Enzymes of Clinical Diagnostic Importance Lecture Note PDF

UNIVERSITY OF CAPE COAST Cape Coast, Ghana Cape Coast, Ghana Enzymes of clinical diagnostic importance  INTRODUCTION Injury or death of tissues can cause the release of tissue-specific enzymes into the bloodstream. Elevated enzyme levels are often in...

UNIVERSITY OF CAPE COAST Cape Coast, Ghana Cape Coast, Ghana Enzymes of clinical diagnostic importance  INTRODUCTION Injury or death of tissues can cause the release of tissue-specific enzymes into the bloodstream. Elevated enzyme levels are often indicators of tissue problems, and are used in the diagnosis of diseases. Enzyme activities in the body fluids are altered by pathological processes so, its measurement is used for disease investigation. www.ucc.edu.gh 2  Measurement of serum enzymes Enzymes are normally intracellular and LOW concentration in blood. Enzyme release (leakage)in the blood indicates cell damage (cell –death, hypoxia, intracellular toxicity) Quantitative measure of cell/tissue damage Organ specificity- but not absolute specificity in spite of same gene content. Most enzymes are present in most cells-differing amounts Time course of disease www.ucc.edu.gh 3  What to know … Enzymes – main features, properties Coenzymes – structures, functions Enzyme kinetics Enzyme activity 4 Isoenzymes – General Features Isozymes (also known as isoenzymes or more generally as multiple forms of enzymes) are enzymes that differ in amino acid sequence but catalyze the same chemical reaction. Genetically determined differences in primary structure Catalyze the same reaction May have different subcellular distribution 5 Isoenzymes – General Features May have different tissue distribution May be combined from more subunits (quarternary structure) May differ in kinetic properties (KM) Usually are determined by electrophoresis 6 proenzyme, isoenzyme, isoform Proenzyme (zymogen) – inactive form of enzyme that becomes active after partial proteolysis example: pepsinogen pepsin Isoenzyme – see previous page Isoform – more general term, includes true isoenzymes and pseudoisoenzymes (posttranslational variations) 7  Enzymes routinely measured NAME OF THE ENZYME PRESENT IN Aspartate Amino Heart and Liver transferase (AST) Serum glutamate- oxaloacetate transaminase (SGOT) Alanine Amino transferase Heart and Liver (ALT) Serum glutamate-pyruvate transaminase (SGPT)  Enzymes routinely NAME OF THE measured ENZYME PRESENT IN Alkaline Phosphatase (ALP) Bone, intestine and other tissues Acid Phosphatase (ACP) Prostate  glutamyl Transferase ( GT) Liver Creatine kinase (CK) Muscle Including cardiac muscle Lactate Dehydrogenase Heart, liver, muscle, (LDH) RBC  Amylase Pancreas Myocardial Infarction Enzyme Assays that are Carried out in Mayocardial Infarction  Commonly done: Creatine kinase (CK) Aspartate Amino transferase (AST) Lactate Dehydrogenase (LDH)  Myocardial Infarction ( MI ) Necrosis of the myocardium, but not angina pectoris release of CK, AST and LDH into the circulation. CK is the first to rise (activity within 6 h of MI ). Total CK reaches a peak at 24-36 h. In uncomplicated cases, CK returns to normal within 3 days. Necrosis is the death of body tissue. It occurs when too little blood flows to the tissue. Apoptosis is a type of cell death in which a series of molecular steps in a cell lead to its death. This is one method the body uses to get rid of unneeded or abnormal cells. www.ucc.edu.gh 13  Myocardial Infarction ( MI ) Serum AST  more slowly ( maximum activity within 48 h) and returns to normal in 4-5 days. No significant elevation in LDH seen for the 1st 24 h (reaches maximum at about 3 days & remain  for up to 8 days). The enzyme is relatively non specific to myocardial tissue.  Liver Diseases Hepatic Necrosis Hepatitis Cholestasis Jaundice Hepatocellular Damage  Liver Enzymes ( ALT, AST, GGT, ALP, LDH) Measurement of serum enzyme activities for : a - Differential Diagnosis of Jaundice. b - Monitoring of drug toxicity. ALT is more specific than AST. Hepatocellular disease has only modest effect on ALP & GGT (up to 3 times the upper limit of normal) In Cholestasis, Higher values of ALP & GGT due to  synthesis  Alanine aminotransferase (ALT) Widely distributed, although the largest amounts found in the liver. Smaller amounts occur in the heart but usually remains normal after MI. Congestive cardiac failure release from the liver More specific for liver disease than AST.  Aspartate aminotransferase (AST) This enzyme is widely distributed in the body. Main sources: Heart, liver, skeletal muscle, and kidney. Useful in the diagnosis of MI, liver disorders and muscle damage. Causes of serum AST levels:. Liver diseases: Hepatitis, hepatic necrosis , cholestasis Cardiac disease: Myocardial Infarction. Diseases of skeletal muscle: Crush injury, trauma, myopathy From Erythrocytes: Hemolysis  Alkaline phosphatase (ALP) Widely distributed, high concentrations in intestines, liver, bone, spleen, placenta and kidney. The main sources of serum ALP are the hepatobiliary tree and bone disorders. Elevated levels during healing of fractures , active growth and during the 3rd trimester of pregnancy.  serum ALP activity in liver disease is mainly due to Cholestasis. Decreased levels are found in the inherited  Alkaline phosphatase (ALP) Causes of increased serum alkaline phosphatase enzyme activity: Infancy - Physiological : Puberty - Pregnancy - Intestinal isoenzymes - Hyperparathyroidism - Bone disease: Osteomalacia, rickets - Paget’s disease of bone - Osteomyelitis - Hepatitis - Hepatobiliary disease: Cholestasis - Cirrhosis - Others: Carcinoma of the bronchus  Acid phosphatase (ACP) Found in prostate, bone, liver, spleen, kidney, RBCs and platelets Primarily used to diagnose prostate cancer .  In other prostatic conditions e.g. prostatitis, benign prostatic hypertrophy. In other non prostatic conditions e.g. hemolysis, Paget’s disease, metastatic carcinoma of the breast & Gaucher’s disease. Prostate- Specific Antigen(PSA): an enzyme occurs in prostatic tissue and  in cases of metastatic carcinoma  Amylase (AMS) HYDROLASES THAT SPLIT COMPLEX POLYSACCHARIDES. - CA+2 REQUIRING METALLOENZYME NORMAL LEVEL: 50-120 U/L INCREASES DRASTICALLY IN ACUTE PANCREATITIS ALSO IN MUMPS SOURCES : 1. PANCREAS (P-TYPE) 2. SALIVARY GLANDS (S-TYPE) 3. INTESTINAL MALIGNANCY CLINICAL SIGNIFICANCE : DIAGNOSIS AND MONITORING OF PANCREATITIS  Lipase (LPS) Breaks down fat into monoacylglycerol and free fatty acids. Primarily from the pancreas. Used to diagnose acute pancreatitis. Pancreatic lipases : Almost exclusively used clinically in the investigation of pancreatitis. - Increase within 2 - 12 hours of acute attack. - May remain elevated for many days. - More specific to acute pancreatitis than amylase. : Specificity of Enzymes Greater specificity is achieved in three ways: 1. Interpreting investigations in the light of clinical features 2. Isoenzyme determination: -  AST may be due to MI or Hepatitis so, it makes confusion in diagnosis to be confirmed by LDH levels. -  ALP in Cholestasis & bone diseases : - Differentiated by  bilirubin & transaminase levels in Cholestasis. - Confirmed by  GGT in Cholestasis.  Conditions in which level of NAME OF THE ENZYME activity in serum is elevated Aspartate Amino Myocardial infarction, Liver disease transferase (AST) especially with liver cell damage Serum glutamate- oxaloacetate transaminase (SGOT) Alanine Amino transferase Liver disease especially with liver (ALT) cell damage Serum glutamate-pyruvate transaminase (SGPT) Alkaline Phosphatase Liver disease- biliary obstruction (ALP) Osteoblastic bone disease-rickets Acid Phosphatase Prostatic carcinoma (ACP)  glutamyl Transferase Liver disorder like liver ( GT) cirrhosis Creatine kinase (CK) Myocardial infarction and skeletal muscle disease(muscular dystrophy Lactate Myocardial infarction, other Dehydrogenase (LDH) diseases like liver disease.some blood diseases  Amylase Acute pancreatitis  ISOENZYMES Catalyze the same reaction Two or more polypeptide chains Different polypeptide chains are products of different genes Differ in AA sequence and physical properties May be separable on the basis of charge Are tissue specific Diagram illustrating the origin of Isoenzymes Lactate dehydrogenase (LD) Tetramer and cytoplasmic Two different chains (H - heart, M - muscle) Five isoenzymes: LD1 (H4), LD2 (H3M), LD3 (H2M2), LD4 (HM3), LD5 (M4) Widely distributed in body Total activity determination – nonspecific finding 30  LACTATE DEHYDROGENASE (LDH) converts pyruvate to lactate (and vice versa) LDH occurs as a tetramer of 2 different subunits H & M: (product of 2 diff. gene) normal LDH2 is high in serum but after MI LDH1 rises increase of total is seen in hemolytic anemia, hepatocellular damage, carcinomas, leukemias & any condition of necrosis. ,five isoenzymes of LDH that occurs as a dimer of 2 different subunits H &M Isoenzyme Composition Composition Present in Elevated in name LDH1 ( H 4) HHHH Myocardium, myocardial RBC infarction LDH2 (H3M1) HHHM Myocardium, RBC LDH3 (H2M2) HHMM Kidney, Skeletal muscle LDH4 (H1M3) HMMM Kidney, Skeletal muscle LDH5 (M4) MMMM Skeletal Skeletal muscle muscle, Liver and liver diseases  Creatine kinase (CK) Dimer Two different chains (M – muscle, B – brain) Three isoenzymes: MM (muscle), MB (heart), BB (brain) Major isoenzyme in blood is MM (95 %) MB form in blood: 0 – 6 % BB in blood: traces (BB cannot pass across blood- brain barrier) 34  Creatine kinase (CK) Creatine kinase is associated with ATP regeneration in muscle and nervous tissue. Elevated blood levels of CK are used as indicators of MI, muscular dystrophy, and stroke. CK occurs as a dimer of 2 different subunits, M and B. - CK-BB: Brain. - CK-MB: cardiac muscle. - CK-MM: Muscle type. CK-MB is released from cardiac muscle cells after MI. Isoenzyme name Composition Present in Elevated in CK-1 BB Brain CNS diseases Acute CK-2 MB Myocardium myocardial / Heart infarction Skeletal CK-3 MM muscle, Myocardium  ENZYMES IN THERAPY Substitution of missing production of digestive enzymes – digestive enzymes – pepsin trypsin… Removal of deposits of death tissue or fibrin (e.g. in lungs, eyes), treatment of skin defects – proteinases, nucleases, collagenase Acceleration of fibrinolysis in lungs embolization (activation of plasmin and plasminogen) streptokinase, 38 www.ucc.edu.gh Metabolites and endogenous biomolecules 39 www.ucc.edu.gh  METABOLITE a metabolite is an intermediate or end product of metabolism. The term metabolite is usually used for small molecules. Metabolites are reaction intermediates and products of physiological metabolism which drive significant biological activities in human body such as signaling, stimulatory and inhibitory functions. 40 www.ucc.edu.gh  ary metabolites Diff. B/n Primary and secondary metabolites Primary Metabolites Secondary Metabolites Same in every species. Different in every species. Perform physiological Derivatives of functions in the body. primary metabolites. Eg., carbohydrates, Eg., Phenolics, steroids, vitamins, ethanol, lactic antibiotics, pigments. acid. 41 www.ucc.edu.gh  BIOMOLECULE Biomolecule, also called biological molecule, any of numerous substances that are produced by cells and living organisms. Biomolecules have a wide range of sizes and structures and perform a vast array of functions. The four major types of biomolecules are carbohydrates, lipids, nucleic acids, and proteins. 42 www.ucc.edu.gh  Endogenous biomolecules of clinical diagnostic importance Analyzing biomolecules is essential for disease diagnostics, food safety inspection, environmental monitoring and pharmaceutical development. 43 www.ucc.edu.gh  Creatinine (50-110 mmol/L) Most reliable biochemical test of glomerular function End product of nitrogen metabolism Changes can occur independently of renal function – Muscle mass changes – Immediately after surgery – Steroid treatment – Re-feeding  Creatinine Increased Decreased Renal disease Low muscle mass Indicates a fall in GFR Children Impaired renal perfusion Reduced muscle bulk Reduced blood pressure Starvation Fluid depletion Wasting disease Renal artery stenosis Steroid therapy Loss of functioning nephrons Glomerulonephritis Pressure increases in tubules Urinary tract obstruction Drugs Compete with creatinine Transient GFR – Glomerular filtration rate Reflects the number of functioning glomeruli Estimate of renal impairment Serum sample for U&E’s Calculation of glomerular filtration rate using the following formula: – 186 x (Creat / 88.4)-1.154 x (Age)-0.203 x (0.742 if female) x (1.210 if black) eGFR should not be used in identifying Acute Kidney Injury but for monitoring chronic kidney disease and function.  GFR – Glomerular filtration rate More reliable that creatinine clearance – Removes inaccuracies of urine collections Separate formula for children and those with renal failure  GFR stages Stage GFR* Description Treatment stage 1 90+ Normal kidney function Observation, control of Any urine findings or structural blood pressure. abnormalities or genetic trait may indicate kidney disease 2 60-89 Mildly reduced kidney function, Observation, control of and other findings (as for stage blood pressure and risk 1) may indicate kidney disease factors. 3A 45-59 Moderately reduced kidney Observation, control of 3B 30-44 function blood pressure and risk factors. 4 15-29 Severely reduced kidney function Planning for end stage renal failure. 5