Lipids and Dyslipoproteinemia PDF

LIPIDS AND DYSLIPOPROTEINEMIA Professor: Dr. Michelle Joy M. Cauan Trans by: Cabacungan, Calsiyao, Carag, Carig, Suyu FUNCTIONS: TOPIC OVERVIEW...

LIPIDS AND DYSLIPOPROTEINEMIA Professor: Dr. Michelle Joy M. Cauan Trans by: Cabacungan, Calsiyao, Carag, Carig, Suyu FUNCTIONS: TOPIC OVERVIEW - Source of energy A. Lipids - Serves as insulator B. Lipoproteins - Absorption of fat-soluble a. Major Lipoproteins vitamins i. Chylomicrons ii. Very-Low-Density Lipoproteins - Cell constituents iii. Low-Density Lipoproteins - transport of important iv. High-Density Lipoproteins substances b. Minor Lipoproteins i. Intermediate-Density Lipoproteins c. Abnormal Lipoproteins LIPOPROTEINS i. Lipoprotein(a) ii. LpX Lipoprotein Molecules composed of lipids and proteins in varying C. Lipoprotein Metabolism proportion D. Blood Sampling and Storage a. Biologic Variation The function of lipoprotein particle is to transport lipids b. Fasting around the body in the blood c. Posture Contain cholesterol in two forms: d. Venous vs Capillary Samples e. Plasma vs Serum ✓ free cholesterol f. Storage ✓ Cholesterol ester E. Estimation of Plasma Lipids Have a micellar structure a. Cholesterol Measurements b. Triglyceride Measurements c. Phospholipids LIPOPROTEIN STRUCTURE F. Estimation of Lipoproteins and Lipoprotein Cholesterol spherical particles with a hydrophobic core and a. Ultracentrifugation Methods amphiphilic surface b. Electrophoretic Methods c. Polyanion precipitation methods Surface - single layer of phospholipids with proteins and d. Determining HDL-C values free cholesterol e. LDL-C Measurements Hydrophobic core - TG and cholesterol ester f. Selective Chemical precipitation g. Beta quantification h. Direct LDL-C measurement FOUR MAJOR CLASSES: G. Additional methods in the study of Dyslipidemia Chylomicrons H. Guidelines for cholesterol testing and management VLDL I. Metabolic Syndrome J. Hypertriglyceridemia LDL K. High cholesterol HDL a. Children MINOR CLASS: b. With high LDL-C L. High triglycerides IDL a. With normal cholesterol b. With high cholesterol M. Low total cholesterol and triglyceride N. Isolated Low HDL-C O. Isolated High HDL-C LIPIDS biological compounds which are soluble in nonpolar organic solvents, but relatively insoluble in polar solvents Cholesterol and triglycerides – not as free- floating molecules in the plasma ○ Part of water-soluble complexes called lipoprotein MAJOR CLASSES: - fatty acids - triglycerides - Cholesterol - phospholipids PAGE 1 BATCH TALAGHAY  Low-Density Lipoproteins Produced by the metabolism of VLDL The particles do not scatter light or alter the clarity of plasma Removed by the liver and macrophages High levels → increased CHD risk High-Density Lipoproteins Consists mostly of protein, cholesterol and phospholipids ApoA-I is the major lipoprotein which is synthesized in the liver and intestine Involved in reverse cholesterol transport 2 subclasses: HDL2 and HDL3 (smaller) HDL2 is more cardioprotective MINOR LIPOPROTEIN INTERMEDIATE-DENSITY LIPOPROTEINS Formed through the metabolism of VLDL Removed from circulation through interaction with the LDLR or further metabolized to LDL Concentrations contribute to the development of CHD MAJOR LIPOPROTEINS ABNORMAL LIPOPROTEIN Chylomicrons Lipoprotein (a) Produced by the intestine that transport lipids of dietary Similar to LDL in terms of density and overall composition origin to the tissues of the body Synthesized in the liver Rich in triglycerides but poor in free cholesterol, Binds to LDLR by its apoB-100 component but lower phospholipids and protein affinity than LDL Secreted into mesenteric lymphatics and reach the Associated with increased risk of CHD, cerebrovascular circulation at the thoracic duct disease and stroke Apolipoproteins: apoB-48, apoA-I, apoA-IV, apoC-I, apoC-II, apoC-III, and apoE LpX Lipoprotein apoC-II serves as an activator of lipoprotein lipase Abnormal lipoprotein found in patients with obstructive biliary disease and with familial lecithin/cholesterol acyltransferase (LCAT) deficiency Very-Low-Density Lipoproteins Produced by the liver β-VLDL (“FLOATING β” LIPOPROTEIN) Supplies the tissues of the body with triglycerides of Accumulates in type 3 hyperlipoproteinemia endogenous origin and cholesterol Richer in cholesterol than VLDL Produce turbid plasma when present in excessive Results from the defective catabolism of VLDL amounts PAGE 2 BATCH TALAGHAY | Surnames  IMPORTANT PROTEINS IN Blood Sampling and Storage LIPOPROTEIN METABOLISM 1. Biologic Variation APOLIPOPROTEINS Cholesterol level rises with age Apos constitute the major protein component of Women have lower levels than men (except in lipoproteins childhood and after the early 50’s) Age related variation is the basis of NCEP LIPOLYTIC ENZYMES recommendation that cholesterol screening be Major enzymatic systems (including LCAT, LPL, HPL, repeated every 5 years. and EL) that are known to participate in plasma 2. Fasting lipoprotein metabolism Ideally, fast for 12 hours before venipuncture Chylomicrons are completely cleared within 6 to 9 hours, their presence after 12-hour fast is considered abnormal NCEP Adult Treatment panel III (ATP III), recommends patients to fast for at least 9 hours. 3. Posture Standing to the recumbent position decreases TG for about 50% and 10% in the concentrations of TC, LDL-C, HDL-C, apoA-a, and apoB. Current NCEP guidelines recommend patients be seated for 5 minutes to prevent hemoconcentration Prolonged venous occlusion can lead to hemoconcentration and cholesterol increases for about 10% to 15%. Tourniquets should not be applied for longer than 1-2 minutes 4. Venous versus Capillary Samples Capillary blood samples are little lower than venous samples 5. Plasma versus Serum Either plasma or serum can be used when only TAG, cholesterol and HDL are measured and LDL-C is calculated from these 3 measurements Plasma is preferred when lipoproteins are measured by ultracentrifugation or electrophoretic methods EDTA is the preferred anticoagulant Citrate – exerts large osmotic effects causing false low concentrations of plasma lipid and lipoproteins Heparin – can alter the electrophoretic mobilities of the lipoproteins. 6. Storage When serum or plasma must be stored for long periods, it should be maintained at a temperature of -70ºC For short term storage (1-2 months) the samples can be kept at -20ºC Estimation of Plasma Lipids CHOLESTEROL MEASUREMENT PAGE 3 BATCH TALAGHAY | Surnames 1. Chemical PHOSPHOLIPIDS Abell-kendall method – reference method used by CDC and measured with Most of the phospholipids in human plasma is Liebermann-Burchardt reagent phosphatidylcholine (70% to 75%) or sphingomyelin Liebermann-Burchardt reaction – uses sulfuric (18% to 20%) acid and acetic anhydride to produce an unstable Measurement is NOT routinely performed green cholestadienyl mono sulfonic acid; color ○ Analysis provides a little information in cases of stabilized by sodium sulfate dyslipoproteinemia 2. Enzymatic ○ In various pathologic conditions, levels are not The most common method of quantifying the markedly altered as that of cholesterol and TG cholesterol oxidase reaction is to measure the amount of hydrogen peroxide produced 1. Ultracentrifugation Methods Lipoprotein can be readily separated on the basis of their densities using ultracentrifugation Two advantage properties of Lipoproteins: ○ Have lower densities than other plasma macromolecules ○ Each class of lipoproteins has a different density 2. Electrophoretic Methods Not required for diagnosis of dyslipoproteinemia 3. Interfering substances Most commonly Sterols - increases cholesterol values supported medium is Ascorbic acid - ↓ total cholesterol levels agarose gel because Bilirubin - increases cholesterol values of its speed, sensitivity Hemoglobin - false low or false high and ability to resolve the lipoprotein classes. TRIGLYCERIDE MEASUREMENT Chylomicrons remain at the origin; HDL 1. Chemical migrates fastest; LDL CDC reference method uses a chloroform is slowest; and VLDL between HDL and LDL. extraction procedure followed by silicic acid chromatography to isolate TG. 3. Polyanion Precipitation Methods ○ formaldehyde produced in the reaction Some lipoproteins are precipitated with polyanions is measured by reaction with a sulfuric such as heparin sulfate, dextran sulfate, acid to produce a pink chromophore. phosphotungstate in the presence of divalent 2. Enzymatic cations such as Ca, Mg, and Mn. The most common method is the hydrolysis of The more dissimilar the lipoproteins proteins from triglyceride to free fatty acids and glycerol, one another the better the separation followed by the phosphorylation of glycerol to 4. Determining HDL-C Values glycerophosphate NCEP recognizes HDL-C as an independent risk factor for CHD ○ 60 mg/dL = protective Homogeneous assays are the most popular method for measuring HDL-C 5. LDL-C Measurement Separated by ultracentrifugation >160 mg/dL with no risk factors is an indication for therapy >130 mg/dL, therapy is initiated if with 2 or more CHD 3. Interfering substances risk factors Ascorbic acid - decreases TG Measured using Friedewald formula Bilirubin - increases TG Hemolysis - decreases TG PAGE 4 BATCH TALAGHAY | Surnames  FRIEDEWALD FORMULA 2. MEASUREMENT OF LIPOPROTEIN SUBCLASSES Total cholesterol= HDL + LDL + VLDL NMR technology is used to identify lipoprotein VLDL= TG (mg/dl) /5 or TG (mmol/L) / 2.2 subclasses Particle subclass numbers tend to increase with Note: formula not valid if Triglycerides (TG) are > 400 mg/dl (4.5 increasing particle size; thus LDL particles of the L2 mmol/L) subclass are larger than L1 particles 3. STANDING PLASMA TEST LDL cholesterol calculation (Friedewald Formula) An aliquot of plasma (2mL) is placed into a 10 x LDL-C= Total Cholesterol -HDL-C- (TG/5) 75-mm test tube and allowed to stand in the Not valid if TG > 400 refrigerator at 4ºC undisturbed overnight Chylomicrons accumulate as a floating “cream” layer and can be detected visually A plasma sample that remains turbid after standing overnight contains an excessive amount of VLDL; if a floating “cream” layer also forms, CMs are present. 4. DETECTION OF β-VLDL AND Lp(a) β-VLDL has a density of VLDL but migrates electrophoretically with LDL in the β region; its presence is usually associated with dysbetalipoproteinemia Lp(a) has a density similar to LDL but migrates similarly to VLDL on electrophoresis 5. VLDL-C/PLASMA TRIGLYCERIDE RATIO 6. SELECTIVE CHEMICAL PRECIPITATION Useful in evaluation of type 3 hyperlipoproteinemia LDL-C is calculated as the difference between TC and Normal ratio: 0.230 to 0.575 the amount remaining after precipitation of LDL-C Type 3 subjects have ratios greater than 0.689 Disadvantage: Allows only for the separation of apoB- (usually in the range of 0.689 to 0.919 containing lipoproteins separating HDL from non-HDL 6. APOLIPOPROTEIN ANALYSIS particles Apolipoproteins are usually measured by 7. BETA QUANTIFICATION immunoassay, such as immunonephelometry which In this method the plasma is ultracentrifuged for at rely on measurement of turbidity caused by least 18 hours at 105K x g apolipoprotein-antibody complexes VLDL and CMs accumulate as a floating layer, leaving Limitation: inherent turbidity of lipemic samples or predominantly LDL and HDl in the solution even non lipemic samples after repeated freezing and The solution is measured for cholesterol and is thawing precipitated for LDL lipoproteins. LDL-C is calculated 7. MODIFIED LIPOPROTEIN according to the difference between these two Includes oxidized LDL and oxidized HDL measurements Sensitive ELISA with monoclonal antibodies specific 8. DIRECT LDL-C MEASUREMENT to oxLDL has been developed to quantify its Useful when triglycerides are elevated because they concentration in the plasma are not subject to interference by triglycerides even at Measurement of oxidized lipoproteins might be a relatively high concentrations (600 mg/dL) useful marker of CHD Uses combination of 2 reagents First reagent selectively removes non-LDL GUIDELINES FOR CHOLESTEROL TESTING AND lipoproteins and inhibits LDL from reacting with MANAGEMENT other enzymes Classifies statin therapy as: Second reagent releases cholesterol from LDL so ○ High intensity – lowers LDL-C by an average of that it can be measured enzymatically ≥50% ○ Moderate intensity - lowers LDL-C by an ADDITIONAL METHODS IN THE STUDY OF DYSLIPIDEMIA average of 30-50% ○ Low intensity - lowers LDL-C by an average of 1. MEASUREMENT OF LIPOPROTEIN PARTICLE NUMBER 500 HIGH CHOLESTEROL IN CHILDREN Current recommendation is screening starting 2 years of age if there is family history of premature cardiovascular disease or those with at least one parent with high blood cholesterol Familial Defective ApoB Autosomal dominant disorder of the apoB gene on chromosome 2 that interferes with the recognition of apoB-100 by the LDLR Results from missense mutation in the LDLR-binding domain of apoB-100 Statin drugs are effective PAGE 6 BATCH TALAGHAY | Surnames  Sitosterolemia Diabetic Dyslipidemia Extremely rare autosomal recessive disorder where Consists of atherogenic dyslipidemia (high TG, low HDL; phytosterols (plant sterols) are absorbed and accumulate and small, dense LDL) in persons with type 2 diabetes in plasma and peripheral tissues Patients with diabetes who are 40 to 75 years of age Results from mutations in the ABCG8 or ABCG5 gene have LDL-C of 70 to 189 mg/dL are recommended to both located at chromosome 2p21 take moderate-intensity statins to reduce ASCVD risk Most patients have had high plasma LDL-C levels during Familial Hypertriglyceridemia childhood Hemolytic anemia, stomatocytes and Usually occurs along with other lipoprotein abnormalities macrothrombocytopenia can be the initial presentation Isolated hypertriglyceridemia (type 4 hyperlipidemia) is Diagnosis considered if patient has xanthomatosis and defined by the familial occurrence of isolated high VLDL hypercholesterolemia while parents have normal levels with TG values most commonly in the 200 to 500 cholesterol mg/dL range Tx: Diet low in sterol and Ezetimibe (sterol absorption Pathophysiology: VLDL triglyceride production is inhibitor) increased in the setting of normal apoB production resulting in the formation of “fluffy,” TG-rich VLDL particles Lipoprotein Lipase Deficiency (Hyperlipoproteinemia Type 1 or Hyperchylomicronemia) Rare autosomal recessive disorder that presents in childhood with abdominal pain and pancreatitis Defective/absent LPL creates an inability to clear chylomicrons, creating classic “type 1” chylomicronemia syndrome Fasting TG levels may be over 100 mg/dL and may rise Autosomal Dominant Hypercholesterolemia to 10,000 mg/dL postprandially Patients do not develop premature CHD, implying AD disorder of the PCSK9 gene on chromosome 1 that is chylomicrons themselves are not atherogenic involved in cholesterol homeostasis in the liver Tx: Low-fat diet, fat soluble supplements and drug Individuals with gain-of-function mutations present therapy clinically with increased plasma levels of LDL-C and exhibit a higher risk of CHD Autosomal Recessive Hypercholesterolemia Involves the ARH gene found on chromosome 1 Also known as LDLRAP1 In these patients, LDLR expression is normal but LDL clearance rates are low Onset of atherosclerotic disease presents later than in homozygous FH patients Can present with large, bulky xanthomas Respond to lipid lowering medications HIGH TRIGLYCERIDES WITH NORMAL CHOLESTEROL These disorders are related to elevations of triglyceride-rich particles – namely, chylomicrons or VLDL (Fredrickson types 1 and 4) Usually due to hyperbetalipoproteinemia (VLDL) and may be due to secondary causes such as excess alcohol or a high-carbohydrate diet LDL and LDL-C are typically normal PAGE 7 BATCH TALAGHAY | Surnames  ApoC-II Deficiency Dysbetalipoproteinemia (Type 3) ApoC-II is an activating cofactor for LPL Primarily affects adults and is more common in men Absence of apoC-II creates a functional LPL deficiency Pathognomonic feature: broad abnormal band between which presents similarly to LPL deficiency VLDL and LDL known as “abnormal migrating beta Clinical manifestations: children and young adults lipoprotein,” or β-VLDL. presents with abdominal pain and pancreatitis Screening method: VLDL-C/Triglyceride ratio Patients can be treated with plasma transfusions during ○ Normal = 0.2 severe hypertriglyceridemia providing apoC-II, which will ○ Type 3 patients = ≥3 activate endogenous LPL Premature atherosclerosis is highly prevalent ApoC-III Excess ApoC-III excess interferes with the activity of LPLand binds to the carboxy-terminal portion of apolipoprotein B, thereby preventing the binding of lipoproteins to the LDLR May be an independent risk factor for CHD Increased apoC-III are seen in patients with T2DM, hyperbilirubinemia, kidney deficiency, thyroid dysfunction and oral contraceptive use in women ApoA-V Highly hydrophobic protein that has a preference for binding to lipids and HDL particles Function is hypothesized to be involved in VLDL assembly and activation of LPL-mediated TG hydrolysis Hepatic Lipase Deficiency Low levels promote hypertriglyceridemia Familial disorder resulting from mutations of HL gene Associated w/ combined hyperlipidemia characterized by: ○ TC levels of 250 to 1500 mg/dL HIGH CHOLESTEROL WITH HIGH TRIGLYCERIDES ○ TG levels of 400 to 8000 mg/dL These disorders are related to elevations of LDL and Physical stigmata include palmar and tuberoeruptive triglycerides (Fredrickson types 2B and 3) xanthoma Familial combined hyperlipidemia (2B) is the most Risk of atherosclerosis is increased common primary hyperlipoproteinemia TC/TG ratio is NOT increased The relatively rare dysbetalipoproteinemia (type 3) is HDL are normal or increased; large and enriched with TG characterized by abnormal LDL (IDL) that appears as broad beta electrophoretic band and distinguishes it from Cholesterol 7-Alpha-Hydroxylase Deficiency familial combined hyperlipidemia Recessive disorder of the CYP7A1 gene Associated with increased cardiac risk because of Very little is known about the disease elevated LDL Few patients are resistant to stain therapy Lack of cholesterol 7-alpha-hydroxylase is thought to Familial Combined Hyperlipidemia (Type 2B) reduce hepatic LDLR activity Relatively common disorder where affected individuals may have simple hypercholesterolemia, simple hypertriglyceridemia, or a mixed defect. LOW TOTAL CHOLESTEROL AND TRIGLYCERIDE Lacks a definitive biochemical marker These disorders are associated with defective apoB Affected families must have more than one pattern of lipid synthesis or metabolism leading to low or nonexistent disorder to meet the diagnostic criteria levels of apo-B lipoproteins such as CM, VLDL, and LDL Triglycerides and cholesterol are low Acquired Combined Hyperlipidemia Fat soluble vitamins are common Common in patients who have metabolic syndrome Pathophysiology: Liver increases production of VLDL Abetalipoproteinemia which eventually matures to LDL until this process is Also known as “Bassen-Kornzweig Syndrome” saturated. When LDL gets high enough, VLDL follows Rare AR disorder involving mutations in MTTP gene causing both hypercholesterolemia and located on chromosome 4 hypertriglyceridemia. Laboratory testing: PAGE 8 BATCH TALAGHAY | Surnames  ○ Decreased apoB, TG and TC (typically < 50 Familial Hypoalphalipoproteinemia mg/dL) Patients develop fat-soluble vitamin deficiencies caused Affected men have HDL-C level