Purification and partial characterization of an aminopeptidase from the midgut tissue of Dysdercus peruvianus

The surface of midgut cells in Hemiptera is ensheathed by a lipoprotein membrane (the perimicrovillar membrane), which delimits a closed compartment with the microvillar membrane, the so-called perimicrovillar space. In Dysdercus peruvianus midgut perimicrovillar space a soluble aminopeptidase maybe involved in the digestion of oligopeptides and proteins ingested in the diet. This D. peruvianus aminopeptidase was purified to homogeneity by ion-exchange chromatography on an Econo-Q column, hydrophobic interaction chromatography on phenyl-agarose column and preparative polyacrylamide gel electrophoresis. The results suggested that there is a single molecular species of aminopeptidase in D. peruvianus midgut. Molecular mass values for the aminopeptidase were estimated to be 106 kDa (gel filtration) and 55 kDa (SDS-PAGE), suggesting that the enzyme occurs as a dimer under native conditions. Kinetic data showed that D. peruvianus aminopeptidase hydrolyzes the synthetic substrates LpNA, RpNA, A beta NA and AsnMCA (K(m)s 0.65, 0.14, 0.68 and 0.74 mM, respectively). The aminopeptidase activity upon LpNA was inhibited by EDTA and 1,10-phenanthroline, indicating the importance of metal ions in enzyme catalysis. One partial sequence of BLAST-identified aminopeptidase was found by random sequencing of the D. peruvianus midgut cDNA library. Semi-quantitative RT-PCR analysis showed that the aminopeptidase genes were expressed throughout the midgut epithelium, in the epithelia of V1, V2 and V3. Malphigian tubules and fat body, but it was not expressed in the salivary glands. These results are important in furthering our understanding of the digestive process in this pest species. (c) 2010 Elsevier Inc. All rights reserved.

Morphology of glandular stomach (Ventriculus glandularis) and muscular stomach (Ventriculus muscularis) of the partrigde Rhynchotus rufescens

Vinte perdizes Rhynchotus rufescens adultas foram utilizadas para estudo morfológico do proventrículo e ventrículo gástricos da perdiz Rhynchotus rufescens. Os materiais foram coletados e os comprimentos do proventrículo e do ventrículo gástricos foram avaliados. Para o estudo histológico, fragmentos dos estômagos foram corados pelas técnicas de ácido periódico de Schiff (PAS) e tricromo de Masson. O proventrículo gástrico é alongado, com formato fusiforme direcionado no sentido craniocaudalmente e para a esquerda, e apresenta um comprimento médio 3,20cm nas fêmeas e 3,65cm nos machos. Histologicamente, o proventrículo gástrico é composto por vários lobos e glândulas. A mucosa é formada por epitélio cúbico, sendo bastante pregueada. O ventrículo gástrico tem o formato de uma lente biconvexa, com comprimento médio de 4,30cm nas fêmeas e 4,35cm nos machos. A mucosa é formada por pregas revestidas por células cilíndricas e pelo muco formador da cutícula. Há criptas na base das pregas. em seguida, há uma lâmina própria e uma espessa camada muscular lisa, que se encontra direcionada de acordo com o formato do ventrículo gástrico. A serosa é constituída por uma densa porção de tecido conjuntivo, entremeado por algumas células musculares lisas.

Effects of feed restriction on yield, retail cuts and tissue composition of carcass of Saanen kids

Com o objetivo de estudar o efeito da restrição alimentar sobre as características de carcaça de caprinos leiteiros, realizou-se um experimento utilizando 27 cabritos machos Saanen distribuídos nos tratamentos 0 (alimentação à vontade), 30 ou 60% de restrição. Os animais apresentavam 5 kg de PV inicial e foram abatidos quando atingiram 20 kg de PV. Foram avaliados os rendimentos comercial e biológico, os cortes comerciais, a composição tecidual da perna, a área de olho-de-lombo (AOL) e a compacidade da carcaça. Utilizaram-se o delineamento inteiramente ao acaso e a análise de regressão em função da restrição alimentar. A restrição alimentar provocou redução do peso da carcaça e dos cortes comerciais, aumento da proporção do pescoço e diminuição da proporção do lombo. A proporção de ossos aumentou e a do tecido muscular e da gordura total diminuiu com o aumento da restrição. A proporção de gordura subcutânea diminuiu com o aumento da restrição alimentar. A área de olho-de-lombo (AOL) e a compacidade da carcaça foram afetadas pela restrição alimentar, mas ambas as medidas podem ser utilizadas para predizer a proporção de músculo da carcaça. O tratamento com 30% de restrição alimentar não prejudica a qualidade da carcaça de cabritos leiteiros.

Toxoplasma gondii: Evaluation of an intranasal vaccine using recombinant proteins against brain cyst formation in BALB/c mice

The purpose of this work was to evaluate protective activity against brain cyst formation in BALB/c mice intranasally vaccinated with recombinant proteins from Toxoplasma gondii. The recombinant proteins rROP2, rGRA5 and rGRA7 were used in vaccine preparation. Thirty-three female mice were divided into three groups, these animals received two doses by intranasal route at days 0 and 21 as follows; group 1 (G1, n = 11) received 12.5 mu g of each recombinant protein plus 0.5 mu g of cholera toxin, group 2 (G2, n = 11) received phosphate buffer saline (PBS) plus 0.5 mu g of cholera toxin, and group 3 (G3, n = 11) received PBS only. At challenge day (day 33) three animals from each group were euthanatized for IgA measure from intestine. Mice were infected orally with 50 cysts from the VEG strain at day 33. At challenge day the G1 animals had high immunoglobulin A levels, however, they only showed IgG antibody titers against rROP2 and rGRAT Animals from G1 also exhibited strong resistance to cyst formation compared with the control group (G3, P < 0.05). However, we did not observe differences in protection against brain cyst formation between G1 and G2 (P > 0.1). These results indicate that intranasal immunization in BALB/c mice with recombinant proteins rROP2, rGRA5 and rGRA7 associated with cholera toxin induced partial protection, when compared with G3, against tissue cyst formation after oral infection with tissue cysts from T gondii. (c) 2007 Elsevier B.V. All rights reserved.

Acute 2,4-dichlorophenoxyacetic acid intoxication in broiler chicks

The acute toxicity of 2,4-dichlorophenoxyacetic acid (2,4-D), a herbicide, was studied in chicks dosed with 100, 300, 500, or 600 mg 2,4-D/kg BW, by the oral route. Clinical, laboratory, and histopathological methods were used as indicators of toxicity. After acute exposure, the herbicide decreased motor activity and induced muscular weakness and motor incoordination; decreased weight gain; increased serum creatine kinase (CK) and alkaline phosphatase (AP) activities and serum uric acid (UA), creatinine (CR), and total proteins (TP) levels; and did not change serum aspartate aminotransferase (AST) or alanine aminotransferase (ALT) activities. These changes were time-and dose-dependent and reversible. The LD50 (lethal dose 50%) calculated for oral 2,4-D in chicks was 420 mg/kg BW (385 to 483). Chromatographic analysis of the serum of the intoxicated chicks showed the presence of the herbicide; the amount found was dose-and time-dependent, increasing from 2 to 8 h after exposure and decreasing afterwards. Histopathological post-mortem studies conducted on intoxicated chicks showed hepatic (vacuolar degeneration of the hepatocytes), renal (tubular nephrosis), and intestinal (hemorragic) lesions. Taken together, the observed alterations mainly reflected kidney and muscle tissue damage, although hepatic toxicity may also have occurred after acute 2,4-D intoxication.

Analytical approach to the metallomic of Nile tilapia (Oreochromis niloticus) liver tissue by SRXRF and FAAS after 2D-PAGE separation: Preliminary results

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

Fat body in some genera of leaf-cutting ants (Hymenoptera : Formicidae). Proteins, lipids and polysaccharides detection

The comparative histochemical analysis of the fat body of workers belonging to the basal species Cyphomyrmex rimosus and Mycetarotes parallelus and to derived species Acromyrmex disciger and Atta laevigata revealed that this tissue is constituted mainly by cells denominated trophocytes and oenocytes. The trophocytes of all species studied here were characterized mainly by the proteins and lipids synthesis and storage, being the derived species the ones who have presented higher quantity of lipids in the trophocytes when compared to the trophocytes of basal species. In workers M. parallelus and A. laevigata, besides proteins and lipids, there has being observed the presence of polysaccharides, however, in C. rimosus and A. disciger these elements were detected in lower quantities. The histochemical studies of the oenocytes of basal and derived species revealed significant presence of proteins as well as lipids in these cells. In the oenocytes of derived species A. disciger and A. laevigata a higher quantity of lipidic inclusions has being observed, when compared to the basal species. (c) 2005 Elsevier Ltd. All rights reserved.

Ultrastructural detection of proteins, lipids and carbohydrates in oocytes of Amblyomma triste (Koch, 1844) (Acari; Ixodidae) during the vitellogenesis process

The ovary of the tick Ainblyomma triste is classified as panoistic, which is characterized by the presence of oogonia without nurse and follicular cells. The present study has demonstrated that the oocytes in all developmental stages (I-IV) are attached to the ovary through a pedicel, a cellular structure that synthesizes and provides carbohydrate, lipids and proteins supplies for the oocytes during the vitellogenesis process. The lipids are deposited during all oocyte stages; they are freely distributed as observed in stages II, III and IV or they form complexes with other elements. The proteins are also deposited in all stages of the oocytes, however, in lower concentration in the stage IV. There is carbohydrate deposition from oocytes in the stage II as well as in stages III and IV. In addition, the present work has demonstrated that the oocyte yolk of A. triste has a glycolipoprotein nature and the elements are deposited in the following sequence: firstly the lipids and proteins, and finally the carbohydrates. (c) 2007 Elsevier Ltd. All rights reserved.

Cyclosporin A causes impairment of the ventral prostate tissue structure of Wistar rats

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

Assessment of bovine biomaterials containing bone morphogenetic proteins bound to absorbable hydroxyapatite in rabbit segmental bone defects

OBJETIVO: Avaliar a capacidade osteo-regenerativa de dois biomateriais utilizando um modelo de defeito segmentar efetuado nas diáfises do rádio de coelhos. MÉTODOS: O defeito direito foi preenchido com pool de proteínas morfogenéticas ósseas (pBMPs) e hidroxiapatita em pó ultrafina absorvível (HA) combinada com matriz óssea inorgânica desmineralizada e colágeno, derivados do osso bovino (Grupo A). O defeito esquerdo foi preenchido com matriz óssea desmineralizada bovina com pBMPs e hidroxiapatita em pó ultrafina absorvível (Grupo B). em ambos os defeitos utilizou-se membrana reabsorvível de cortical bovina desmineralizada para reter os biomateriais no defeito ósseo e guiar a regeneração tecidual. Os coelhos foram submetidos à eutanásia aos 30, 90 e 150 dias após a cirurgia. Foram efetuados exames radiográficos, tomográficos e histológicos em todos os espécimes. RESULTADOS: Aos 30 dias de pós-cirúrgico, o osso cortical desmineralizado foi totalmente reabsorvido em ambos os grupos. A HA tinha reabsorvido nos defeitos do Grupo A, mas persistiu nos do Grupo B. Uma reação de corpo estranho foi evidente com ambos os produtos, porém mais pronunciada no Grupo B. Aos 90 dias os defeitos do grupo B tinham mais formação óssea que os do Grupo A. Entretanto, aos 150 dias após a cirurgia, nenhum tratamento havia promovido o completo reparo do defeito. CONCLUSÃO: Os biomateriais testados contribuíram pouco ou quase nada para a reconstituição do defeito segmentar.

Mecanismos celulares e moleculares que controlam o desenvolvimento e o crescimento muscular

O músculo estriado esquelético é formado pela associação de fibras musculares com a matriz extracelular. Esse tecido possui alta plasticidade e o conhecimento das características morfológicas, da miogênese, e da dinâmica do crescimento é importante para o entendimento da morfofisiologia bem como para a seleção de animais visando a melhoria na produção de carne. A maioria dos músculos estriados originam-se de células precursoras do mesoderma a partir dos somitos do embrião e o controle da diferenciação ocorre pela ação de fatores indutores ou inibidores. Um grupo de fatores transcricionais, pertencentes à família MyoD tem um papel central na diferenciação muscular. Coletivamente chamados de Fatores de Regulação Miogênica (MRFs), são conhecidos quatro tipos: MyoD, myf-5, miogenina e MRF4. Esses fatores ligam-se à seqüências de DNA conhecidas como Ebox (CANNTG) na região promotora de vários genes músculo-específicos, levando à expressão dos mesmos. As células embrionárias com potencial para diferenciação em células musculares (células precursoras miogênicas) expressam MyoD e Myf-5 e são denominadas de mioblastos. Essas células proliferam, saem do ciclo celular, expressam miogenina e MRF4, que regulam a fusão e a diferenciação da fibra muscular. Uma população de mioblastos que se diferencia mais tardiamente, as células miossatélites, são responsáveis pelo crescimento muscular no período pós natal, que pode ocorrer por hiperplasia e hipertrofia das fibras. As células satélites quiescentes não expressam os MRFs, porém, sob a ação de estímulos como fatores de crescimento ou citocinas, ocorre a ativação desse tipo celular que prolifera e expressa os MRFs de maneira similar ao que ocorre com as células precursoras miogênicas durante a miogênese. Os mecanismos de crescimento muscular são regulados pela expressão temporal dos (MRFs), que controlam a expressão dos genes relacionados com o crescimento muscular.

Reaction of the human gingival tissue to different suture materials used in periodontal surgery.

The gingival reaction to 4 different suture materials used in periodontal surgery was studied in 36 patients. The gingiva was sutured prior to surgery and biopsies were taken at 3, 7 and 14 days to observe the tissue reaction. The histological examination showed that silk caused the most intense and longest inflammatory response. Polyester and perlon provoked shorter, less intense tissue reactions than silk, and nylon caused the least inflammatory response, with earlier tissue repair.

Acute, subchronic and chronic 2,4-dichlorophenoxyacetic acid (2,4-D) intoxication in rats

The acute, subchronic and chronic toxicities of 2,4- dichlorophenoxyacetic acid (2,4-D) were studied in rats. Animals were exposed acutely (600 mg/kg), subchronically (200 ppm for 30 d) and chronically (200 ppm for 180 d) to 2,4-D by the oral route. Clinical, laboratory and histopathological methods were used as indicators of toxicity. After acute exposure, the herbicide decreased locomotor activity and induced ataxia, sedation, muscular weakness (mainly of the hind quarters) and gasping for breath; increased aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH), alkaline phosphatase (AP), amylase activities and creatinine levels; decreased total protein (TP) and glucose levels; and increased hematocrit values. Subchronic and chronic 2,4-D exposures did not induce overt clinical signs or symptoms of intoxication. However, subchronic herbicide exposure increased AST activity and albumin and hematocrit values, and chronic exposure increased AST, AP and LDH activities, decreased amylase and glucose levels, but did not change hematocrit values. Chromatographic analysis of the serum of chronically exposed rats showed the presence of the herbicide; the amount found (3.76 ± 1.16 mg/ml) suggested the absence of 2,4-D accumulation within the body. Although macroscopic or histopathological lesions were not observed in acutely, subchronically or chronically 2,4-D exposed rats, the laboratory data obtained suggest tissue injuries after dosing, since the results are considered early indicators of primarily hepatic and muscle tissue damage.

Localization of total proteins and lactate dehydrogenase in hamster epididymis

The concentration of total protein measured by photocolorimetric methodology and reported as units per 100 mg of tissue decreased from the initial segment to the cauda epididymidis of the Golden hamster, being significant the numeric difference observed between these two regions. This observation was related with an increased synthesis and secretion of proteins to the lumen in proximal segments of the epididymidis duct, mainly in initial segment, as proposed for other rodents. LDH activity was higher in initial segment and distal cauda than in the caput and corpus epididymidis, although no significant differences in mean values had been observed. The high LDH activity observed in initial segment and cauda epididymidis of hamster had been related to an expressive epithelium metabolic activity presented in these regions. This metabolic activity help to guarantee the survival of spermatozoa stored in cauda epididymidis. Furthermore, lower LDH activity noted in the caput and corpus epididymidis might be related with a progressive reduction of glycolysis in initial maturation step of spermatozoa mainly verified in corpus epididymidis. © 2007 Sociedad Chilena de Anatomía.

Acute phase proteins in canine lymphoma during antineoplastic chemotherapy

The lymphoma is the main hematopoietic tumor in dogs and it is characterized by the proliferation of cells from lymphoid tissue, histiocytes and its precursors. Animals with lymphoma often show changes in biochemical and hematological parameters such as non-regenerative normochromic normocytic anemia, hemolytic anemia, hypocalcaemia and monoclonal gammopathy. The development of tumor can cause alterations in serum concentrations of acute phase proteins (APPs), consequent of hepatocytes stimulus by cytokines of inflammatory action. This study aimed to quantify and qualify APPs in dogs with lymphoma, at diagnosis time and during the time of chemotherapy sessions. After syneresis, centrifugation and fractioning the serum samples of 10 healthy and 10 dogs with lymphomas, the proteins fractions were separated by polyacrilamide gel electrophoresis (SDS-PAGE) and its concentrations were determined by computer densitometry. Between 18 and 30 proteins were separated by eletrophoresis, with molecular weights ranging from 18 to 245 kDa (kilodaltons). The alpha-1-glicoprotein acid (AGP) and transferrin serum concentration showed significantly higher in dogs with lymphoma, when compared with healthy dogs at diagnosis. The alpha-1-antitripsin (AAT) serum concentrations showed significantly higher in healthy dogs, when compared with dogs with lymphoma at diagnosis. The dogs with lymphoma the albumin did not appear as negative APP. On the other hand, transferrin appeared as positive AAP at diagnosis time and during the chemotherapy sessions. Healthy dogs had AAT serum concentrations significantly higher when compared to dogs with lymphoma at diagnosis. So, in this trial, it is suggested that this protein has been shown as a negative APP in the dogs with lymphoma. These dogs presented significantly higher AGP serum concentrations, in relation to healthy dogs at diagnosis, evidencing this protein APP positive behavior in neoplasm.