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The influence of operative trauma on blood levels of oxytocinase
398
CLINICA CHIMICA ACTA
THE
INFLUENCE
OF OPERATIVE
TRAUMA
ON BLOOD
LEVELS
OF OXYTOCINASE RUDOLF
KLIMEK,
MARIA
PIETRZYCKA
AND
JAN
GROCHOWSKI
1st Obstetrical and Gynecological Clinic* and the 2nd Surgical Clinic * *, Medical Academy, Cracow (Poland)
(Received
October
qth,
1961)
Biochemical biopsy in the form of determinations of enzymic activity of body and tissue fluids has found wide application in medical diagnosis; moreover, the method often provides essential information regarding the severity of the morbid condition, or is helpful in the evaluation of therapeutic results. For clinical and methodological reasons, determinations of enzymic activity are usually made in the blood, and more than twenty such methods have now been adopted routinely. A special position among these is occupied by oxytocinase, the levels of which fluctuate in physiological states. The estimation of oxytocinasaemia has found a permanent place in obstetrical differential diagnosis, especially for predicting the term of pregnancy on the basis of the continuously rising level of oxytocinase in the blood of the pregnant female2-4T e, 9* lo, Ia. Besides the dynamic oxytocin test, such determinations form part of the biochemical method of fixing the date of labor, which is now possible with an error of days, instead of weeks as hithertos. The great interest accorded to oxytocinase is due in the first place to the practical advantage of being able to follow the development of the fetus on the basis of the behavior of oxytocinasaemia. During a myomectomy of a large myoma performed by SCHWARZ on a patient in the third month of pregnancy, our attention was drawn to the difficulty of interpretation of raised levels of oxytocinasaemia due to lack of knowledge of the influence of operative trauma on the levels of this enzyme in blood. Such an influence could be expected in view of the work of OSZACKI~ and OSZACKI et aLs on post-operative changes in the water-electrolyte balance which is associated with the production of the peptide hormones of the posterior lobe of the hypophysis. We therefore decided to investigate the effect of operative trauma on the oxytocinase level in blood. The study was performed on patients operated on in our clinics with the hope of obtaining practical data, and also some theoretical information concerning the possibility of inducing synthesis of oxytocinase in the body by an effect of operative trauma. METHODS
Blood oxytocinase levels were assayed by the method of TUPPY AND NESVADBA~*, which has been described and confirmed in our previous publications47 6. The activity of 0.1 ml of serum splitting off I pug of /?-naphthylamine in standard conditions from * Director Prof. Dr. STEFAN SCHWARZ. ** Director Prof. Dr. JAN OSZACKI.
Clin. Chim. Acta, 7 (1962) 398-402
BLOOD LEVELS
OF OXYTOCINASE
399
L-cystine-di+naphthylamide as substrate represents one unit of the enzyme. The normal Ieve is 4.4 f 1.3 units3$P. Patients of the 1st Obstetrical and Gynecological Clinic and the 2nd Surgical Clinic of the Medical Academy in Cracow were examined. RESULTS
The influence of operative trauma on level of oxytocinasaemia was investigated in 40 patients. Three determinations were made in each patient : before, and 2 and IO days after, operation. The patients were divided into two groups. The first comprised patients in whom hepatic impairment was not expected, and the second group those in whom such impairment was likely, including patients with jaundice or disease of the biliary tract of Iong standing. As is known, in diseases of the liver the level of oxytocinase in the blood is elevated.
I t
operation
1
2
3
4
Fig.
5
6
7
8 Days
9 after
10 operation
I.
Fig. I depicts the behavior of oxytocinasaemia in patients of the first group, which included rg females and 8 males ranging in age from IO to 60 years. In this group 4 stomach resections, 3 thyroidectomies, 4 appendectomies, 3 plastic hemiotomies, 3 trial laparatomies, 3 total hysterectomies, 3 removals of the body uterus, 3 plastic perineal operations, and one ovarectomy were performed. The data from each case are connected by solid lines, and the dashed lines connect the mean values. The arithmetic means and standard deviations are shown in Table I. TABLE
-
Oxytocinasaemia
Before
units
o@eratiola
IMean arithmetic Standard deviation Range
4.74 I.54 2.1-7.7
I
After 2 days
7.24 2.14 2.5-12.7
operation --SOT&G
5.75 2.48 3.6-12
CL&. Chim Acta, 7 (rg6z) 3g8-qoz
w
w
w
w
m
69
49
34
52
62
62
49
38
61
33
3o
5o
G.G.
K.R.
D.J.
Z.J.
SE.
M.J.
K.M.
W.S.
P.P.
H.M.
B.Z.
W.K.
2
3
4
5
6
7
8
9
10
II
12
13
w
w
w
w
m
w
w
w
32
S.W.
I
Sex
Age
No
Patient
--
11
12
7 10.7
4.1 9 I2
Cholecystostomia
Cholecystectomia Choledochotomia Cholecystectomia Choledochotomia Cholecystectomia Choledochotomia Splenectomia Op.m.Luiscot-Blak Cholecystectomia Cholecystectomia
Icterus mechanicus Odditis sclerosans
Icterus mechanicus Choledocholithiasis
Icterus mechanicus Choledocholithiasis
Icterus mechanicus Choledocholithiasis
Cirrhosis hepatis Splenomegalia
Cholelithiasis
Peritonitis
6.4 5 5.8
Cholecystectomia Cholecystectomia Cholecystectomia
Cholelithiasis
Cholelithiasis
Cholelithiasis
-
13
Excisio polypus
Icterus mechanicus Polypus duodeni
biliaris
15
Cholecystectomia Choledochotomia
Icterus mechanicus Choledocholithiasis
____
operation
8
8
18
12
8.6
16
2 days
_---
7
6.2
IO
-
14
II
10
11.7
11
15.5
IO
14
16.5
____
IO days
Oxytocinase units ___ &fO?Y? After operation
18
II
Cholecystectomia Choledochotomia
Operation
TABLE
Icterus mechanicus Choledocholithiasis
Diagnosis
__~~_
II
6.57
7.35
9.68
6.54
10.82
2.64
3.77
7.8
days
7.8
2
8.34
4.40
7.17
8.82
operation
____~
ime%)
days
4.64
I .63
2.10
3.14
1.76
IO ____
After opevation
Bilhbinaemia Before
?
0
%
BLOOD LEVELS OF OXYTOCINASE
401
The elevation in the blood oxytocinase level on the second post-operative day was statistically significant, and persisted for up to IO days. In Table II are listed 13 patients of the second group, including II females and 2 males aged from 3o-69 years. The diagnoses were based on operative findings in the liver and bile tract, supported by microscopic examinations of sections, electrophoretic studies of the serum proteins, determinations of transaminase activity, and the Van den Bergh reaction. A statistical presentation of the mean concentrations of oxytocinase and bilirubin (in cases of mechanical jaundice) is given in Table III. TABLE
III
Before operatkm Oxytocinasaemia units Mean arithmetic Standard deviation Range
9.9 4.17 4.1-18
Bilirubinaemia mg% Mean arithmetic Range
8.17 4.4-x I
After 2
days
12.1
3.61 7-17 6.71 2.64-9.68
operation IO
days
11.4 3.15 6.2-16.5
3.73 1.767.3
A transient rise in the blood level of oxytocinase on the second day after operation was not statistically significant; after IO days a return to the initial level had still not taken place. Nevertheless, these values continued on a high level above normal, which was statistically significant. On the other hand, the continuous decline in the level of bilirubinaemia which was noted, is normal in such cases’. CONCLUSIONS
From our findings it can be concluded that in patients with an intact liver, operative trauma causes a statistically significant rise in the blood level of oxytocinase on the second day, which persists for IO days. These investigations confirm our previous observations, as well as those of other writers, according to whom oxytocinasaemia is distinctly elevated in jaundice6p6. In these cases operative trauma causes a further rise, although this is not statistically significant. Hence, when the blood level of oxytocinase in pregnant females is evaluated, the effect of trauma on the body should be taken into account. SUMMARY
The influence of operative trauma on the level of oxytocinasaemia was investigated in 40 patients. Determinations were made before, and 2 and IO days after, operation. In patients with an intact liver, operative trauma caused a statistically significant rise in the blood level of oxytocinase on the second day, which persists for IO days. The oxytocinasaemia is distinctly elevated in jaundice, but operative trauma causes a further rise; this is not statistically significant. Hence, when the blood level of oxytocinase is evaluated in pregnant females, the effect of trauma must be considered. Clin. Chim. Acta, f (1962) 3g8-px
R. KLIMEK et al.
402
REFERENCES J. GROCHOWSKI AND R. KLIMEK, Polski Przeglad Chir., 33 (1961) 527. R. KLIMEK. Ginekol. Polska. 30 (IQW) 541. R. KLIMEK; Ginekol. Polskaj & the press (1962). R. KLIMEK, 1. MADEJ AND M. PIETRZYCKA, Polski Tymdnik Lek., 16 (1961) 1309. _ R. KLIM&~ND M. PIETRZYCKA, Clin.Chim. Acta, 6iig61) 326. 6 W. MILLER-HARTBURG,H.TUPPY AND H.NESVADBA, Arch.GyniikoZ., 191 (Ig5g)442. 7 J. OSZACKI, Postepy Chirurgii, I (1954) I. 8 J. OSZACKI, A. SOWI&SKA AND B. ADaMCZYK, Polski Przeglad Chir., 2 (1959) 159. 9 E. W. PAGE, Science,105 (1947) 292. 10 C. SMITH, Triangel, 3 (1957) 150. l1 H. TUPPY AND H. NESVADBA, Monatsh.Chem., 88 (1957) 977. I2 E. WERLE AND K. SEMM, Arch. Gyniikol., 187 (1956) 449. 1 B 3 4 5
-__I
-
.I
Clin. Chim. Acta, 7 (1962) 3g8-hoz
ENZYMES
OF THE
RED
ADENYLATE PAOLO
BLOOD KINASE
CERLETTI
AND
CELL
MEMBRANE:
ACTIVITY
GIASCARLO
DE RITIS
Institute of Biological Chemistry of the University of Rome and National Research Council Unit for Enzyme Studies, Rome (Italy) (Received
August 7th, 1961)
Considerable amounts of ATP* and ADP are present within the erythrocyte, the life span of which has been related to its ATP contentl. The mature mammalian red blood cell is however unable to carry out the biosynthesis de lzovo of purine nucleotides2. Although it disposes of the enzymes catalyzing the final reactions required for completion of the purine ring and for the formation of purine nucleotide triphosphates3, this ability may well represent merely a non-functional remnant 2. The main route by which the content of ATP is restored seems therefore to be the phosphorylation of ADP and AMP in the glycolytic and hexose monophosphate pathways. In previous research, adenylate kinase activity has been found within the red blood cells of several mammalian species4 and its partial purification has been achieved5. The presence of an enzyme that preserves the balance between all three adenosine phosphates seems to be most interesting, the more so as it appears only in the cytoplasm of the ce114. Enzyme systems that metabolize adenosine polyphosphates are located both on the cell membrane and in the cytoplasm. It is therefore valuable to have further information on the possibility of some interaction between adenylate kinase and the cell membrane, which would allow a closer functional interconnection with the enzymes located there. EXPERIMENTAL
Materials
and methods
Proteins were determined by the biuret reactions and/or by the absorbancy at 280 m,u. Hb was determined by the absorbancy of the Soret band at 418 mp. Chro* The following abbreviations are used: diphosphate ; AMP: adenosine monophosphate
ATP : adenosine triphosphate ; ADP : adenosine ; Hb: haemoglobin; TCA : trichloroacetic acid.
CLINICA CHIMICA ACTA
THE
INFLUENCE
OF OPERATIVE
TRAUMA
ON BLOOD
LEVELS
OF OXYTOCINASE RUDOLF
KLIMEK,
MARIA
PIETRZYCKA
AND
JAN
GROCHOWSKI
1st Obstetrical and Gynecological Clinic* and the 2nd Surgical Clinic * *, Medical Academy, Cracow (Poland)
(Received
October
qth,
1961)
Biochemical biopsy in the form of determinations of enzymic activity of body and tissue fluids has found wide application in medical diagnosis; moreover, the method often provides essential information regarding the severity of the morbid condition, or is helpful in the evaluation of therapeutic results. For clinical and methodological reasons, determinations of enzymic activity are usually made in the blood, and more than twenty such methods have now been adopted routinely. A special position among these is occupied by oxytocinase, the levels of which fluctuate in physiological states. The estimation of oxytocinasaemia has found a permanent place in obstetrical differential diagnosis, especially for predicting the term of pregnancy on the basis of the continuously rising level of oxytocinase in the blood of the pregnant female2-4T e, 9* lo, Ia. Besides the dynamic oxytocin test, such determinations form part of the biochemical method of fixing the date of labor, which is now possible with an error of days, instead of weeks as hithertos. The great interest accorded to oxytocinase is due in the first place to the practical advantage of being able to follow the development of the fetus on the basis of the behavior of oxytocinasaemia. During a myomectomy of a large myoma performed by SCHWARZ on a patient in the third month of pregnancy, our attention was drawn to the difficulty of interpretation of raised levels of oxytocinasaemia due to lack of knowledge of the influence of operative trauma on the levels of this enzyme in blood. Such an influence could be expected in view of the work of OSZACKI~ and OSZACKI et aLs on post-operative changes in the water-electrolyte balance which is associated with the production of the peptide hormones of the posterior lobe of the hypophysis. We therefore decided to investigate the effect of operative trauma on the oxytocinase level in blood. The study was performed on patients operated on in our clinics with the hope of obtaining practical data, and also some theoretical information concerning the possibility of inducing synthesis of oxytocinase in the body by an effect of operative trauma. METHODS
Blood oxytocinase levels were assayed by the method of TUPPY AND NESVADBA~*, which has been described and confirmed in our previous publications47 6. The activity of 0.1 ml of serum splitting off I pug of /?-naphthylamine in standard conditions from * Director Prof. Dr. STEFAN SCHWARZ. ** Director Prof. Dr. JAN OSZACKI.
Clin. Chim. Acta, 7 (1962) 398-402
BLOOD LEVELS
OF OXYTOCINASE
399
L-cystine-di+naphthylamide as substrate represents one unit of the enzyme. The normal Ieve is 4.4 f 1.3 units3$P. Patients of the 1st Obstetrical and Gynecological Clinic and the 2nd Surgical Clinic of the Medical Academy in Cracow were examined. RESULTS
The influence of operative trauma on level of oxytocinasaemia was investigated in 40 patients. Three determinations were made in each patient : before, and 2 and IO days after, operation. The patients were divided into two groups. The first comprised patients in whom hepatic impairment was not expected, and the second group those in whom such impairment was likely, including patients with jaundice or disease of the biliary tract of Iong standing. As is known, in diseases of the liver the level of oxytocinase in the blood is elevated.
I t
operation
1
2
3
4
Fig.
5
6
7
8 Days
9 after
10 operation
I.
Fig. I depicts the behavior of oxytocinasaemia in patients of the first group, which included rg females and 8 males ranging in age from IO to 60 years. In this group 4 stomach resections, 3 thyroidectomies, 4 appendectomies, 3 plastic hemiotomies, 3 trial laparatomies, 3 total hysterectomies, 3 removals of the body uterus, 3 plastic perineal operations, and one ovarectomy were performed. The data from each case are connected by solid lines, and the dashed lines connect the mean values. The arithmetic means and standard deviations are shown in Table I. TABLE
-
Oxytocinasaemia
Before
units
o@eratiola
IMean arithmetic Standard deviation Range
4.74 I.54 2.1-7.7
I
After 2 days
7.24 2.14 2.5-12.7
operation --SOT&G
5.75 2.48 3.6-12
CL&. Chim Acta, 7 (rg6z) 3g8-qoz
w
w
w
w
m
69
49
34
52
62
62
49
38
61
33
3o
5o
G.G.
K.R.
D.J.
Z.J.
SE.
M.J.
K.M.
W.S.
P.P.
H.M.
B.Z.
W.K.
2
3
4
5
6
7
8
9
10
II
12
13
w
w
w
w
m
w
w
w
32
S.W.
I
Sex
Age
No
Patient
--
11
12
7 10.7
4.1 9 I2
Cholecystostomia
Cholecystectomia Choledochotomia Cholecystectomia Choledochotomia Cholecystectomia Choledochotomia Splenectomia Op.m.Luiscot-Blak Cholecystectomia Cholecystectomia
Icterus mechanicus Odditis sclerosans
Icterus mechanicus Choledocholithiasis
Icterus mechanicus Choledocholithiasis
Icterus mechanicus Choledocholithiasis
Cirrhosis hepatis Splenomegalia
Cholelithiasis
Peritonitis
6.4 5 5.8
Cholecystectomia Cholecystectomia Cholecystectomia
Cholelithiasis
Cholelithiasis
Cholelithiasis
-
13
Excisio polypus
Icterus mechanicus Polypus duodeni
biliaris
15
Cholecystectomia Choledochotomia
Icterus mechanicus Choledocholithiasis
____
operation
8
8
18
12
8.6
16
2 days
_---
7
6.2
IO
-
14
II
10
11.7
11
15.5
IO
14
16.5
____
IO days
Oxytocinase units ___ &fO?Y? After operation
18
II
Cholecystectomia Choledochotomia
Operation
TABLE
Icterus mechanicus Choledocholithiasis
Diagnosis
__~~_
II
6.57
7.35
9.68
6.54
10.82
2.64
3.77
7.8
days
7.8
2
8.34
4.40
7.17
8.82
operation
____~
ime%)
days
4.64
I .63
2.10
3.14
1.76
IO ____
After opevation
Bilhbinaemia Before
?
0
%
BLOOD LEVELS OF OXYTOCINASE
401
The elevation in the blood oxytocinase level on the second post-operative day was statistically significant, and persisted for up to IO days. In Table II are listed 13 patients of the second group, including II females and 2 males aged from 3o-69 years. The diagnoses were based on operative findings in the liver and bile tract, supported by microscopic examinations of sections, electrophoretic studies of the serum proteins, determinations of transaminase activity, and the Van den Bergh reaction. A statistical presentation of the mean concentrations of oxytocinase and bilirubin (in cases of mechanical jaundice) is given in Table III. TABLE
III
Before operatkm Oxytocinasaemia units Mean arithmetic Standard deviation Range
9.9 4.17 4.1-18
Bilirubinaemia mg% Mean arithmetic Range
8.17 4.4-x I
After 2
days
12.1
3.61 7-17 6.71 2.64-9.68
operation IO
days
11.4 3.15 6.2-16.5
3.73 1.767.3
A transient rise in the blood level of oxytocinase on the second day after operation was not statistically significant; after IO days a return to the initial level had still not taken place. Nevertheless, these values continued on a high level above normal, which was statistically significant. On the other hand, the continuous decline in the level of bilirubinaemia which was noted, is normal in such cases’. CONCLUSIONS
From our findings it can be concluded that in patients with an intact liver, operative trauma causes a statistically significant rise in the blood level of oxytocinase on the second day, which persists for IO days. These investigations confirm our previous observations, as well as those of other writers, according to whom oxytocinasaemia is distinctly elevated in jaundice6p6. In these cases operative trauma causes a further rise, although this is not statistically significant. Hence, when the blood level of oxytocinase in pregnant females is evaluated, the effect of trauma on the body should be taken into account. SUMMARY
The influence of operative trauma on the level of oxytocinasaemia was investigated in 40 patients. Determinations were made before, and 2 and IO days after, operation. In patients with an intact liver, operative trauma caused a statistically significant rise in the blood level of oxytocinase on the second day, which persists for IO days. The oxytocinasaemia is distinctly elevated in jaundice, but operative trauma causes a further rise; this is not statistically significant. Hence, when the blood level of oxytocinase is evaluated in pregnant females, the effect of trauma must be considered. Clin. Chim. Acta, f (1962) 3g8-px
R. KLIMEK et al.
402
REFERENCES J. GROCHOWSKI AND R. KLIMEK, Polski Przeglad Chir., 33 (1961) 527. R. KLIMEK. Ginekol. Polska. 30 (IQW) 541. R. KLIMEK; Ginekol. Polskaj & the press (1962). R. KLIMEK, 1. MADEJ AND M. PIETRZYCKA, Polski Tymdnik Lek., 16 (1961) 1309. _ R. KLIM&~ND M. PIETRZYCKA, Clin.Chim. Acta, 6iig61) 326. 6 W. MILLER-HARTBURG,H.TUPPY AND H.NESVADBA, Arch.GyniikoZ., 191 (Ig5g)442. 7 J. OSZACKI, Postepy Chirurgii, I (1954) I. 8 J. OSZACKI, A. SOWI&SKA AND B. ADaMCZYK, Polski Przeglad Chir., 2 (1959) 159. 9 E. W. PAGE, Science,105 (1947) 292. 10 C. SMITH, Triangel, 3 (1957) 150. l1 H. TUPPY AND H. NESVADBA, Monatsh.Chem., 88 (1957) 977. I2 E. WERLE AND K. SEMM, Arch. Gyniikol., 187 (1956) 449. 1 B 3 4 5
-__I
-
.I
Clin. Chim. Acta, 7 (1962) 3g8-hoz
ENZYMES
OF THE
RED
ADENYLATE PAOLO
BLOOD KINASE
CERLETTI
AND
CELL
MEMBRANE:
ACTIVITY
GIASCARLO
DE RITIS
Institute of Biological Chemistry of the University of Rome and National Research Council Unit for Enzyme Studies, Rome (Italy) (Received
August 7th, 1961)
Considerable amounts of ATP* and ADP are present within the erythrocyte, the life span of which has been related to its ATP contentl. The mature mammalian red blood cell is however unable to carry out the biosynthesis de lzovo of purine nucleotides2. Although it disposes of the enzymes catalyzing the final reactions required for completion of the purine ring and for the formation of purine nucleotide triphosphates3, this ability may well represent merely a non-functional remnant 2. The main route by which the content of ATP is restored seems therefore to be the phosphorylation of ADP and AMP in the glycolytic and hexose monophosphate pathways. In previous research, adenylate kinase activity has been found within the red blood cells of several mammalian species4 and its partial purification has been achieved5. The presence of an enzyme that preserves the balance between all three adenosine phosphates seems to be most interesting, the more so as it appears only in the cytoplasm of the ce114. Enzyme systems that metabolize adenosine polyphosphates are located both on the cell membrane and in the cytoplasm. It is therefore valuable to have further information on the possibility of some interaction between adenylate kinase and the cell membrane, which would allow a closer functional interconnection with the enzymes located there. EXPERIMENTAL
Materials
and methods
Proteins were determined by the biuret reactions and/or by the absorbancy at 280 m,u. Hb was determined by the absorbancy of the Soret band at 418 mp. Chro* The following abbreviations are used: diphosphate ; AMP: adenosine monophosphate
ATP : adenosine triphosphate ; ADP : adenosine ; Hb: haemoglobin; TCA : trichloroacetic acid.