RESUMO
We propose a bioinformatics pipeline in which we use an ESTs database to predict and validate single-nucleotide polymorphisms (SNPs) directly linked to gene-coding regions at the HLA class I genes (HLA-A, HLA-B and HLA-C). Annotation originated from our analysis revealed various classes of possible new variations that may indicate possible new alleles. Thus, bioinformatics pipelines seem to be useful approaches to help screening for novel genetic variations at the HLA panel, and further analysis will foster this aim to provide celerity at the massive analysis of data currently generated in large-scale high-throughput experiments.
Assuntos
Bases de Dados Genéticas , Etiquetas de Sequências Expressas , Antígenos HLA/genética , Polimorfismo de Nucleotídeo Único , Sequência de Bases , Biologia Computacional/métodos , Éxons/genética , Antígenos HLA-A/genética , Antígenos HLA-B/genética , Antígenos HLA-C/genética , Humanos , Reprodutibilidade dos Testes , Alinhamento de Sequência/métodos , Homologia de Sequência do Ácido NucleicoRESUMO
BACKGROUND: Fructose 1,6-biphosphate (FBP) has been shown to exert therapeutic effects in models of ischemia-reperfusion in organs other than the liver. This study compared FBP and University of Wisconsin (UW) solution during cold storage and reperfusion, among mitochondria of adult male Wistar rat livers. METHODS: Adult male Wistar rats were assigned to two groups according to the preservation solution used; UW or FBP Aspartate transaminase (AST), alanine transferase (ALT); and lactic dehydrogenase (LDH) were measured in samples of the storage solution obtained at 2, 4 and 6 hours of preservation. After 6 hours of cold storage, we reperfused the liver, taking blood samples to measure AST, ALT, LDH, and throbarbituric acid reactive substances (TBARS). Hepatic fragments were processed for histologic analysis; for determinations of TBARS, catalase, and nitric oxide as well as for mitochondrial evaluation by infrared spectroscopy. RESULTS: During cold preservation, levels of AST and LDH in the storage solution were lower among the FBP group, but after reperfusion, serum levels of AST, ALT, and LDH were higher in this group, as was catalase activity. TBARS and nitric oxide were comparable between the groups. In the UW group there was a higher amide I/amide II ratio than in the FBP group, suggesting an abnormal protein structure of the mitochondrial membrane. No signs of preservation injury were observed in any liver biopsy, but sinusoidal congestion was present in livers preserved with FBP. CONCLUSION: FBP showed a protective effect for preservation during cold storage seeming to protect the mitochondrial membrane although it did not prevent reperfusion injury.
Assuntos
Frutosedifosfatos/administração & dosagem , Fígado , Mitocôndrias Hepáticas/efeitos dos fármacos , Preservação Biológica , Animais , Frutosedifosfatos/farmacologia , Masculino , Mitocôndrias Hepáticas/patologia , Ratos , Ratos Wistar , SoluçõesRESUMO
BACKGROUND AND PURPOSE: D-Fructose-1,6-bisphosphate (FBP) is an intermediate in the glycolytic pathway, exerting pharmacological actions on inflammation by inhibiting cytokine production or interfering with adenosine production. Here, the possible antinociceptive effect of FBP and its mechanism of action in the carrageenin paw inflammation model in mice were addressed, focusing on the two mechanisms described above. EXPERIMENTAL APPROACH: Mechanical hyperalgesia (decrease in the nociceptive threshold) was evaluated by the electronic pressure-metre test; cytokine levels were measured by elisa and adenosine was determined by high performance liquid chromatography. KEY RESULTS: Pretreatment of mice with FBP reduced hyperalgesia induced by intraplantar injection of carrageenin (up to 54%), tumour necrosis factor alpha (40%), interleukin-1 beta (46%), CXCL1 (33%), prostaglandin E(2) (41%) or dopamine (55%). However, FBP treatment did not alter carrageenin-induced cytokine (tumour necrosis factor alpha and interleukin-1 beta) or chemokine (CXCL1) production. On the other hand, the antinociceptive effect of FBP was prevented by systemic and intraplantar treatment with an adenosine A(1) receptor antagonist (8-cyclopentyl-1,3-dipropylxanthine), suggesting that the FBP effect is mediated by peripheral adenosine acting on A(1) receptors. Giving FBP to mice increased adenosine levels in plasma, and adenosine treatment of paw inflammation presented a similar antinociceptive mechanism to that of FBP. CONCLUSIONS AND IMPLICATIONS: In addition to anti-inflammatory action, FBP also presents an antinociceptive effect upon inflammatory hyperalgesia. Its mechanism of action seems dependent on adenosine production but not on modulation of hyperalgesic cytokine/chemokine production. In turn, adenosine acts peripherally on its A(1) receptor inhibiting hyperalgesia. FBP may have possible therapeutic applications in reducing inflammatory pain.
Assuntos
Analgésicos/farmacologia , Frutosedifosfatos/farmacologia , Hiperalgesia/tratamento farmacológico , Inflamação/tratamento farmacológico , Adenosina/metabolismo , Animais , Cromatografia Líquida de Alta Pressão , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Hiperalgesia/fisiopatologia , Inflamação/fisiopatologia , Masculino , Camundongos , Medição da Dor , Receptor A1 de Adenosina/metabolismoRESUMO
Isolated liver perfusion has been used to evaluate the beneficial effects of several agents. In the present study, we developed a model using a recipient rat to reperfuse harvested livers in an ex situ, in vivo recirculating system. A total of 25 reperfusion procedures using adult male Wistar rats as donors and recipients were done. The preservation of the livers was performed with University of Wisconsin solution for 6 hours. Thereafter, the liver was reperfused with blood from another rat. We believe that the model presented herein offers an alternative method to evaluate early hepatocellular damage or hepatic microcirculation.
Assuntos
Circulação Hepática , Sistema Porta/fisiologia , Traumatismo por Reperfusão/fisiopatologia , Reperfusão/métodos , Animais , Modelos Animais de Doenças , Masculino , Modelos Biológicos , Sistema Porta/patologia , Sistema Porta/fisiopatologia , Ratos , Ratos Wistar , Traumatismo por Reperfusão/patologiaRESUMO
The temperature and pH effects on the equilibrium of a blood plasma model have been studied on the basis of artificial neural networks. The proposed blood plasma was modeled considering two important metals, calcium and magnesium, and six ligands, namely, alanate, carbonate, citrate, glycinate, histidinate and succinate. A large data set has been used to simulate different concentrations of magnesium and calcium as a function of temperature and pH and these data were used for training the neural network. The proposed model allowed different types of analyses, such as the effects of pH on calcium and magnesium concentrations, the competition between calcium and magnesium for ligands and the effects of temperature on calcium and magnesium concentrations. The model developed was also used to predict how the variation of calcium concentration can affect magnesium concentrations. A comparison of neural network predictions against experimental data produced errors of about 3%. Moreover, in agreement with experimental measurements (Wang et al. in Arch. Pathol. 126:947-950, 2002; Heining et al. in Scand. J. Clin. Lab. Invest. 43:709-714, 1983), the artificial neural network predicted that calcium and magnesium concentrations decrease when pH increases. Similarly, the magnesium concentrations are less sensitive than calcium concentrations to pH changes. It is also found that both calcium and magnesium concentrations decrease when the temperature increases. Finally, the theoretical model also predicted that an increase of calcium concentrations will lead to an increase of magnesium concentration almost at the same rate. These results suggest that artificial neural networks can be efficiently applied as a complementary tool for studying metal ion complexation, with especial attention to the blood plasma analysis.
Assuntos
Cálcio/sangue , Magnésio/sangue , Redes Neurais de Computação , Humanos , Concentração de Íons de Hidrogênio , Ligantes , Compostos Organometálicos/sangue , TemperaturaRESUMO
Fructose-1,6-bisphosphate (FBP) has been reported to have a protective effect on liver injury following ischemic/reperfusion periods because it maintains ATP levels during cold preservation. In the present study, we evaluated the effects of addition of FBP to storage solutions for cold liver preservation during 12 or 36 hours. Adult male Wistar rats were randomly divided into three experimental groups. The hepatic perfusion and preservation were performed with these solutions: UW; UW plus 10 mmol/L FBP; and FBP 10 mmol/L (FBPS) alone. The biochemical measurements of AST and ALT were performed on samples of the cold storage solution after 12- or 36-hour preservation. UW and FBPS solutions showed similar preservation grades at 12 hours. Addition of 10 mmol/L of FBP to UW solution induced liver injury and a poor preservation grade during 12 or 36 hours. UW solution was better than FBPS after 36 hours preservation. UW solution continues to offer a superior performance for liver preservation during long times; however, FBPS may be an alternative for short cold preservation times.
Assuntos
Frutosedifosfatos/farmacologia , Fígado , Soluções para Preservação de Órgãos , Adenosina , Alopurinol , Animais , Glutationa , Insulina , Fígado/efeitos dos fármacos , Masculino , Modelos Animais , Preservação de Órgãos/métodos , Rafinose , Ratos , Ratos WistarRESUMO
Fructose-1,6-bisphosphate (FBP) has been reported to have a protective effect on liver injury following ischemic/reperfusion periods. FBP maintains ATP levels and thereby cellular energy metabolism, which is important to the liver during cold preservation. In the present study, we evaluated the effects of FBP on the composition of storage solutions for cold liver preservation. Adult male Wistar rats were randomly divided into three experimental groups. Hepatic perfusion and preservation were performed with UW, UW plus 10 mmol/L FBP (UWM), and FBP 10 mmol/L (FBPS) alone solutions. Biochemical measurements of AST, ALT, and TBARS were performed on samples of the cold storage solution at 0, 12, 18, and 24 hours preservation. FBPS and UW solutions showed similar preservation grades during 18 hours. Addition of 10 mmol/L of FBP to UW solution induced liver injury and a poor preservation grade. FBP appears to protect the liver from injury caused by free radicals when the preservation time is less than 18 hours. Therefore, FBP may exert a protective effect for the preservation of livers during cold storage, and could represent an important component of new cold storage solutions.
Assuntos
Frutosedifosfatos , Transplante de Fígado/fisiologia , Fígado , Soluções para Preservação de Órgãos , Adenosina , Alanina Transaminase/análise , Alopurinol , Animais , Glutationa , Insulina , Fígado/fisiologia , Masculino , Rafinose , Ratos , Ratos Wistar , Substâncias Reativas com Ácido Tiobarbitúrico/análiseRESUMO
It has been suggested that the serotonin transporter (5-hydroxytryptamine-transporter or 5-HTT) may be involved in the pathogenesis of affective disorders. Recently, Collier et al. (1996) found that the frequency of the low-activity short variant (s) of the 5-HTT-linked polymorphic region (5-HTTLPR) was higher among patients with affective disorders than in normal controls. However, since the observed level of significance was not high, they suggest that these findings should be replicated in independent samples. We have analyzed 86 unrelated patients (47 with bipolar disorder and 39 with schizophrenia) and 98 normal controls from the Brazilian population for the 5-HTTLPR. Statistical analysis revealed that the genotypes (LL, Ls, ss) as well as the estimated allele frequencies (L,s) did not differ significantly among the three studied groups or between bipolar and normal controls. In addition, although not statistically significant, the genotype ss in our sample was less frequent among our bipolar patients than in our normal controls (12.8% versus 16.3%) which is the opposite of what was found by Collier et al. (24% versus 18%) in the European study. Although it will be important to extend the present analysis in a larger sample, our preliminary results suggest that the 5-HTTLPR does not seem to play a major role in the genetics of bipolar and schizophrenic disorders at least in this group of Brazilian psychiatric patients.