Integrity of neural extracellular matrix is required for microglia-mediated synaptic remodeling.

Microglia continuously remodel synapses, which are embedded in the extracellular matrix (ECM). However, the mechanisms, which govern this process remain elusive. To investigate the influence of the neural ECM in synaptic remodeling by microglia, we disrupted ECM integrity by injection of chondroitinase ABC (ChABC) into the retrosplenial cortex of healthy adult mice. Using in vivo two-photon microscopy we found that ChABC treatment increased microglial branching complexity and ECM phagocytic capacity and decreased spine elimination rate under basal conditions. Moreover, ECM attenuation largely prevented synaptic remodeling following synaptic stress induced by photodamage of single synaptic elements. These changes were associated with less stable and smaller microglial contacts at the synaptic damage sites, diminished deposition of calreticulin and complement proteins C1q and C3 at synapses and impaired expression of microglial CR3 receptor. Thus, our findings provide novel insights into the function of the neural ECM in deposition of complement proteins and synaptic remodeling by microglia.

Real-time mechanisms of exacerbated synaptic remodeling by microglia in acute models of systemic inflammation and tauopathy.

Remodeling of synapses by microglia is essential for synaptic plasticity in the brain. However, during neuroinflammation and neurodegenerative diseases, microglia can induce excessive synaptic loss, although the precise underlying mechanisms are unknown. To directly observe microglia-synapse interactions under inflammatory conditions, we performed in vivo two-photon time-lapse imaging of microglia-synapse interactions after bacterial lipopolysaccharide administration to model systemic inflammation, or after inoculation of Alzheimer's disease (AD) brain extracts to model disease-associated neuroinflammatory microglial response. Both treatments prolonged microglia-neuron contacts, decreased basal surveillance of synapses and promoted synaptic remodeling in response to synaptic stress induced by focal single-synapse photodamage. Spine elimination correlated with the expression of microglial complement system/phagocytic proteins and the occurrence of synaptic filopodia. Microglia were observed contacting spines, then stretching and phagocytosing spine head filopodia. Thus, in response to inflammatory stimuli microglia exacerbated spine remodeling through prolonged microglial contact and elimination of spines 'tagged' by synaptic filopodia.

Experimental brain infection with cysticercosis in sheep. / Infección experimental cerebral con cisticercosis en ovejas.

OBJECTIVE.: To explore the feasibility of developing a sheep model of neurocysticercosis (NCC) by intracranial infection with T. solium oncospheres. MATERIALS AND METHODS.: We carried out an experimental infection model of NCC in sheep. Approximately 10 T. solium oncospheres previously cultured for 30 days were inoculated intracranially into ten sheep. The oncospheres, in 0.1 mL of physiological saline, were injected into the parietal lobe through an 18-gauge needle. RESULTS.: After three months, granulomas were found in two sheep. In a third sheep we identified a 5 mm diameter cyst in the right lateral ventricle and histological evaluation confirmed that the cyst corresponded to a T. solium larva. Immunohistochemistry with monoclonal antibodies directed against membrane components and excretory/secretory antigens of the T. solium cyst was also used to confirm the etiology of the found granulomas. One of them showed reactivity to the monoclonal antibodies used, thus confirming that it was a cysticercus. CONCLUSION.: This experiment is the proof of concept that it is possible to infect sheep with cysticercosis by intracranial inoculation.

OBJETIVO: . Explorar la viabilidad de desarrollar un modelo de neurocisticercosis (NCC) de oveja mediante infección intracraneal de oncosferas de T. solium. MATERIALES Y MÉTODOS.: Se realizó un modelo de infección experimental de NCC en ovejas. Se inocularon aproximadamente 10 posoncósferas de T. solium cultivadas previamente por 30 días por vía intracraneal en diez ovejas. Las oncósferas, en 0,1 mL de solución salina fisiológica, se inyectaron en el lóbulo parietal a través de una aguja de calibre 18. RESULTADOS.: Después de tres meses, en dos ovejas se encontraron granulomas y en una tercera identificó un quiste de 5 mm de diámetro en el ventrículo lateral derecho y la evaluación histológica confirmó que el quiste corresponde a una larva de T. solium. También se utilizó inmunohistoquímica con anticuerpos monoclonales dirigidos contra componentes de membrana y antígenos excretorios/secretorios del quiste de T. solium para confirmar la etiología de los granulomas encontrados. Uno de ellos mostro reactividad ante los anticuerpos monoclonales utilizados, confirmando así que se trató de un cisticerco. CONCLUSIÓN.: Este experimento es la prueba de concepto de que es posible infectar ovejas con cisticercosis por inoculación intracraneal.

Infección experimental cerebral con cisticercosis en ovejas / Experimental brain infection with cysticercosis in sheep

RESUMEN Objetivo . Explorar la viabilidad de desarrollar un modelo de neurocisticercosis (NCC) de oveja mediante infección intracraneal de oncosferas de T. solium. Materiales y métodos. Se realizó un modelo de infección experimental de NCC en ovejas. Se inocularon aproximadamente 10 posoncósferas de T. solium cultivadas previamente por 30 días por vía intracraneal en diez ovejas. Las oncósferas, en 0,1 mL de solución salina fisiológica, se inyectaron en el lóbulo parietal a través de una aguja de calibre 18. Resultados. Después de tres meses, en dos ovejas se encontraron granulomas y en una tercera identificó un quiste de 5 mm de diámetro en el ventrículo lateral derecho y la evaluación histológica confirmó que el quiste corresponde a una larva de T. solium. También se utilizó inmunohistoquímica con anticuerpos monoclonales dirigidos contra componentes de membrana y antígenos excretorios/secretorios del quiste de T. solium para confirmar la etiología de los granulomas encontrados. Uno de ellos mostro reactividad ante los anticuerpos monoclonales utilizados, confirmando así que se trató de un cisticerco. Conclusión. Este experimento es la prueba de concepto de que es posible infectar ovejas con cisticercosis por inoculación intracraneal.

ABSTRACT Objective. To explore the feasibility of developing a sheep model of neurocysticercosis (NCC) by intracranial infection with T. solium oncospheres. Materials and methods. We carried out an experimental infection model of NCC in sheep. Approximately 10 T. solium oncospheres previously cultured for 30 days were inoculated intracranially into ten sheep. The oncospheres, in 0.1 mL of physiological saline, were injected into the parietal lobe through an 18-gauge needle. Results. After three months, granulomas were found in two sheep. In a third sheep we identified a 5 mm diameter cyst in the right lateral ventricle and histological evaluation confirmed that the cyst corresponded to a T. solium larva. Immunohistochemistry with monoclonal antibodies directed against membrane components and excretory/secretory antigens of the T. solium cyst was also used to confirm the etiology of the found granulomas. One of them showed reactivity to the monoclonal antibodies used, thus confirming that it was a cysticercus. Conclusion. This experiment is the proof of concept that it is possible to infect sheep with cysticercosis by intracranial inoculation.

Microglia Depletion-Induced Remodeling of Extracellular Matrix and Excitatory Synapses in the Hippocampus of Adult Mice.

The extracellular matrix (ECM) plays a key role in synaptogenesis and the regulation of synaptic functions in the central nervous system. Recent studies revealed that in addition to dopaminergic and serotoninergic neuromodulatory systems, microglia also contribute to the regulation of ECM remodeling. In the present work, we investigated the physiological role of microglia in the remodeling of perineuronal nets (PNNs), predominantly associated with parvalbumin-immunopositive (PV+) interneurons, and the perisynaptic ECM around pyramidal neurons in the hippocampus. Adult mice were treated with PLX3397 (pexidartinib), as the inhibitor of colony-stimulating factor 1 receptor (CSF1-R), to deplete microglia. Then, confocal analysis of the ECM and synapses was performed. Although the elimination of microglia did not alter the overall number or intensity of PNNs in the CA1 region of the hippocampus, it decreased the size of PNN holes and elevated the expression of the surrounding ECM. In the neuropil area in the CA1 str. radiatum, the depletion of microglia increased the expression of perisynaptic ECM proteoglycan brevican, which was accompanied by the elevated expression of presynaptic marker vGluT1 and the increased density of dendritic spines. Thus, microglia regulate the homeostasis of pre- and postsynaptic excitatory terminals and the surrounding perisynaptic ECM as well as the fine structure of PNNs enveloping perisomatic-predominantly GABAergic-synapses.

Attenuation of the extracellular matrix restores microglial activity during the early stage of amyloidosis.

In the advanced stages of Alzheimer's disease (AD), microglia are transformed to an activated phenotype with thickened and retracted processes, migrate to the site of amyloid-beta (Aß) plaques, and proliferate. In the early stages of AD, it is still poorly understood whether the microglial function is altered and which factors may regulate these changes. Here, we focused on studying microglia in the retrosplenial cortex (RSC) in 3- to 4-month-old 5xFAD mice as a transgenic mouse model of AD. At this age, there are neither Aß plaques, nor activation of microglia, nor dysregulation in the expression of genes encoding major extracellular matrix (ECM) molecules or extracellular proteases in the RSC. Still, histochemical evaluation of the fine structure of neural ECM revealed increased levels of Wisteria floribunda agglutinin labeling in holes of perineuronal nets and changes in the perimeter of ECM barriers around the holes in 5xFAD mice. Two-photon vital microscopy demonstrated normal morphology and resting motility of microglia but strongly diminished number of microglial cells that migrated to the photolesion site in 5xFAD mice. Enzymatic digestion of ECM by chondroitinase ABC (ChABC) ameliorated this defect. Accordingly, the characterization of cell surface markers by flow cytometry demonstrated altered expression of microglial CD45. Moreover, ChABC treatment reduced the invasion of myeloid-derived mononuclear cells into the RSC of 5xFAD mice. Hence, the migration of both microglia and myeloid cells is altered during the early stages of amyloidosis and can be restored at least partially by the attenuation of the ECM.

Light-induced engagement of microglia to focally remodel synapses in the adult brain.

Microglia continuously monitor synapses, but active synaptic remodeling by microglia in mature healthy brains is rarely directly observed. We performed targeted photoablation of single synapses in mature transgenic mice expressing fluorescent labels in neurons and microglia. The photodamage focally increased the duration of microglia-neuron contacts, and dramatically exacerbated both the turnover of dendritic spines and presynaptic boutons as well as the generation of new filopodia originating from spine heads or boutons. The results of microglia depletion confirmed that elevated spine turnover and the generation of presynaptic filopodia are microglia-dependent processes.

TNF-α blockade suppresses pericystic inflammation following anthelmintic treatment in porcine neurocysticercosis.

BACKGROUND: Neurocysticercosis (NCC) is an infection of the brain with the larval cyst of the tapeworm, Taenia solium. Cysticidal treatment induces parasite killing resulting in a post inflammatory response and seizures, which generally requires corticosteroid treatment to control inflammation. The nature of this response and how to best control it is unclear. We investigated the anti-inflammatory effects of pretreatment with etanercept (ETN), an anti-tumor necrosis factor agent, or dexamethasone (DEX), a high potency corticosteroid, on the post treatment inflammatory response in naturally infected pigs with neurocysticercosis after a single dose of the cysticidal drug praziquantel (PZQ). METHODOLOGY/PRINCIPAL FINDINGS: We followed the methods from a previously developed treatment model of NCC in naturally infected swine. The four study groups of infected pigs included 3 groups treated with PZQ on day 0: PZQ-treated alone (100 mg/kg PO; n = 9), pretreated with dexamethasone (DEX, 0.2 mg/kg IM administered on days -1, +1 and +3; n = 6), and pretreated with etanercept (ETN, 25 mg IM per animal on days -7 and 0; n = 6). The fourth group remained untreated (n = 3). As measured by quantitative RT-PCR, ETN pretreatment depressed transcription of a wide range of proinflammatory, regulatory and matrix protease encoding genes at 120 hr post PZQ treatment in capsules of cysts that demonstrated extravasated Evans Blue (EB) (a measure of blood brain barrier dysfunction) compared to animals not receiving ETN. Transcription was significantly depressed for the proinflammatory genes tumor necrosis factor (TNF)-α, and interferon (IFN)-γ; the inflammation regulating genes cytotoxic T-lymphocyte-associated protein (CTLA)4, interleukin (IL)-13 and transforming growth factor (TGF)-ß; the tissue remodeling genes matrix metalloprotease (MMP)1 and 9, tissue inhibitors of metalloproteases (TIMP)1 and 2, and the genes regulating endothelial function vascular endothelial growth factor (VEGF)1, angiopoietin (Ang)1, Ang 2, and platelet endothelial cell adhesion molecule (PECAM)-1. In contrast, transcription was only modestly decreased in the DEX pretreated pigs compared to PZQ alone, and only for TNF-α, IL-6, IFN-γ, TGF-ß and Ang1. IL-10 was not affected by either ETN or DEX pretreatments. The degree of inflammation, assessed by semi-quantitative inflammatory scores, was modestly decreased in both ETN and DEX pretreated animals compared to PZQ treated pigs whereas cyst damage scores were moderately decreased only in cysts from DEX pretreated pigs. However, the proportion of cysts with EB extravasation was not significantly changed in ETN and DEX pretreated groups. CONCLUSIONS/SIGNIFICANCE: Overall, TNF-α blockade using ETN treatment modulated expression of a large variety of genes that play a role in induction and control of inflammation and structural changes. In contrast the number of inflammatory cells was only moderately decreased suggesting weaker effects on cell migration into the inflammatory capsules surrounding cysts than on release of modulatory molecules. Taken together, these data suggest that TNF-α blockade may provide a viable strategy to manage post-treatment pericystic inflammation that follows antiparasitic therapy for neurocysticercosis.

Radiological evolution of porcine neurocysticercosis after combined antiparasitic treatment with praziquantel and albendazole.

BACKGROUND: The onset of anthelmintic treatment of neurocysticercosis (NCC) provokes an acute immune response of the host, which in human cases is associated with exacerbation of neurological symptoms. This inflammation can occur at the first days of therapy. So, changes in the brain cysts appearance may be detected by medical imaging. We evaluated radiological changes in the appearance of brain cysts (enhancement and size) on days two and five after the onset of antiparasitic treatment using naturally infected pigs as a model for human NCC. METHODS AND RESULTS: Contrast T1-weighted magnetic resonance imaging with gadolinium was performed before and after antiparasitic treatment. Eight NCC-infected pigs were treated with praziquantel plus albendazole and euthanized two (n = 4) and five (n = 4) days after treatment; another group of four infected pigs served as untreated controls. For each lesion, gadolinium enhancement intensity (GEI) and cyst volume were measured at baseline and after antiparasitic treatment. Volume and GEI quantification ratios (post/pre-treatment measures) were used to appraise the effect of treatment. Cysts from untreated pigs showed little variations between their basal and post treatment measures. At days 2 and 5 there were significant increases in GEI ratio compared with the untreated group (1.32 and 1.47 vs 1.01, p = 0.021 and p = 0.021). Cyst volume ratios were significantly lower at days 2 and 5 compared with the untreated group (0.60 and 0.22 vs 0.95, p = 0.04 and p = 0.02). Cysts with lower cyst volume ratios showed more marked post-treatment inflammation, loss of vesicular fluid and cyst wall wrinkling. CONCLUSION/SIGNIFICANCE: A significant and drastic reduction of cyst size and increased pericystic enhancement occur in the initial days after antiparasitic treatment as an effect of acute perilesional immune response. These significant changes showed that early anthelmintic efficacy (day two) can be detected using magnetic resonance imaging.

Mathematical algorithm for the automatic recognition of intestinal parasites.

Parasitic infections are generally diagnosed by professionals trained to recognize the morphological characteristics of the eggs in microscopic images of fecal smears. However, this laboratory diagnosis requires medical specialists which are lacking in many of the areas where these infections are most prevalent. In response to this public health issue, we developed a software based on pattern recognition analysis from microscopi digital images of fecal smears, capable of automatically recognizing and diagnosing common human intestinal parasites. To this end, we selected 229, 124, 217, and 229 objects from microscopic images of fecal smears positive for Taenia sp., Trichuris trichiura, Diphyllobothrium latum, and Fasciola hepatica, respectively. Representative photographs were selected by a parasitologist. We then implemented our algorithm in the open source program SCILAB. The algorithm processes the image by first converting to gray-scale, then applies a fourteen step filtering process, and produces a skeletonized and tri-colored image. The features extracted fall into two general categories: geometric characteristics and brightness descriptions. Individual characteristics were quantified and evaluated with a logistic regression to model their ability to correctly identify each parasite separately. Subsequently, all algorithms were evaluated for false positive cross reactivity with the other parasites studied, excepting Taenia sp. which shares very few morphological characteristics with the others. The principal result showed that our algorithm reached sensitivities between 99.10%-100% and specificities between 98.13%- 98.38% to detect each parasite separately. We did not find any cross-positivity in the algorithms for the three parasites evaluated. In conclusion, the results demonstrated the capacity of our computer algorithm to automatically recognize and diagnose Taenia sp., Trichuris trichiura, Diphyllobothrium latum, and Fasciola hepatica with a high sensitivity and specificity.

Perilesional Inflammation in Neurocysticercosis - Relationship Between Contrast-Enhanced Magnetic Resonance Imaging, Evans Blue Staining and Histopathology in the Pig Model.

BACKGROUND: Disease manifestations in neurocysticercosis (NCC) are frequently due to inflammation of degenerating Taenia solium brain cysts. Exacerbated inflammation post anthelmintic treatment is associated with leakage of the blood brain barrier (BBB) using Evans blue (EB) staining. How well EB extravasation into the brain correlates with magnetic resonance imaging (MRI) using gadolinium (Gd) enhancement as a contrast agent and pericystic inflammation was analyzed in pigs harboring brain cysts of Taenia solium. METHODOLOGY/PRINCIPAL FINDINGS: Three groups of 4 naturally infected pigs were assessed. The first and second groups were treated with both praziquantel plus albendazole and sacrificed two and five days post treatment, respectively. A third untreated group remained untreated. Pigs were injected with EB two hours prior to evaluation by Gd-enhanced T1-MRI, and euthanized. The EB staining for each cyst capsule was scored (EB grades were 0: 0%; 1: up to 50%; 2: over 50% but less than 100%; 3: 100%). Similarly, the Gd enhancement around each cyst was qualitatively and quantitatively scored from the MRI. The extent of pericystic inflammation on histology was scored in increasing severity as IS1, IS2, IS3 and IS4. Grade 3 EB staining and enhancement was only seen in treated capsules. Also, treated groups had higher Gd intensity than the untreated group. Grades of enhancement correlated significantly with Gd enhancement intensity. EB staining was correlated with Gd enhancement intensity and with IS4 in the treated groups. These correlations were stronger in internally located cysts compared to superficial cysts in treated groups. SIGNIFICANCE: EB staining and Gd enhancement strongly correlate. The intensity of enhancement determined by MRI is a good indication of the degree of inflammation. Similarly, EB staining highly correlates with the degree of inflammation and may be applied to study inflammation in the pig model of NCC.

Inflammation Caused by Praziquantel Treatment Depends on the Location of the Taenia solium Cysticercus in Porcine Neurocysticercosis.

BACKGROUND: Neurocysticercosis (NCC), infection of the central nervous system by Taenia solium cysticerci, is a pleomorphic disease. Inflammation around cysticerci is the major cause of disease but is variably present. One factor modulating the inflammatory responses may be the location and characteristics of the brain tissue adjacent to cysticerci. We analyzed and compared the inflammatory responses to cysticerci located in the parenchyma to those in the meninges or cysticerci partially in contact with both the parenchyma and the meninges (corticomeningeal). METHODOLOGY/PRINCIPAL FINDINGS: Histological specimens of brain cysticerci (n = 196) from 11 pigs naturally infected with Taenia solium cysticerci were used. Four pigs were sacrificed after 2 days and four after 5 days of a single dose of praziquantel; 3 pigs did not receive treatment. All pigs were intravenously injected with Evans Blue to assess disruption of the blood-brain barrier. The degree of inflammation was estimated by use of a histological score (ISC) based on the extent of the inflammation in the pericystic areas as assessed in an image composed of several photomicrographs taken at 40X amplification. Parenchymal cysticerci provoked a significantly greater level of pericystic inflammation (higher ISC) after antiparasitic treatment compared to meningeal and corticomeningeal cysticerci. ISC of meningeal cysticerci was not significantly affected by treatment. In corticomeningeal cysticerci, the increase in ISC score was correlated to the extent of the cysticercus adjacent to the brain parenchyma. Disruption of the blood-brain barrier was associated with treatment only in parenchymal tissue. SIGNIFICANCE: Inflammatory response to cysticerci located in the meninges was significantly decreased compared to parenchymal cysticerci. The suboptimal inflammatory response to cysticidal drugs may be the reason subarachnoid NCC is generally refractory to treatment compared to parenchymal NCC.

Post-treatment vascular leakage and inflammatory responses around brain cysts in porcine neurocysticercosis.

Cysticidal treatment of neurocysticercosis, an infection of humans and pig brains with Taenia solium, results in an early inflammatory response directed to cysts causing seizures and focal neurological manifestations. Treatment-induced pericystic inflammation and its association with blood brain barrier (BBB) dysfunction, as determined by Evans blue (EB) extravasation, was studied in infected untreated and anthelmintic-treated pigs. We compared the magnitude and extent of the pericystic inflammation, presence of EB-stained capsules, the level of damage to the parasite, expression of genes for proinflammatory and regulatory cytokines, chemokines, and tissue remodeling by quantitative PCR assays between treated and untreated infected pigs and between EB-stained (blue) and non stained (clear) cysts. Inflammatory scores were higher in pericystic tissues from EB-stained cysts compared to clear cysts from untreated pigs and also from anthelmintic-treated pigs 48 hr and 120 hr after treatment. The degree of inflammation correlated with the severity of cyst wall damage and both increased significantly at 120 hours. Expression levels of the proinflammatory genes for IL-6, IFN-γ, TNF-α were higher in EB-stained cysts compared to clear cysts and unaffected brain tissues, and were generally highest at 120 hr. Additionally, expression of some markers of immunoregulatory activity (IL-10, IL-2Rα) were decreased in EB-stained capsules. An increase in other markers for regulatory T cells (CTLA4, FoxP3) was found, as well as significant increases in expression of two metalloproteases, MMP1 and MMP2 at 48 hr and 120 hr post-treatment. We conclude that the increase in severity of the inflammation caused by treatment is accompanied by both a proinflammatory and a complex regulatory response, largely limited to pericystic tissues with compromised vascular integrity. Because treatment induced inflammation occurs in porcine NCC similar to that in human cases, this model can be used to investigate mechanisms involved in host damaging inflammatory responses and agents or modalities that may control damaging post treatment inflammation.

Identificación de células proliferativas en quistes de Taenia solium / Identification of proliferating cells in Taenia solium cysts

Los neoblastos son células totipotentes, únicas responsables de la proliferación y maduración de tejidos en platelmintos de vida libre. Células similares se han aislado en platelmintos parásitos como Echinococcus. Taenia solium causa la teniasis humana (intestinal) y la cisticercosis en humanos y cerdos. La infección del cerebro con larvas (quistes) de T. solium resulta en neurocisticercosis, hiperendémica en el Perú; su tratamiento se asocia a síntomas neurológicos graves. La capacidad proliferativa y el desarrollo de los estadios de T. solium aún no se describen, y no se ha caracterizado los neoblastos de este parásito. Se buscó células proliferativas en quistes de T. solium colectados de un cerdo infectado, que fueron identificadas al replicarse e incorporar el nucleótido bromodesoxiuridina, detectado con un anticuerpo monoclonal. Una línea celular estable de neoblastos sería útil para estudios sistemáticos in vitro sobre eficacia de drogas y sobre la biología de T. solium.

Neoblasts are totipotent cells, solely responsible for the proliferation and maturation of tissues in free-living flatworms. Similar cells have been isolated from parasitic flatworms such as Echinococcus. Taenia solium causes human taeniasis (intestinal) and cysticercosis in humans and pigs. Brain infection with larvae (cysts) of T. solium results in neurocysticercosis which is hyperendemic in Peru, and its treatment is associated with serious neurological symptoms. The proliferative capacity and development stages of T. solium have not been described and the neoblasts of this parasite have not been characterized We looked for cell proliferation in T. solium cysts collected from an infected pig, which were identified when replicating and incorporating bromodeoxyuridine nucleotide detected with a monoclonal antibody. A stable cell line of neoblasts would be useful for systematic in vitro studies on drug efficacy and the biology of T. solium.

Evans blue staining reveals vascular leakage associated with focal areas of host-parasite interaction in brains of pigs infected with Taenia solium.

Cysticidal drug treatment of viable Taenia solium brain parenchymal cysts leads to an acute pericystic host inflammatory response and blood brain barrier breakdown (BBB), commonly resulting in seizures. Naturally infected pigs, untreated or treated one time with praziquantel were sacrificed at 48 hr and 120 hr following the injection of Evans blue (EB) to assess the effect of treatment on larval parasites and surrounding tissue. Examination of harvested non encapsulated muscle cysts unexpectedly revealed one or more small, focal round region(s) of Evans blue dye infiltration (REBI) on the surface of otherwise non dye-stained muscle cysts. Histopathological analysis of REBI revealed focal areas of eosinophil-rich inflammatory infiltrates that migrated from the capsule into the tegument and internal structures of the parasite. In addition some encapsulated brain cysts, in which the presence of REBI could not be directly assessed, showed histopathology identical to that of the REBI. Muscle cysts with REBI were more frequent in pigs that had received praziquantel (6.6% of 3736 cysts; n = 6 pigs) than in those that were untreated (0.2% of 3172 cysts; n = 2 pigs). Similar results were found in the brain, where 20.7% of 29 cysts showed histopathology identical to muscle REBI cysts in praziquantel-treated pigs compared to the 4.3% of 47 cysts in untreated pigs. Closer examination of REBI infiltrates showed that EB was taken up only by eosinophils, a major component of the cellular infiltrates, which likely explains persistence of EB in the REBI. REBI likely represent early damaging host responses to T. solium cysts and highlight the focal nature of this initial host response and the importance of eosinophils at sites of host-parasite interaction. These findings suggest new avenues for immunomodulation to reduce inflammatory side effects of anthelmintic therapy.

[Identification of proliferating cells in Taenia solium cysts]. / Identificación de células proliferativas en quistes de Taenia solium.

Neoblasts are totipotent cells, solely responsible for the proliferation and maturation of tissues in free-living flatworms. Similar cells have been isolated from parasitic flatworms such as Echinococcus. Taenia solium causes human taeniasis (intestinal) and cysticercosis in humans and pigs. Brain infection with larvae (cysts) of T. solium results in neurocysticercosis which is hyperendemic in Peru, and its treatment is associated with serious neurological symptoms. The proliferative capacity and development stages of T. solium have not been described and the neoblasts of this parasite have not been characterized We looked for cell proliferation in T. solium cysts collected from an infected pig, which were identified when replicating and incorporating bromodeoxyuridine nucleotide detected with a monoclonal antibody. A stable cell line of neoblasts would be useful for systematic in vitro studies on drug efficacy and the biology of T. solium.

Disruption of the blood-brain barrier in pigs naturally infected with Taenia solium, untreated and after anthelmintic treatment.

Neurocysticercosis is a widely prevalent disease in the tropics that causes seizures and a variety into of neurological symptoms in most of the world. Experimental models are limited and do not allow assessment of the degree of inflammation around brain cysts. The vital dye Evans Blue (EB) was injected to 11 pigs naturally infected with Taenia solium cysts to visually identify the extent of disruption of the blood-brain barrier. A total of 369 cysts were recovered from the 11 brains and classified according to the staining of their capsules as blue or unstained. The proportion of cysts with blue capsules was significantly higher in brains from pigs that had received anthelmintic treatment 48 and 120h before the EB infusion, indicating a greater compromise of the blood-brain barrier due to treatment. The model could be useful for understanding the pathology of treatment-induced inflammation in neurocysticercosis.