Textbook - BIOLOGICAL CHEMISTRY - Hubskyi Yu.I. - 2000

Chapter VI. BIOCHEMISTRY OF PHYSIOLOGICAL FUNCTIONS AND SPECIALIZED TISSUES

CHAPTER 31. BIOCHEMICAL FUNCTIONS OF THE LIVER. DETOXIFICATION PROCESSES

31.3. BILE PIGMENT METABOLISM. BIOCHEMISTRY OF JAUNDICE

Mature erythrocytes undergo continuous destruction in The Human Body at a rate of (1-2) · 10⁸/h; their average lifespan is 100-120 days.

Bile pigments (bilirubin, biliverdin) are catabolic products of erythrocyte Hemoglobin and, partially, of other heme-containing Proteins. The destruction of erythrocytes and The breakdown of hemoglobin, which accounts for 90-95 % of the dry residue of erythrocytes, take place in the Cells of the reticuloendothelial system across various human Organs and Tissues—primarily the Spleen, Liver Kupffer cells, Bone Marrow, and Connective Tissue histiocytes.

Class="center">Hemoglobin Catabolism and bile pigment METABOLISM

Heme, a component of hemoglobin, constitutes about 80 % of the total body heme pool. In an adult weighing 70 kg, approximately 6 g of hemoglobin is turned over (broken down and resynthesized) daily. The degradation of Hemoproteins involves proteolysis of the protein moiety, release of iron ions—which are reutilized for the synthesis of iron-containing proteins—and irreversible catabolism of the heme porphyrin ring, yielding bilirubin (a yellow-red bile pigment) and biliverdin (a green pigment) that are subsequently excreted from the body.

The catabolism of hemoglobin and its porphyrin prosthetic group (heme) proceeds through the following stages.

1. Cleavage of the heme tetrapyrrole ring (within hemoglobin) via oxidative splitting of the methene bridge between rings I and II of the protoporphyrin cycle; As a result, the red erythrocyte pigment hemoglobin is converted into the green Blood pigment verdoglobin (choleglobin):

This reaction is catalyzed by the NADPH-dependent enzyme heme oxygenase, which is structurally an isoform of cytochrome P-450, and is accompanied by the release of carbon monoxide.

The conversion of hemoglobin to verdoglobin via heme oxidation causes a sequential color change in the areas of hematomas that form "bruises".

2. Breakdown of verdoglobin with the cleavage of the protein moiety, release of the iron ion, and Formation of the tetrapyrrole molecule biliverdin.

3. Conversion of biliverdin to bilirubin via the reduction of the methene bridge (between pyrroles III and IV). The reaction is catalyzed by the NADPH-dependent enzyme biliverdin reductase:

4. The aforementioned stages of bile pigment formation (1-3) occur within the cells of the reticuloendothelial system, from which bilirubin enters the blood and is adsorbed by serum albumin molecules. The serum albumin-bilirubin complex is transported to the liver, where the pigment is taken up by hepatocytes and undergoes further transformations.

5. Bilirubin is a lipid-soluble substance and, at high concentrations, exhibits membrane toxicity, especially toward Brain cells. Bilirubin detoxification—which involves converting the pigment into a Water-soluble (and less toxic) form, bilirubin glucuronide—takes place in the membranes of the hepatocyte Endoplasmic reticulum. The interaction of bilirubin with UDP-glucuronic acid (UDPGA) yields bilirubin mono- and diglucuronides:

The reaction is catalyzed by UDP-glucuronosyltransferase. The bulk of bilirubin is excreted into bile in the form of diglucuronides; when hepatocyte enzymatic Functions are impaired (parenchymal jaundice — see below), bilirubin monoglucuronides predominantly accumulate in the blood of patients.

In the blood serum of a healthy individual, the concentration of bilirubin is low, ranging from 0.1-1.0 mg % (1-10 mg/L, or 1.7-17 μmol/L). This bilirubin (serum "total bilirubin") consists of two fractions ("forms"):

(1) free bilirubin (accounting for about 75% of total bilirubin)—bilirubin that has not undergone conjugation with glucuronic acid (unconjugated bilirubin); this form circulates in the blood in a complex with serum albumin;

(2) bound bilirubin (accounting for up to 25% of total bilirubin)—bilirubin that has undergone conjugation with glucuronic acid (conjugated bilirubin); this form is secreted by normal hepatocytes into the bile, and enters the blood only partially and in insignificant amounts.

To determine the levels of total bilirubin and its fractions in human blood serum, the Van den Bergh method is used. It is based on The Use of diazosulfanilic acid (Ehrlich's diazo reagent), which reacts with bilirubin to form a pink diazo compound. Because free bilirubin is complexed with albumin, it yields a positive reaction with Ehrlich's diazo reagent only after protein precipitation with ethanol (the "indirect diazo reaction"), hence the designation indirect bilirubin. Bound (conjugated) bilirubin gives an immediate ("direct") reaction with the diazo reagent and is referred to in clinical and biochemical practice as direct bilirubin. Alterations in the quantitative ratios between the fractions of indirect and direct plasma bilirubin serve as an important differential diagnostic marker for various types of jaundice.

6. Bilirubin glucuronides ("conjugated bilirubin") are excreted by hepatocytes into bile and enter the intestine as part of bile, where they undergo biotransformation by microbial Enzymes, partial reabsorption, and excretion with feces (the "enterohepatic Circulation" of bile pigments).

The biotransformation of bilirubin glucuronides in the intestine involves the cleavage of glucuronic acid (mediated by microbial $eta$-glucuronidase) and the consecutive formation of tetrapyrrole compounds such as mesobilirubin and mesobilinogen (in the Small Intestine) and stercobilinogen (a product formed in the Large Intestine and excreted with feces):

A certain amount of mesobilinogen is already produced in The Liver and enters the Gallbladder along with bilirubin glucuronides. Urobilinogen and stercobilinogen are colorless compounds that, upon entering feces and urine, are oxidized into yellow pigments — urobilin and stercobilin.

The general metabolic pathway of hemoglobin degradation and bile pigment transformation is illustrated in the diagram (Fig. 31.6).

Fig. 31.6. Diagram of hemoglobin catabolism and bile pigment biotransformation.

Intestinal absorption of tetrapyrroles

The majority of bilirubin transformation products in the intestine—about 200-300 mg/day (approximately 95% of all tetrapyrrole compounds)—are eliminated from the human body via feces. However, a fraction of bile Pigments and Their biotransformation products is reabsorbed from the intestine into the blood and undergoes further transformations (Fig. 31.7):

Fig. 31.7. Intestinal absorption and subsequent biotransformation of bile pigment transformation products (T.T. Berezov, B.F. Korovkin, 1983; modified).

(a) stercobilinogen (the bulk of which is excreted in feces as stercobilin) is partially reabsorbed in the lower sections of the large intestine, from where it enters the systemic circulation via the Vessels of the pl. haemorroidalis, thus bypassing the liver. From the blood, this water-soluble stercobilinogen is excreted into urine as urobilin (0-4 mg/day); these trace concentrations of the pigment may not be detected in urine by standard clinical and Biochemical Methods, and therefore it is generally assumed that "urobilin" is typically absent in the urine of a healthy person;

(b) mesobilinogen (urobilinogen) is reabsorbed by the mucous membrane of the small intestine and transported via the v. porta Vascular System to the liver, where it is cleaved by hepatocyte enzymes into dipyrrole compounds that are ultimately excreted from the body via bile. When the barrier function of the liver is impaired (parenchymatous jaundice — see below), mesobilinogen is not cleaved in the liver, causing this pigment to enter the bloodstream and be excreted by the Kidneys, also referred to as urinary urobilin, which adds to the urobilin resulting from stercobilinogen absorption described in item (a).

Pathobiochemistry of Jaundice

When the total serum bilirubin concentration in humans exceeds 2-4 mg %, it manifests as a characteristic yellowing of the Skin and mucous membranes (especially the sclera) and is referred to as jaundice (icterus in Latin).

The causes of Hyperbilirubinemia and The Development of jaundice include excessive bilirubin production in the body, liver damage impairing its detoxification and excretory functions, or mechanical obstructions in the biliary tract that prevent the normal flow of bile into the intestine. Accordingly, several types of jaundice are distinguished.

Prehepatic (hemolytic) jaundice develops as a result of pathologically enhanced destruction (hemolysis) of erythrocytes, hemoglobin degradation, and excessive accumulation of bilirubin in the blood. Causes of this condition may include Rhesus incompatibility in newborns, incompatible blood transfusions, radiation injury, exposure to hemotoxic poisons, etc.

Prehepatic jaundice is characterized by an elevated concentration of total bilirubin in the blood, primarily due to the indirect fraction, i.e., free bilirubin that fails to be conjugated in the liver under conditions of its excessive production.

Under these conditions, the influx of large amounts of bilirubin into the intestine leads to enhanced formation of stercobilinogen (in some cases up to 10 g), which is excreted in increased quantities with feces (stercobilin) and (following reabsorption in the large intestine) in urine (urinary urobilin).

Hepatic (parenchymatous) jaundice develops as a result of impaired hepatocyte Structure and enzymatic properties caused by damaging factors of viral, bacterial, or chemical origin (viral, infectious, toxic hepatitis). This type of jaundice is characterized by marked hyperbilirubinemia (elevated total bilirubin concentration) due to the following factors:

a) impaired bilirubin conjugation resulting from damage to The endoplasmic reticulum membranes of hepatocytes and decreased activity of UDP-glucuronosyltransferase; this factor leads to an elevated blood level of indirect bilirubin;

b) impaired secretory function of hepatocytes, i.e., their ability to transport bilirubin glucuronide into bile (transport against a concentration gradient); this circumstance, along with necrosis of liver cells, leads to an excessive influx of direct bilirubin into blood serum.

An increase in direct bilirubin (bilirubin glucuronide) in the blood, which is capable of crossing renal membranes, is accompanied (in severe cases of the disease) by the appearance of bilirubin glucuronide in the urine (Laboratory tests for bile pigments in urine become positive). Due to the impaired ability of hepatocytes to cleave tetrapyrroles (absorbed as mesobilinogen), these compounds also enter the urine (yielding an elevated reaction for urinary "urobilin").

Posthepatic (obstructive) jaundice is caused by the inability of bile to enter the duodenum due to obstruction of the biliary tract (tumors, cholelithiasis). This type of jaundice is characterized by acholic feces due to the absence of stercobilinogen ("acholic stool") and the complete absence of urobilin in the urine. Due to impaired passage of bilirubin glucuronide into the bile (increased hydrostatic pressure in the bile ducts), the conjugated pigment is reabsorbed into the blood in increased amounts, leading to elevated direct bilirubin levels; under these conditions, bile pigments may appear in the urine, similar to parenchymatous jaundice.

Enzymatic (hereditary) jaundice arises from genetic enzymopathies caused by disruptions in the expression of genes responsible for the synthesis of bilirubin conjugation enzymes in hepatocytes (UDP-glucuronosyltransferase and/or UDPG dehydrogenase—the enzyme that forms UDPGA from UDP-glucose), its absorption from the blood, or its excretion into bile. Unconjugated bilirubin, which accumulates in blood serum in excessive amounts in these types of jaundice, can cross the blood-brain barrier into the brain and deposit in the Basal Ganglia and Brainstem nuclei, causing severe neurological disorders ("kernicterus").

Crigler-Najjar syndrome is a type of jaundice caused by a deficiency in the synthesis of UDP-glucuronosyltransferase ("conjugation jaundice").

Gilbert's syndrome is a pathological condition representing a heterogeneous group of disorders caused either by a block in the synthesis of UDP-glucuronosyltransferase or by impaired hepatocyte capacity to uptake bilirubin from the blood ("absorption jaundice").

Dubin-Johnson syndrome is a type of jaundice associated with impaired transport of bilirubin glucuronide from hepatocytes into the bile ("excretion jaundice").

Enzymatic jaundices can also occur in newborns as a temporary condition caused by delayed activation of genes encoding UDP-glucuronosyltransferase. A therapeutic effect can be achieved by administering Phenobarbital to infants, which acts as a universal inducer of hepatic detoxification enzymes—both for microsomal oxidation and substrate glucuronidation.

The main mechanisms underlying hyperbilirubinemia in various types of jaundice are illustrated in Fig. 31.8.

Fig. 31.8. Diagram of bilirubin (B) and bilirubin glucuronide (BGlu) metabolism and transport between blood, hepatocytes, and bile canaliculi, along with their impairments in various types of jaundice: (1) hemolytic jaundice — increased hemoglobin breakdown; (2) parenchymal and enzymatic jaundice — impaired conjugation of bilirubin with UDPGA; (3) obstructive jaundice — impaired normal bile outflow.