RESUMO
Canine mammary tumours (CMTs) are the most common neoplasms affecting female dogs. There is an urgent need for molecular biomarkers that can detect early stages of the disease in order to improve accuracy of CMT diagnosis. The aim of this study was to examine whether caveolin-1 (Cav-1) and matrix metalloproteinase 14 (MMP14) are associated with CMT histological malignancy and invasion. Sixty-five benign and malignant CMT samples and six normal canine mammary glands were analysed using quantitative reverse transcription-polymerase chain reaction. Cav-1 and MMP14 genes were highly expressed in CMT tissues compared to normal tissues. Cav-1 especially was overexpressed in malignant and invasive CMT tissues. When a CMT cell line was cultured on fluorescent gelatin-coated coverslips, localisation of Cav-1 was observed at invadopodia-mediated degradation sites of the gelatin matrix. These findings suggest that Cav-1 may be involved in CMT invasion and that the markers may be useful for estimating CMT malignancy.
Assuntos
Caveolina 1/metabolismo , Doenças do Cão/metabolismo , Regulação Neoplásica da Expressão Gênica/fisiologia , Neoplasias Mamárias Animais/metabolismo , Metaloproteinase 14 da Matriz/metabolismo , Animais , Neoplasias da Mama/metabolismo , Estudos de Casos e Controles , Caveolina 1/genética , Linhagem Celular Tumoral , Cães , Feminino , Regulação Enzimológica da Expressão Gênica , Humanos , Metaloproteinase 14 da Matriz/genéticaAssuntos
Mastite Bovina/microbiologia , Infecções por Mycoplasma/veterinária , Mycoplasma/isolamento & purificação , Animais , Bovinos , Contagem de Células/veterinária , Feminino , Glândulas Mamárias Animais/microbiologia , Leite/citologia , Leite/microbiologia , Infecções por Mycoplasma/microbiologiaAssuntos
Mastite Bovina/epidemiologia , Leite/microbiologia , Infecções por Mycoplasma/veterinária , Mycoplasma/isolamento & purificação , Animais , Antibacterianos/uso terapêutico , Bovinos , Farmacorresistência Bacteriana , Feminino , Japão , Mastite Bovina/tratamento farmacológico , Mycoplasma/efeitos dos fármacos , Infecções por Mycoplasma/tratamento farmacológico , Infecções por Mycoplasma/epidemiologia , PrevalênciaRESUMO
Solid tumors are composed of a heterogeneous population of cells surviving in various concentrations of oxygen. In a hypoxic environment, tumor cells generally up-regulate glycolysis and, therefore, generate more lactate that must be expelled from the cell through proton transporters to prevent intracellular acidosis. Monocarboxylate transporter 1 (MCT1) is a major proton transporter in mammalian cells that transports monocarboxylates, such as lactate and pyruvate, together with a proton across the plasma membrane. Melanocytic neoplasia occurs frequently in dogs, but the prognosis is highly site-dependent. In this study, 50 oral canine melanomas, which were subdivided into 3 histologic subtypes, and 17 ocular canine melanocytic neoplasms (14 melanocytomas and 3 melanomas) were used to examine and compare MCT1 expression. Immunohistochemistry using a polyclonal chicken anti-rat MCT1 antibody showed that most oral melanoma exhibited cell membrane staining, although there were no significant differences observed among the 3 histologic subtypes. In contrast, the majority of ocular melanocytic tumors were not immunoreactive. Additionally, we documented the presence of a 45-kDa band in cell membrane protein Western blots, and sequencing of a reverse transcriptase polymerase chain reaction band of expected size confirmed its identity as a partial canine MCT1 transcript in 3 oral tumors. Increased MCT1 expression in oral melanomas compared with ocular melanocytic tumors may reflect the very different biology between these tumors in dogs. These results are the first to document canine MCT1 expression in canine tumors and suggest that increased MCT1 expression may provide a potential therapeutic target for oral melanoma.
Assuntos
Doenças do Cão/metabolismo , Neoplasias Oculares/veterinária , Melanoma/veterinária , Transportadores de Ácidos Monocarboxílicos/metabolismo , Neoplasias Bucais/veterinária , Simportadores/metabolismo , Animais , Cães , Neoplasias Oculares/metabolismo , Regulação Neoplásica da Expressão Gênica , Melanoma/metabolismo , Transportadores de Ácidos Monocarboxílicos/genética , Neoplasias Bucais/metabolismo , Simportadores/genética , Regulação para CimaRESUMO
This study investigated the distribution and expression of monocarboxylate transporter 1 (MCT1) in the livers of pre-ruminant calves and adult bovines (bulls and cows), using different molecular biological techniques. Reverse transcription-polymerase chain reaction (RT-PCR) verified the presence of mRNA encoding for MCT1 in both pre-ruminant and adult bovine livers. Immunohistochemically, MCT1 was clearly demonstrated on the sinusoidal surfaces of bovine hepatocytes but its expression varied widely between pre-ruminants and adult bovines. In pre-ruminants, a faint hepatocellular expression of MCT1 was observed in a few hepatocytes, whereas an intense immunoreactive staining for MCT1 was shown in the majority of adult bovine hepatocytes. Western blot analysis also confirmed the results of the immunohistochemistry. Quantitative immunoblotting, as estimated by densitometric analysis, showed that the level of MCT1 in the liver of adult bovines was 8-9-fold greater (P<0.01) than that in pre-ruminant calf livers although no significant differences were detected between bulls and cows. The results demonstrated that MCT1 may play a crucial role in the transport of propionate in bovine liver, suggesting that MCT1 expression may be influenced by developmental and metabolic regulations.
Assuntos
Bovinos/crescimento & desenvolvimento , Bovinos/metabolismo , Fígado/metabolismo , Transportadores de Ácidos Monocarboxílicos/metabolismo , Simportadores/metabolismo , Envelhecimento , Animais , Feminino , Regulação da Expressão Gênica , MasculinoRESUMO
In this study, we investigated the tissue distribution and expression of monocarboxylate transporter 1 (MCT1) along the gastrointestinal tract of sheep. Western blot analysis suggested the presence of MCT1 as a 43-kDa protein in immunoblots of membranes from the various tissues examined. The results of Western blotting were further confirmed by immunohistochemical studies, which revealed intense immunoreactivity for the MCT1 protein in the forestomach (rumen, reticulum and omasum) and large intestine (caecum, proximal and distal colon). Moderate reactivity, however, was detected in the abomasum, while no immunoreactivity could be seen in any regions of the small intestine examined. Furthermore, MCT1 was expressed at the mRNA level as determined by reverse transcriptase polymerase chain reaction (RT-PCR), which showed a band of the expected size (300 bp) in all tissues examined. From these results we concluded that MCT1 protein is highly expressed and distributed in the stomach and large intestine of sheep suggesting that MCT1 may play a significant role in the transport of short chain fatty acids and their metabolites in the gastrointestinal tract of ruminants.
Assuntos
Trato Gastrointestinal/metabolismo , Expressão Gênica/fisiologia , Transportadores de Ácidos Monocarboxílicos/metabolismo , Ovinos , Simportadores/metabolismo , Animais , Western Blotting/veterinária , Regulação da Expressão Gênica/fisiologia , Imuno-Histoquímica/veterinária , Masculino , Peso Molecular , Transportadores de Ácidos Monocarboxílicos/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Simportadores/genéticaRESUMO
In the present study the expression and distribution of monocarboxylate transporter 1 (MCT1) along the gastrointestinal tract (rumen, reticulum, omasum, abomasum, duodenum, jejunum, ileum, cecum and colon) of calves were investigated on both mRNA and protein levels. The expression of MCT1 protein and its distribution were determined by Western blotting and immunohistochemical staining, respectively by using antibody for MCT1. MCT1 protein was visualized as a 43-kDa band on immunoblots of the membrane proteins prepared from the various regions examined, and it was more highly expressed in forestomach and large intestine than in abomasum and small intestine. With the use of reverse transcriptase-polymerase chain reaction, mRNA encoding for MCT1 was demonstrated in the different tissues examined. The immunohistochemical study confirmed the Western blot findings and showed strong MCT1 immunopositive staining in the stratified squamous epithelia of the forestomach as well as the epithelial cells lining the digestive tract in the cecum, proximal colon, and distal colon. The results suggest that MCT1 may play a role in the transport of SCFA and their metabolites in the gastrointestinal tract of bovines.
Assuntos
Bovinos/metabolismo , Trato Gastrointestinal/metabolismo , Expressão Gênica/fisiologia , Transportadores de Ácidos Monocarboxílicos/metabolismo , Simportadores/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Bovinos/genética , Masculino , Dados de Sequência Molecular , Transportadores de Ácidos Monocarboxílicos/genética , RNA Mensageiro/metabolismo , Simportadores/genéticaRESUMO
Immunological interaction between the host and Helicobacter pylori seems to play a critical role in follicular formation in gastric mucosa. We reported H. pylori-induced follicular gastritis model using neonatally thymectomized mice. In this study, we investigated the involvement of various cytokines in this model. BALB/c mice were thymectomized on the third day after birth (nTx). At 6 weeks old, these mice were orally infected with H. pylori. Histological studies showed that follicular formation occurred from 8 weeks after the infection and that most of the infiltrating lymphocytes were CD4(+) and B cells. Neutrophils increased transiently at 1 week after the infection. Gamma interferon, interleukin-7 (IL-7), and IL-7 receptor were expressed in the stomach of the nTx mice irrespective of the infection. In contrast, expressions of the tumor necrosis factor alpha, IL-4 and lymphotoxin-alpha genes were remarkably upregulated by the infection. Our findings suggest that follicular formation may require cooperative involvement of a Th2-type immune response, tumor necrosis factor alpha and lymphotoxin-alpha in addition to the Th1-type immune response in H. pylori-induced gastritis in nTx mice.
Assuntos
Citocinas/genética , Expressão Gênica , Infecções por Helicobacter/imunologia , Helicobacter pylori/imunologia , Estômago/imunologia , Animais , Animais Recém-Nascidos , Feminino , Citometria de Fluxo , Infecções por Helicobacter/patologia , Técnicas Imunoenzimáticas , Tecido Linfoide/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Timectomia , Fatores de TempoRESUMO
A full-length cDNA encoding a phenol UDP-glucuronosyltransferase was isolated by plaque hybridization, RT-PCR and 5'-RACE from a cDNA library prepared from the bovine liver. The deduced amino acid sequence (529 amino acid residues) has A signal sequence (23 amino acid residues) at the amino terminus and a transmembrane-anchoring domain (17 amino acid residues) at the carboxyl terminus. The encoded protein has a potential asparagine-linked glycosylation site (Asn291). The cloned cDNA was named bovUGT1A6 on the basis of the amino acid similarity. BovUGT1A6 cloned in the pAAH5 expression vector was transformed into Saccharomyces cerevisiea AH22 cells to obtain an active 54-kDa bovUGT1A6 enzyme. The expressed enzyme represented UDP-glucuronosyltransferase activities toward 1-naphthol and 4-methylumbelliferone, confirming that the isolated cDNA is an isoform of bovine phenol UDP-glucuronosyltransferase. Microsomal UDP-glucuronosyltransferase activity toward 1-naphthol in the bovine kidney cortex was found to be higher than that in the liver and other organs, and mRNA of bovUGT1A6 was more strongly detected in the kidney on Northern blotting analysis. These results suggest that the bovine kidney, which strongly expresses bovUGT1A6, is a significant organ for xenobiotics glucuronidation.
Assuntos
Clonagem Molecular , DNA Complementar/genética , Glucuronosiltransferase/genética , Sequência de Aminoácidos , Animais , Northern Blotting , Bovinos , DNA Complementar/análise , Expressão Gênica , Glucuronosiltransferase/biossíntese , Himecromona/metabolismo , Córtex Renal/enzimologia , Fígado/enzimologia , Masculino , Microssomos/enzimologia , Dados de Sequência Molecular , Naftóis/metabolismo , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
1. To analyse the mechanisms of muscle layer-dependent inhibition of porcine myometrial contractility by 5-hydroxytryptamine (5-HT), the effects of 5-HT, 5-carboxamidotryptamine(5-CT), 5-methoxytryptamine (5-MeOT), forskolin and cyclic adenosine 3', 5'-monophosphate (cyclic AMP) analogues on spontaneous and stimulant-induced contractions were examined in longitudinal (LM) and circular muscles (CM). In addition, accumulation of cyclic AMP by 5-HT and distribution of 5-HT(7) receptors in LM and CM layers were compared using biochemical and molecular approaches. 2. 5-HT receptor agonists inhibited the spontaneous contractions of LM and CM (5-CT>5-HT>5-MeOT), but CM was more sensitive than was LM. The inhibition by the agonists was antagonized by methiothepin (100 nM). 3. Carbachol-, high-K(+)-, histamine- and Ca(2+)-induced contractions were inhibited by 5-HT with different responses (CM>LM). Even in the presence of 3-isobutyl-1-methylxanthine (IBMX), the inhibition by 5-HT in the CM was still more conspicuous than that in the LM. 4. Compared with the CM, the inhibition of spontaneous contraction by forskolin, dibutyryl-cyclic AMP and 8-bromo-cyclic AMP was marked in the LM. 5. 5-HT (1 nM - 1 microM) increased the cyclic AMP in both muscle layers, but the increment in the CM was higher than that in the LM whether IBMX was present or not. 6. LM and CM layers contained a single class of [(3)H]-5-CT binding sites with a similar K(d) value (0.21 - 0.24 nM). However, B(max) (5-HT(7) receptor concentration) in the CM (120.6 fmol mg(-1) protein) was higher than that in the LM (30.4 fmol mg(-1) protein). 7. The molecular study (reverse transcription polymerase chain reaction) demonstrated the expression of 5-HT(7) receptor mRNA in the CM was higher than that in the LM. 8. These results suggest that the muscle layer-dependent difference in inhibition by 5-HT is not restricted to spontaneous contraction but applies to various contractions in the porcine myometrium. Different inhibition of the contractility by 5-HT is caused by muscle layer-related accumulation of cyclic AMP (CM>LM), due to smooth muscle-layer dependent distribution (CM>LM) of 5-HT(7) receptors.
Assuntos
Contração Muscular/fisiologia , Miométrio/metabolismo , Receptores de Serotonina/metabolismo , Serotonina/análogos & derivados , 5-Metoxitriptamina/farmacologia , Animais , AMP Cíclico/metabolismo , Feminino , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Músculo Liso/metabolismo , Miométrio/efeitos dos fármacos , Receptores de Serotonina/efeitos dos fármacos , Serotonina/metabolismo , Serotonina/farmacologia , Agonistas do Receptor de Serotonina/farmacologia , Inibidores Seletivos de Recaptação de Serotonina/farmacologia , SuínosRESUMO
Nosema tyriae n.sp. was found in 63% of a population of Cinnabar moth larvae (Tyria jacobaeae). The infection was found in the gut wall, silk glands, and fat body and was probably generalized but appeared to be of low pathogenicity. Merogony and sporogony were by binary fission of diplokaryotic stages. Fresh spores were elongate, slightly pointed at the anterior end, and measured 4.7 x 2.0 microm. Ultrastructural features of special interest were 20-nm tubules connecting the surface of sporonts with host cell cytoplasm and, in the spores, a deeply domed polar sac, polaroplast consisting of closely packed longitudinally arranged membranes and loosely packed horizontally arranged membranes, and 10.5-14 coils of the polar tube in a single rank. The 16S rRNA genes of N. tyriae and Nosema bombycis from silkworms, Bombyx mori, differed by only six nucleotides and N. tyriae spores gave a moderately positive reaction with a monoclonal antibody raised to N. bombycis. N. tyriae was infective to B. mori but was less virulent than N. bombycis. However, no amplification product was obtained by PCR using N. tyriae DNA and primers considered to be specific for N. bombycis. Also, the spores of the two species are of entirely different shapes. A second diplokaryotic microsporidium, Nosema sp., found as a light infection in only one of the larvae had much smaller developmental stages and spores measuring 3.8 x 2.0 microm (fixed). Ultrastructurally it was distinguished by an abundance of dense membranes in cytoplasmic vesicles in both meronts and sporonts. Spores with up to 15 coils of the polar tube in irregular clusters or with about 12 coils in a single rank were observed in the tissues fixed from the one larva infected with this parasite. As this larva had been kept with N. tyriae-infected larvae for a few days before examination, it is possible that the two types of spores resulted from a double infection.
Assuntos
Mariposas/parasitologia , Nosema , Animais , Sequência de Bases , DNA de Protozoário , Dados de Sequência Molecular , Nosema/genética , Nosema/ultraestruturaRESUMO
Bisphenol A, an environmental oestrogenic chemical, was found to conjugate highly with glucuronic acid in male rat liver microsomes studied in vitro. In the various isoforms tested (1A1, 1A3, 1A5, 1A6, 1A7 and 2B1), glucuronidation of bisphenol A and of diethylstilboestrol, a synthetic crystalline compound possessing oestrogenic activity and known to be glucuronidated by liver microsomes, was catalysed by an isoform of UDP-glucuronosyltransferase (UGT), namely UGT2B1, which glucuronidates some endogenous androgens. UGT activity towards bisphenol A in liver microsomes and in UGT2B1 expressed in yeast AH22 cells (22.9 and 0.58 nmol/min per mg of microsomal proteins respectively) was higher than that towards diethylstilboestrol (75.0 and 4.66 pmol/min per mg of microsomal proteins respectively). UGT activities towards both bisphenol A and diethylstilboestrol were distributed mainly in the liver but were also observed at substantial levels in the kidney and testis. Northern blot analysis disclosed the presence of UGT2B1 solely in the liver, and about 65% of the male rat liver microsomal UGT activities towards bisphenol A were absorbed by the anti-UGT2B1 antibody. These results indicate that bisphenol A, in male rat liver, is glucuronidated by UGT2B1, an isoform of UGT.
Assuntos
Estrogênios não Esteroides/metabolismo , Glucosiltransferases/metabolismo , Glucuronatos/metabolismo , Fenóis/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Compostos Benzidrílicos , Primers do DNA , Feminino , Glucosiltransferases/genética , Glucosiltransferases/imunologia , Masculino , Dados de Sequência Molecular , RatosRESUMO
Asp446 in rat UDP-glucuronosyltransferase (UGT), UGT1A6, is an essential amino acid residue for its enzymatic activity (H. Iwano et al. Biochem. J. 325, 587-591, 1997). The role of Asp446 in UGT1A6 was investigated by comparing some properties of UGT mutant proteins that have a single mutation (D446K, D446E, D446N, D446Q, D446A, and D446T). These mutants, except D446K, had catalytic activities toward 1-naphthol and 4-methylumbelliferone. The UGT activities of D446E and D446N were about half of that of the wild type, and the activities of the other mutants were only about 1/5-1/10 of that of the wild type. The Km values for 1-naphthol of these mutants were similar to that of the wild type, while the values for UDP-glucuronic acid were slightly higher. The mutants were unstable in a low-pH buffer solution and were dramatically inactivated by heat treatments. Interestingly, after 30 min of treatment at 37 degrees C in the presence of UDP-glucuronic acid, the UGT activities of all functional mutants were elevated. These results suggest that Asp446 is an indispensable residue for folding a functional conformation of rat UGT1A6 by cooperation with UDP-glucuronic acid.
Assuntos
Ácido Aspártico/fisiologia , Glucuronosiltransferase/genética , Glucuronosiltransferase/metabolismo , Sequência de Aminoácidos , Substituição de Aminoácidos , Animais , Cromatografia Líquida de Alta Pressão , Sequência Conservada/genética , Estabilidade Enzimática , Fluorometria , Expressão Gênica , Immunoblotting , Microssomos/metabolismo , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Ratos , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/genética , Alinhamento de Sequência , Temperatura , Fatores de TempoRESUMO
Inhibitory effects of uridine diphosphate on the enzymatic activity of UDP-glucuronosyltransferase (UGT) were investigated. Pyrimidine nucleotides such as UDP, UTP and cytidine diphosphate reduced the activity of rat purified UGT (phenol UGT) to about 10%, 48% and 46% of the control, respectively, at the same concentration as a donor substrate, UDP-glucuronic acid. Purine nucleotides, uridine monophosphate, glucuronic acid and some UDP-sugars were only slightly inhibitory toward the transferase. Similar effects were observed in the expressed UGT (UGT1A6; corresponding to phenol UGT) in yeast cells and rat liver microsomal membrane-binding UGT, indicating that uracil and diphosphate residues are essential for the UDP inhibition. Interestingly, 2'-deoxy UDP was found to be a less effective inhibitor (about 50% inhibition) than UDP on the purified, the expressed (UGT1A6 and UGT2B1) and microsomal membrane-binding UGTs. These results indicate that not only uracil and diphosphate residues but also 2'-hydroxyl residue of UDP ribose participates in the interactions between UDP and UDP-glucuronosyltransferase.
Assuntos
Inibidores Enzimáticos/farmacologia , Glucuronosiltransferase/antagonistas & inibidores , Difosfato de Uridina/farmacologia , Animais , Clonagem Molecular , Glucuronosiltransferase/biossíntese , Glucuronosiltransferase/metabolismo , Ratos , Leveduras/metabolismoRESUMO
In the rat intestine, UDP-glucuronosyltransferase (UGT) isoforms were highly induced by oral administration of 2-naphthoflavone, as shown by intestinal UGT activity toward 1-naphthol (1-NA). The greatest increase in UGT activity occurred in the duodenum. Using UGT1A6 cDNA as a probe, we obtained three types of clones corresponding to UGT1A2, UGT1A6 and UGT1A7, in a ratio of 1:1:8, from a cDNA library constructed from the 2-naphthoflavone-treated rat intestine. The induction of each isoform was evaluated by means of Northern blotting with isoform-specific probes. The mRNAs of UGT1A6 (glucuronizing various phenolic xenobiotics) and the mRNAs of UGT1A7 (glucuronizing the ultimate carcinogenic metabolite of benzo(a)pyrene) were expressed constitutively and were highly induced in the duodenum and proximal jejunum. S1 mapping showed that induction of the isoforms of the UGT1 family was more pronounced in the liver than in the small intestine and that UGT1A7 was the major UGT1 isoform in the small intestine of vehicle-treated rats and in that of 2-naphthoflavone-treated rats. These results indicate that, in rats, UGT1A7 is expressed constitutively and is particularly inducible in the small intestine. In the light of these results, we believe that the UGT1A7 isoform would play an important role in glucuronidation in the small intestinal mucosa of rats.
Assuntos
Glucuronosiltransferase/biossíntese , Mucosa Intestinal/enzimologia , Intestino Delgado/enzimologia , beta-Naftoflavona/administração & dosagem , Administração Oral , Processamento Alternativo/efeitos dos fármacos , Processamento Alternativo/genética , Animais , Northern Blotting , DNA Complementar/isolamento & purificação , Indução Enzimática/efeitos dos fármacos , Glucuronosiltransferase/genética , Mucosa Intestinal/efeitos dos fármacos , Intestino Delgado/efeitos dos fármacos , Isoenzimas/biossíntese , Isoenzimas/genética , Masculino , RNA Mensageiro/análise , Ratos , Ratos Wistar , Endonucleases Específicas para DNA e RNA de Cadeia Simples/metabolismoRESUMO
An amino acid residue, Asp446, was found to be essential for the enzymic activity of UDP-glucuronosyltransferase (UGT). We obtained a rat phenol UGT (UGT1*06) cDNA (named Ysh) from male rat liver by reverse-transcription (RT)-PCR using pfu polymerase. A mutant Ysh having two different bases, A1337G and G1384A (named Ysh A1337GC1384A), that result in two amino acid substitutions, D446G and V462M, was obtained by RT-PCR using Taq polymerase. Ysh was expressed functionally in microsomes of Saccharomyces cerevisiae strain AH22. However, the expressed protein from YshA1337GG1384A had no transferase activity. Two other mutant cDNAs with YshA1337G having one changed base, A1337G, resulting in one amino acid substitution, D446G, and YshG1384A having a changed base, G1384A, resulting in an amino acid substitution, V462M, were constructed and expressed in the yeast. The expressed protein from YshG1384A (named YshV462M) exhibited enzymic activity, but the one from YshA1337G (named YshD446G) did not show any activity at all. Asp446 was conserved in all UGTs and UDP-galactose:ceramide galactosyltransferases reported, suggesting that Asp446 plays a critical role in each enzyme.
Assuntos
Ácido Aspártico/metabolismo , Glucuronosiltransferase/metabolismo , Animais , Ácido Aspártico/genética , DNA Complementar , Glucuronosiltransferase/química , Glucuronosiltransferase/genética , Masculino , Dados de Sequência Molecular , Mutação , Ratos , Ratos Wistar , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/genética , Especificidade por SubstratoRESUMO
Phenol UDP-glucuronosyltransferase activity was highly induced in the microsomes of the kidney medulla of rats by beta-naphthoflavone treatment. In the medulla, phenol UDP-glucuronosyltransferase and its mRNA were greatly increased in both immunoblotting and Northern blot analyses following beta-naphthoflavone treatment of the rats. In untreated rat kidneys, phenol UDP-glucuronosyltransferase was detected by immunohistochemical analysis only in proximal convolution tubular cells located in the cortex. After beta-naphthoflavone treatment of the rats, UDP-glucuronosyltransferase appeared in the epithelial cells in the straight portion of the distal tubules located in the medulla. In conclusion, the medullary distal tubular cells have high latent glucuronidation activity and are thought to play an important role in drug excretion.
Assuntos
Glucuronosiltransferase/biossíntese , Medula Renal/enzimologia , Fenóis/metabolismo , beta-Naftoflavona/farmacologia , Animais , Hidrocarboneto de Aril Hidroxilases/biossíntese , Indução Enzimática , Masculino , Fenol , Ratos , Ratos WistarRESUMO
The prefrontal cortex (PFC) has anatomical and functional relationships with the striatum. In a previous study we showed that dopamine (DA) turnover in the PFC of rats is enhanced during the performance of a discrimination task. In the present study, we used an in vivo microdialysis method to examine whether DAergic activity in the striatum could also be altered by the discrimination task. The results showed a substantial and sustained suppression of DAergic activity during and after the discriminative behaviour. The fact that the discriminative performance induced opposite changes in DAergic activity in the striatum and the PFC is consistent with the results of biochemical studies, suggesting that the suppressed DA turnover in the striatum may be induced by the enhanced DAergic activity in the PFC during the discrimination task.
Assuntos
Corpo Estriado/fisiologia , Aprendizagem por Discriminação/fisiologia , Dopamina/fisiologia , Córtex Pré-Frontal/fisiologia , Ácido 3,4-Di-Hidroxifenilacético/metabolismo , Análise de Variância , Animais , Dopamina/metabolismo , Ácido Homovanílico/metabolismo , Masculino , Microdiálise , Ratos , Ratos Endogâmicos F344RESUMO
Acetylcholine (ACh) release in the hippocampus of Wistar strain rats with permanent bilateral occlusion of the common carotid arteries was examined during a discrimination learning task using a microdialysis method. Such occlusion resulted in obvious impairment of the discrimination performance. The state, the basal value and released patterns, of ACh in the hippocampus differed in the sham-operated control and the experimental group, while ACh release was elevated during the dialysis experiment in both groups. These findings suggested that the bilateral occlusion produced persistent learning deficits from an early stage after the operation and that the impaired discrimination learning performance might be related to the diffusion of ACh in the hippocampus.
Assuntos
Acetilcolina/metabolismo , Isquemia Encefálica/metabolismo , Discriminação Psicológica/fisiologia , Hipocampo/metabolismo , Aprendizagem/fisiologia , Animais , Modelos Animais de Doenças , Masculino , Ratos , Ratos WistarRESUMO
To investigate regulation of the septo-hippocampal cholinergic system by dopaminergic inputs to the septum in rats which performed a discrimination learning task, an in vivo microdialysis method with the dual-probe approach was used. Rats were trained to discriminate between lamp-on and -off states under an operant-type learning procedure. After stable discriminative behavior was established, dialysis probes were implanted into the hippocampus and the lateral septum area of each rat. The concentration of dopamine (DA) in the septum rapidly increased within 20 min after the beginning of a learning session. However, another group of rats trained on a similar but non-discriminative task showed no such increase. The concentration of acetylcholine (ACh) in the hippocampus was significantly enhanced during the learning session and rapidly returned to the basal value after the session, but showed a delayed and diminished increase in the non-discrimination group. These results suggest that DAergic inputs to the septum may be involved in control of the septo-hippocampal cholinergic system which is of importance for discrimination learning behavior.