Lecture 18 Chemistry Proteins and Hepatic Markers PDF

Chemistry: Proteins PLASMA PROTEINS  Plasma Proteins Most plasma proteins are produced by the liver can provide insight into how well the liver if working Alterations of protein concentrations occur in a variety of disease conditions PLN, liver failure, DIC ...

Chemistry: Proteins PLASMA PROTEINS  Plasma Proteins Most plasma proteins are produced by the liver can provide insight into how well the liver if working Alterations of protein concentrations occur in a variety of disease conditions PLN, liver failure, DIC  Protein Functions Help form the structural matrix of all cells, organs, and tissues Maintain osmotic pressure Serve as enzymes for biochemical reactions Act as buffers in acid–base balance Serve as hormones Function in blood coagulation Defend the body against pathogenic microorganisms Serve as transport/carrier molecules for most constituents of plasma  Total Protein Total plasma protein includes fibrinogen* Total serum protein does not include fibrinogen* *TPP and TSP may differ slightly as a result TP concentrations can be affected by: Altered hepatic synthesis Altered protein distribution Altered protein breakdown or excretion Dehydration Overhydration  Total Protein Valuable in determining patient hydration status Dehydrated—hyperproteinemia (elevated) Overhydrated—hypoproteinemia (decreased) Screening patients with: Edema Ascites Diarrhea Weight loss Hepatic and renal disease Blood clotting problems  Total Protein Testing Two primary methods for testing Refractometer Estimates the refractive index of serum or plasma Often obtained as part of a PCV Fast, inexpensive, and accurate Biuret Measures the number of molecules containing more than three peptide bonds in serum or plasma Used in analytic instruments Simple and accurate Other Testing Methods Performed in Research/Reference Labs Separation & Measurement of Protein Populations Gel Electrophoresis Salt fractionation Chromatography Schematic diagram showing the results of the electrophoresis of whole serum. Four major peaks develop consistently: the albumin and three globulin peaks.  Albumin One of the most important proteins in plasma or serum Makes up 35% to 50% of the total plasma in most animals Any state of hypoproteinemia likely caused by albumin loss Hepatocytes synthesize albumin Any diffuse liver disease may result in decreases Renal disease, dietary intake, and intestinal protein absorption may also influence levels.  Albumin Function Major binding and transport protein Maintains osmotic pressure of plasma Aka. Oncotic pressure or Colloid- osmotic pressure  Globulins Alpha globulins— Gamma globulins synthesized in the liver and (immunoglobulins) primarily transport and bind Synthesized by proteins plasma cells High-density lipoproteins Responsible for (HDL) antibody production Very low-density (immunity) lipoproteins (LDL) Immunoglobulins in Beta globulins—include animals complement (C3, C4), IgM transferrin, and ferritin IgG Responsible for iron IgD transport, heme binding, IgE and fibrin formation and lysis IgA  Albumin/Globulin Ratio An alteration in the albumin/globulin ratio is often the first indication of a protein abnormality. Ratio is analyzed in conjunction with the protein profile. Most pathologic conditions alter the A/G. However, if the concentrations are reduced in equal amounts—no alteration in the ratio. Hemorrhage  Albumin/Globulin Ratio Calculated by dividing the albumin concentration by the globulin concentration In dogs, horses, sheep, and goats Albumin is greater than globulins. A/G is greater than 1.0 In cattle, pigs, and cats Albumin is equal to or lower than globulins. A/G is less than 1.0  Acute-phase Proteins Produced by hepatocytes immediately following injury or inflammation. Albumin and transferrin are negative acute-phase proteins because their plasma concentration decreases following injury or inflammation. 30 Recognized acute-phase proteins Serum amyloid A (SAA) Impt in cats, cattle, horses Rise w/in hours following injury/inflammation C-reactive protein (CRP) levels rise w/ variety of diseases (cardiac, sepsis, neoplasia) Levels increased w/in 6hrs of inflammatory event and beak @24-48hrs Fibrinogen Haptoglobin (HP) Ceruloplasmin α1-Acid glycoprotein (AGP) Major acute-phase protein (MAP) Acute-phase Proteins Different species produce different ones at different levels. Magnitude of increase in some acute-phase proteins indicates specific disease states in some species. Most of the acute-phase proteins of significance in domestic animals are measured with immunoassays. A chemical test is available for measurement of haptoglobin. A handheld portable analyzer is available for measurement of SAA in horses.  Fibrinogen Synthesized by hepatocytes Insoluble protein that forms the matrix of blood clots Converted to fibrin by thrombin Decreased fibrinogen levels Little to no clotting Make up 3% to 6% of total plasma proteins Not found in serum  Fibrinogen Acute inflammation or tissue damage may elevate levels. Calculated Value Some automated analyzers may provide fibrinogen results Not routinely included in small animal chemistries  Conditions Associated with Altered Serum Proteins Hyperproteinemia Hemodilution and/or (overhydration) hyperalbuminemia Hemoconcentration Blood loss (dehydration) Glomerulonephriti Inflammatory disease s Plasmacytoma Hepatic Lymphoma insufficiency Hypoproteinemia Malabsorption and/or hypoalbuminemia Malnutrition Chemistry: Hepatic Markers  Hepatic Markers Hepatobiliary clearance markers Bilirubin Bile Acids Ammonia Hepatic Enzymes Leakage enzymes: ALT, AST, Inducible enzymes Hepatic synthesis markers BUN, Cholesterol, Albumin (proteins), Glucose  The liver The liver is the largest internal organ. Complex structure, function, and pathologic characteristics Functions— metabolism of amino acids, carbohydrates, and lipids synthesis of albumin, cholesterol, plasma proteins, and clotting factors digestion and absorption of nutrients secretion of bilirubin or bile elimination or detoxification of toxins and catabolism of certain drugs.  Gallbladder Gallbladder—closely associated with liver Both anatomically and functionally Primary function is bile storage.  Malfunctions in the Liver or Gallbladder can lead to…. Hyperbilirubinemia (jaundice) Hypoalbuminemia Hemostasis problems Hypoglycemia Hyperlipoproteinemia Hepatoencephalopathy HEPATOBILIARY CLEARANCE MARKERS  Hepatobiliary Assays Liver cells compartmentalize so that damage in one zone will not affect all liver functions. Diverse function Over 100 tests available to detect problems. No single test is superior to any other. Tests being developed to detect problem sooner.  Hepatocyte Function Tests Test primarily run in veterinary settings. Bilirubin Bile acids Other hepatocyte function tests are less sensitive and test may not indicate a problem until ⅔ to ¾ of tissue is damaged. Albumin and cholesterol  Bilirubin Hyperbilirubinemia- increase of bilirubin in the blood Jaundice/Icterus- clinical manifestation of hyperbilirubinemia Insoluble molecule derived from the breakdown of hemoglobin by macrophages in the spleen. Binds to albumin for transport to the liver. Hepatic cells metabolize and conjugate the bilirubin to the molecule bilirubin glucuronide. This is secreted and is a major component of bile. Bacteria in the GI tract act on the bilirubin glucuronide and produce urobilinogens. Urobilinogens are broken down and excreted in feces or may be absorbed in the blood and excreted through the kidney.  Bilirubin Measurements of unconjugated and conjugated bilirubin can help pinpoint cause of jaundice. Difference in the relative solubility of these molecules help identify them. Unconjugated —bound to albumin Comprises approximately two-thirds of the total bilirubin in serum. Increases indicate problems with uptake or liver ability to conjugate bilirubin Conjugated—direct Increases indicate bile duct obstruction.  Bile Acids* Serve many functions Aid in fat absorption Modulate cholesterol levels Synthesized in the hepatic cells from cholesterol and are conjugated with glycine and taurine. Secreted across the canalicular membrane and reach the duodenum via the biliary system. Stored in the gallbladder until contraction associated with feeding. Except in horses, rats Once bile acids reach the ileum, transported back to the portal circulation and back to the liver.  Bile Acids 90% to 95% are actively resorbed in the ileum. Remainder excreted in feces Circulation of bile acids  Bile Acids Postprandial serum BA concentrations are higher than fasting concentrations. Any process that interferes with the circulation of bile acids results in elevated numbers. Elevated SBA levels usually indicate liver diseases such as congenital portosystemic shunts, chronic hepatitis, hepatic cirrhosis, cholestasis, or neoplasms can also result from extrahepatic diseases that secondarily affect the liver Decreased SBA Levels may be seen with intestinal malabsorptive diseases.  Bile Acids Testing Paired serum samples performed after 12 hours of fasting and 2 hours after eating Not specific for type of liver problem A screening test may also be used to follow the progress of liver disease during treatment  Ammonia (NH3)* Metabolism Majority of ammonia is generated in intestinal tract by bacterial flora Liberated ammonia is absorbed from GI into portal circulation → removed by hepatocytes & converted inti urea → urea is released into circulation and cleared by kidneys Mechanism of hyperammonemia Present w/ decreased functional hepatic mass Present w/ decrease in presentation of ammonia to liver (PSS) Urea toxicosis in ruminants (dietary urea too high) LIVER ENZYMOLOGY Liver Enzymology Leakage enzymes-enzymes released from damaged hepatocytes These enzymes are free within the cytoplasm and thus increase faster with direct hepatocyte damage Alanine transaminase (ALT) Aspartate transaminase (AST) Dehydrogenase enzymes Iditol dehydrogenase (ID) Glutamate dehydrogenase (GLDH) Inducible Enzymes Membrane bound within the cell and thus slower to increase within blood Alkaline Phosphatase (ALP/ALKP) Aspartate transaminase (AST) Gamma Glutamyltransferase (GGT)  Alanine Transaminase (ALT) Major source of ALT is the hepatocyte. Found free in the cytoplasm Elevations seen within 12 hours of liver damage Considered a liver specific test in dogs, cats, and primates. Horses, ruminants, pigs, and birds do not have enough ALT to be considered a liver specific test. ALT can be elevated by damage to renal cells, cardiac muscle, skeletal muscle, and the pancreas. Screening test, as it is not specific enough to identify specific liver disease.  Aspartate Transaminase (AST)* Present in hepatocytes—both in cytoplasm and bound to the mitochondrial membrane More severe liver damage required to release the membrane-bound AST. Levels rise slower than ALT but return to normal faster. Found in other tissues RBCs, cardiac muscles, skeletal muscles, kidneys, and pancreas Common cause for elevation Liver disease, muscle inflammation or necrosis, hemolysis  Iditol Dehydrogenase * (Sorbitol Dehydrogenase- SDH) Primary source is the hepatocyte. Also found in kidney, small intestine, skeletal muscles, and RBCs Especially useful in evaluating liver disease in large animals Large animals do not have diagnostic levels of ALT, so ID is the diagnostic test. Not a readily available veterinary test  Alkaline Phosphatase (ALP/ALKP) Inducible Present in hepatocytes, osteoblasts, chondroblasts, intestines, and placenta and liver cells Largest increase seen with cholestasis in small animals Gamma glutamyltransferase (GGT) Found in many tissues—primary source is the liver. Cattle, horses, sheep, goats, and birds have higher GGT than dogs and cats. Elevated levels in liver disease Especially with obstructive liver disease MARKERS OF HEPATIC SYNTHESIS  Hepatic Synthesis Liver is responsible for the synthesis of various components BUN Synthesized by hepatocatyes from ammonia → as ammonia ↑ BUN ↓ Glucose Cholesterol Albumin/Globulin Coagulation factors