Clinical Biochemistry of Domestic Animals (Sixth Edition) - UMK ...

III. Factors Affecting Serum Enzyme Activity
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diseased cells that remain viable, perhaps through membrane
pores or tears. However, it is difficult to envision that a cell
could develop a pore or tear large enough to allow “ leakage
” of macromolecules such as enzymes while maintaining
the intracellular-to-extracellular electrolyte ratios necessary
to remain viable. An alternative mechanism by which
a cell could sustain damage from which it survives and yet
allows the release of cytoplasmic enzymes is the formation
of membrane blebs ( Coltran et al. , 1999 ; Gores et al. , 1990 ;
Lemasters et al. , 1983 ; Mair, 1999 ). These blebs then are
ruptured or are released as vesicles into the blood where they
are eventually broken down, releasing their contents, including
cytoplasmic enzymes. The body of knowledge supporting
this concept has been growing since the 1980s (Gore
et al. , 1990; Kristensen, 1994 ; Mair, 1999 ; Solter, 2005 ).
Cell membrane bleb formation has been recognized following
hypoxic insults and likely reflects two sequential
developments. Depletion in energy stores in the form of ATP
is followed by several events including the influx of calcium
into the cell ( Coltran et al. , 1999 ). This calcium influx results
in activation of intracellular phospholipases, endonucleases,
and proteases and ultimately in disruption in the phosphorylation
state of cytoskeletal proteins and an alteration in lipid
membrane content. A combination of the altered cytoskeletal
proteins, lipid membrane content, and osmotic swelling
of the cell leads to bleb formation, release of these blebs,
and resealing of the cell membrane ( Fig. 12-1 ). Hepatocyte
bleb formation, projection of these blebs through the fenestrations
of endothelial cells, and release of these blebs
during hypoxia are clearly shown in scanning electron
micrographs ( Lemasters et al. , 1983 ). Bleb formation
has been described with many conditions including ischemia,
shock, viral infections, toxemia, and cholestasis.
The magnitude of serum enzyme increase with reversible
cell injury and bleb formation is not clearly understood,
but it is likely that the magnitude of release and resultant
increase activity in serum are considerably less than what
might be observed with cell necrosis. Hence, it is reasonable
to assume that the greater the magnitude of serum
enzyme increase, the greater likelihood of some irreversible
cell death, whereas mild serum enzyme activity increases
may be associated with reversible cell injury.
Although cytoplasmic enzymes may be released from
cells into blood because of bleb formation, enzymes
associated with the mitochondria are not released by this
mechanism ( Kamiike et al. , 1989 ). Appreciable loss of
membrane integrity and presumably cell death are necessary
for release of mitochondrial aspartic aminotransferase
(mAST) from hepatocytes. Ischemic liver does not lose
mAST until almost all cytoplasmic aspartic aminotransferase
(cAST) is lost.
Regardless of the mechanism of release of the enzymes,
there is evidence that the enzymes are released into the
interstitial space where the greater portion is carried by
lymphatics to the thoracic duct and emptied into the blood
ALT
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SDH
AST
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SDH
AST
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Sinusoid
SDH
AST
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FIGURE 12-1 Membrane bleb formation in reversible injury, allowing
the release of cytoplasmic enzymes either directly into blood or into the
interstitial space where they can be carried by lymphatics to blood.
( Bolter and Critz, 1976 ; Lindena et al. , 1986 ). Lymph-toserum
ratios of most enzymes are greater than 1:1, providing
support for the delivery of enzymes to blood via the
lymphatics. However, the direct delivery of the enzyme
from the injured cell to blood cannot be discounted and is
supported by the electron micrographs of cell blebs extending
through the fenestrations of the endothelium as discussed
earlier. This delivery of the enzyme from the injured
cell to blood, whether directly or indirectly via lymphatics,
likely affects the time of maximum serum increase of
the enzyme after injury and duration of the presence of the
increase of the enzyme in blood as suggested by the longer
half-life of CK when injected intramuscularly as opposed
to intravenous injection ( Aktas et al. , 1995 ).
D . Mechanisms of Release of Membrane-
Bound Enzymes
Those enzymes attached to the external surface of cell
membranes such as alkaline phosphatase (ALP), gammaglutamyl
transferase (GGT), and 5 nucleotidase (5 N) are
released from cells to blood by distinctly different mechanisms
than enzymes derived from the cytoplasm.
Of historical interest, increases of serum ALP activity
were perhaps first thought to result from a failure of excretion
of bone ALP by the liver. This concept was put to rest
many years ago and followed by the concept that cholephilic
enzymes were shed from the bile canalicular surface
of hepatocytes or biliary epithelial cells into bile and then
regurgitated into blood through tight junctions. Support for