Lipids in the blood

pATHOLOGY

1. Determination of Lipids in the blood or L ipid profile

2. Lipid profile It includes the following tests: Total Lipids Total cholesterol Triglycerides HDL cholesterol LDL cholesterol VLDL- cholesterol IDL- Cholestrol

3. Cholesterol Cholesterol is a type of fat, found in your blood. It is produced by your body and also comes from the foods you eat (animal products). Cholesterol is needed by your body to maintain the health of your cells. Too much cholesterol leads to coronary artery disease . Your blood cholesterol level is related to the foods you eat or to genetic conditions (passed down from other generations of family members).

4. Cont.…. Has important function in body: important part in membrane of cells, organs and tissues in the body is used to make hormones, forms acids that are needed to absorb nutrients from food. Therefore, cholesterol deficiency is not good. Source: 70% synthesized in body, 30% from food (animal source as meat, eggs and dairy products)

5. Cholesterol Mostly found in Brain Bile Kidney Blood In blood it exist in two forms : In free form = 20-40% In esterified form = 60-80%

6. Estimation Methods Liebermann- Burchardt reaction: Abell method Iron-salt acid method: Toluene Sulfonic acid method Enzymatic end point method

7. Estimation Specimen: Serum or plasma is stable at 2-8°C for 7 days. Reagents and procedure: Follow the detailed procedure as per standard instructions provided by the manufacturer of the commercial kit in use . Reference Range: 3.8 – 5.2 mmol /L mg/dl x 0.026= mmol /L mmol /L X38.7=mg/dl

8. Normal Ranges Child :170 mg/ dL Adult :200 mg/ dL

9. Sources of Cholesterol Exogenous: brain, egg yolk, kidney, liver, milk and butter Endogenous: it is synthesized in all tissues but mostly in liver, intestine the liver daily synthesize about 900mg cholesterol

10. Determination( enzymatic reaction ) cholesterol ester → cholesterol+ Fatty Acid Cholesterol+O2 → cholesterol + H2O2 2 H2O2 + 4-aminoantipyrine → chromogen(red)

11. Clinical applications Hyper cholesterolaemia: Hyperthyroidism Nephritic syndrome Atherosclerosis Multiple myeloma….etc Hypercholesterolemia: Anemia's Starvation Hyperthyroidism…..etc

12. TRIGLYCERIDE Triglycerides are composed of three fatty acids and a glycerol. Mainly synthesized in small intestine and liver

13. Triglycerides → 3 fatty acids + glycerol Glycerol ATP → glycerol-3-phosphate + ADP Glycerol phosphate+ Di-hydroxy acetone phosphate + + Chromogen Red colour

14. Normal Values

15. Specimen Serum, fasting 10 to 12 hours. Reference Ranges Male = 45–204 mg/ dL Females = 42–159 mg/ dL National Cholesterol Education Program risk factors Optimal 150 mg/ dL High 150–199 mg/ dL Hyper triglyceridemic 200–499 mg/ dL Very high 499 mg/ dL

16. Clinical applications Hyperglyceridaemia: -after meal -alcoholism -atherosclerosis -pancreatitis -liver diseases …..etc

17. HDL CHOLESTEROL Good cholesterol, carry cholesterol from organs and blood to liver to get rid of it. It removes excess cholesterol from tissues (it cleans blood). High levels linked to a reduced risk of heart and blood vessel disease . The higher your HDL level, the better

18. HDL VS LDL

19. HDL (high density lipoprotein ) Synthesized in the liver and small intestine Transfer cholesterol from peripheral cells to the liver Act as a scavengers of cholesterol Inversely related to the risk of CHD Named as good of beneficial cholesterol

20. Measurement The Precipitation Reaction This method use phosphotungstic acid with magnesium chloride (MgCl2) to precipitate LDL and VLDL lipoproteins from the sample, leaving HDL in the supernatant. It is measured by ultracentrifugation, column chromatography, electrophoresis and enzymatic methods Centrifuged for at least 15 min

21. Cont…. The HDL supernatant is then assayed for cholesterol. The resulting answer (in mg/ dL ) represents the amount of HDL in the serum sample. The supernatant is tested for cholesterol concentration.

22. The Specimen Serum, plasma 10 to 12 hours fasting. Reference Ranges: Men = 31–63 mg/ dL Women = 37–83 mg/Dl National Cholesterol Education Program risk factors (adult male): Low risk 59 mg/ dL High risk 40 mg/ dL

23. Clinical applications Increased level of HDL: Estrogens Exercise Moderate consumption of alcohol Decrease level of HDL: Obesity Tension Hepatocellular diseases Heavy cigarette smoking

24. LDL CHOLESTEROL It transmit cholesterol to tissue directly related to the risk of CHD Named as bad cholesterol

25. Measurement LDL-C involves a calculation that includes total cholesterol, HDL cholesterol and triglyceride (TG) values formula: LDL-Cholesterol = total cholesterol – HDL- Ch + (TG/2.2) Or LDL=Total Cholestrol -(TG/5 -HDL) Cannot be used for TG over 400 mg/ dL

26. Example Calculation Total cholesterol 350 mg/ dL triglycerides 150 mg/ dL HDL-C 30 mg/ dL LDL-C = total cholesterol – [HDL-C + (TG/5)] LDL-C = 350– [30 + (150/5)] LDL-C = 350 – (30 30) LDL-C = 350 – 60 LDL-C=290 mg/dL

27. Normal values Reference Ranges: Male = 70–165 mg/ dL Females = 71–164 mg/ dL National Cholesterol Education Program risk factors (adult male): Optimal 100 mg/ dL Near optimal 100–129 mg/ dL Borderline high 130–159 mg/ dL High 160 mg/ dL Very high 190 mg/ dL

28. VLDL It is measured by ultracentrifugation, column chromatography, electrophoresis, enzymatic and calculation methods . Calculations : VLDL CHOLESTROL =TG/2.2

29. IDL CHOLESTEROL ESTIMATION Intermediate Density Lipoprotein It is measured by ultracentrifugation, column chromatography, electrophoresis , enzymatic and calculation methods. Formula : TG/5

30. Total Lipids Formula: Total Lipids = (TG + Total Cholesterol)x2 Also performed by Gravimetric method ,photometric method. Mostly its calculated

31. Limited clinical significance FATTY ACID ESTIMATION Fats are extracted by ethanol-ether mixture. The extract is saponified. Fatty acids are liberated by acidification, which are finally titrated with NaOH to calculate the concentration of fatty acids . Due to technical limitations as well as limited clinical significance, fatty acid estimation is not practical anymore. PHOSPHOLIPIDS ESTIMATION Fats are extracted by ethanol-ether mixture. After treating the supernatant with H2O2, the phosphates are determined calorimetrically to derive the values of phospholipid

32. ELECTROPHORESIS : The lipoproteins can be separated by electrophoresis on agarose or cellulose acetate membrane. Fractions are visualises by fat stains. Following bands are seen: • α-band for HDL • Pre-β band for VLDL • β-band for LDL and • Chylomicrons at application site ULTRACENTRIFUGATION: Lipoproteins have lower density so can be isolated from other plasma proteins by ultracentrifugation in a salt solution of specified density. The instrument is expensive and technique requires expertise.

33. LIPOPROTEINS ASSESSMENT 1. Apo A-I (HDL, Chylomicron) 2 . Apo A-II (HDL, Chylomicron) 3 . Apo B-48 (Chylomicron) 4 . Apo B-100 (LDL, IDL, VLDL) 5 . Apo C-I (Chylomicron, HDL, VLDL) 6 . Apo C-II (Chylomicron, HDL, VLDL) 7 . Apo C-IH (Chylomicron, HDL, VLDL) 8 . Apo E (chylomicron, HDL, VLDL) 9 . Apo (a) (LDL, IDL)