Practical Lab Physiology I - 2015

EXPLORATION OF THE LIVER

The liver is the organ with the most complex metabolic activity. The main metabolic functions
of the liver are: formation and excretion of bile, homeostasy of carbohydrates, synthesis and
secretion into the plasma of lipids, lipoproteins, controlling the cholesterol metabolism,
forming urea, seric albumines, coagulation factors, enzymes and many other proteins. The liver
is also involved in metabolisation and detoxification of some internal toxines, drugs or other
external substances.
The liver may be affected by many factors: infections, toxines, ischemia, drugs, autoimune
diseases.All these factors determine lesions and functional degeneration of hepatocytes
followed by symptoms or/and changes in hepatic tests. These changes reflect the intensity and
complexity of certain lesions.

I. FUNCTIONAL EXPLORATION OF THE LIVER

It is done for diagnosing liver disease → main biological liver syndromes detection. Biological
liver syndromes are a group of biochemical tests used to explore the structure or function of
impaired liver.
1. Hepatocitolitic syndrome
2. Extrahepatic cholestasis syndrome
3. Jaundice syndrome
4. Hepatopriv syndrome
5. Inflammatory syndrome → mesenchymal hyperreactivity

1. ENZYMES THAT REFLECT DAMAGE TO HEPATOCYTES

Hepatic cytolysis syndrome represents the disturbance in membrane function due to hepatic
cells injuries(replacement of active transport with passive diffusion and increased cell
permeability).The following enzymes are released into the blood in greater amounts.

1.1. TRANSAMINASES. These enzymes are involved in proteic methabolism, in transferring

an amino group (-NH2) from an aminoacid to alfaketoglutarate.

The best known are:

 Aspartataminotransferase (AST) is a mitochondrial enzyme, that can be found in liver,
heart, muscles, brain and kidneys. Acute tissue distruction releases the enzyme from the
cells leading to an increased plasma level.
 Alaninaminotransferase (ALT) is a cytoplasmatic enzyme, present mostly in liver, but
also in the heart and muscles. Its concentration is bigger in liver, therefore a plasma
increase is more specific for hepatic diseases than AST increase.

Normal values: 0-35 IU/L

Pathological variations of ALT and AST. Increased values in:
 Liver diseases (acute hepatitis, intoxications, ischaemia, hepatic steathosis)
 Cholestasis (choledocholithiasis)
 Alcohol abuse
 Miocardial infarction
 Muscle diseases
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AST/ALT Ratio is called Ritis index and has a normal value of 1,3. Values over 2 suggest
hepatitis and alcoholic cirrhosis. Its increase is due to hepatic cell distruction and due to the
lack of vitamine B6. Early increasing of transaminases in acute hepatitis are proportional to
tissue distruction intensity, without having a prognostic significance. The values stated are not
available for all laboratories, and from the standpoint of the clinician, it's important the report
to normal value than the absolute value. Their value can increase up to 100 fold. Their values
increase proportional, but there can be single ALT elevation with normal AST (ie. Chronic C
Virus Hepatitis). In chronic liver disease transaminases can be normal or elevated (only in acute
episodes). Normal values in advanced cirrhosis are due to increased liver fibrosis.

1.2. GLUTAMATDEHIDROGENASE is a mitochondrial enzyme, involved in urea synthesis.

Normal values: below 1.5 mIU/ml
Pathological variations. Increased values in:
 Severe acute hepatitis
 Acute episodes of chronic hepatitis and cirrhosis
 Biliary duct obstructions (elevations precede and overcome those of Alkaline
phosphatase)

1.3. LACTATDEHIDROGENASE (LDH) transforms lactic acid in piruvic acid. There are five known
isoenzymes, number 5 being characteristic to the liver.
Normal values: 100-190 IU/L
Pathological variations. Increased values:
LDH 5: LDH 1 and LDH 2:
 Acute hepatitis cardiovascular disease
 Chronic hepatitis kidney disease
 Cirrhosis anemias
 Liver cancer

Note: in hepatic jaundice increased of enzymes that reflecting hepatocytolisis is more intense
than those of cholestasis.

2. CHOLESTATIC ENZYMES

Cholestasis syndrome is defined as disturbances in biliary function of the liver (biliary excretion
or flow). It is characterised by increased values of the above mentioned enzymes associated
with elevated conjugated bilirubin.
Cholestasis can be intrahepatic (ie. primary biliary cirrhosis) or extrahepatic (ie.
choledocholithiasis).

2.1. ALKALINE PHOSPHATASE (AP) hydrolises phosphoric acid esters and is active at an optimal
pH between 8.6 and 9.1. It has three permanent isoenzymes: liver, bone and intestinal, and a
temporary one during pregnancy. The liver isoenzyme is involved in transports from the biliary
and sinusoidal pole of hepatocytes and biliary ducts.
Normal values: 30-120 IU/L
Pathological variations. Increased values in:
 Intra and extra hepatic cholestasis (GGT also increased)
 Primary and secondary liver tumors (metastases, values >1000 IU/L), primary biliary
cirrhosis
 Acute hepatitis (moderate)
 Bone tumor (increase is isolated)

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2.2. GAMAGLUTAMILTRANSPEPTIDASE (GGT) is a microsomal enzyme nonspecific for the liver.

Normal values: 1-94 IU/L
Pathological variations. Increased values in:
 Cholestasis (increases proportional with AP)
 Alcohol abuse
 Acute hepatitis
 Pancreatic diseases, Diabetes mellitus

2.3. 5-NUCLEOTIDASE is present in all tissues and mostly in billiary ducts and liver sinusoides. It
increases in cholestasis proportional with AP. In bone disease its values remain normal.

2.4. LEUCIN AMINOPEPTIDASE (LAP) is present in biliary ducts and its plasma concentration
elevates in duct obstruction.

2.5. COLINESTERASE is a ribosomal enzyme, with several isoenzymes, non specific for the liver.
Normal values: 5-12 IU/ml
Pathological variations. Decreased values in:
 Acute and chronic hepatitis
 Malnutrition

Note: in the mechanical jaundice increase of cholestatic enzymes is more intense than those
that reflect hepatocytolisis.

3. EXCRETO-BILIARY FUNCTION OF THE LIVER

Bile is the secretion product of the hepatocytes. It is continuously produced and collected into
the gallbladder between meals. During eating it is relished into the duodenum.
The main bile components are: water, mucin, biliary pigments, biliary salts, lecithin and
cholesterol.
Bilirubin is the main biliary pigment, generated from hem degradation (a non proteic group
that contains iron and belongs to hemoproteins). Hemoproteins represent a class of proteins
involved in oxygen transport and metabolism (i.e. Hemoglobin and mioglobin). 70 to 90% of
bilirubin comes from red blood cells hemoglobin, destroyed in the reticulo-endothelial system
(spleen, liver and bone marrow). The difference up to 100% comes from the degradation of
myoglobin, P-450 cytochrome, catalase and peroxidase especially in the liver and also from
inefficient erythropoiesis. The generated bilirubin is called unconjugated, is not soluble in water
and transported in plasma binded to albumin.

Unconjugated bilirubin enters the hepatocyte through facilitated difussion, then it is conjugated
with glucuronic acid resulting conjugated bilirubin. It is water soluble and is excreted in the
bile.

In the large intestine under the influence of bacteria conjugated bilirubin is transformed in
urobilinogen (UBG), 20% of it being reabsorbed in blood. A part of urobilinogen is eliminated in
urine by kidney filtration and the other part reaches the liver from were it will be excreted into
the bile. Unabsorbed UBG (80%) will be eliminated into the faeces as stercobilin.

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Plasma concentration of bilirubin is determined through diazo van den Berg reaction. Bilirubin
that directly reacts is the conjugated bilirubin, the difference being represented by the indirect
fraction (unconjugated bilirubin). The total amount of bilirubin is called total bilirubin.

Normal values:
Total Bilirubin 0.3-1 mg %
Conjugated Bilirubin (CB) 0.1-0.3 mg %
Unconjugated Bilirubin (UB) 0.2-0.7 mg %

Jaundice is defined as yellow coloration of skin and mucosae due to the increase of bilirubin
plasma values. It is taking place when total bilirubin plasma concentration is above 2-3 mg%
(see Table I). There are various causes generating jaundice, predominating conjugated or
unconjugated bilirubin.(see Table II).

Table I. Biochemical diagnosis of jaundice

Blood
Test Prehepatic Jaundice Hepatic Jaundice Posthepatic Jaundice
TB < 5 mg% 10-20mg% > 20 mg%
UB ↑↑ ↑ ↑
CB ↑ ↑ ↑↑
UB/TB > 0,8 0,25 – 0,8 0,15 – 0,25
BS abs + +++
Urine
UBG ++ ++ abs
B + ++ +++
BS abs + +++

Table II. Main causes of Jaundice and increased bilirubin levels(after Cecil Medicine ed. 23,
2008).
Elevated unconjugated bilirubin
1 Increased bilirubin production
2 Decrease in liver uptake of unconjugated bilirubin (rifampicin, furosemide, etc.)
3 Decrease in hepatic conjugation and generation of CB (newborn jaundice)
Liver disease
1 Acute or sub-acute hepatitis
2 Chronic hepatitis
3 Cholestatic liver diseases
 Intrahepatic biliary ducts inflammation
 Liver Metastases
 Contraceptives
 Steroids
 Primary biliary cirrhosis
 Drugs (eritromicin)

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Extra hepatic biliary obstruction

1 Choledocholithiasis
2 Biliary ducts inflammation
3 Tumor (cholangiocarcinoma)
4 External Compression of biliary ducts (Pancreas Carcinoma, Lymph nodes)

4. METABOLIC FUNCTION OF THE LIVER

Hepatopriv syndrome defines the protein synthesis deficit accompanied by defective synthesis
of other metabolites.

4.1. METABOLISM OF PROTEINS

Liver maintains the normal plasma concentration of proteins through de novo synthesis of
proteins from aminoacids and by food and tissue proteins metabolism. A decrease in proteins
plasma level signs a decrease of liver function.

Evaluation of liver function in proteins metabolism is realised through the following

determinations (Table III):
 Total serum proteins (NV: 55 – 80 g/l)
 Plasma proteins types:
- Albumins (NV = 35 – 55 g/l)
- Globulins (NV = 20 – 35 g/l)
- Fibrinogen (NV = 2 – 4 g/l)
 Fibrinogen (NV = 2 – 4 g/l) ↓ in chronic liver disease, cirrhosis

Table III. Changes of ELFO proteins in hepatic diseases

Test Normal values Variations
Absolute values Relative values
(g/l) (%)
Albumin 35 – 55 50 - 60 ↓ in chronic liver disease, cirrhosis
Alfa 1- globulins 2–4 4.2 – 7.2 ↓ in liver disease, ↑ in tumors
Alfa 2- globulins 5–9 6.8 - 12 ↑ in cholestasis
Beta - globulins 6 – 11 9.3 - 15 ↑ in cholestasis
Gamma - 7 – 17 13 – 23 ↑ in hepatitis, primary biliary
globulins cirrhosis

Quick time (NV: 11.1-13.1 sec) evaluates the protrombinic complex, represents a sensitive
marker of syntesis function of the liver with prognostic value in acute and chronic diseases. It
increases in cirrhosis.

4.2. METABOLISM OF LIPIDS

Liver has an essential role in cholesterol metabolism, in biliary acids synthesis, but also in the
metabolism of triglycerides, phospholipids and lipoproteins. The following tests are used (Table
IV):

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Table IV. Exploration of lipid metabolism in liver diseases

TEST NORMAL VALUES VARIATIONS
Total cholesterol 150 - 200 mg% ↑ in cholestasis , PBC, steatosis
↓ in cirrhosis
Triglycerides 50 - 150 mg% ↑ in cholestasis, hepatitis
↓ in cirrhosis
Lipoproteins
LDL cholesterol < 115 mg% ↑ in cholestasis, ↓ in cirrhosis
HDL cholesterol ↓ in cirrhosis
- femei > 40 mg%
- barbati > 50 mg%

Although the level of biliary acids is not currently determined it increases in hepatic diseases
(acute viral and toxic hepatitis) and biliary diseases (cholestasis) due to the decrease in their
clearence. Decreased secretion of biliary acids affects lipids digestion and absorbtion
generating steatorrhea (lipids in faeces). On the other hand increased biliary salts in the large
bowel (after cholecistectomy) generates diarrhea with water and electrolytes losses.

4.3. CARBOHYDRATES METABOLISM

Liver has an important anabolic and catabolic role in the carbohydrates metabolism. The
following tests are used in this way:

4.3.1 Blood Glucose (NV:70-110 mg%). Its blood level decreases in cirrhosis due to increased
tissue degradation of glucose as source of energy. Liver deposits of glicogen decrease,too.

4.3.2 Oral glucose tolerance test (OGTT). 75 g of glucose is administrated, a jeun, in 250 ml of
water and after 2 hours we determine the blood glucose. The test is modified (>140 mg%) in
cirrhosis due to the decreased capacity of liver to regulate glucose homeostasis.

4.3.3 Galactose tolerance test (Bauer test). It shows hepatocytes capacity to convert galactose
in glucose and deposit it as glicogen. 40 g of galactose is administered, a jeun, and we
determine the galactose eliminated in the urine in 24 hours. Normal it is below 2 g, and it is
increased in chronic liver diseases.

5. EXPLORATION OF LIVER MESENCHYMAL REACTIVITY

Inflammatory syndrome reflects mesenchymal hyperreactivity. It is investigated by usual

biochemical tests: ESR (↑), normal values: male: 1-10 mm/h, woman: 2-13 mm/h, ELFO - γ-
globulins (↑), normal values: 13 - 23%. Other tests: C-reactive protein (↑), serum fibrinogen -
↓¯ in chronic liver disease, cirrhosis.

II. MORPHOLOGICAL EXPLORATION OF THE LIVER

1. Ultrasonography allows liver or other organs exploration, as well as body regions with the
help of the ultrasounds. Succesive ultrasounds beam is focused over the investigated region
and then reflected ultrasounds are intercepted and projected on a screen. Different colour
tones between white and black are the result of ultrasound reflection due to different tissue
density. Through this method we evaluate the liver structure or its changes (ie. Hemangioma,
liver metastases).

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2. Contrast ultrasonography, uses an iv administrated substance (ie.Levovist), and makes a

better differential diagnosis of liver formations based on different substance captation
(depends on vessels type).

3. Computer tomography (CT) and Magnetic resonace investigation (MRI) are very precise and
elective methods, used when an accurate diagnosis cannot be provided otherwise.

4. Liver biopsy (performed under ultrasonographic guidance and with a special needle) allows
the taking of a liver sample for histological exam. Chronic hepatitis are evaluated in this way
(inflammation and fibrosis).

5. FibroScan (elastography) a modern technique that allows quantification of liver fibrosis using
a shock wave propagated through the liver tissue. It is noninvasive and harmless, easy to
perform and repeat. It is used for the staging of liver fibrosis. Values over 13 kPa indicates the
presence of cirrhosis.

6. Laparoscopy allows the visualisation of abdominal cavity and liver using a laparoscope
introduced in the abdomen through the ombilicus after the generation of a
pneumoperitoneum.

7. Laparotomy allows the visualisation of abdominal cavity and liver after a transversal or
vertical incision of the abdominal wall.

Bibliography:
1. Textbook of Medical Physiology, Arthur C. Guyton, John E. Hall.—11th ed.W.B Saunders
Company 2006, ISBN 0-7216-0240-1
2. Harrison’s Principles of Internal Medicine, D.L.Kasper, A.S.Fauci – 16 th ed. 2005, ISBN
0-07-140235-7
3. Oxford Handbook of Gastroenterologz and Hepatology, S.Bloom, G.Webster – edit.
Oxford University Press 2006, ISBN 0-19-856652-2
4. Esenţialul în Gastroenterologie şi Hepatologie, O.Pascu – editura Naţional 2003, ISBN
973-659-045

BULLETINS:

Evaluate the liver function knowing:

1. Total Proteins = 4 g%
ELFO: Albumins = 40%
Alfa 1 Globulins = 5%
Alfa 2 Globulins = 10%
Beta Globulins = 12%
Gamma Globulins = 33%
Quick time = 21 sec
Fibrinogen = 1.1 g/l
ESR = 40 mm/h

Hepatopriv, inflammatory syndrome

Chronic liver disease

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2. AST = 105 U/L

ALT = 140 U/L
TB = 21 mg%
AP = 290 U/L
GGT = 250 U/L
Urine ex: Ubg absent
B (++)
BS (++)

Hepatocytolitic syndrome, jaundice, cholestasis

Obstructive jaundice

3. AST = 451 U/L

ALT = 367 U/L
TB = 13 mg
AP = 135 U/L
GGT = 112 U/L
Urine ex: Ubg (++)
B (++)
BS (+)

Hepatocytolitic syndrome, jaundice, cholestasis

Acute liver disease

MULTIPLE CHOICE QUESTIONS

1. Which of the following is true:

A. AST does not increase in hepatic cytolysis syndrome
B. ALT increase in hepatic cytolysis syndrome
C. LDH decrease in hepatic cytolysis syndrome
D. Glutamatdehidrogenase does not increase in hepatic cytolysis syndrome

2. Which of the following enzymes increase in hepatic cytolysis syndrome:

A. AST
B. Colinesterase
C. AP
D. GGT

3. Which of the following enzymes increase in cholestasis:

A. AP and GGT
B. Colinesterase
C. TB
D. Cholesterol

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4. All of the follwing are true except:

A. Transaminases remain normal in acute hepatitis
B. Transaminases might be normal in cirrhosis
C. Transaminases could increase in hemochromatosis
D. AST could be increased in muscle injuries

5. Which of the following is true:

A. UB is water soluble
B. CB is water soluble
C. UB is the result of conjugation process of CB in the hepatocyte
D. UB is filtered and eliminated in urine under normal conditions

6. All of the follwing are true except:

A. UBG is reabsorbed in the gut
B. CB is water soluble
C. Bilirubin is the result of globin metabolisation from the hemoglobin molecule
D. TB is not over 1 mg%, under normal conditions.

7. Which of the following is false:

A. Jaundice means yellow coloration of skin and mucosa
B. Jaundice appears when TB is over 4-5 mg%
C. Jaundice may appear in cirrhosis
D. Jaundice may appear in acute hepatitis

8. Chronic hepatic diseases are characterized by:

A. Decreased gamma globulins
B. Decreased albumin
C. Decreased Quick T.
D. Increased Fibrinogen

9. All of the follwing are true except:

A. Total Cholesterol is decreased in cirrhosis
B. LDL is decreased in cirrhosis
C. HDL is decreased in cirrhosis
D. Triglicerides are icreased in cirrhosis.

10. Choose the false statement:

A. hepatocytolisis syndrome is assessed by determining transaminase
B. cholestasis syndrome is assessed by determining bilirubinâ
C. hepatopriv syndrome is assessed by investigating protein metabolism
D. hepatopriv syndrome is assessed by investigating lipid metabolism

Answers: 1-B, 2-A, 3-A, 4-A, 5-B, 6-C, 7-B, 8-B, 9-D, 10-B