1. A 65-year-old male presented with fever, abdominal pain, distension, and jaundice for 4 weeks. Imaging showed a diffuse process in the liver. Liver biopsy revealed adenocarcinoma infiltration of the liver. 2. A 58-year-old female with diabetes and elevated liver enzymes was evaluated. She had a history of elevated enzymes attributed to lipid medication years ago. Current labs showed elevated AST and ALT with normal ALP and GGT. She had weight gain and abnormal lipid profile. 3. The first case describes a patient with diffuse liver lesions found to be metastatic adenocarcinoma on biopsy. The second case involves a patient with metabolic risk factors and elevated amin

1. ENZYMES IN LIVERENZYMES IN LIVER
DISORDERSDISORDERS
Presenter – Dr.Preeti
Moderator- Dr.Hemalatha A

2. ObjectivesObjectives
 Anatomy
 Functions of liver
 Liver function tests
 Introduction to Enzymes
 Liver enzymes
 Case discussion

3. Anatomy LiverAnatomy Liver
The liver is the largest
organ in the body
It is located below the
diaphragm and extended
approximately from the
right 5th rib to the lower
border of the rib cage.

4. Anatomy LiverAnatomy Liver

5. Microscopy of LiverMicroscopy of Liver
Functional unit – Acini

7. Exogenous Endogenous
Bile and Urine
1.Carbohydrate metabolism
2.Harmone metabolism
3.Lipid metabolism
4.Protein and drug

8. LFTs are classified asLFTs are classified as::
 Excretory function tests: Bile pigments, salts,
acids, bilirubin
Metabolic functions tests : Carbohydrates,
Protiens, Fats
Synthetic capabilities : Protiens(albumin),
coagulation factors
Detoxification : Ammonia, drugs
Tests of liver injury : Enzyme assays, autoimmune
markers, markers of hepatitis virus infections

9. Uses of LFTsUses of LFTs
Diagnosis of type of jaundice- etiology
Assess severity & follow trend of liver disease
Detect latent liver disease
Screening of infective hepatitis
Screen drug hepatotoxicity

10. Indication and limitation of LFTIndication and limitation of LFT
Indication-
◦ Screen for liver diseases
◦ Identifying the nature of liver
diseases( hepatocellular, cholestatic, or
infiltrative.)
◦ Assess severity and prognosis of liver disease
◦ Follow up the course of liver disease.
Limitations –
◦ Do not necessarily assess liver function.
◦ Lack sensitivity
◦ Lack specificity

11. Tests included under LFTTests included under LFT
1. Total Bilirubin
2. Conjugated Bilirubin
3. Total protien
4. Albumin
5. Coagulation factors.

12. EnzymesEnzymes
Introduction
1. Enzymes are biological catalysts that speed up the
rate of the biochemical reaction.
2. Most enzymes are three dimensional globular
proteins (tertiary and quaternary structure).
3. The molecule for which enzyme catalyzes reaction is
called Substrate.

13. Structure of enzymesStructure of enzymes

14. How do enzyme work??How do enzyme work??
 By lock and key mechanism.
 Converts the complex structures into simple structures

15. Concentration & ActivityConcentration & Activity
Depends on rate of synthesis and degradation
Control occurs at transcriptional and translation
levels
Do not effect the value of equilibrium – constant
between reactants and products.
The 3 dimensional shape of the active site is a vital
determinant in recognition and specificity of
process.

16. Enzyme classificationEnzyme classification
Class type Reaction catalyzed
Oxidoreductases Oxidation–reduction reactions
Transferases Transfer of functional groups
Hydrolases Hydrolysis reactions
Lyases Group elimination to form double
bonds
Isomerases Isomerizations
Ligases Bond formation coupled with
ATP hydrolysis

17. Factors affecting plasma activityFactors affecting plasma activity
1. Increased release
 Necrosis of cell
 Increased permeability of the membrane
 Increased synthesis
 Increase in tissue source of enzyme in
malignancy.
2. Impaired deposition
 Increased levels in obstructive jaundice
 Increased levels in renal failure.

18. Liver enzymesLiver enzymes
5’ Nucleotidase
GGT

19. Liver enzymesLiver enzymes
Aspartate Aminotransferase(AST)
Alanine Aminotransferase(ALT)
Alkaline phosphatase
Gamma glutamyl transferase.
5’ Nucleotidase

20. Cellular location of enzymesCellular location of enzymes

21. Serum emzyme tests are grouped inSerum emzyme tests are grouped in
2 categories2 categories
1.Enzymes whose elevation reflects damage to
hepatocytes
2. Enzymes whose elevation reflects cholestasis

22. ENZYMES WHOSE ELEVATION
REFLECTS DAMAGE TO
HEPATOCYTES

23. Aspartate Aminotransferase(AST)
Alanine Aminotransferase(ALT)

24. Aspartate transaminaseAspartate transaminase
Biochemistry & Physiology
1. Normal range – 5- 40 IU/Lt.
2. Half life – 17hrs.
3. Its 7000 times more than the plasma
4. Catalyzes the reversible transfer of aspartate
group
 Alpha ketoglutarate Glutamate +
Oxaloacetate
AST

25. Aspartate transaminaseAspartate transaminase
Found in
1. Liver
2. Cardiac muscle
3. Skeletal muscle
4. Kidney
5. Brain
Released - Damage to liver cell membrane.
1. Alcohol –related
2. Drug -induced liver injury,
3. Hemolysis
4. Myopathic processes

26. 2 forms of AST are known-
1. Cytosolic
2. Mitochondrial (mAST) – it is synthesized in
precursor form (pre-mAST)
converted to mature mAST
•mAST/total AST ratio – marker of chronic alcohol
consumption

27. Alanine transaminaseAlanine transaminase
Biochemistry & Physiology
1. Normal range – 5- 40 IU/Lt.
2. Half life – 47hrs.
3. Its 3000 times more than the plasma
4. Catalyzes the reversible transfer of Alanine group
 Alpha ketoglutarate Glutamate +
Pyruvate
ALT

28. Alanine transaminaseAlanine transaminase
Found in
1. Liver
2. Kidney.
Impaired synthesis seen in
1. Pyridoxine deficiency
2. Hepatic fibrosis
3. Hepatic cirrhosis

29. Diagnostic significance ofDiagnostic significance of
TransaminasesTransaminases
Levels of >1000 IU/L occurs in –
Acute viral hepatitis
Toxin and drug induced hepatitis
Ischaemic liver injury
In most acute hepatocellular disorders ALT is
higher or equal to AST

30. ALT is usually normal in alcoholic liver disease ;
can be sometimes low due to an alcohol
induce deficiency of pyridoxal phosphate
 AST/ALT <1 is seen in Non Alcoholic Staeto
Hepatitis and viral hepatitis

31. Determination of these enzymes are helpful in
distinguishing hepatocellular from cholestatic
jaundice
Increase in AST and ALT is much more ( >500
IU/L)in hepatocellular jaundice than in cholestatic
jaundice (>200 IU/L)
Persistence of elevated ALT and AST beyond 6
months in a case of hepatitis indicates
development of chronic hepatitis

32. AST/ALT RATIOAST/ALT RATIO
Normal AST/ALT ratio is 0.8.
A ratio > 2 is seen in
1. Alcoholic hepatitis
2. Hepatitis with cirrhosis
3. Non alcoholic steatohepatitis
4. Liver metastasis.
5. Myocardial infarction
6. Erythromycin treatment

33. Continu…Continu…
A low ratio (ALT is higher) is seen in
1. Acute hepatocellular injury
2. Toxic exposure
3. Extrahepatic obstruction (cholestasis)

34. Assay for AST & ALTAssay for AST & ALT
 Vit B6 important requirement for AST and ALT assays
 Normal serum levels upto 40 IU/dl for both
↑ Aspartate in reaction glutamate→ α ketoglutararte
Glutamate
dehydrogenase
NAD NADH(Colour indicator)
Oxaloacetate malate
malate
dehydrogenase
NAD NADH(Colour indicator)
Measured
spectrophotometrically

35. Alanine ALT Puruvate
NA
D NADH

37. Mild Chronic Elevation inMild Chronic Elevation in
Serum AminotransferasesSerum Aminotransferases
Step one
◦ Medications and supplements
◦ Alcohol use
◦ Viral hepatitis B and C
◦ Hemochromatosis
◦ Fatty liver (hepatic steatosis and
steatohepatitis )

38. Mild Chronic Elevation inMild Chronic Elevation in
Serum AminotransferaseSerum Aminotransferasess
Step two
◦ Muscle disorders
◦ Thyroid disease
◦ Celiac disease (less)
◦ Adrenal insufficiency (less)
◦ Anorexia nervosa (less)

39. Mild Chronic Elevation inMild Chronic Elevation in
Serum AminotransferaseSerum Aminotransferasess
Step three
◦ Autoimmune hepatitis
◦ Wilson's disease
◦ Alpha-1-antitrypsin deficiency

40. Mild Chronic Elevation inMild Chronic Elevation in
Serum AminotransferaseSerum Aminotransferasess
Step four
◦ A liver biopsy is often considered in patients
in whom all of the above testing has been
unyielding.
◦ However, in some settings, the best course
may be observation.

41. Enzymes whose elevation
reflects cholestasis

42. 3 enzyme activities are important –
1.Alkaline phosphatase (ALP)
2. 5’nucleotidase (5’NT)
3. Gamma glutamyl transferase (GGT)

44. Alkaline phosphatesAlkaline phosphates
• Biochemistry and Physiology
1. Type of hydrolase
2. Plasma ALP forms are coded by single gene on
chromosome 1- producing tissue nonspecific
enzymes.
3. Two other genes on chromosome 2 – Code for
placental & intestinal origin of ALP.

45. 4. Bound to cell membranes.
5. Facilitate the movement of substances across cell
membrane.
By cleavage of pyrophosphate
6. Released by ingestion of Fatty foods.
7. The half life of isoenzymes of ALP
• Intestine – 1 minute
• Bine – 1 day
• Liver – 3 days
• Placenta – 7days.

46. Reference range and preanalyticalReference range and preanalytical
variationvariation
Normal - 39-117 IU /Lt
Peak in Teens
Increases in postmenopausal women – Bone
isoenzymes
Increase in ALP seen in
1. High body mass index
2. Antiepileptic agents
3. Smoking

47. Continu….Continu….
 Decrease in ALP is seen in
1. Oral contraceptives
2. Fibric acid derivatives.
3. Blood transfusion
4. Cardiopulmonary bypass

48. • Increases in > 10 times of nomal
1. Obstruction of biliary tract – by stones in ducts.
2. Space occupying lesions
3. Ascending cholangitis

49. Assay of ALPAssay of ALP
• By using p- nitrophenyl phosphate as a substrate
in alkaline pH
Inorganic phosphate + highly colored para –
nitrophenoxide anion.
Measured by spectrophotometer at 340nm.
Hydrolysis

51. Gamma glutamyl transferaceGamma glutamyl transferace
• Regulates the transport of aminoacids across the
membrane.
Glutathion
GGT
Free amino acid
• Clinically used to differentiate the increased level
of ALP in biliary tract disease from bone
disease.
• Half life – 10days.

52. • Serum GGT is increased in = >10times of the
normal
1. Alcoholism
2. Chronic Cholestasis
3. Malignant infiltration of the liver
4. Drugs - acetaminophen and phenytoin and
carbamazepine ( 5 times the normal)
• GGT and 5’NT is especially used to assess the
nature of ALP

53. Assay of GGTAssay of GGT
• Most assays for GGT utilize the substrate -γ
glutamyl–p-nitroanilide.
• In the reaction catalyzed by GGT
• p-nitroaniline is liberated
chromogenic
measured by spectrophotometer

54. 5’Nucleotidase5’Nucleotidase
• Biochemistry and physiology
1. Is a phosphoric monoester hydrolase.
2. Aslo called 5’ Ribonucleotide phosphohydrolase
3. 5’ Ribonucleotide + H2O = Ribonucleoside + PO4
4. It’s a cytoplasmic membrane bound phosphatase.

55. FunctionsFunctions
Extracellular adenosine production
Nutrient absorption
Cell proliferation
Is derived predominantly in liver.

56. Reference range and preanalyticalReference range and preanalytical
variationvariation
5 -NT is normally present at low activities in′
children, rises in adolescence and plateaus until
age 40
Increase in 5’ NT - 2nd
and 3rd
trimester of
pregnancy.
Increase in Antiepileptic drug intake.

57. • Assay of 5’ Nucleotidase
1. By measurement of nucleoside released by action
of 5’NT is required.
• Causes of abnormal result
1. Acute hepatitis
2. Ovarian carcinoma
3. Rheumatoid arthritis

58. Lactate dehydrogenaseLactate dehydrogenase
• Biochemistry and physiology
1. Is a Oxidoreductase enzyme
2. Acts on CH-OH group of donors with NAD+ as
acceptor.
Lactate + NAD+ Pyruvate +NADH+ H+
3. Major isoenzyme – 4 tetramers
H(lactate)
M (pyruvate)

59. 4. 5 isoenzymes are seen – LD 1 – LD 5
5. LD1 & LD2 – Cardiac muscle , kidney &
erythrocyte.
6. LD4 & LD5 - Liver & skeletal muscle.
7. Total LD activity – 150IU/Lt
8. Half life – 4 hrs.

60. Significance of LDHSignificance of LDH
> 500 IU / Lt with ALP with normal AST and ALT
Indicate space occupying lesions.

61. Algorithm for Diagnosis of Liver DiseasesAlgorithm for Diagnosis of Liver Diseases

62. Case 1Case 1
65 yr old male with history of fever and altered
bowel habits , abdominal pain and distention pof
abdomen sincev 4 weeks.
tender hepatomegaly
Ascitis , Icterus - +
Bilirubin – 11 Albumin – low
ALT - 30 Cultures - negative
AST - 84 Antibiotics – No help
ALP - 820 PT - High
GGT – 468

63. USG - Possible diffuse process
Repeat CT – no focal lesions.
Became coagulopathic and tachycardiac.
DIAGNOSIS ??

64. Liver biopsy - Multiple lesions seen –
Adenocarcinoma.
Liver infiltration by carcinoma.

68. Case 2Case 2
58-year-old with type 2 diabetes was sent for
evaluation to the hepatology clinic with persistent
elevation of liver enzymes and complaints of
fatigue, dull right upper quadrant abdominal pain,
nausea and mild leg swelling for the past 3–4
months.
From routine laboratory data 3–4 years ago the
patient was found to have mildly elevated liver
enzymes, which was attributed to a lipid-lowering
agent (niacin), which was stopped at that time.
 she has not been on lipid-lowering medication
since then

69. . Laboratory work was not repeated until recently,
when she was found again to have elevated
aminotransferase levels.
Past history – uneventful except for the 6kg
wieght gain .
CBC –normal
Albumin - 3.2 g/dL
AST - 67 U/L
ALT - 89 U/L
ALP -166 U/L
GGT - 70 U/L

70. Total cholesterol - 212 mg/dL
LDL cholesterol - 144 mg/dL
Triglycerides - 316 mg/dL
USG abdomen – hepatomegaly
Liver biopsy - significant fatty change (60%) .Portal
areas showed mild chronic lymphocytic infiltration
without interface changes. Hepatic lobules showed
significant hepatocyte degeneration (balloon
degeneration and Mallory hyaline) and moderate
inflammatory infiltrate consisting of lymphocytes
and neutrophils.

71. Nonalcoholic Fatty Liver Disease (NAFLD)