The effect of anti oxidant drugs on platelet Enzymes

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2 The effect of anti oxidant drugs on platelet Enzymes ( xanthineoxidase and lipid peroxidase) in MI patients Mohsen Hamidpour (MSc, PhD ) Shahid Beheshti University of Medical Science Paramedical Faculty

3 Five risk factors involved in Heart and vascular disease: 1 Precipitate of lipids in vein endothelial cells. 2 High Shear stress 3 Systemic factors (hyper lipidemia, Diabetic) 4 platelet p dysfunction & Coagulation inhibitor s dysfunction 5 Genetic

4 Platelet Function

5 The Platelet roll in MI The h platelets l play an important roll in ischemic i myocardial disease in many way Consequence ischemic of the myocardial is huge generation oxygen derived free radicals that have the huge pathologic consequences. Platelet xanthine oxidase is on of the free radical generation sources.

6 Xanthine oxidoreductase the metabolism of purines, catalyzing the conversion of both hypoxanthine and xanthine to uric acid under normal conditions i exists in dehyrogenase form and uses NAD+ and there is no or very little production of superoxide anion Under ischemic conditions depletion of ATP and subsequent loss of membrane Ca2+ gradient Increased Ca2+ levels activates Ca2+ dependent proteases cause selective proteolysis of the dehydrogenase to convert it into xanthine which expense of oxygen to produce superoxide ion Thus xanthine oxidase in ischemic conditions of the heart, may play an important role in MI

7 Materials & methods

8 Patients and controls 20 patients with MI, 10 voluntary healthy people were under taken as Normal controls The Patients were divided in two groups Group 1 administrate their routine drugs and aspirin after myocardial infraction Group 2 anti oxidant durgs such as asprin with vitamin E

9 Methods collected 10 ml anticoagulated blood from patients and controls Anticoagulated blood was centrifuged at 200 g for 15 min to obtained Plasma Rich platelet( PRP) containing 2x10 9 /ul platelets.

10 Detection of Xanthine oxidase 0.30 ml Tris HCl buffer, 50 mm ph ml CuSO4, 10 mm 0.05 ml. Xanthine, 2.58 mm per ml. in 0.05 M glycine gy buffer, ph ml. of diluted PRP and water to make up the volume to 3 ml.

11 Detection of Xanthine oxidase Absorbance the mixture at 290 nm at 15 seconds interval for one minute. One unit of activity has been defined as change in absorbance b at 290 nm in 1 minute by 1 ml. enzyme preparation.

12 Detection of Lipid peroxidase The Lipid peroxidase measured as the levels of Malondialdehyde (MDA ) To 0.20 ml. of PRP was added d 0.8 ml. SDS 8.1% 0.50 ml acetic acid and thiobarbituric acid, 0.8% to make up the volume to 3.0 ml. Contents of the tubes were mixed and heated over at 90 C for 1 hr Cooled under running water. Added To each tube, 1.0 ml. water and 5.0 ml solution of n butanol and pyridine (15:1 v/v) centrifuged at 800 g for 10 min. The upper layer was aspirated out and color intensity measured at 532 nm. The 1, 1,3, 3 tetra ethoxy propane was used as STANDARD

13 Data Analysis The data was analyzed using the statistical tools like SPSS.

14 Results

15 The CK and CK MB patients G1a: Enzyme activity of patients G1 (3 hrs after MI) G1a: Enzyme activity of patients G1 (6 days after MI) G2a: Enzyme activity of patients G2 (3 hrs after MI) G1b: Enzyme activity of patients G2 (6 days after MI)

16 Thecomparison of MDA (lipid peroxidase and xanthin oxidase) Between two groups of patients and control Groups (MDA (mean± SD ) xanthine oxidation(mean± SD ) Normal (n=10) ± ± (n=10) ± ± (n=10)) ± ± G1 Recivec B blocker and Asprin G2 B blockers + Asprin and Vitimin E The P value of platelet l t X ox Normal control with G1 is : P< The P value of platelet X ox G1 and G2 P< 0.001

17 The P value of platelet Lipid peroxidase Normal control with G1 is : P< The P value of platelet Lipid peroxidase G1 and G2 P< 0.005

18 The P value of platelet Xanthine oxidase Normal control with G1 is : P< The P value of platelet Xanthine oxidase G1 and G2 P< 0.001

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20 discussion 1 administration of Vitamin E alone asprin after 6 days gave us a good result as an anti oxidant drugs effect as evidence by reducing platelet xanthine oxidase activity P< Lowering lipid peroxidase metabolite (MDA) P<

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22 suggestion Using the Vitamin E alone the other B blocker drugs would protect patients with MI from oxidant and further heart fouler Detection of xanthine oxidase and MAD as a MI Biomarker in Heart clinical centers

23 Future works Use more anti oxidant drugs and supplements like B glucan and Search with multicenter (Hospitals) and more patients

24 References eee 1. Pandey, NR., et al. Enzymatic oxidant and anti oxidants of human blood platelets in unstable angina and myocardial infarction. Int. J. Cardiol 2010 ; 7 : Haseruck N, et al. The plaque lipid lysophosphatidic p acid stimulates platelet activation and platelet monocyte aggregate formation in whole blood: involvement of P2Y1 and P2Y12 receptors. Blood 2004; 103: Vivek K. Dwivedi, et al. Effect of viamin E on platelet enzymatic anti oxidant in the patients of myocardial infraction. i Indian Journal of Clinical i l Biochemistry, i 2005; 20 4 Chandra, M..effect of vitamin E on the platelet xanthine oxldase and lipid peroxidation in the patients of myocardial infarction. J. Clinical Biochemistry 2008 ; 21: 26 5 Bogani, P., et al.postprandial anti inflammatory and antioxidant effects of extra virgin olive oil. Atherosclerosis 2010; 191: Lee TM, et al. Differential effects of NADPH oxidase and xanthine oxidase inhibition on sympathetic reinnervation in postinfarct rat hearts. Free Radic Biol Med. 2011; 50: