Peripheral blood fibrocytes: new information to explain the dynamics of Leishmania infection

Fibrocytes are important for understanding the progression of many diseases because they are present in areas where pathogenic lesions are generated. However, the morphology of fibrocytes and their interactions with parasites are poorly understood. In this study, we examined the morphology of peripheral blood fibrocytes and their interactions with Leishmania (L.) amazonensis . Through ultrastructural analysis, we describe the details of fibrocyte morphology and how fibrocytes rapidly internalise Leishmania promastigotes. The parasites differentiated into amastigotes after 2 h in phagolysosomes and the infection was completely resolved after 72 h. Early in the infection, we found increased nitric oxide production and large lysosomes with electron-dense material. These factors may regulate the proliferation and death of the parasites. Because fibrocytes are present at the infection site and are directly involved in developing cutaneous leishmaniasis, they are targets for effective, non-toxic cell-based therapies that control and treat leishmaniasis.

The protective immunity of a DNA vaccine encoding Schistosoma japonicum Chinese strain triose-phosphate isomerase in infected BALB/C mice.

The development of a DNA vaccine for schistosomiasis japonica and testing the protective efficacy after challenge in BALB/c mice were performed. Thirty-nine female BALB/c mice were divided into three groups. Each mouse of the control group was injected intramuscularly with 100 microg of pcDNA3.1 DNA. In the TPI group, each mouse was injected with 100 microg of pcDNA3.1-SjCTPI DNA. The TPI+IL-12 group was injected with 100 microg of pcDNA3.1-SjCTPI DNA and 100 microg of the mixture of pcDNA3.1-P35 and pcDNA3.1-P40 DNA. Each mouse was immunized three times at two-week intervals and challenged with 45 cercariae of Schistosoma japonicum Chinese strain four weeks post-immunization. Then the mice were sacrificed and perfused at 45 days after challenge; the recovered worms and hepatic eggs were counted. Cytotoxic T lymphocyte (CTL) activity mediated by SjCTPI was detected with the 51Cr release assay. ELISA was performed for the detection of anti-rTPI antibodies. Anti-rTPI antibody detection with ELISA after immunization showed ten serum samples from the control group were negative, five of ten serum samples from the TPI group were weakly positive, six of ten from the TPI+IL-12 group were also weakly positive. The CTL activity of the control group was 9.1%, while CTL activities of the TPI group and the TPI+IL-12 group were 27.6% and 54.4%, respectively. The worm and egg reduction rates of TPI group and the TPI+IL-12 group were 30.2%, 52.9%, 32.7%, and 47.0%, respectively in comparison with the control group. This study further proved the possibility of the SjCTPI DNA vaccine as a potential DNA vaccine for schistosomiasis.

Niveles de parasitemia y alteraciones histopatológicas en Mus musculus BALB/c infectado con Trypanosoma cruzi obtenido de Panstrongylus chinai del Valle Chamán, La Libertad - Perú / Parasitemia levels and histopathologycal changes in Mus musculus balb/c infected with Typanosoma cruzi obtained from Panstrongylus chinai of Chaman Valley, la libertad - Peru

Se determinó los niveles de parasitemia y las alteraciones histopatológicas en el corazón y el encéfalo de Mus musculus cepa BALB/c infectado experimentalmente con Trypanosoma cruzi obtenido de Panstrongyius chinai capturados en el valle de Chamán, La Libertad-Perú. El trabajo se hizo en 10 ejemplares de M. musculus machos (nueve experimentales y un control) de 20 días de edad. A cada uno de los ejemplares del grupo experimental se les inoculó intraperitonealmente 5x104 tripomastigotes (0,1 mL inoculo original) suspendidos en sangre, y al control 0,1 mL de solución salina fisiológica estéril. La parasitemia se inició a partir del quinto día de la inoculación del T. cruzi, elevándose hasta niveles comprendidos entre 2,5x106 y 7,0x106 tripomastigotes/mL, antes de la muerte de los animales (18 días como máximo). El corazón y encéfalo fueron separados y conservados en formol al 10 por ciento para realizar cortes y colorearlos con la técnica H/E. Todos los ratones parasitados presentaron: erizamiento de pelo, incontinencia urinaria, parálisis de extremidades posteriores y convulsiones. A nivel histopatológico, tanto en el corazón como en el encéfalo, se observaron áreas de edema, congestión vascular, hipercromasia y nidos de amastigota. En el encéfalo se presentó, además, focos de inflamación crónica y difusa y en el corazón infiltrado inflamatorio linfocitario.

A comparative study on mouse MHC class I sequences detected in Schistosoma japonicum recovered from BALB/c (H-2d) and C57BL/6 (H-2b) mice.

The mouse major histocompatibility complex (MHC) class I sequence was detected in all the 8-week-old Schistosoma japonicum recovered from BALB/c (H-2d) and C57BL/6 (H-2b) mice by in situ polymerase chain reaction (in situ PCR). The signals of the mouse class I MHC sequence were observed in the nuclei of the mesenchymal and reproductive cells of 8-week-old S. japonicum. Furthermore, the class I MHC sequence was detected in each DNA extracted from S. japonicum cercariae maintained in BALB/c and C57BL/6 mice by nested PCR. To prove both horizontal and vertical transmission of this sequence in schistosomes, we have used cercariae obtained from parasites maintained in BALB/c mice to infect C57BL/6 and BALB/c mice, and vice versa. The MHC sequences from adult worms were compared to the cercarial MHC and host MHC sequences. Nucleotide sequence comparisons between adult worm DNA, host (H-2d and H-2b mice) DNA and cercarial DNA used for the infection suggested that the sequence of mouse class I MHC was incorporated into schistosome adults and inherited throughout their life-cycle.

Production of monoclonal antibodies to the cuticle of advanced third-stage larva of Gnathostoma spinigerum.

This study has demonstrated that sera from Balb/c mice infected with live advanced third-stage larvae (aL3), but not those immunized with crude larval extract, immunoprecipitated the 25-kDa protein from surface-iodinated extract of aL3. Hybridoma cell lines derived from spleen cells of an infected mouse secreted antibodies that reacted with several tissue of aL3 including the esophagus, intestine, muscle and cuticle by immunofluorescence assay. However, none of the cuticle-positive hybridoma cell lines produced antibodies that recognized surface-iodinated protein of aL3 by immunoprecipitation. Western blot analysis showed that monoclonal antibodies (MAbs) secreted by clones derived from one of the cuticle-positive hybridoma lines recognized proteins of molecular weights ranging from 55-96 kDa. The MAbs most likely reacted with the collagenous component of the cuticle.

Tripomastigotes de sangre y de cultivo celular de trypanosoma cruzi Y: diferencia en la infectividad para ratones Balb/c / Blood and culture trypomastigotes forms of trypanosoma cruzi Y: difference on the infectivity in Balb/c mice

La variación del inóculo de las formas tripomastigotes de sangre (Tr.s) y de cultivo celular (Tr.cc.) de la cepa T. cruzi Y, en ratones BALB/c hembras y machos de distintas edades, presentó distintos grados de susceptibilidad a la infección. Aún cuando, se observó un patrón similar de comportamiento, evaluado por los niveles de máxima parasitemia, porcentajes y períodos de mortalidad. El estudio de la evolución de la enfermedad en los ratones permitió de acuerdo a la presencia o ausencia de la fase aguda y al porcentaje de mortalidad, establecer las categorías de ratones Altamente Susceptibles, Susceptibles, Parcialmente Susceptibles y Poco Susceptibles. Los ratones BALB/c inoculados con Tr.s se ubicaron en las categorías Altamente Susceptibles y Susceptibles. Mientras que los inoculados con Tr.cc. ocuparon todas las categorías (Altamente Susceptibles, Susceptibles, Parcialmente Susceptibles y Poco Susceptibles)

Life cycle of Trypanosoma cruzi (y strain) in mice

Since 1958, we have studied experimental Chagas' disease (CD) by subcutaneous inoculation of 1,000 blood forms of Trypanosoma cruzi (Y strain) in Balb/C. mice. Evolution of parasitemia remained constant, beginning on the 5th and 6th day of the disease, increasing progressively, achieving a maximum on about the 30th day. After another month, only a few forms were present, and they disappeared from the circulation after the third month, as determined from direct examination of slides and the use of a Neubauer Counting Chamber. These events coincided with the appearance of amastigote nests in the tissues (especially the cardiac ones), starting the first week, and following the Gauss parasitemia curve, but they were not in parallel until the chronic stage. In 1997, we began to note the following changes: Parasites appeared in the circulation during the first week and disappeared starting on the 7th day, and there was a coincident absence of the amastigote nests in the tissues. A careful study verified that young forms in the evolutionary cycle of T. cruzi (epi + amastigotes) began to appear alongside the trypomastigotes in the circulation on the 5th and 7th post-inoculation day. At the same time, rounded, oval, and spindle shapes were seen circulating through the capillaries and sinusoids of the tissues, principally of the hematopoietic organs. Stasis occurs because the diameter of the circulating parasites is greater than the vessels, and this makes them more visible. Examination of the sternal bone marrow revealed young cells with elongated forms and others truncated in the shape of a "C" occupying the internal surface of the blood cells that had empty central portions (erythrocytes?). We hypothesize that there could be a loss of virulence or mutation of the Y strain of Trypanosoma cruzi

Resolution of an infection with Leishmania braziliensis confers complete protection to a subsequent challenge with Leishmania Major in BALB/c mice

Both Leishmania major and L. braziliensis induce cutaneous leishmaniasis in BALB/c mice. Whereas BALB/c mice die of infection with L. major, they cure an infection with L. braziliensis. We report here that after curing an infection with L. braziliensis, BALB/c mice are resistant to challenge with L. major. When challenged with L. major, L. braziliensis pre-treated BALB/c mice mounted a delayed-type hypersensitivity response to L. major and produced high amounts of interferon-gamma (IFN-gamma) but low amounts of interleukin-4. The IFN-gamma produced by the L. braziliensis pre-infected mice was involved in the protection seen against L. major challenge since treating the mice with a neutralizing anti-IFN-gamma abrogated the protection. This suggests that cross-reactive antigen epitopes exist between L. braziliensis and L. major and that pre-infection with L. braziliensis primes BALB/c mice to epitopes on L. major that can elicit a protective Th1 response to the parasite.

Helminth parasites of conventionally maintained laboratory mice - II. Inbred strains with an adaptation of the anal swab technique

Worm burdens recovered from inbred mice strains, namely C57B1/6, C57B1/10, CBA, BALB/c, DBA/2 and C3H/He, conventionnally maintained in two institutional animal houses in the State of Rio de Janeiro, RJ, Brazil, were analysed and compared, regarding their prevalences and mean intensities. Three parasite species were observed: the nematodes Aspiculuris tetraptera, Syphacia obvelata and the cestode Vampirolepis nana. A modification of the anal swab technique is also proposed for the first time as an auxiliary tool for the detection of oxyurid eggs in mice.

Schistosoma mansoni: relocation of parasites to lungs from hepatic portal system in rodents.

The prevalence and development of adult worms in the lungs of mice and gerbils infected with Schistosoma mansoni was investigated. All infected BALB/c mice harbored the schistosomes in their lungs at 10-12 weeks post-infection, showing the distinct relocation of adult worms to the lungs, from the hepatic portal system. The male and female flukes from lungs of BALB/c mice were significantly smaller than those from livers. The percentage of gravid females in lungs was considerably lower than that in the livers. The number of eggs recovered from lungs of BALB/c mice and gerbils having lung female worms, however, was higher than that from animals without lung females, indicating egg deposition of lung females. The number of eggs detected in the brains correlated well with the number of eggs from the lungs in BALB/c and ICR mice. Out of 119 infected gerbils at 8 weeks post-infection, only two animals had egg-emboli in the brain vessels, although many eggs embolized in the lungs of those animals. These data suggest that transfer of worms to the lungs from livers involves reduction of worm recovery from the portal circulation, and also pulmonary pathology of the disease.

La caracterización de los antígenos de superficie del nemátodo parásito Trichinella spiralis: estudio de su participación en los mecanismos de protección y su utilidad en el diagnóstico de la triquinosis / Charaterization of surface antigens of the parasite nematode Trichinella spiralis: mechanisms of protection and use in diagnosis of trichinosis

Entre los aspectos más importantes en el estudio de la triquinosis, se encuentra el desarrollo de los métodos de diagnóstico específicos y confiables que permitan la detección oportuna de la infección, así como la identificación y caracterización de antígenos del parásito que previenen la infección. En este contexto, la caracterización de los antígenos de superficie esticosoma y de excreción secreción de la larva muscular de Trichinella spiralis, ha sido muy relevante, dada su alta inmunogenicidad prácticamente en todos los huéspedes hasta ahora estudiados. Así, durante el desarrollo de este trabajo se ha logrado identificar y caracterizar las moléculas de estos dos compartimentos de la larva muscular de T. spiralis que se han empleado tanto en la detección específica de la infección por T. spiralis (particularmente en cerdos), como en los ensayos para evaluar su potencial en la inducción de protección modelos murinos

Cura biológica de la criptococosis experimental murina con la asociación de anfotericina B y 5 fluorocitosina / Biological cure of murine experimental cryptococcosis with the association of anphotericin B and 5 fluorocytosine

Se evaluó la capacidad de la asociación de anfotericina B (AMB) (6mg/kg/día por medio) con 5-fluorocitosina (5FC)(300mg/kg/día), durante 2 semanas, para producir la cura biológica de la criptococosis esperimental murina. Nueve de 12 (75 por ciento) ratones Balb/C tratados, sacrificados a los 90, 180 y 210 días de evolución (3 en cada oportunidad), presentaron hepatosplenomegalia en forma inconstante y macro, microscopia, histopatología y siembra masiva de cerebro negativas. Los 3 ratones restantes (25 por ciento) murieron espontáneamente a los 81, 148 y 150 días post infección, con presencia, en la mayor parte de ellos, de macro, microscopia, histopatología y siembra masiva de cerebro positivas. Según lo apreciado en este modelo experimental, estudiado hasta los 210 días postinfección, la asociación de AMB con 5FC logró la cura biológica de esta infección experimental por lo menos a nivel de su localización más importante, el sistema nervioso central

Caracterizaçäo biológica de cepas de Leishmania amazonensis isoladas de pacientes com diferentes formas clínicas de Leishmaniose / Caracteriztion biologic of L. Amazone meuses leishmania ceps isolated of patients from diffuse clinicas forms of leishmaniose

No presente estudo, foi avaliado o comportamento biológico de 5 cepas de L. amazonensis isoladas de pacientes com leishmaniose tegumentar (3 cepas) e leishmaniose visceral (2 cepas). Foram utilizados camundongos isogênicos BALB/c, os quais säo susceptíveis à infecçäo por L. amazonensis, e desta maneira afastamos a interferência da variaçäo genética na determinaçäo da forma de doença. Os animais foram infectados com 5 x 10**6 promastigotas em fase estacionária de cada cepa. Foram realizadas medidas das lesöes das patas semanalmente, visceralizaçäo do parasita com cultivo dos linfonodos popliteos e baços e o peso destes, com 3, 8 e 15 semanas pós-infecçäo, resposta linfoproliferativa das células esplênicas 7 semanas pós-infecçäo, dosagem, dos níveis séricos de IgG, IgG1 e IgG2a anti-leishmania com 3, 7 e 10 semanas pós-infecçäo, além de avaliaçäo do reconhecimento de fraçöes antigênicas com os soros de cada cepa contra o antígeno homólogo e os das demais cepas. O comportamento da infecçäo nas patas dos camundongos BALB/c infectados com as cepas tegumentares foi semelhante ao descrito na literatura. Os animais infectados com as cepas viscerais, tiveram evoluçäo lenta das lesöes até a 15ª semana pós-infecçäo. Os aspectos histopatológicos das lesöes destes animais evidenciaram um padräo semelhante aos descritos para animais resistentes à infecçäo por leishmania, com presença de macrófagos, linfócitos, plasmócitos, formaçäo de granuloma e poucos parasitas no interior de macrófagos. Outro aspecto histológico encontrado foi a miosite e perineurite, até entäo näo descrito na leishmaniose experimental. A disseminaçäo do parasita também foi diferente no grupo de animais infectados com as cepas viscerais em relaçäo ao tegumentar. Só foi observado a presença de leishmania no cultivo dos baços dos animais infectados com as cepas viscerais na 8ª semana pós-infecçäo, enquanto nos infectados com as cepas tegumentares era detectado desde a 3ª semana. Nos animais do grupo tegumentar, os baços aumentaram de peso com o evoluir da doença e no grupo visceral diminuiram. Os níveis séricos de IgG específica para leishmania foi menor no grupo visceral, em relaçäo ao tegumentar, na 10ª semana pós-infecçäo (p < 0,05). O reconhecimento de fraçöes antigênicas também foi diferente entre os soros das cepas tegumentares e os das cepas viscerais. A banda de 63 KD foi a mais reconhecida, e os antígenos maiores do que 63 KD só foram reconhecidas em uma das cepas tegumentares (Teg 3). Os soros das cepas viscerais da 3ª semana pós-infecçäo näo reconheceram nenhuma banda no antígeno homólogo, enquanto reconheceram nos antígenos das cepas tegumentares...

Nota sobre a variaçäo do comportamento da patogenicidade de imunogecidade de cepas de T. cruzi variedade hastatus E y mutante após longo período de manutençäo em cultura / Note about variation of the patogenicity and immunogenicity of strains of T. cruzi, hastatus and Y, maintained in cultures for longer periods

Neste trabalho nos reportamos a falta absoluta de proteçäo de camundongos isogênicos Balc-C, desafiados com cepa Y de T. cruzi patogênica, que se deu tanto com formas de cultivo epimastigotas e tripomastigotas induzidas em cultivo e delas associadas, quanto com as glicoproteínas de superfície do T.hastatus mais adjuvantes. A cepa do T. hastatus foi a mesma que há anos atrás havia protegido camundongos näo isogênicos desafiados com a mesma cepa Y patogênica. Estes dados sugeriram a perda da imunogenicidade da cepa de T. hastatus, já näo infectante, ao longo de sua manutençäo em cultura. Na mesma ocasiäo, foi constatada a perda da infectividade da cepa Y mutante de T. cruzi mantida por 15 anos em cultura. Consideraçöes de natureza teórica foram feitas admitindo-se que as variaçöes biológicas e de natureza bioquímica ocorrentes com T. cruzi se desenvolvem permanente e evolutivamente, provavelmente em funçäo de mutaçöes na dependência da interaçäo entre o parasita e os componentes do meio em que ele vive

Protection of athymic (NU/NU) BALB/c mice against Plasmodium berghei by splenocytes from normal (NU/+) BALB/c mice

Camundongos atímicos BALB/c (Nu/Nu) sucumbem entre 7-13 dias após a inoculaçäo (DAI) da cepa NK65 de Plasmodium berghei. Todavia, seus singenêicos heterozigotos (Nu/+) morrem em 7-8 DAI. Camundongos nude (Nu/Nu) reconstituídos com 2x10 esplenócitos de camundongos heterozigotos singenêicos normais näo infectados (Nu/+) 20 dias antes de inoculaçäo a (DBI) do parasita, sucumbem 2 dias antes que os animais controles. Camundongos nude reconstituídos 10 ou 2 DBI, vivem 2-4 dias a mais que os animais controles e alguns deles sobrevivem. Esses achados indicam que a cepa NK65 de P. berghei induz, no mínimo, dois imunofenômenos dependentes de linfócitos T; um supressivo e outro estimulatório. A reconstituiçäo de camundongos nude com células T de camundongos BALB/c (Nu/+) parece reduzir ou "Bypass" a atividade supressora das células T, o qual leva à formaçäo de uma resposta imune protetora por alguns dos camundongos nude