Enzymes in health and diseas Flashcards
Energy depletion as a mechanism leading to the release of cytosolic enzymes:
Reduce ATP –>efflux of K+ and influx of Na+ (inhibition of Na-K ATPase –> cell swelling –> reversible damage –> over time: irreversible damage –> influx of calcium and formation of free radicals
Effects of energy depletion
Stretching (due to swelling),
Degradation of phospholipids,
Decomposition of the cytoskeleton
Generalized septicemia
Irreversible membrane damage causing massive increases in serum enzyme activities
Cell necrosis causes
liberation of organelle enzymes
Presence of enzymes in serum is evidence for
severe cell injury
Enzymes released from intra–>extracellular space frist appear in…
Interstitial fluid
Enzymes released from blood/endothelial cells first appear in
Intravascular space
Pathway from interstitial fluid –>blood
through LYMPH** OR capillary wall (depends on permeability) ** important part of movement from intracellular–> intravascular space. Not always immediately present in blood as lymphatic flow is slow
Mechanisms of enzyme elimination from the blood (2):
- Renal excretion (not of general importance as most enzymes are larger than this method supports [ex. ~70-160 kDa])
- Uptake by cells that degrade enzymes
Kidney excretion is available for enzymes w/ mw lower than:
60 kDa
Mechanism for degradation of enzymes among cells:
Receptor mediated endocytosis
Myocardial infarction
Heart attack, occlusion of a coronary artery
MI occurs as a result of… (2):
lack of O2 delivery to the myocardial tissue
lack of removal of metabolites
MI is influenced by:
severity of ischemia (inadequate blood supply),
age,
sex
First few seconds of MI
Anoxia –> depletion of energy stores –> OxPhos shutdown –> Anaerobic respiration –> leakage of intracellular electrolytes (espec. K+)
First few minutes (reversible damage)
Ischemia –> impaired clearnce of metabolites (ex. inorganic P, lactate, adenosine, H+)
Irreversible damage associated with MI:
Release of macromolecules
Activation of lipoprotein lipases
Loss of mitochondrial control
Spillage of mitochondrial components
Fate of released cellular components
Ions/metabolites: intracellular space –> interstitial space –> circulation
Macromolecules: delayed clearance as they are released only through irreversible injury and are large and require LYMPHATIC DRAINAGE
CK equilibrium reaction
Creatine + ATP Creatine phosphate + ADP
CK
important in diagnosis of MI, two subunits either M or B type producing 3 different isoenzymes: BB, MB, and MM
Stats on CK
High levels in neonate, fall w/in first few wks of life
Total CK is elevated in MI and injury
Males have higher values than females
Activities of CK increase w/ age + body weight
Injuries elevating CK
Skeletal muscle injury and disease IM injections Hypothyroidism Generalized convulsions Cerebral injury Infectious disease Prolonged hypothermia
CK: MB (CK-2)
has advantages over other CK isoenzymes, found in trace concentrations in skeletal muscles and higher in the myocardium
CK nomenclature
due to electrophoretic mobility, most anodal receiving the lowest number
CK: MM (CK-3)
found in skeletal muscle and myocardium
CK: BB (CK-1):
found in the brain and gastrointestinal tract
Method of measurement
Antibody based CK-MB mass assays (ug/L)
Double monoclonal antibody-based sandwich type assay
Lactate dehydrogenase (LD)
responsible for catalysing the reversible reduction of pyruvate to lactate
NADH + pyruvate lactate + NAD
late marker of MI as it stays in the blood longer
Important in glycolysis when O2 is limiting
widespread tissue distribution (NONSPECIFICITY)
Individuals who exercise heavily have…
high LD
LD results for MI are obscured by…
Hemolysis, due to its presence in red cells
The heart contains high amounts of
LD1
Myoglobin and Troponin
cardiac proteins of diagnostic value in MI
Myoglobin
found in skeletal muscle and heart
does not exist as isoenzymes
low specificity
small size
Troponin
3 subunits: C, I, and T
Sequence unique to the cardiac muscle (high specificity)
assays have monoclonal antibodies binding to the most stable region of the cTnI molecule
exists in many forms in the blood
Biochemical markers in ischemic injury
Initial lag phase… and then:
CK: elevates after 20 hours and decreases rapidly (specifically MB), not a good early marker (NOT A GOOD TEST)
Myoglobin: spike 10h after onset and rapid decrease (NOT USED)
TroponinI: longer response time, elevates after 20 hours and stays elevated (GOLD STANDARD FOR MI). Stay elevated. Sensitivity higher, specificity higher.
LD, LD-1: extremely useful late marker due to prolonged half life in serum. LD-1 is better specificity than LD, sensitivity similar in both. A ratio >1 is indicative of MI in LD-1>LD-2. Hemolysis gives a flipped pattern of this (NOT USED ANYMORE)
High sensitivity troponins
Usually hsTnT
Can measure levels in almost 100% of healthy individuals reliably
10x lower detection limit
Early diagnosis
Problem: number of false positives increases as we move decision threshold to the left, causes other than MI
Rising/falling pattern is critical to distinguish between non-cardiac conditions and acute cardiac conditions
Universal definition of MI
a rise and or fall in cTn in patients w/ evidence of myocardial ischemia with at least one cTn value above the 99th percentile of reference group
Pancreatic enzymes of diagnostic value (2):
Amylase
Lipase
Acute pancreatitis
inflammatory disease of the pancreas which when the attack is resolved permits restoration of normal function
Causes: alcohol, biliary tract disease, post-surgery, and sever hypertriglycerdemia
Symptom: acute abdominal pain, hypotension, respiratory failure, hyperglycemia, hypocalcemia, hypoalbuminemia
Amylase
highest concentration in saliva and pancreas
secreted into the duodenum –> hydrolysis of macromolecular carbohydrates
Two types: pancreatic type and salivary type, present in about equal concentrations in normal serum and urine
low mw
The only serum enzyme normally found in urine
isoenzyme p-amylase has high sensitivity and specificity
Lipase
hydrolysing triglycerides
pancreatic isoenzyme interacts with long chain triglycerides
colipase is required for full activity (coenzyme)
Rises to a higher extent than amylase
sensitivity and specificity are higher than amylase
Trypsin
20-30x more costly than amylase/lipase
similar results to same
not routinely used
