Biomedical subjects

Isabel Freitas

Publications and source records attributed to Isabel Freitas.

11 recordsLinked to original sources

PubMed2006

Evaluation of Rana snk esculenta blood cell response to chemical stressors in the environment during the larval and adult phases.

The assessment of the biological effects on aquatic vertebrate species is frequently employed to monitor water pollution, as it provides significant information on bioavailability and actual concentration levels. In anamniote vertebrates (fish and amphibians), significant correlations have been observed between exposure to contaminants - both natural and experimental - and blood modification. We investigated the changes in some circulating blood cell parameters of green frog (Rana snk esculenta) tadpoles and adults collected at two sample rice fields, one heavily polluted and the other relatively unpolluted. The frequency of eosinophilic leucocytes, mitotic, anucleated and micronucleated erythrocytes was evaluated also regarding the haemopoietic/haemocatheretic and NOS expression of the liver. Haematological indicators in polluted samples were found to be significantly different from controls as regards both larval and adult exposure, and provided information on long-term background pollution of the habitats under investigation. The population of the polluted area showed evident effects of chronic exposure to contaminants, to a degree which could lead to sub-lethal alterations of their health status. The general nature of responses to this kind of stress emphasizes the role of amphibian peripheral blood as a sensitive indicator regarding contamination in aquatic environments.

Animals↗

PubMed2005

[Idiopathic central precocious puberty in girls in Bahia, Brazil].

INTRODUCTION: Although the therapeutic effects of GnRH analogues (GnRHa) in central precocious puberty (CPP) have been established, clinical aspects may vary according to racial groups. OBJECTIVE AND METHODS: To describe the profile of 175 girls from Bahia, Brazil, treated with GnRHa for idiopathic CPP. RESULTS: The mulatto racial group predominated in the sample. The chronological age at puberty was 6.3 +/- 0.1 years. At diagnosis, height was 2.2 +/- 0.1 SD and body mass index (BMI) was 1.5 +/- 0.1 SD (> 2 SD in 32%), this one, inversely associated with age at onset of puberty (R = -0.20, p = 0.008). At treatment, chronological and bone ages were 8.4 +/- 0.1 years and 10.3 +/- 0.1 years, respectively. At the end of treatment (n = 52) age, BMI and height were 10.6 +/- 0.1 years, 1.6 +/- 0.2 SD and 1.9 +/- 0.2 SD, respectively. CONCLUSION: Clinical presentation and evolution were similar to other studies, independently of racial characteristics. In this sample, treatment with GnRHa was not associated to significant weight gain.

Brazil↗

PubMed2005

Autofluorescence spectroscopy of rat liver during experimental transplantation procedure. An approach for hepatic metabolism assessment.

Ischemia-reperfusion injury, a major cause of organ metabolic alterations and consequent dysfunction in liver transplantation, could be overcome by optimizing organ preservation procedures. The potential of autofluorescence analysis was investigated with the aim to define parameters suitable for in vivo monitoring tissue functionality. Spectrofluorometric analysis was performed on explanted rat livers during cold storage, under standard (4 degrees C University of Wisconsin medium for 20 h) and purposely damaging (4 degrees C Eurocollins medium for 20, 43 and 72 h) preservation conditions, and reperfusion (rewarming-reoxygenation). For both preservation conditions, cold hypoxia caused a signal amplitude increase, mainly attributable to NAD(P)H, and a spectral shape modification, ascribable to changes in the relative contributions of NAD(P)H and flavins, as a result of the tissue reduced state enhancement. Upon rewarming-reoxygenation the autofluorescence signal decreased with a rate depending on the preservation conditions. The time constant changed according to the extent of the liver functionality impairment, as assessed by conventional biochemical and histochemical analyses, thus providing a parameter exploitable for an in situ, real time monitoring of the efficacy of experimental preservation procedures.

Animals↗

PubMed2005

Exogenous melatonin enhances bile flow and ATP levels after cold storage and reperfusion in rat liver: implications for liver transplantation.

Although the use of melatonin in the transplantation field has been suggested, it has not been previously tested in a liver cold-storage model. We used a rat liver model to study (a) the dose-dependent effect of melatonin on bile production, and (b) the potential of melatonin to improve liver function after cold-storage. Male Wistar rats were perfused with Krebs-Henseleit bicarbonate buffer (KHB) at 37 degrees C without or with 25, 50, 100 and 200 microM melatonin. Each dose of melatonin stimulated bile production. For cold-storage studies, livers were flushed with either University of Wisconsin (UW) or Celsior solution and stored for 20 hr at 4 degrees C. Reperfusion (120 min) was performed with KHB at 37 degrees C. In subsequent studies, 100 microM melatonin were added to the perfusate during the reperfusion period. ATP and melatonin levels in the tissue were measured. Bile analysis was performed by measuring melatonin, bilirubin and gamma-glutamyl transpeptidase (gamma-GT) levels in the fluid. A dose-dependent increase in bile secretion, associated with an enhanced melatonin and bilirubin levels in the bile were observed. Also, tissue levels of melatonin increased in a dose-dependent manner. When melatonin was added during the reperfusion period, bile production and bile bilirubin levels increased both with UW and Celsior solutions. The analysis of gamma-GT in the bile showed an increase in the Celsior-preserved liver and the addition of melatonin to the perfusate reduced this effect. Tissue ATP levels were higher when melatonin was added to the perfusion medium. Higher levels of melatonin in bile than in tissue were found. In conclusion, we demonstrate that melatonin improves significantly the restoration of liver function after cold-storage and reperfusion.

Adenosine Triphosphate↗

PubMed2004

Apoptosis vs. necrosis: glutathione-mediated cell death during rewarming of rat hepatocytes.

Hypothermia induces injury in its own right, but the mechanisms involved in the cell damage are still unclear. The aim of this study was to test the effects that glutathione (GSH) depletion induces on cell death in isolated rat hepatocytes, kept at 4 degrees C for 20 h, by modulating intracellular GSH concentration with diethylmaleate and buthionine sulfoximine (DEM and BSO). Untreated hepatocytes showed Annexin V stained cells (AnxV(+)), scarce propidium iodide stained cells (PI(+)) and presented a low level of lactate dehydrogenase (LDH) leakage after 20 h at 4 degrees C and rewarming at 37 degrees C. When DEM and BSO were added before cold storage, we observed a few AnXV(+) cells and an increase in PI(+) cells associated with LDH release in the incubation medium. Conversely, the addition of DEM and BSO only during rewarming caused a marked increase in cell death by apoptosis. Production of reactive oxygen species (ROS) and thiobarbituric acid species (TBARS), associated with a decrease in GSH concentrations, was higher when DEM and BSO were added before cold storage. Cells treated with DEM and BSO before cold storage showed lower ATP energy stores than hepatocytes treated with DEM and BSO only during rewarming. Pretreatment of hepatocytes with deferoxamine protected against apoptotic and necrotic morphology in conditions of GSH depletion. These results suggest that pretreatment of hepatocytes with DEM and BSO before cold storage induces necrosis, while the treatment of hepatocytes only during rewarming increases apoptosis. In both conditions, iron represents a crucial mediator of cell death.

Analysis of Variance↗

PubMed2004

Autofluorescence properties of isolated rat hepatocytes under different metabolic conditions.

The contribution of endogenous fluorophores - such as proteins, bound and free NAD(P)H, flavins, vitamin A, arachidonic acid - to the liver autofluorescence was studied on tissue homogenate extracts and on isolated hepatocytes by means of spectrofluorometric analysis. Autofluorescence spectral analysis was then applied to investigate the response of single living hepatocytes to experimental conditions resembling the various phases of the organ transplantation. The following conditions were considered: 1 h after cells isolation (reference condition); cold hypoxia; rewarming-reoxygenation after cold preservation. The main alterations occurred for NAD(P)H and flavins, the coenzymes strictly involved in energetic metabolism. During cold hypoxia NAD(P)H, mainly the bound form, showed an increase followed by a slow decrease, in agreement with the inability of the respiratory chain to reoxidize the coenzyme, and a subsequent NADH reoxidation through alternative anaerobic metabolic pathways. Both bound/free NAD(P)H and total NAD(P)H/flavin ratio values were altered during cold hypoxia, but approached the reference condition values after rewarming-reoxygenation, indicating the cells capability to restore the basal redox equilibrium. A decrease of arachidonic acid and vitamin A contributions occurred after cold hypoxia: in the former case it may depend on the balance between deacylation and reacylation of fatty acids, in the latter it might be related to the vitamin A antioxidant role. An influence of physico-chemical status and microenvironment on the fluorescence efficiency of these fluorophores cannot be excluded. In general, all the changes observed for cell autofluorescence properties were consistent with the complex metabolic pathways providing for energy supply.

Animals↗

PubMed2003

Selective blockade of mGlu5 metabotropic glutamate receptors is protective against acetaminophen hepatotoxicity in mice.

BACKGROUND/AIMS: mGlu5 metabotropic glutamate receptor antagonists protect rat hepatocytes against hypoxic death. Here, we have examined whether mGlu5 receptor antagonists are protective against liver damage induced by oxidative stress. METHODS: Toxicity of isolated hepatocytes was induced by tert-butylhydroperoxide (t-BuOOH) after pretreatment with the mGlu5 receptor antagonists, MPEP, SIB-1757 and SIB-1893. The effect of these drugs was also examined in mice challenged with toxic doses of acetaminophen. RESULTS: Addition of tBuOOH (0.5 mM) to isolated hepatocytes induced cell death (70+/-5% at 3 h). Addition of MPEP or SIB-1893 to hepatocytes reduced both the production of reactive oxygen species (ROS) and cell toxicity induced by t-BuOOH (tBuOOH=70+/-5%; tBuOOH+MPEP=57+/-6%; tBuOOH+SIB-1893=40+/-4%). In mice, a single injection of acetaminophen (300 mg/kg, i.p.) induced centrilobular liver necrosis, which was detectable after 24 h. MPEP (20 mg/kg, i.p.) substantially reduced liver necrosis and the production of ROS, although it did not affect the conversion of acetaminophen into the toxic metabolite, N-acetylbenzoquinoneimine. MPEP, SIB-1893 and SIB-1757 (all at 20 mg/kg, i.p.) also reduced the increased expression and activity of liver iNOS induced by acetaminophen. CONCLUSIONS: We conclude that pharmacological blockade of mGlu5 receptors might represent a novel target for the treatment of drug-induced liver damage.

Acetaminophen↗

PubMed2003

Tumor microenvironment and hemorheological abnormalities.

This article reviews the various mechanisms in a malignant tumor that lead to hypoxia, production of tumor interstitial fluid, and rheological changes in its microenvironment. In addition, the associated procoagulant effects are described. The latter phenomenon is the result of complex and dynamic interplay among tumor cells, endothelial cells, circulating blood cells, and angiogenic, clotting, and hemorheological factors. The implications of these abnormalities on therapeutic approach are discussed.

Extracellular Fluid↗

PubMed2002

In situ detection of reactive oxygen species and nitric oxide production in normal and pathological tissues: improvement by differential interference contrast.

Reactive oxygen species (ROS), among which nitric oxide (NO) is currently included, play a plethora of (patho)physiological roles. Harman's free radical theory of aging put forth over 40 years ago received full support since then. A nitric oxide hypothesis of aging recently proposed by McCann, is very likely to be the object of widespread investigation in the near future. Therefore, the possibility of localizing at the (sub)cellular level under the light microscope the sites of ROS and NO production with simple and reliable methods appears as a powerful tool for analytic cytology and pathology. Various histochemical methods were developed for visualizing ROS production; a recently improved version to localize superoxide (and possibly also singlet oxygen), based on a DAB-Mn2+ -Co2+ reaction, appears very promising. Since the direct detection of NO is still very difficult, the action sites of NO are currently localized by the identification of NO synthase (NOS). The most widespread method to reveal the catalytic activity of NOS is that of demonstrating the fixation-resistant NADPH diaphorase activity with the tetrazolium salt method. We have improved this method by using a tetrazolium salt whose formazan particles are very thin and lipid insoluble (tetranitroblue tetrazolium, TNBT) and by including a tissue protectant, polyvinyl alcohol, in the incubation medium. Here significant examples of application of the DAB-Mn2+ -Co2+ technique for ROS and the TNBT-PVA method for NOS to normal liver and brain and to solid tumors are presented. We further document the usefulness of Nomarkis's differential interference contrast (DIC) to analyze wide tissue areas where ROS production or NOS activity is low or even nil. The improved version for NOS allowed for the first time to demonstrate NOS activity in liver fat-storing cells and in astrocyte-like cells in the brain.

Aging↗

PubMedDate not supplied

Stem cell recruitment and liver de-differentiation in MMTV-neu (ErbB-2) transgenic mice.

The liver of tumor-bearing hosts manifests fetal phenotypes. We investigated the expression of differentiation markers on the liver in MMTV-neu (ErbB-2) transgenic mice, in the period from incipient neoangiogenesis to lung metastatization. We report AFP expression by hepatocytes in all lobular zones, CD34 cell arrest and subsequent hemopoiesis in periportal and mid-zone areas, oval-like cells (CD34+, CK19+, AFP+) and ductular reaction in portal tracts, portal CK19+ and GGT+ hepatoblast-like cells, and midzonal large dysplastic hepatocytes. We hypothesize that CD34 cells are recruited by the tumor from the marrow for angiogenic purposes and that their differentiation in the liver is influenced by altered liver microenvironment(s). AFP may act as a growth factor and biological response modifier for these cells and for the tumor. Dysplasia might be enhanced by metabolic stress. We conclude that the liver differentiation potential is lobular-zone-dependent and that the risk for eventually developing a pre-malignant lesion is not negligible.

Adenocarcinoma↗

PubMedDate not supplied

In situ demonstration of improvement of liver mitochondria function by melatonin after cold ischemia.

In a previous investigation, reperfusion with a melatonin-containing medium was demonstrated to enhance bile production and tissue ATP levels in rat livers, cold-preserved with University of Wisconsin (UW) or Celsior solutions, with respect to melatonin-free reperfusion; lipid peroxidation products in the perfusate were not influenced by the indole. This was ascribed to an increased efficiency of the hepatocyte mitochondria induced by melatonin. Reactive oxygen species (ROS) normally leak from the electron transfer chain in mitochondria and excessive ROS production is presumed to mediate ischemia-reperfusion (I/R) damage. A histochemical reaction was used to demonstrate ROS on the same model. Compared to the lobular zonation of ROS in control livers, the stained area of cold-preserved livers reperfused without melatonin was restricted to a narrow portal region, in keeping with the much lower ATP content. When reperfusion was performed with melatonin, the liver morphology was improved and the ROS reaction in hepatocytes more intense, though not reaching the control liver pattern. Sinusoidal cells were poorly-stained in both cases. In conclusion, with this different approach, melatonin was confirmed to improve mitochondrial performance and to discriminate parenchymal from sinusoidal cell behavior. Our observations confirm that melatonin mitigates I/R injury and support its potential in liver transplantation.

Adenosine↗