Bilirubin Metabolism Introduction

[오세효]

INTRODUCTION

Bilirubin is the potentially toxic catabolic product of heme metabolism. Fortunately, there are elaborate physiologic mechanisms for its detoxification and disposition. Understanding these mechanisms is necessary for interpretation of the clinical significance of high serum bilirubin concentrations. Furthermore, because bilirubin shares its metabolic pathway with various other sparingly water soluble substances that are excreted in bile, understanding bilirubin metabolism also provides insight into the mechanisms of transport, detoxification, and elimination of many other organic anions.

An overview of the major aspects of bilirubin formation and disposition will be reviewed here. The settings in which bilirubin disposition is impaired will also be discussed briefly. Clinical aspects of serum bilirubin determination, the eval‎uation of patients with hyperbilirubinemia, and the classification of causes of jaundice are presented separately. (See "Clinical aspects of serum bilirubin determination" and "Diagnostic approach to the patient with jaundice or asymptomatic hyperbilirubinemia" and "Classification and causes of jaundice or asymptomatic hyperbilirubinemia".)

FORMATION OF BILIRUBIN

Bilirubin is formed by breakdown of heme present in hemoglobin, myoglobin, cytochromes, catalase, peroxidase and tryptophan pyrrolase. Eighty percent of the daily bilirubin production (250 to 400 mg in adults) is derived from hemoglobin [1]; the remaining 20 percent being contributed by other hemoproteins and a rapidly turning-over small pool of free heme. Enhanced bilirubin formation is found in all conditions associated with increased red cell turnover such as intramedullary or intravascular hemolysis (eg, hemolytic, dyserythropoietic, and megaloblastic anemias).

Heme consists of a ring of four pyrroles joined by carbon bridges and a central iron atom (ferroprotoporphyrin IX). Bilirubin is generated by sequential catalytic degradation of heme mediated by two groups of enzymes:

• Heme oxygenase
• Biliverdin reductase

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Chinese Medical Information Portal Website (Taiwan Medical Network-TMN) - OldDoc Wu's Series
(Established in March 1996)

Bilirubin

Dr. Wu's Liver diseases
for professionals (medical students and residents) and consumers

by Jau-Shin WU, M.D., Ph.D.

(Posted Sep. 16, 1996; Updated May 14, 2009)

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General consideration:

• Bilirubin--bile pigment--accumulate in body tissues causing greenish-yellow discoloration called jaundice (icterus).
• Usually, jaundice is linked to liver diseases, and jaundice is thought to indicate liver diseases.
• Actually, not all jaundice is induced by or linked to liver diseases and liver diseases are not always accompanied with jaundice.
• Bilirubin, a physiological product of RBC, is metabolized in the liver and excreted into bile ducts, therefore an appearance of jaundice means that there is a breakdown of balance of bilirubin metabolism and the patient may have a problem of liver or RBC production and destruction.
• Except for liver diseases, hemolytic disease, a blood disease, will also present with jaundice. Although, the incidence of this condition is rather rare, you must always keep it in mind when you are managing a case with jaundice.
• You need an experienced eye to diagnose jaundice with naked eye. Experienced physician can diagnose jaudice with minimum of 2.0 mg/dl serum bilirubin level. For people, non-professional, to notice jaudice, the serum bilirubin level needs to be at least 5~6 mg/dl or higher.
• Therefore, the detection of serum bilirubin level is one of the basic test for the detection of the presence and extent of liver diseases.
• Urobilirubinogen:
  After the excretion to the alimentary tract, the bilirubin is reduced to colorless urobilinogen by intestinal pathogens, followed by oxidation to colored urobilin (stercobilin) which is then excreted out of the body through the alimentary tract. A part of urobilinogen is reabsorbed from the alimentary tract and a part of the reabsorbed urobilirubinogen is excreted out of the body by the way of the kidney, whille most of the reabsorbed urobilinogen return to the liver and re-excreted into bile. Therefore, the detection of urinary bilirubin and urobilinogen is also of an important significance in the diagnosis of liver diseases.

Metabolism of bilirubin:

• Bilirubin is an end product of heme metabolism, coming mainly, 70 ~ 80 %, from hemoglobin of senescent red blood cells and the others from shunt pathway, myoglobin and respiratory enzymes with the structure of heme.
• The daily amount of bilirubin originated from destruction of red blood cells is around 250 ~ 300 mg.
• Bilirubin split to heme and globin at first, then further split to iron and biliverdin, and the biliverdin converts to bilirubin finally.
• The bilirubin combines tightly with albumin in the blood stream, and is separated just before being uptaken into liver cells.
• The bilirubin uptaken into liver cells combine with ligandin in the hepatocyte (cytosolic protein ligandin, Y-protein) to be transferred to smooth endoplasmic reticulum.
• The bilirubin in the hepatocytes conjugates with glucuronic acid to become conjugated bilirubin, which is excreted from heptocytes to the biliary tract and intestines and finally excreted out of the body.
• The original bilirubin from hemoglobin is free unconjugated bilirubin in the blood stream and is not soluble in water. After being taken into hepatocytes, it is converted to soluble conjugated form and is able to be excreted into bile ducts.
• The bilirubin is divided into two types, direct reacting bilirubin and indirect reacting bilirubin, according to its mode of reaction during the test process. Although, it is not completely the same, it can be recognized as that the direct reacting bilirubin is the conjugated bilirubin and the indirect reacting bilirubin the unconjugated bilirubin.
• The conjugated bilirubin is not absorbed in the bile ducts and intestinal tract, and is absorbed in the distal portion of the ileum after hydrolyzed and converted to urobilinogen by the intestinal pathogens.
• The urobilinogen is then oxidized to orange-color urobilin (stercobilin) and excreted in the stool out of the body. The yellowish color of stool is the color of the sterocobilin. Therefore, the clay color stool means that there is no bilirubin coming from biliary ducts owing to the obstruction of the bile ducts.
• About 15 ~ 20 % of the urobilinogen is reabsorbed from the intestine into portal veins and finally 90 % of them return to the liver and is re-excreted in the bile, it is called entero-hepatic circulation of bilirubin. The remainding 10 % gets into the systemic circulation and finally exreted in the urine through kidney. Thus the urine urobilinogen increases in the case of hemolytic disease, hepatocellular disease and porto-systemic shunt.

Pathogenesis of hyperbilirubinemia

• The balance of the bilirubin between production from the hemoglobin and excretion from the bile duct is kept well in normal condition.
• Hyperbilirubinemia -- jaundice occurs when the bilirubin balance between production and excretion breaks.
• From mechanism of the bilirubin metabolism we can learn the possible causes of hyperbilirubinemia:
  1. over production of bilirubin ----- hemolysis
  2. the impairment in bilirubin uptake by the liver, conjugation in the liver, and excretion from the liver cells
  3. the unconjugated and conjugated bilirubin is obstructed or leak back into the blood stream in the liver cells or from bile ducts.
• Therefore, the increase in serum unconjugated bilirubin is caused by over-production of bilirubin -- hemolysis or failure of uptake of unconjugated bilirubin by the liver or impairement in the conjugation process in the liver cells
  and the icrease in conjugated bilirubin is caused by impairment in the excretion process of the liver cells or failure of bilirubin excretion due to the obstruction of the bile ducts.

Method of determination

• Although there are many kinds of methods for the determiantion of bilirubin, most of the methods are according to the principle of Diazo reaction method reported by Hymans van den Bergh in 1913.
• The bilirubin concentration is determined by the detection of Azobilirubin produced when bilirubin reacts with diazotized sulfanic acid. By this method, the conjugated bilirubin reacts without the addition of hydrolysis promotor, alcohol, thus named direct bilirubin; and indirect bilirubin reacts only after addition of alcohol, so it is called indirect bilirubin.

Reference value

The reference value measured by autoanalyzer differs somewhat by the reaction time, temperature, and promotor of the diazotization.

• Total bilirubin: 0.3 ~ 1.2 mg/dl
• Direct bilirubin: 0.0 ~ 0.3 mg/dl
  Direct type bilirubin actually does not exist in the serum, however, a small portion of indirect reacting bilirubin may presents direct reaction, thus the result of direct bilirubin may show the maximum value of 0.3 mg/dl, but never above. It is about 20 ~ 30 % of the total bilirubin concentration.
  In some liver diseases, the total bilirubin will be within 1.2 mg/dl, but when the direct bilirubin concentration is above 0.3 mg/dl, the existence of liver disease should be considered.
• Starvation, pregnancy, pill intake, post-operation, hemolysis, alcohol intake, and steroid administation will show the increase in bilirubin level.
• Administration of sulfa drug, and phenobarbital will decrease the level of serum bilirubin concentration.

Change of the serum level in diseases

• We can determine two types of bilirubin, direct and indirect.
• Each kind of bilirubin represents the product of different stage of bilirubin meatbolism.
• The change of serum level in each kind of bilirubin represents the damage of bilirubin metabolism in some specific stage.
• Although different diseases may show the same pattern of change in two types of bilirubin, we still can differentiate the diseases by the different combination of serum concentration of these two bilirubin.

(A) Hyper-unconjugated-bilirubinemia

• (1) Overproduction:

  • Normal liver can handle the amount of seven times of normal daily bilirubin production.
  • When the production of bilirubin is increased due to hemolysis and to the amount over the ability of normal liver to handle, the serum indirect bilirubin will increase and is called prehepatic jaundice. The other liver tests, such as GOT, GPT, Alk-P, that will reflect the damage of hepatocytes will remain to be normal and only indirect bilirubin is increased.
  (a) Hemolysis:
  • When there is congenital or acquired shortage of life span of red blood cells, there will be increase in indirect bilirubin due to overproduction.
  • In chronic hepatitis or cirrhosis, there will be change in life span of red blood cells due to change in red blood cell membrane induced by the change in lipid metabolism.
  • Although, this degree of change will be able to be managed by the normall liver, but it will fail when the liver cells are damaged.
  (b) Increase in early bilirubin:
  • Ineffective erythropoiesis due to blood diseases or primary shunt hyperbilirubinemia will induce increase in indirect bilirubin.

• (2) Abnormality in uptake and conjugation:

  • Serum indirect bilirubin may increase in damage of uptake by the liver cells or conjugation in the liver cells of bilirubin due to the failure of change of indirect bilirubin to conjugate bilirubin. This condition is called chronic non-hemnolytic unconjugated hyperbilirubinemia.
  • Such as Crigler-Najjar syndrome (congenital non-hemolytic jaundice) is caused by the deficiency congenital glucuronyl transferase. The symptoms will appear in the infant stage, and there are two types, Type I is more severe than Type II, and may induce kernicterus.
  • Gilbert's syndrome (constitutional hepatic dysfunction) or idiopathic unconjugated hyperbilirubinemia are also considered to be caused by the similar mechanism as Crigler-Najjar syndrome, and only different in degree. The serum bilirubin level rarely exceed 3 mg/dl.

(B) Hyper-conjugated-bilirubinemia:

• The unconjugated-bilirubin conjuagte with glucuronic acid to become the conjugated-bilirubin.
• Usually, when there is obstruction of bile flow, the condition is called cholestasis, including stasis of bile acids, cholesterol, alkaline-phosphatase, and electrolytes in addition to bilirubin, however, sometimes not all these components are retained.
• When transportation of conjugated-bilirubin is impaired in the liver and/or during excretion process from liver cells or obstructed during passage from biliary capillary, bile ductules, intra-hepatic duct throughout to the orifice of duodenum, the condition is called as cholestatic jaundice.
• As in the case of Dubin-Johnson syndrome, only bilirubin is retained and no other components as cholesterol, bile acids and alkaline-phopatase are retained. Therefore, sometimes the Dubin-Johnson syndrone is classified as a condition of bilirubin retension, and the meaning of cholestasis in this context denotes bilirubin retension.

(1) Intrahepatic cholestasis:

• In liver diseases, the increase in serum bilirubin is mixed in type that means increase in both direct and indirect bilirubin.
• In liver diseases, the life span of red blood cells is usually shortened, thus the load of liver cells also increases owing to the increase in bilirubin production.
• In hepatocyte diseases, i.e. acute and chronic liver diseases including cirrhosis, the uptake, conjugation and excretion of blirubin in the hepatocytes are impaired and induce an intra-hepatic cholestasis.
• Therefore, the serum bilirubin elevation presents mixed type.
• The jaundice in the other condition such as drug-induced hepatitis and jaundice in pregnancy are also an intra-hepatic cholestasis.
  Usually, the elevation of serum bilirubin in the intra-hepatic cholestasis is higher than extra-hepatic cholestasis. Occasionally, it is higher than 30 mg/dl.
• Someone reported that the ratio of direct and indirect may be of use in differential diagnosis of intra- and extra-hepatic jaundice, but I don't think so from my experience.
• Many acute hepatitis cases may not present with jaundice, therefore they are often misdiagnosed as just a case of common cold.
• In icteric hepatitis cases, the jaundice usually appear 3 ~ 5 days after the onset of prodromal symptoms of general malaise and nauseation, and reach the peak in one to two weeks, and subside within two weeks to two months.
• In my 67 biopsy verified cases, the average serum bilirubin level is 11.7 mg/dl (1.4 ~ 26.4 mg/dl) for the total bilirubin and for the direct bilirubin component it is 6.4 mg/dl (0.2 ~ 15.6 mg/dl). Of them, 49 (73 %) cases are with direct bilirubin over 50 % of total bilirubin.
• The elevation of serum bilirubin is less in cases of chronic persitent hepatitis than chronic active hepatitis, and usually not to be notified by naked eyes. In chronic active hepatitis and cirrhosis cases the serum bilirubin level used to fluctuate.
• In chronic alcoholics, the serum bilirubin level is slightly elevated in 55 % of cases. In acute alcoholic hepatitis cases, the serum bilirubin level is usually over 5 mg/dl.
• Dubin-Johnson Syndrome (chronic idiopathic jaundice) and Rotor Syndrome (chronic familial jaudice) are considerd as congenital intraheptic cholestsis. The increase of serum bilirubin is mainly conjugated-bilirubin, and Rotor syndrome is considered as a variant of Dubin-Johnson syndrome. Morphologically, melanin pigments deposit in the liver cells are noted in Dubin-Johnson syndrome but not in Rotor Syndrome. They show different type of BSP retension clinically.
• Primary biliary cirrhosis shows obstruction of biliary ductules and inter-lobular bile ductules.
• Primary/secondary sclerosing cholangitis, primary/secondary hepatic cancer and other malignancies, and benign tumor and intrahepatic biliary stone will induce hyper-conjugated-bilirubinemia.

(2) Extrahepatic cholestasis:

• Stones, benign or malignant tomor in the biliary tract, and/or biliary obstruction due to external compression such as pancreatic head tumor or swelling due to pancreatitis will induce elevation in serum conjugated-bilirubin.
• Generally, the serum bilirubin elevation due to biliary tract stones used to fluctuate, and that due to malignancy used to show steady increase day by day.

Urine bilirubin

• No bilirubin is present in normal urine. When urine bilirubin becomes positive, it means something wrong in the biliary tract or the liver.
• Only conjugated-bilirubin will pass through renal glomeruli, and it is still unclear what factors will influence the renal clearance of bilirubin.
• Serum level of bilirubin does not parallel to the amount of urinary bilirubin. The urinary bilirubin becomes positive in prodromal stage of acute hepatitis when the serum bilirubin is still within normal level, and on the contrary the urine bilirubin can not be detected at the peak stage of serum conjugated-bilirubin.
• Therefore, you can detect acute hepatitis by urine bilirubin test before the jaudice appears.

Urobilinogen

• As stated in the previous paragraph, only a small part of urobilinogen absorbed from the intestinal tract is excreted out of the body through the kidney, and most of the urobilinogen return to the liver and are reexcreted to the intestinal tract.
• The amount of urinary urobilinogen is affected by the amount of conjugated-bilirubin in the biliary duct and also intestinal pathogens that convert bilirubin to urobilinogen.
• The amount of 24-hr urinary urobilinogen in normal person is 0 ~ 4 mg. In hemolytic diseases the urinary urobilinogen will increase because of production of a large amount of bilirubin.

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