Hamster IFN-γ+CD4+ and IL-4+CD4+ T cell responses against leptospires are significantly higher than those of mice.

BACKGROUND: Leptospirosis is a bacterial disease caused by the Leptospira interrogans. The hamster is considered a susceptible host while the mouse is resistant. The knowledge of hamster T cell immunity is limited compared to the mouse. The reason why the hamster and the mouse give different responses to leptospires remains unclear. OBJECTIVE: To determine the differential responses of CD4+ T cells between hamsters and mice using Leptospira interrogans as an infectious model. METHODS: The CD4+ T-cell reactivity and their intracellular cytokine responses after infection with live L.interrogans serovar Autumnalis or leptospiral antigens, or injection with recombinant LipL32 protein (rLipL32) were elucidated. For secondary immune responses, mononuclear cells were re-stimulated with leptospiral crude antigens (LAg) or rLipL32. Intracellular cytokines and CD4+ T cells were determined using flow cytometry. RESULTS: There were no significant differences between the percentages of hamster and mouse CD4+ and CD25+CD4+ T cell responses to live bacteria. Mouse CD4+ (24.50±1.98%) and CD25+CD4+ T cells (3.83±0.88) responded significantly higher than those of hamster (15.07±2.82% and 2.00±0.37%) when infected and re-stimulated with LAg. The numbers of IFN-γ and IL-4 producing cells in hamsters at 1.76±0.10% and 0.82±0.25% for IFN-γ+CD4+ and IL-4+CD4+ T cells were significantly higher than those in resistant mice at 0.10±0.02% and 0.23±0.03% for IFN-γ+CD4+ and IL-4+CD4+ T cells. CONCLUSION: Hamsters responded significantly higher in secondary stimulation especially in the levels of the IFN-γ+ and IL-4+CD4+ T cells. The mechanisms of this dissimilarity remain to be elucidated.

Characterisation of monoclonal antibodies specific for hamster leukocyte differentiation molecules.

Flow cytometry was used to identify mAbs that recognize conserved epitopes on hamster leukocyte differentiation molecules (hLDM) and also to characterize mAbs developed against hLDM. Initial screening of mAbs developed against LDMs in other species yielded mAbs specific for the major histocompatibility (MHC) II molecule, CD4 and CD18. Screening of sets of mAbs developed against hLDM yielded 22 new mAbs, including additional mAbs to MHC II molecules and mAbs that recognize LDMs expressed on all leukocytes, granulocytes, all lymphocytes, all T cells, a subset of T cells, or on all B cells. Based on comparison of the pattern of expression of LDMs expressed on all hamster leukocytes with the patterns of expression of known LDMs in other species, as detected by flow cytometry (FC), four mAbs are predicted to recognize CD11a, CD44, and CD45. Cross comparison of mAbs specific for a subset of hamster T cells with a cross reactive mAb known to recognize CD4 in mice and one recognising CD8 revealed they recognize CD4. The characterization of these mAbs expands opportunities to use hamsters as an additional model species to investigate the mechanisms of immunopathogenesis of infectious diseases.

Correlations between phytohemagglutinin response and leukocyte profile, and bactericidal capacity in a wild rodent.

Phytohemagglutinin (PHA)-induced swelling is widely used to investigate cell-mediated and innate immunity across different vertebrate taxa. However, its physiological mechanism is still an open question due to the complexity of the involved immune components. In the present study, we measured the synchronous variations of PHA response, the proportion of different subtypes of leukocytes, as well as serum bactericidal capacity in circulation blood at 6, 12 and 24 h after PHA versus PBS injection in striped hamster, Cricetulus barabensis. First, the results showed that PHA responses reached a peak at 6 h postinjection, then sharply declined at 12 h and 24 h postinjection. Serum bactericidal capacity was higher at 6 h and 12 h than at 24 h. The proportion of different subtypes of leukocytes, as well as the ratio of neutrophils to lymphocytes did not display significant changes across different time points. Second, PHA response was positively correlated with the proportion of neutrophils and serum bactericidal capacity. The proportion of monocytes was negatively correlated with that of eosinophils and neutrophils. The proportion of basophils was negatively correlated with that of lymphocytes. Our results indicate that earlier enhanced PHA response is important for the striped hamster to cope with changing environmental conditions due to its small body mass, and the increased components of innate immunity in circulation blood may contribute to the enhancement of PHA swelling response.

Delayed hypersensitivity reaction after initial dose of infliximab: a case report.

We report here an unusual case of delayed hypersensitivity reaction in a young woman with ulcerative colitis after the first administration of infliximab (IFX). The patient developed severe serum-sickness-like reaction, and her anti-IFX antibody titer increased rapidly after a single infusion of IFX. The possible reason for the delayed hypersensitivity reaction to a single IFX exposure might be the presensitization of the patient by murine antigens as she had been keeping mice and hamsters as pets for several years.

A novel small animal model to study the replication of simian foamy virus in vivo.

Preclinical evaluation in a small animal model would help the development of gene therapies and vaccines based on foamy virus vectors. The establishment of persistent, non-pathogenic infection with the prototype foamy virus in mice and rabbits has been described previously. To extend this spectrum of available animal models, hamsters were inoculated with infectious cell supernatant or bioballistically with a foamy virus plasmid. In addition, a novel foamy virus from a rhesus macaque was isolated and characterised genetically. Hamsters and mice were infected with this new SFVmac isolate to evaluate whether hamsters are also susceptible to infection. Both hamsters and mice developed humoral responses to either virus subtype. Virus integration and replication in different animal tissues were analysed by PCR and co-cultivation. The results strongly indicate establishment of a persistent infection in hamsters. These studies provide a further small animal model for studying FV-based vectors in addition to the established models.

Vaccination with leishmania hemoglobin receptor-encoding DNA protects against visceral leishmaniasis.

Leishmaniasis is a severe infectious disease. Drugs used for leishmaniasis are very toxic, and no vaccine is available. We found that the hemoglobin receptor (HbR) of Leishmania was conserved across various strains of Leishmania, and anti-HbR antibody could be detected in kala-azar patients' sera. Our results showed that immunization with HbR-DNA induces complete protection against virulent Leishmania donovani infection in both BALB/c mice and hamsters. Moreover, HbR-DNA immunization stimulated the production of protective cytokines like interferon-γ (IFN-γ), interleukin-12 (IL-12), and tumor necrosis factor-α (TNF-α) with concomitant down-regulation of disease-promoting cytokines like IL-10 and IL-4. HbR-DNA vaccination also induced a protective response by generating multifunctional CD4(+) and CD8(+) T cells. All HbR-DNA-vaccinated hamsters showed sterile protection and survived during an experimental period of 8 months. These findings demonstrate the potential of HbR as a vaccine candidate against visceral leishmaniasis.

Koch's postulate of Arcanobacterium pyogenes and its immunogenicity in local and imported Saanen goats.

The aim of this survey was to study Koch's postulate of Arcanobacterium pyogenes recovered from the necrotic lung of a kid and to compare the immunogenicity of this isolate in local and imported Saanen goats. The disease was successfully reproduced in intrathoracically challenged hamsters which showed lung congestion and liver abscesses, while hamsters that were intraperitoneally challenged showed only the formation of intestinal abscesses. The percentage of histopathologic legions in 12 observed microscopic fields per lung of three groups of hamsters (unchallenged controls, challenged intrathoracically and challenged intraperitoneally) showed a significant increase in lung necrosis of the intrathoracically challenged group, followed by intraperitoneally challenged hamsters, in comparison to unchallenged controls (p<0.05). In addition, the frequency of mucus accumulation in alveolar ducts followed the same respective pattern (p>0.05), while there was no significant difference in the frequency of neutrophil infiltration (p>0.05). The isolate was successfully recovered from the lungs and livers of hamsters challenged by both routes. Saanen does showed significant seroconversion using the indirect haemagglutination (HA) test and slide agglutination test (SAT) and at three weeks following priming and boosting with A. pyogenes antigens (p<0.05); however, only SAT showed significant seroconversion in local does at three weeks post booster (p<0.05). The possible causes and impact of the greater immunogenicity to A. pyogenes antigens in Saanen goats compared to local does are discussed.

A combination DNA vaccine encoding nucleoside hydrolase 36 and glycoproteine 63 protects female but not male hamsters against Leishmania mexicana.

Leishmaniasis is a group of diseases caused by protozoan parasites of the Leishmania genus. Previous studies have shown that a DNA vaccine encoding Leishmania donovani antigen nucleoside hydrolase 36 and L. mexicana glycoprotein 63 is protective in mice. We investigated here the efficacy of this DNA vaccine to induce protection in golden hamsters. Male hamsters were more susceptible to infection by Leishmania mexicana than females. Following immunization with two doses of the DNA vaccine, only females resulted protected while males developed normal lesions.

Production of polyclonal antisera.

Much of modern biology and biochemistry relies on the availability of highly specific antibodies for use in such ubiquitous techniques as immunohistochemistry, ELISAs, immunoprecipitation, and immunoblotting. Thus, the generation of large quantities of specific antibodies directed against proteins or peptides of interest is essential to the success of both basic and applied research programs. In addition, with the advent of antibody-based proteomic strategies for profiling protein expression and post-translational modification, a requirement for timely production of specific antibodies has been exemplified. Polyclonal antibodies derived from animals immunized with purified proteins or peptides are particularly valuable for use in the laboratory. This unit provides protocols for the production of polyclonal antisera specific for protein antigens using rabbits, rats, mice, and hamsters.

Severe respiratory syndrome induced by allergic mono-sensitization to European hamster (Cricetus cricetus) in a older woman.

Although the increase in the rate of hamster ownership, no report of allergic sensitization to common hamster (Cricetus cricetus)-derived allergens as a consequence of domestic exposure has been published in Italy. A 64-year-old woman was referred to our Allergy Centre for the recent onset of conjunctival and severe respiratory symptoms (rhinitis, cough, wheezing and dyspnea). About three months ago she had purchased a common hamster as home pet. Another hamster had lived at patient's home for about four months nine years ago. The results of SPT revealed allergic sensitization to Cricetus cricetus dander only (wheal 6x7 mm, positive control 7x7 mm). Total IgE were 59.3 kU/L. Specific IgE only to Cricetus cricetus epithelia (2.10 kUA/L), were also detected. Spirometry revealed a moderate degree of bronchial obstruction. Some important considerations can be drawn from our report: a) few months of hamster ownership are probably sufficient to induce an allergic sensitization and clinical symptoms, b) older age of sensitization in comparison to other studies, c) rapid remission of clinical symptoms after the removal of hamster d) skin prick tests and/or evaluation of specific IgE for hamster allergens should be performed in all potentially susceptible individuals.

Detection of the antibody to lymphocytic choriomeningitis virus in sera of laboratory rodents infected with viruses of laboratory and newly isolated strains by ELISA using purified recombinant nucleoprotein.

An enzyme-linked immunosorbent assay (ELISA) was developed to detect the antibody against lymphocytic choriomeningitis virus (LCMV) in sera of laboratory animals. In this ELISA system, LCMV-nucleoprotein (NP) expressed by recombinant baculovirus and purified with high molar urea was used as the antigen. Sera from laboratory animals experimentally infected with the Armstrong strain or the newly isolated M1 strain of LCMV were examined to detect anti-LCMV antibody by the ELISA system, and the reactivity was compared with that of IFA test. Regardless of LCMV strain, all the sera of adult mice infected with LCMV were positive with very high optical density (OD). Also, the sera from mice neonatally infected with LCMV M1 strain were positive with slightly lower OD than adult mice. In contrast, all the sera of uninfected mice were negative to LCMV-NP antigen. Similarly, anti-LCMV antibodies were detected in all the sera of hamsters, mastomyses, and gerbils infected with the LCMV Armstrong strain. The results of the ELISA were in complete agreement with those of IFA, and indicate the high sensitivity and specificity of the ELISA system in the detection of anti-LCMV antibody. Because this ELISA system does not require handling infectious LCMV in the course of the antigen preparation and serological assay, there is no risk of contamination in the laboratory or nearby animal facility. In addition, by using negative control antigen in parallel with positive antigen in ELISA, we can exactly check the LCMV contamination in laboratory animals.

Essential role of TNF receptor superfamily 25 (TNFRSF25) in the development of allergic lung inflammation.

We identify the tumor necrosis factor receptor superfamily 25 (TNFRSF25)/TNFSF15 pair as critical trigger for allergic lung inflammation, which is a cardinal feature of asthma. TNFRSF25 (TNFR25) signals are required to exert T helper cell 2 (Th2) effector function in Th2-polarized CD4 cells and co-stimulate interleukin (IL)-13 production by glycosphingolipid-activated NKT cells. In vivo, antibody blockade of TNFSF15 (TL1A), which is the ligand for TNFR25, inhibits lung inflammation and production of Th2 cytokines such as IL-13, even when administered days after airway antigen exposure. Similarly, blockade of TNFR25 by a dominant-negative (DN) transgene, DN TNFR25, confers resistance to lung inflammation in mice. Allergic lung inflammation-resistant, NKT-deficient mice become susceptible upon adoptive transfer of wild-type NKT cells, but not after transfer of DN TNFR25 transgenic NKT cells. The TNFR25/TL1A pair appears to provide an early signal for Th2 cytokine production in the lung, and therefore may be a drug target in attempts to attenuate lung inflammation in asthmatics.

Avaliação da imunização de hamsters com plasmídeos que codificam componentes salivares de Lutzomiya longipalpis (LJM19) e/ou antígeno parasitário (KMP11) contra a infecção com Leishmania chagasi / Evaluation of immunization of hamsters with plasmids encoding salivary components of Lutzomyia longipalpis (LJM19) and / or parasite antigen (KMP11) against infection with Leishmania chagasi

Hamsters têm sido utilizados como modelos experimentais visando compreender os mecanismos de respostas imunes contra espécies de Leishmania do complexo donovani. Estes modelos são capazes de reproduzir muitas das manifestações clínicas da leishmaniose visceral humana. Estudos recentes demonstraram que a imunização de hamsters com plasmídeos codificantes para proteínas salivares (LJM19) de Lutzomyia longipalpis, vetor de L. chagasi, bem como antígenos parasitários (KMPl1) protege hamsters contra um desafio letal com Leishmania chagasi. Neste trabalho, hamsters foram utilizados para avaliar o efeito protetor contra uma infecção por L. chagasi utilizando imunização com os plasmídeos que codificam as proteínas LJM19 e KMP11 administrados em conjunto. A imunização com os plasmídeos induziu a produção de IFN-y nos linfonodos drenantes do local das imunizações quando os animais foram avaliados 7, 14 e 21 dias após a última imunização. Uma vez imunizados e desafiados com L. chagasi mais saliva do vetor, os animais mostraram maiores relações (...) nos linfonodos drenantes quando mensuradas 7 e 14 dias após o desafio. Quando avaliados 2 e 5 meses após o desafio, os animais imunizados mostraram menores cargas parasitárias no baço e no fígado e maiores relações (...) no baço 2 meses após o desafio. Além disso, os hamsters imunizados apresentaram maior conservação da arquitetura histológica do baço e do fígado nos tempos avaliados e não desenvolveram distúrbios hematológicos quando comparados com animais controles sadios.

Contudo, efeito protetor adicional pela imunização com os diferentes plasmídeos administrados em conjunto não foi observado em relação às imunizações com os plasmídeos separados. Comparações entre rotas de administração de plasmídeos foram estudadas utilizando as vias intradérmica e intramuscular. Os grupos de animais que receberam a imunização intradérmica apresentaram uma proteção mais prolongada quando comparados aos animais imunizados intramuscularmente. Estes resultados mostram que apesar da combinação de plasmídeos não induzir maior proteção que os plasmídeos separados, a via de imunização intradérmica pode conferir uma proteção mais duradoura quando comparada com a imunização pela via intramuscular.

Molecular cloning and sequences of interleukin-10 in the Djungarian (Phodopus sungorus), Chinese (Cricetulus griseus), and Syrian (Mesocricetus auratus) hamster.

Interleukin 10 (IL-10) genes of Djungarian, Chinese, and Syrian hamsters were cloned. The clones of IL-10 consisted of 537 bp nucleotides and 178 amino acids in full length, and the nucleotide and amino acid sequences exhibited a high degree of homology with those of the mouse and human. Since the number and position of signal sequences, N-glycosylations and cysteine sites in the IL-10 amino acid sequences of the hamsters were the same as those of the mouse, we suggest that the IL-10 molecular structures of the hamster are closer to that of the mouse than human.

Histological evaluation of the lesion induced by inoculation of Leishmania mexicana in the cheek pouch of the hamster / Avaliação histológica da lesão induzida pela inoculação de Leishmania mexicana na bolsa jugal do hamster

We have studied the role of the immune response in the morphology of the leishmaniotic granuloma induced in the cheek pouch of hamsters, an immunologically privileged site, after inoculation of 3 x 105 Leishmania mexicana. Animals were histologically and immunologically evaluated until 120 days after inoculation. Independent of the time of sacrifice, the animals were always non-reactors to the footpad test (FPT). At histology, the introduction of L. mexicana in the cheek pouch leads to an abscess that evolves to a granulomatous reaction rich in amastigote forms, and later it leads to resolution, even in the absence of immune response detectable by FPT. Our results demonstrate that the development of immune response is not preponderant for the control of infection induced by L. mexicana inoculated subcutaneously in the cheek pouch of the hamster. It also suggests that the macrophages present in the leishmaniotic granuloma are capable of eliminating this parasite, even in the absence of immune response evaluated by FPT.

No presente estudo, investigamos o papel da resposta imune na morfologia do granuloma leishmaniótico induzido na bolsa jugal do hamster, um local imunologicamente privilegiado, após inoculação de 3x105 Leishmania mexicana. Os animais foram avaliados histológica e imunologicamente até os 120 dias da inoculação. Independente da época do sacrifício, os animais foram sempre não reatores ao teste do coxim plantar. Histologicamente, a inoculação de Leishmania mexicana na bolsa jugal resultou na formação de abcesso que evoluiu para reação granulomatosa rica em formas amastigotas e, posteriormente, para resolução. Esses resultados sugerem que o desenvolvimento da resposta imune não é preponderante no controle da infecção induzida pela Leishmania mexicana inoculada subcutaneamente na bolsa jugal do hamster. Sugerem ainda que os macrófagos que compõe os granulomas leishmanióticos são capazes de eliminar esse parasita, independente da presença de resposta imune avaliável pelo teste do coxim plantar.

[Clinical usefulness of the histamine release test in assessing adult asthmatic patients who keep hamster].

Recently, the prevalence of keeping pets, such as hamsters and guinea pigs, at home has been increasing in Japan. The number of adult asthmatic patients who keep hamster (HKA) has increased, and accounted for 20% of all pet owners in 1997. Histamine release tests (HRT) were performed on 28 patients of HKA, who consulted with the outpatient clinic of our department, and the results were compared with CAP-RAST. HRT were performed with peripheral blood obtained from each subject, and histamine content was measured by fluorescent assay. 7 in 12 RAST positive patients and 4 in 16 RAST negative patients in HKA had histamine release titer reaction over 15 ng/ml. HRT and CAP-RAST scores were correlated in HKA, and HRT could be evaluated before and after stopping keeping hamsters in five subjects to find decreased HRT scores in all. The results obtained indicate that those tests were useful for the clinical diagnosing and monitoring of HKA.

Leishmania major infections in Phlebotomus duboscqi fed on murine models immunized with L. major subcellular antigens and sandfly gut antigens.

The ability of antibodies in bloodmeals of mice and hamsters immunized with Leishmania major subcellular fractions and sandfly (Phlebotomus duboscqi) gut antigens to inhibit development of L. major in its vector P. duboscqi was examined. Antibodies from animals immunized with either L. major subcellular fractions alone or sandfly gut antigen alone were not very effective in inhibiting development of L. major in the sandfly. When P. duboscqi were fed on blood from animals immunized with both parasite flagella and sandfly gut antigen, development of L. major was significantly inhibited (P<0,05). Control sandflies fed on naive animals displayed a normal pattern of parasite development to the metacyclic stage. Electron microscopy studies showed that one of the mechanisms through which antisandfly gut antibody can cause inhibition of parasite development is by lysing sandfly gut epithelium. This study has demonstrated that it is possible to reduce transmission of leishmaniosis through immunization against both the parasite and its sandfly vector.