirofilariasis/diagnóstico , Enfermedades de los Perros/diagnóstico , Animales , Biomarcadores/sangre , Dirofilariasis/parasitología , Enfermedades de los Perros/parasitología , Perros , Inmunoglobulina G/sangreRESUMEN
No approved therapies are available for nonalcoholic steatohepatitis (NASH). Adenosine monophosphate-activated protein kinase (AMPK) is a central regulator of cell metabolism; its activation has been suggested as a therapeutic approach to NASH. Here we aimed to fully characterize the potential for direct AMPK activation in preclinical models and to determine mechanisms that could contribute to efficacy for this disease. A novel small-molecule direct AMPK activator, PXL770, was used. Enzyme activity was measured with recombinant complexes. De novo lipogenesis (DNL) was quantitated in vivo and in mouse and human primary hepatocytes. Metabolic efficacy was assessed in ob/ob and high-fat diet-fed mice. Liver histology, biochemical measures, and immune cell profiling were assessed in diet-induced NASH mice. Direct effects on inflammation and fibrogenesis were assessed using primary mouse and human hepatic stellate cells, mouse adipose tissue explants, and human immune cells. PXL770 directly activated AMPK in vitro and reduced DNL in primary hepatocytes. In rodent models with metabolic syndrome, PXL770 improved glycemia, dyslipidemia, and insulin resistance. In mice with NASH, PXL770 reduced hepatic steatosis, ballooning, inflammation, and fibrogenesis. PXL770 exhibited direct inhibitory effects on pro-inflammatory cytokine production and activation of primary hepatic stellate cells. Conclusion: In rodent models, direct activation of AMPK is sufficient to produce improvements in all core components of NASH and to ameliorate related hyperglycemia, dyslipidemia, and systemic inflammation. Novel properties of direct AMPK activation were also unveiled: improved insulin resistance and direct suppression of inflammation and fibrogenesis. Given effects also documented in human cells (reduced DNL, suppression of inflammation and stellate cell activation), these studies support the potential for direct AMPK activation to effectively treat patients with NASH.
Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Enfermedad del Hígado Graso no Alcohólico/enzimología , Animales , Glucemia/metabolismo , Modelos Animales de Enfermedad , Activación Enzimática/efectos de los fármacos , Fibrosis/fisiopatología , Hepatocitos/metabolismo , Humanos , Inflamación/fisiopatología , Insulina/sangre , Lipogénesis/efectos de los fármacos , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/sangre , Enfermedad del Hígado Graso no Alcohólico/patología , Enfermedad del Hígado Graso no Alcohólico/fisiopatología , Piridonas/farmacología , Tetrahidronaftalenos/farmacologíaRESUMEN
It is possible that parasites may influence the course of COVID-19 infection, as either risk factors or protective agents; as such, the current coronavirus pandemic may affect the diagnosis and prevention of parasitic disease, and its elimination programs. The present review highlights the similarity between the symptoms of human parasitoses and those of COVID-19 and discuss their mutual influence. The study evaluated selected human parasitoses with similar symptoms to COVID-19 and examined their potential influence on SARS-CoV-2 virus invasion. The available data suggest that at least several human parasitoses could result in misdiagnosis of COVID-19. Some disorders, such as malaria, schistosomiasis and soil-transmitted helminths, can increase the risk of severe infection with COVID-19. It is also suggested that recovery from parasitic disease can enhance the immune system and protect from COVID-19 infection. In addition, the COVID-19 pandemic has affected parasitic disease elimination programs in endemic regions and influenced the number of diagnoses of human parasitoses.
RESUMEN
Proinflammatory activation of macrophages in metabolic tissues is critically important in the induction of obesity-induced metaflammation. Here, we demonstrate that the soluble mannose receptor (sMR) plays a direct functional role in both macrophage activation and metaflammation. We show that sMR binds CD45 on macrophages and inhibits its phosphatase activity, leading to an Src/Akt/NF-κB-mediated cellular reprogramming toward an inflammatory phenotype both in vitro and in vivo. Remarkably, increased serum sMR levels were observed in obese mice and humans and directly correlated with body weight. Importantly, enhanced sMR levels increase serum proinflammatory cytokines, activate tissue macrophages, and promote insulin resistance. Altogether, our results reveal sMR as regulator of proinflammatory macrophage activation, which could constitute a therapeutic target for metaflammation and other hyperinflammatory diseases.
Asunto(s)
Regulación de la Expresión Génica/efectos de los fármacos , Activación de Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Receptor de Manosa/química , Proteínas de la Membrana/farmacología , Alimentación Animal , Animales , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Dieta Alta en Grasa , Microbioma Gastrointestinal , Inflamación , Activación de Macrófagos/fisiología , Masculino , Receptor de Manosa/metabolismo , Ratones , Ratones Noqueados , Distribución AleatoriaRESUMEN
BACKGROUND/OBJECTIVES: The worldwide prevalence of obesity, metabolic syndrome and type 2 diabetes (T2D) is reaching epidemic proportions that urge the development of new management strategies. Totum-63 is a novel, plant-based polyphenol-rich active principle that has been shown to reduce body weight, fasting glycemia, glucose intolerance, and fatty liver index in obese subjects with prediabetes. Here, we investigated the effects and underlying mechanism(s) of Totum-63 on metabolic homeostasis in insulin-resistant obese mice. METHODS: Male C57Bl6/J mice were fed a high-fat diet for 12 weeks followed by supplementation with Totum-63 for 4 weeks. The effects on whole-body energy and metabolic homeostasis, as well as on tissue-specific inflammation and insulin sensitivity were assessed using a variety of immunometabolic phenotyping tools. RESULTS: Totum-63 decreased body weight and fat mass in obese mice, without affecting lean mass, food intake and locomotor activity, and increased fecal energy excretion and whole-body fatty acid oxidation. Totum-63 reduced fasting plasma glucose, insulin and leptin levels, and improved whole-body insulin sensitivity and peripheral glucose uptake. The expression of insulin receptor ß and the insulin-induced phosphorylation of Akt/PKB were increased in liver, skeletal muscle, white adipose tissue (WAT) and brown adipose tissue (BAT). Hepatic steatosis was also decreased by Totum-63 and associated with a lower expression of genes involved in fatty acid uptake, de novo lipogenesis, inflammation, and fibrosis. Furthermore, a significant reduction in pro-inflammatory macrophages was also observed in epidydimal WAT. Finally, a potent decrease in BAT mass associated with enhanced tissue expression of thermogenic genes was found, suggesting BAT activation by Totum-63. CONCLUSIONS: Our results show that Totum-63 reduces inflammation and improves insulin sensitivity and glucose homeostasis in obese mice through pleiotropic effects on various metabolic organs. Altogether, plant-derived Totum-63 might constitute a promising novel nutritional supplement for alleviating metabolic dysfunctions in obese people with or without T2D.
Asunto(s)
Composición Corporal/efectos de los fármacos , Inflamación/tratamiento farmacológico , Obesidad/tratamiento farmacológico , Extractos Vegetales/farmacología , Polifenoles/farmacología , Animales , Composición Corporal/fisiología , Modelos Animales de Enfermedad , Inflamación/prevención & control , Resistencia a la Insulina/fisiología , Ratones , Ratones Endogámicos C57BL/metabolismoRESUMEN
Macrophages are highly plastic, key regulators of inflammation. Deregulation of macrophage activation can lead to excessive inflammation as seen in inflammatory disorders like atherosclerosis, obesity, multiple sclerosis and sepsis. Targeting intracellular metabolism is considered as an approach to reshape deranged macrophage activation and to dampen the progression of inflammatory disorders. ATP citrate lyase (Acly) is a key metabolic enzyme and an important regulator of macrophage activation. Using a macrophage-specific Acly-deficient mouse model, we investigated the role of Acly in macrophages during acute and chronic inflammatory disorders. First, we performed RNA sequencing to demonstrate that Acly-deficient macrophages showed hyperinflammatory gene signatures in response to acute LPS stimulation in vitro. Next, we assessed endotoxin-induced peritonitis in myeloid-specific Acly-deficient mice and show that, apart from increased splenic Il6 expression, systemic and local inflammation were not affected by Acly deficiency. Also during obesity, both chronic low-grade inflammation and whole-body metabolic homeostasis remained largely unaltered in mice with Acly-deficient myeloid cells. Lastly, we show that macrophage-specific Acly deletion did not affect the severity of experimental autoimmune encephalomyelitis (EAE), an experimental model of multiple sclerosis. These results indicate that, despite increasing inflammatory responses in vitro, macrophage Acly deficiency does not worsen acute and chronic inflammatory responses in vivo. Collectively, our results indicate that caution is warranted in prospective long-term treatments of inflammatory disorders with macrophage-specific Acly inhibitors. Together with our earlier observation that myeloid Acly deletion stabilizes atherosclerotic lesions, our findings highlight that therapeutic targeting of macrophage Acly can be beneficial in some, but not all, inflammatory disorders.
Asunto(s)
ATP Citrato (pro-S)-Liasa/metabolismo , Encefalomielitis Autoinmune Experimental/enzimología , Inflamación/enzimología , Macrófagos/enzimología , Peritonitis/enzimología , ATP Citrato (pro-S)-Liasa/genética , Animales , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Dieta Alta en Grasa , Encefalomielitis Autoinmune Experimental/inducido químicamente , Encefalomielitis Autoinmune Experimental/genética , Encefalomielitis Autoinmune Experimental/inmunología , Inflamación/etiología , Inflamación/genética , Inflamación/inmunología , Mediadores de Inflamación/metabolismo , Lipopolisacáridos , Macrófagos/inmunología , Ratones Endogámicos C57BL , Ratones Noqueados , Glicoproteína Mielina-Oligodendrócito , Obesidad/complicaciones , Fragmentos de Péptidos , Peritonitis/inducido químicamente , Peritonitis/genética , Peritonitis/inmunología , Fenotipo , Transducción de SeñalRESUMEN
Dirofilariarepens is a parasitic nematode causing a vector-borne zoonotic infection (dirofilariosis), considered an emerging problem in human and veterinary medicine. Currently, diagnosis is based on the detection of the adult parasite and microfilariae in the host tissues. However, the efficacy of tests relying on microfilariae detection is limited by microfilariae periodic occurrence. Therefore, a new reliable and affordable serological diagnostic method is needed. Better characteristic of the parasite biology and its interaction with host immune system should help to achieve this goal. This study analyzes adult and microfilariae proteomes, and the use of one-dimensional electrophoresis (1-DE) and two-dimensional electrophoresis (2-DE) proteomics, immunoproteomics, and LC-MS/MS mass spectrometry allowed us to identify 316 potentially immunogenic proteins (75 belong to adult stage, 183 to microfilariae, and 58 are common for both). Classified by their ontology, the proteins showed important similarities and differences between both parasite stages. The most frequently identified proteins are structural, metabolic, and heat shock proteins. Additionally, real-time PCR analysis of some immunogenic targets revealed significant differences between microfilariae and adult life stages. We indicated molecules involved in parasite-host interactions and discussed their importance in parasite biology, which may help to reveal potential diagnostic antigens or select drug and vaccine targets.
RESUMEN
Trichinella spiralis is a foodborne zoonotic nematode, which causes trichinellosis. During the infection, parasite evades the host immune responses by direct and indirect (through excretory-secretory products) contact with host immune cells. One of the main targets for immunomodulation induced by helminths are macrophages. In this study, we examined whether direct contact of different stages of T. spiralis can affect the polarization of human THP-1 macrophages. Co-culture of adult parasite stage and cells in direct contact without LPS addition had a significant impact on TNFα levels. Interestingly, in settings with the addition of LPS, the levels of IL-1ß and TNFα significantly increased in adult parasite and newborn larvae (NBL) but not for muscle larvae (ML). While we tested muscle larvae ESP products to compare its effect with whole ML parasite, we detect an increase of pro-inflammatory cytokines like IL-1ß and TNFα in no LPS conditions. Whereas, muscle larvae ESP significantly suppressed the inflammatory response measured by IL-1ß, TNFα, and IL-6 levels and anti-inflammatory IL-10 compared to LPS control. Our findings indicate the anti-inflammatory potential of T. spiralis muscle larvae excretory-secretory products and propose signaling pathways which might be engaged in the mechanism of how muscle larvae ESP affect human macrophages.
Asunto(s)
Citocinas/inmunología , Interacciones Huésped-Parásitos , Inmunomodulación , Activación de Macrófagos , Trichinella spiralis/inmunología , Triquinelosis/inmunología , Animales , Antígenos Helmínticos/inmunología , Femenino , Humanos , Recién Nacido , Larva/inmunología , Macrófagos/inmunología , Macrófagos/parasitología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Músculos/parasitología , Transducción de Señal , Células THP-1 , Trichinella spiralis/fisiología , Triquinelosis/parasitologíaRESUMEN
Type 2 immunity plays an essential role in the maintenance of metabolic homeostasis and its disruption during obesity promotes meta-inflammation and insulin resistance. Infection with the helminth parasite Schistosoma mansoni and treatment with its soluble egg antigens (SEA) induce a type 2 immune response in metabolic organs and improve insulin sensitivity and glucose tolerance in obese mice, yet, a causal relationship remains unproven. Here, we investigated the effects and underlying mechanisms of the T2 ribonuclease omega-1 (ω1), one of the major S mansoni immunomodulatory glycoproteins, on metabolic homeostasis. We show that treatment of obese mice with plant-produced recombinant ω1, harboring similar glycan motifs as present on the native molecule, decreased body fat mass, and improved systemic insulin sensitivity and glucose tolerance in a time- and dose-dependent manner. This effect was associated with an increase in white adipose tissue (WAT) type 2 T helper cells, eosinophils, and alternatively activated macrophages, without affecting type 2 innate lymphoid cells. In contrast to SEA, the metabolic effects of ω1 were still observed in obese STAT6-deficient mice with impaired type 2 immunity, indicating that its metabolic effects are independent of the type 2 immune response. Instead, we found that ω1 inhibited food intake, without affecting locomotor activity, WAT thermogenic capacity or whole-body energy expenditure, an effect also occurring in leptin receptor-deficient obese and hyperphagic db/db mice. Altogether, we demonstrate that while the helminth glycoprotein ω1 can induce type 2 immunity, it improves whole-body metabolic homeostasis in obese mice by inhibiting food intake via a STAT6-independent mechanism.
Asunto(s)
Ingestión de Alimentos , Endorribonucleasas/uso terapéutico , Glicoproteínas/uso terapéutico , Proteínas del Helminto/uso terapéutico , Obesidad/tratamiento farmacológico , Tejido Adiposo/efectos de los fármacos , Tejido Adiposo/metabolismo , Animales , Células Cultivadas , Endorribonucleasas/farmacología , Glicoproteínas/farmacología , Proteínas del Helminto/farmacología , Locomoción , Macrófagos/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas Recombinantes/farmacología , Proteínas Recombinantes/uso terapéutico , Schistosoma mansoni/enzimología , Linfocitos T Colaboradores-Inductores/efectos de los fármacos , Termogénesis , Nicotiana/genética , Nicotiana/metabolismoRESUMEN
14-3-3 proteins are present in all eukaryotic organisms and are ubiquitously expressed in a broad range of tissues and cellular compartments. They are regulatory adapter proteins that play key roles in a variety of signaling pathways, and have been proposed as suitable targets for the control and detection of certain parasites. Trichinella britovi is a widely-distributed parasitic nematode, transmitted through ingestion of meat products containing invasive larvae. The present study describes the cloning and expression of Tb14-3-3, and investigates the immunological and protective potential of the recombinant protein. Immunization of mice with rTb14-3-3 triggered an IgG response, and significant differences, in the profiles of secreted cytokines observed in vitro, between experimental groups. Nonetheless, neither specific antibodies, nor increased secretion of IFNγ, IL-4, and IL-10 cytokines, conferred greater protection against infection. No reduction in larval burden was observed during recovery at 48 dpi. Additionally, rTb14-3-3 was not recognized by sera from the infected control mice, except for one, suggesting some mismatch between native and recombinant Tb14-3-3 antigenic sites. Therefore, before 14-3-3 can be considered a potential tool for Trichinella detection and vaccination, more research regarding its target proteins, and actual specific function, is needed.
RESUMEN
Trichinellosis is a globally-distributed zoonotic parasitic disease caused by nematode worms of the genus Trichinella. One of the most common species of Trichinella known to affect human health is T. britovi; however, it is relatively poorly investigated. A thorough knowledge of the proteins expressed by Trichinella is important when developing immunological detection methods and vaccines and studying its interactions with the host. The present study uses the Pichia pastoris expression system to produce a soluble TbCLP antigen which induces strong antibody responses in the host during natural infection. Our results demonstrate the feasibility of TbCLP antigen production in yeasts, which are able to carry out post-translational modifications such as glycosylation and disulfide bond formation; they also indicate that the glycosylated TbCLP antigen had immunogenic effects in the tested mice and induced a mixed Th1/Th2 response, and was associated with a reduced larval burden after challenge with T. britovi. Subsequent in vitro stimulation of mice splenocytes revealed that TbCLP most likely possesses immunomodulatory properties and may play a significant role in the early phase of infection, affecting host immunological responses.