lecture no. 4 part 2 clinical Biochemistry

clinical Biochemistry

1. Clinical
Enzymology

2. •Enzymes are the protein catalysts that increase the rate of specific
chemical reaction in the body.
•Enzymes are found in small amounts mainly within cells,clotting
factors and digestive enzymes function naturally after secretion:
-Plasma specific –Thrombin
-Secreted-Lipase, Amylase
-Intracellular-transaminases, creatine kinase
•Injury or death of tissues can cause the release of tissue-specific
enzymes into the bloodstream.
•Elevated enzyme levels are oftenindicators of tissue problems, and
are used in the diagnosis of diseases.
•Enzyme activities in the body fluids are altered by pathological
processes so, its measurement is used for disease investigation.
INTRODUCTION

3. •All known enzymes are proteins.
•They are high molecular weight compounds made up principally
of chains of amino acids linked together by peptide bonds.
•Enzymes can be denatured and precipitated with salts, solvents
and other reagents.
•They have molecular weights ranging from 10,000 to 2,000,000.
•Many enzymes require the presence of other compounds -
cofactors -before their catalytic activity can be exerted.
•This entire active complexis referred to as the HOLOENZYME ;
i.e., APOENZYME (protein portion) plus the COFACTOR
(coenzyme, prosthetic group or metal-ion-activator) .
Chemical Nature of Enzymes

4. Apoenzyme+ Cofactor= Holoenzyme
The Cofactormay be:
1 •A coenzyme-a non-protein organic substance
which is dialyzable, thermostable and loosely
attached to the protein part.
2 •A prosthetic group-an organic substance which
is dialyzable and thermostable which is firmly
attached to the protein or apoenzyme portion.
3 •A metal-ion-activator-these include K
+
, Fe
++
,
Fe
+++
, Cu
++
, Co
++
, Zn
++
, Mn
++
, Mg
++
, Ca
++
, and
Mo
+++
.
Chemical Nature of Enzymes ……contd.

6. Enzymes can be classified by the kind of chemical reaction
catalyzed.
A.Addition or removal of water :
1.Hydrolases-these include esterases, carbohydrases, nucleases,
deaminases, amidases, and proteases
2. Hydrases such as fumarase, enolase, aconitaseand carbonic
anhydrase
B.Transfer of electrons :
1.Oxidases
2.Dehydrogenases
Classification of Enzymes :

7. Classification of Enzymes …… contd.
C. Transfer of a radical:
1. Transglycosidases-of monosaccharides
2. Transphosphorylasesand phosphomutases-of a phosphate
group
3. Transaminases -of amino group
4. Transmethylases-of a methyl group
5. Transacetylases-of an acetyl group
D.Splitting or forming a C-C bond:
1. Desmolases
2. Changing geometry or structure of a molecule
3. Isomerases
E. Joining two molecules through hydrolysis of
pyrophosphate bond in ATP or other tri-phosphate
1. Ligases

8. •One of the properties of enzymes that makes them so
important as diagnostic and research tools is the specificity
they exhibit relative to the reactions they catalyze.
•Greater specificity is achieved in three ways:
1.Interpreting investigations in the light of clinical features
2.Test pattern recognition
3.Isoenzyme determination:
-ASTmay be due toMIorHepatitis so, itmakes
confusion in diagnosis to be confirmedby LDH levels.
-ALP inCholestasis& bone diseases:
-Differentiatedbybilirubin & transaminase levels in
Cholestasis .
-Confirmed by GGT in Cholestasis.
Specificity of Enzymes :

9. ISOENZYMES
•Catalyze the same reaction
•Two or more polypeptide chains
•Different polypeptide chains are products of
different genes
•Differ in AA sequence and physical properties
•May be separable on the basis of charge
•Are tissue specific

10. Aminotransferases:
•ALT
•AST
•GGT
ALT and AST:
1.Pyridoxal dependent
2.Indicates Hepatitis, Myocardial infarction:
-Elevations take > 4 hours to develop, last 4 days
-ALT elevation lasts longer than AST
-AST elevations are higher than ALT
-Elevations occur in most definite infarctions
3.Skeletal muscle damage
4.Hemolysis
5.Pancreatitis
Isoenzymes and multiple forms

11. GGT
•Hepatobiliary enzyme -highly inducible in 75% of the
population
•Highest increases in intra or posthepatic biliary
obstruction
•Higher and more persistent increases than AlP (Alkaline
Phosphatase)
Isoenzymes and multiple forms

14. Hepatocytedamage
changes in plasma enzyme activity generally indicate liver cell membrane damage
Aminotransferases
aspartate
aminotransferase (AST)
alanine aminotransferase
(ALT),
A rise in plasma aminotransferase
activities is a sensitive indicator
of damage to cytoplasmic and/or
mitochondrial membranes.

15. Raised plasma transaminase
concentrationsareindicativeof
hepatocytedamage,
Ininflammatoryorinfectiveconditions,
suchasviralhepatitis,thecytoplasmic
membranesustainsthemaindamage;
leakageofcytoplasmiccontentscausesa
relativelygreaterincreaseinplasma
ALTthanASTactivities.

16. •AST(SGOT)–foundinliver>cardiac
muscle>skeletalmuscle>kidneys
>brain
•ALT(SGPT)–foundprimarilyin
liver
•Normallypresentinseruminlow
concentration(0-40IU/L)
•Whenthereisdamagetolivercell
membrane–increasedpermeability
andsoincreasedserumconcentration
•alsopresentinredbloodcells,
andcardiacandskeletal
muscleandisthereforenot
specifictotheliver.

17. TranminaseLevels of >1000 IU/L occurs in –
•Acute viral hepatitis
•Toxin and drug induced hepatitis
•Ischaemicliver injury

18. LDH-Lactate Dehydrogenase in MI
•LDHis a tetramer of two non-identical subunits (LDH5(M4),
LDH4(M3H), LDH3(M2H2), LDH2(MH3), LDH1(H4)

19. •Acid phosphatase (ACP)
•Amylase (AMS)
•Alanine aminotransferase (ALT)
•Alkaline phosphatase (ALP)
•Aspartate aminotransferase (AST)
•Creatine kinase (CK)
•Gamma-glutamyltransferase (GGT)
•Glucose 6-phosphate dehydrogenase (G6PD)
•Lactate dehydrogenase (LDH or LD)
•Lipase (LPS)
ENZYMES OF CLINICAL INTEREST

20. Diagnostically Important Enzymes 1/3
Principle Clinical
Applications
Principal Sources Enzyme
Carcinoma of prostateProstate, erythrocytesAcid
Phosphatase (ACP)
Hepatic parenchymal
disease
Liver, Skeletal muscle,
Heart
Alanine
aminotransferase(ALT)
Bone diseases,
hepatobiliary diseases
Liver, bone, intestinal
mucosa, placenta,
kidney
Alkaline
Phosphatase (ALP)
Pancreatic diseasesSalivary glands,
pancreas, ovaries
Amylase(AMS)
Myocardial infarction,
hepatic parenchymal
disease, muscle disease
Liver, skeletal muscle,
heart, kidney,
erythrocytes
Aspartate
aminotransferase(AST)

21. Diagnostically Important Enzymes 2/3
Principle Clinical
Applications
Principal Sources Enzyme
Myocardial
infarction, muscle
diseases
Skeletal muscle,
brain, heart, smooth
muscle
Creatinekinase(CK)
Hepatic parenchymal
disease
LiverGlutamate
dehydrogenase

22. Diagnostically Important Enzymes 3/3
Principle Clinical
Applications
Principal Sources Enzyme
Myocardial
infarction, hemolysis,
hepatic parenchymal
disease
Heart, liver, skeletal
muscle, erythrocytes,
platelets, lymph
nodes
Lactate
dehydrogenase(LDH)
Pancreatic diseasesPancreasTrypsin(ogen)

23. Alanine aminotransferase(ALT)
•Widely distributed, although the largest amounts
found in the liver.
•Smaller amounts occur in the heart but usually
remains normal after MI .
•More specific for liver disease than AST.

24. •Widely distributed, high concentrations in intestines,
liver, bone, spleen, placenta and kidney.
•The main sources of serum ALP are the hepatobiliary
tree and bone disorders.
•Elevated levelsduring healing of fractures , active
growth and during the 3
rd
trimester of pregnancy.
•serum ALP activity in liver disease is mainly due to
Cholestasis.
Alkaline phosphatase (ALP)

25. Alkaline phosphatase (ALP)
Causes of increased serum alkaline phosphatase enzyme activity:
Physiological :
Bone disease:
Hepatobiliary disease:
-Infancy
-Puberty
-Pregnancy
-Osteomalacia, rickets
-Osteomyelitis
-Hepatitis
-Cholestasis
-Cirrhosis

26. •This enzyme is widely distributed in the body.
•Main sources: Heart, liver, skeletal muscle, and kidney.
•Useful in the diagnosis of MI, liver disorders and muscle damage.
•Causes of serum AST levels:
•Physiological: Neonates.
•Liver diseases: Hepatitis, hepatic necrosis, cholestasis
•Cardiac disease:Myocardial Infarction.
•Diseases of skeletal muscle:Crush injury,trauma,myopathy
•From Erythrocytes:Hemolysis
Aspartate aminotransferase (AST)

27. •Creatine kinase is associated with ATP regeneration in
muscle and nervous tissue.
•Elevated blood levels of CK are used as indicators of MI,
muscular dystrophy, and stroke.
•CK occurs as a dimer of 2 different subunits, M and B.
-CK-BB: Brain type.
-CK-MB: Hybrid type.
-CK-MM: Muscle type.
•These can be separated by electrophoresis.
•CK-MB is released from cardiac muscle cells after MI.
Creatine kinase (CK)

28. •A microsomal enzyme its synthesis induced by ethanol
and anticonvulsant drugs.
•Found mainly in the kidney and significant amounts in
liver, brain, prostate, and pancreas.
•Used primarily for diagnosis of hepatobiliary problems .
•ALT, AST and GGT are the main liver function tests .
•Marked elevation of serum GGT level is seen in
alcoholic liver disease.
Gamma-glutamyltransferase (GGT)

29. •Converts pyruvate to lactate (and vice versa) during and
after anaerobic metabolism.
•LDHoccurs as a tetramerof 2 different subunits:
LD-1 (HHHH)from the heart:
Elevated after MI.
LD-2 (HHHM)from the kidney:
Elevated after renalinfarction.
LD-3 (HHMM)from the lung, spleen and pancreas:
Elevated in pulmonary embolism.
LD-4 (HMMM)and LD-5 (MMMM), both from the liver
and skeletal muscle:
Elevated in injury to liver or skeletal muscle.
Lactate dehydrogenase (LDH or LD)

30. Control
LDH Isoenzymes
1 2 3 4 5

31. Myocardial Infarction ( MI )
•CK from skeletal muscle may be following intramuscular
injection,
•CKspecificity is by measuring CK-MB.
•Cardiac enzyme measurements are very sensitive indicators of
MI because it is in over 95% of cases.

32. Measurement of serum enzyme activities for :
a -Differential Diagnosis of Jaundice.
b -Monitoring of drug toxicity.
•ALTis more specificthan AST.
•Hepatocellular disease has only modest effect on ALP &
GGT (up to 3 times the upper limit of normal)
•In Cholestasis, Higher values of ALP & GGT due to
synthesis ( the values are 5-10 times the upper normal
level) .
Liver Enzymes( ALT, AST, GGT, ALP, LDH)

33. •ALPenzyme is usually normalin Osteoporosis
ماظعلا ةشاشهas osteoblastic activity is not increased
•Modestof ALPin Osteomalacia ماظعلا نيلتvand
Ricketsحاسكلا
•Healing fracturesTransient of ALP
•1
ry
& 2
ry
bone tumorsof ALP (5 times normal)
Bone Enzymes -( Alkaline Phosphatase)ALP

34. A55-year-oldfemalepatientpresentstothe
emergencydepartmentwithchestpainand
shortnessofbreath.Thepatient'sphysician
suspectsapossiblemyocardialinfarction(MI)
A 50-year-old male patient presents with
jaundice and pruritus. Laboratory tests indicate
liver dysfunction, including elevated bilirubin
levels.

36. Case Study 1: Liver Enzymes in Hepatocellular Injury
A 40-year-old male patient presents with fatigue, abdominal pain, and jaundice.
Laboratory tests reveal elevated liver enzymes, including ALT, AST, and ALP. The
patient's physician suspects hepatocellular injury as the underlying cause.
1.Which liver enzyme is more specific for hepatocellular injury?
1.ALT (Alanine aminotransferase)
2.What are the normal functions of ALT and AST in the liver?
1.ALT: Primarily found in the liver, it is involved in amino acid metabolism.
2.AST: Found in various tissues, including the liver, it plays a role in energy
metabolism.
3.What does an elevation in ALP levels indicate in this case?
1.An elevation in alkaline phosphatase (ALP) levels suggests cholestasis or
biliary obstruction.

37. Case Study 2: Cardiac Enzymes in Myocardial Infarction (MI)
A 55-year-old female patient presents to the emergency
department with chest pain and shortness of breath. The patient's
physician suspects a possible myocardial infarction (MI) and
orders cardiac enzyme measurements.
Which enzyme is specifically measured to indicate cardiac
muscle damage?
Creatine kinase (CK) is measured to indicate cardiac muscle
damage.
What are LDH1 and LDH5, and how are they related to cardiac
conditions?
LDH1 and LDH5 are different isoforms of lactate
dehydrogenase (LDH). LDH5 is predominantly found in
cardiac muscle, while LDH1 is found in red blood cells. An
elevated LDH5 level can indicate cardiac muscle damage.

38. Case Study 2: Cholestasis
A 50-year-old male patient presents with jaundice and pruritus.
Laboratory tests indicate liver dysfunction, including elevated
bilirubin levels.
3.How do ALP and gamma-glutamyl transferase (GGT) levels
change in cholestasis?
3.In cholestasis, higher values of ALP and GGT are observed
due to an increase in their synthesis. The values can be 5-
10 times the upper normal level.
4.What is the main reason for the elevated ALP activity in liver
disease associated with cholestasis?
3.The main reason for the elevated ALP activity in liver
disease associated with cholestasis is the impaired flow of
bile.

39. Case Study 1: Bone Disorder -Osteoporosis
A 65-year-old postmenopausal female patient presents with chronic
back pain and a history of recurrent fractures. Laboratory tests reveal
abnormal bone markers.
1.What is the typical effect of osteoporosis on alkaline phosphatase
(ALP) levels?
1.ALP enzyme is usually normal in osteoporosis as osteoblastic
activity is not increased.
2.How is ALP affected in osteomalacia and rickets?
1.Modest elevation of ALP can be observed in both osteomalacia
and rickets.