Clinical Biochemistry

Course & Instructor Information

• Module: CEM3133 Clinical Biochemistry (Biomedical Science – Medical Biology)

• Programme: Foundation in Science

• Institution: MAHSA University, Centre for Pre-University Studies

• Lecturer: Dr. Vinibha Rajakumari

• Contact: vinibharajakumari@mahsa.edu.my

Learning Outcomes

• Explain the diagnostic importance of key enzymes (AST, ALT, LDH, CK).

• Explain the diagnostic significance of selected body-fluid analyses (CSF, Synovial, Amniotic).

• Apply and explain concepts of etiology, pathogenesis, morphology for common human diseases.

Clinical Biochemistry & Enzymology

• Also termed Chemical Pathology – analysis of body fluids for diagnosis/therapy.

• Clinical Enzymology: application of enzyme measurements for disease diagnosis & management.

• Utility of serum-enzyme profiling:
  • Myocardial infarction
  • Liver, muscle, bone & GI diseases
  • Cancers

• Unit of enzyme activity: International Unit (IU)
  • $1\,\text{IU} = 1\,\mu\text{mol of substrate converted}\;\text{min}^{-1}\,\text{L}^{-1}$ (optimal conditions, defined temperature).
  • Routine reporting: $\text{IU\,L}^{-1}$.

• Definition of Serum: plasma after coagulation – fibrinogen → fibrin retained in clot.

Key Diagnostic Enzymes (overview)

• Alanine Transaminase (ALT, SGPT)

• Aspartate Transaminase (AST, SGOT)

• Lactate Dehydrogenase (LDH)

• Creatine Kinase (CK/CPK)

Aspartate & Alanine Transaminases (AST / ALT)

• Reactions (require pyridoxal-5'-phosphate):
  • AST: $\text{Oxaloacetate} + \text{Glutamate} \rightleftharpoons \text{Aspartate} + \alpha\text{-Ketoglutarate}$
  • ALT: $\text{Pyruvate} + \text{Glutamate} \rightleftharpoons \text{Alanine} + \alpha\text{-Ketoglutarate}$

• Equilibria favour aspartate & alanine production respectively.

• Reference ranges (adult):
  • AST: 8 – 48 IU/L
  • ALT: 7 – 55 IU/L

Tissue Distribution & Clinical Significance

• Highest activities: liver, heart, skeletal muscle.

• Raised in cell injury of these tissues; mild rise post-alcohol intake.

• ALT is more liver-specific (rarely elevated by non-hepatic causes).

• AST less specific – rises in liver, heart, muscle disorders.

Differential Patterns

• Possible conditions when both ↑: extrahepatic cholestasis, cirrhosis, hepatocellular carcinoma.

• Acute hepatic injury → sharp ALT spike.

• Chronic/secondary organ damage → gradual AST rise.

AST / ALT Ratio Guidelines (normal ≈ 0.8)

Creatine Kinase (CK)

• Catalyses reversible phosphorylation:
  $\text{Creatine} + \text{ATP} \;\xrightleftharpoons[\;pH\,6.7\;]{\text{CK},\,Mg^{2+}}\; \text{Phosphocreatine} + \text{ADP}$

• At neutral pH phosphocreatine has higher phosphoryl-potential → reaction favours ATP regeneration during muscle contraction.

Clinical Significance

• Elevated total CK signifies tissue damage: skeletal muscle, myocardium, brain.

• Isoenzyme differentiation (CK-MB, CK-BB, CK-MM) aids diagnosis of:
  • Myocardial infarction
  • Muscular dystrophies / rhabdomyolysis
  • CNS trauma or stroke

Lactate Dehydrogenase (LDH)

• Catalyses: $\text{Pyruvate} + \text{NADH} + H^{+} \;\rightleftharpoons\; \text{Lactate} + \text{NAD}^{+}$

• Ubiquitous; high in muscle.

• Reaction direction depends on cellular redox state.

Clinical Significance

• Serum LDH released upon tissue damage; rises as cell lysis begins.

• Elevated in:
  • Anemia
  • Infections (mono, meningitis, encephalitis, HIV)
  • Sepsis
  • Infarctions: intestinal, myocardial, pulmonary
  • Acute kidney, liver, muscle injuries
  • Cancer – high LDH predicts poor prognosis & failed chemotherapy

• CSF LDH: high → bacterial meningitis; low/normal → viral meningitis.

Body-Fluid Compartments & Composition

• Total body water ≈ 70 % body weight.

• Extracellular fluid (ECF):
  • Interstitial fluid (~80 % of ECF)
  • Plasma (~20 %)
  • Also lymph, synovial, aqueous humour, CSF, pleural, peritoneal fluids.

• Organic solutes: glucose, amino acids, fatty acids, hormones, enzymes.

• Inorganic ions: Na⁺, K⁺, Ca²⁺, Mg²⁺, Cl⁻, $\text{PO}<em>4^{3-}$, $\text{SO}</em>4^{2-}$.

Laboratory Examination of Fluids

• Physical properties (colour, viscosity, volume).

• Biochemical analysis (analytes, enzymes, metabolites).

• Morphologic/cellular elements.

• Microbial culture.

• Ancillary studies (e.g., fetal anomaly screening).

Cerebrospinal Fluid (CSF)

Composition & Formation

• Third major body fluid.

• Adults: 125 – 150 mL; Neonates: 10 – 60 mL.

• Formed by choroid plexus & ependymal cells; fills sub-arachnoid space.

Functions

• Mechanical cushioning of brain/spinal cord.

• Circulates nutrients & chemicals filtered from blood.

• Removes metabolic waste & toxins.

Protein Analysis

• Normal total protein: 15 – 45 mg/dL (<< plasma).

• >80 % derives from plasma (but <1 % of plasma concentration).

• Elevated when:
  • Blood–brain barrier (BBB) damage
  • Intrathecal IgG synthesis
  • Neural tissue degeneration.

Detailed Protein Fractions (mg L⁻¹)

• Pre-albumin 17.3; Albumin 155; Transferrin 14.4; IgG 12.3; etc.

Causes of ↑ CSF Protein

• Traumatic tap (blood contamination).

• ↑ BBB permeability: arachnoiditis (e.g., post-methotrexate), meningitis (bacterial/viral/fungal/TB), subarachnoid or intracerebral hemorrhage.

• Local IgG synthesis: neurosyphilis, multiple sclerosis (MS), subacute sclerosing panencephalitis.

• Combined BBB + IgG: Guillain-Barré, collagen vascular disease (SLE, periarteritis).

• Drug toxicity: ethanol, phenothiazines, phenytoin.

• CSF circulation defects: obstruction (tumour, abscess, herniated disc), loculated effusions.

Albumin Indices

• Albumin ratio (BBB evaluation):
  $\text{CSF Albumin} : \text{Serum Albumin} = 1:230\; \text{(normal)}$

• Albumin index:
  $\text{Index} = \frac{\text{CSF Albumin (mg/dL)}}{\text{Serum Albumin (g/dL)}}$
  • <9 → intact barrier
  • 9 – 14 → slight impairment
  • 14 – 30 → moderate
  • >30 → severe
  • Traumatic tap invalidates calculation.

IgG Index

• Detects intrathecal IgG production (e.g., MS):
  $\text{IgG Index} = \frac{\text{CSF IgG (mg/dL)} \times \text{Serum Albumin (g/dL)}}{\text{Serum IgG (g/dL)} \times \text{CSF Albumin (mg/dL)}}$

• Normal upper limit 0.8; >0.8 indicates CNS IgG synthesis.

CSF Protein Electrophoresis Patterns

• Compare CSF vs serum IgG bands:
  • Type 1 Normal (polyclonal both).
  • Type 2 Intrathecal oligoclonal bands (MS).
  • Type 3 Intrathecal + systemic disease.
  • Type 4 Systemic inflammation (mirror oligoclonal).
  • Type 5 Monoclonal gammopathy (e.g., myeloma).

Glucose

• Fasting CSF glucose 50 – 80 mg/dL (≈ 60 % of plasma).

• CSF/plasma ratio 0.3 – 0.9.

• <40 mg/dL (hypoglycorrhachia) → bacterial/TB/fungal meningitis.

• Causes: ↑ anaerobic glycolysis by brain & leukocytes, impaired transport.

• Normalises earlier than protein/cell counts during recovery, useful for monitoring treatment.

Additional Chemical Markers

• Lactate ↑ → CNS hypoxia/anaerobic metabolism.

• F₂-isoprostanes ↑ → Alzheimer’s disease marker.

• Myelin basic protein ↑ → chronic demyelination.

• ADA ↑ → tuberculous meningitis.

• LDH ↑ → leukemia, lymphoma, bacterial meningitis, hemorrhage.

• CSF Glutamine reflects brain ammonia; ↑ in hypercapnia or sepsis.

Procedure Snapshot – Lumbar Puncture

1. Patient in fetal position.

2. Local anaesthetic to lower back.

3. Needle inserted between lumbar vertebrae.

4. Small CSF volume collected.

Synovial Fluid (SF)

• Viscous joint fluid secreted by synovial membrane; contains hyaluronic acid & low-level plasma proteins.

Functions

• Nutrient supply to cartilage.

• Lubrication for articulating surfaces.

Biochemical Tests

• Glucose (fasting): normal <0.55 mmol/L; large drop vs serum suggests infection/inflammation.

• Protein: normal ≈ ⅓ of plasma; ↑ indicates hemorrhagic or inflammatory joint disease (membrane permeability altered).

• LDH: elevated in rheumatoid arthritis & other inflammatory arthropathies.

• Uric acid: crystallisation confirms gout.

Amniotic Fluid (AF)

Composition & Formation

• Clear/slightly yellow liquid in amniotic sac.

• Origins: fetal-cell metabolism, maternal trans-placental water flow, fetal urine (later gestation).

• Volume grows with gestation (≈ 500 – 2500 mL).

Functions

• Cushions fetus from trauma.

• Medium for nutrient/waste exchange.

• Allows fetal movement → symmetrical musculoskeletal development.

Indications & Tests

• Sample volume ≤20 mL (amniocentesis).

• Indications: suspected chromosomal/metabolic disorders, neural-tube defects, hemolytic disease of newborn (HDN), infection, assessment of fetal maturity/gestational age.

Key Analytes

• Alpha-fetoprotein (AFP) – neural tube defects; measured in both AF and maternal serum.

• Cytogenetics – karyotyping for chromosomal anomalies (e.g., trisomy 21).
  • Indicated in mothers >35 yr, recurrent miscarriages, abnormal maternal AFP, parental carrier status.

• Other chemistry:
  • Bilirubin – degree of hemolysis (HDN).
  • Creatinine – fetal kidney maturity & gestational age.

General Pathology Concepts

• Pathology bridges basic science & clinical medicine; studies disease manifestations, causes, mechanisms.

Knowledge Checklist for Any Disease

• Definition, Epidemiology (where/when), Etiology (cause), Pathogenesis (evolution), Morphology (structural change), Management, Prognosis, Prevention.

Manifestations

• Signs – objective examiner findings.

• Symptoms – patient-perceived functional disturbances.

• Lesions – visible tissue/organ changes (benign, malignant, gross, occult, primary).

• Exacerbation – sudden worsening during disease course.

Etiological Factors

• Environmental: physical, chemical, nutritional, infectious, immunological, psychological.

• Genetic traits, Age.

• Multifactorial diseases: diabetes, hypertension, cancer.

Pathogenesis Definition

• Sequence of cellular/tissue events from initial etiologic stimulus → disease expression → outcome (recovery/death).

Key Terminology

• Acute (rapid onset), Fulminating (acute & fatal), Chronic (slow, long), Intercurrent (superimposed disease), Idiopathic (unknown cause), Teratogenic (drug-induced fetal harm), Contagious (direct contact), Venereal (sexual), Infectious, Communicable (via vectors/fomites).

Case Illustration – Chickenpox (Varicella-Zoster Virus, VZV)

Etiology & Transmission

• DNA herpesvirus; inhalation of infected respiratory droplets or contact with vesicular fluid.

Pathogenesis Timeline

• Incubation ~10-21 days; contagious 1-2 days pre-rash until crusting.

Morphology of VZV Particle

• Core: double-stranded DNA genome.

• Capsid: icosahedral protein shell.

• Tegument: enzymes for commandeering host metabolism.

• Envelope: outer lipid bilayer with glycoprotein spikes.

• Size: 150 – 200 nm diameter.

Ethical & Practical Implications

• Correct enzyme/body-fluid interpretation guides critical decisions (e.g., MI thrombolysis timing, antimicrobial choice for meningitis, prenatal counseling).

• Misinterpretation (e.g., traumatic tap artefacts) may lead to overtreatment or missed diagnoses.

• Prenatal testing requires genetic counseling to address ethical concerns of selective termination.

Quick Reference Values & Equations

• AST 8-48 IU/L, ALT 7-55 IU/L.

• AST/ALT ratio diagnostic thresholds (see table above).

• Albumin ratio 1:230, Albumin index <9 normal.

• IgG index ≤0.8 normal.

• CSF glucose 50-80 mg/dL; abnormal <40 mg/dL.

• Synovial glucose <0.55 mmol/L (fasting).

• IU definition: $1\,\mu\text{mol min}^{-1}\,\text{L}^{-1}$.

End-of-Lecture Integration

• Enzyme assays + body-fluid analysis form a diagnostic matrix: correlating biochemical markers with etiology, pathogenesis, morphology gives a holistic understanding, essential for evidence-based practice.