Chem chapter 8

• Lipids are hydrophobic molecules vital for energy storage, cell membrane construction, and hormone creation.
• Clinically significant lipids encompass fatty acids, triglycerides, phospholipids, cholesterol, and lipoproteins.
• Coronary heart disease (CHD) risk factors increase the likelihood of developing CHD, in which plaque builds up inside the coronary arteries.
• Plaque is made up of cholesterol, fat, calcium, and other substances and can narrow the arteries.
• Reduced blood flow can cause chest pain (angina), shortness of breath, heart attack, and other issues.

Fatty Acids

• Fatty acids consist of a hydrocarbon chain with a carboxyl group.
• Fatty acids are carboxylic acids with a long aliphatic tail, which can be saturated or unsaturated.
• Classification is based on hydrocarbon chain length and the presence of double bonds.
• Fatty acids are key components of lipids and are crucial for energy storage, cell membrane structure, and signaling.
• Saturated fatty acids lack double bonds, while unsaturated ones contain at least one.
• Unsaturated fatty acids include monounsaturated (one double bond) and polyunsaturated (multiple double bonds).
• Fatty acids serve as key energy sources and building blocks for complex lipids.
• Elevated saturated fatty acid levels correlate with increased cardiovascular disease risk.
• Essential fatty acids, such as omega-3 and omega-6, must come from the diet.
• They are vital in brain function, inflammation, and blood clotting.

Saturated Fatty Acids

• Saturated fatty acids contain only single bonds between carbon atoms in their hydrocarbon chain.
• Each carbon atom is bonded to the maximum number of hydrogen atoms.
• The general formula is CH3(CH2)nCOOH, where n is an even number.
• Saturated fatty acids are typically solid at room temperature due to their ability to pack tightly together.
• Examples include lauric acid (12 carbon atoms), myristic acid (14 carbon atoms), palmitic acid (16 carbon atoms), and stearic acid (18 carbon atoms).
• Saturated fats are commonly found in animal products like meat and dairy, as well as some plant-derived oils such as coconut oil and palm oil.
• Their structure gives them higher melting points compared to unsaturated fatty acids.
• The straight, unkinked chains allow for close packing, increasing intermolecular (Van der Waals) forces.
• High intake is associated with increased levels of LDL cholesterol, potentially increasing the risk of cardiovascular diseases.

Unsaturated Fatty Acids

• Unsaturated fatty acids contain one or more double bonds between carbon atoms in their hydrocarbon chain.
• The presence of double bonds introduces "kinks" or bends in the chain.
• Unsaturated fatty acids are typically liquid at room temperature.
• Monounsaturated fatty acids (MUFAs) contain only one double bond.
  • Oleic acid (18 carbon atoms, one double bond) is common in olive oil.
• Polyunsaturated fatty acids (PUFAs) contain two or more double bonds.
  • Linoleic acid (18 carbon atoms, two double bonds).
  • Alpha-linolenic acid (18 carbon atoms, three double bonds).
  • Arachidonic acid (20 carbon atoms, four double bonds).
• Omega-3 and omega-6 fatty acids are types of polyunsaturated fatty acids, named based on the position of the last double bond from the omega (methyl) end of the fatty acid chain.
• Unsaturated fats are commonly found in plant-based oils, such as olive oil, sunflower oil, and flaxseed oil, as well as in fish.
• The presence of double bonds reduces their melting points because the kinks in the chain disrupt the packing.
• This decreases the intermolecular forces, making them liquid at room temperature.
• The configuration around the double bond can be either cis or trans.
  • Cis double bonds are more common and create a more significant bend in the fatty acid chain.
  • Trans double bonds are less common and result in a straighter chain, which is similar to saturated fatty acids.
• Trans fats are primarily produced industrially through partial hydrogenation of vegetable oils and are associated with adverse health effects.
• Unsaturated fats, particularly MUFAs and PUFAs, are generally considered healthier than saturated fats.
• They can help lower LDL cholesterol and reduce the risk of heart disease.
• Omega-3 fatty acids, found in fish oil, are known for their anti-inflammatory properties and benefits for brain health.

Triglycerides

• Triglycerides are composed of glycerol and three esterified fatty acids.
• They are the primary form of fat storage.
• High blood levels (hypertriglyceridemia) are linked to increased cardiovascular disease and pancreatitis risk.
• Hypertriglyceridemia and hypotriglyceridemia refer to abnormally high and low levels of triglycerides in the blood.
• Triglycerides are a type of fat (lipid) in the blood that the body uses for energy.
• Influenced by diet, genetics, and conditions like diabetes.
• Measurement is part of a standard lipid panel.
• High levels can increase heart disease risk.
• Triglyceride tests measure the amount of triglycerides in a blood sample.

Methodologies for Measuring Triglycerides

• Enzymatic methods are the most common for measuring triglycerides.
• These methods involve a series of enzymatic reactions that break down triglycerides into glycerol and fatty acids.
• Glycerol is converted into a measurable product, such as hydrogen peroxide, which is detected spectrophotometrically.
• Common enzymatic methods include:
  • Glycerol-3-phosphate oxidase (GPO) method: This is the most widely used method.
  • Glycerol dehydrogenase (GPDH) method: This method is less susceptible to interference from free glycerol.
• Chemical methods, such as the Van Handel and Zilversmit method, are older and less specific.
• These methods involve saponification of triglycerides, followed by extraction and colorimetric determination of glycerol.
• Chemical methods are rarely used in clinical laboratories due to their complexity and interference.
• Point-of-care testing (POCT) devices are available for rapid triglyceride measurement.
• These devices typically use enzymatic methods with reflectance photometry for detection.
• POCT devices are useful for immediate results but may have lower accuracy and precision compared to laboratory methods.

Steps in Enzymatic Triglyceride Measurement

• Lipase hydrolyzes triglycerides into glycerol and free fatty acids.
• Glycerol kinase phosphorylates glycerol to glycerol-3-phosphate (G-3-P).
• Glycerol-3-phosphate oxidase oxidizes G-3-P to dihydroxyacetone phosphate and hydrogen peroxide (H2O2).
• Peroxidase catalyzes the reaction of H2O2 with a chromogen to produce a colored product.
• The absorbance of the colored product is measured spectrophotometrically.

Factors Affecting Triglyceride Measurements

• Lipemia (high levels of lipids in the blood) can interfere with triglyceride measurements.
• Lipemic samples may scatter light, leading to falsely elevated results.
• Fasting is required for accurate triglyceride measurement.
• Non-fasting samples can have significantly higher triglyceride levels.
• Certain medications, such as corticosteroids, beta-blockers, and diuretics, can increase triglyceride levels.
• Improper storage of blood samples can affect triglyceride levels.
• Samples should be stored at 2-8°C and analyzed within 7 days or frozen for longer storage.

Reference Range for Triglycerides

• The reference range can vary slightly depending on the laboratory and the method used.
• Generally accepted reference ranges for fasting adults include:
  • Normal: Less than 150 mg/dL (1.7 mmol/L).
  • Borderline High: 150-199 mg/dL (1.7-2.2 mmol/L).
  • High: 200-499 mg/dL (2.3-5.6 mmol/L).
  • Very High: 500 mg/dL or above (5.6 mmol/L or above).
• Desirable levels are below 150 mg/dL.
• These ranges are based on fasting samples.

Clinical Significance of Triglyceride Levels

• High triglyceride levels are associated with an increased risk of cardiovascular disease (CVD).
• Elevated triglycerides can contribute to the development of atherosclerosis, the buildup of plaque in arteries.
• Very high triglyceride levels (above 500 mg/dL) can increase the risk of acute pancreatitis.
• High triglycerides are often seen in individuals with metabolic syndrome.
• Metabolic syndrome is a cluster of conditions that increase the risk of heart disease, stroke, and type 2 diabetes.
• Low triglyceride levels are less common but may be seen in individuals with malnutrition or certain genetic disorders.

Factors Influencing Triglyceride Levels

• Diet: High intake of saturated and trans fats, cholesterol, and simple carbohydrates can increase triglyceride levels.
• Physical activity: Lack of exercise can lead to higher triglyceride levels.
• Weight: Obesity is associated with increased triglyceride levels.
• Alcohol consumption: Excessive alcohol intake can raise triglyceride levels.
• Age: Triglyceride levels tend to increase with age.
• Genetics: Some individuals are genetically predisposed to having higher triglyceride levels.
• Medical conditions: Certain conditions, such as diabetes, kidney disease, and hypothyroidism, can affect triglyceride levels.

Managing High Triglyceride Levels

• Lifestyle modifications are the first-line treatment for high triglycerides.
• These include:
  • Following a healthy diet low in saturated and trans fats, cholesterol, and simple carbohydrates.
  • Regular physical activity.
  • Weight loss, if overweight or obese.
  • Limiting alcohol consumption.
• Medications may be prescribed if lifestyle changes are not sufficient to lower triglyceride levels.
• Common medications include:
  • Fibrates: These drugs are effective at lowering triglyceride levels and increasing HDL cholesterol.
  • Niacin: Niacin can lower triglyceride levels and LDL cholesterol and raise HDL cholesterol.
  • Omega-3 fatty acids: High doses of omega-3 fatty acids can help lower triglyceride levels.
  • Statins: While primarily used to lower LDL cholesterol, statins can also have a modest effect on triglyceride levels.

Hypertriglyceridemia

• Hypertriglyceridemia is a condition characterized by elevated levels of triglycerides in the blood.
• Commonly defined as triglyceride levels above 150 mg/dL.
• Severe hypertriglyceridemia is defined as levels above 500 mg/dL.
• It is a risk factor for cardiovascular disease and acute pancreatitis.

Genetic Causes

• Familial Hypertriglyceridemia: A genetic disorder characterized by elevated triglycerides from birth.
• Familial Combined Hyperlipidemia: Characterized by high cholesterol and high triglycerides.
• Lipoprotein Lipase Deficiency: Rare genetic disorder leading to very high triglyceride levels due to a deficiency in the enzyme that breaks down triglycerides.
• Apolipoprotein CII Deficiency: Deficiency in a protein needed for lipoprotein lipase activation, causing hypertriglyceridemia.
• Other genetic mutations affecting lipid metabolism can contribute to hypertriglyceridemia.

Dietary Factors

• High intake of saturated and trans fats: These fats can increase triglyceride production in the liver.
• Excessive alcohol consumption: Alcohol can increase triglyceride synthesis and reduce their breakdown.
• High carbohydrate intake: Especially refined carbohydrates and sugars, which can be converted into triglycerides.
• Overeating: Consuming more calories than the body needs, leading to excess triglyceride production.
• Frequent snacking: Especially on processed and sugary foods, contributing to increased triglyceride levels.

Medical Conditions and Diseases

• Obesity: Associated with increased triglyceride production and reduced clearance.
• Type 2 Diabetes: Insulin resistance can lead to increased triglyceride levels.
• Metabolic Syndrome: A cluster of conditions including high blood pressure, high blood sugar, excess abdominal fat, and abnormal cholesterol levels, which often includes hypertriglyceridemia.
• Hypothyroidism: Underactive thyroid can decrease the breakdown of triglycerides.
• Kidney Disease: Impaired kidney function can affect lipid metabolism.
• Liver Disease: Conditions like non-alcoholic fatty liver disease (NAFLD) can increase triglyceride synthesis.
• Polycystic Ovary Syndrome (PCOS): Hormonal imbalances in PCOS can contribute to hypertriglyceridemia.
• Human Immunodeficiency Virus (HIV): HIV infection and certain antiretroviral medications can raise triglyceride levels.
• Autoimmune Diseases: Certain autoimmune conditions like lupus can be associated with hypertriglyceridemia.

Medications

• Thiazide Diuretics: Used to treat high blood pressure, can increase triglyceride levels.
• Beta-Blockers: Commonly prescribed for heart conditions, may elevate triglycerides.
• Oral Contraceptives: Some formulations can increase triglyceride levels.
• Glucocorticoids: Steroid medications that can raise triglyceride levels.
• Retinoids: Used to treat skin conditions, may affect lipid metabolism.
• Antiretroviral Medications: Some drugs used to treat HIV can increase triglyceride levels.
• Immunosuppressants: Medications used after organ transplants can sometimes cause hypertriglyceridemia.
• Estrogens: Hormone replacement therapy, can increase triglyceride levels in some individuals.

Lifestyle Factors

• Sedentary Lifestyle: Lack of physical activity can reduce triglyceride breakdown.
• Smoking: Can elevate triglyceride levels and negatively affect lipid metabolism.
• Stress: Chronic stress can affect hormone levels and contribute to hypertriglyceridemia.

Hypotriglyceridemia

• Hypotriglyceridemia is a condition characterized by abnormally low levels of triglycerides in the blood.
• Generally defined as triglyceride levels below 50 mg/dL.
• While less common than hypertriglyceridemia, it can be indicative of underlying health issues.

Genetic Causes

• Abetalipoproteinemia: A rare genetic disorder that prevents the body from properly absorbing dietary fats, leading to very low triglyceride levels.
• Hypobetalipoproteinemia: A genetic condition where the body cannot produce enough apolipoprotein B, which is essential for forming lipoproteins like VLDL and LDL, resulting in low triglyceride and cholesterol levels.
• Chylomicron Retention Disease: A rare genetic disorder where the body cannot properly transport chylomicrons (lipoproteins that carry dietary fats) from the intestines into the bloodstream.

Malabsorption Syndromes

• Celiac Disease: An autoimmune disorder triggered by gluten consumption, which can damage the small intestine and impair nutrient absorption, including fats.
• Cystic Fibrosis: A genetic disorder that affects the lungs and digestive system, leading to malabsorption of fats and fat-soluble vitamins.
• Short Bowel Syndrome: Occurs when a significant portion of the small intestine is removed or damaged, leading to reduced nutrient absorption, including triglycerides.
• Inflammatory Bowel Disease (IBD): Chronic inflammation of the digestive tract, as seen in Crohn's disease and ulcerative colitis, which can impair fat absorption.

Dietary Factors

• Very Low-Fat Diets: Diets that severely restrict fat intake can lead to lower triglyceride levels.
• Malnutrition: Insufficient intake of calories and essential nutrients can reduce triglyceride production.
• Starvation: Prolonged periods without food can lead to the depletion of fat stores and low triglyceride levels.

Medical Conditions and Diseases

• Hyperthyroidism: Overactive thyroid can increase the breakdown of triglycerides.
• Cachexia: A wasting syndrome associated with chronic diseases like cancer, leading to severe weight loss and low triglyceride levels.
• Acquired Immunodeficiency Syndrome (AIDS): Advanced stages of HIV/AIDS can cause malabsorption and reduced triglyceride synthesis.

Medications

• Fibrates: While typically used to lower high triglyceride levels, excessive use can sometimes lead to hypotriglyceridemia.
• Statins: Medications used to lower cholesterol can also reduce triglyceride levels, though usually not to the point of hypotriglyceridemia.
• Fish Oil Supplements: High doses of omega-3 fatty acids can lower triglyceride levels.
• Orlistat: A weight-loss medication that reduces the absorption of dietary fats, potentially leading to lower triglyceride levels.

Other Factors

• Severe Infections: Systemic infections can lead to metabolic changes that reduce triglyceride levels.
• Post-Surgery: Major surgical procedures can temporarily lower triglyceride levels due to metabolic stress and reduced food intake.

Phospholipids

• Similar to triglycerides, but one fatty acid is replaced by a phosphate group linked to another molecule (e.g., choline, serine, or ethanolamine).
• Major cell membrane components, forming a lipid bilayer.
• Amphipathic properties come from a polar (hydrophilic) head and nonpolar (hydrophobic) tails.
• Examples include phosphatidylcholine (lecithin), phosphatidylserine, and sphingomyelin.
• Also involved in cell signaling and lipid transport.

Cholesterol Metabolism Overview

• Cholesterol methodology and reference ranges are crucial in clinical diagnostics for assessing cardiovascular risk.
• Different methods yield varying results, and understanding principles and limitations is essential for accurate interpretation.
• Reference ranges provide a context for interpreting cholesterol levels, but they can vary based on population characteristics and laboratory practices.
• Cholesterol is an essential structural component of mammalian cell membranes and a precursor for steroid hormones, bile acids, and vitamin D.
• Cholesterol metabolism involves absorption, synthesis, transport, and excretion.
• The liver plays a central role in regulating cholesterol levels.
• Cholesterol is transported in the blood via lipoproteins, including LDL (low-density lipoprotein) and HDL (high-density lipoprotein).
• Cholesterol is a sterol with a four-ring structure.
• Essential for cell membranes, providing support and regulating fluidity.
• Precursor for steroid hormones (e.g., cortisol, estrogen, testosterone) and bile acids.
• Obtained from dietary sources and liver synthesis.
• High levels, especially LDL-cholesterol, are strongly linked to atherosclerosis and cardiovascular disease.
• Cholesterol levels are a key therapeutic target, influenced by statins and other lipid-lowering drugs.

Cholesterol Absorption

• Dietary cholesterol is absorbed in the small intestine.
• Cholesterol absorption is enhanced by bile acids, which emulsify fats and cholesterol, facilitating their uptake.
• The cholesterol absorption inhibitor ezetimibe reduces cholesterol absorption in the intestine.
• Plant sterols (phytosterols) compete with cholesterol for absorption, reducing cholesterol uptake.
• After absorption, cholesterol is incorporated into chylomicrons for transport in the lymphatic system and then the bloodstream.

Enzymatic methods

• Enzymatic methods are widely used because of their high accuracy, specificity, and ease of automation.
• Most common methods involve a series of coupled enzymatic reactions.
• Cholesterol esterase hydrolyzes cholesterol esters into free cholesterol and fatty acids.
• Free cholesterol is oxidized by cholesterol oxidase, producing cholest-4-en-3-one and hydrogen peroxide (H2O2)
• Peroxidase detects hydrogen peroxide in a chromogenic reaction where a colorless substrate is oxidized to a colored product.
• The colored product is measured spectrophotometrically, and it's directly proportional to cholesterol concentration.
• Variations exist, including different chromogens and optimized reaction conditions.
• These assays are adaptable to high-throughput analyzers.

Cholesterol Synthesis

• Cholesterol synthesis occurs primarily in the liver, with smaller amounts in other tissues.
• The synthesis pathway is complex, involving multiple enzymes and steps.
• Acetyl-CoA is the primary building block for cholesterol synthesis.
• HMG-CoA reductase is the rate-limiting enzyme in cholesterol synthesis.
• Statins inhibit HMG-CoA reductase, leading to decreased cholesterol synthesis.
• Cholesterol synthesis is regulated by cellular cholesterol levels; high cholesterol inhibits synthesis, while low cholesterol stimulates it.
• SREBPs (sterol regulatory element-binding proteins) play a key role in regulating the transcription of genes involved in cholesterol synthesis.
• Insulin promotes cholesterol synthesis.
• Glucagon inhibits cholesterol synthesis.

Cholesterol Catabolism

• Cholesterol catabolism primarily occurs in the liver.
• Cholesterol is converted into bile acids, which are secreted into the bile and aid in fat digestion.
• Some bile acids are reabsorbed in the ileum and returned to the liver via enterohepatic circulation.
• Cholesterol can also be excreted directly in the bile.
• A small amount of cholesterol is converted into steroid hormones, which are then excreted in urine.
• The excretion of cholesterol in feces represents a major route of cholesterol removal from the body.
• Cholesterol cannot be broken down into smaller molecules for elimination.
• The steroid nucleus of cholesterol is eliminated from the body by conversion to bile acids and excretion.

Chemical methods

• Chemical methods for cholesterol measurement, such as the Abell-Kendall method, are older but still relevant as reference methods.
• The Abell-Kendall method involves saponification of cholesterol esters, extraction of cholesterol with organic solvents, and reaction with Liebermann-Burchard reagent.
• Liebermann-Burchard reagent contains sulfuric acid, acetic acid, and acetic anhydride, which reacts with cholesterol to produce a colored complex.
• The intensity of the color is measured spectrophotometrically, and it's proportional to the cholesterol concentration.
• Chemical methods are more labor-intensive and have lower throughput compared to enzymatic methods.
• They are less specific and prone to interferences from other compounds in the sample.
• They are used in reference laboratories for validating other methods.

Lipoproteins and Cholesterol Transport

• Lipoproteins are responsible for transporting cholesterol and other lipids in the blood.
• Lipoproteins transport lipids (fats and cholesterol) in the blood.
• They are spherical particles with a core of triglycerides and cholesterol esters, surrounded by a shell of phospholipids, free cholesterol, and apolipoproteins.
• Lipoproteins are classified based on their density, which is determined by the ratio of lipids to proteins.
• Major lipoprotein classes include chylomicrons, VLDL (very-low-density lipoprotein), LDL, and HDL.
• Major lipoprotein classes include chylomicrons, VLDL (very-low-density lipoprotein), LDL, and HDL.
• LDL transports cholesterol from the liver to peripheral tissues.
• High levels of LDL cholesterol are associated with an increased risk of atherosclerosis.
• HDL transports cholesterol from peripheral tissues back to the liver.
• High levels of HDL cholesterol are associated with a decreased risk of atherosclerosis.
• Lipoprotein lipase (LPL) hydrolyzes triglycerides in VLDL and chylomicrons, releasing fatty acids for tissue uptake.
• CETP (cholesteryl ester transfer protein) mediates the exchange of cholesterol esters and triglycerides between lipoproteins.
• ABCA1 (ATP-binding cassette transporter A1) is involved in the efflux of cholesterol from cells to HDL.
• Lipoproteins transport lipids (cholesterol, triglycerides, and phospholipids) in the bloodstream.
• They consist of a hydrophobic lipid core surrounded by phospholipids, cholesterol, and apolipoproteins.
• Apolipoproteins are proteins that bind to lipids and have structural and functional roles in lipoprotein metabolism.
• The major classes are chylomicrons, VLDL, LDL, and HDL.

Chylomicrons

• Transport dietary triglycerides from the intestines to other tissues.
• They are the largest and least dense lipoproteins.
• Largest and least dense of the lipoproteins.
• Transport dietary triglycerides from the intestines to the rest of the body.
• Synthesized in the endoplasmic reticulum of enterocytes.
• Contain apolipoprotein B-48, apolipoprotein A-I, apolipoprotein A-II, apolipoprotein C, and apolipoprotein E.
• High percentage of triglycerides (85%).
• After delivering triglycerides, chylomicron remnants are taken up by the liver.
• Major components include:
  • Apolipoproteins: ApoB-48, ApoA-I, ApoA-II, ApoC-II, ApoE
  • Lipids: Primarily triglycerides, some cholesterol, phospholipids.
• Chylomicrons are metabolized in the capillaries of adipose and muscle tissue.
• Triglycerides are removed by lipoprotein lipase (LPL), releasing fatty acids that are taken up by the tissues.
• After the triglycerides are removed, the chylomicron remnants are taken up by the liver.

Very-Low-Density Lipoproteins (VLDL)

• Transport triglycerides synthesized in the liver to peripheral tissues.
• VLDL is catabolized, releasing triglycerides and becoming LDL.
• Synthesized in the liver.
• Transport endogenous triglycerides from the liver to the rest of the body.
• Contain apolipoprotein B-100, apolipoprotein C, and apolipoprotein E.
• Primarily composed of triglycerides (50-70%).
• Major components include:
  • Apolipoproteins: ApoB-100, ApoC-I, ApoC-II, ApoC-III, ApoE
  • Lipids: Primarily triglycerides, cholesterol, and phospholipids.
• VLDL particles are metabolized in the capillaries of adipose and muscle tissue.
• Triglycerides are removed by LPL, releasing fatty acids that are taken up by the tissues.
• As triglycerides are removed, VLDL particles become smaller and denser, eventually becoming LDL.

Low-Density Lipoproteins (LDL)

• The primary carrier of cholesterol.
• Delivers cholesterol to cells.
• Formed from the metabolism of VLDL.
• Primary carrier of cholesterol in the blood.
• Contain apolipoprotein B-100.
• Rich in cholesterol (40-50% of its mass).
• Elevated levels are a major risk factor for cardiovascular disease because LDL can accumulate in artery walls, leading to plaque formation (atherosclerosis).
• Major components include:
  • Apolipoproteins: ApoB-100
  • Lipids: Primarily cholesterol, some triglycerides and phospholipids.
• LDL particles are taken up by cells via the LDL receptor.
• LDL receptor recognizes apolipoprotein B-100 on the surface of the LDL particle.
• After binding, the LDL particle is internalized by endocytosis.
• In the lysosome, the LDL particle is broken down, releasing cholesterol.
• High levels of LDL cholesterol in the blood can lead to the development of atherosclerosis.

High-Density Lipoproteins (HDL)

• Transports cholesterol from peripheral tissues back to the liver, a process known as reverse cholesterol transport.
• Often referred to as "good" cholesterol because higher levels are associated with a lower risk of cardiovascular disease.
• HDL helps remove cholesterol from plaques and has anti-inflammatory and antioxidant properties.
• Synthesized in the liver and intestine.
• Participate in reverse cholesterol transport, removing cholesterol from cells and transporting it back to the liver.
• Contain apolipoprotein A-I, apolipoprotein C, and apolipoprotein E.
• Relatively high in protein (about 50%).
• Major components include:
  • Apolipoproteins: ApoA-I, ApoA-II, ApoC-I, ApoE
  • Lipids: Primarily phospholipids, cholesterol, some triglycerides.
• HDL picks up cholesterol from cells and other lipoproteins.
• Cholesterol is esterified by the enzyme lecithin-cholesterol acyltransferase (LCAT), forming cholesteryl esters which are stored in the core of the HDL particle.
• HDL can transfer cholesterol to the liver directly or transfer it to other lipoproteins (VLDL and LDL) via cholesteryl ester transfer protein (CETP).
• High levels of HDL cholesterol are associated with a reduced risk of cardiovascular disease.

HDL-C Methodologies

• HDL-C methodologies involve different techniques to measure high-density lipoprotein cholesterol in blood.

Direct Methods

• Direct methods selectively quantify HDL-C without prior precipitation.
• These assays employ specific antibodies that target apolipoprotein A-I (apoA-I) on HDL particles.
• Some direct methods use polymers or detergents, which selectively solubilize non-HDL lipoproteins, leaving HDL for quantification.
• Enzymatic cholesterol assays are commonly coupled with these separation or selective solubilization techniques to measure HDL-C concentration.

Precipitation Methods

• Precipitation involves separating HDL from other lipoproteins (VLDL, LDL, and chylomicrons) through precipitation.
• Polyanions (heparin, phosphotungstate) in the presence of divalent cations (magnesium, manganese) are used to precipitate non-HDL lipoproteins.
• After centrifugation, the supernatant containing HDL is analyzed for cholesterol using enzymatic methods.

Regulation of Cholesterol Levels

• Cholesterol levels are regulated by a combination of dietary intake, synthesis, absorption, and excretion.
• The liver plays a central role in maintaining cholesterol homeostasis.
• High dietary cholesterol intake can suppress cholesterol synthesis in the liver.
• Bile acid sequestrants increase bile acid excretion, leading to increased cholesterol synthesis in the liver.
• Fibrates increase the expression of lipoprotein lipase (LPL), leading to increased fatty acid oxidation and decreased VLDL levels.
• Niacin inhibits lipolysis in adipose tissue, leading to decreased VLDL and LDL levels and increased HDL levels.

Cholesterol and Disease

• Elevated LDL cholesterol levels are a major risk factor for atherosclerosis and cardiovascular disease.
• Atherosclerosis is characterized by the accumulation of cholesterol-rich plaques in the arteries.
• High levels of triglycerides can also increase the risk of cardiovascular disease.
• Genetic disorders, such as familial hypercholesterolemia, can cause severely elevated cholesterol levels.
• Statins are commonly used to lower LDL cholesterol levels and reduce the risk of cardiovascular disease.
• Lifestyle modifications, such as diet and exercise, can also help to lower cholesterol levels.

Clinical Significance of Lipid Measurement

• Lipid panels are commonly used to assess cardiovascular risk.
• A standard lipid panel includes measurements of total cholesterol, triglycerides, HDL-cholesterol, and LDL-cholesterol (calculated or directly measured).
• The ratio of total cholesterol to HDL-cholesterol is also used as a risk indicator.
• Abnormal lipid levels (dyslipidemia) are treated with lifestyle modifications (diet and exercise) and/or medications (e.g., statins, fibrates, niacin, and PCSK9 inhibitors).
• Monitoring lipid levels is important for managing and preventing cardiovascular disease.
• Lipid measurements are also useful in diagnosing and managing other conditions, such as pancreatitis and certain genetic disorders.

Point-of-care testing (POCT)

• POCT devices provide rapid cholesterol measurement at the point of care, such as clinics, pharmacies, and home settings.
• These devices use dry chemistry methods or electrochemical techniques.
• Dry chemistry methods use enzymatic reactions with reagents impregnated on a test strip or cartridge.
• The sample is applied to the strip, the reaction occurs, and a colored product is measured by reflectance spectrophotometry.
• Electrochemical methods detect hydrogen peroxide produced enzymatically using an electrode.
• POCT devices are convenient and provide rapid results but may have lower accuracy and precision compared to laboratory-based methods.
• They require regular quality control and maintenance to ensure reliable results.

Lipid Metabolism

• Lipid metabolism includes exogenous, endogenous, and reverse cholesterol transport pathways.

Exogenous Pathway

• The exogenous pathway involves the digestion, absorption, and transport of dietary lipids.
• Dietary triglycerides are emulsified by bile acids in the small intestine.
• Pancreatic lipase hydrolyzes triglycerides into monoglycerides and fatty acids.
• Monoglycerides, fatty acids, and cholesterol are absorbed by enterocytes.
• Inside enterocytes, triglycerides and cholesterol are re-esterified and packaged into chylomicrons.
• Chylomicrons are secreted into the lymphatic system, then enter the bloodstream.
• Lipoprotein lipase (LPL) on the endothelial surface of capillaries hydrolyzes triglycerides in chylomicrons.
• This releases fatty acids for uptake by tissues, such as muscle and adipose tissue.
• Chylomicron remnants, depleted of triglycerides, are taken up by the liver via receptor-mediated endocytosis.

Endogenous Pathway

• The endogenous pathway involves the synthesis and transport of lipids produced by the body, primarily in the liver.
• The liver synthesizes triglycerides and cholesterol.
• These lipids are packaged into very-low-density lipoproteins (VLDL).
• VLDL is secreted into the bloodstream.
• Lipoprotein lipase (LPL) hydrolyzes triglycerides in VLDL, releasing fatty acids for tissue uptake.
• As VLDL loses triglycerides, it is converted into intermediate-density lipoprotein (IDL).
• IDL can either be taken up by the liver or further metabolized into low-density lipoprotein (LDL).
• LDL delivers cholesterol to peripheral tissues via LDL receptors.
• Excess LDL can be taken up by macrophages in arterial walls, contributing to atherosclerosis.

Reverse Cholesterol Transport

• Reverse cholesterol transport (RCT) is the process by which excess cholesterol from peripheral tissues is transported back to the liver for excretion.
• High-density lipoprotein (HDL) plays a central role in RCT.
• HDL removes cholesterol from cells via the ABCA1 transporter.
• Lecithin-cholesterol acyltransferase (LCAT) esterifies cholesterol within HDL, forming cholesteryl esters.
• Cholesteryl esters are transferred from HDL to VLDL and LDL via cholesteryl ester transfer protein (CETP).
• HDL can also directly deliver cholesterol to the liver via the SR-B1 receptor.
• The liver excretes cholesterol in bile, either directly or after converting it to bile acids.
• RCT is considered anti-atherogenic because it reduces cholesterol accumulation in arterial walls.

Clinical Significance of Hypercholesterolemia and Hypocholesterolemia

Conditions Associated with Hypercholesterolemia:

• Familial hypercholesterolemia: A genetic disorder causing high LDL cholesterol levels.
• Diet high in saturated and trans fats: Increases LDL cholesterol.
• Obesity: Often associated with higher levels of LDL cholesterol and triglycerides.
• Diabetes mellitus: Can lead to increased LDL cholesterol and decreased HDL cholesterol.
• Hypothyroidism: Can cause elevated LDL cholesterol levels.
• Nephrotic syndrome: Associated with hyperlipidemia, including hypercholesterolemia.
• Cholestasis (Biliary obstruction): Can lead to increased cholesterol due to impaired bile excretion.
• Certain medications: Such as thiazide diuretics, beta-blockers, and anabolic steroids.
• Metabolic syndrome: A cluster of conditions including high blood pressure, high blood sugar, unhealthy cholesterol levels, and abdominal fat.
• Anorexia nervosa.

Reference range

• Optimal HDL-C levels are generally considered to be ≥ 60 mg/dL.
• An HDL-C level < 40 mg/dL for men and < 50 mg/dL for women is considered a major risk factor for coronary heart disease.
• These values are based on the Adult Treatment Panel (ATP) III guidelines, which emphasize the inverse relationship between HDL-C levels and cardiovascular risk.
• Reference ranges can vary slightly between laboratories due to differences in assay methods, patient populations, and calibration standards.
• Factors such as age, sex, genetics, and lifestyle can influence an individual's HDL-C level.
• HDL-C reference ranges should be interpreted in conjunction with other lipid measurements (total cholesterol, LDL-C, triglycerides) for comprehensive cardiovascular risk assessment.
• Reference ranges for cholesterol vary depending on factors such as age, sex, population, and laboratory methods.
• Guidelines recommend desirable, borderline high, and high cholesterol levels to assess cardiovascular risk.
  • Desirable total cholesterol: less than 200 mg/dL (5.17 mmol/L)
  • Borderline high total cholesterol: 200-239 mg/dL (5.17-6.18 mmol/L)
  • High total cholesterol: 240 mg/dL (6.21 mmol/L) or higher
  • Desirable LDL cholesterol: less than 100 mg/dL (2.59 mmol/L) for high-risk individuals, less than 130 mg/dL (3.36 mmol/L) for others
  • Desirable HDL cholesterol: 60 mg/dL (1.55 mmol/L) or higher
  • Borderline low HDL cholesterol: 40-60 mg/dL (1.03-1.55 mmol/L) for men, 50-60 mg/dL (1.29-1.55 mmol/L) for women
  • Low HDL cholesterol: less than 40 mg/dL (1.03 mmol/L) for men, less than 50 mg/dL (1.29 mmol/L) for women
• These values are general guidelines, and individual treatment goals may vary based on overall cardiovascular risk assessment and clinical judgment.
• Laboratories should establish their own reference ranges based on their specific methods and local population.

Conditions Associated with Hypocholesterolemia:

• Malnutrition: Reduced cholesterol synthesis due to lack of substrates.
• Malabsorption syndromes: Impaired absorption of fats and cholesterol in the intestine (e.g., celiac disease, Crohn's disease).
• Hyperthyroidism: Can lead to increased cholesterol metabolism and lower levels.
• Liver disease: Severe liver damage can impair cholesterol synthesis.
• Abetalipoproteinemia: A rare genetic disorder that prevents the formation of chylomicrons and VLDL, leading to very low cholesterol levels.
• Hypobetalipoproteinemia: A genetic disorder characterized by low levels of LDL cholesterol.
• Sepsis and severe infections: Cholesterol levels may decrease during acute illness.
• Certain medications: Such as statins (intended effect), ezetimibe, and bile acid sequestrants.
• Cancer: Some cancers may be associated with hypocholesterolemia.
• Smith-Lemli-Opitz syndrome (SLOS): A genetic disorder affecting cholesterol synthesis, leading to low cholesterol levels.

Factors affecting cholesterol measurement

• Several preanalytical and analytical factors can affect cholesterol measurement and influence result accuracy.
• Preanalytical factors include patient preparation, sample collection, handling, and storage.
• Patients should fast for 9-12 hours before blood collection to minimize the effects of dietary fat on cholesterol levels.
• Samples should be collected in tubes containing EDTA or serum separator tubes.
• Samples should be stored at 2-8°C and analyzed within 7 days.
• Analytical factors include method performance, calibration, quality control, and interferences.
• Methods should be calibrated regularly using appropriate calibrators.
• Quality control samples should be run daily to monitor method performance and ensure accuracy and precision.
• Interferences from substances such as bilirubin, hemoglobin, and triglycerides can affect cholesterol measurement.
• Lipemic samples with high triglyceride levels can cause falsely