RESUMO
Several evidences have suggested the involvement of enzymes belonging to the phosphotransfer network, formed by creatine kinase (CK), pyruvate kinase (PK) and adenylate kinase (AK), as well the oxidative stress on the pathogenesis of infectious diseases associated with the central nervous system (CNS). Thus, the aim of this study was to evaluate whether listeriosis alters the brain energy metabolism and/or causes oxidative stress in different brain structures of cattle experimentally infected by Listeria monocytogenes. The cytosolic CK activity was inhibited in the cerebral cortex, cerebellum, brainstem and hippocampus of infected animals compared to uninfected animals, while the mitochondrial CK activity was increased. The PK activity was inhibited in all brain structures of infected animals, while the AK activity was unchanged. Na+, K+-ATPase activity decreased in the cerebral cortex, cerebellum and hippocampus of animals infected by L. monocytogenes. Regarding the oxidative strees variables, the cerebellum and brainstem of infected animals showed increased thiobarbituric acid reactive substances, while the catalase activity was inhibited. Glutathione S-transferarase was inhibited in the cerebral cortex and brainstem of infected animals, and it was increased in the cerebellum. L. monocytogenes was quantified in the liver (nâ¯=â¯5/5) and cerebral cortex (nâ¯=â¯4/5) of the infected cattle. Based on these evidences, the nucleocytoplasmic communication between CK isoenzymes was insufficient to avoid an impairment of cerebral bioenergetics. Moreover, the inhibition on brain PK activity caused an impairment in the communication between sites of ATP generation and ATP utilization. The lipid peroxidation and alteration on antioxidant status observed in some brain structures were also involved during the disease. In summary, these alterations contribute to disease pathogenesis linked to CNS during cattle listeriosis.
Assuntos
Adenilato Quinase/metabolismo , Encéfalo/enzimologia , Doenças dos Bovinos/enzimologia , Creatina Quinase/metabolismo , Listeria monocytogenes/fisiologia , Listeriose/veterinária , Piruvato Quinase/metabolismo , Adenilato Quinase/genética , Animais , Antioxidantes/metabolismo , Encéfalo/metabolismo , Encéfalo/microbiologia , Bovinos , Doenças dos Bovinos/metabolismo , Doenças dos Bovinos/microbiologia , Creatina Quinase/genética , Creatina Quinase Mitocondrial/genética , Creatina Quinase Mitocondrial/metabolismo , Metabolismo Energético , Listeriose/enzimologia , Listeriose/metabolismo , Listeriose/microbiologia , Oxidantes/metabolismo , Estresse Oxidativo , Fosforilação , Piruvato Quinase/genéticaRESUMO
The enzymatic activities of NTPDase, 5'-nucleotidase and adenosine deaminase (ADA) are important in regulating the concentration of adenine nucleotides, molecules known to be involved on platelet aggregation. Fasciolosis causes coagulation disorders that have not been completely elucidated. Taking into consideration the association between the purinergic system and hemostasis, this study aimed to evaluate the enzymatic activities of NTPDase (hydrolyze ATP and ADP), 5'-nucleotidase (hydrolyze AMP) and ADA (deamination of adenosine) in platelets from cattle experimentally infected by Fasciola hepatica on days 20, 40, 60 and 80 post-infection (PI). For this study, 10 healthy Friesian steers were separated into two groups: the group A (n = 5) was used as uninfected control, and the group B was composed of steers experimentally infected by F. hepatica (n = 5). The number of platelets did not differ between groups in the periods evaluated. Reduction of NTPDase (p < 0.05) hydrolysing ATP (days 20, 40 and 60 PI), and ADP (days 40, 60 and 80 PI), and on 5'-nucleotidase hydrolyzing AMP (days 40 and 60 PI) was observed. A reduction (p < 0.05) in ADA activity on day 20 PI, as well as an increase (p < 0.05) in ADA activity on days 40 and 60 PI was observed when compared to the control. Based on these results, we can conclude that ATP, ADP and AMP hydrolysis and adenosine deamination were altered in platelets of cattle infected by F. hepatica. Considering the importance of the purinergic system in hemostasis, it is believed that those changes may contribute to the coagulation impairment observed in acute fasciolosis described in the literature.
Assuntos
Adenosina Desaminase/sangue , Plaquetas/enzimologia , Doenças dos Bovinos/parasitologia , Fasciolíase/veterinária , Nucleotidases/sangue , Animais , Bovinos , Doenças dos Bovinos/sangue , Doenças dos Bovinos/enzimologia , Fasciola hepatica/fisiologia , Fasciolíase/sangue , Fasciolíase/enzimologia , Fezes/parasitologia , Fígado/parasitologia , Fígado/patologia , Masculino , Contagem de Ovos de Parasitas/veterinária , Contagem de Plaquetas/veterináriaRESUMO
The enzyme adenosine deaminase (ADA) is critical for modulating the immune system, and in the presence of zinc, its activity is catalyzed. The aim of this study was to evaluate the ADA activity in pancreas of cattle naturally infected by Eurytrema coelomaticum in relation to the results of zinc levels, pathological findings and parasite load. For this study 51 slaughtered cattle were used. The animals were divided into two groups: Group A consisting of animals naturally infected by E. coelomaticum (n=33) and Group B of uninfected animals (n=18). Blood and pancreas were collected of each animal for analysis of zinc and ADA, respectively. Infected cattle showed a reduction on seric levels of zinc, and decreased ADA activity in the pancreas (P>0.05). A positive correlation between zinc levels and ADA activity was observed. Thus, high parasite load and severity of histopathologic lesions affect the ADA activity in pancreas, as well as the zinc levels in serum of infected animals (negative correlation between these variables). Therefore, we can conclude that cattle infected by E. coelomaticum have low ADA activity in pancreas, which can be directly related to zinc reduction, responsible for ADA activation and catalyzes.
Assuntos
Adenosina Desaminase/metabolismo , Doenças dos Bovinos/metabolismo , Dicrocoeliidae/fisiologia , Pâncreas/parasitologia , Carga Parasitária/veterinária , Infecções por Trematódeos/veterinária , Zinco/sangue , Animais , Bovinos , Doenças dos Bovinos/enzimologia , Doenças dos Bovinos/parasitologia , Pâncreas/metabolismo , Espectrofotometria/veterinária , Infecções por Trematódeos/enzimologia , Infecções por Trematódeos/metabolismo , Infecções por Trematódeos/parasitologiaRESUMO
The aim of this study was to evaluate the oxidative stress in serum and liver and adenosine deaminase (ADA) activity of cattle experimentally infected by Fasciola hepatica. The group A consisted of five healthy animals (uninfected), and the group B was composed of five animals orally infected with 200 metacercariae of F. hepatica. On days 20, 40, 60 and 80 post-infection (PI) serum was collected to measure oxidative stress variables. On day 100 PI, animals were humanely euthanized and liver samples were collected. Infected animals showed lower (P < 0·05) seric ADA activities on days 40 and 60 PI but higher (P < 0·05) in the liver tissue compared with uninfected animals. Seric and hepatic reactive oxygen species (ROS) were higher (P < 0·05) in infected compared with uninfected animals. Hepatic thiobarbituric acid reactive substances were higher (P < 0·05) in infected animals. Catalase and glutathione S-transferase activities were lower in liver tissue of infected animals, while glutathione peroxidase was higher compared with uninfected (P < 0·05). In summary, we conclude that oxidative stress occurs in cattle experimentally infected by F. hepatica, mainly due to excessive ROS production in the course of fasciolosis, contributing to hepatic damage, and that increased in hepatic ADA activity may contribute to the inflammatory process.
Assuntos
Adenosina Desaminase/metabolismo , Doenças dos Bovinos/parasitologia , Fasciola hepatica , Fasciolíase/veterinária , Estresse Oxidativo/fisiologia , Adenosina Desaminase/sangue , Adenosina Desaminase/genética , Animais , Biomarcadores , Bovinos , Doenças dos Bovinos/enzimologia , Doenças dos Bovinos/metabolismo , Fasciolíase/enzimologia , Fasciolíase/parasitologia , Regulação Enzimológica da Expressão Gênica , Fígado/enzimologia , Fígado/patologia , Masculino , Espécies Reativas de OxigênioRESUMO
The most important regulators of tissue remodelling during ovarian follicular growth, development, ovulation and atresia are gonadotropins, steroid hormones, growth factors and different proteolytic enzymes. Matrix metalloproteinases (MMPs) such as collagenase or gelatinase (i.e. MMP-1, -8, -2 and -9) and associated tissue inhibitors of metalloproteinases (TIMP-1, -2, -3 and -4) control connective tissue remodelling during follicular rupture. In this study, we hypothesized that an imbalance in the MMP-TIMP system may be an intra-ovarian component that contributes to the pathogenesis of cystic ovarian disease (COD) in cows. Taking into account that the control of MMP activity by TIMPs could determine their effects in both physiological and pathological conditions, MMP and TIMP mRNA and protein expression was examined by real-time polymerase chain reaction (RT-PCR) and immunohistochemistry (IHC), respectively, in ovaries from control cows and cows with COD. Expression of mRNA encoding MMP-2, TIMP-1 and TIMP-2 was lower in follicular cysts than in control pre-ovulatory follicles, while the results by IHC showed this imbalance only for TIMP-2 protein expression. Additional analysis by zymography to evaluate the gelatinase activity of MMP-2 and MMP-9 demonstrated higher MMP-2 activity in follicular fluid (FF) of cysts than in FF of pre-ovulatory follicles. On the other hand, MMP-9 activity was increased in follicular cysts and absent in the FF of pre-ovulatory follicles. These findings suggest that the altered mRNA expression and activities of the MMP-TIMP system may be related to the failure in ovulation and follicular development observed in COD.
Assuntos
Doenças dos Bovinos/enzimologia , Metaloproteinases da Matriz/metabolismo , Cistos Ovarianos/veterinária , Inibidores Teciduais de Metaloproteinases/metabolismo , Animais , Western Blotting , Bovinos , Feminino , Processamento de Imagem Assistida por Computador , Imuno-Histoquímica , Reação em Cadeia da Polimerase em Tempo RealRESUMO
The aim of this study was to evaluate seric NTPDase and 5'nucleotidase activities of cattle naturally infected by Eurytrema coelomanticum, as well as to correlate them to histopathological lesions in the pancreas and the degree of parasitism. Blood samples and pancreas of 51 bovines were collected on a slaughterhouse in Southern Brazil: 33 from cattle naturally infected by E. coelomanticum (the Group A), and 18 from uninfected animals (the Group B). Infected animals showed an average of 532 parasites per pancreas. In the pancreatic histology, ducts displayed hyperplasia, stenosis, proliferation of fibrous tissue, and interstitial inflammatory infiltration of lymphocytes. The serum from infected animals showed an increase in NTPDase activity when ATP was used as substrate (P<0.001). For the ADP substrate, there was no difference between groups regarding NTPDase activity (P=0.37), as well as 5'-nucleotidase activity (P=0.27). Correlating NTPDase activity (ATP substrate) with the degree of histopathological lesions (rho=0.66, P<0.001) and the parasitic load on the pancreas (rho=0.65, P<0.001), a positive correlation was observed. Similar results were found between the degree of histopathological lesions and NTPDase activity (ADP substrate; rho=0.29, P=0.03), and 5'nucleotidase activity (rho=0.35, P=0.01). Based on the results of NTPDase and 5'nucleotidase enzymes in cattle naturally infected by E. coleomanticum, it is possible to suggest that these enzymes are involved in the modulation of inflammation, and they can act as markers of inflammatory response.
Assuntos
5'-Nucleotidase/sangue , Apirase/sangue , Doenças dos Bovinos/patologia , Dicrocoeliidae , Inflamação/veterinária , Pâncreas/parasitologia , Infecções por Trematódeos/veterinária , Matadouros , Animais , Antígenos CD , Biomarcadores/sangue , Brasil , Bovinos , Doenças dos Bovinos/enzimologia , Doenças dos Bovinos/parasitologia , Inflamação/enzimologia , Inflamação/patologia , Linfócitos , Pâncreas/patologia , Carga Parasitária , Infecções por Trematódeos/enzimologia , Infecções por Trematódeos/parasitologia , Infecções por Trematódeos/patologiaRESUMO
UNLABELLED: Rotaviruses (RVs) enter cells through different endocytic pathways. Bovine rotavirus (BRV) UK uses clathrin-mediated endocytosis, while rhesus rotavirus (RRV) employs an endocytic process independent of clathrin and caveolin. Given the differences in the cell internalization pathway used by these viruses, we tested if the intracellular trafficking of BRV UK was the same as that of RRV, which is known to reach maturing endosomes (MEs) to infect the cell. We found that BRV UK also reaches MEs, since its infectivity depends on the function of Rab5, the endosomal sorting complex required for transport (ESCRT), and the formation of endosomal intraluminal vesicles (ILVs). However, unlike RRV, the infectivity of BRV UK was inhibited by knocking down the expression of Rab7, indicating that it has to traffic to late endosomes (LEs) to infect the cell. The requirement for Rab7 was also shared by other RV strains of human and porcine origin. Of interest, most RV strains that reach LEs were also found to depend on the activities of Rab9, the cation-dependent mannose-6-phosphate receptor (CD-M6PR), and cathepsins B, L, and S, suggesting that cellular factors from the trans-Golgi network (TGN) need to be transported by the CD-M6PR to LEs to facilitate RV cell infection. Furthermore, using a collection of UK × RRV reassortant viruses, we found that the dependence of BRV UK on Rab7, Rab9, and CD-M6PR is associated with the spike protein VP4. These findings illustrate the elaborate pathway of RV entry and reveal a new process (Rab9/CD-M6PR/cathepsins) that could be targeted for drug intervention. IMPORTANCE: Rotavirus is an important etiological agent of severe gastroenteritis in children. In most instances, viruses enter cells through an endocytic pathway that delivers the viral particle to vesicular organelles known as early endosomes (EEs). Some viruses reach the cytoplasm from EEs, where they start to replicate their genome. However, other viruses go deeper into the cell, trafficking from EEs to late endosomes (LEs) to disassemble and reach the cytoplasm. In this work, we show that most RV strains have to traffic to LEs, and the transport of endolysosomal proteases from the Golgi complex to LEs, mediated by the mannose-6-phosphate receptor, is necessary for the virus to exit the vesicular compartment and efficiently start viral replication. We also show that this deep journey into the cell is associated with the virus spike protein VP4. These findings illustrate the elaborate pathway of RV entry that could be used for drug intervention.
Assuntos
Catepsinas/metabolismo , Doenças dos Bovinos/enzimologia , Doenças dos Bovinos/virologia , Endossomos/virologia , Doenças dos Macacos/enzimologia , Receptor IGF Tipo 2/metabolismo , Infecções por Rotavirus/veterinária , Rotavirus/fisiologia , Animais , Catepsinas/genética , Bovinos , Doenças dos Bovinos/genética , Doenças dos Bovinos/metabolismo , Endossomos/enzimologia , Endossomos/metabolismo , Macaca mulatta , Camundongos , Doenças dos Macacos/genética , Doenças dos Macacos/metabolismo , Doenças dos Macacos/virologia , Receptor IGF Tipo 2/genética , Rotavirus/genética , Infecções por Rotavirus/enzimologia , Infecções por Rotavirus/metabolismo , Infecções por Rotavirus/virologia , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/metabolismo , Internalização do VírusRESUMO
Cystic ovarian disease (COD) is an important cause of infertility in cattle, and ACTH has been involved in regulatory mechanisms related to ovarian function associated with ovulation, steroidogenesis, and luteal function. Here, we examined the localization of 11ß-hydroxysteroid dehydrogenase type 1 (11ßHSD1) and 11ßHSD2 proteins in the ovary of healthy cows and animals with spontaneous and ACTH-induced COD and the in vitro response of the follicular wall exposed to ACTH. After stimulation by ACTH, we documented changes in 11ßHSD expression and cortisol secretion by the follicular wall of large antral and follicular cysts. Follicular cysts showed a higher constitutive expression of both enzymes, whereas ACTH induced an increase in 11ßHSD1 in tertiary follicles and follicular cysts and a decrease in 11ßHSD2 in follicular cysts. Moderate expression of 11ßHSD1 was observed by immunohistochemistry in granulosa of control animals, with an increase (P < 0.05) from primary to secondary, tertiary, and atretic follicles. The level of immunostaining in theca interna was lower than that in granulosa. The expression of 11ßHSD2 was lower in the granulosa of primary follicles than in that of secondary, tertiary, and atretic follicles and was lower in the theca interna than in the granulosa. In ACTH-induced and spontaneously occurring follicular cysts, differences from controls were observed only in the expression of 11ßHSD1 in the granulosa, being higher (P < 0.05) than in tertiary follicles. These findings indicate that follicular cysts may be exposed to high local concentrations of active glucocorticoids and indicate a local role for cortisol in COD pathogenesis and in regulatory mechanisms of ovarian function.
Assuntos
11-beta-Hidroxiesteroide Desidrogenases/análise , Hormônio Adrenocorticotrópico/farmacologia , Doenças dos Bovinos/enzimologia , Cistos Ovarianos/veterinária , Ovário/enzimologia , Animais , Bovinos , Doenças dos Bovinos/patologia , Estradiol/sangue , Feminino , Células da Granulosa/enzimologia , Hidrocortisona/sangue , Imuno-Histoquímica , Microscopia Eletrônica de Varredura/veterinária , Cistos Ovarianos/induzido quimicamente , Cistos Ovarianos/enzimologia , Folículo Ovariano/enzimologia , Folículo Ovariano/metabolismo , Folículo Ovariano/patologia , Progesterona/sangue , Testosterona/sangue , Ultrassonografia/veterináriaRESUMO
In this study, we describe the interaction between Araçatuba virus (ARAV), a naturally occurring Brazilian vaccinia virus isolated from an outbreak at a dairy farm, and the host cell's signal transduction pathways. Even though ARAV infection led to phosphorylation of MAPKs MEK/ERK, JNK, and p38MAPK, genetic or pharmacological inhibition of these pathways had no impact on viral replication. We also provide evidence that ARAV stimulated the phosphorylation of Akt (PKB) at serine 473 (S473-P), a signaling event that is required for full activation of Akt during the infectious cycle. Furthermore, pharmacological inhibition of PI3K (LY294002) abrogated ARAV-induced Akt activation (S473-P) and affected early and late viral gene expression, which was followed by a decrease in virus yield (~1 log). Taken together, our data shed some light onto the biological differences between ARAV and vaccinia virus strain WR (VACV-WR), which could contribute, at least in part, to the low-virulence phenotype displayed by ARAV. Thus, while the requirement for the PI3K/Akt pathway for successful ARAV replication is also shared with VACV-WR and cowpox virus strain BR (CPXV-BR), ARAV showed a lower replicative capacity, as well as a smaller plaque-size phenotype after infection of A31 cells when compared to VACV-WR.
Assuntos
Doenças dos Bovinos/enzimologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Vaccinia virus/fisiologia , Vacínia/veterinária , Replicação Viral , Animais , Bovinos , Doenças dos Bovinos/virologia , Linhagem Celular , Interações Hospedeiro-Patógeno , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Vacínia/enzimologia , Vacínia/virologia , Vaccinia virus/genéticaRESUMO
The present investigation was intended to show a different immunohistochemical profile of matrix metalloproteinase-2 and Tissue inhibitor metalloproteinase-2 in bovine uteri with adenomyosis during follicular phase. Uterine samples of 32 cows in reproductive age were taken from the medial third of one of the uterine horns and grouped according to the adenomyosis degree (superficial and deep). Tissue sections (4 µm) were incubated overnight at 4°C with monoclonal antibody for matrix metalloproteinase-2 and Tissue inhibitor metalloproteinase-2. Staining intensities were evaluated in the luminal epithelium, ectopic and dystopic endometrial tissue (stroma, capillaries and glands), endometrial-myometrial border, myometrium, myometrial vessels (middle tunic and endothelium). The matrix metalloproteinase-2 expression was higher for deep adenomyosis samples, showing a differential mean reactivity in superficial endometrium, myometrial vessels, myometrium adjacent to adenomyotic focus and endometrial-myometrial border (P < 0.05). Moreover, matrix metalloproteinase-2 expression was higher in deep adenomyosis samples than that of Tissue inhibitor metalloproteinase-2 in almost all uterine structures analyzed (except for the endometrial and myometrial vessels and endometrial-myometrial border). The opposite was observed in the follicular phase, for both normal specimens and with superficial adenomyosis, where Tissue inhibitor metalloproteinase-2 expression was higher than that of matrix metalloproteinase-2. In conclusion, a differential pattern of matrix metalloproteinase-2 and Tissue inhibitor metalloproteinase-2 was observed in cow uteri with adenomyosis.
Assuntos
Doenças dos Bovinos/enzimologia , Endometriose/veterinária , Endométrio/patologia , Metaloproteinase 2 da Matriz/metabolismo , Miométrio/patologia , Inibidor Tecidual de Metaloproteinase-2/metabolismo , Animais , Bovinos , Doenças dos Bovinos/metabolismo , Doenças dos Bovinos/patologia , Endometriose/enzimologia , Endometriose/metabolismo , Endometriose/patologia , Endométrio/metabolismo , Feminino , Fase Folicular , Imuno-Histoquímica , Miométrio/metabolismoRESUMO
In the winter of 2008, cattle on a farm in the province of Neuquen, Argentina died from subacute and chronic liver fluke disease despite four previous treatments with Triclabendazole (TCBZ). In the spring of 2009, a preliminary efficacy test revealed good performance using nitroxynil, whereas TCBZ efficacy was only 18% by egg counts of Fasciola eggs in the faeces. Resistance to Fasciola hepatica to TCBZ has never been reported in South America, so in January of 2010 a controlled trial was conducted to confirm and to define the degree of resistance in this herd. In a clinical trial, the fluke egg output was monitored on Days 14 and 21 and serum enzymes gamma-glutamyl transpeptidase (GGT) and glutamic-oxaloacetic transaminase (GOT) on Days 0 and 21 in 36 calves treated with TCBZ or with closantel. The results showed a reduction of 100% in fluke egg output at Days 14 and 21 for closantel. The mean epg in the TCBZ-treated groups did not decrease. Because of the fact that in this study TCBZ treatment in cattle had no effect, even at double the recommended dose, it is highly indicative that resistance of F. hepatica against TCBZ is present on this farm. The GGT and GOT levels decreased in the closantel-treated group as a result of the treatment at 21 days after dosing. To evaluate the importance of TCBZ resistance in F. hepatica in Argentina, a study on more farms from endemic areas is needed.
Assuntos
Anti-Helmínticos/farmacologia , Benzimidazóis/farmacologia , Doenças dos Bovinos/parasitologia , Fasciola hepatica/crescimento & desenvolvimento , Fasciolíase/veterinária , Hepatopatias/veterinária , Salicilanilidas/farmacologia , Animais , Anti-Helmínticos/uso terapêutico , Argentina , Aspartato Aminotransferases/sangue , Benzimidazóis/uso terapêutico , Bovinos , Doenças dos Bovinos/tratamento farmacológico , Doenças dos Bovinos/enzimologia , Resistência a Medicamentos , Fasciolíase/tratamento farmacológico , Fasciolíase/enzimologia , Fasciolíase/parasitologia , Fezes/parasitologia , Hepatopatias/tratamento farmacológico , Hepatopatias/enzimologia , Hepatopatias/parasitologia , Contagem de Ovos de Parasitas/veterinária , Salicilanilidas/uso terapêutico , Triclabendazol , gama-Glutamiltransferase/sangueRESUMO
Cattle in western Uruguay that were eating Solanum bonariense developed periodic episodes of ataxia, hypermetria, hyperesthesia, head and thoracic limb extension, opisthotonus, nystagmus, and falling to the side or backward. Similar clinical signs were experimentally reproduced in cattle by administration of S. bonariense via rumen cannula at a dose of 1,024 g/kg body mass. No significant gross lesions were observed in field cases or experimentally induced cases. Spontaneous and induced histologic lesions were similar and included vacuolation, degeneration, and loss of Purkinje cells. Axonal spheroids, microcavitations, and other changes of wallerian-type degeneration in cerebellar white matter were also observed. Ultrastructural changes included increased number of electron-dense residual storage bodies in membrane-bound vesicles in affected Purkinje cells, and similar vesicles and mitochondria in axonal spheroids. No histologic lesions were detected in the other examined tissues. The Purkinje-cell swelling and vacuolation with subsequent cerebellar degeneration are suggestive of Purkinje-cell specific toxin that produces abnormal lysosome function and cell specific axonal transport. This is the first report of S. bonariense toxicity.
Assuntos
Encéfalo/patologia , Doenças dos Bovinos/etiologia , Doenças Cerebelares/veterinária , Doenças Neurodegenerativas/veterinária , Plantas Tóxicas , Solanum , Fosfatase Alcalina/sangue , Animais , Aspartato Aminotransferases/sangue , Bovinos , Doenças dos Bovinos/enzimologia , Doenças dos Bovinos/patologia , Doenças Cerebelares/enzimologia , Doenças Cerebelares/etiologia , Doenças Cerebelares/patologia , Feminino , Histocitoquímica/veterinária , Masculino , Microscopia Eletrônica de Transmissão/veterinária , Doenças Neurodegenerativas/enzimologia , Doenças Neurodegenerativas/etiologia , Doenças Neurodegenerativas/patologia , Células de Purkinje/patologia , Células de Purkinje/ultraestrutura , gama-Glutamiltransferase/sangueRESUMO
As part of a general project aimed at elucidating the initiation of mucin-type O-glycosylation in helminth parasites, we have characterized a novel ppGalNAc-T (UDP-N-acetyl-D-galactosamine:polypeptide N-acetylgalactosaminyltransferase) from the cestode Echinococcus granulosus (Eg-ppGalNAc-T1). A full-length cDNA was isolated from a library of the tissue-dwelling larval stage of the parasite, and found to code for a 654-amino-acid protein containing all the structural features of ppGalNAc-Ts. Functional characterization of a recombinant protein lacking the transmembrane domain showed maximal activity at 28 degrees C, in the range 6.5-7.5 pH units and in the presence of Cu2+. In addition, it transferred GalNAc to a broad range of substrate peptides, derived from human mucins and O-glycosylated parasite proteins, including acceptors containing only serine or only threonine residues. Interestingly, the C-terminal region of Eg-ppGalNAc-T1 bears a highly unusual lectin domain, considerably longer than the one from other members of the family, and including only one of the three ricin B repeats generally present in ppGalNAc-Ts. Furthermore, a search for conserved domains within the protein C-terminus identified a fragment showing similarity to a recently defined domain, specialized in the binding of organic phosphates (CYTH). The role of the lectin domain in the determination of the substrate specificity of these enzymes suggests that Eg-ppGalNAc-T1 would be involved in the glycosylation of a special type of substrate. Analysis of the tissue distribution by in situ hybridization and immunohistochemistry revealed that this transferase is expressed in the hydatid cyst wall and the subtegumental region of larval worms. Therefore it could participate in the biosynthesis of O-glycosylated parasite proteins exposed at the interface between E. granulosus and its hosts.