Evaluation of short-interfering RNAs treatment in experimental rabies due to wild-type virus

ABSTRACTWe have evaluated the efficacy of short-interfering RNAs targeting the nucleoprotein gene and also the brain immune response in treated and non-treated infected mice. Mice were inoculated with wild-type virus, classified as dog (hv2) or vampire bat (hv3) variants and both groups were treated or leaved as controls. No difference was observed in the lethality rate between treated and non-treated groups, although clinical evaluation of hv2 infected mice showed differences in the severity of clinical disease (p = 0.0006). Evaluation of brain immune response 5 days post-inoculation in treated hv2 group showed no difference among the analyzed genes, whereas after 10 days post-inoculation there was increased expression of 2',5'-oligoadenylate synthetase 1, tumor necrosis factor alpha, interleukin 12, interferon gamma, and C-X-C motif chemokine 10 associated with higher expression of Ngene in the same period (p < 0.0001). In hv2 non-treated group only higher interferon beta expression was found at day 5. The observed differences in results of the immune response genes between treated and non-treated groups is not promising as they had neither impact on mortality nor even a reduction in the expression of N gene in siRNA treated animals. This finding suggests that the use of pre-designed siRNA alone may not be useful in rabies treatment.

Minimally traumatic alveolar ridge augmentation with a tunnel injectable thermo-sensitive alginate scaffold

Injectable bone substitutes and techniques have been developed for use in minimally invasive procedures for bone augmentation. Objective : To develop a novel injectable thermo-sensitive alginate hydrogel (TSAH) as a scaffold to induce bone regeneration, using a minimally invasive tunnelling technique. Material and Methods : An injectable TSAH was prepared from a copolymer solution of 8.0 wt% Poly(N-isopropylacrylamide) (PNIPAAm) and 8.0 wt% AAlg-g-PNIPAAm. In vitro properties of the material, such as its microstructure and the sustained release of recombinant human bone morphogenetic protein-2 (rhBMP-2), were investigated. Then, with the subperiosteal tunnelling technique, this material, carrying rhBMP-2, was injected under the labial periosteum of the maxillary anterior alveolar ridge in a rabbit model. New bone formation was evaluated by means of X-ray, micro-computed tomography (micro-CT), fluorescence labelling, histological study, and immunohistochemistry study. Results : The material exhibited good injectability and thermo-irreversible properties. SEM showed an interconnected porous microstructure of the TSAH. The result of ALP activity indicated sustained delivery of BMP-2 from the TSAH from days 3 to 15. In a rabbit model, both TSAH and TSAH/rhBMP-2 induced alveolar ridge augmentation. The percentage of mineralised tissue in the TSAH/rhBMP-2 group (41.6±3.79%) was significantly higher than in the TSAH group (31.3±7.21%; p<0.05). The density of the regenerating tissue was higher in the TSAH/rhBMP-2 group than in the other groups (TSAH group, positive control, blank control; p<0.05). Conclusions : The TSAH provided convenient handling properties for clinical application. To some extent, TSAH could induce ridge augmentation and mineral deposition, which can be enhanced when combined with rhBMP-2 for a minimally invasive tunnelling injection. .

Percepção dos graduandos de enfermagem sobre o seu envelhecimento / Percepción de los estudiantes de enfermería sobre lo proprio envejecimiento / Perception of nursing undergraduate students on self-aging

RESUMO Objetivo: analisar a percepção dos graduandos de enfermagem sobre o próprio envelhecimento. Método: pesquisa de abordagem qualitativa, realizada em agosto e setembro de 2011, com 18 graduandos de enfermagem de uma Universidade pública de Salvador (Bahia). Os depoimentos foram analisados por meio da Análise de Conteúdo. Resultados: apreendeu-se o núcleo temático: Percepção do graduando de enfermagem sobre o próprio envelhecimento e, a partir deste, emergiram duas subcategorias: A) O Não Pensar; B) O contexto influenciando no processo. Conclusão: os graduandos revelam que o envelhecimento está intrínseco ao desenvolvimento humano, e possui o vínculo familiar, a espiritualidade e atividade física como ferramentas fundamentais para um envelhecimento ativo. Entretanto, os mesmos relatam que, o modo de vida acelerado e estressante vivido na sociedade possibilita inserir hábitos considerados inadequados, como o consumo de “fast food” e álcool, que trazem influências negativas para o próprio processo de envelhecimento. .

RESUMEN Objetivo: analizar la percepción de los estudiantes de enfermería sobre su proprio envejecimiento. Método: estudio cualitativo, realizado en agosto y septiembre de 2011, con 18 estudiantes de enfermería de una universidad pública en Salvador/Bahia. Los datos fueron analizados através de análisis de contenido. Resultado: incautados el tema central: Percepción de alumnos de enfermería sobre su propio envejecimiento y de esto surgieron dos subcategorías: A) No creo; B) El contexto influye en el proceso. Conclusión: los estudiantes revelan que el envejecimiento es intrínseco al desarrollo humano, y tiene los vínculos familiares, la espiritualidad y la actividad física como herramienta clave para el envejecimiento activo. Sin embargo, el mismo informe que, debido a la forma de vida que se vive en la sociedad de ritmo rápido y estresante permite insertar hábitos considerados inadecuados, como el consumo de “comida rápida” y el alcohol y convertirse en influencias negativas para su propio proceso tuvo como objetivo analizar de los estudiantes de enfermería su propio envejecimiento. .

ABSTRACT Objective: to analyze the perceptions of nursing undergraduate students on their self-aging process. Method: qualitative study carried out between August and September, 2011 with 18 nursing undergraduate students of a public university in Salvador, Bahia. The interviews were analyzed by means of the Content Analysis method. Results: the following thematic concept was apprehended: Perceptions of nursing undergraduates on their self-aging, which generated two subcategories: A) The “don’t think about it” process; B) The context infl uencing the process. Conclusion: undergraduates reveal that the aging process is an intrinsic factor to human development. Family ties, spirituality and physical activity would be key mechanisms toward active aging. However, students also reported that their accelerated and stressed social lifestyles led to inadequate habits, such as the consumption of fast food and alcohol, which become negative infl uences in their aging process. .

Pro- and anti-inflammatory cytokine expression and histopathological characteristics in canine brain with traumatic brain injury

We analyzed the expression level and cellular localization of pro- and anti-inflammatory cytokines and histopathologically characterized canine traumatic brain injury (TBI). Canine TBI brains revealed subarachnoid and cerebral cortical hemorrhage, neutrophilic infiltration, neuronal necrosis, astrocytosis, and vasogenic edema. Immunohistochemical evaluations suggested that both pro-inflammatory cytokines [interleukin (IL)-1beta, IL-6, and tumor necrosis factor-alpha] and anti-inflammatory cytokines [IL-10 and transforming growth factor-beta (TGF-beta)] were highly expressed in neurons and neutrophils. In particular, the highest magnitude of expression was identified for IL-1beta and TGF-beta. This data helps describe the pathologic characteristics of canine TBI, and may help in the design of potential therapeutic approaches to control secondary damage by inflammatory cytokines.

The presence of antiautonomic membrane receptor antibodies do not correlate with brain lesions in Chagas' disease / A presença de anticorpos anti-autonômicos não se correlaciona a lesões cerebrais na doença de Chagas

We previously demonstrated correlation between parasympathetic dysfunction and brain white matter lesions in chronic chagasic patients. OBJECTIVE: To correlate serum functional circulating antibodies with beta adrenergic (Ab-β), muscarinic (Ab-M) or muscarinic and beta adrenergic (Ab-Mβ) activity, the autonomic system function and brain lesions in chronic chagasic patients. METHOD: In fifteen consecutive chagasic patients, the autonomic nervous system was evaluated and brain magnetic resonance imaging (MRI) was performed. The sera of all patients were tested to the presence of circulating functional antibodies. RESULTS: Sera from 11 of 15 chronic chagasic patients had some activity (Ab-β: 7; Ab-M: 1; Ab-Mβ: 3); however, there was no significant correlation between the presence of antibodies and the autonomic system function or the presence of hyperintensities in MRI. CONCLUSION: The mechanism involved in the genesis of hyperintense lesions seen in brain MRI of chronic chagasic patients is still unresolved, although apparently related to parasympathetic dysfunction.

A correlação entre disfunção parassimpática e lesões de substância branca cerebral em pacientes chagásicos já foi previamente demonstrada. OBJETIVO: Correlacionar a presença de anticorpos circulantes funcionais com atividade beta-adrenérgica (Ab-β), muscarínica (Ab-M) ou muscarínica e beta adrenérgica (Ab-Mβ), a presença de disautonomia e lesões de substância branca cerebral em pacientes chagásicos crônicos. MÉTODO: Em quinze pacientes chagásicos consecutivos, foram realizados a avaliação do sistema nervoso autônomo e ressonância magnética (RM) do crânio. O soro dos pacientes foi testado para a presença de anticorpos funcionais circulantes. RESULTADOS: O soro de 11 dos 15 pacientes chagásicos apresentou alguma atividade (Ab-β: 7; Ab-M: 1; Ab-Mβ: 3); porém não houve correlação significativa entre a presença de anticorpos circulantes e disautonomia ou de hiperintensidades à RM. CONCLUSÃO: O mecanismo envolvido na gênese das lesões hiperintensas à RM do crânio dos pacientes chagásicos crônicos não está esclarecida ainda, apesar de aparentemente relacionada à disfunção parassimpática.

Evaluation of IgG4 and total IgG antibodies against cysticerci and peptide antigens for the diagnosis of human neurocysticercosis by ELISA.

To support the clinical diagnosis of human neurocysticercosis (NCC), we evaluated two peptides, HP6-3 and Ts45W-1, as well as crude saline extract (SE) of Tenia solium cysticerci as antigens for the detection of specific IgG4 subclass and total IgG antibodies by an enzyme-linked immunosorbent assay (ELISA). The sera of definitive diagnosed NCC patients, patients infected with other parasitoses and healthy controls were examined. The diagnostic sensitivity for IgG4 and total IgG detection of the ELISA against SE antigen was 100% and 64.3% with a high amount of cross-reactions to taeniasis saginata at 88.9% (8/9) and 100% (9/9), respectively. The SE-based IgG4-ELISA showed the highest specificity (80.9%). Both peptide-based IgG4-ELISAs provided a superior sensitivity (78.6%) to the total IgG tests whereas their specificity was 66.7% for HP6-3 and 69.8% for Ts45W-1 only. The SE-based ELISA for the detection of specific IgG4 antibody can be used for the diagnosis of neurocysticercosis as well as for serological surveys of NCC endemic areas. The peptide-based IgG4 ELISAs potentially provide a reliable and cost effective alternative method independent from live parasite supply.

Astrocytic and microglial response and histopathological changes in the brain of horses with experimental chronic Trypanosoma evansi infection / Resposta astrocítica e microglial e alterações histopatológicas no sistema nervoso central de eqüinos infectados cronicamente com Trypanosoma evansi

This study aimed to characterize astrocytic and microglial response in the central nervous system (CNS) of equines experimentally infected with T. evansi. The experimental group comprised males and females with various degrees of crossbreeding, ages between four and seven years. The animals were inoculated intravenously with 10(6) trypomastigotes of T. evansi originally isolated from a naturally infected dog. All equines inoculated with T. evansi were observed until they presented symptoms of CNS disturbance, characterized by motor incoordination of the pelvic limbs, which occurred 67 days after inoculation (DAI) and 124 DAI. The animals in the control group did not present any clinical symptom and were observed up to the 125th DAI. For this purpose the HE histochemical stain and the avidin biotin peroxidase method was used. Lesions in the CNS of experimentally infected horses were those of a wide spread non suppurative meningoencephalomyelitis.The severity of lesions varied in different parts of the nervous system, reflecting an irregular distribution of inflammatory vascular changes. The infiltration of mononuclear cells was associated with anisomorphic gliosis and reactive microglia was identified. The intensity of the astrocytic response in the CNS of the equines infected by T. evansi characterizes the importance of the performance of these cells in this trypanosomiasis. The characteristic gliosis observed in the animals in this experiment suggests the ability of these cells as mediators of immune response. The parasite, T. evansi, was not identified in the nervous tissues.

Este estudo objetivou caracterizar a participação astrocítica e microglial no sistema nervoso central (SNC) de eqüinos experimentalmente infectados com T. evansi. O grupo experimental foi formado por machos e fêmeas com vários graus de cruzamentos e idade variando entre quatro e sete anos. Os animais foram inoculados com 10(6) tripomastigotas de T. evansi, originalmente isolada de um cão infectado naturalmente. Todos os eqüinos inoculados foram observados até o aparecimento dos sintomas neurológicos, caracterizados por incoordenação motora dos membros pélvicos, o qual ocorreu entre 67 e 124 dias após a inoculação (DPI). Os animais do grupo controle não apresentaram sinais clínicos e foram observados até o 125º DPI. Para este propósito, foram utilizados os métodos histoquímicos (HE) e imunoistoquímicos do complexo avidina-biotina peroxidase (ABC). A lesão no sistema nervoso central (SNC) dos eqüinos infectados com T. evansi foi caracterizada como meningoencefalomielite não supurativa. A gravidade das lesões variou em diferentes segmentos do SNC, refletindo distribuição irregular das alterações vasculares. Infiltrado perivascular e meníngeo foi associado a gliose anisomórfica e microgliose reativa. A intensidade da resposta astrocítica no SNC dos equinos infectados com T. evansi caracteriza a importância da performance destas células nas tripanossomíases. A gliose observada nos animais deste experimento sugerem a habilidade destas células como mediadoras da resposta imune. T. evansi não foi identificado no parênquima do SNC.

Immunohistochemical characterization of mononuclear cells and MHC II expression in the brain of horses with experimental chronic Trypanosoma evansi infection / Caracterização imunoistoquímica de células mononucleares e expressão de CMH II no sistema nervoso central de eqüinos com infecção crônica experimental por Trypanosoma evansi

An histochemical and immunohistochemical study was carried out to evaluate the mechanisms of immune response of horses experimentally infected by Trypanosoma evansi. For this purpose the HE histochemical stain and the avidin biotin peroxidase method were used. To determine the presence and immunoreactivity of immune cells we used anti-major histocompatibility complex II antibodies. Cellular infiltration fenotype was characterized with the aid of anti-CD3 antibody for T lymphocytes and by anti-BLA 36 antibodies for B lymphocytes. Macrophages were marked with an antibody against myeloid/histyocites antigen (clone Mac387). Lesions in the CNS of experimentally infected horses were those of a wide spread non suppurative encephalomyelitis and meningomyelitis. The severity of lesions varied in different parts of the nervous system, reflecting an irregular distribution of inflammatory vascular changes. Lymphoid perivascular cuffs and meningeal infiltrations were of predominantly composed of T and B cells. The parasite, T. evansi, was not identified in these horses tissues.

Este estudo objetivou caracterizar a resposta imune celular no sistema nervoso central (SNC) de eqüinos com infecção crônica experimental por Trypanosoma evansi. Para este propósito, foram utilizados os métodos histoquímicos (HE) e imunoistoquímicos do complexo avidina-biotina peroxidase (ABC). O fenótipo do infiltrado celular foi caracterizado com o auxílio de anticorpos anti - CD3, para linfócitos T e antiBLA36 para linfócitos B. Os macrófagos foram marcados com anticorpo antiantígenos da linhagem mielóide/histiócitos (Clone Mac387). A lesão no sistema nervoso central (SNC) dos eqüinos infectados com T. evansi foi caracterizada como meningoencefalite e meningomielite não supurativa. A gravidade das lesões variou em diferentes segmentos do SNC, refletindo distribuição irregular das alterações vasculares. A distribuição de células T e B e antígenos do complexo maior de histocompatibilidade classe II foram avaliados dentro do SNC de eqüinos cronicamente infectados com T. evansi. O infiltrado perivascular e meníngeo eram constituídos predominantemente por células T e B. Macrófagos foram raramente visualizados. T.evansi não foi identificado no parênquima do SNC dos eqüinos.

Interações imunocerebrais e implicações nos transtornos psiquiátricos / Brain-immune interactions and implications in psychiatric disorders

OBJETIVO: Nesta revisão será focado o papel das citocinas no sistema nervoso central e suas implicações para o quadro depressivo. Posteriormente, serão discutidos os principais achados sobre medidas de citocinas em pacientes com depressão maior. MÉTODO: Foi realizada uma pesquisa no Pubmed selecionando estudos entre 1999-2007, utilizando as seguintes palavras-chave: "depression, cytokine"; "depressive disorder, cytokine". Focou-se nos estudos de medidas de citocinas pró-inflamatórias em pacientes com síndrome depressiva que utilizaram critérios DSM. RESULTADOS: Várias linhas de evidência sugerem que as citocinas possam exercer um papel na depressão. Entre elas, destacam-se: citocinas induzindo a "comportamento doentio"; doenças clínicas relacionadas com citocinas também apresentam associação com quadros depressivos; uso de imunoterapia levando ao desenvolvimento de depressão. Além disso, níveis elevados de citocinas pró-inflamatórias em pacientes com depressão foram relatados, apesar de resultados contraditórios. CONCLUSÃO: O papel das citocinas na fisiopatologia em alguns casos de depressão é descrito; porém, uma relação causal não foi ainda estabelecida. Novos estudos são necessários para determinar padrões específicos de citocinas em pacientes com depressão, levando em consideração outros fatores associados à ativação imunológica. Além disso, medidas simultâneas de múltiplos marcadores biológicos podem gerar informações importantes para a compreensão dos mecanismos fisiopatológico da depressão e em doenças relacionadas à produção de citocinas.

OBJECTIVE: This review will focus on the role of cytokines in the central nervous system and its implications to depressive disorder. We will then discuss the main findings of cytokine measurements in patients with major depressive disorder. METHOD: We searched Pubmed for studies published from 1999-2007, using the keywords depression and cytokine; and depressive disorder and cytokine. We have focused on pro-inflammatory cytokine measurements in patients with depression syndrome using DSM-criteria. RESULTS: Several lines of evidence suggest that cytokines have effects on depression, such as the induction of sickness behavior; clinical conditions related to cytokines that also overlap depressive symptoms; and immunotherapy that can lead to depressive symptoms attenuated by antidepressant treatment. Finally, patients with depression exhibit increased levels of pro-inflammatory cytokines, although conflicting results have been described. CONCLUSION: Cytokines may play a role in the pathophysiology of some cases of depression, although a causal link has not been established yet. Further longitudinal studies are needed to determine patterns of cytokine in patients with major depressive disorder, taking into account confounding factors closely associated with the activation of pro-inflammatory cytokines. In addition, simultaneous measurements of multiple biomarkers could provide critical insights into mechanisms underlying major depressive disorder and a variety of common cytokine-related diseases.

Development and evaluation of an enzyme immunoassay for rapid diagnosis of rabies in humans and animals.

The presently advocated tests for rapid diagnosis of rabies such as fluorescent antibody test (FAT) is expensive and requires expertise to carry out and interpret the results. In this study we have developed and evaluated a simple enzyme immuno-assay (EIA) to detect rabies antigen in the brain specimens of animals and humans. We have also evaluated the utility of this test in ante mortem diagnosis of human rabies. The brain homogenates of suspected rabid animals (n=250), humans (n=16) and clinical samples like saliva (n=16) and cerebrospinal fluid (CSF, n=16) applied on to ELISA plates coated with rabies antinucleoprotein antibody and the absorbed rabies nucleoprotein antigen was detected using biotinylated anti-nucleoprotein antibody followed by treatment with streptavidin peroxidase conjugate and colour development with OPD. Rabies infected and normal mouse brain homogenates were used as positive and negative controls respectively. The results of this test was evaluated with fluorescent antibody technique (for brain samples) and mice inoculation test (for saliva and CSF samples). A distinct dark brown color was seen in positive control and all positive samples and there was no color development in negative control and samples. The concordance between FAT and EIA was 98.4%. With brain samples, 83.3% with saliva and 91.6% with CSF samples. The specificity of the test was found to be 100%. It can be concluded that the EIA described here is a sensitive, specific and rapid test for post mortem diagnosis of rabies in animals and humans. The utility of this test for ante mortem diagnosis of rabies needs to be further evaluated.

Expression of TNF-alpha and TGF-beta1 in the Rat Brain After a Single High-Dose Irradiation

Cytokines and growth factors are important regulatory proteins controlling the growth and differentiation of normal and malignant glial cells. In this study, we investigated the expression and origin of tumor necrosis factor-alpha (TNF-alpha) and transforming growth factor-beta 1 (TGF-beta 1) in the subacute brain injury after a single high-dose irradiation using 60 Sprague-Dawley rats. The right cerebral hemispheres of rats were exposed to a single 10 Gy dose of gamma rays using Ir-192. The radiation effect was assessed at 1 week, 2 weeks, 4 weeks, 6 weeks, and 8 weeks after irradiation, and the results were compared with those in sham operation group. Histological changes characteristic of radiation injury were correlated with the duration after the single dose irradiation. The loss of cortical thickness also increased with the lapse of time after irradiation. The TNF-alpha expression in the irradiated cerebral hemispheres was significantly increased compared with that in the sham operation group. TGF-beta 1 expression was also increased in the irradiated hemispheres. Immunohistochemical study revealed that TGF-beta 1 was expressed predominantly by infiltrating macrophages and astrocytes around the necrotic areas. These findings indicate that TNF-alpha and TGF-beta 1 may play prominent roles in the radiation injuries after a single high-dose irradiation.

Correlation between mouse intracerebral potency & 50 per cent histamine sensitizing dose of pertussis vaccine.

A good correlation was observed between mouse intracerebral potency in terms of International Units and 50 per cent histamine sensitizing dose (HSD50) of various types of pertussis vaccine. The correlation was significant irrespective of the inactivating agent used for the preparation of pertussis vaccine. If the potency of the vaccine increased, the HSD50 decreased and vice versa showing a negative correlation between potency and HSD50. The relative histamine sensitizing (HS) activities of various types of pertussis vaccines in comparison to that of heat inactivated pertussis vaccine (HIPV) obtained in different experiments were very similar although the HSD50 values of same types of vaccines obtained in these experiments varied greatly. The conventional HIPV preparations having potency around 8 IU per 40 x 10(9) organisms had HSD50 values in the range of 1.0 to 2.2 x 10(8) organisms. The estimation of HSD50 of pertussis vaccine has been found to be a useful measure of the toxicity of the vaccine and also correlates well with the potency of the vaccine.