Bilirubin_metabolism_and_excretion_FA23_LEO.pptx

BILIRUBIN METABOLISM AND EXCRETION Vanessa De La Rosa, PhD CVI FA 2023 • Overview heme catabolism, including primary sources and site of its conversion to bilirubin, a potential toxin and diagnostic marker for abnormal bilirubin metabolism. Session Objectiv es • Describe the catabolism of heme t...

BILIRUBIN METABOLISM AND EXCRETION Vanessa De La Rosa, PhD CVI FA 2023 • Overview heme catabolism, including primary sources and site of its conversion to bilirubin, a potential toxin and diagnostic marker for abnormal bilirubin metabolism. Session Objectiv es • Describe the catabolism of heme to yield bilirubin by using NADPH and discuss how it is normally processed by the liver and excreted in the bile. • Discuss the role of UDP-glucuronyl-transferase in heme catabolism. • Describe the catabolism of conjugated bilirubin and how it is converted back into unconjugated bilirubin by intestinal bacteria. • Distinguish between direct and indirect measurements of bilirubin and its implications. • Outline the biochemical basis for problems associated with neonatal jaundice and infant kernicterus. • Explain the biochemical basis of genetically inherited disorders of hyperbilirubiemia, such as Gilbert & Crigler-Najjar syndromes (type I and II), and acquired conditions, and predict the effects of these disorders on bilirubin lab values. Overview • Heme is degraded to bilirubin • ~80% derived from senescent erythrocytes • Avg. lifespan of a mature erythrocyte: 90-120 days • senescent erythrocytes removed from circulation by reticuloendothelial system (occurs in liver & spleen) • remaining 20% results from turnover of immature RBCs & nonerythroid tissues and cells (various cytochromes) • Bilirubin is a toxic waste product • detoxified in liver & excreted into bile for conjugation & elimination • Many clinical conditions caused by abnormal bilirubin metabolism Heme degradation • Occurs in macrophages • Heme ring opening catalyzed by heme oxygenase (HO-1) and nonenzymatic oxidation • HO-1 Requires O2 or NADPH (reducing agents) • Only known reaction in human tissues and cells that produces carbon monoxide (CO) as a by-product of metabolism • Iron is released and biliverdin produced (greenish pigment) • Biliverdin converted to bilirubin (yelloworange pigment) • Catalyzed by biliverdin reductase • Bilirubin less polar than biliverdin Bilirubin metabolism • Liver is primary site of bilirubin metabolism and excretion • Transported to the liver by albumin • Binds noncovalently to albumin (reversible) • Binding disrupted by certain anionic drugs • Bilirubin rapidly removed from circulation by liver • Bilirubin kept soluble in liver through interactions with cytosolic proteins (ligandins) • Efficient excretion across bile canaliculus requires conversion to polar conjugates Conjugation of bilirubin • Glucuronidated to be more polar; facilitate excretion • Glucuronic acid is major conjugating group (requires UDP-glucuronic acid) • Source of negative charges • Glucuronidation catalyzed by uridine diphospho-glucuronosyltransferase (UDPGT) ** remember activated sugars? See PPP session** • UDPGT important enzyme in Phase II metabolism of drugs/xenobiotics • MRP2 (multidrug resistance protein 2) transports conjugated bilirubin across hepatocyte canalicular (apical) membrane Fate of conjugated bilirubin • Conjugated bilirubin in bile is secreted into the intestinal tract • Glucuronic acid is removed • Bilirubin is converted to urobilinogen (colorless) • Some urobilinogen reabsorbed into blood and converted to urobilin (makes urine yellow) • Urobilinogen is oxidized by intestinal bacteria into stercobilin (makes feces brown) Disorders of bilirubin metabolism • Abnormalities at any stage of hepatic bilirubin processing may result in hyperbilirubinemia, jaundice, kernicterus • Broadly divided into two classes (most present with jaundice [yellow skin discoloration]): • disorders of unconjugated bilirubin • disorders of conjugated bilirubin • Van den bergh assay standard clinical laboratory test that measures serum bilirubin Clinical biochemistry Van den bergh assay used to measure conjugated and total bilirubin A. yields conjugated bilirubin B. yields total bilirubin A. water soluble (conjugated) bilirubin reacts rapidly B. electrophilic attack of reagent diazonium salt (ethy anthranilate) on bilirubin duplicate assay performed in water & methanol both forms of bilirubin soluble in methanol= total bilirubin Indirect bilirubin (unconjugated) calculated: TOTAL – DIRECT= INDIRECT water insoluble (unconjugated) bilirubin NOT detected in short incubations Disorders of unconjugated (indirect) hyperbilirubinemia • neonatal jaundice affects ≈50% of newborns (majority of cases innocuous) • due to elevated unconjugated bilirubin • caused by delayed maturation of the liver • if untreated, can result in yellow discoloration of basal ganglia (kernicterus) • phototherapy (exposure to blue light [400-450nm]) most common treatment • changes bilirubin configuration  excreted in bile without conjugation • bilirubin important antioxidant defense for newborn for inhibiting lipid peroxidation Inherited disorders of unconjugated hyperbilirubine mia Decreasing activity Indirect (unconjugated) bilirubin Increasing severity • 3 syndromes related to UDPGT expression (UGT1A1 gene) • Results in decreased hepatic conjugation of bilirubin • Continuous spectrum of altered bilirubin conjugation • Gilbert Syndrome most common & clinically benign • UDPGT conjugates other drugs/xenobiotics • Decreased UGT1A1 activity predisposes these individuals to toxicity Inherited Disorders of Predominantly Conjugated Hyperbilirubinemia • 2 benign disorders of mostly direct hyperbilirubinemia (up to 20mg/dl) • Characterized by altered hepatobiliary transport or storage of bilirubin • Dubin-Johnson Syndrome (DJS) is inherited deficiency of MRP2 transporter (also known as ABCC2) • Rotor Syndrome (RS) caused by unknown molecular defect; nonspecific symptoms • Elevated direct bilirubin • Differentiated by urinary coproporphyrin excretion Summary Gilbert-Meulengracht Dubin-Johnson Canalicular multispecific UDP-glucuronosyltransferase organic anion transporter Genetic basis UGT1A1 ; chromosome 2q37 ABCC2 ; chromosome 10q23 Biliary transport deficiency Inappropriate conjugation activity Underlying mechanism of non-bile acid organic (uptake? transport?) anions Inheritance Autosomal recessive Autosomal recessive Conjugated = Hyperbilirubinemia Unconjugated unconjugated Aminotransferases Normal Normal Gross pathology: black Histology Normal liver, histology: lysosomal pigment Normal total urine Normal urine coproporphyrin Urine features coproporphyrin, 80% (75% coproporphyrin III) coproporphyrin I Prognosis Benign Benign Rotor Unknown Defective hepatic storage of conjugated bilirubin Autosomal recessive Conjugated > unconjugated Normal Normal 2–5-fold excretion of coproporphyrin, 65% coproporphyrin I Benign Other causes of jaundice Hemolytic jaundice • excessive erythrocyte destruction • bilirubin levels in liver exceed conjugating capacity & ability to excrete it into bile • results in increased circulating levels of unconjugated bilirubin Sickle cell anemia or deficiency of pyruvate kinase or glucose 6phosphate dehydrogenase Obstructive jaundice • partial or complete block of bile ducts • impaired excretion of conjugated bilirubin • results in increased circulating levels of predominantly conjugated bilirubin Hepatocellular jaundice • liver or gall bladder damage • impaired liver capacity to conjugate & excrete bilirubin • caused by toxins, poisons, cardiac failure, acute & chronic liver disease, gall bladder disease • results in increased circulating levels of predominantly Bilirubin produced fast than conjugated