A model for preservation of thymocyte-depleted thymus

DiGeorge syndrome is a disorder caused by a microdeletion on the long arm of chromosome 22. Approximately 1% of patients diagnosed with DiGeorge syndrome may have an absence of a functional thymus, which characterizes the complete form of the syndrome. These patients require urgent treatment to reconstitute T cell immunity. Thymus transplantation is a promising investigational procedure for reconstitution of thymic function in infants with congenital athymia. Here, we demonstrate a possible optimization of the preparation of thymus slices for transplantation through prior depletion of thymocytes and leukocyte cell lineages followed by cryopreservation with cryoprotective media (5% dextran FP 40, 5% Me2SO, and 5% FBS) while preserving tissue architecture. Thymus fragments were stored in liquid nitrogen at -196°C for 30 days or one year. The tissue architecture of the fragments was preserved, including the distinction between medullary thymic epithelial cells (TECs), cortical TECs, and Hassall bodies. Moreover, depleted thymus fragments cryopreserved for one year were recolonized by intrathymic injections of 3×106 thymocytes per mL, demonstrating the capability of these fragments to support T cell development. Thus, this technique opens up the possibility of freezing and storing large volumes of thymus tissue for immediate transplantation into patients with DiGeorge syndrome or atypical (Omenn-like) phenotype.

T cell receptor excision circles as a tool for evaluating thymic function in young children

The thymus is a primary lymphoid organ responsible for the maturation of T cells as well as the immunological central tolerance. It is in the antenatal period and infancy that it plays its major role. In clinical practice, T cell receptor excision circles (TRECs) are considered a direct and reliable measure of the thymic function. TRECs are a by-product of DNA formation in gene rearrangement of T cell receptors. They are stable and they do not duplicate during mitosis, representing the recent emigrant T cells from the thymus. Despite their importance, TRECs have been neglected by physicians and there is a lack of data regarding thymic function during infancy of healthy children. In order to evaluate thymic function in the first years of life, we propose measuring TRECs as a valuable tool. One hundred and three blood samples from children and adolescents between 3 months and 20 years of age were analyzed. The mean TRECs count was 136.77±96.7 copies of TRECs/μL of DNA. The individuals between 0 and 5 years of age had significantly higher TRECs values than those between 10 and 20 years of age. No significant difference was observed in TRECs values among age groups below 5 years of age. An inverse correlation between TRECs and age was found (r=0.3 P=0.003). These data highlight and validate the evidence of decreased thymus function with age, even during infancy. Awareness should be raised with this important albeit ignored organ.

Bipartição escrotal associada à má formação prepucial em ovino Morada Nova: relato de caso / Scrotal bipartition associated with preputial malformation in sheep Morada Nova: case report

As alterações anatômicas do aparelho reprodutor masculino de ovinos são pouco relatadas, especialmente a condição de bipartição escrotal. Devido à importância do esclarecimento sobre esta alteração anatômica, objetivou-se relatar o caso de um ovino da raça Morada Nova que apresentava bipartição escrotal associada à má formação prepucial. Observou-se que essa alteração pode ocorrer como condição de adaptabilidade a climas tropicais.

The anatomical changes in the male reproductive system of sheep are rarely described, especially the scrotal bipartition condition. Due to the importance of clarification of this anatomical change, we aimed to describe a case of a Morada Nova sheep that had scrotal bipartition associated with preputial malformation. It was observed that this change may occur as a adaptability condition to tropical climates.

Armazenamento de Trichogramma pretiosum Riley (Hymenoptera: Trichogrammatidae) em baixas temperaturas / Cold storage of Trichogramma pretiosum Riley (Hymenoptera: Trichogrammatidae)

Avaliou-se o efeito do armazenamento em baixa temperatura de pupas de Trichogramma pretiosum, em ovos de Sitotroga cerealella, sobre a emergência e a viabilidade reprodutiva do parasitoide. Os testes foram conduzidos em câmara climática a 5, 8 e 10° C, 70% UR, escotofase constante, com 10 tratamentos (testemunha, 4, 6, 8, 10, 12, 14, 16, 18 e 20 dias de armazenamento) e 10 repetições. Foram distribuídos 1.000 ovos contendo pupas em tubos de ensaio (10 x 3 cm), 100 foram mantidos a 25º C, 70% UR e 14 horas de fotofase (testemunha) e os demais armazenados. Após cada período de armazenamento, 100 ovos foram transferidos para 25º C e avaliados quanto à emergência e funções reprodutivas. A emergência de T. pretiosum não foi influenciada pelo período de armazenamento nas três temperaturas (Tukey P ≤ 0,05) e as menores taxas de emergência foram de 85,6%, 84,5% e 77,6%, respectivamente para os parasitoides armazenados a 5, 8 e 10° C. Não houve perda da viabilidade reprodutiva dos parasitoides provenientes da estocagem nas temperaturas avaliadas, pois todas as fêmeas acasaladas produziram indivíduos de ambos os sexos. Estes resultados auxi-liarão no planejamento da criação massal, embalagem e transporte de T. pretiosum do laboratório para o local de liberação.

The objective of this work was to evaluate the effect of storage at low temperature of Trichogramma pretiosum pupae in eggs of Sitotroga cerealella on the emergence and reproduction rates of the parasitoid. The trials were conducted in a climatic chamber at 5, 8 and 10º C, 70% R.H., constant scotophase, with 10 treatments: 4, 6, 8, 10, 12, 14, 16, 18, 20 days of storage and a control not kept in storage, all in 10 replicates. A total of 1,000 eggs containing pupae were distributed in glass tubes (10 x 3 cm), 100 of which were kept at 25° C, 70% RH and 14 h photophase (control), while the remaining ones were stored. After each storage period 100 eggs were transferred to 25º C and the emergence and reproductive capacity were observed. The T. pretiosum emergence rate was not affected by the storage time at 3 temperatures (Tukey P ≤ 0.05) and the lowest emergence rates were 85.6%, 84.5% and 77.6%, respectively for parasitoid storage at 5, 8 and 10º C. The reproductive capacity of T. pretiosum was not affected after the exposure periods in the temperatures evaluated. This result will aid in the planning of mass production, package and transport of T. pretiosum to the place of release.

Evaluation of the immune humoral response of Brazilian patients with Rubinstein-Taybi syndrome

Rubinstein-Taybi syndrome (RTS) is a rare developmental disorder characterized by craniofacial dysmorphisms, broad thumbs and toes, mental and growth deficiency, and recurrent respiratory infections. RTS has been associated with CREBBP gene mutations, but EP300 gene mutations have recently been reported in 6 individuals. In the present study, the humoral immune response in 16 RTS patients with recurrent respiratory infections of possible bacterial etiology was evaluated. No significant differences between patients and 16 healthy controls were detected to explain the high susceptibility to respiratory infections: normal or elevated serum immunoglobulin levels, normal salivary IgA levels, and a good antibody response to both polysaccharide and protein antigens were observed. However, most patients presented high serum IgM levels, a high number of total B cell and B subsets, and also high percentiles of apoptosis, suggesting that they could present B dysregulation. The CREBBP/p300 family gene is extremely important for B-cell regulation, and RTS may represent an interesting human model for studying the molecular mechanisms involved in B-cell development.

The first report in Brazil of severe infection caused by community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA)

Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) is an emergent pathogen in Brazil. However, there are no data on the prevalence of CA-MRSA. We report here the first well-characterized case of severe life-threatening CA-MRSA infection in a child living in Rio de Janeiro city. The patient had many complications including hematogenous osteomyelitis and involvement of multiple sites requiring drainage of soft-tissue abscess, and pleural and pericardial empyema. The MRSA isolates recovered were genotyped using PFGE, SCCmec typing and multilocus sequence typing. Disk diffusion tests were performed following Clinical and Laboratory Standards Institute recommendations. In addition, the presence of Panton-Valentine leukocidin (PVL) was assessed by PCR amplification, using specific primers for lukF-pv (encoding for the F subunit of the PVL). The bacterial isolates were related to the ST30-SCCmecIV lineage (Oceania Southwest Pacific clone), a PVL producer CA-MRSA previously detected in Porto Alegre, RS, Brazil. Also, the isolates analyzed were susceptible to all non-â-lactam antibiotics tested. The present report demonstrates that disseminated CA-MRSA disease is also occurring in Rio de Janeiro. Thus, the empirical treatment of moderate or severe infections suspected of being associated with CA-MRSA needs to be reviewed in order to allow prompt initiation of an effective therapy that also covers these microorganisms.

Cytotoxicity of current adhesive systems: in vitro testing on cell culture of L929 and balb/c 3T3 fibroblasts / Citotoxicidade de sistemas adesivos atuais: teste in vitro em cultura de células fibroblásticas L929 e balb/c 3T3

Purpose: The aim of this study was to evaluate the cytotoxicity of three current adhesives: Prime&Bond NT (PBNT), Single Bond (SB) and XENO III (XENO). Methods: After embedding and curing circles of filter paper with the tested adhesives, the filters were placed in contact with the solidified agar surface over L929 monolayer cells plated in 6-well cell culture plate and incubated for 24 h. The inhibition zone around the filter papers was measured in mm. MTT assay was performed using fibroblasts Balb/c 3T3 cell lines in multiwell culture plates. All assays were done in triplicate. Results: All materials were cytotoxic (Kruskal-Wallis, P<0.05) in a similar level to latex (P>0.05). For intra-groups analysis, SB presented the lowest cytotoxicity (P<0.01), while there was no statistical difference between PBNT and XENO (P>0.05). MTT assay confirmed the cytotoxicity of the tested adhesives. Conclusion: Considering the limits of this work, all adhesives tested were as cytotoxic as latex.

Objetivo: O objetivo deste estudo foi avaliar a citotoxicidade de três adesivos: Prime & Bond NT (PBNT), Single Bond (SB) e XENO III (XENO). Metodologia: Após embebição e polimerização de filtros de papel com os referidos adesivos, estes foram colocados em contato com a superfície de agar solidificada sobre a monocamada de células L929 plaqueadas em cultura celular de 6-poços e incubadas por 24 h. A zona de inibição formada ao redor dos filtros de papel foi medida em milímetros. Outro teste realizado foi o do MTT, utilizando fibroblastos Balb / c 3T3 em placas de multi-poços, sendo os ensaios realizados em triplicatas. Resultados: Todos os materiais testados foram citotóxicos (Kruskal-Wallis, P < 0,05) e semelhantes ao látex (P > 0,05). Para a análise intra-grupos, o SB apresentou a mais baixa citotoxicidade (P < 0,01), enquanto não houve diferença estatística entre PBNT e XENO (P > 0,05). O ensaio de MTT confirmou a citotoxicidade dos adesivos. Conclusão: Considerando as limitações deste trabalho, todos os adesivos testados foram tão citotóxicos quanto o látex.

Dientes supernumerarios y atresia maxilar. Tratamiento orto-quirúrgico en Odontopediatría. Relato de un caso clínico: 5 años de seguimiento / Supernumeraty teeth: surgical treatment in children

Se describe a los dientes supernumerarios o hiperodoncia como el aumento del número de órganos dentarios comparado con la dentición normal. Todos los dientes supernumerarios deben ser extraídos, respetando el momento ideal para su remoción quirúrgica, evaluando siempre el momento oportuno para intervenir. En el presente trabajo las autoras presentan un caso clínico de una paciente de sexo femenino, que tuvo un seguimiento durante cinco años a partir de los 6 meses de edad asistida en el Curso de Extensión Universitaria Bebé Clínica de la Universidad Federal de Río Grande del Sur. Durante sus visitas periódicas de revisión odontológica correspondientes al programa de la Bebé Clínica, se diagnosticó clínicamente la presencia de mordida cruzada anterior, atresia maxilar y radiográficamente la presencia de dos elementos supernumerarios. Frente a ese diagnóstico, es necesario un planeamiento multidisciplinario, o sea, orto-quirúrgico para la realización del tratamiento, destacando una vez más, la importancia de la atención odontológica del bebé y el diagnóstico precoz de posibles alteraciones como en el caso clínico en cuestión: la presencia de elementos supernumerarios y maloclusión dentaria.

Antibody response to pneumococcal capsular polysaccharide vaccine in Down syndrome patients

The majority of children with Down syndrome (DS) tend to have frequent bacterial infections including recurrent respiratory infections. Our objective was to evaluate the production of antibodies to pneumococcal polysaccharide antigens after active immunization in DS subjects. IgG antibodies to pneumococcal serotypes (1, 3, 6B, 9V, and 14) were measured before and 6 weeks after immunization with a 23-valent pneumococcal vaccine (Pneumo23®, Pasteur-Merrieux) in 6- to 13-year-old DS children (N = 17) and in aged-matched normal controls (N = 30). An adequate response was defined as a 4-fold increase over baseline or a post-immunization level of specific pneumococcal serotype antibody > or = 1.3 æg/mL. After immunization, all DS children had an increase in post-immunization levels against all serotypes analyzed. A 4-fold or more increase was observed in all DS children concerning serotypes 1 and 14, in 90 percent of subjects for serotypes 3 and 9V, and in 65 percent for serotype 6B. Regarding this increase, 8 of the 17 DS children had an adequate response to all serotypes analyzed, 8/17 patients to 4 serotypes and 1/17 to 3 serotypes. However, when we compared post-immunization levels between DS children and controls, we observed lower levels in the former group (P < 0.05) for all serotypes except serotype 3. We conclude that pneumococcal polysaccharide immunization could be beneficial for these DS children.

The use of reverse transcription-PCR for the diagnosis of X-linked chronic granulomatous disease

Chronic granulomatous disease (CGD) is an inherited disorder of the innate immune system characterized by a defective oxidative burst of phagocytes and subsequent impairment of their microbicidal activity. Mutations in one of the NADPH-oxidase components affect gene expression or function of this system, leading to the phenotype of CGD. Defects in gp91-phox lead to X-linked CGD, responsible for approximately 70 percent of CGD cases. Investigation of the highly heterogeneous genotype of CGD patients includes mutation analysis, Northern blot or Western blot assays according to the particular case. The aim of the present study was to use reverse transcription (RT)-PCR for the analysis of molecular defects responsible for X-linked CGD in eight Brazilian patients and to assess its potential for broader application to molecular screening in CGD. Total RNA was prepared from Epstein B virus-transformed B-lymphocytes and reverse transcribed using random hexamers. The resulting cDNA was PCR-amplified by specific and overlapping pairs of primers designed to amplify three regions of the gp91-phox gene: exons 1-5, 3-9, and 7-13. This strategy detected defective gp91-phox expression in seven patients. The RT-PCR results matched clinical history, biochemical data (nitroblue tetrazolium or superoxide release assay) and available mutation analysis in four cases. In three additional cases, RT-PCR results matched clinical history and biochemical data. In another case, RT-PCR was normal despite a clinical history compatible with CGD and defective respiratory burst. We conclude that this new application of RT-PCR analysis - a simple, economical and rapid method - was appropriate for screening molecular defects in 7 of 8 X-linked CGD patients.

Antioxidant dietary deficiency induces caspase activation in chick skeletal muscle cells

Apoptosis and necrosis are two distinct forms of cell death that can occur in response to different agents and stress conditions. In order to verify if the oxidative stress induced by dietary selenium and vitamin E deficiencies can lead muscle cells to apoptosis, one-day-old chicks were reared using diets differing in their vitamin E (0 or 10 IU/kg) and selenium (0 or 0.15 ppm) supplementation. Chick skeletal muscle tissue was obtained from 28-day-old animals and used to verify apoptosis occurrence based on caspase activity detection and DNA fragmentation. Antioxidant deficiency significantly increased caspase-like activity assessed by the hydrolysis of fluorogenic peptide substrates (Abz-peptidyl-EDDnp) at lambdaexc = 320 nm and lambdaem = 420 nm. Proteolytic activation was not accompanied by typical internucleosomal DNA fragmentation detected by field inversion gel electrophoresis. Although the general caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp(O-Me) fluoromethyl ketone (Z-VAD-fmk) (0 to 80 muM) did not block caspase-like activity when preincubated for 30 min with muscle homogenates, the hydrolyzed substrates presented the same cleavage profile in HPLC (at the aspartic acid residue) when incubated with the purified recombinant enzyme caspase-3. These data indicate that oxidative stress causes caspase-like activation in muscle cells and suggest that cell death associated with exudative diathesis (dietary deficiency of selenium and vitamin E) can follow the apoptotic pathway

Glycosaminoglycans affect the interaction of human plasma kallikrein with plasminogen, factor XII and inhibitors

Human plasma kallikrein, a serine proteinase, plays a key role in intrinsic blood clotting, in the kallikrein-kinin system, and in fibrinolysis. The proteolytic enzymes involved in these processes are usually controlled by specific inhibitors and may be influenced by several factors including glycosaminoglycans, as recently demonstrated by our group. The aim of the present study was to investigate the effect of glycosaminoglycans (30 to 250 æg/ml) on kallikrein activity on plasminogen and factor XII and on the inhibition of kallikrein by the plasma proteins C1-inhibitor and antithrombin. Almost all available glycosaminoglycans (heparin, heparan sulfate, bovine and tuna dermatan sulfate, chondroitin 4- and 6-sulfates) reduced (1.2 to 3.0 times) the catalytic efficiency of kallikrein (in a nanomolar range) on the hydrolysis of plasminogen (0.3 to 1.8 æM) and increased (1.9 to 7.7 times) the enzyme efficiency in factor XII (0.1 to 10 æM) activation. On the other hand, heparin, heparan sulfate, and bovine and tuna dermatan sulfate improved (1.2 to 3.4 times) kallikrein inhibition by antithrombin (1.4 æM), while chondroitin 4- and 6-sulfates reduced it (1.3 times). Heparin and heparan sulfate increased (1.4 times) the enzyme inhibition by the C1-inhibitor (150 nM)

The 245 kb amplified chromosome of Leishmania (V. ) braziliensis contains a biopterin transporter gene

Leishmania (V.) braziliensis M2903 presents a small linear and stable 245 kb chromosome originating from a genomic amplification. Similar amplifications present in other species of Leishmania contain a gene coding for a biopterin transporter. Since Leishmania is auxotrophic for this metabolite, this amplification could result from the need to better capture biotpterin from growth media under specific circumstances. In this paper we show that this gene is also present in L. (V.) braziliensis small chromosome, which shares sequences with other genomic amplifications already described

Periódicos científicos: características e exigências / Scientific periodicals: challenges and perspectives

Apresenta uma rápida reflexão acerca do periódico científico, enquanto veículo de comunicação e registro da produção intelectual. Desde que surgiu, esse tipo de publicação apresentou um expressivo aumento no número de títulos publicados, culminando com a marca de 1.000.000 de títulos editados no mundo em 2000. A criação de índices bibliográficos e, posteriormente bases de dados, levou à criação de indicadores para avaliação das revistas científicas nem sempre reconhecidos pelos pesquisadores e estudiosos. No Brasil, vários esforços têm sido empregados buscando imprimir qualidade às publicações nacionais, gerando efetivos programas de avaliação. Outra preocupação em relação às revistas científicas é o aumento excessivo do custo para manutenção das assinaturas internacionais, que tem levado à procura de alternativas que garantam o acesso à informação e ao documento, como por exemplo, a publicação eletrônica e os consórcios institucionais.

Estudo da populaçäo microbiana e da liberaçäo de amônia de cama de frangos tratada com gesso agrícola / Study of the microbial population and ammonia release of treated broiler litter with agricultural gypsum

O presente trabalho foi realizado com o objetivo de avaliar a populaçäo microbiana (contagem padräo) e a liberaçäo de amônia da cama de frangos de maravalha tratada com gesso agrícola, durante o ciclo de criaçäo das aves. Foram utilizados 1440 pintos de um dia para corte, criados em galpäo convencional dividido em boxes, sob densidade de nove aves/m². Os dados das variáveis analisadas foram coletados no início, no 25º dia e ao final do experimento (49º dia de vida). O delineamento experimental adotado foi o inteiramente ao acaso, com a distribuiçäo de nove tratamentos em esquema fatorial 4 x 2 + 1 (níveis de gesso x formas de aplicaçäo), com quatro repetiçöes e 40 aves por parcela. Os resultados evidenciaram a capacidade inibidora do gesso na volatilizaçäo de amônia de cama de frangos no 25º dia e ao final do experimento, principalmente para a aplicaçäo parcelada, implicando no decréscimo da contagem padräo de microrganismos

Immunization of persons with HIV infection and other secondary immunodeficiencies

Patients with secondary immunodeficiencies are at a high risk of infection. Currently some of these infections are preventable through specific immunization. Prevention of these diseases can diminish morbidity and mortality amongst these patients. In this review we describe the use of vaccines in persons with secondary immunodeficiencies.

Imaturidade imunológica fetal e neonatal: implicaçöes na evoluçäo clínica da infecçäo pelo HIV-1 em crianças / Immunologic fetal and neonatal immaturity: influence on the clinical evolution of HIV-1 infection in chindren

Crianças infectadas pelo HIV-1, por via vertical, apresentam uma evoluçao clínica mais grave do que crianças infectadas por outras vias e adultos. A imaturidade fisiológica dos sistemas imunitários fetal e neonatal, no momento da infecçao, parece ter papel crucial na progressao da infecçao pelo HIV-1 em crianças. Neste artigo, fazemos revisao da ontogenia do sistema imunológico humano, correlacionando-a com a imunopatogenia da síndrome de imunodeficiência adquirida (AIDS), em crianças infectadas por transmissao vertical, em suas diferentes fases.

Análises microbiológicas da cama de frango utilizada na alimentaçäo de ruminantes / Microbiological analyses of poultry litter used for ruminant feeding

O presente trabalho teve por objetivos avaliar a variaçäo da contagem total de bactérias em amostras de "cama" de frangos compostas por casca de arroz, após o armazenamento, assim como identificar aquelas bactérias potencialmente patogênicas presentes. Para as contagens foi utilizado o ágar para contagem padräo (Difco), os meios utilizados na identificaçäo foram ágar sangue, MacConkey, Baird-Parker, e para as anaeróbicas tubos com infusäo de cérebro e coraçäo, meio de carne cozida (Difco) e placas de ágar Clostrisel, BHI ágar e ágar gema de ovo, incubados em anerobiose com o sistema Gas-Pak (BBL). A metodologia para a identificaçäo bacteriana foi dirigida para a procura de Staphylococcus aureus, Salmonella sp, Clostridium perfringens, C. botulinum, C. chauvoei, Campylobacter sp, Escherichia coli, e Corynebacterium sp. Foram colhidas amostras da "cama" provenientes de galpöes de frangos de corte. O delineamento experimental utilizado foi inteiramente casualizado com quatro tratamentos e quatro repetiçöes, totalizando dezesseis amostras. Os resultados da avaliaçäo demonstraram haver decréscimo no número total de bactérias (NTB) quando se compararam os tratamentos T1(zero dias de estocagem) e T2 (14 dias de estocagem). Em contraposiçäo os tratamentos T3 (28 dias de estocagem) e T4 (42 dias de estocagem) apresentaram contagens de NTB significativamente superiores em relaçäo ao tratamento T1. Constatou-se a presença de bactérias patogênicas primárias ou secundárias da família das enterobactérias (Escherichia coli, Proteus, Arizona, Providencia, Edwardsiella), assim como Staphylococcus aureus, S. epidermidis, além de diversas espécies de Clostridium como C. perfringens, C. sordelli, C. chauvoei, C. tetani e C. novyi, e algumas cepas de Corynebacterium pyogenes. Entre toda a flora bacteriana determinada, o grupo das enterobactérias, demonstrou maior sensibilidade às modificaçöes do meio, em funçäo dos diversos tempos de estocagem testados

Mucosal immunology and models of mucosal HIV infection

The mucosa associated lymphoid tissue regulates and coordinates immune responses against mucosal pathogens. Mucosal tissues are the major targets exposed to HIV during transmission. In this paper we describe in vitro models of HIV mucosal infection using human explants to investigate target cells within this tissue.

Gastrointestinal immune responses in HIV infected subjects

The gut associated lymphoid tissue is responsible for specific responses to intestinal antigens. During HIV infection, mucosal immune deficiency may account for the gastrointestinal infections. In this review we describe the humoral and cellular mucosal immune responses in normal and HIV-infected subjects.