Subsection: Heme synthesis
Case Scenario 1:
A 4-year-old boy is brought to the pediatric clinic with complaints of abdominal pain, dark-colored urine, and photosensitivity. His mother also reports behavioral changes such as irritability and episodes of confusion. Urine examination reveals elevated levels of δ-aminolevulinic acid (ALA) and porphobilinogen (PBG).
- Describe the normal pathway of heme synthesis.
- Name the disorder likely affecting this child and explain the enzymatic defect involved.
- Mention two clinical features and the biochemical basis of these manifestations.
Answer:
- Normal Pathway of Heme Synthesis:
Heme synthesis occurs partly in the mitochondria and partly in the cytosol, primarily in liver and bone marrow cells. The steps are:
- Condensation of glycine and succinyl-CoA (in mitochondria) → δ-ALA, catalyzed by ALA synthase (rate-limiting enzyme, requires pyridoxal phosphate).
- Two ALA molecules condense (in cytosol) → Porphobilinogen (PBG) via ALA dehydratase.
- Four PBG units polymerize to form Hydroxymethylbilane → Uroporphyrinogen III, then modified to Coproporphyrinogen III.
- Coproporphyrinogen III enters mitochondria → Protoporphyrin IX.
- Ferrochelatase adds Fe²⁺ to Protoporphyrin IX to form Heme.
- Diagnosis and Enzymatic Defect:
The likely diagnosis is Acute Intermittent Porphyria (AIP).
This is due to a deficiency of the enzyme Porphobilinogen deaminase (also called hydroxymethylbilane synthase), leading to accumulation of ALA and PBG.
Porphyrias are metabolic disorders caused by defects in enzymes of the heme biosynthetic pathway. They are classified into:
- Acute (Hepatic) Porphyrias – Present with neurovisceral symptoms.
Eg: Acute Intermittent Porphyria (AIP), Variegate Porphyria, Hereditary Coproporphyria.
- Cutaneous Porphyrias – Present with photosensitivity and skin lesions.
Eg: Porphyria Cutanea Tarda (PCT), Erythropoietic Protoporphyria.
- Clinical Features and Biochemical Basis:
- Abdominal pain and neuropsychiatric symptoms – Due to accumulation of neurotoxic intermediates (ALA and PBG) affecting the nervous system.
- Dark-colored urine – PBG and ALA oxidize on standing, giving urine a red or dark color (described as “port wine” urine).
Subsection: Hemoglobin Abnormalities
Case Scenario 2:
A 10-year-old boy from a tribal area presents with complaints of fatigue, joint pain, and recurrent episodes of limb swelling. His mother reports that he has had repeated episodes of fever and jaundice. On examination, he is pale, mildly icteric, and has a palpable spleen. A peripheral blood smear shows sickle-shaped red blood cells. Hemoglobin electrophoresis reveals increased levels of HbS.
a. Describe the genetic and molecular defect in sickle cell disease and its effect on hemoglobin structure.
b. Explain the pathophysiology of the major clinical features seen in this condition.
c. List two diagnostic tests and two complications associated with sickle cell disease.
Answer:
- Genetic and Molecular Defect:
Sickle cell disease is caused by a point mutation in the β-globin gene on chromosome 11.
The mutation results in substitution of valine for glutamic acid at the 6th position of the β-globin chain, forming abnormal hemoglobin S (HbS).
In deoxygenated states, HbS polymerizes, causing red blood cells to assume a sickle shape, making them rigid and less deformable.
- Pathophysiology of Clinical Features:
- Hemolytic anemia: Sickled RBCs have a shorter lifespan and are destroyed in the spleen → leads to anemia and jaundice.
- Vaso-occlusive crises: Sickled cells obstruct small blood vessels → causes painful episodes, swelling of limbs, and organ damage.
- Splenomegaly: Due to sequestration and destruction of sickled cells in the spleen, especially in children.
- Diagnostic Tests and Complications:
Diagnostic Tests:
- Peripheral blood smear – shows sickle-shaped RBCs.
- Hemoglobin electrophoresis – confirms presence of HbS.
Complications:
- Stroke or organ infarction – due to vaso-occlusion.
- Increased risk of infections – due to functional asplenia.
Case Scenario 3:
A 6-year-old boy is brought to the pediatric OPD with complaints of weakness, pallor, and poor growth. His parents report that he has required frequent blood transfusions since the age of one year. On examination, the child has frontal bossing, hepatosplenomegaly, and signs of anemia. A peripheral blood smear shows microcytic, hypochromic RBCs with target cells. Hemoglobin electrophoresis reveals elevated levels of HbF and HbA₂.
a. Classify thalassemias and describe the molecular defect in β-thalassemia major.
b. Explain the pathophysiology behind the clinical features seen in this child.
c. List two laboratory diagnostic features and two complications of thalassemia major.
Answer:
- Classification and Molecular Defect:
Thalassemias are inherited disorders of hemoglobin synthesis characterized by reduced or absent production of α or β globin chains.
- α-thalassemia – reduced or absent α-globin chain synthesis.
- β-thalassemia – reduced or absent β-globin chain synthesis.
In β-thalassemia major, mutations in the β-globin gene (usually point mutations) lead to absent (β⁰) or reduced (β⁺) synthesis of β chains. This causes an imbalance between α and β chains, with excess α chains precipitating in RBC precursors → ineffective erythropoiesis.
- Pathophysiology of Clinical Features:
- Anemia: Ineffective erythropoiesis and increased destruction of abnormal RBCs lead to severe anemia.
- Bone deformities (e.g., frontal bossing): Due to marrow hyperplasia as the body tries to compensate for anemia.
- Hepatosplenomegaly: Due to extramedullary hematopoiesis and RBC destruction in liver and spleen.
- Growth retardation: Chronic anemia and iron overload impair growth.
- Laboratory Features and Complications:
Diagnostic Features:
- Peripheral blood smear – shows microcytic, hypochromic anemia with target cells and nucleated RBCs.
- Hemoglobin electrophoresis – shows elevated HbF and HbA₂, reduced or absent HbA.
Complications:
- Iron overload (secondary hemochromatosis) – due to repeated transfusions.
- Skeletal deformities and growth failure – from marrow expansion and chronic anemia.
Subsection: Heme Degradation
Case Scenario 4:
A 9-year-old girl presents with complaints of yellowing of the eyes and dark-colored urine. Her mother reports that she often looks pale and tires easily. On examination, the child has pallor, icterus, and mild splenomegaly. Laboratory investigations reveal anemia, increased reticulocyte count, elevated unconjugated bilirubin, and increased urobilinogen in urine. Liver function tests are normal. Peripheral blood smear shows schistocytes.
a. Explain the normal pathway of hemoglobin degradation and bilirubin formation.
b. Describe how increased hemolysis leads to hemolytic jaundice, relating it to hemoglobin breakdown.
c. Mention two distinguishing lab findings of hemolytic jaundice and their biochemical basis.
Answer:
a. Normal Hemoglobin Degradation and Bilirubin Formation:
- Senescent RBCs are phagocytosed by macrophages in spleen and liver.
- Hemoglobin → heme + globin.
- Globin → amino acids.
- Heme → iron (reused) + protoporphyrin → biliverdin → unconjugated bilirubin.
- Unconjugated bilirubin (lipid-soluble) binds to albumin and is transported to the liver.
- In hepatocytes, it is conjugated with glucuronic acid by UDP-glucuronyl transferase, forming conjugated bilirubin (water-soluble).
- Conjugated bilirubin is secreted into bile, passes into the intestine, and is converted to urobilinogen and stercobilin, which give stool its brown color,
- Hemolytic Jaundice and Hemoglobin Breakdown:
- In hemolytic jaundice, excessive destruction of RBCs (hemolysis) leads to overproduction of unconjugated bilirubin.
- The liver’s capacity to conjugate bilirubin is exceeded, so unconjugated bilirubin accumulates in plasma.
- Since unconjugated bilirubin is not water-soluble, it is not excreted in urine.
- However, more bilirubin reaches the intestine, increasing urobilinogen formation, part of which is reabsorbed and excreted in urine, leading to increased urinary urobilinogen.
- Distinguishing Lab Findings of Hemolytic Jaundice:
- Elevated Unconjugated Bilirubin
- Due to excessive heme breakdown and limited conjugation capacity of the liver.
- Increased Urinary Urobilinogen
- More bilirubin enters the gut → increased conversion to urobilinogen → more reabsorption and urinary excretion.
Case Scenario 5:
A 15-year-old boy presents with complaints of yellowish discoloration of eyes, fatigue, and loss of appetite for one week. On examination, he is icteric and has a tender, enlarged liver. Laboratory findings show elevated total bilirubin with a mixed increase in conjugated and unconjugated bilirubin, raised liver enzymes (ALT, AST), and mild elevation of alkaline phosphatase. Urine shows the presence of bilirubin and decreased urobilinogen.
- Describe the normal pathway of hemoglobin degradation and bilirubin metabolism.
- Explain how liver dysfunction leads to hepatic jaundice in relation to bilirubin handling.
- Mention two key laboratory features of hepatic jaundice and their biochemical basis.
Answer:
- Normal Hemoglobin Degradation and Bilirubin Metabolism: Pathway
- Mechanism of Hepatic Jaundice:
In hepatic jaundice, damage to liver cells (e.g., viral hepatitis, hepatotoxicity) impairs:
- Uptake of unconjugated bilirubin.
- Conjugation due to defective enzyme activity.
- Excretion of conjugated bilirubin into bile canaliculi.
As a result:
- Both unconjugated and conjugated bilirubin accumulate in blood (mixed hyperbilirubinemia).
- Conjugated bilirubin, being water-soluble, appears in urine.
- Less bilirubin reaches the intestine → decreased urobilinogen formation.
- Laboratory Features of Hepatic Jaundice:
- Mixed hyperbilirubinemia (both unconjugated and conjugated):
- Due to impaired uptake, conjugation, and excretion of bilirubin.
- Presence of bilirubin in urine, decreased urobilinogen:
- Conjugated bilirubin leaks into blood and is filtered by kidneys (appears in urine).
- Reduced bilirubin excretion into gut leads to less urobilinogen formation.
Case Scenario 6:
A 50-year-old woman presents with yellowish discoloration of the skin and eyes for 10 days, along with pale stools and dark-colored urine. She also complains of itching and right upper abdominal discomfort. On examination, she is icteric and has a palpable gallbladder. Liver function tests show markedly elevated conjugated (direct) bilirubin, raised alkaline phosphatase, and mildly elevated transaminases. Ultrasound reveals gallstones obstructing the common bile duct.
- Describe the normal pathway of hemoglobin degradation leading to bilirubin formation and excretion.
b. Explain how bile duct obstruction leads to obstructive jaundice in relation to bilirubin metabolism.
c. Mention two distinguishing laboratory findings of obstructive jaundice and explain their biochemical basis.
Answer:
- Normal Hemoglobin Degradation and Bilirubin Excretion: Pathway
- Mechanism of Obstructive Jaundice:
- In obstructive jaundice, bile flow is blocked due to gallstones, tumors, or strictures.
- Conjugated bilirubin cannot be excreted into the intestine.
- It accumulates in the liver and refluxes into the bloodstream.
- Since it is water-soluble, conjugated bilirubin is excreted in urine, causing dark urine.
- Lack of bilirubin in the gut results in pale-colored stools and reduced urobilinogen formation.
- Laboratory Findings in Obstructive Jaundice:
- Elevated conjugated (direct) bilirubin:
- Due to backflow of bile into blood when excretion is blocked.
- Raised alkaline phosphatase (ALP):
- Due to cholestasis and bile duct injury.
Additional features:
- Bilirubin present in urine (dark urine)
- Absent or low urobilinogen in urine and pale stools
Case Scenario 7:
A 3-day-old full-term newborn is brought to the pediatric clinic with yellowish discoloration of the skin and eyes. The baby is feeding well and shows no signs of lethargy or irritability. On examination, the baby is alert, active, and afebrile, with mild jaundice visible up to the chest. Total serum bilirubin is elevated, predominantly in the unconjugated form. There is no evidence of hemolysis or infection.
Questions:
- Describe the normal pathway of hemoglobin degradation leading to bilirubin formation and excretion.
b. Explain the causes of physiological jaundice in newborns in relation to bilirubin metabolism.
c. Mention two differences between physiological and pathological jaundice in neonates.
Answer:
- Normal Hemoglobin Degradation and Bilirubin Metabolism: Pathway
- Causes of Physiological Jaundice in Newborns:
- Increased RBC turnover:
- Newborns have a higher red cell mass and shorter RBC lifespan.
- Breakdown of fetal hemoglobin (HbF) releases large amounts of bilirubin.
- Immature liver enzymes:
- The enzyme UDP-glucuronyl transferase is underdeveloped in neonates, leading to impaired conjugation of bilirubin.
- Increased enterohepatic circulation:
- Delayed passage of meconium increases reabsorption of unconjugated bilirubin from the intestine.
These factors lead to transient, mild unconjugated hyperbilirubinemia, peaking by day 3–5 and resolving by day 7–10 in full-term infants.
- Differences Between Physiological and Pathological Jaundice:
| Feature | Physiological Jaundice | Pathological Jaundice |
| Onset | After 24 hours of birth | Within 24 hours of birth |
| Type of bilirubin | Unconjugated | May be unconjugated or conjugated |
| Peak bilirubin level | < 15 mg/dL (usually <12 in term) | Often >15 mg/dL |
| Duration | Resolves within 7–10 days | Persists >14 days or worsens |