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Determination of enzyme activities in serum for the detection of xenobiotic effects on the liver
Exp. Pathol. 1990; 39: 157-164 Gustav Fischer Verlag Jena
Department of Clinical Chemistry and Laboratory Medicine, Friedrich Schiller University lena, GDR (Head: Prof. Dr. sc. nat. E. KEIL)
Determination of enzyme activities in semm for the detection of xenobiotic effects on the liver) By E. KEIL With 5 figures Address for correspondence: Prof. Dr. E. KEIL, Department of Clinical Chemistry and Laboratory Medicine, BachstraBe 18, DDR - 6900 lena Key words: xenobiotic effects; liver; enzyme patterns; serum
Summary
The determinations of enzyme actIvltles in the serum are of considerable importance in detecting xenobiotic effects on the liver. After a brief introduction to the basics of enzyme diagnostics, the enzymes ALAT, ASAT, ICDH, LDH, SDH, GLDH, AP, y-GT, CHE are characterized with regard to their occurrence, their half-life periods in the serum and their clinical value. They are followed by enzyme levels and the presentation of the dynamics of enzyme activities in the serum after xenobiotic influences on the liver in humans.
Introduction The determination of enzyme activities in the serum or plasma is of major importance to diagnosis, clinical course and prognosis of diseases as well as to the control of therapy. Diseases of the heart, the pancreas, the skeletal muscles and, particularly, the liver have to be considered the domains of enzyme diagnostics. Enzyme determinations account for 10-15 % of laboratory tests. The development of diagnostic enzymology begins with the detection of increased activities of diastase (
SEIDEL
on the occasion of his 75th birthday Exp. Pathol. 39 (1990) 3-4
157
in the amount of enzyme activities as well as in the interrelations of the main chain enzymes; in the content of enzymes specific for the organ and, with respect to the functional heterogeneity of the liver, also for certain cell groups; in the distribution of the isoenzymes (11). In the cells each enzyme is differently localized (see table 1).
Table 1. Enzyme localization in the liver cell (modified from HASCHEN, R.: Enzymdiagnostik, 1981, p. 23); (5).
Cytosol
Bile canaliculi
Aldolase Alanine aminotransferase Aspartate aminotransferase Arginase Isocitrate dehydrogenase Lactate dehydrogenase Leucine Aminopeptidase Ornithine carbamyltransferase Sorbitol dehydrogenase
Alanine aminopeptidase Alkaline phosphatase y-Glutamyltransferase 5' -Nucleotidase
Mitochondria Alanine aminotransferase Aspartate aminotransferase Glutamate dehydrogenase
Ribosomes Cholinesterase
Lysosomes ~-Glucuronidase
Acid phosphatase
Between the intracellular space with high enzyme activities and the extracellular space with physiologically low activities, there is a great concentration gradient which is maintained by permanent expenditure of energy. Normally, the individual levels of cellular enzymes in blood plasma are, apart from changes in childhood and sometimes also in older age, fairly constant over years. The levels of enzymes are regulated by a complex system of flux equilibrium between the intracellular, interstitial and intravascular spaces (3). In enzyme diagnostics, it is postulated that in case of reversible or irreversible disorders of cellular integrity, no matter what etiology, the enzymes leave the intracellular space and, on an organ-dependent pathway (directly, lymph, interstitial space), enter the blood at varying speed. From the enzyme activities and enzyme patterns detected in the blood as well as from their dynamics, conclusions can be drawn about the damaged organ, the intracellular localization as well as the extent and severity of the damage. Here, it must be taken into consideration that only under ideal conditions a "copy" of the enzyme pattern of the damaged cell can be found in the blood. As causes for occurring distortions and overJappings of blood enzyme patterns typical of organs and cells, we have to take into account the partial inactivation of the enzymes as well as their absorption and degradation after entry into the extracellular space, perhaps already in the intracellular space; the differing half-life periods and distribution quotients of each enzyme in the extracellular compartments; the varying intracellular localization of the enzymes - in the presence of slight cell damage such as increase in permeability of the cell membrane, the cell is left predominantly by the enzymes localized in the cytoplasma and only in case of severe cell damage (necrosis) also by the enzymes localized in the organelles; the involvement of other organs in the disease process (2, 5, 11). 158
Exp. Pathol. 39 (1990) 3-4
In the following, we will briefly characterize some enzymes that are utilized for the detection of xenobiotic effects on the liver. The order of organs chosen under "Occurrence" corresponds to the level of enzyme activities in decreasing direction. The "EC data" are in accordance with the Enzyme Nomenclature 1978 (1).
Enzymes in the cytosol
Alanine aminotransferase (ALA T), EC: 2.6. 1. 2 tl2 in serum: 47 ± 10 h. Occurrence: liver, skeletal muscles, heart, kidney, pancreas, erythrocytes; cytoplasmic (85 %) and mitochondrial (15 %) localization. Clinical significance: Owing to its high activity in the liver, ALAT is largely regarded as liverspecific and predominantly used to diagnose liver diseases and to follow their course. The enzyme is considered a sensitive indicator for demonstrating disturbances of the permeability in the cell of the liver parenchyma. Elevated activities in the serum are found, e.g., in acute and chronic hepatitides, cirrhoses, intoxications, congested liver due to right ventricular failure (4, 5, 8, 11).
Aspartate aminostransferase (ASAT), EC: 2.6.l.1 tl2 in serum: 17 ± 5 h. Occurrence: liver, heart, skeletal muscle, brain, kidney, pancreas, lung, erythrocytes; localization in the mitochondria (80 % m-ASAT -isoenzyme) and in cytosol (20 %, cASAT -isoenzyme). Clinical significance: Preferential employment in the diagnosis of heart and liver diseases. Elevated activities in the serum, for instance, in myocardial and pulmonary infarction, liver diseases, haemolysis, myopathies (4, 5, 8, 11).
Isocitrate dehydrogenase (NADP+) (ICDH), EC: 1.1.1.42 tl2 in serum: 60 min. Occurrence: ubiquitous, e.g., liver, skeletal muscles, kidney; localization in the cytoplasma and in the mitochondria. Detection of isoenzymes. Clinical significance: The determination of cytoplasmic ICDH is chiefly employed in the diagnosis ofliver diseases. High activities in the serum are notably to be expected in the acute phase in hepatitides and following intoxications. Due to the short half-life period ofthe enzyme, slight and transient rises in serum activity can hardly be registered (4, 5).
Lactate dehydrogenase (LDH), EC: l.1.l.27 tl2 in serum: LDH j isoenzyme 113 ± 60 h; LDH2 isoenzyme 10 ± 2 h. Occurrence: ubiquitous, e.g., liver, skeletal muscles, heart, kidney, pancreas, erythrocytes; identification of 5 isoenzymes, distinct isoenzyme pattern in the organs. Clinical significance: The determination of serum enzyme activity is mainly performed to diagnose and control liver , blood and muscle diseases, in myocardial infarction as well as in tumour diagnosis. High activities are to be expected in intoxications (carbon tetrachloride, death-head) (5, 6,8,11).
Sorbitol dehydrogenase (SDH), EC: 1.1.l.l4 Occurrence: liver, kidney, cardiac and skeletal muscles. Clinical significance: Depending on its distribution, SDH is regarded as liver-specific. The low stability of the enzyme limits its diagnostic applicability. Raised serum activities are found in the acute phase, for example, in liver diseases and shock (haemorrhagic, cardiogenic) (5).
Enzymes in the mitochondria ALAT and ASAT (see above); escape from the mitochondria in severe liver damage. Exp. Pathol. 39 (1990) 3-4
159
Glutamate dehydrogenase (GLDH), EC: 1.4.1.3
± 1 h. Occurrence: liver, kidney, brain, lung, cardiac and skeletal muscles. Clinical significance: Due to the high enzyme activity in the liver as compared to other organs, GLDH is considered relatively liver-specific. Owing to its exclusive localization in the mitochondria, a rise in the serum activity is to be expected only in severe liver cell damage. Increased activities are found, for instance, in liver diseases (acute and chronic hepatitis, cirrhosis, metastatic liver, obstructive jaundice, intoxications) (5, 6, 11). tl2 in serum: 18
Enzymes in the bile canaliculi Alkaline phosphatase (AP), EC: 3.1.3.1
tl2 in serum: placental AP 3-4 d, biliary AP 9.7 ± 1 d. Occurrence: small bowel, bone, liver, kidney, placenta. Isoenzymes: intestinal, placental, hepatic, renal, bone AP (posttranscriptional forms?). Clinical significance: Elevated activities in the serum are observed, for example, in processes of bone remodelling and absorption as well as in hepatobiliary processes (intra- and extrahepatic occlusions of the bile duct). Drugs may lead to increased (e.g. antiepileptics, hypnotics, peroral antidiabetics) and decreased activities (antilipaemics, peroral contraceptives) (4, 5, 8, 11). y-Glutamyl transferase (y-GT), EC: 2.3.2.2
tl2 in serum: 4.1 d (3). Occurrence: kidney (brush border of the proximal tubules), pancreas, liver (epithelial cells of the bile duct); several multiple forms identified. Clinical significance: The activity of y-GT in serum is almost exclusively determined by the liver. The enzyme is considered a sensitive indicator of a hepatobiliary disease (cholestasisindicating enzyme). Raised activities in the serum after impact of xenobiotics are chiefly based on an induction of enzyme synthesis (4, 5, 8, 12). Ribosomal enzymes Cholinesterase (CHE), pseudocholinesterase (PCHE), EC: 3.1.1.8
tl2 in serum: about 10 d. Occurrence: liver, pancreas; nomerous genetically-induced variants. Clinically significance: The enzyme is synthesized in the liver and, therefore, mainly serves to examine the synthetic performance of the liver or to establish the extent of a liver cell failure. Increased serum activities, induced by an elevated enzyme synthesis, are found in case of chronic loss of protein by the kidney and the intestine, decreased values in hepatopathies (e.g. chronic hepatitis, liver cirrhosis, liver malignomas). Lowered activities in the serum caused by xenobiotic effects can be due to a restriction of biosynthesis or to an inhibition of the enzyme (e.g. organic phosphoric-acid esters, chlorinated hydrocarbons) (4, 5, 8).
Table 2. Relative degree of increase of several serum enzymes in toxic hepatic injury (from ZIMMERMANN, H. 1., 1982) (15). Degree of increase of serum enzymes
Lesion Toxin
Necrosis
Steatosis
AS AT
ALAT
ICDH MDH
GLDH
CCL4 Thioacetamide Tetracycline Ethionine
+ +
+
4+ 4+ 2+ 1+
3+ 3+
4+ 4+ 2+
2+ 2+
160
Exp. Pathol. 39 (1990) 3-4
+ +
1+
1
conlracept i ves
acetylsalicyliC acid
U/l
U/t CHE
CHE
5000
2000 1000 ASAT AlAr
.sao
AP
0
Q AP
1GT~
200 100
ALAT
so
]~ T~
GLOH
lO
-to 5
GLDH
Z
~
~
5000
2000 1000
SOO 200 100
so 20 -10
5
2.
Fig.l. Enzyme patterns in serum in drug-induced liver damage. D normal range I!ZZZJ intoxication (from E. and F. W.
SCHMIDT,
Kleine Enzym-Fibel, 1976, p. 31); (13).
U/L 3000
U/t
ALAT
Q
LDH
1000
CHE
300 AP
100 30 10
3
EJ
Q
3000
1000
300
100
30 10
3
1
Fig.2. Enzyme pattern in serum in halothane necrosis. Mean peak values, U/l, 25 °C, optimized methods. D normal range I!ZZZJ halothan-intoxication (from E. and F. W. 11
SCHMIDT,
1979); (10).
Exp. Pathol. 39 (1990) 3-4
161
Pot. B. J. ~ absent from home: i. Z. 3.
U/l
a:> •
'to 6 ' 5. 10 • 6. 5 ' 7. 20 ' 8. 11t
35
1.
Z5
't ooys
10 •
2..
3.
+ +
It.
5.
~ ~
~
6.
8.
7.
~
••
15
10
5
W 1971
ron
1972
normal
1910
Fig. 3. GLDH activities in serum in chronic pentachlorophenol intoxication, depending on the stay in the own apartment (wood impregnated with a preserving agent) (from SCHMIDT, E. and I. VIDO, 1976); (14).
There is a wide variety of changes in enzyme activities or enzyme patterns found in the serum after xenobiotic effects on the liver (table 2, figs. 1, 2, 3, 4). This is certainly connected with the fact that the xenobiotics have different intracellular sites of action and that, along with primary reactions, also secondary reactions lead to exit of enzyme out of the cell. Depending on their effect, the xenobiotics can be dichotomized into the obligatory hepatoxins, which produce a liver damage in every exposed person, and the facultative toxins which cause a damage solely in a part of the exposed. With regard to the liver diseases induced by xenobiotics, diverse morphological pictures may dominate, e.g., necrosis, hepatitis, cholestasis as well as the steatosis type (fig. 5). For the detection of liver and bile duct diseases, based on screening diagnostics, ALAT and y-GT and, for further differentiation of the disease process, ASAT, GLDH, AP, CHE are recommended, along with other laboratory examinations (7). According to SCHMIDT and SCHMIDT (10), there is suspicion of a liver damage caused by drugs or toxins: 162
Exp. Pathol. 39 (1990) 3-4
NB
'5- GT 17
GL DH
16
ALAr
15
ASAT CH~
1't
13
12
,
11
10
\
9
\
8 7
6 5
It
3 2 1
I\.
r~,
," I "
I :
fi
J
\.
\ \.
\'.. ', "-
".'.
\ .." '-.... ..... ... _'. '-...
\'--. -- ..... ...:::::-:::-....... ~ - - - - - - - - - - ....:..:.-=:::--:.:. ...
,----- ---- ------
=-=
::.:-- - - -
oL--,..,--,------r------.----,---- d 01 Z
15
8
2.3
Fig. 4. Dynamics of enzyme activities in serum in a man who painted a swimming pool for 3.5 h; composition of the paint and the solvent could not be ascertained. Initial values were known from analytical value a control examination. NB = --'--,.---,-normal value
Hepatotaxic slbstances
Carbcn tetrachloride Thioacetamide Phalloidin Amanitin Cyclophosphamide
Iproniazid Phenytoin Phenylbutazone Allopurinol Indomethacin
Chlorpromazine Imipramin Methylestosterone Ethinylestradiol Tolbutamide
Tetracycline Methotrexote Nitrosomine Azoserin Puromycin
Fig. 5. II *
Exp. Pathol. 39 (1990) 3-4
163
if y-GT is disproportionally high and three factors have been ruled out: alcohol, biliary obstruction and space-occupying processes in the liver; if y-GT is normal, but the rest of the enzyme pattern is pathological; if CHE activity falls rapidly or is solitarily low; if there are markedly pathological enzyme activities in serum but a normal histological picture of the liver. Finally, I would like to summarize that the determination of enzyme activities in the serum can be of great value for the detection of xenobiotic effects on the liver, but that maximal information can be gained only in association with the medical history, the clinical findings, with immunological, histological as well as other paraclinical investigations.
References 1. Enzyme Nomenclature. Recommendations (1978) of the Nomenclature Committee of the International Union of Biochemistry. Academic Press, New York-San Francisco-London 1979. 2. FRIEDEL, R.: Der Mechanismus des Ubertritts von Enzymen in das Blut. Med. unserer Zeit 1978; 2: 177-184. 3. - DIEDERICHS, F., LINDENA, J.: Release and extracellular turnover of cellular enzymes. In: Advances in Clinical Enzymology (eds. SCHMIDT, E., SCHMIDT, F. W., TRAUTSCHOLD, I., FRIEDEL, R.). S. Karger, BaselMiinchen-Paris-London-New York-Sydney 1979; pp. 70-105. 4. GRESSNER, A. M.: Leber und Gallenwege. In: GREILING, H., GRESSNER, A. M. (Hrsg.): Lehrbuch der Klinischen Chemie und Pathobiochemie. Schattauer, Stuttgart-New York 1987; pp. 421-507. 5. HASCHEN, R. 1.: Enzymdiagnostik. 2. Auflage, VEB Gustav Fischer Verlag, Jena 1981. 6. - Pathobiochemie der Leber. VEB Verlag Volk und Gesundheit, Berlin 1981. 7. IRRGANG, B., PATZOLD, K., THIELE, H.-J.: Labordiagnostisches Stufenprogramm zur gezielten Unterstiitzung der Diagnostik von Lebererkrankungen. Z. Klin. Med. 1987; 42: 735-740. 8. KELLER, H.: Klinisch-chemische Labordiagnostik fUr die Praxis. Georg Thieme Verlag, Stuttgart-New York 1986. 9. SCHMIDT, E., SCHMIDT, F. W.: Clinical enzymology. FEBS Lett. 1976; 62 (Supp!.): E62-E79. 10. - - Enzyme diagnosis in diseases of the liver and the biliary system. In: Advances in clinical enzymology (eds. SCHMIDT, E., SCHMIDT, F. W., TRAUTSCHOLD, I., FRIEDEL, R.). S. Karger, Basel-Miinchen-Paris-London-New York-Sydney 1979; pp. 239-292. 11. Enzym-Fibe!. Biochemica, Boehringer Mannheim GmbH, 1969. 12. - y-Glutamyl-transpeptidase. Dt. med. Wochenschr. 1973; 98: 1572-1578. 13. - Kleine Enzym-Fibe!. Diagnostics, Boehringer Mannheim GmbH, 1976. VIDO, I.: Exogen-toxische Leberschiiden. In: Toxische Leberschiiden (Medikamente und Leber) (Hrsg. 14. WANNAGAT, L.). Georg Thieme Verlag, Stuttgart 1976, S. 168. 15. ZIMMERMANN, H. J.: Chemical hepatic injury and its detection. In: Toxicology of the liver (eds. PLAA, G. L., HEWITT, W. R.). Raven Press, New York 1982, pp. 32. (Accepted January 17, 1990)
164
Exp. Pathol. 39 (1990) 3-4
Department of Clinical Chemistry and Laboratory Medicine, Friedrich Schiller University lena, GDR (Head: Prof. Dr. sc. nat. E. KEIL)
Determination of enzyme activities in semm for the detection of xenobiotic effects on the liver) By E. KEIL With 5 figures Address for correspondence: Prof. Dr. E. KEIL, Department of Clinical Chemistry and Laboratory Medicine, BachstraBe 18, DDR - 6900 lena Key words: xenobiotic effects; liver; enzyme patterns; serum
Summary
The determinations of enzyme actIvltles in the serum are of considerable importance in detecting xenobiotic effects on the liver. After a brief introduction to the basics of enzyme diagnostics, the enzymes ALAT, ASAT, ICDH, LDH, SDH, GLDH, AP, y-GT, CHE are characterized with regard to their occurrence, their half-life periods in the serum and their clinical value. They are followed by enzyme levels and the presentation of the dynamics of enzyme activities in the serum after xenobiotic influences on the liver in humans.
Introduction The determination of enzyme activities in the serum or plasma is of major importance to diagnosis, clinical course and prognosis of diseases as well as to the control of therapy. Diseases of the heart, the pancreas, the skeletal muscles and, particularly, the liver have to be considered the domains of enzyme diagnostics. Enzyme determinations account for 10-15 % of laboratory tests. The development of diagnostic enzymology begins with the detection of increased activities of diastase (
SEIDEL
on the occasion of his 75th birthday Exp. Pathol. 39 (1990) 3-4
157
in the amount of enzyme activities as well as in the interrelations of the main chain enzymes; in the content of enzymes specific for the organ and, with respect to the functional heterogeneity of the liver, also for certain cell groups; in the distribution of the isoenzymes (11). In the cells each enzyme is differently localized (see table 1).
Table 1. Enzyme localization in the liver cell (modified from HASCHEN, R.: Enzymdiagnostik, 1981, p. 23); (5).
Cytosol
Bile canaliculi
Aldolase Alanine aminotransferase Aspartate aminotransferase Arginase Isocitrate dehydrogenase Lactate dehydrogenase Leucine Aminopeptidase Ornithine carbamyltransferase Sorbitol dehydrogenase
Alanine aminopeptidase Alkaline phosphatase y-Glutamyltransferase 5' -Nucleotidase
Mitochondria Alanine aminotransferase Aspartate aminotransferase Glutamate dehydrogenase
Ribosomes Cholinesterase
Lysosomes ~-Glucuronidase
Acid phosphatase
Between the intracellular space with high enzyme activities and the extracellular space with physiologically low activities, there is a great concentration gradient which is maintained by permanent expenditure of energy. Normally, the individual levels of cellular enzymes in blood plasma are, apart from changes in childhood and sometimes also in older age, fairly constant over years. The levels of enzymes are regulated by a complex system of flux equilibrium between the intracellular, interstitial and intravascular spaces (3). In enzyme diagnostics, it is postulated that in case of reversible or irreversible disorders of cellular integrity, no matter what etiology, the enzymes leave the intracellular space and, on an organ-dependent pathway (directly, lymph, interstitial space), enter the blood at varying speed. From the enzyme activities and enzyme patterns detected in the blood as well as from their dynamics, conclusions can be drawn about the damaged organ, the intracellular localization as well as the extent and severity of the damage. Here, it must be taken into consideration that only under ideal conditions a "copy" of the enzyme pattern of the damaged cell can be found in the blood. As causes for occurring distortions and overJappings of blood enzyme patterns typical of organs and cells, we have to take into account the partial inactivation of the enzymes as well as their absorption and degradation after entry into the extracellular space, perhaps already in the intracellular space; the differing half-life periods and distribution quotients of each enzyme in the extracellular compartments; the varying intracellular localization of the enzymes - in the presence of slight cell damage such as increase in permeability of the cell membrane, the cell is left predominantly by the enzymes localized in the cytoplasma and only in case of severe cell damage (necrosis) also by the enzymes localized in the organelles; the involvement of other organs in the disease process (2, 5, 11). 158
Exp. Pathol. 39 (1990) 3-4
In the following, we will briefly characterize some enzymes that are utilized for the detection of xenobiotic effects on the liver. The order of organs chosen under "Occurrence" corresponds to the level of enzyme activities in decreasing direction. The "EC data" are in accordance with the Enzyme Nomenclature 1978 (1).
Enzymes in the cytosol
Alanine aminotransferase (ALA T), EC: 2.6. 1. 2 tl2 in serum: 47 ± 10 h. Occurrence: liver, skeletal muscles, heart, kidney, pancreas, erythrocytes; cytoplasmic (85 %) and mitochondrial (15 %) localization. Clinical significance: Owing to its high activity in the liver, ALAT is largely regarded as liverspecific and predominantly used to diagnose liver diseases and to follow their course. The enzyme is considered a sensitive indicator for demonstrating disturbances of the permeability in the cell of the liver parenchyma. Elevated activities in the serum are found, e.g., in acute and chronic hepatitides, cirrhoses, intoxications, congested liver due to right ventricular failure (4, 5, 8, 11).
Aspartate aminostransferase (ASAT), EC: 2.6.l.1 tl2 in serum: 17 ± 5 h. Occurrence: liver, heart, skeletal muscle, brain, kidney, pancreas, lung, erythrocytes; localization in the mitochondria (80 % m-ASAT -isoenzyme) and in cytosol (20 %, cASAT -isoenzyme). Clinical significance: Preferential employment in the diagnosis of heart and liver diseases. Elevated activities in the serum, for instance, in myocardial and pulmonary infarction, liver diseases, haemolysis, myopathies (4, 5, 8, 11).
Isocitrate dehydrogenase (NADP+) (ICDH), EC: 1.1.1.42 tl2 in serum: 60 min. Occurrence: ubiquitous, e.g., liver, skeletal muscles, kidney; localization in the cytoplasma and in the mitochondria. Detection of isoenzymes. Clinical significance: The determination of cytoplasmic ICDH is chiefly employed in the diagnosis ofliver diseases. High activities in the serum are notably to be expected in the acute phase in hepatitides and following intoxications. Due to the short half-life period ofthe enzyme, slight and transient rises in serum activity can hardly be registered (4, 5).
Lactate dehydrogenase (LDH), EC: l.1.l.27 tl2 in serum: LDH j isoenzyme 113 ± 60 h; LDH2 isoenzyme 10 ± 2 h. Occurrence: ubiquitous, e.g., liver, skeletal muscles, heart, kidney, pancreas, erythrocytes; identification of 5 isoenzymes, distinct isoenzyme pattern in the organs. Clinical significance: The determination of serum enzyme activity is mainly performed to diagnose and control liver , blood and muscle diseases, in myocardial infarction as well as in tumour diagnosis. High activities are to be expected in intoxications (carbon tetrachloride, death-head) (5, 6,8,11).
Sorbitol dehydrogenase (SDH), EC: 1.1.l.l4 Occurrence: liver, kidney, cardiac and skeletal muscles. Clinical significance: Depending on its distribution, SDH is regarded as liver-specific. The low stability of the enzyme limits its diagnostic applicability. Raised serum activities are found in the acute phase, for example, in liver diseases and shock (haemorrhagic, cardiogenic) (5).
Enzymes in the mitochondria ALAT and ASAT (see above); escape from the mitochondria in severe liver damage. Exp. Pathol. 39 (1990) 3-4
159
Glutamate dehydrogenase (GLDH), EC: 1.4.1.3
± 1 h. Occurrence: liver, kidney, brain, lung, cardiac and skeletal muscles. Clinical significance: Due to the high enzyme activity in the liver as compared to other organs, GLDH is considered relatively liver-specific. Owing to its exclusive localization in the mitochondria, a rise in the serum activity is to be expected only in severe liver cell damage. Increased activities are found, for instance, in liver diseases (acute and chronic hepatitis, cirrhosis, metastatic liver, obstructive jaundice, intoxications) (5, 6, 11). tl2 in serum: 18
Enzymes in the bile canaliculi Alkaline phosphatase (AP), EC: 3.1.3.1
tl2 in serum: placental AP 3-4 d, biliary AP 9.7 ± 1 d. Occurrence: small bowel, bone, liver, kidney, placenta. Isoenzymes: intestinal, placental, hepatic, renal, bone AP (posttranscriptional forms?). Clinical significance: Elevated activities in the serum are observed, for example, in processes of bone remodelling and absorption as well as in hepatobiliary processes (intra- and extrahepatic occlusions of the bile duct). Drugs may lead to increased (e.g. antiepileptics, hypnotics, peroral antidiabetics) and decreased activities (antilipaemics, peroral contraceptives) (4, 5, 8, 11). y-Glutamyl transferase (y-GT), EC: 2.3.2.2
tl2 in serum: 4.1 d (3). Occurrence: kidney (brush border of the proximal tubules), pancreas, liver (epithelial cells of the bile duct); several multiple forms identified. Clinical significance: The activity of y-GT in serum is almost exclusively determined by the liver. The enzyme is considered a sensitive indicator of a hepatobiliary disease (cholestasisindicating enzyme). Raised activities in the serum after impact of xenobiotics are chiefly based on an induction of enzyme synthesis (4, 5, 8, 12). Ribosomal enzymes Cholinesterase (CHE), pseudocholinesterase (PCHE), EC: 3.1.1.8
tl2 in serum: about 10 d. Occurrence: liver, pancreas; nomerous genetically-induced variants. Clinically significance: The enzyme is synthesized in the liver and, therefore, mainly serves to examine the synthetic performance of the liver or to establish the extent of a liver cell failure. Increased serum activities, induced by an elevated enzyme synthesis, are found in case of chronic loss of protein by the kidney and the intestine, decreased values in hepatopathies (e.g. chronic hepatitis, liver cirrhosis, liver malignomas). Lowered activities in the serum caused by xenobiotic effects can be due to a restriction of biosynthesis or to an inhibition of the enzyme (e.g. organic phosphoric-acid esters, chlorinated hydrocarbons) (4, 5, 8).
Table 2. Relative degree of increase of several serum enzymes in toxic hepatic injury (from ZIMMERMANN, H. 1., 1982) (15). Degree of increase of serum enzymes
Lesion Toxin
Necrosis
Steatosis
AS AT
ALAT
ICDH MDH
GLDH
CCL4 Thioacetamide Tetracycline Ethionine
+ +
+
4+ 4+ 2+ 1+
3+ 3+
4+ 4+ 2+
2+ 2+
160
Exp. Pathol. 39 (1990) 3-4
+ +
1+
1
conlracept i ves
acetylsalicyliC acid
U/l
U/t CHE
CHE
5000
2000 1000 ASAT AlAr
.sao
AP
0
Q AP
1GT~
200 100
ALAT
so
]~ T~
GLOH
lO
-to 5
GLDH
Z
~
~
5000
2000 1000
SOO 200 100
so 20 -10
5
2.
Fig.l. Enzyme patterns in serum in drug-induced liver damage. D normal range I!ZZZJ intoxication (from E. and F. W.
SCHMIDT,
Kleine Enzym-Fibel, 1976, p. 31); (13).
U/L 3000
U/t
ALAT
Q
LDH
1000
CHE
300 AP
100 30 10
3
EJ
Q
3000
1000
300
100
30 10
3
1
Fig.2. Enzyme pattern in serum in halothane necrosis. Mean peak values, U/l, 25 °C, optimized methods. D normal range I!ZZZJ halothan-intoxication (from E. and F. W. 11
SCHMIDT,
1979); (10).
Exp. Pathol. 39 (1990) 3-4
161
Pot. B. J. ~ absent from home: i. Z. 3.
U/l
a:> •
'to 6 ' 5. 10 • 6. 5 ' 7. 20 ' 8. 11t
35
1.
Z5
't ooys
10 •
2..
3.
+ +
It.
5.
~ ~
~
6.
8.
7.
~
••
15
10
5
W 1971
ron
1972
normal
1910
Fig. 3. GLDH activities in serum in chronic pentachlorophenol intoxication, depending on the stay in the own apartment (wood impregnated with a preserving agent) (from SCHMIDT, E. and I. VIDO, 1976); (14).
There is a wide variety of changes in enzyme activities or enzyme patterns found in the serum after xenobiotic effects on the liver (table 2, figs. 1, 2, 3, 4). This is certainly connected with the fact that the xenobiotics have different intracellular sites of action and that, along with primary reactions, also secondary reactions lead to exit of enzyme out of the cell. Depending on their effect, the xenobiotics can be dichotomized into the obligatory hepatoxins, which produce a liver damage in every exposed person, and the facultative toxins which cause a damage solely in a part of the exposed. With regard to the liver diseases induced by xenobiotics, diverse morphological pictures may dominate, e.g., necrosis, hepatitis, cholestasis as well as the steatosis type (fig. 5). For the detection of liver and bile duct diseases, based on screening diagnostics, ALAT and y-GT and, for further differentiation of the disease process, ASAT, GLDH, AP, CHE are recommended, along with other laboratory examinations (7). According to SCHMIDT and SCHMIDT (10), there is suspicion of a liver damage caused by drugs or toxins: 162
Exp. Pathol. 39 (1990) 3-4
NB
'5- GT 17
GL DH
16
ALAr
15
ASAT CH~
1't
13
12
,
11
10
\
9
\
8 7
6 5
It
3 2 1
I\.
r~,
," I "
I :
fi
J
\.
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Fig. 4. Dynamics of enzyme activities in serum in a man who painted a swimming pool for 3.5 h; composition of the paint and the solvent could not be ascertained. Initial values were known from analytical value a control examination. NB = --'--,.---,-normal value
Hepatotaxic slbstances
Carbcn tetrachloride Thioacetamide Phalloidin Amanitin Cyclophosphamide
Iproniazid Phenytoin Phenylbutazone Allopurinol Indomethacin
Chlorpromazine Imipramin Methylestosterone Ethinylestradiol Tolbutamide
Tetracycline Methotrexote Nitrosomine Azoserin Puromycin
Fig. 5. II *
Exp. Pathol. 39 (1990) 3-4
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if y-GT is disproportionally high and three factors have been ruled out: alcohol, biliary obstruction and space-occupying processes in the liver; if y-GT is normal, but the rest of the enzyme pattern is pathological; if CHE activity falls rapidly or is solitarily low; if there are markedly pathological enzyme activities in serum but a normal histological picture of the liver. Finally, I would like to summarize that the determination of enzyme activities in the serum can be of great value for the detection of xenobiotic effects on the liver, but that maximal information can be gained only in association with the medical history, the clinical findings, with immunological, histological as well as other paraclinical investigations.
References 1. Enzyme Nomenclature. Recommendations (1978) of the Nomenclature Committee of the International Union of Biochemistry. Academic Press, New York-San Francisco-London 1979. 2. FRIEDEL, R.: Der Mechanismus des Ubertritts von Enzymen in das Blut. Med. unserer Zeit 1978; 2: 177-184. 3. - DIEDERICHS, F., LINDENA, J.: Release and extracellular turnover of cellular enzymes. In: Advances in Clinical Enzymology (eds. SCHMIDT, E., SCHMIDT, F. W., TRAUTSCHOLD, I., FRIEDEL, R.). S. Karger, BaselMiinchen-Paris-London-New York-Sydney 1979; pp. 70-105. 4. GRESSNER, A. M.: Leber und Gallenwege. In: GREILING, H., GRESSNER, A. M. (Hrsg.): Lehrbuch der Klinischen Chemie und Pathobiochemie. Schattauer, Stuttgart-New York 1987; pp. 421-507. 5. HASCHEN, R. 1.: Enzymdiagnostik. 2. Auflage, VEB Gustav Fischer Verlag, Jena 1981. 6. - Pathobiochemie der Leber. VEB Verlag Volk und Gesundheit, Berlin 1981. 7. IRRGANG, B., PATZOLD, K., THIELE, H.-J.: Labordiagnostisches Stufenprogramm zur gezielten Unterstiitzung der Diagnostik von Lebererkrankungen. Z. Klin. Med. 1987; 42: 735-740. 8. KELLER, H.: Klinisch-chemische Labordiagnostik fUr die Praxis. Georg Thieme Verlag, Stuttgart-New York 1986. 9. SCHMIDT, E., SCHMIDT, F. W.: Clinical enzymology. FEBS Lett. 1976; 62 (Supp!.): E62-E79. 10. - - Enzyme diagnosis in diseases of the liver and the biliary system. In: Advances in clinical enzymology (eds. SCHMIDT, E., SCHMIDT, F. W., TRAUTSCHOLD, I., FRIEDEL, R.). S. Karger, Basel-Miinchen-Paris-London-New York-Sydney 1979; pp. 239-292. 11. Enzym-Fibe!. Biochemica, Boehringer Mannheim GmbH, 1969. 12. - y-Glutamyl-transpeptidase. Dt. med. Wochenschr. 1973; 98: 1572-1578. 13. - Kleine Enzym-Fibe!. Diagnostics, Boehringer Mannheim GmbH, 1976. VIDO, I.: Exogen-toxische Leberschiiden. In: Toxische Leberschiiden (Medikamente und Leber) (Hrsg. 14. WANNAGAT, L.). Georg Thieme Verlag, Stuttgart 1976, S. 168. 15. ZIMMERMANN, H. J.: Chemical hepatic injury and its detection. In: Toxicology of the liver (eds. PLAA, G. L., HEWITT, W. R.). Raven Press, New York 1982, pp. 32. (Accepted January 17, 1990)
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