Resistance of Santa Ines and crossbred ewes to naturally acquired gastrointestinal nematode infections

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Immune responses in sheep naturally infected with Oestrus ovis (Diptera: Oestridae) and gastrointestinal nematodes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Obtenção e caracterização de anticorpo monoclonal murino anti-fator VIII da coagulação sangüínea

Entre os avanços da engenharia celular e biotecnologia nas últimas décadas, destaca-se a produção de anticorpos monoclonais murinos (AcMm) utilizados no aprimoramento diagnóstico nas rotinas laboratoriais. A produção de fator VIII de alta pureza sempre foi o desejo e a preocupação das indústrias de hemoderivados para tratamento de pacientes portadores de hemofilia A, porém este produto inexiste no Brasil, sendo necessária sua obtenção no mercado internacional a custos elevados. O trabalho tem por objetivo a produção de AcMm anti-fator VIII humano (FVIII H) através da expansão dos clones e caracterização imunoquímica do anticorpo. Camundongos Balb/c foram imunizados com FVIII H purificado como também proveniente de crioprecipitado e as células esplênicas dos animais foram fusionadas com células mielomatosas murinas segundo o método descrito por Kohler e Milstein para produção de híbridos em cultura. Foram testados 1.983 híbridos dos quais 105 foram submetidos à clonagem. Destes, 39 obtiveram monoclonalidade e 7 destes clones foram caracterizados através de técnicas de immunoblotting. Foram submetidas à purificação por cromatografia três imunoglobulinas de diferentes classes pertencentes aos clones LAMB1-10A1A4, LAMB1-17A1A1 e LAMB1-24A2A1. A imunoglobulina purificada pertencente ao clone LAMB1-10A1A4 foi adsorvida em coluna de imunoafinidade para purificação de concentrado de FVIII proveniente de crioprecipitado plasmático.

Immunological disturbance in West and Lennox-Gastaut syndromes

Investigamos a imunidade humoral e celular em 18 pacientes com sindrome de West, 12 com sindrome de Lennox-Gastaut e 19 controles. Os exames realizados foram os seguintes: percentagem de linfócitos Ô e  no sangue periférico, niveis séricos de IgG, IgA e IgM, sensibilização cutânea com o DNCB, PHA intra dérmica, teste de inibição de migração de leucócitos e transformação blástica de linfócitos em presença de PHA. Detectamos deficiência de imunidade celular em 28 crianças, (18 com sindrome de West e 10 com sindrome de Lennox-Gastaut) e baixos níveis de imunoglobulinas em apenas 6. A depressão imunitária mostrou-se mais intensa nas crianças que apresentam sindrome de West.

Immunologic aspects of West syndrome and evidence of plasma inhibitory effects on T cell function

OBJETIVO: O objetivo deste estudo foi determinar o perfil da deficiência imune em um grupo bem definido de epilepsia: crianças com síndrome de West (SW) e seus padrões EEG de evolução, idade-dependentes, como os complexos onda-aguda- onda lenta generalizadas da síndrome de Lenox-Gastaut (SLG) e as pontas multifocais independentes (PMI). MÉTODO: Um grupo de 50 crianças, 33 com SW, 10 com SLG, 7 com PMI e 20 crianças sadias (controle) foram avaliadas em relação aos seguintes parâmetros:determinação de subpopulações de linfócitos T (CD1, CD3, CD4 e CD8), relação CD4/CD8 e resposta proliferativa de linfócitos frente a fitohemaglutinina (PHA), na presença de plasma autólogo ou de plasma AB (homólogo). A prova cutânea de sensibilização ao Dinitroclorobenzeno (DNCB) foi realizada apenas nos pacientes. Os níveis séricos de IgG, IgA e IgM foram comparados aos valores normais em crianças Brasileiras, em diferentes faixas etárias. RESULTADOS: A resposta ao DNCB foi ausente ou fracamente reativa em 76% dos pacientes. Níveis séricos elevados de IgG (45,7%) e de IgM (61,4%) e baixos de IgA (23,9%) foram detectados nos pacientes. A determinação das subpopulações de linfócitos T em sangue periférico mostrou: deficiência nas proporções de células CD3+ (p<0,05) e de CD4+ (p<0,05), aumento de CD8+ (p<0,01) e diminuição da relação CD4 / CD8 (p<0,001). A proporção de células CD1+ no grupo controle manteve-se menor que 3%, enquanto que em 18% dos pacientes esses níveis variaram entre 3 e 11%. A resposta proliferativa de linfócitos frente a PHA revelou índices blastogênicos significativamente mais baixos apenas quando células dos pacientes foram cultivadas na presença do próprio plasma (plasma autólogo). Quando estas células foram cultivadas na presença de plasma AB, não se evidenciou diferença significativa em relação ao grupo controle. CONCLUSÃO: A imunodeficiência na SW caracterizou-se por: anergia, alteração de imunidade mediada por células e dos níveis de imunoglobulinas, presença de timócitos imaturos na circulação periférica e deficiência funcional de linfócitos T induzida por fatores plasmáticos inibidores. Discutem-se as principais evidências de disfunção imune como imunodeficiência e autoimunidade.

Renal involvement in canine leishmaniasis: a morphological and immunohistochemical study

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Imunoglobulinas e C3 no granuloma paracoccidióidico

Utilizou-se o modelo experimental de paracoccidioidomicose, em camundongos, induzida pela inoculação endovenosa de suspensão de formas cerebriformes do P. brasiliensis (cepa Bt2; 1x10(6) formas viáveis/animal), para avaliar, após 2, 4, 8, 16 e 20 semanas: 1. A presença de imunoglobulinas e C3 nos granulomas pulmonares, por imunofluorescência direta; 2. A resposta imune humoral (imunodifusão) e celular (teste do coxim plantar), e 3. A histopatologia das lesões. Os camundongos apresentaram resposta imunocelular positiva desde a 2a. semana, com depressão transitória na 16a. semana, e anticorpos desde a 4a. semana, com pico na 16a. semana. Os granulomas pulmonares foram epitelióides, com numerosos fungos e microabscesses; a extensão das lesões foi progressiva até a 16a. semana, com regressão discreta na 20a. semana. Desde a 2a. semana, houve deposição de IgG e C3 na parede dos fungos no interior dos granulomas e a presença de células IgG positivas no halo linfomononuclear periférico; estes achados foram máximos entre a 4a. e 16a. semanas. Não se detectou depósito de IgG e C3 no interstício dos granulomas. IgG e C3 parecem exercer papel precoce e importante na defesa do hospedeiro contra o P. brasiliensis, contribuindo possivelmente para a destruição dos fungos e bloqueando a difusão de antígenos para fora dos granulomas.

Identification of enteropathogens from buffalo calves with and without diarrhoea in the Ribeira Valley, State of São Paulo, Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Agarose gel electrophoresis of cerebrospinal fluid proteins of dogs after sample concentration using a membrane microconcentrator technique

Background: Cerebrospinal fluid (CSF) is produced in the cerebral ventricles through ultrafiltration of plasma and active transport mechanisms. Evaluation of proteins in CSF may provide important information about the production of immunoglobulins within the central nervous system as well as possible disturbances in the blood-brain barrier. Objective: the objective of this study was to measure the concentration and fractions of protein in CSF samples using a membrane microconcentrator technique followed by electrophoresis, and to compare the protein fractions obtained with those in serum. Methods: CSF samples from 3 healthy dogs and 3 dogs with canine distemper virus infection were concentrated using a membrane microconcentrator having a 0.5 to 30,000 d nominal molecular weight limit (Ultrafree, Millipore, Billerica, MA, USA). Protein concentration was determined before and after concentration. Agarose gel electrophoresis was done on concentrated CSF samples, serum, and serial dilutions of one of the CSF samples. Results: Electrophoretic bands were clearly identified in densitometer tracings in CSF samples with protein concentrations as low as 1.3 g/dL. The higher CSF protein concentration in dogs with distemper was mainly the result of increased albumin concentration. Conclusion: the microconcentrating method used in this study enables characterization of the main protein fractions in CSF by routine electrophoresis and may be useful for interpreting the underlying cause of changes in CSF protein concentrations

Histological and immunological reaction of cattle skin to first-instar larvae of Dermatobia hominis

Six cattle that had earlier exposure to Dermatobia hominis were infested experimentally with first-instar larvae of the parasite. Skin biopsies taken at intervals were studied in wax and in plastic sections. The avidin-biotin-peroxidase method was used to detect the presence and localization of host immunoglobulins (Igs) G and M and antigens of first and second instar larvae of Dermatobia hominis. The larvae penetrated actively through the skin and migrated towards the subcutaneous tissues. The great numbers of eosinophils suggest that they are the most important cell in mediating damage to D.hominis larvae. The immunoglobulins bound only to dead or moulting larvae in which access to binding sites may have been altered. This could represent a morphological manifestation of a mechanism that protects larvae from the host immune response. Large amounts of soluble antigens detected along the fistulous tract may be important in the maintenance of this tract by disturbing the normal cicatrization process.

Condição imunológica de bovinos das raças Holandesa e Nelore frente a Babesia bovis e B. bigemina em duas regiões do Estado de São Paulo

The immunological reply of a population to an infectious agent can vary between races and handling of this population. Regional research becomes important, in order to know the interrelation between the agent and its host. In this way, the occurrence of immunoglobulins of class G, anti-Babesia bovis and anti-Babesia bigemina in the Nelore (Bos indicus) and Hostein breed (Bos taurus), was investigated in two regions of the State of São Paulo, 300 km distant from each other. For the indirect method of ELISA, 1, 161 bovine serum samples were tested. The medium frequencies of antibodies showed that in the two regions exists an enzootic stability for B. bovis in both breeds studied; even so there was a tendency of marginal area for the Nelore breed in one of the regions. Regarding B. bigemina, in both regions exists enzootic stability for the Hostein and enzootic instability for the Nelore breed. Therefore, acute cases of the disease or specific outbreaks by B. bigemina infection in the Nelore breed may occur in these regions.

CROSS-REACTIVITY BETWEEN HARD TICK ANTIGENS

1. The present study was carried out to determine the target cells and tissues for anti-tick immunoglobulins using an indirect immunohistochemical technique.2. Sections in triplicate prepared from unfed ticks Rhipicephalus appendiculatus, R. evertsi and Amblyomma variegatum were used to assess the cross-reactivity of serum from guineapigs naturally infested with these tick species or immunized against them.3. The sections showed slight (+) to strong (++++) labelling of several structures in the tick body, e.g. salivary gland, gut lumen and malpighian tubules, depending on the serum used.4. The immune serum resulting from the immunization of guinea pigs with an extract of unfed nymphs of R. appendiculatus ticks showed the most intense cross-reactivity with the sections examined.

Kinetics of B cell response during Yersinia enterocolitica infection in resistant and susceptible strains of mice

In this study we analyze the B-cell response in murine yersiniosis. To this end, we determined whether polyclonal activation of B-lymphocytes occurs during infection of susceptible (BALB/c) and resistant (C57BL/6) mice with Y. enterocolitica 0:8 and compared the immunoglobulin (Ig) isotypes produced in response to the infection by the two strains. The number of splenic cells secreting nonspecific and specific immunoglobulins was determined by ELISPOT. The presence of anti-Yersinia antibodies in serum was detected by ELISA. In both strains, the number of specific Ig-secreting cells was relatively low. Polyclonal B-cell activation was observed in both strains of mice, and the greatest activation was observed in the BALB/c mice, mainly for lgG(1)- and IgG(3)- secreting cells. The C57BL/6 mice showed a predominance of IgG(2a)-secreting cells. The peak production of anti-Yersinia IgG antibodies in the sera of BALB/c mice was seen on the 28th day after infection. The greatest increase in IgM occurred on the 14th day. A progressive increase of specific IgG antibodies was observed in C57BL/6 mice up to the 28th day after infection while IgM increased on the 21st day after infection. The production of specific IgA antibodies was not detected in either BALB/c or C57BL/6 mice. We conclude that polyclonal. activation of B lymphocytes occurs in both the Yersinia resistant and Yersinia-susceptible mice and that the more intense activation of B lymphocytes observed in the susceptible BALB/c mice does not enhance their resistance to Y. enterocolitica infection.

Biodisponibilidade de fontes orgânicas e inorgânicas de zinco em ovinos

This research was done to compare the effects of different zinc sources and doses in the Santa Ines sheep diet. Forty lambs at weaning, with 18,4kg BW were randomly allotted and fed 10 treatments: 1- base diet without zinc supplementation; 2- base diet + 200mg Zn/kg of DM as zinc oxide; 3- base diet + 400mg Zn/kg of DM as zinc oxide; 4- base diet + 600mg Zn/kg of DM as zinc oxide; 5- base diet + 200mg Zn/kg of DM as amino acid zinc; 6- base diet + 400mg Zn/kg of DM as amino acid zinc; 7- base diet + 600mg Zn/kg of DM as amino acid zinc; 8- base diet + 200mg Zn/kg of DM as proteinato zinc; 9- base diet + 400mg Zn/kg of DM as proteinato zinc; 10- base diet + 600mg Zn/kg of DM as proteinato zinc. The animals were weighed and sampled for blood zinc analysis, phosphatase alkaline analysis and immunoglobulins G and M analysis. At the end of the experiment liver samples were collected to study the zinc hepatic levels. There was no difference in phosphatase alkaline levels, hepatic zinc levels and weight gain (P>0,05) but differences (P<0,05) in plasmatic zinc levels and in IgG and IgM levels were observed. Based on liver tissue uptake, estimates of the zinc bioavailability, through the regression equations showed that the organic and inorganic sources of zinc did not differ.

Aspectos imunogeneticos na pesquisa da etiologia da catarata senil

The immunogenetic study of senile cataract showed that the association with the HLA antigens and Ia antigens suggests an influence of the antigenicity for cataractogenesis, but not a peculiar specificity. However, the investigation of the immunoglobulins in lens suggest that the lens pathology may be produced by antibodies reacting with the lens capsule and consequently interfering with the lens epithelial metabolism, and contributes to the complex mechanism of cataract formation.