1. Biochemistry of liver
Alice Skoumalová

2. The liver-introduction
importance in intermediary metabolism and in the detoxification and elimination of toxic substances
one of the largest organs in the human body
it constitutes only 2-3% of the body mass, it accounts for 20-30% of the total oxygen consumption

3. Diagram of a hepatocyte
There are approximately 300 billion cells in the liver (80% of them are hepatocytes)
The hepatocytes are central to the intermediary metabolism of the body

4. Functions of the liver
The uptake of nutrients delivered from the digestive tract via the portal vein
The synthesis, storage, interconversion, and degradation of metabolites (metabolism)
The regulated supply of energy-rich intermediates and building blocks for biosynthetic reactions
The detoxification of harmful compounds by biotransformation
The excretion of substances with the bile, as well as the synthesis and degradation of many blood plasma constituens
Fighting infections (Kuppfer cells-macrophages)
The liver plays a vital role in fighting infections by mobilising the macrophage systém. The liver contains over half of the body‘s supply of macrophages (Kuppfer cells).

5. Liver metabolism The primary function is the maintenance of sufficient plasma levels of the metabolites (homeostasis)
Carbohydrate metabolism
Lipid metabolism
Amino acid and protein metabolism
Biotransformations
The liver plays an important role in hormonal modification and inactivation. The steroid hormones are inactivated by the liver (hydroxylation and conjugation). Most plasma proteins (except antibodies) are synthesized and secreted by the liver. Hemostasis- most of the 12 clotting factors are plasma proteins produced by the liver. Storage- the liver stores glucose in the form of glycogen, and also fat-soluble vitamins, vitamins B6 and B12, and minerals such as copper and iron. Vitamin D is hydroxylated in the liver.

6. Liver metabolism The primary function is the maintenance of sufficient plasma levels of the metabolites (homeostasis)
Carbohydrate metabolism:
1. The liver takes up glucose and other monosaccharides from the blood plasma -These sugars are then converted to glucose 6-phosphate and other intermediates of glycolysis (subsequently, they are either stored as the reserve carbohydrate glycogen or degraded)
-Another large part is converted into fatty acids
-Only a small fraction is used for the generation of ATP
2. The liver secretes glucose when there is a major decline in the blood glucose level
-This glucose is derived from the glycogen store
-If the glycogen store is already exhausted, glucose can also be synthesized by gluconeogenesis from lactate, glycerol, or the backbones of amino acids
Lipid metabolism
Amino acid and protein metabolism
Biotransformations

7. 1. The liver synthesizes fatty acids from acetate units
Liver metabolism The primary function is the maintenance of sufficient plasma levels of the metabolites (homeostasis)
Carbohydrate metabolism
Lipid metabolism:
1. The liver synthesizes fatty acids from acetate units
-The fatty acids formed are then used for the synthesis of fats and phospholipids (these are subsequently released into the blood as complexes with proteins-lipoproteins)
2. The liver can take up fatty acids from the plasma
-The conversion of fatty acids into ketone bodies, which are also excreted
3. The liver also uses acetate units to synthesize cholesterol
-It is then transported to other organs as a constituent of plasma lipoproteins
4. Excess cholesterol is converted into bile acids or excreted with the bile
Amino acid and protein metabolism
Biotransformations

8. Liver metabolism The primary function is the maintenance of sufficient plasma levels of the metabolites (homeostasis)
Carbohydrate metabolism
Lipid metabolism
Amino acid and protein metabolism:
1. The liver controls the amino acid level of the plasma
-It breaks down excess amino acids
-The nitrogen thus released is converted to urea and transported to the kidneys
-The carbon skeletons of the amino acids enter intermediary metabolism, where they are consumed either in the synthesis of glucose or in the production of energy
2. The liver is the site of the synthesis and degradation of most proteins and peptides in the blood plasma
Biotransformations

9. Liver metabolism The primary function is the maintenance of sufficient plasma levels of the metabolites (homeostasis)
Carbohydrate metabolism
Lipid metabolism
Amino acid and protein metabolism
Biotransformations
1. Steroid hormones and degradation products of the blood pigment hemoglobin are taken up by the liver
-They are inactivated and converted into highly polar metabolites
2. Drugs, ethanol, and other foreign substances (xenobiotics) are metabolized by similar reactions
-Preparation for excretion

10. The role of the liver in nutrition
The tissues are dependent on a constant supply of energy-rich metabolites to provide energy and as precursors for the synthesis of complex macromolecules
The liver:
1. It compensates the variation of the level of these fuels in the diet (buffering organ)
2. It supplies with metabolites (storage organ)
Biochemistry of nutrition:
1. Well-fed (= absorptive state)
-The energy requirements of the tissues are predominantly met by glucose
2. Starvation (= postresorptive state)
- The energy requirements of the tissues are met mainly by fatty acids, amino acids, and ketone bodies (exceptions: nerve tissue and erythrocytes)
The transition between these two states depends on the levels of energy-rich metabolites in the blood plasma (it is brought about by the joint action of hormones and signals from the nervous system)

11. Absorptive state (lasts for about 2 to 4 hours after a meal)
Anabolická fáze ve tkáních: 1. Játra-metabolity jsou přeměňovány na glykogen a tuky. 2. Svaly-syntéza glykogenu a proteinů. 3. Tuková tkáň- syntéza tuků. Tvorba energetických zásob.

12. Liver metabolism in the well-fed state

13. Postresorptive state (starvation)

14. Liver metabolism during starvation

15. The role of the liver in carbohydrate metabolism
The concentration of glucose in the blood is maintained constant (at 4-6 mM)
-this is achieved by the precise regulation of glucose- forming and glucose-utilizing pathways
In the liver, glucose is derived from:
1. Glycogenolysis
2. Gluconeogenesis
3. Fructose and galactose metabolism
4. Cori cycle (from lactate)
5. Alanine cycle (from alanine)

16. Liver glycogen
Glycogen serves as a carbohydrate reserve, from which glucose can be released
The human body can store up to 450 g of glycogen (a third in the liver)
Liver glycogen serves in the maintenance of the blood glucose level, it declines to zero in periods of starvation that last more than one day
Glykogen ve svalech, pouze pro energii, svaly nemají glukosa-6-fosfatasu

17. Liver gluconeogenesis
The glycogen reserves are already depleted after one day
The blood glucose level begins to fall
An increase of gluconeogenesis

18. Liver gluconeogenesis
Gluconeogenesis occurs predominantly in the liver (90%)
Precursors:
1. Amino acids derived from the muscles (prolonged fasting results in a massive degradation of muscle protein)
2. Lactate formed in erythrocytes and in muscles
3. Glycerol produced from the degradation of fats
Cortisol, glucagon, epinephrine: promote gluconeogenesis
Insulin: inhibits gluconeogenesis

19. Cori and alanine cycles
Degradation of proteins: -the amino groups are transferred to pyruvate, giving rise to alanine
Alanine is transported to the liver: -the carbon skeleton is converted into glucose, -nitrogen is converted into urea
Cori cycle:
Lactate (anaerobic glycolysis)
-transported to the liver
-converted back into glucose via gluconeogenesis
Glucose- is sent back to its site of utilization

20. The role of the liver in lipid metabolism
The liver is the most important site for the formation of fatty acids, fats, ketone bodies, and cholesterol
Absorptive state:
-The liver converts glucose via acetyl-CoA into fatty acids
-The liver can also retrieve fatty acids from lipids supplied with chylomicrons from the intestine
-The fatty acids are converted into neutral fats and phospholipids, VLDL are formed (transport of fatty acids to other tissues)
Postresorptive state:
-The adipose tissue releases fatty acids
-Fatty acids are taken up by the liver and oxidatively degraded to acetyl-CoA
-Acetyl-CoA is converted to ketone bodies

21. Biosynthesis of ketone bodies
Acetyl-CoA (ß-oxidation):
-can be oxidized by the citric acid cycle
-when the acetyl-CoA production exceeds the energy requirements of the hepatocytes (starvation, diabetes mellitus) they form ketone bodies (supply other tissues with energy)
Ketone bodies:
They are released by the liver into the blood, in which they are readily soluble (their levels are elevated during period of starvation)
3-Hydroxybutyrate and acetoacetate serve as the key metabolites in energy production (after 1-2 weeks of starvation, the nerve tissue also begins to utilize ketone bodies as energy sources)
Acetone is exhaled via the lungs
Ketonemia, ketosuria, ketoacidosis: when the production of ketone bodies exceeds their use outside the liver (prolonged starvation, diabetes mellitus)

22. Cholesterol metabolism
Sources of cholesterol: 1. the diet, 2. de novo synthesis from acetyl-CoA (liver)
Utilization of cholesterol: 1. the synthesis of bile acids, 2. building block for cell membranes, 3. stored in the form of lipid droplets, following esterification with fatty acids, 4. formation of VLDL (supply other tissues)
The liver takes up from the blood and degrades lipoprotein complexes containing cholesterol (HDL)

23. The role of the liver in amino acid metabolism
The main site of amino acid degradation is the liver (significant quantities of ammonia are released)
Ammonia:
-is toxic to cells (at higher concentrations, it is especially damaging to the nerve cells)
-has to be rapidly inactivated and excreted (in human, this occurs primarily through the formation of urea)
NH3 metabolism of the liver:
Amino nitrogen, which accumulates in the tissues, is transported in the blood incorporated into glutamine (Gln) and alanine (Ala)
Gln is deaminated by glutaminase to yield NH3 and glutamate (Glu)
The amino group of alanine is transferred to 2-oxoglutarate by alanine transaminase
NH3 is released from glutamate by oxidative deamination (catalyzed by glutamate dehydrogenase)
Aspartate (Asp), the second amino group donor in the urea cycle, also arises from glutamate (catalyzed by aspartate transaminase)
glutamate dehydrogenase- an enzyme typical of the liver
aspartate transaminase, like alanine transaminase, is present with high activity in the liver

24. Urea cycle Urea: -It is neutral and non-toxic
-It can cross membranes by diffusion (small, uncharged molecule)
-It is highly soluble in water
-It can be easily transported by the blood and excreted in the urine
Urea synthesis:
-It occurs predominantly in the liver
-Urea is formed in a cyclic series of reactions; its nitrogen atoms are derived from ammonia and aspartate, and the carbonyl carbon from hydrogen carbonate

25. Bile acid metabolism Bile acids:
-steroids with 24 carbons (from one to three α-hydroxyl groups and a side chain with a carboxylate group)
-synthesized in the liver from cholesterol
-amphipathic (have a polar and an apolar side); act as detergents
-linked with an amino acid (glycine or taurine)- bile salts
-primary bile acids are formed in the liver, secondary bile acids in the intestine (by dehydroxylation of the primary bile acids)
Lipid digestion:
-facilitate the solubilization of dietary lipids during the process of digestion by promoting micelle formation (this action increases the effectiveness of pancreatic lipases, and promotes the absorption of lipids from the intestine)

26. Metabolism of bile salts (exlusively in the liver)
4. a 5. Intestinal bacteria in the colon produce enzymes that can attack and alter the bile salts
6. Most of the bile acids are reabsorbed from the intestine and, following transport to the liver, returned once again into the bile (enterohepatic circulation)
1. The biosynthesis of bile acids from cholesterol
2. The conjugation with the amino acids
3. The concentration in the gallbladder by removal of water

27. Transformation products
Biotransformations (= the detoxification and elimination; predominantly in the liver)
Foreign substances: Self-substances
Xenobiotics (natural origin) Steroid hormones
Pharmaceuticals Bile pigments
Synthetics
Transformation reactions
Either incorporate new functional groups into apolar compounds, or alter groups that are already present in the molecule
The result is an increase in the polarity and a decrease in the biological activity or toxicity of the substance
The most important reactions:
Oxidative reactions: hydroxylation, epoxide formation, dealkylation, deamination
Reductive reactions
Methylation
Desulfuration
Poorly soluble, biologically active, some toxic
Phase I
Foreign substances can be of natural origin (xenobiotics), or may have been synthetically produced by man.
The chemical reactions involved are similar to those used for the detoxification and elimination of the body‘s own wastes.
Transformation products

28. Transformation products
Biotransformations
Transformation products
Conjugate formation:
These reactions couple substrates to highly polar, often negatively charged molecules
The enzymes involved are transferases
1. Glucuronidation
2. Esterification with sulfate
3. Amidation with glycine and glutamate
Phase II
Conjugate
Bile
Urine

29. Cytochrome P450 system Monooxygenases that contain a heme group
Enzymatic hydroxylation (phase I reactions): drugs, ethanol
Also involved in the biosynthesis of steroid hormones, bile acids, eicosanoids, and unsaturated fatty acids
Low specificity
Found in the liver, adrenal cortex
Cytochrome P450-dependent monooxygenases: reactions
The reductive cleavage of molecular oxygen (one atom is incorporated into the substrate, the other is released as water)
The reducing equivalents are derived from NADPH + H+
Hem váže CO, absorpce při 450nm (proto název P450). Léky i etanol jsou degradovány stejným systémem, proto společné požití vede ke škodlivému nárůstu efektu.

30. Examples of the Cyt P450-dependent reactions:
Cytochrome P450 system
Examples of the Cyt P450-dependent reactions:
The hydroxylation of aromatic rings (drugs, steroids)
Aliphatic methyl groups can be oxidized to hydroxyl functions
The epoxidation of aromatics yields products that are toxic (benzopyrene)
Dealkylations

31. The alcohol content of alcoholic beverages
Ethanol metabolism
The alcohol content of alcoholic beverages
Given as percent by volume (conversion to gram ethanol, density = 0,79 kg/l)
A bottle of beer: 0,5l with 4% alcohol contains 16g of ethanol
A bottle of wine: 0,7l with 12% alcohol contains 66g of ethanol
Ethanol is rapidly taken up from the digestive tract by diffusion
The maximum blood alcohol level is reached within minutes Alcohol is rapidly distributed throughout the organism (a large amount is taken up by muscles and the brain)
The lethal blood alcohol level is %

32. Ethanol degradation (in the liver)
Other enzymes are also involved in ethanol degradation (catalase, Cyt P450)
Alcohol dehydrogenase oxidizes ethanol to ethanal (acetaldehyde)
It is futher oxidized to acetate by aldehyde dehydrogenase
Acetate is then converted to acetyl-CoA by acetate-CoA ligase in an ATP-dependent reaction

33. Liver damage
Elevated consumption of ethanol over many years may lead to liver damage (the limit for a man is about 60g daily, and for a woman about 50g); these values are strongly dependent on body weight, health and the use of medication
The high levels of NADH and acetyl-CoA inhibit citric acid cycle activity and ketogenesis, whereas they exert a stimulatory effect on the synthesis of neutral fats and cholesterol (an increase in the storage of lipids leads to a fatty liver- it is reversible)
In chronic alcoholism- liver cell death (cirrhosis of the liver, an irreversible, progressive loss of liver function)

34. Bilirubin metabolism and excretion
Bilirubin (degraded product of hemoglobin) is transported in the blood bound to albumin
The liver takes up bilirubin and conjugates it with glucuronic acid
Conjugated bilirubin is excreted via the bile ducts into the gut
It is converted to urobilinogen (most of it is oxidized to urobilin in the colon and excreted)
Some urobilinogen is absorbed from the small intestine and enters the enterohepatic circulation (most is excreted in the bile, some reached the systemic circulation and is excreted in the urine)

35. Hyperbilirubinemia
Elevated bilirubin levels in the blood (>10 mg/l); bilirubin may diffuse into peripheral tissues, giving them a yellow color (jaundice)
Cause:
1. Pre-hepatic: excessive formation of bilirubin by increased degradation of erythrocytes (icterus neonatus, hemolytic anemia)
2. Hepatic: insufficient processing of bilirubin as a result of liver defects (hepatitis, liver toxic damage, cirrhosis, hepatic failure)
3. Post-hepatic: by impaired excretion of gall (obstructive jaundice due to gallstones, inflammation of biliary tract)
Unconjugated bilirubin can cross the blood-brain barrier, leading to brain damage
Jaundice in neonates (increased bilirubin degradation+immaturity of the conjugation enzymes): phototerapy – isomerization of bilirubin to more soluble pigments

36. ↑↑(UC) N ↑ ↑↑(both) ↑ N ↑↑(C) ↑ ↓ Bilirubin
blood urine deriv. in feces
Urobilinogen
blood urine
Prehepatic
↑↑(UC) N ↑
↑ ↑
Intrahepatic
↑↑(both) ↑ N
↑↑ ↑↑
Posthepatic
↑↑(C) ↑ ↓
↓ ↓

37. Biochemical assessment of liver function
Basic tests: albumin, ALT, AST, GGT, ALP, bilirubin, U-urobilinogen, U-bilirubin
Special tests: serum protein electrophoresis, prealbumin, glutamate dehydrogenase, cholinesterase, lactate dehydrogenase, oral glucose tolerance test (oGTT), P-Fe
Very special tests: ceruloplasmin, S-Cu, α1-antitrypsin, ammonia, P-bile acids, serological tests in hepatitis B, assessment of clotting factors

38. ALT (alanine aminotransferase):
increased when cells of the liver are inflamed or undergo cell death
rises dramatically in acute liver damage, such as viral hepatitis
AST (aspartate aminotransferase):
less specific for liver disease
raises in acute liver damage
GGT (gamma-glutamyltransferase):
produced by the bile duct (sensitive marker for cholestatic damage)
raises in alcohol toxicity
ALP (alkaline phosphatase):
associated with the biliary tract
raises in biliary tract damage and inflammation

39. Bilirubin
-total and direct (=conjugated)
Urine urobilinogen:
-insufficient function of the liver
Urine bilirubin:
-when plasma direct bilirubin is elevated (obstructive and hepatocellular icterus)