The incidences of multiple sclerosis have risen worldwide, yet neither the trigger nor efficient treatment is known. Some research is dedicated to looking for treatment by parasites, mainly by helminths. However, little is known about the effect of helminths that infect the nervous system. Therefore, we chose the neurotropic avian schistosome Trichobilharzia regenti, which strongly promotes M2 polarization and tissue repair in the central nervous system, and we tested its effect on the course of experimental autoimmune encephalomyelitis (EAE) in mice. Surprisingly, the symptoms of EAE tended to worsen after the infection with T. regenti. The infection did not stimulate tissue repair, as indicated by the similar level of demyelination. Eosinophils heavily infiltrated the infected tissue, and the microglia number increased as well. Furthermore, splenocytes from T. regenti-infected EAE mice produced more interferon (IFN)-γ than splenocytes from EAE mice after stimulation with myelin oligodendrocyte glycoprotein. Our research indicates that the combination of increased eosinophil numbers and production of IFN-γ tends to worsen the EAE symptoms. Moreover, the data highlight the importance of considering the direct effect of the parasite on the tissue, as the migrating parasite may further tissue damage and make tissue repair even more difficult.

• Klíčová slova
• EAE, IFN-γ, Trichobilharzia regenti, demyelination, eosinophilia, experimental autoimmune encephalomyelitis, neurotropic parasite, neurotropic schistosome,
• MeSH
• encefalomyelitida autoimunitní experimentální * imunologie   patologie   MeSH
• eozinofily imunologie   MeSH
• interferon gama * metabolismus   MeSH
• myši inbrední C57BL * MeSH
• myši MeSH
• roztroušená skleróza imunologie   patologie   MeSH
• Schistosomatidae fyziologie   MeSH
• slezina patologie   parazitologie   imunologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• interferon gama * MeSH

BACKGROUND: Avian schistosomes, the causative agents of human cercarial dermatitis (or swimmer's itch), die in mammals but the mechanisms responsible for parasite elimination are unknown. Here we examined the role of reactive nitrogen species, nitric oxide (NO) and peroxynitrite, in the immune response of mice experimentally infected with Trichobilharzia regenti, a model species of avian schistosomes remarkable for its neuropathogenicity. METHODS: Inducible NO synthase (iNOS) was localized by immunohistochemistry in the skin and the spinal cord of mice infected by T. regenti. The impact of iNOS inhibition by aminoguanidine on parasite burden and growth was then evaluated in vivo. The vulnerability of T. regenti schistosomula to NO and peroxynitrite was assessed in vitro by viability assays and electron microscopy. Additionally, the effect of NO on the activity of T. regenti peptidases was tested using a fluorogenic substrate. RESULTS: iNOS was detected around the parasites in the epidermis 8 h post-infection and also in the spinal cord 3 days post-infection (dpi). Inhibition of iNOS resulted in slower parasite growth 3 dpi, but the opposite effect was observed 7 dpi. At the latter time point, moderately increased parasite burden was also noticed in the spinal cord. In vitro, NO did not impair the parasites, but inhibited the activity of T. regenti cathepsins B1.1 and B2, the peptidases essential for parasite migration and digestion. Peroxynitrite severely damaged the surface tegument of the parasites and decreased their viability in vitro, but rather did not participate in parasite clearance in vivo. CONCLUSIONS: Reactive nitrogen species, specifically NO, do not directly kill T. regenti in mice. NO promotes the parasite growth soon after penetration (3 dpi), but prevents it later (7 dpi) when also suspends the parasite migration in the CNS. NO-related disruption of the parasite proteolytic machinery is partly responsible for this effect.

• Klíčová slova
• 3-Nitrotyrosine, Cathepsin B, Nitric oxide, Nitric oxide synthase, Peroxynitrite, Schistosomatidae, Trichobilharzia,
• MeSH
• centrální nervový systém parazitologie   MeSH
• guanidiny farmakologie   MeSH
• infekce červy třídy Trematoda farmakoterapie   MeSH
• kůže parazitologie   MeSH
• kyselina peroxydusitá farmakologie   MeSH
• lidé MeSH
• mícha parazitologie   MeSH
• myši MeSH
• oxid dusnatý farmakologie   MeSH
• proteasy účinky léků   metabolismus   MeSH
• proteiny červů účinky léků   metabolismus   MeSH
• ptáci parazitologie   MeSH
• Schistosoma účinky léků   růst a vývoj   patogenita   MeSH
• Schistosomatidae účinky léků   růst a vývoj   patogenita   MeSH
• schistosomóza farmakoterapie   MeSH
• synthasa oxidu dusnatého účinky léků   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• guanidiny MeSH
• kyselina peroxydusitá MeSH
• oxid dusnatý MeSH
• pimagedine MeSH Prohlížeč
• proteasy MeSH
• proteiny červů MeSH
• synthasa oxidu dusnatého MeSH

Schistosomula (the post-infective stages) of the neurotropic schistosome Trichobilharzia regenti possess multiple isoforms of cathepsin B1 peptidase (TrCB1.1-TrCB1.6) with involvement in nutrient digestion. The comparison of substrate preferences of TrCB1.1 and TrCB1.4 showed that TrCB1.4 had a very narrow substrate specificity and after processing it was less effective toward protein substrates when compared to TrCB1.1. Self-processing of both isoforms could be facilitated by sulfated polysaccharides due to a specific binding motif in the pro-sequence. Trans-activation by heterologous enzymes was also successfully employed. Expression profiling revealed a high level of transcription of genes encoding the enzymatically inactive paralogs TrCB1.5 and TrCB1.6. The transcription level of TrCB1.6 was comparable with that of TrCB1.1 and TrCB1.2, the most abundant active isoforms. Recombinant TrCB1.6wt, a wild type paralog with a Cys29-to-Gly substitution in the active site that renders the enzyme inactive, was processed by the active TrCB1 forms and by an asparaginyl endopeptidase. Although TrCB1.6wt lacked hydrolytic activity, endopeptidase, but not dipeptidase, activity could be restored by mutating Gly29 to Cys29. The lack of exopeptidase activity may be due to other mutations, such as His110-to-Asn in the occluding loop and Asp224-to-Gly in the main body of the mature TrCB1.6, which do not occur in the active isoforms TrCB1.1 and TrCB1.4 with exopeptidase activity. The catalytically active enzymes and the inactive TrCB1.6 paralog formed complexes with chicken cystatin, thus supporting experimentally the hypothesis that inactive paralogs could potentially regulate the activity of the active forms or protect them from being inhibited by host inhibitors. The effect on cell viability and nitric oxide production by selected immune cells observed for TrCB1.1 was not confirmed for TrCB1.6. We show here that the active isoforms of TrCB1 have different affinities for peptide substrates thereby facilitating diversity in protein-derived nutrition for the parasite. The inactive paralogs are unexpectedly highly expressed and one of them retains the ability to bind cystatins, likely due to specific mutations in the occluding loop and the enzyme body. This suggests a role in sequestration of inhibitors and protection of active cysteine peptidases.

• Klíčová slova
• cathepsin B, cystatin, helminth, occluding loop, peptidase, processing, schistosome, substrate specificity,
• MeSH
• astrocyty metabolismus   MeSH
• cystatiny metabolismus   MeSH
• hydrolýza MeSH
• izoenzymy metabolismus   MeSH
• kathepsin B chemie   genetika   metabolismus   MeSH
• makrofágy metabolismus   MeSH
• myši MeSH
• oxid dusnatý metabolismus   MeSH
• prekurzory enzymů metabolismus   MeSH
• proteolýza MeSH
• RAW 264.7 buňky MeSH
• rekombinantní proteiny metabolismus   MeSH
• Schistosomatidae enzymologie   patogenita   MeSH
• substituce aminokyselin MeSH
• substrátová specifita MeSH
• vazba proteinů MeSH
• viabilita buněk MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• cystatin, egg-white MeSH Prohlížeč
• cystatiny MeSH
• izoenzymy MeSH
• kathepsin B MeSH
• oxid dusnatý MeSH
• prekurzory enzymů MeSH
• rekombinantní proteiny MeSH

Trichobilharzia species are parasitic flatworms (called schistosomes or flukes) that cause important diseases in birds and humans, but very little is known about their molecular biology. Here, using a transcriptomics-bioinformatics-based approach, we explored molecular aspects pertaining to the nutritional requirements of Trichobilharzia szidati ('visceral fluke') and T. regenti ('neurotropic fluke') in their avian host. We studied the larvae of each species before they enter (cercariae) and as they migrate (schistosomules) through distinct tissues in their avian (duck) host. Cercariae of both species were enriched for pathways or molecules associated predominantly with carbohydrate metabolism, oxidative phosphorylation and translation of proteins linked to ribosome biogenesis, exosome production and/or lipid biogenesis. Schistosomules of both species were enriched for pathways or molecules associated with processes including signal transduction, cell turnover and motility, DNA replication and repair, molecular transport and/or catabolism. Comparative informatic analyses identified molecular repertoires (within, e.g., peptidases and secretory proteins) in schistosomules that can broadly degrade macromolecules in both T. szidati and T. regenti, and others that are tailored to each species to selectively acquire nutrients from particular tissues through which it migrates. Thus, this study provides molecular evidence for distinct modes of nutrient acquisition between the visceral and neurotropic flukes of birds.

• MeSH
• cerkárie klasifikace   genetika   patogenita   MeSH
• DNA helmintů klasifikace   genetika   MeSH
• fylogeneze * MeSH
• kachny genetika   parazitologie   MeSH
• lidé MeSH
• nemoci ptáků genetika   parazitologie   MeSH
• ptáci genetika   parazitologie   MeSH
• Schistosomatidae genetika   patogenita   MeSH
• schistosomóza genetika   parazitologie   MeSH
• Trematoda klasifikace   genetika   patogenita   MeSH
• výpočetní biologie MeSH
• živiny MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA helmintů MeSH

Cercarial dermatitis (CD) is an allergic skin disease that rises in consequence of infection by invasive stages (cercariae) of trematodes of the family Schistosomatidae. CD has been considered a re-emerging disease, human cases have been reported from all continents, and tourism-threatening outbreaks occur even in frequented recreational areas. Although the symptoms of CD are generally known, the data on immune response in human patients are sporadic and incomprehensive. In the present study, we attempted to correlate the symptoms, personal history, and time course of CD in human patients with differential cell counts, dynamics of selected cytokines, and dynamics and quality of antibody response. By a systematic follow-up, we obtained a uniquely complex dataset from ten persons accidentally and concurrently infected by the same parasite species in the same locality. The onset of CD was significantly faster, and the symptoms were heavier in participants with a history of CD if compared to naive ones, who, however, also developed some of the symptoms. The repeatedly infected persons had elevated proportion of eosinophils 1 week post exposure (p.e.) and a stronger specific IgG but not IgM response, whereas specific IgE response was not observed. Increased serum levels of IL-4 occurred 1 and 3 week(s) p.e. in all participants. There was high variability in individual immunoblot patterns of IgG response, and no antigen with a universal diagnostic potential was confirmed. The presented analyses suggested that a complex approach can improve the accuracy of the diagnosis of CD, but component data should be interpreted carefully.

• Klíčová slova
• Allergy, Diagnosis, Immunity, Schistosome, Skin, Trichobilharzia,
• MeSH
• dermatitida imunologie   parazitologie   MeSH
• dospělí MeSH
• epidemický výskyt choroby MeSH
• imunoglobulin E krev   MeSH
• imunoglobulin G krev   MeSH
• imunoglobulin M krev   MeSH
• infekce červy třídy Trematoda diagnóza   imunologie   parazitologie   MeSH
• interleukin-4 krev   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• následné studie MeSH
• protilátky protozoální krev   MeSH
• průzkumy a dotazníky MeSH
• rybníky parazitologie   MeSH
• Schistosomatidae imunologie   MeSH
• zvířata MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• IL4 protein, human MeSH Prohlížeč
• imunoglobulin E MeSH
• imunoglobulin G MeSH
• imunoglobulin M MeSH
• interleukin-4 MeSH
• protilátky protozoální MeSH

BACKGROUND: Helminth neuroinfections represent a serious health problem, but host immune mechanisms in the nervous tissue often remain undiscovered. This study aims at in vitro characterization of the response of murine astrocytes and microglia exposed to Trichobilharzia regenti which is a neuropathogenic schistosome migrating through the central nervous system of vertebrate hosts. Trichobilharzia regenti infects birds and mammals in which it may cause severe neuromotor impairment. This study was focused on astrocytes and microglia as these are immunocompetent cells of the nervous tissue and their activation was recently observed in T. regenti-infected mice. RESULTS: Primary astrocytes and microglia were exposed to several stimulants of T. regenti origin. Living schistosomulum-like stages caused increased secretion of IL-6 in astrocyte cultures, but no changes in nitric oxide (NO) production were noticed. Nevertheless, elevated parasite mortality was observed in these cultures. Soluble fraction of the homogenate from schistosomulum-like stages stimulated NO production by both astrocytes and microglia, and IL-6 and TNF-α secretion in astrocyte cultures. Similarly, recombinant cathepsins B1.1 and B2 triggered IL-6 and TNF-α release in astrocyte and microglia cultures, and NO production in astrocyte cultures. Stimulants had no effect on production of anti-inflammatory cytokines IL-10 or TGF-β1. CONCLUSIONS: Both astrocytes and microglia are capable of production of NO and proinflammatory cytokines IL-6 and TNF-α following in vitro exposure to various stimulants of T. regenti origin. Astrocytes might be involved in triggering the tissue inflammation in the early phase of T. regenti infection and are proposed to participate in destruction of migrating schistosomula. However, NO is not the major factor responsible for parasite damage. Both astrocytes and microglia can be responsible for the nervous tissue pathology and maintaining the ongoing inflammation since they are a source of NO and proinflammatory cytokines which are released after exposure to parasite antigens.

• Klíčová slova
• Anti-inflammatory cytokines, Astrocytes, Avian schistosome, Cathepsin B, Microglia, Neuroinfection, Nitric oxide, Proinflammatory cytokines, Trichobilharzia regenti,
• MeSH
• astrocyty imunologie   parazitologie   MeSH
• interleukin-6 metabolismus   MeSH
• kultivované buňky MeSH
• myši MeSH
• neuroglie imunologie   parazitologie   MeSH
• oxid dusnatý metabolismus   MeSH
• Schistosomatidae imunologie   MeSH
• TNF-alfa metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• interleukin-6, mouse MeSH Prohlížeč
• interleukin-6 MeSH
• oxid dusnatý MeSH
• TNF-alfa MeSH