Clinical Enzymology Notes

Clinical Enzymology

Learning Goals

• Understand isoenzymes.
• Understand how isoenzymes (LDH, CPK, ALP) and other enzymes (HBDH, ALT, AST, GGT, 5’-nucleotidase) can be used diagnostically:
  • Determine organ damage.
  • Timeline of the patient after a myocardial infarct.
  • Indicating liver function.
  • Gastrointestinal diseases (e.g., pancreatitis).

Clinical Enzymology

• Measurement of enzyme or enzyme function in biological fluids to determine if a process is working correctly.
• Measure activity of enzymes in blood (most common) or urine to diagnose disease.
• Functional enzymes are always present.
• Non-functional enzymes have no known function in plasma (focus of the lecture).

Functional Enzymes

• Certain enzymes, zymogens, and their substrates are always found in the plasma in normal individuals.
• Perform a function in the plasma.
• Examples:
  • Blood clotting enzymes and enzymes involved in blood clot dissolution.
  • Lipoprotein lipase.
• The majority of these enzymes are synthesised and secreted by the liver into the plasma.

Non-Functional Enzymes

• Cell-derived enzymes.
• No known physiological function in plasma.
• Presence in plasma indicates:
  • Cell damage.
  • Cell turnover (normal levels?).
  • Secretion.
• Tissue damage or necrosis resulting from injury or disease is accompanied by increases in non-functional plasma enzymes.
• Decreased levels can indicate:
  • Enzyme inhibition.
  • Lack of cofactors.
  • Genetic or acquired deficiency.
• Variety of enzymes measured:
  • Myocardial infarction.
  • Liver function.
  • Cancer.
  • Gastrointestinal tract diseases.

Isoenzymes

• Isoenzyme: Different enzymes that catalyse the same reaction.
  • Slightly different amino acid sequence (isoform).
  • Kinetic parameters different (different K_M).
  • Can be regulated differently.
• Permits fine-tuning to meet the needs of tissue or developmental stage.
• Can be used diagnostically (i.e., clinically).

Measurement of LDH Isoenzymes

• Lactate dehydrogenase (LDH) - 5 isozymes with 4 subunits = combinations of Heavy (H) and Medium (M) chains.
• Different isozymes present in different tissues.
• Converts lactate to pyruvate (liver) OR pyruvate to lactate (low oxygen).
• Increased and persistent levels of LDH-1 and LDH-2 indicates myocardial damage.
• Proportions of isozymes give an idea of timing.
• LDH-1 > LDH-2 between 12 and 24 hours post-infarction.
• High LDH-5 indicates liver disease and liver failure.
• High LDH-4 indicates pancreatitis.

Release of Enzymes Following Myocardial Infarction

• Determines if it has happened and the timing.
• CPK = creatine phosphokinase (isozyme in heart that acts as energy reservoir).
• LDH = lactate dehydrogenase.
• HBDH = α-hydroxybutyrate dehydrogenase (mitochondria involved in fatty acid metabolism).
• Based on proportions, indicates timing.

Liver Function Tests - Liver Damage

• Enzymes present in blood plasma.
• Liver enzymes:
  • Alanine transaminase (ALT) (alanine to glutamate + pyruvate), also known as alanine aminotransferase.
  • Aspartate transaminase (AST) (aspartate to oxaloacetate + glutamate), also known as aspartate aminotransferase.
  • Low levels in blood normally.
• Increased levels of ALT and AST indicate liver damage or viral hepatitis.

Liver Function Tests - Cholestasis

• Liver enzymes:
  • Alkaline phosphatase (ALP).
  • 5’-nucleotidase.
  • γ-glutamyl transpeptidase (GGT).
  • Low levels in blood normally in adults.
  • ALP is elevated in children and adolescents (rapid bone growth) and in normal pregnancy.
• Increased levels of ALP, GGT, and 5’-nucleotidase indicate cholestasis (flow of bile from liver is reduced or blocked).

Measurement of ALP Isoenzymes

• Alkaline phosphatase
  • 4 isozymes (intestinal, placental, germ cell, liver/bone/kidney).
  • Different isozymes present in different tissues (hepatic, bone, placental, intestinal).
  • Catalyses the hydrolysis of phosphate monoesters at basic pH values.
• Can detect different isozymes (indicate different conditions).
• Increased levels can indicate:
  • Hepatic biliary obstruction.
  • Active growth or bone disease (bone isoenzyme elevated in children during active growth).
  • Last 6 weeks of pregnancy (placental isoenzymes).
  • Recent ingestion of fatty meal or various gastrointestinal disorder (intestinal isoenzyme).
• Regan isoenzyme present in plasma of ~15% of patients with lung, liver, or gastrointestinal carcinoma (similar to placental ALP).
• Nagao isoenzyme is a variant of Regan isoenzyme, found in metastatic pleural carcinoma and pancreatic and bile duct adenocarcinoma.

Enzymes in Gastrointestinal Tract Diseases

• Liver enzymes:
  • Amylase
  • Lipase
• Increased levels of amylase and lipase in the serum are seen in acute pancreatitis.
• Lipase is also elevated in perforated ulcers and intestinal obstruction.

Practice Exam Questions

• Q: What is an isoenzyme?
  • A. Different enzyme isoform that performs the same function with different kinetic and regulatory properties.
• Q: Which of the following enzymes are NOT used to measure liver function?
  • A. Lactate dehydrogenase and Creatinine phosphokinase
• Q: What does an increased level of ALT and AST indicate?
  • A. Liver damage

Practice Exam Question Part 2

• Q: A serum sample contains 1000 U/L amylase (normal level: 20-300 U/L) and 300 U/L lipase (normal level: 0-160 U/L). The presence of which of the following diseases are indicated by this result?
  • A. Acute Pancreatitis

Summary

• Enzyme activity can be used as a clinical measure of disease/organ function (e.g., timing of myocardial infarction, measure of liver function, and presence of gastrointestinal disease).
• Isoenzymes are different proteins that perform the same function but have different kinetic and regulatory properties.