Effect of Glutathione and N-Acetylcysteine on in Vitro and in VIVO Cardiac Toxicity of Doxorubicin

The effects of two sulfhydryl compounds, glutathione (GSH) and N-acetylcysteine (NAC), on the cardiotoxicity of doxorubicin (DXR) were tested on in vitro and in vivo models. DXR was administered to rats as 4 weekly i.v. doses of 3mg/kg. GSH (1.5 mmoles/kg), given i.v. 10 min before and 1 hr after DXR, was found to prevent the development of the delayed cardiotoxic effects of DXR, as assessed by electrocardiographic and mechanical parameters, as well as by histological examination of left ventricular preparations. In contrast, equimolar oral doses of NAC (1 hr before and 2hrs after DXR) were found to be ineffective. Both GSH and NAC prevented the negative inotropic effect produced by DXR on isolated rat atria. A good correlation exists between the cardioprotective effects of the two agents and their ability to enhance the non-protein sulfhydryl group content of the myocardium. Differences observed in vivo between GSH and NAC might be accounted for by pharmacokinetic factors.     

Effect of menthol in experimentally induced ulcers: Pathways of gastroprotection

Based on ethnopharmacological indications that Mentha species may be used in the treatment of gastrointestinal diseases, this study aimed to characterize the gastroprotective mechanisms of menthol (ME), the major compound of the essential oil from species of the genus Mentha. The gastroprotective action of ME was analyzed in gastric ulcers that were induced by ethanol or indomethacin in Wistar male rats. The mechanisms responsible for the gastroprotective effect were assessed by analyzing the amount of mucus secreted, involvement of non-protein sulfhydryl (NP-SH) compounds, involvement of calcium ion channels and NO/cGMP/K⁺_ATP pathway, gastric antisecretory activity and the prostaglandin E₂ (PGE₂) production. The anti-diarrheal activity and acute toxicity of ME were also evaluated. Oral treatment with ME (50 mg/kg) offered 88.62% and 72.62% of gastroprotection against ethanol and indomethacin, respectively. There was an increased amount of mucus and PGE₂ production. The gastroprotective activity of ME involved NP-SH compounds and the stimulation of K⁺_ATP channels, but not the activation of calcium ion channels or the production of NO. The oral administration of ME induced an antisecretory effect as it decreased the H⁺ concentration in gastric juice. ME displayed anti-diarrheal and antiperistaltic activity. There were no signs of toxicity in the biochemical analyses performed in the rats’ serum. These results demonstrated that ME provides gastroprotective and anti-diarrheal activities with no toxicity in rats.     

The Gastroprotective Effect of Menthol: Involvement of Anti-Apoptotic,Antioxidant and Anti-Inflammatory Activities

The aim of this research was to investigate the anti-apoptotic, antioxidant and anti-inflammatory properties of menthol against ethanol-induced gastric ulcers in rats. Wistar rats were orally treated with vehicle, carbenoxolone (100 mg/kg) or menthol (50 mg/kg) and then treated with ethanol to induce gastric ulcers. After euthanasia, stomach samples were prepared for histological slides and biochemical analyses. Immunohistochemical analyses of the cytoprotective and anti-apoptotic heat-shock protein-70 (HSP-70) and the apoptotic Bax protein were performed. The neutrophils were manually counted. The activity of the myeloperoxidase (MPO) was measured. To determine the level of antioxidant functions, the levels of glutathione (GSH), glutathione peroxidase (GSH-Px), glutathione reductase (GR) and superoxide dismutase (SOD) were measured using ELISA. The levels of the pro-inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) and the anti-inflammatory cytokine interleukin-10 (IL-10) were assessed using ELISA kits. The menthol treated group presented 92% gastroprotection compared to the vehicle-treated group. An increased immunolabeled area was observed for HSP-70, and a decreased immunolabeled area was observed for the Bax protein in the menthol treated group. Menthol treatment induced a decrease in the activity of MPO and SOD, and the protein levels of GSH, GSH-Px and GR were increased. There was also a decrease in the levels of TNF-α and IL-6 and an increase in the level of IL-10. In conclusion, oral treatment with menthol displayed a gastroprotective activity through anti-apoptotic, antixidant and anti-inflammatory mechanisms.     

Numerical modeling of hemodynamics scenarios of patient-specific coronary artery bypass grafts

A fast computational framework is devised to the study of several configurations of patient-specific coronary artery bypass grafts. This is especially useful to perform a sensitivity analysis of the hemodynamics for different flow conditions occurring in native coronary arteries and bypass grafts, the investigation of the progression of the coronary artery disease and the choice of the most appropriate surgical procedure. A complete pipeline, from the acquisition of patient-specific medical images to fast parameterized computational simulations, is proposed. Complex surgical configurations employed in the clinical practice, such as Y-grafts and sequential grafts, are studied. A virtual surgery platform based on model reduction of unsteady Navier–Stokes equations for blood dynamics is proposed to carry out sensitivity analyses in a very rapid and reliable way. A specialized geometrical parameterization is employed to compare the effect of stenosis and anastomosis variation on the outcome of the surgery in several relevant cases.     

Thermodynamic and NMR Study of Proton Complex Formation of 2′-Deoxyadenylyl-(3′→5′)-2′-Deoxyadenosine in Aqueous Solution

Abstract

A thermodynamic and proton NMR study has been carried out in order to characterize the behaviour of 2′-deoxyadenylyl-(3′→5′)-2′-deoxyadenosine towards the protonation. Conformational changes occurring following two protonation steps are discussed.     

Gastroprotective mechanisms of Citrus lemon (Rutaceae) essential oil and its majority compounds limonene and β-pinene: involvement of heat-shock protein-70, vasoactive intestinal peptide, glutathione, sulfhydryl compounds, nitric oxide and prostaglandin E₂

Citrus lemon (CL) belongs to Rutaceae family and is popularly known in Brazil as limão siciliano. The phytochemical analysis of CL fruit bark essential oil showed two majority components, limonene (LIM) and β-pinene (PIN). This study aimed to evaluate the gastroprotective mechanism of action from CL, LIM and PIN in ethanol- and indomethacin-induced gastric ulcers and its in vitro anti-Helicobacter pylori activity. After ethanol-induced gastric ulcer, the ulcer area was measured and the stomachs were destined to histology (HE and PAS), immunohistochemistry for HSP-70 and VIP and glutathione (GSH) measurement. The involvement of nitric oxide (NO) and sulfhydryl (SH) compounds was determined. The ulcer area for indomethacin-induced gastric ulcers was measured. PGE₂ concentration was biochemically measured. The minimum inhibitory concentration (MIC) against H. pylori was determined in vitro. In ethanol model, CL and LIM demonstrated 100% of gastroprotection, while PIN did not exert effective gastroprotection (53.26%). In the indomethacin model, CL and LIM offered effective gastroprotection but PIN did not show gastroprotective effect. The gastric ulcer area of rats pretreated with NO-synthase inhibitor or SH-blocker was decreased in comparison to the control group. The MIC obtained for CL was 125 μg/mL, for LIM was 75 μg/mL and for PIN was 500 μg/mL. The gastroprotective effect of CL and LIM was involved with increasing in mucus secretion, HSP-70 and VIP, but not with GSH, NO or SH compounds. CL gastroprotective mechanism is involved with PGE₂. PIN did not present gastroprotective activity.     

Effect of lanthanum on the turnover of calcium in rat uterus.

Previous investigations suggest that lanthanum might enter uterine smooth muscle cells and work as intracellular calcium displacing agent. The present investigation had been carried out in order to confirm if lanthanum develops an intracellular effect. Experiments show that lanthanum brings about a marked increase of the intracellular calcium; the comparison of the uptake and of the wash-out curve of 45Ca shows that lanthanum induces a lowering of the rapid phase of 45Ca release from rat uterus, while the uptake of the labelled ion is not modified or is even enhanced. The present data demonstrate that the action of lanthanum in rat uterus is limited to the cell membrane, whose calcium extruding properties are inhibited.     

Identification of New Autoantigens by Protein Array Indicates a Role for IL4 Neutralization in Autoimmune Hepatitis

Autoimmune hepatitis (AIH) is an unresolving inflammation of the liver of unknown cause. Diagnosis requires the exclusion of other conditions and the presence of characteristic features such as specific autoantibodies. Presently, these autoantibodies have relatively low sensitivity and specificity and are identified via immunostaining of cells or tissues; therefore, there is a diagnostic need for better and easy-to-assess markers. To identify new AIH-specific autoantigens, we developed a protein microarray comprising 1626 human recombinant proteins, selected in silico for being secreted or membrane associated. We screened sera from AIH patients on this microarray and compared the reactivity with that of sera from healthy donors and patients with chronic viral hepatitis C. We identified six human proteins that are specifically recognized by AIH sera. Serum reactivity to a combination of four of these autoantigens allows identification of AIH patients with high sensitivity (82%) and specificity (92%). Of the six autoantigens, the interleukin-4 (IL4) receptor fibronectin type III domain of the IL4 receptor (CD124), which is expressed on the surface of both lymphocytes and hepatocytes, showed the highest individual sensitivity and specificity for AIH. Remarkably, patients'' sera inhibited STAT6 phosphorylation induced by IL4 binding to CD124, demonstrating that these autoantibodies are functional and suggesting that IL4 neutralization has a pathogenetic role in AIH.Autoantibodies specific for proteins or nonprotein antigens (dsDNA, snRNP, carbohydrates) are often the serological hallmark of autoimmune diseases. Autoantibodies can be simply an epiphenomenon secondary to a chronic inflammatory milieu (1), but they can also play a direct pathogenetic role, as antithyroglobulin antibodies do in Hashimoto''s thyroiditis (2).Autoimmune hepatitis (AIH)¹ is a chronic necro-inflammatory disease of unknown etiology that affects predominantly women with an incidence of 1 to 2 per 100,000 per year and a prevalence of 10 to 20 out of 100,000 (3, 4). AIH is subdivided into two major types on the basis of autoantibody reactivity (5). Antibodies to nuclei and/or to smooth muscle characterize type 1 AIH, whereas antibodies to a liver-kidney microsomal constituent define patients with type 2 AIH. Because the detection of these autoantibodies is done by means of immunofluorescence on rodent multi-organ sections (liver, kidney, stomach), there are problems with the standardization and interpretation of the immunostaining patterns (6). To overcome these methodological problems, the International Autoimmune Hepatitis Group established an international committee to define guidelines and develop procedures and reference standards for more reliable testing (7, 8). Although ELISA and bead assays with purified or recombinant autoantigens are under development (9), they actually represent a complementary, rather than alternative, approach to traditional immunofluorescence. Moreover, serological overlap is frequently observed between AIH and other non-autoimmune liver diseases such as chronic viral hepatitis (10). Therefore, new, highly specific markers represent an unmet medical need for the more accurate diagnosis and classification of AIH.Besides the potential diagnostic application, the discovery of novel AIH autoantigens could provide insights into the disease pathogenicity mechanism. Although some AIH target-autoantigens have been identified and characterized, little is known about their pathogenetic role, and other autoantigens are probably still unknown. Autoantibodies, to be considered pathogenetic, must have at least two features: (i) the target-autoantigen should be either expressed on the plasma membrane of target cells or secreted by cells (i.e. should be exposed to autoantibodies), and (ii) binding of the autoantibodies to the target antigen should disturb a cellular function directly or indirectly. A possible pathogenetic role in AIH has been put forward for autoantibodies specific for cytochrome P450 2D6 (CYP2D6) or Asialoglycoprotein receptor 1 (AGPR-1), which are both present on the hepatocyte cell membrane (10).Protein microarrays are a powerful technology, as they allow the simultaneous screening of thousands of analytes (11). In the present study, to identify new autoantigens with potential diagnostic and/or pathogenetic roles in AIH, we printed a microarray with 1626 human proteins whose main feature was being either secreted or membrane associated (i.e. potentially exposed to autoantibody recognition). We used this microarray to screen panels of sera from patients with AIH and identified six new protein antigens that are recognized with high sensitivity and specificity. One of these six autoantigens is the interleukin-4 (IL4) receptor fibronectin type III (FNIII) domain of the IL4 receptor (CD124), and, interestingly, patients'' autoantibodies specific for CD124 neutralize IL4 signaling, suggesting a possible pathogenetic role for IL4 neutralization in AIH.     

The effects of ethanol on the responses of the vascular musculature to norepinephrine

Intravenous infusion of ethanol (EtOH) (up to 0.1 ml kg-1 min-1) does not modify the blood pressure in conscious rabbits. In this range of doses, EtOH brings about an increase of the pressure responses induced by norepinephrine (NE) with respect to values obtained in the absence of EtOH. Similar experiments have been performed on the isolated rat tail artery: EtOH perfusion does not modify the basal tone of the preparation up to 2% concentration, while the contractile response to NE administration is potentiated. The supersensitivity of the arteries to NE induced by EtOH, is a function of EtOH concentration and can be represented by a bellshaped curve reaching the maximal value at 1% EtOH, whereby concentrations larger than 2% induce an inhibition of NE activity. The present experiments suggest that the potentiation of the contractile effects of NE induced by EtOH might be related to an enhancement of the rate of calcium transport due to the increase in cell membrane plasticity induced by EtOH.