Need for the study



RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES,

BANGALORE, KARNATAKA

ANNEXURE II

PROFORMA FOR REGISTRATION OF SUBJECTS

FOR DISSERTATION

|1. |NAME OF THE CANDIDATE |Dr. RAHIMTAJ BASARIKATTI |

| |AND ADDRESS |POST GRADUATE STUDENT, |

| |(in block letters) |DEPARTMENT OF BIOCHEMISTRY, |

| | |J.J.M. MEDICAL COLLEGE, |

| | |DAVANGERE – 577004, |

| | |KARNATAKA. |

|2. |NAME OF THE INSTITUTION |J.J.M. MEDICAL COLLEGE, |

| | |DAVANGERE- 577004. |

|3. |COURSE OF STUDY AND SUBJECT |M.D. BIOCHEMISTRY |

|4. |DATE OF ADMISSION TO COURSE |26-07-2010 |

| | |“RELATIONSHIP BETWEEN INSULIN RESISTANCE AND HYPOTHYROIDISM” |

|5. |TITLE OF THE TOPIC | |

| | |

|6. |BRIEF RESUME OF THE INTENDED WORK : |

| | |

| |6.1 Need for the study : |

| |Hypothyroidism is defined as a deficiency of thyroid activity. It results from reduced secretion either of total thyroxine(T4) and |

| |triiodothyronine(T3).Decreased T4 and T3 concentrations lead to hypersecretion of pituitary thyroid stimulating hormone(TSH) and an |

| |amplified increase in serum TSH levels. |

| |Subclinical hypothyroidism is term used for a condition in which there is elevation in TSH, yet normal circulating levels of thyroid|

| |hormones.1 |

| | |

| |Hypothyroidism affects over 1% of general population and 5%of individual over age of 60 years.2 |

| |In Colorado study 9.5% of participants were hypothyroid, most of whom were subclinically hypothyroid vary according to age and sex,4%|

| |in 18-24 years old,16-22% in men and women over age of 74 not taking L-thyroxine respectively.3 |

| |Clinical and subclinical hypothyroidism are established risk for insulin resistance, hyperlipidemia, hypercoagulability and low grade|

| |inflammation4 |

| |.Insulin-stimulated glucose transport in monocyte from patients with clinical and subclinical hypothyroidism was found to be |

| |decreased due to impaired translocation of GLUT4 glucose transports on the plasma membrane. T3 and insulin have synergistic role in |

| |glucose homeostasis since these hormones possess similar action sites in the regulation of glucose metabolism both at cellular and |

| |molecular levels. Decreased T3 could lead to impaired insulin stimulated glucose disposal. Even subtle decrease in the levels of |

| |thyroid hormones within the physiological range have shown to correlate inversely with HOMA index(homeostasis model |

| |assessment).Insulin resistance is observed in both fasting and post glucose state. These findings are consistent with the recent |

| |studies reporting an increased cardiovascular risk in these conditions.5 Insulin resistance is due to defect in the ability of |

| |insulin to increase glucose utilization in peripheral tissue, mainly muscles.6 |

| |Insulin resistance leads to an increased production of hepatic cholesterol.4Patients with mild thyroid failure and even subjects with|

| |high normal serum TSH value have evidence of comparable atherogenic factors and high serum cholesterol.5 |

| |The body mass index(BMI) of subjects both in clinical and Subclinical hypothyroidism were significantly more than in euthyroid |

| |controls.4 |

| |According to studies correlation between TSH and insulin as well as TSH and HOMA-IR in female population suffering from both clinical|

| |and subclinical hypothyroidism is found. TSH and HOMA-IR showed significant correlation with total cholesterol in clinical |

| |hypothyroidism. Development of insulin resistance leads to many of metabolic abnormalities.4 |

| |This study is undertaken to evaluate relationship between fasting glucose, fasting insulin, insulin resistance, BMI and total |

| |cholesterol levels in hypothyroidism. |

| | |

| |6. 2 Review of literature : |

| |Studies have shown that patients with subclinical hypothyroidism having insulin resistance is comparable with clinical |

| |hypothyroidism. This findings could justify increased risk for insulin resistance associated disorders cardiovascular disorders as |

| |observed in these patients.5 |

| |Studies have shown that despite increase in plasma insulin level in hypothyroidism net glucose uptake in forearm muscles and adipose |

| |tissue was increased after meals suggesting the presence of insulin resistance.6 |

| |Studies have shown that elevated TSH levels were associated with deleterious effects on serum lipids in large population based sample|

| |of older women.7 |

| |Studies have shown that subclinical hypothyroidism in about one sixth of metabolic syndrome patients8 |

| |Studies have shown that subclinical hypothyroidism is highly prevalent among elderly women and is associated with a greater frequency|

| |of aortic atherosclerosis and myocardial infarction.9 |

| |Studies have shown that glucose metabolism in human is dependent on thyroid status. Hypothyroidism induced post-absorptive decrease |

| |in both glucose disposal and endogenous glucose appearance impaired the ability of insulin to stimulate glucose disposal related to |

| |insulineamia10 |

| | |

| |6.3 Objectives of the study : |

| |To study the levels of serum T3,T4,TSH and to evaluate the relationship between fasting glucose ,fasting insulin, insulin |

| |resistance(HOMA-IR), BMI and total cholesterol levels in clinical and subclinical hypothyroidism patients. |

| | |

| | MATERIALS AND METHODS |

| |7.1.Materials and methods |

| | |

| |Source of data |

| |The study will be carried out for a period of one year in Bapuji Hospital and Chigateri General Hospital Davangere(Both hospitals are|

| |attached to J.J.M.medical college) |

| |The study will be carried out in clinically proven cases of hypothyroidism after taking informed consent. |

| |30 clinical hypothyroidism, 30 subclinical hypothyroidism patients and 30 age and sex matched euthyroid healthy individuals as |

| |controls will be selected based on inclusion and exclusion criteria. |

| |Inclusion Criteria |

| |Cases-newly diagnosed, clinically proved cases of hypothyroidism in the age group of 18-70 years. |

| |Patients with signs and symptoms of goiter and myxedema. |

| |Clinical hypothyroidism patients with (increased serum TSH and decreased T3 and T4 levels) |

| |Subclinical hypothyroidism patients with((increased serum TSH and normal T3 and T4 levels) |

| |Controls-will include healthy euthyroid age and sex matched individuals without any major illness and not on any medications. |

| |Exclusion Criteria |

| |Diabetes, hypertension. |

| |Patients on thyroxine treatment, hypolipidemics, anti epileptic drugs, women on oral contraceptives. |

| |Pregnant women, previous thyroid surgeries, and other systemic illness like liver disorders and kidney disorders. |

| | |

| |7.2 Method of data collection: |

| |Under all aseptic precautions about 4ml of venous blood will be collected in a sterile bulb after overnight fasting of 12 hours. 2ml |

| |will be collected in EDTA vial(for plasma) 2 ml will be collected in plain vial (for serum),it will be subjected for centrifugation ,|

| |serum and plasma will be separated and T3,T4,TSH, insulin, and cholesterol will be estimated from serum and fasting glucose from |

| |plasma . |

| |(a)Estimation of serum T3,T4,andTSH by immunoenzymometric assay(chemiluminescence immunoassay).11 |

| |Principle: In this procedure, the immobilization takes place during the assay at the surface of a microplate well through the |

| |interaction of streptavidin coated on the well and exogenously added biotinylated monoclonal antibody coupled to anylate of |

| |interest. Upon mixing monoclonal biotinylated antibody, the enzyme-labeled antibody and a serum containing the native antigen, |

| |reaction results between the native antigen and the antibodies, without competition or steric hindrance, to form a soluble sandwich |

| |complex. Simultaneously, the complex is deposited to the well through the high affinity reaction of streptavidin and biotinylated |

| |antibody. After equilibrium is attained, the antibody-bound fraction is separated from unbound antigen by washing or aspiration. |

| |The enzyme activity, determined by the reaction with substrate that generates light in the antibody-bound fraction is directly |

| |proportional to the native antigen concentration. |

| |(b) Estimation of serum fasting insulin by solid phase two-site enzyme immunoassay12 |

| |Principle: In this procedure, the immobilization takes place during the assay at the surface of a microplate well through the |

| |interaction of streptavidin coated on the well and exogenously added biotinylated monoclonal insulin antibody. Upon mixing |

| |monoclonal biotinylated antibody, the enzyme-labeled antibody and a serum containing the native antigen, reaction results between the|

| |native antigen and the antibodies, without competition or steric hindrance, to form a soluble sandwich complex. Simultaneously, the |

| |complex is deposited to the well through the high affinity reaction of streptavidin and biotinylated antibody. After equilibrium is |

| |attained, the antibody-bound fraction is separated from unbound antigen by washing or aspiration. The enzyme activity determined by |

| |the reaction with substrate that generates light, in the antibody-bound fraction is directly proportional to the native antigen |

| |concentration. |

| |(c) Estimation of fasting plasma glucose by Glucose oxidase-peroxidase method12 |

| |Principle: Glucose oxidase enzyme oxidizes glucose to gluconic acid and hydrogen peroxide. The colorimetric indicator, quinonemine |

| |is generated from 4-aminoantipyrine and phenol by hydrogen peroxide under catalytic action of peroxidase. Intensity of colour |

| |generated is directly proportional to glucose concentration. |

| | |

| |(d) Calculation of insulin resistance by HOMA & HOMA2 model13 |

| |Formula: HOMA-IR = [ fasting plasma glucose(mmol/l) x fasting insulin(mU/l) ] / 22.5 |

| |HOMA2 is the updated computer model of HOMA which is available from ocdem.ox.ac.uk. |

| |(e) Estimation of total cholesterol by cholesterol oxidase/phenol aminoantipyrine method 14 |

| |Principle: Cholesterol esterase hydrolyses cholesterol esters to free cholesterol and fatty acids. Then cholesterol is oxidized by |

| |cholesterol oxidase forming cholesterol-4-ene-3-one and hydrogen peroxide. In presence of enzyme peroxidase, hydrogen peroxide causes|

| |oxidative coupling of phenol and antipyrine to form red colored quinoneimine dye which is |

| |measured at 520nm. |

| |(f) Calculation of body mass index(BMI)15 |

| |Formula: BMI=weight in kgs/(height in meter)2 |

| |STATISTICAL ANALYSIS : |

| |Statistical analysis: |

| |One way ANOVA will be used for multiple groups comparison and student ‘t’ test for group wise comparison. |

| |Correlation analysis will be done to assess any relationship between insulin resistance, serum cholesterol, and BMI with |

| |hypothyroidism . |

| |7.3 Does the study require any investigations or interventions to be conducted on patients or other human or animals? if so, please |

| |describe briefly |

| |Yes |

| |Estimation of serum T3, T4,TSH, fasting glucose, fasting insulin, total cholesterol and BMI in hypothyroidism patients and normal |

| |euthyroid individuals using human venous blood sample. |

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| |7.4 Has ethical clearance been obtained from your institution in case of 7.3 |

| |Yes |

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|8. |LIST OF REFERENCES: |

| |1.Reza A,Mansourian, Ezatollah et al.Serum lipid level alteration in subclinical hypothyroid patients in Gorgan(South East of |

| |Caspian Sea).Journal of Chinese Clinical Medicine;2008,4 Vol 3 No.4 |

| |2.Fitzgerald AP.Endocrine disorders.In:Mcphee JS, Papadakis, Laurance M,Tierney Jr.editors.Current Medical Diagnosis and |

| |Treatment.47th eds.LANGE CMDT 2008;p 960 |

| |Canaris JG, Monowitz RN, and Ridgway CE.The Colorado Thyroid Disease Prevalence Study.Arch Internal Med.2000;160:526-534. |

| |B M Singh,B Gowswami and Mallika.Association between insulin resistance and hypothyroidism in females attending a tertiary care |

| |hospital.Indian Journal of clinical Biochemistry 2010;25(2):141-145 |

| |Maratou E, Hadjidakis JD, Kollias A et al.Studies of insulin resistance in patients with clinical and subclinical hypothyroidism. |

| |European Journal of Endocrinology.2009;160.785-790. |

| |Dimitriadis G,Mitrou P,Lambadiari V et al. Insulin action in adipose tissue and muscle in hypothyroidism.J Clinical Endocrinol metab |

| |2006;92:4930-4937. |

| |Bauer CD, Ettinger Band Browner SW.Thyroid functions and serum lipids in older women:A Papulation based study. The American Journal |

| |of Medicine.1998;104:546-551. |

| |Uzunlulu M, Yorulmz E and Ogus A.Prevalence of subclinical hypothyroidism in patients with metabolic syndrome. Endocrine |

| |Journal.2007;54:71-76 |

| |Hak AE, Pouls HAP, Visser JT et al. Subclinical hypothyroidism is an independent risk factor for atherosclerosis and myocardial |

| |infarction in elderly women: Rottendam study. Annals of Internal Medicine.Society2000;132:270-278. |

| |Rochon C, Tauveron I, Dejax C et al. Response of glucose disposal to hyper insulineamia in human hypothyroidism and hyperthyroidism. |

| |The biochemical society and medical research society 2003;1047:7-15 |

| |Demers LM and Spencer C.The Thyroid, pathophysiology and thyroid function testing.In:Burtis CA, Ashwood ER and Bruns DE,eds.Tietz |

| |textbook of clinical chemistry and molecular diagnostics,4th edn. New Delhi:Elsiever Co,2006;2053-2095 |

| |David B Sacks.Carbohydrates. In:Burtis CA, Ashwood ER and Bruns DE,eds.Tietz textbook of clinical chemistry and molecular |

| |diagnostics,4th edn.New Delhi:Elsiever Co,2006;837-901 |

| |Wallace TM, Levy JC and Mathews DR.use and abuse of HOMA modeling .Diabetes Care.2004;27:1487-95. |

| |Nader R, Warnick GR.Lipids,lipoprotiens and other cardiovascular risk factors. In:Burtis CA, Ashwood ER and Bruns DE,eds.Tietz |

| |textbook of clinical chemistry and molecular diagnostics,4th edn.New Delhi:Elsiever Co,2006;916-52 |

| |Park.Epidemiology of chronic non communicable diseases and conditions Park’s Textbook of Preventive and Social Medicine.19th eds. p |

| |332-336 |

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|9. |SIGNATURE OF THE CANDIDATE | |

| | |This study is recommended to know the biochemical alterations |

|10 |REMARKS OF THE GUIDE |of T3, T4,TSH, insulin resistance(HOMA),BMI and total |

| | |cholesterol levels in hypothyroidism. |

|11 | | |

| |NAME & DESIGNATION. |Dr. REKHA M.,MBBS MD |

| | |READER, |

| |11.1 GUIDE |DEPARTMENT OF BIOCHEMISTRY |

| | |J.J.M.MEDICAL COLLEGE |

| | |DAVANGERE.577 004 |

| | | |

| | | |

| |11.2 SIGNATURE | |

| |11.3 CO-GUIDE |Dr. DHANANJAY P E MD |

| |(if any) |PROFESSOR, |

| | |DEPARTMENT OF MEDICINE |

| | |J.J.M.MEDICAL COLLEGE DAVANGERE.577 004 |

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| |11.4 SIGNATURE | |

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| |11.5 HEAD OF THE DEPARTMENT |Dr. D.S. JAYAPRAKASH MURTHY |

| | |B.Sc.,M.B.B.S.,M.D., |

| | | |

| | |Professor & HOD, |

| | |Department of Biochemistry, |

| | |JJM Medical College, |

| | |Davangere. |

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| |11.6 SIGNATURE | |

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|12 |REMARKS OF THE CHAIRMAN & | |

| |PRINCIPAL | |

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| |12.1 SIGNATURE | |

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