Post on 28-Jan-2020
1 | P a g e
LIVER FUNCTION TESTS
A- Metabolic Functions of the Liver:
1. The liver plays a major role in carbohydrate, lipid and protein homeostasis,
with the processes of glycolysis, the Krebs cycle, gluconeogenesis, glycogen synthesis and glycogenolysis, lipogenesis, ketogenesis, amino acid synthesis and degradation, and protein synthesis; all taking place in the hepatocytes.
2. Hepatocytes also metabolize and detoxify endogenous (haem) and exogenous
products (drugs), which are then excreted via the biliary tree.
B- Importance and Classification of Liver Function Tests: Importance:
Liver function test are valuable for:
1. Diagnosis of liver disease. 2. Categorize the liver disease into its appropriate category. 3. Following the progress of liver disease.
Classification: Liver function tests can be classified according to many categories:
A- According to its function of the test:
1. Basic Metabolic function tests: include estimation of serum levels of liver
enzymes e.g. sGPT (ALT) and sGOT (AST).
2. Excretory function tests: include estimation of serum levels of total and direct
bilirubin.
3. Synthetic function tests: include estimation of serum levels of albumin and
cholesterol.
B- According to the type of variables:
1. Hepatic anion transport tests: as bilirubin excretion and plasma bile acid
conjugation.
2. Plasma protein abnormalities: they include
a) Determination of plasma albumin: (see section No. 3).
2 | P a g e
Clinical Chemistry of Liver
b) Determination Coagulation factors: Through estimation of: Vit. K (fat soluble vitamin): where the decrease in the level of this
vitamin (as in cholestasis) leads to increase the tendency of the patient to bleeding and failure of lipids absorption.
Prothrombin time: due to deficiency of one or more coagulation factors
as in hepato cellular damage.
c) Plasma Ig: they not have large clinical significance but important in
chronic cases as in liver cirrhosis ↑↑ IgA, ↑IgM and ↑IgG
d) Ceruloplasmin: it is metalo-protein formed if Cu bind with protein and
act as α1–protease inhibitor.
3. Plasma enzyme tests:
a) Estimation of the soluble enzymes released from cytoplasm and
mitochondria.
b) Estimation of the enzymes released from membrane where they are bind
to it. They include: o γ-glutamyl transferase (GGT). o Alkaline phosphatase (ALP).
c) Certain enzymes that their synthesis is impaired when hepato cellular
function is greatly impaired e.g. cholinesterase enzyme
4. Metabolic capacity tests:
Galactose clearance. Caffeine clearance.
C- According to type of liver disease:
1. Tests of hepatic fibrosis:
Collagen: that deposited in space on basement membrane. N-terminal peptide.
2. Liver cirrhosis induced by alcoholic drink: estimation of desialated
transferin.
3. Cholestasis tests: e.g. Estimation of total and direct bilirubin.
3 | P a g e
Some of routine liver function test group:
Clinical Chemistry of Liver
4. Liver tumors: e.g. Estimation of alpha-fetoprotein as liver tumor marker.
Liver Function Tests: Albumin
The liver also makes albumin, an essential protein that circulates in blood.
Albumin levels are low in people with severe chronic liver disease,
because the liver does not make normal amounts of albumin.
However, albumin levels also may fall in a variety of medical conditions.
A low albumin level is often temporary, so it is not a reliable way to
diagnose liver disease.
Liver Function Tests: Bilirubin
Bilirubin is a waste product from the breakdown of red blood cells. The liver
processes bilirubin so it can be excreted in stool. Bilirubin flows through the liver's
bile ducts, dissolved in bile.
Bilirubin blood levels may be elevated in people with impaired bile flow. This can
occur in severe liver disease, gallbladder disease, or other bile system conditions.
Very high bilirubin levels cause jaundice, in which the skin and whites of
the eyes turn yellow. Bilirubin can be a useful liver function test in people with a
known bile flow problem. An elevated bilirubin may also be present in
people with a type of anemia, called hemolytic anemia.
Test Function
Plasma albumin. Protein synthesis.
Plasma total and direct bilirubin. Hepatic anion transport.
Plasma enzyme activities
ALT/AST. Hepatocellular integrity.
Alkaline phosphatase and GGT. Presence of Cholestasis.
4 | P a g e
1- BILIRUBIN METABOLISM
(Excretory and Anion Transport Hepatic Function)
Sources of Bilirubin: Hemoglobin;
o RBC breakdown; 90% of RBC breakdown occurs within reticulo-
endothelial system (RES) cells (mainly in spleen).
o Ineffective erythropoiesis (in bone marrow).
Other haem containing proteins e.g. myoglobin and cytochrome P450 (mainly
in liver).
70 to 80% of daily bilirubin production is derived from the breakdown of
senescent red blood cells, while the remainder is derived from ineffective erythropoiesis and the breakdown of other haem-containing proteins.
Total daily bilirubin production is 450 up to 550 µmol/day.
Formation of Bilirubin: Hemoglobin is broken down to globin and haem. Globin (a protein) is broken down to its constituent amino acids.
Haem (a 4 ring structure containing Fe2+ at its center) is broken down (via
biliverdin) to carbon monoxide, iron and bilirubin.
Biliverdin gives the green colour sometimes seen in a resolving bruise. The bilirubin at this stage is termed un-conjugated bilirubin because it has not yet been processed by conjugation in the liver.
5 | P a g e
Clinical Chemistry of Liver
Un-Conjugated Bilirubin (Hemo-Bilirubin, Indirect Bilirubin):
A hydrophobic molecule. Strongly bound (high affinity) to hydrophobic sites on albumin. Does not appear in urine. Free un-conjugated bilirubin normally <3 µmol/l. Can be displaced from binding sites on albumin by drugs (salicylates,
sulphonamides).
Free un-conjugated bilirubin is neurotoxic.
o At high concentrations it deposits in cell membranes (esp. basal ganglia)
causing kernicterus.
Uptake by the Liver: The un-conjugated bilirubin - albumin complex is carried in the plasma to the
hepatic sinusoids, enters the space of Disse and dissociates at the hepatocyte membrane.
The un-conjugated bilirubin is taken up by the hepatocytes by a carrier-
mediated process.
Within the hepatocyte the un-conjugated bilirubin is bound to ligandin.
6 | P a g e
Carried
Clinical Chemistry of Liver
Conjugation by the Liver: The bilirubin is then conjugated with glucuronic acid by UDP-glucuronyl
transferase (UDPGT-I) to bilirubin mono-glucuronide (BMG) and by UDPGT- II to bilirubin diglucuronide (BDG).
Conjugated bilirubin is more water soluble and can be excreted in bile or urine.
Under normal circumstances there is no conjugated bilirubin present in plasma.
Excretion into Bile: Conjugated bilirubin is transported out of the liver cells into the bile canaliculi
by an energy-dependant carrier-mediated process which is sensitive to cell injury.
This canalicular excretion step rather than conjugation is thought to be the rate-
limiting step in bilirubin metabolism.
Bile flows through the canaliculi, into the bile ducts, and finally into the
duodenum.
Reticuloendothelial cells
Spleen - Bone Marrow
Biliverdin Bilirubin
Iron Amino acids
Heme
Systemic circulation
Bilirubin
Albumin
on albumin
Kidney
Urobilin, brownish
to Urine
Old RBCs
Hb Globin Liver
Bilirubin
Conjugation with
Glucuronic acid
Intestine
Free Bilirubin
Dr. Kakul Husain Firoz 5
Reabsorption
Urobilinogens,
colorless
Urobilin, brownish
to feces
8 | P a g e
BILIRUBIN:
Bilirubin is usually measured by the Jendrassik Grof modification of the van
den Bergh reaction: conjugated bilirubin (direct-reacting) (water soluble). unconjugated bilirubin (indirect-reacting) (water insoluble).
Normal serum total bilirubin <17 µmol/l (all unconjugated). Normal urine contains no unconjugated bilirubin, since it is albumin-bound and
not filtered.
Bilirubin only appears in urine when conjugated bilirubin is elevated in
plasma.
Increased amounts of unconjugated bilirubin are found in plasma in: TWO
o Increased bilirubin production. o Decreased uptake or conjugation of bilirubin. o in generalised hepatocellular dysfunction. o in specific rare inherited syndromes (Gilbert’s and Criggler-Najjar
syndromes).
Increased amounts of conjugated bilirubin are found in the plasma (and urine)
in:
Decreased excretion of bilirubin:
o in obstructive liver disease.
LIVER DISORDERS AND JAUNDICE
Jaundice or icterus is the yellow appearance of skin and sclerae due to the
presence of an excessive amount of bilirubin (jaundice becomes clinically visible when serum bilirubin is > 40 µmol/l).
The liver has a large reserve capacity- jaundice only appears with severe
impairment of liver function. CLASSIFICATION OF JAUNDICE ACCORDING TO ITS CAUSE:
A. Prehepatic: Excess bilirubin production.
B. Intrahepatic:
9 | P a g e
Decreased conjugation of bilirubin by liver cells. Decreased excretion of bilirubin into bile canaliculi.
C. Posthepatic: Biliary obstruction.
A. PREHEPATIC JAUNDICE
(Increased Production of Bilirubin).
1- Haemolytic Disorders: Abnormal haemoglobins (e.g. sickle cell anaemia). RBC membrane defects (e.g. hereditary spherocytosis). RBC enzyme defects (e.g. G6PD deficiency). Autoimmune haemolytic anaemias. Blood group incompatibility; o foetal / maternal (newborns) o transfusion.
Bacterial toxins causing haemolysis. Malaria.
2- Ineffective Erythropoiesis:
Megaloblastic anaemias.
Results of laboratory tests in cases of excess bilirubin production:
Increased un-conjugated bilirubin (rate of bilirubin production exceeds
maximum rate of conjugation). Haemolytic jaundice is never deep (except in newborns).
Normal conjugated bilirubin (no obstruction). Urine bilirubin negative. Urine urobilinogen increased (increased flux through pathway). Other liver function tests normal. Haematological tests: o increased reticulocyte count. o tests to identify cause of haemolysis (RBC morphology, abnormal
haemoglobin, Coombs test, etc.).
10 | P a g e
B. INTRAHEPATIC JAUNDICE
(Decreased Handling of Bilirubin by the Liver)
Specific Inherited Defects In Bilirubin Metabolism (Rare)
1- Defect In Uptake / Conjugation Of Bilirubin:
A- Gilbert’s Syndrome: Inherited disorder - decreased expression of UDPGT I due to a TA insertion in
the TATA box.
Mildly increased levels of unconjugated bilirubin (levels up to 85 µmol/l),
made worse by viral infections and fasting.
B- Criggler-Najjar Syndromes :
Criggler-Najjar type I syndrome :
Inherited disorder - severe mutation of UDPGT I gene leading to absent
UDPGT I activity.
Severe disorder with very high levels of unconjugated bilirubin.
Criggler-Najjar type II syndrome (Arias syndrome):
Inherited disorder - milder mutation of UDPGT I gene leading to
reduced UDPGT I activity.
Milder than type I.
2- Defect In Excretion Of Bilirubin Into Bile Canaliculi: Dubin-Johnson and Rotor’s Syndromes :
Benign inherited disorders. Increased levels of conjugated bilirubin in serum and urine. Due to impaired excretion.
3. POSTHEPATIC JAUNDICE
Causes of extrahepatic obstruction (“cholestasis”):
In the lumen (gallstones, worms). In the wall (bile duct carcinoma, stricture, atresia).
Outside the wall (carcinoma of head of pancreas, lymph node enlargement at
11 | P a g e
porta hepatis).
Clinical and biochemical features of obstruction (intrahepatic or extrahepatic):
Jaundice, dark urine, pale stools. The jaundice may be very deep, despite a generally well patient. Increased conjugated bilirubin in serum and urine.
Types of Gallstones: 1- Cholesterol stones. Common gallstone. Predisposing factors:Obesity, increasing age, female sex, diabetes, hyperlipidaemia. Chronic cholecystitis. Relative deficiency of tri- and dihydroxy bile salts leading to "lithogenic bile".
o Oral chenodeoxycholic acid has been used therapeutically.
2- Pigment stones.
Excess bilirubin production in haemolytic anaemias.
Presentation: Intermittent passage of gallstone: Severe right upper quadrant abdominal pain (biliary colic). Intermittent jaundice and bilirubinuria. Intermittent elevations of ALP, GGT. Prolonged painless obstructive jaundice. Acute cholecystitis.
CLASSIFICATION OF JAUNDICE ACCORDING TO TYPE OF BILIRUBIN
A- Unconjugated Hyperbilirubinaemia
Types
Physiological jaundice
Jaundice develops on days 3-5 (never on day 1). Levels of bilirubin may reach 150 µmol/l, all unconjugated.
Usually phototherapy is all the treatment that is required.
Blood group incompatibility ("haemolytic disease of the newborn”):
Jaundice develops on day 1. Rhesus or ABO systems commonly responsible, others rarely. A few foetal RBCs cross into maternal circulation, mother makes antibodies if
12 | P a g e
blood groups are incompatible. These IgGs then pass into foetal circulation, causing haemolysis of foetal RBCs. Excessive bilirubin production is not a problem for the foetus, since it crosses placenta and is conjugated, excreted by mother's liver. However, bilirubin starts rising rapidly after birth. Severe haemolysis in utero causes foetal anaemia and heart failure ("hydrops foetalis").
Inherited haemolytic disorders (as in adults).
Features: - Lethargy, hypotonia, poor feeding, spasticity and death. Treatment
• Phototherapy
Light at 440-470 nm causes formation in skin of isomers of unconjugated
bilirubin which are more soluble, excreted in urine.
B- CONJUGATED HYPERBILIRUBINAEMIA:
Types Neonatal hepatitis (Jaundice):
o Many infective agents can cause hepatitis in neonates (e.g. rubella,
CMV, herpes simplex, toxoplasmosis).
o In utero, bilirubin is transported across the placenta, conjugated and
excreted by the mother's liver.
o After birth the neonate must excrete the bilirubin. o Factors contributing to neonatal jaundice:**
Increased relative RBC mass. Shorter RBC lifespan. Immature liver, especially in premature neonates. Takes 2-
4 weeks to develop fully.
Many infective agents can cause hepatitis in neonates (e.g.
rubella, CMV, herpes simplex, toxoplasmosis).
13 | P a g e
2- BILE ACID (= BILE SALT) METABOLISM
Source of Bile Acids: Bile acids are soluble metabolites of cholesterol.
Formation of Bile Acids: The products of cholesterol metabolism are chenodeoxycholic acid and cholic
acid, known as primary bile acids because of their hepatic origin.
The rate limiting and regulated step in their formation is 7-α-hydroxylation of
cholesterol.
Prior to secretion into the bile canaliculi, the primary bile acids are conjugated
to glycine or taurine (this increases their solubility).
Conjugated bile acids are transported out of the liver cells into the bile
canaliculi by an energy-dependant carrier-mediated process (which is independent of the bilirubin transport mechanism).
In the GIT, bacterial enzymes deconjugate and α dehydroxylate the primary
bile acids and convert them to the secondary bile acids lithocholic acid and deoxycholic acid.
Most of the bile acids in the GIT are reabsorbed into the portal circulation
(75% in the ileum and 10% in the colon), taken up by the liver again and re- excreted (enterohepatic circulation).
Re-uptake of bile acids by the liver is highly efficient, but sensitive to liver
damage.
14 | P a g e
PROTHROMBIN TEST
Besides its functions in metabolism, the liver makes proteins that are essential to
normal blood clotting. True liver function tests check the liver's ability to make these
proteins. They include:
Prothrombin time (PT): A test of the time it takes for a blood sample to clot, under
specific conditions in a lab; if low levels of clotting factors are present, the
prothrombin time is longer.
International normalized ratio (INR): Not really a test, but a standardized way for all
labs to report PT, so their results can be compared accurately with each other.
PT and INR rise in people with severe liver disease because the liver fails to make
normal amounts of certain clotting factors. An elevated PT can have many other
causes besides liver disease, however.
PT is often checked together with PTT (partial thromboplastin time), which is not a
liver function test. If PT and/or PTT are elevated, a problem with bleeding or clotting
may be present.
***************