RAJIVGANDHIUNIVERSITY OF HEALTH SCIENCES, KARNATAKA

BANGALORE

ANNEXURE-II

PROFORMA FOR REGISTRATION OF SUBJECTS FOR DISSERTATION

1 / Name of the candidate and address (in block letters) / Dr. SHILPASREE.A.S
POST GRADUATE STUDENT,
DEPARTMENT OF BIOCHEMISTRY,
J.J.M.MEDICALCOLLEGE,
DAVANGERE
2 / Name of the institution / J.J.M.MEDICALCOLLEGE,
DAVANGERE-577004
3 / Course of the study and subject / M.D. BIOCHEMISTRY
4 / Date of admission to the course / 30-05-2009
5 / Title of the topic / “STUDY OF SERUM APOLIPOPROTEINS AI, B AND LIPID PROFILE IN STROKE”
6
7.
8. / BRIEF RESUME OF THE INTENDED WORK
6.1 Need for the study
Stroke is defined as rapidly developing clinical signs of focal or global disturbances of cerebral function lasting for more than 24 hours leading to death with no apparent cause other than vascular origin.1
Stroke incidence ranges from 0.2 to 2.5/1000 population per year. There is no reliable information from India, but analysis of data suggests that 2% of all hospital cases, 4.5% of medical and 20% of neurological cases are from stroke. Stroke is the leading cause of death. It accounts for 10-12% of total deaths.1
Abnormalities in plasma lipoproteins and derangements in lipid metabolism rank
among the most firmly established and best understood risk factors for atherosclerosis.
Abnormal lipid parameters like total cholesterol,LDL, HDL and triglycerides are probable risk factors for ischemic stroke, largely by their link to atherosclerosis.2 Atherogenic index defined as log (TG/HDL-c in mmol/l) is an indicator to predict the risk for the development of athersclerosis.
Apolipoproteins are protein components of lipoproteins.Apolipoprotein AI andB
are the protein components of HDL and LDL cholesterol respectively. Apolipoprotein AI and HDL are antiatherogenic whereas apolipoprotein B and LDL are atherogenic.
Dyslipidemia, low apolipoprotein AI and high apolipoprotein B are widely accepted as risk factors for Coronary heart disease, in contrast the relation is not well established for stroke.3
So the present study is undertaken to evaluate the relationship between apolipoprotein AI, B and lipid profile with stroke.
6.2 Review of literature:
Various studies have demonstrated positive and significant association between total cholesterol, LDL cholesterol and risk of stroke and inverse relationship between HDL cholesterol and fatal stroke.3,4,5
Studies have also shown that linear increase in the levels of nonfasting triglycerides leads to stepwise increase in the risk of stroke.6
Analysis of lipid and lipoprotein profile in young patients with stroke revealed that total cholesterol, LDL cholesterol, Triglycerides, Apo AI, Apo B and Apo B/ Apo AI ratio were significantly altered as compared to normal healthy controls.7
Among the patients with preexisting cardiovascular disease high levels of ApoB and low ApoA1 levels are associated with increased risk of ischemic stroke. Apolipoproteins remained independent risk factors upon adjustment for traditional risk factors. Apo B/Apo AI ratio as a useful indicator of ischemic stroke risk in patients with preexisting atherothrombotic disease.8, 12
Prospective cohort study from Taiwan showed that Apo AI but not Apo B levels serve as an effect modifier of hypertension for the risk of stroke.9
In contrast few studies have shown no difference between total cholesterol, triglycerides, Apolipoprotein AI and B in patients compared to controls but higher LDL cholesterol and lower HDL cholesterol was observed in stroke patients.11
6.3 Objectives of the study:
a)To study the levels of apolipoprotein AI and B in stroke patients and healthy subjects.
b)To study the ratio of Apo B/Apo AI in stroke patients and healthy subjects.
c)To study the lipid profile (Total cholesterol, LDL, HDL and triglycerides) and atherogenic index – log (TG/HDL-c in mmol/l) in stroke patients and healthy subjects.
MATERIALS AND METHODS:
7.1 Source of data:
A crosssectional study will be carried out for a period of one year. The patients will be selected from Chigateri general hospital and Bapuji hospital Davangere (both hospitals are attached to the teaching institute JJM medical college Davangere).
Inclusion criteria:
Cases - 50 clinically proven cases of stroke in the age group of 30 - 70 years.
Cases diagnosed by computed tomography.
Controls - 50 age and sex matched healthy individuals without any major illness and not on any medications.
Exclusion criteria:
Stroke patients with
  • Hepatic and renal disease
  • Sepsis
  • Malignancy
  • Patients on hypolipidemic drugs
7.2 Methods of collection of data
About 6 ml of venous blood will be drawn under aseptic precaution in a sterile bulb from selected subjects after overnight fasting. Serum separated by centrifugation will be used for analysis. All biochemical parameters will be estimated by semiautoanalyser.
a) Estimation of total cholesterol by cholesterol oxidase/phenol aminoantipyrine method.10
Principle: cholesterol esterase hydrolyses cholesterol esters to free cholesterol and fatty acids. Then cholesterol is oxidised by cholesterol oxidase forming cholesterol4ene-3one and hydrogen peroxide. In presence of enzyme peroxidase, hydrogen peroxide causes oxidative coupling of phenol and antipyrine to form red coloured quinoneimine dye which is measured at 520nm.
b) Estimation of HDL cholesterol by cholesterol oxidase/phenol aminoantipyrine method.10
Principle:Chylomicrons, VLDL and LDL are precipitated by adding phosphotungstic acid and magnesium ions to the sample. Centrifugation leaves only HDL in the supernatant. Cholesterol content in it is determined enzymatically.
c)Estimation of LDL cholesterol by Friedewald formula.10


d)Estimation of triglycerides by glycerol phosphate oxidase-phenol aminoantipyrine method.10
Principle: Triglycerides are hydrolysed by lipoprotein lipase to glycerol and fatty acids. Glycerol is then phosphorylated to glycerol-3-phosphate followed by oxidation to hydrogen peroxide and dihydroxyacetone phosphate by glycerol phosphate oxidase. Hydrogen peroxide in presence of peroxidase causes oxidative coupling of 4 chlorphenol and 4 amonophenazone to form red coloured quinoneimine dye which is measured at 512nm.
e) Estimation of apolipoprotein B and AI by immunoturbidimetric method.10
Principle:Immunoturbidimetric method is based on the reaction of a sample containing human Apolipoprotein A1 or B and specific antiserum to form an insoluble complex which can be measured turbidimetrically at 340 nm.
Statistical analysis:
Results will be subjected for stastical analysis. Students‘t’ test will be used to compare different biochemical parameters between cases and controls.
7.3 Does the study require any investigation or intervention to be conducted on patients or other humans or animals? If so, please describe briefly.
Yes
Estimation of Apoli[poprotein AI, B, total cholesterol, LDL cholesterol, HDL cholesterol and triglycerides in stroke patients and normal healthy controls.
7.4 Has ethical clearance been obtained from your institution in case of 7.3?
Yes
LIST OF REFERENCES:
  1. Park: Park’s textbook of preventive and social medicine. 19th edition. Jabalpur: Barasidas Bhanot; 2008. P314
  2. Peterlibby. The pathogenesis prevention and treatment of atherosclerosis. In:Fauci, Braunwald, K Asper, Hauser, Longo, Jameson, Loscalzo editors. Harrison’s principal of internal medicine. Vol 2, 17th edition. USA: Mc Graw Hill Company; 2008. P 2513
  3. Tsuyoshi Imamura, Yasufumi Doi, Hisatomy Arima, Koji Yonemoto, Jun Hata, Michiaki kubo, et al. LDL cholesterol and the development of stroke subtypes and coronary heart disease in a general Japanese population: The Hisayama study. Stroke 2009; 40: 382-388
  4. Wanna methee, Goya S, Shaper, Gerald A, Ebrahim S. HDL- cholesterol, Total cholesterol and risk of stroke in middle aged British men. Stroke2000; 31:1882-1888.
  5. Tanne, David, Yaari, Shlomit, Goldbourt, Uri. High density lipoprotein cholesterol and risk of ischemic stroke mortality: A 21 year follow up of 8586 men from isreli ischemic heart disease study. Stroke 1997; 28: 83-87.
  6. Jacob J Freiberg, Anne Tybjaerg-Hansen, Jan Skov Jensen,Borge G Nordestgaard. Non-fasting triglycerides and risk of ischemic stroke in general population. JAMA 2008;300(18):2142-2152.
  7. Adriano paula sabino, Marinez De Oliveira Sousa, Luciana Mo Moreira Lima, Daniel Dias Ribeiro, Luci Maria Santano Dusse, Marie Das Gracas Carvalho et al. Apo B/ApoAI ratio in young patients with ischemic cerebral stroke or peripheral arterial disease. Transitional research 2008; 152: 113-118.
  8. Nira Koren morag, Uri Goldbourt, Eran Graff, David Tanne. Apolipoproteins B and AI and the risk of ischemic cerebrovascular events in patients with preexisting atherthrombotic disease, Journal of neurological sciences 2008; 270: 82-87.
  9. Kuo-liong chien, Fung-Chang-Sang, Hsiu-Ching Hsu, Ta-chen Su, Ruey-S Lin, Yuan-The Lee. Apolipoprotein AI and B and stroke events in a community based cohort in thaiwan. Report of Chin-Shan community cardiovascular study. Stroke 2002; 33: 39-44.
  10. Nader Rifai, Russel Warnick G.Lipids, lipoproteins, apolipoproteins and other cardiovascular risk factors. In: BrutisCA, Ashwood ER, Bruns DE.editors. Teitz textbook of clinical chemistry and molecular diagnostics. 4th edition. New Delhi: Elesvier;2006. P 942-960
  11. Pedro- Botet J, Senti M, Nogues X, Rubies-Prat J, Roquer L D’Olhaberriaque J et al. Lipoproteins and Apolipoprotein profile in men with ischemic stroke. Role of lipoprotein (a) triglyceride rich lipoproteins and apolipoprotein E polymorphism.Stroke 1992; 23: 1556-1562.
  12. Bhatia M, Howard S C, Clark T G, Neale R, Qizilbash N, Murphy M F G et al. Apolipiproteins as predictors of ischemic stroke in patients with a previous transient ischemic attack. Cerebrovascular dis 2006; 21: 323-328.

9. / Signature of the Candidate
10. / Remarks of the Guide / Study will help in identifying the individuals at risk and to prevent the stroke attack by normalizing the lipid profile.
11. / Name and Designation of
(in block letters)
11.1 Guide
11.2 Signature
11.3 Co-Guide
11.4 Signature
11.5 Head of the Department
11.6 Signature / DR. SAVITRI. R. SAHUKAR,MD
READER,
DEPT. OF BIOCHEMISTRY
JJMMEDICALCOLLEGE, DAVANGERE.
DR. JAYAPRAKASH MURTHY. D.S.
Bsc, MBBS, MD
PROFESSSOR AND HEAD
DEPT. OF BIOCHEMISTRY
JJMMEDICALCOLLEGE, DAVANGERE.
12 / 12.1 Remarks of the Chairman & the principal
12.2 Signature