Emphasis Type="Italic"N/Emphasisacetylcystein

Authors: Erfan Nur Dees P Brandjes Tom Teerlink HansMartin Otten Ronald P J Oude Elferink Frits Muskiet Ludo M Evers Hugo ten Cate Bart J Biemond Ashley J Duits JohnJohn B Schnog on behalf of the CURAMA study group

Publish Date: 2012/02/10

Volume: 91, Issue: 7, Pages: 1097-1105

PDF Link

Abstract

Oxidative stress is of importance in the pathophysiology of sickle cell disease SCD In this open label randomized pilot study the effects of oral Nacetylcysteine NAC on phosphatidylserine PS expression as marker of cellular oxidative damage primary end point and markers of hemolysis coagulation and endothelial activation and NAC tolerability secondary end points were studied Eleven consecutive patients ten homozygous HbSS sickle cell patients one HbSβ0thalassemia patient were randomly assigned to treatment with either 1200 or 2400 mg NAC daily during 6 weeks The data indicate an increment in whole blood glutathione levels and a decrease in erythrocyte outer membrane phosphatidylserine exposure plasma levels of advanced glycation endproducts AGEs and cellfree hemoglobin after 6 weeks of NAC treatment in both dose groups One patient did not tolerate the 2400 mg dose and continued with the 1200 mg dose During the study period none of the patients experienced painful crises or other significant SCD or NAC related complications These data indicate that Nacetylcysteine treatment of sickle cell patients may reduce SCD related oxidative stressThe CURAMA study group is a collaborative effort studying sickle cell disease in the Netherlands Antilles and the Netherlands Participating centers The Red Cross Blood Bank Foundation Curaçao Netherlands Antilles The Antillean Institute for Health Research Curaçao Netherlands Antilles The Department of Internal Medicine Slotervaart Hospital Amsterdam the Netherlands the Department of Vascular Medicine and the Department of Hematology Academic Medical Center Amsterdam the Netherlands the Department of Hematology Erasmus Medical Center Rotterdam the Netherlands Pathology and Laboratory Medicine University Medical Center Groningen the Netherlands the Department of Internal Medicine the Laboratory of Clinical Thrombosis and Hemostasis and the Cardiovascular Research Institute Academic Hospital Maastricht the NetherlandsOxidative stress plays a role of major importance in the development of organ damage in sickle cell disease SCD 1 2 3 4 Oxidative stress in SCD results from factors such as the unstable autooxidative sickle hemoglobin HbS 5 6 chronic intravascular hemolysis 7 8 9 recurrent ischemia reperfusion injury 4 and low grade inflammation 10 Increased levels of reactive oxygen species ROS lead to further acceleration of hemolysis 8 hypercoagulability 11 12 decreased nitric oxide NO bioavailability 13 and endothelial damage 14Given the fact that oxidative stress is a likely major contributing factor in the development of both acute and chronic complications in SCD the potential of antioxidants as therapeutics for SCD should be explored A major intracellular antioxidant is the reduced form of the aminothiol glutathione GSH 15 16 Due to increased consumption by excessive levels of ROS sickle cell patients have decreased levels of plasma and erythrocyte total glutathione and the ratio of GSH to its oxidized form glutathione disulfide GSSG is reduced 17 18 Nacetylcysteine NAC the rate limiting substrate for GSH generation is an important antioxidant with pleiotropic effects on inflammation and vasomotor function 19 NAC readily enters cells and within the cytoplasm it is converted to lcysteine which is a precursor to GSH 20 Treatment of sickle cell patients with NAC has been demonstrated to have an inhibitory effect on the formation of dense red cells 21 Augmenting the antioxidant capacity in sickle red blood cells by NAC may reduce oxidative red cell membrane damage and reduce its many downstream pathophysiological effects such as hemolysis endothelial damage and activation activation of the coagulation cascade and the decrease of NO bioavailability 13 22 23 In this randomized open label pilot study the effects of oral NAC treatment on markers of oxidative stress and hemolysis in sickle cell patients were investigatedConsecutive adult age ≥18 years homozygous sickle cell anemia HBSS or HbSβ0thalassemia outpatients high performance liquid chromatography HPLC confirmed at the Academic Medical Center AMC Amsterdam The Netherlands were eligible for the study Exclusion criteria were painful crisis and blood transfusion in the preceding 4 weeks and 4 months respectively pregnancy or the desire to get pregnant in the following 3 months calculated glomerular filtration rate of 90 ml/min Cockcroft and Gault formula known gastric/duodenal ulcers active infections autoimmune inflammatory diseases and use of hydroxyurea vitamin K antagonists or other oral anticoagulants and contraindications for NAC use Ten race and agematched healthy volunteers were included as controls for baseline values All participants received verbal and written explanation of the objectives and procedure of the study and subsequently provided written informed consent The study was approved by the AMC Medical Ethical Commission and experiments were performed in accordance with the Declaration of Helsinki The study was registered in the Dutch Trial Registry wwwtrialregisternl trial ID number NTR1013The primary end point of the study was reduction of erythrocyte phosphatidylserine PS expression as a direct indicator of erythrocyte membrane oxidative damage Changes in markers of hemolysis hemoglobin reticulocytes lactate dehydrogenase LDH and bilirubin hypercoagulability endothelial activation and inflammation and tolerability of oral NAC were secondary end pointsAfter baseline measurements and randomization to either 1200 or 2400 mg of NAC per day patients started taking NAC acetylcysteine 600 mg tablets dissolved in water Pharmachemie BV Haarlem The Netherlands orally twice daily during 6 weeks followed by another 6 weeks of followup after NAC cessation Both during NAC treatment visits 0–3 and in the posttreatment period visits 4–6 patients were seen twoweekly for followup visits during which questionnaires pertaining to side effects were completed weight blood pressure and pulse were measured and a blood sample was drawn via venipuncture Patients kept a daily pain score diary visual analogue scale pain score Patient compliance was monitored by history taking and pill counts If possible NAC treatment would not be discontinued in case of an hospital admission due to acute vasoocclusive pain crisis or other clinical eventStandard blood counts were performed in EDTA anticoagulated blood CellDyn 4000 Abbott Illinois USA LDH and total and direct bilirubin levels were measured in heparinized plasma with spectrophotometry P800 Modular Roche Basel Switzerland Citrate serum and EDTA samples were centrifuged immediately for 15 min at 3000 rpm 4°C Aliquots were stored at −80°C until further analysis larginine immediate precursor of NO asymmetric dimethylarginine ADMA and symmetric dimethylarginine SDMA levels were measured in EDTA plasma using reversedphase HPLC as described elsewhere 24 25 NTproBNP levels were measured in EDTA plasma with electrochemiluminescence immunoassay Roche Diagnostics Ultrasensitive Creactive protein USCRP was measured with ELISA according to the manufacturers instructions Biokit Barcelona Spain Prothrombin fragment 1 + 2 F1 + 2 and thrombin–antithrombin TAT complexes were determined using sandwich enzymelinked immunosorbent assay ELISA Enzygnost Dade Behring Marburg GmbH Germany Plasma levels of von Willebrand Factor antigen vWFag were determined in citrate plasma with in an inhouse ELISA Soluble vascular adhesion molecule1 sVCAM1 levels were determined in serum RD Systems Minneapolis MN USA

Keywords: