Resuscitation 7, 185-198
Alterations in the enzyme profile in intensive care patients undergoing total parenteral nutrition
M. ZANELLO,
E. CASTELLI, J. BERGER and C. CETRULLO
Institute of Anaesthesiology and Reanimation of the University of Bologna, Policlinico S. Orsola, Via Massarenti, 9, 40138, Bologna, Italy
Summary
Total parenteral nutrition (TPN) has been demonstrated to be an effective therapeutic means in improving the clinical course of the critically ill patient. Various metabolic complications are described; the cause of some of these remain unclear. The changes in some plasma enzyme indices (GOT, GPT, GIDH, LDH, HBDH, CPK, ChE, AP, gamma-GT) in two groups of critically ill patients undergoing TPN (group with more marked enzyme alterations and group with less marked alteration) were examined. Two types of alterations were found: (1) early increase of some enzymes (GOT, GPT, GIDH); (2) constant increase of plasma enzyme level during TPN (AP, gamma GT). These two evolutionary patterns were more evident in the complicated group and the enzyme changes were statistically significant for GOT and GPT (P=O.O5) and not significant for initial values of GlDH, AP and gamma-GT. Both groups presented constant elevated plasma values of LDH, HBDH, CPK and depressed constant ChE value during treatment; the difference was not significant in both groups for the same enzymes. The data were interpreted from a functional point of view; that is they were related to both the metabolic post-aggressive state and TPN. A relationship between the rate of protein catabolism and the inductive increase of some enzymes (GOT, GPT, GlDH) was found. Whereas a final induction in the energy metabolism is suggested for other enzymes (LDH, HBDH), the alteration of CPK, AP, gamma-GT and ChE was interpreted as dependent on: (1) direct muscular trauma (CPK); (2) functional increase in relation to the duration of TPN (AP and gamma-GT); (3) possible depressed malnutritive synthesis (ChE). The improvement of the enzymatic patterns with the early use of TPN and with the improvement of clinical and nutritional conditions was emphasized. Introduction
The use of total parenteral nutrition (TPN) in intensive care has significantly improved patient survival. With the extension of experience and application of this nutritional approach, a host of technical and metabolic complications have been observed (Dudrich, McFayden, Van Buren, Ruberg & Maynard, 1972; Ryan, 1976; Sanders & Sheldon, 1976). 185
186
M. ZANELLO AND OTHERS
Some of the metabolic alterations, ‘apparent complications’, remain a continuous problem. In previous reports (Cetrullo, Castelli & Zanello, 1976; Castelli, Cetrullo & Zanello, 1978) data were presented concerning the serial changes in certain laboratory indices of hepatic and metabolic function during TPN. However, the cause of these alterations remains substantially unclear. In this report we present data concerning our most recent experience with TPN in critically ill patients in which we have attempted to correlate serial alterations in the laboratory indices of metabolic function with the clinical course of the patients. Special consideration is given to establish those alterations attributable directly to the disease state and those due to TPN. In this study we direct our attention to the following questions: (1) Is there a characteristic pattern to the alterations in the laboratory indices of metabolic function during TPN in critically ill patients? (ii) Are these alterations due to the TPN, to the pathology of the disease state or to a combination of the two? (iii) Do the alterations in the laboratory indices correlate with the clinical course of these patients? (iv) Does early use of TPN affect the laboratory changes? Patients and methods The patients population for this study consisted of 16 females and 18 males, of ages between 16 and 76 years, with one patient of age 5 years. They represent patients who underwent TPN for more than 5 days in the Intensive Care Unit (ICU) of the Institute of Anaesthesiology and Reanimation, University of Bologna, Italy, in the last year and were selected for inclusion without bias as to age (excluding newborn and infants), sex, presenting disease or clinical condition (except for patients with obvious hepatic or cardiac disease). The most frequent TPN solution used consisted of a mixture of 50% glucose and 8.5% amino acids (Freamine) approaching progressively the infusion of 750 g of glucose and 18.75 g of N per day: electrolytes and vitamins were added to the solution according to need. Hypertonic solutions were infused, according to the usual method, by means of a vena caval catheter; correct placement of catheter was controlled radiologically. In addition to the usual laboratory studies, which are necessary for the proper care of critically ill patients and conduction of TPN, such as determination of electrolytes, nitrogen balance, blood gases, urinalysis etc., activities of the plasma enzymes of particular interest in this study were measured: transaminases, more specifically glutamic-pyruvic transaminase (GPT), glutamic-oxaloacetic transaminase (GOT) and glutamic dehydrogenase (GlDH); alkaline phosphatase (AP), gamma-glutamyl transpeptidase (gamma-GT); lactic dehydrogenase (LDH), beta-hydroxybutyrate dehydrogenase (HBDH); creatine phosphokinase (CPK); pseudo-cholinesterase (ChE). Results Summary data of the patient population are presented in Tables 1 and 2. The patients are divided into two groups, designated as a ‘laboratory uncomplicated’ group (LUC) and a ‘laboratory complicated’ (LC), respectively. The criteria used for their separation
ENZYMES IN TOTAL PARENTERAL NUTRITION
187
Table 1. Diagnosis and complications of 15 ‘laboratory uncomplicated’ patients. Key (and for Table 2): B (bleeding), C (cardiocirculatory insufficiency), K (renal insufficiency), J (hyperbilirubinaemia), N (coma/neurologic disorder), 0 (surgical operation), P (peritonitis), R (respiratory insufficiency), S (sepsis), D (deceased), T (transferred).
Patient
Age (years)
Sex
Time after admission to onset of TPN (days)
C.L.
35
F
2
20
M.V. M.L.L.
72 29
F F
1 1
31 15
C.P. M.B.
24 74
M M
0
40 24
G.V. G.P.
52 66
F M
0
3
8 20
G.G.
76
M
2
20
M.V.
76
F
0
17
N.P. A.D.P. I.B.
22 72 51
M M F
2 4
40 30 6
A.L.
72
F
1
6
G.R.
66
F
1
11
G.B.B.
20
M
0
6
X
53,8
M=7 P=8
1,8(+2,3)
9
1
Duration of TPN (days)
20
Diagnosis on admission Ingestion of caustic agent Respiratory insufficiency Ingestion of caustic agent Chest/abdominal traums Cerebral vascular accident Acute pancreatitis Intestinal occlusion/ peritonitis Ingestion of caustic agent Postoperative respiratory insufficiency Multiple trauma Respiratory insufficiency Respiratory insufficiency/ myasthenia Ingestion of caustic agent Postoperative respiratory insufficiency Chest trauma
Clinical complication
Outcome T
R K
D D
O-S-B-K N O-N-K B-S-P T
T
N-R-C
D
O-R-S-B N-K-R C-R
T D T
J-K
D
O-R
T
B-R
D T = n.6 D = n.9
were based on the following observation. Initially all patients were grouped together and a mean value was calculated for each measurement for each TPN day. Although this analysis presented us with a relatively clear picture of altered laboratory indices during TPN, individual values varied very much. It was therefore decided to divide the patients into two groups on the basis of whether they showed marked alterations in at least one or more days in at least three or more of laboratory indices. It is to be emphasized that this division was initially based on the laboratory values, but it was also seen that there was a relationship between the extent of laboratory changes and the frequency of severe clinical complications, that is the LC group also showed more critical clinical course. The two groups differed also in the fact that the LC group was composed of 44% of severe trauma cases, compared with 20% in the ‘laboratory uncomplicated’ group. Analysis of the data in Tables l-2 shows that the number of patients, age and sex distribution were similar in both groups: also in both groups the mortality rate was
188 M. ZANELLO AND OTHERS Table 2.
Diagnosis and complications of 19 ‘laboratory complicated’ patients.
Patient
Age (years)
Sex
Time after admission to onset of TPN (days)
O.P. D.A. G.M. C.M. G.R. I.V. W.C. A.C.
20 12 60 17 39 54 51 54
M M F F F M M M
10 2 5 5 1 6 3 1
22 30 62 6 21 15 6 21
G.F. M.P.
36 73
M F
3 20
14 15
G.T. M.G.
69 43
M M
3 2
12
T.C.
53
F
4
29
G.C. A.L. S.I. A.N. F.D.M. F.M.
5 16 44 34 50 52
M M F F M M
21 31 69 21 10 22
x
44,6
M= 12 4,74 (+4,7) F=7
24,44
See also Table 1.
Duration of TPN Diagnosis on admission (days)
Clinical complications B-O-J N-S R-O-P-S P O-S-J-K O-J-S R O-R-K-S
T T T T D T
O-P-S-K R-C-K
D D
B-N-O S
D D
R-C-B
D
o-c
T T T D D T
I
Ingestion of alkali Head trauma Multiple trauma Chest/abdominal trauma Multiple trauma Chest/abdominal trauma Respiratory insufficiency Postoperative esophagocoloplasty Chest/abdominal trauma Mendelson synd./ megaesophagus Multiple trauma Postoperative esophagocoloplasty Peritonitis/postgastrectomy Cachexia/colectomy CO poisoning Chest/abdominal trauma Coma post-eclampsia Cerebral Vast. act. Ingestion of caustic agent
N-K-R R-O-S-P N-R-J N-B-R B-O
Outcome
:
T=lO D=3
high, as expected in critically ill patients. The LC group on the average had a longer interval between admission to the intensive care unit and the initiation of TPN than did the LUC group (means of 4.7 and 1.8 days respectively); this difference was significant (P =O.Ol). On the average the LC group also had a longer course of TPN (mean of 24.4 and 20 days/patient respectively). The data for SGOT and SGPT are presented in Figs. 1 and 2. In these Figures only the mean values for the two groups have been presented for evidence. Bearing in mind that there was a significant dispersion of individual values within each group, the initial average values for the LC group are seen to be significantly greater than the LUC group for both enzymes (P=O.O5). During the course of TPN the LC group showed an initial period of increasing activities of both enzymes followed by a slower return towards initial, and finally almost normal levels. One notes also that the extent of the increase in values is greater in the initial phase for GPT than for GOT. The LUC group, on the other hand, showed relatively constant values during the entire TPN course, remaining slightly above the normal range.
189
ENZYMES IN TOTAL PARENTERAL NUTRITION mu/ml
100
1
90.
_
i&C.
. ..---.
L.C. .
p=
GROUP GROUP
0.05
s”“‘“-I
\
20. 10.
N.V.
1
5
10
15
20
25
TPN
DAVS
Fig. 1. Plasma activity of glutamic-oxaloacetic transaminase as mean values in the ‘laboratory uncomplicated’ (LUC) and ‘laboratory complicated’ (LC) groups of patients during TPN.
-
Luc.
.----
L.C. .
GROUP GROUP
p= 0.05
N.V. 1
5
10
15
20
25 TPN
DAYS
Fig. 2. Mean plasma activities of glutamic-pyruvic transaminase in the ‘laboratory uncomplicated ‘laboratory complicated’ groups of patients during TPN.
and
Analysis of GlDH values in the same two groups (Fig. 3) showed a marked spread of individual values; because of this the difference between the means of the two groups was not always statistically significant, but the mean values presented by the LC group initially presented values which tended to increase during the first 10 days of TPN, followed by progressive decrease towards initial values in the second 10 days. The LUC group, on the other hand, presented values which tended to remain in the normal range or slightly above during the entire nutritional therapy.
190 mu/ml
M. ZANELLO
AND OTHERS
60 40.
-
L.U.C.
GROUP
c--a
L.C.
GROUP
36. 30. 26.
1
3
6
7
6
11
Fig. 3. Mean plasma activities of glutamic dehydrogenase patients during TPN.
13
16
‘17
16
21
IN.V.
TPN DAYS
and their standard deviation in the two groups of
The data concerning other indices of amino acid metabolism related also to bile excretion are in Figs. 4 and 5, The evolution of AP activity showed similar initial values for both groups of patients which fell within normal range (Fig. 4). The LC group, however, showed a relatively linear increase in AP activity over the entire course of TPN; the LUC group maintained normal levels throughout. The evolution of gamma-GT activity (Fig. 5) in the LC group was similar to that of AP activity. In this case, however, the LUC group also presented increasing values above the normal range. Also for these two enzymes statistically significant difference between the mean values for both groups was not noted. The LDH and HBDH enzymes were followed in this study as indices related to intermediate and energetic metabolism. There was great variability in the mean values in both groups during the course of TPN for both enzymes and no characteristic patterns were observed. The mean values remained generally above the upper normal limits but did not exceed twice the normal values. The initial values of CPK activity also showed marked variability for both groups, which was greater in the LC group: these initial values were significantly elevated above the normal range, especially in the LC group. Both groups also showed a tendency towards decreasing values during the TPN course, with eventual earlier return to normal in the LUC group (Fig. 6). The ChE enzyme values for the two groups are presented in Fig. 7. Both groups had initial values below the normal range and both groups subsequently showed similar evolutionary patterns during the course of therapy, i.e. subsequent return towards the lower limit of the normal range during the following 10-12 days. Surprisingly, this tendency toward normalization was more marked in the LC patients. Finally, the mean nitrogen balance was calculated for the two groups for each day of TPN (Fig. 8). Except for an initial positive peak in the LUC group, both groups showed
ENZYMES
mu/ml
IN TOTAL
PARENTERAL
191
NUTRITION
_
L.U.C. GROUP
.----.
L.C.
GROUP
300-
25O-
Y
T
N.V.
50.
1
Fig. 4. TPN.
mu/ml
Changes
5
10
in the mean alkaline
15
phosphatase
activity
1 25 TPN
20
in the two groups
DAYS
of selected patients
_
t..U.C.
.-e-B.
L.C.
during
GROUP GROUP
200.
150.
100.
50.
l-2
Fig. 5. Changes during TPN.
5-6
in the gamma-glutamyl
9-10
transpeptidase
13-14
plasma
17-16
activity
21-22
in the same groups
TPN
DAYS
of patients
a protracted period of negative balance, which lasted approximately 18 days, with subsequent tendency toward positive balance, reached earlier by LUC group. Discussion Several findings have prompted us to consider the alterations in the enzyme activity during TPN from a functional point of view. First, the patients selected for
192
M. ZANELLO AND OTHERS
mu/ml
700
900
1
500. 400. 300.
8
‘\
‘\,
200.
.... -
i&c.
oooo -----
Lc.
GROUP GROUP
0 ‘\
‘ii
iFi 50. 40. 30. N.V.
20
10
1* 0’
l-2
9-10
5-9
17-19
13-u
21-22
TPN DAYS
Fig. 6. Spread of individual values and mean changes of creating phosphokinase plasma activity in the ‘laboratory uncomplicated’ and ‘laboratory complicated’ groups during TPN.
mu/ml
_
L.U.C.
c--d
L.C.
9J
GROUP GROUP
J
f
I
N.V.
I 1
Fig. 7.
3
5
7
Mean plasma pseudocholinesterase
9
11
13
15
17
q9
21 TPN DAYS
activity in the same two groups of patients during TPN.
ENZYMES IN TOTAL PARENTERAL NUTRITION
9
_
L.u.c.
.-m-d
L.C.
193
GROUP
GROUP
N/day
1
Fig. 8.
5
10
, 15
20
25 TPN
DAYS
Changes in mean nitrogen balance for each TPN day in the two groups of patients.
this study did not have liver disease. Secondly, although both groups of patients showed alterations in plasma enzyme activities, they did not reach the levels generally associated with severe organic lesions or necrosis in either group. Thirdly, notwithstanding the difference in the enzyme patterns noted between the two groups, the mortality and recovery rates were similar. Generally alterations in plasma enzyme activity can be considered to result from four conditions: decrease in synthesis; increase in postinductive synthesis; increase in loss or leakage from cells; defective excretion (Zimmerman & Seeff, 1970; Schmidt & Schmidt, 1971; Schepartz, 1973). Some of these mechanisms come into play, for various functional reasons, in critically ill patients undergoing TPN. One of the principal disturbances in cellular function, resulting from altered energetic post-traumatic metabolism, is abnormal membrane permeability (Flear, 1970, 1974). The subsequent loss or leakage of small molecules (ions, amino acids, substrates etc.) and also larger molecules (such as small proteins) can lead to variations in the normal plasma levels of these substances (McFarlane, 1969; Courtice, Adams, Shannon & Bishop, 1974). Further variations in the plasma activities of various hepatic and muscular enzymes have been described, which reflect the response to changes in nitrogen content of the diet; these include thereonine-serine dehydrogenase (Harper, 1965); some enzymes of the urea cycle (Schimke, 1962; Das & Waterlow, 1974) and several aminotransferases including glutamic acid-oxaloacetate aminotransferase (Steigink & Baker, 1971). Probably an abundance of amino acids entering the liver causes an increase in the amount of transforming enzymes: the enzyme induction seems to be a very fundamental biological reaction. The complicated interaction, also, of multiple
194
M. ZANELLO
AND OTHERS
transaminases in cells and mitochondria has been extensively reviewed by Munro (1964, 1970). Theoretically, the elevated content in amino acids of the TPN results in a condition which is similar to that described above. Further, the increased endogenous turnover of amino acids and protein catabolism (primarily muscular), which characterize the poststress phase in critically ill patients (Cuthberson & Tiltson, 1969; Kinney, Duke, Long & Gump, 1970; Moore & Brennan, 1975) are further inductive factors in this nutritional context. The concentration of amino acids in the blood is also regulated by certain hormones such as the catecholamines, insulin, glucagon, growth hormone and glucocorticosteroids, whose circulation concentrations are also increased in the acute post-traumatic phase (Wilmore, Long, Skreen, Mason & Pruitt, 1974; Johnston, 1974; Moore & Brennan, 1975; Blackburn, Maini & Pearce, 1977). It is also known that some of these hormones have a direct action of enzyme induction (Wannemacher, 1975). Further, it has been demonstrated that there is a reduction in the levels of certain enzymes whose synthesis is depressed in the condition of malnutrition; various hepatic enzymes (which are labile proteins) (Munro, 1964), amylase and alkaline phosphatase (Srikantia, Jacob & Reddy, 1964) and pseudocholinesterase (Waterlow & Stephen, 1970). The serial measurement of plasma activity of certain enzymes in patients undergoinng TPN during the acute phase of recovery in the intensive care unit show two general evolutionary patterns. Some enzymes, e.g., GOT, GPT, GlDH, whose activity was elevated in the plasma in the initial post-acute period or in relation to a worsening clinical course, usually show a global tendency towards return to normal of values with improvement of the clinical and nutritional condition of the patient. Other enzymes, such as AP and gamma-GT, on other hand, tend to show a continuous increase in relation to the duration of TPN. These two patterns of evolution of the enzymes examined appear more pronounced in the LC group than in the LUC group as defined earlier, although both groups were treated similarly with respect to TPN. The TPN itself did not seem to have a direct effect on these enzymatic alterations, since in most cases in the LC group enzyme activities were usually significantly elevated even before the onset of TPN. The increase in plasma activity of those enzymes connected with gluconeogenesis (GOT, GPT), urea cycle (GlDH) and general energetic metabolism (LDH, HBDH, CPK) therefore seems to be related to the particular posttraumatic metabolic response of critically ill patients. The final effect of the post-traumatic catabolic reaction seems to be the production of small molecules, intermediate substrates of glucose metabolism which can be inserted into the multiple anabolic pathways of gluconeogenesis, protein and lipid synthesis or degradative pathways to yield energy (Baruh, Sherman, Kolodny & Singh, 1973; Wannemacher, 1975; Owen, Pate1 & Block, 1976), processes that necessitate various enzyme systems, including those followed in this study. The increase of cellular enzymes in the blood as a result of altered membrane permeability from unbalanced energetic metabolism due to toxic or hypoxic conditions (Flear, 1970; 1974) is supported by the finding that the level of these enzymes remained lower and returned to normal levels more quickly in the patients in which TPN was initiated sooner, possibly because this reduced the degree of enzyme induction or decreased celiular leakage by improving the energetic state of the cells.
ENZYMES
IN TOTAL PARENTERAL
NUTRITION
195
In this context TPN exerts an effect on metabolic homeostasis, which, at least theoretically, modulates the plastic and energetic cellular requirements, and therefore, also changes the orientation of cellular metabolism by reducing those conditions which promote catabolism. The alteration in transaminases activity (GOT, GPT, GlDH) is considered to reflect the extent of protein catabolism, the amount of transformation and transport of NH:, and the rate of gluconeogenesis. It seems that the increase in transaminase activity is the result of enzyme induction in response to endogenous, and perhaps exogenous, increased substrate levels, the latter due to amino acids during total parenteral nutrition. The most elevated levels of enzyme activity observed in the LC group (clearly more catabolic, as demonstrated by the considerable negative nitrogen balance: Fig. 8) seems to confirm this supposition, even though we have not measured the specific substrates of the reactions. The degradation of branched-chain amino acids resulting in the formation of glutamine (Felig, Wahren, Karl, Cerasi, Luft & Kipnis, 1973) and the liberation of alanine (Felig, Pozefsky, Marliss & Cahill, 1970; Marliss, Aoki, Pozetsky, Rost & Cahill, 1971) on the part of muscle tissue seems to be the fundamental mechanisms for the endogenous production of some of those substrates. Various amino acids, but particularly alanine, must first be converted into pyruvate in order to enter the hepatic gluconeogenetic pathway (Wannemacher, 1975), which can explain the obligatory rise in the converting enzyme, GPT (alanine to pyruvate), as noted in the more catabolic patients of the LC group. The increased activity of the converting enzyme GOT (aspartate to oxaloacetate), which was also observed in these patients, must be considered as the result of a more non-specific increase in the degradative process involving many amino acids, but which serves as a fundamental step in subsequent gluconeogenesis, and acts as the metabolic key for glucose metabolism and the Krebs cycle. The increased production of NH: by transamination with secondary acceleration of the urea cycle activity, probably induces synthesis of GlDH, which participates in the transport of ammonium radicles from the muscle to the liver, and catalyses the amination of many amino acids, as demonstrated during starvation (Das & Waterlow, 1974). In the post-traumatic state, in addition to accelerated activities of the glucosealanine cycle (Feling et al., 1970) and the glutamine cycle (Wannemacher, 1975), one notes also the induction of the Cori cycle (glucose-lactate-glucose) (Bassler, 1976). The increases in LDH activity in both groups of patients seems to indicate either an induction of alternative energetic pathways or the permanence of conditions of anaerobic metabolism. These same mechanisms may be similarly active in the elevation of the HBDH activity, the enzyme which catalyses the reaction acetoacetate-betahydroxybutyrate, making ketone bodies as an alternative energy source in post-stress metabolism (Barton, 1970). The elevated levels of CPK observed more markedly in the LC patients because of the magnitude of the values reached, the particular evolutionary patterns observed and the simultaneous presence of negative nitrogen balance (as expression of energy deficit), seem to be more related to muscular trauma than to enzyme induction and accumulation of high-energy substrates as described in other clinical conditions (Clark, 1971; Nevins, Madhukar, Bright & Lyon, 1973; Sklar & Wigand, 1973).
196 M. ZANELLO AND OTHERS
The tendency towards increased activities of other enzymes, such as AP and gammaGT, in relation to the duration of TPN therapy, seems to signify an increased need for amino acid transport in the gamma-glutamyl cycle, or some functional hepatic changes, probably cholestatic in nature, even though hyperbilirubinaemia is an inconstant finding. One cannot exclude the protracted infusion of hypertonic glucose solutions or the eventual deficit or imbalance of amino acid mixtures employed in TPN as probable causes of these cholestatic-like enzyme patterns (Dubrick et al., 1972; Touloukian & Downing, 1973). However, it is important to emphasize that critically ill patients often suffer from clinical conditions which can lead to hepatic damage, cholestasis or changes in the entero-hepatic circulation of bile salts (e.g., peritonitis, septicaemia, intestinal occlusion, prolonged and massive drug treatment etc.), which cannot be neglected as possible aetiological agents for some of these enzyme alterations. The constant observation of depressed plasma ChE values is interpreted as an expression of the general reduced synthesis of this rapidly turning over enzyme probably due to effective malnutrition. This view is supported by the finding of progressively increasing levels of this enzyme related to improvement in nitrogen balance. The relatively more elevated mean values of the LC group are not easily explained. It is possible that the more frequent blood and plasma transfusions given to this group of patients served as an exogenous source of enzyme. On the other hand, the difference between the mean values for the two groups may be only apparent as it was not found to be statistically significant. Finally, comparing the individual enzyme patterns of the two groups of patients with their respective nitrogen balance curves, one notes a definitive relationship between improvement in nitrogen balance and normalization of plasma enzyme levels. The delay in the nitrogen balance becoming positive in the LC group with respect to the LUC group could be dependent on the delay in initiation of TPN in this group, and the lack of positive initial values can be considered as an expression of nitrogen deficit accumulated before initiation of nutritional parenteral support. In view of what has been discussed so far, it is now possible to answer the four questions which were posed in the Introduction to this paper. On the basis of our data, the answers are: (i) In spite of the great variability of enzyme levels between individual patients, each group (LC and LUC) showed a characteristic evolutionary pattern for some enzymes and, above all, of the transaminases. (ii) The altered levels of plasma enzymes observed in these patients were probably due to functional disturbances of the cellular activity. However, at the same time, TPN, as exogenous source of amino acids, could induce specific enzyme synthesis in relation to the metabolic tendency of the post-stress condition. (iii) The patients whose clinical course was more severe seem to have more markedly altered metabolic indices. In addition, there was usually a general improvement indicated by these laboratory findings, with improvement of the clinical conditions. (iv) Early initiation of TPN improves the pattern of laboratory findings in critically ill patients probably by several mechanisms. It is clear that patients receiving early TPN have much more improved nitrogen and energy balances. That probably leads to a decreased leakage of intracellular enzymes even if the absolute enzyme activity has been increased because of induction. According to this point of view, TPN seems to be an effective therapeutic means of control of the metabolic response to injury.
ENZYMES IN TOTAL PARENTERAL NUTRITION
197
Barton, R. N. (1970) Ketone body metabolism after trauma. In: Energy Metabolism in Trauma, pp. 173-181. Ed. Porter, R. & Knight, J. Churchill, London. Baruh, S., Sherman, L., Kolodny, D. & Singh, J. (1973) Fasting hypoglycemia. Med. Clin. North Amer. 57, 144-1449. Biissler, K. H. (1976) Metabolism of the nutrient substances used for parenteral nutrition. In: Parenteral Nutrition, pp. 1-16. Ed. Ahnefeld, F. W., Burri, C., Dick, W. & Halmagyi, M. Springer-Verlag, Berlin. Blackburn, G. L., Maine, B. S. & Pierce, E. C. (1977) Nutrition in the critically ill patient. Anesthesiology 47, 181-194. Castelli, E., Cetrullo, C. & Zanello, M. (1978) Metabolic alterations during prolonged total parenteral nutrition in intensive care. Resuscitation 6, 235-242. Cetrullo, C., Castelli, E. & Zanello, M. (1976) Complications of total parenteral nutrition. In: Total Parenteral Aliment&on, pp. 316-329. Ed. Manni, C., Scrascia, S. I. & Magalini, E. Excerpta Medica, Amsterdam, Oxford. Clark, R. G. (1971) The effect of surgery on the plasma level of creatine phosphokinase. Br. J. Surg. S&304305. Courtice, F. C., Adams, E. P., Shannon, A. D. & Bishop, D. M. (1974) Acid hydrolases in the rabbit in haemorrhagic shock. Quart. .I. Exp. Physiol. 59, 31-41. Cuthbertson, D. P. & Tilstone, W. Y. (1969) Metabolism during the post injury period. Adr. Clin. Chem. 12, l55. Das. T. K. & Waterlow, J. C. (1974) The rate of adaptation of urea cycle enzymes, aminotransferase and glutamic dehydrogenase to changes in dietary protein intake. Br. J. Nutr. 32, 353-361. Dudrick, S. J., McFayden, B. V., van Buren, C. T., Ruberg, R. L. & Maynard, A. T. (1972) Parenteral hyperalimentation, metabolic problems and solutions. Ann. Surg. 176, 259-264. Felig, P., Pozefsky, T., Marliss, E. & Cahill, G., Jr (1970) Alanine: key role in gluconeogenesis. Science 167, 1003-1004. Felig, P., Wahren, J., Karl, I., Cerasi, E., Luft, R. & Kipnis, D. M. (1973) Glutamine and glutamate metabolism in normal and diabetic subjects. Diabetes 22, 573-576. Flear, C. T. G. (1970) Metabolic response to trauma. J. C/in. Path. 23 (Suppl. 4). 16-24. Flear. C. T. G. (1974) Intravenous feeding. Lancer i. 757-758. Harper, A. E. (1965) Effect of variations & protein intake on enzymes of amino acid metabolism. Can. J. Biochem. 43, 1589-1598.
Johnston, I. D. A. (1974) The endocrine response to trauma. In: Parenteral Nutrition in Acute Metabolic Illness, pp. 211-218. Ed. Lee, H. A. Academic Press. London, New York. Kinney, J. M., Duke, J. H., Long, C. L. & Gump, F. E. (1970) Carbohydrate and nitrogen metabolism after injury. J. Chin Pathol. 23 (Suppl. 4), 65-74. Marliss, E., Aoki, T., Pozefsky, T., Rost, A. & Cahill, G., Jr (1971) Muscle and splanchnic glutamine and glutamate metabolism in post-absorptive and starved man. J. Clin. Invest. 50, 814817. McFarlane, R. G. (1969) A discussion on triggered enzyme systems in blood plasma. Proc. Roy. Sot. B. 173, 257445. Moore, F. D. & Brennan, M. G. (1975) Surgical injury: body composition, protein metabolism and neuroendocrinology. In: Manual ofSurgical Nutrition, pp. 169-222. Ed. Ballinger, W. F. Am. Coil. of Surgeons. W. B. Saunders, Philadelphia. Munro. H. N. (1964) General aspects of the regulation of protein metabolism by diet and by hormone. In: Mammalian Protein Metabolism, vol. 1, pp. 396435. Ed. Munro, H. N. & Allison, J. B. Academic Press, New York. Munro, H. N. (1970) In: Balanced Nutrition and Therapy, pp. l-47. Ed. Long, K., Fekl, W. & Berg, G. Georg Thieme Verlag, Stuttgart. Nevins, M. H., Madhukar, S., Bright, M. & Lyon, L. J. (1973) Pitfalls in interpreting serum creatine phosphokinase activity. J. Am. Med. Assn. 224, 1382-1387. Owen, 0. E., Patel, M. S. & Block, B. S. B. (1976) Gluconeogenesis Regulation in Mammalian Species, pp. 533-558. Ed. Hanson, R. W. & Mehenan, M. A. John Wiley, New York. Ryan, J. A., Jr (1976) Complications of total parenteral nutrition. In: Total Parenteral Nutrition. pp. 55-100. Ed. Fischer, J. E. Little, Brown and Co., Boston. Sanders, R. A. & Sheldon, G. F. (1976) Septic complications of total parenteral nutrition: a five year experience. Am. J. Surg. 132, 216220. Schepartz, B. (1973) Introduction to metabolic regulation. In: Regulation of Amino Acid Metabolism i~ Mammals, pp. 1-18. W. B. Saunders, Philadelphia.
198 M. ZANELLO AND OTHERS Schimke, R. T. (1962) Adaptive characteristics of urea cycle enzymes in the rat. J. Biol. Chem. X37,459468. Schmidt, E. & Schmidt, F. W. (1971) Theorie un praxisder Enzymdiagnostik. Wiener Min. Wschr. 20,353369. Sklas, S. H. & Wigand, J. S. (1973)Creatine phosphokinase as indicator of adenosine triphosphate activity. J. Am. Med. Assn. 226, 1464-1465. Srikantia, S. K., Jacob, C. M. & Reddy, V. (1964) Serum enzyme levels in protein-calorie malnutrition, Am. J. Dis. Child. 107, 256-259. Steigink, L. D. & Baker, G. L. (1971) Infusion of protein hydrolysate in the newborn infant: plasma amino acid concentrations. J. Ped. 78, 595-602. Touloukian, R. J. & Downing, S. E. (1973) Cholestasis associated with long-term parenteral hyperalimentation. Arch. Surg. 106, 58-62. Wannemacher, R. W. Jr (1975) Protein metabolism (applied biochemistry). In: Total Parenteral NutritionPremises and Promises, pp. 85-153. Ed. Ghadirni, H. J. Wiley, New York. Waterlow, J. C. & Stephen, J. M. L. (1969) Enzymes and the assessment of protein nutrition. Proc. Nutr. Sot. 28,234246.
Wilmore, D. W., Long, J. A., Skreen, R., Mason, A. D., Jr & Pruitt, B. A., Jr (1974) Catecholamines: mediator of the hypermetabolic response following thermal injury. Ann. Surg. 180,653--669. Zimmerman, H. F. & See&L. B. (1971) Enzymes in hepatic disease. In: Diagnostic Enzymology, pp. 149. Ed. Coodley, E. Lea and Fabiger, Philadelphia.
Alterations in the enzyme profile in intensive care patients undergoing total parenteral nutrition
M. ZANELLO,
E. CASTELLI, J. BERGER and C. CETRULLO
Institute of Anaesthesiology and Reanimation of the University of Bologna, Policlinico S. Orsola, Via Massarenti, 9, 40138, Bologna, Italy
Summary
Total parenteral nutrition (TPN) has been demonstrated to be an effective therapeutic means in improving the clinical course of the critically ill patient. Various metabolic complications are described; the cause of some of these remain unclear. The changes in some plasma enzyme indices (GOT, GPT, GIDH, LDH, HBDH, CPK, ChE, AP, gamma-GT) in two groups of critically ill patients undergoing TPN (group with more marked enzyme alterations and group with less marked alteration) were examined. Two types of alterations were found: (1) early increase of some enzymes (GOT, GPT, GIDH); (2) constant increase of plasma enzyme level during TPN (AP, gamma GT). These two evolutionary patterns were more evident in the complicated group and the enzyme changes were statistically significant for GOT and GPT (P=O.O5) and not significant for initial values of GlDH, AP and gamma-GT. Both groups presented constant elevated plasma values of LDH, HBDH, CPK and depressed constant ChE value during treatment; the difference was not significant in both groups for the same enzymes. The data were interpreted from a functional point of view; that is they were related to both the metabolic post-aggressive state and TPN. A relationship between the rate of protein catabolism and the inductive increase of some enzymes (GOT, GPT, GlDH) was found. Whereas a final induction in the energy metabolism is suggested for other enzymes (LDH, HBDH), the alteration of CPK, AP, gamma-GT and ChE was interpreted as dependent on: (1) direct muscular trauma (CPK); (2) functional increase in relation to the duration of TPN (AP and gamma-GT); (3) possible depressed malnutritive synthesis (ChE). The improvement of the enzymatic patterns with the early use of TPN and with the improvement of clinical and nutritional conditions was emphasized. Introduction
The use of total parenteral nutrition (TPN) in intensive care has significantly improved patient survival. With the extension of experience and application of this nutritional approach, a host of technical and metabolic complications have been observed (Dudrich, McFayden, Van Buren, Ruberg & Maynard, 1972; Ryan, 1976; Sanders & Sheldon, 1976). 185
186
M. ZANELLO AND OTHERS
Some of the metabolic alterations, ‘apparent complications’, remain a continuous problem. In previous reports (Cetrullo, Castelli & Zanello, 1976; Castelli, Cetrullo & Zanello, 1978) data were presented concerning the serial changes in certain laboratory indices of hepatic and metabolic function during TPN. However, the cause of these alterations remains substantially unclear. In this report we present data concerning our most recent experience with TPN in critically ill patients in which we have attempted to correlate serial alterations in the laboratory indices of metabolic function with the clinical course of the patients. Special consideration is given to establish those alterations attributable directly to the disease state and those due to TPN. In this study we direct our attention to the following questions: (1) Is there a characteristic pattern to the alterations in the laboratory indices of metabolic function during TPN in critically ill patients? (ii) Are these alterations due to the TPN, to the pathology of the disease state or to a combination of the two? (iii) Do the alterations in the laboratory indices correlate with the clinical course of these patients? (iv) Does early use of TPN affect the laboratory changes? Patients and methods The patients population for this study consisted of 16 females and 18 males, of ages between 16 and 76 years, with one patient of age 5 years. They represent patients who underwent TPN for more than 5 days in the Intensive Care Unit (ICU) of the Institute of Anaesthesiology and Reanimation, University of Bologna, Italy, in the last year and were selected for inclusion without bias as to age (excluding newborn and infants), sex, presenting disease or clinical condition (except for patients with obvious hepatic or cardiac disease). The most frequent TPN solution used consisted of a mixture of 50% glucose and 8.5% amino acids (Freamine) approaching progressively the infusion of 750 g of glucose and 18.75 g of N per day: electrolytes and vitamins were added to the solution according to need. Hypertonic solutions were infused, according to the usual method, by means of a vena caval catheter; correct placement of catheter was controlled radiologically. In addition to the usual laboratory studies, which are necessary for the proper care of critically ill patients and conduction of TPN, such as determination of electrolytes, nitrogen balance, blood gases, urinalysis etc., activities of the plasma enzymes of particular interest in this study were measured: transaminases, more specifically glutamic-pyruvic transaminase (GPT), glutamic-oxaloacetic transaminase (GOT) and glutamic dehydrogenase (GlDH); alkaline phosphatase (AP), gamma-glutamyl transpeptidase (gamma-GT); lactic dehydrogenase (LDH), beta-hydroxybutyrate dehydrogenase (HBDH); creatine phosphokinase (CPK); pseudo-cholinesterase (ChE). Results Summary data of the patient population are presented in Tables 1 and 2. The patients are divided into two groups, designated as a ‘laboratory uncomplicated’ group (LUC) and a ‘laboratory complicated’ (LC), respectively. The criteria used for their separation
ENZYMES IN TOTAL PARENTERAL NUTRITION
187
Table 1. Diagnosis and complications of 15 ‘laboratory uncomplicated’ patients. Key (and for Table 2): B (bleeding), C (cardiocirculatory insufficiency), K (renal insufficiency), J (hyperbilirubinaemia), N (coma/neurologic disorder), 0 (surgical operation), P (peritonitis), R (respiratory insufficiency), S (sepsis), D (deceased), T (transferred).
Patient
Age (years)
Sex
Time after admission to onset of TPN (days)
C.L.
35
F
2
20
M.V. M.L.L.
72 29
F F
1 1
31 15
C.P. M.B.
24 74
M M
0
40 24
G.V. G.P.
52 66
F M
0
3
8 20
G.G.
76
M
2
20
M.V.
76
F
0
17
N.P. A.D.P. I.B.
22 72 51
M M F
2 4
40 30 6
A.L.
72
F
1
6
G.R.
66
F
1
11
G.B.B.
20
M
0
6
X
53,8
M=7 P=8
1,8(+2,3)
9
1
Duration of TPN (days)
20
Diagnosis on admission Ingestion of caustic agent Respiratory insufficiency Ingestion of caustic agent Chest/abdominal traums Cerebral vascular accident Acute pancreatitis Intestinal occlusion/ peritonitis Ingestion of caustic agent Postoperative respiratory insufficiency Multiple trauma Respiratory insufficiency Respiratory insufficiency/ myasthenia Ingestion of caustic agent Postoperative respiratory insufficiency Chest trauma
Clinical complication
Outcome T
R K
D D
O-S-B-K N O-N-K B-S-P T
T
N-R-C
D
O-R-S-B N-K-R C-R
T D T
J-K
D
O-R
T
B-R
D T = n.6 D = n.9
were based on the following observation. Initially all patients were grouped together and a mean value was calculated for each measurement for each TPN day. Although this analysis presented us with a relatively clear picture of altered laboratory indices during TPN, individual values varied very much. It was therefore decided to divide the patients into two groups on the basis of whether they showed marked alterations in at least one or more days in at least three or more of laboratory indices. It is to be emphasized that this division was initially based on the laboratory values, but it was also seen that there was a relationship between the extent of laboratory changes and the frequency of severe clinical complications, that is the LC group also showed more critical clinical course. The two groups differed also in the fact that the LC group was composed of 44% of severe trauma cases, compared with 20% in the ‘laboratory uncomplicated’ group. Analysis of the data in Tables l-2 shows that the number of patients, age and sex distribution were similar in both groups: also in both groups the mortality rate was
188 M. ZANELLO AND OTHERS Table 2.
Diagnosis and complications of 19 ‘laboratory complicated’ patients.
Patient
Age (years)
Sex
Time after admission to onset of TPN (days)
O.P. D.A. G.M. C.M. G.R. I.V. W.C. A.C.
20 12 60 17 39 54 51 54
M M F F F M M M
10 2 5 5 1 6 3 1
22 30 62 6 21 15 6 21
G.F. M.P.
36 73
M F
3 20
14 15
G.T. M.G.
69 43
M M
3 2
12
T.C.
53
F
4
29
G.C. A.L. S.I. A.N. F.D.M. F.M.
5 16 44 34 50 52
M M F F M M
21 31 69 21 10 22
x
44,6
M= 12 4,74 (+4,7) F=7
24,44
See also Table 1.
Duration of TPN Diagnosis on admission (days)
Clinical complications B-O-J N-S R-O-P-S P O-S-J-K O-J-S R O-R-K-S
T T T T D T
O-P-S-K R-C-K
D D
B-N-O S
D D
R-C-B
D
o-c
T T T D D T
I
Ingestion of alkali Head trauma Multiple trauma Chest/abdominal trauma Multiple trauma Chest/abdominal trauma Respiratory insufficiency Postoperative esophagocoloplasty Chest/abdominal trauma Mendelson synd./ megaesophagus Multiple trauma Postoperative esophagocoloplasty Peritonitis/postgastrectomy Cachexia/colectomy CO poisoning Chest/abdominal trauma Coma post-eclampsia Cerebral Vast. act. Ingestion of caustic agent
N-K-R R-O-S-P N-R-J N-B-R B-O
Outcome
:
T=lO D=3
high, as expected in critically ill patients. The LC group on the average had a longer interval between admission to the intensive care unit and the initiation of TPN than did the LUC group (means of 4.7 and 1.8 days respectively); this difference was significant (P =O.Ol). On the average the LC group also had a longer course of TPN (mean of 24.4 and 20 days/patient respectively). The data for SGOT and SGPT are presented in Figs. 1 and 2. In these Figures only the mean values for the two groups have been presented for evidence. Bearing in mind that there was a significant dispersion of individual values within each group, the initial average values for the LC group are seen to be significantly greater than the LUC group for both enzymes (P=O.O5). During the course of TPN the LC group showed an initial period of increasing activities of both enzymes followed by a slower return towards initial, and finally almost normal levels. One notes also that the extent of the increase in values is greater in the initial phase for GPT than for GOT. The LUC group, on the other hand, showed relatively constant values during the entire TPN course, remaining slightly above the normal range.
189
ENZYMES IN TOTAL PARENTERAL NUTRITION mu/ml
100
1
90.
_
i&C.
. ..---.
L.C. .
p=
GROUP GROUP
0.05
s”“‘“-I
\
20. 10.
N.V.
1
5
10
15
20
25
TPN
DAVS
Fig. 1. Plasma activity of glutamic-oxaloacetic transaminase as mean values in the ‘laboratory uncomplicated’ (LUC) and ‘laboratory complicated’ (LC) groups of patients during TPN.
-
Luc.
.----
L.C. .
GROUP GROUP
p= 0.05
N.V. 1
5
10
15
20
25 TPN
DAYS
Fig. 2. Mean plasma activities of glutamic-pyruvic transaminase in the ‘laboratory uncomplicated ‘laboratory complicated’ groups of patients during TPN.
and
Analysis of GlDH values in the same two groups (Fig. 3) showed a marked spread of individual values; because of this the difference between the means of the two groups was not always statistically significant, but the mean values presented by the LC group initially presented values which tended to increase during the first 10 days of TPN, followed by progressive decrease towards initial values in the second 10 days. The LUC group, on the other hand, presented values which tended to remain in the normal range or slightly above during the entire nutritional therapy.
190 mu/ml
M. ZANELLO
AND OTHERS
60 40.
-
L.U.C.
GROUP
c--a
L.C.
GROUP
36. 30. 26.
1
3
6
7
6
11
Fig. 3. Mean plasma activities of glutamic dehydrogenase patients during TPN.
13
16
‘17
16
21
IN.V.
TPN DAYS
and their standard deviation in the two groups of
The data concerning other indices of amino acid metabolism related also to bile excretion are in Figs. 4 and 5, The evolution of AP activity showed similar initial values for both groups of patients which fell within normal range (Fig. 4). The LC group, however, showed a relatively linear increase in AP activity over the entire course of TPN; the LUC group maintained normal levels throughout. The evolution of gamma-GT activity (Fig. 5) in the LC group was similar to that of AP activity. In this case, however, the LUC group also presented increasing values above the normal range. Also for these two enzymes statistically significant difference between the mean values for both groups was not noted. The LDH and HBDH enzymes were followed in this study as indices related to intermediate and energetic metabolism. There was great variability in the mean values in both groups during the course of TPN for both enzymes and no characteristic patterns were observed. The mean values remained generally above the upper normal limits but did not exceed twice the normal values. The initial values of CPK activity also showed marked variability for both groups, which was greater in the LC group: these initial values were significantly elevated above the normal range, especially in the LC group. Both groups also showed a tendency towards decreasing values during the TPN course, with eventual earlier return to normal in the LUC group (Fig. 6). The ChE enzyme values for the two groups are presented in Fig. 7. Both groups had initial values below the normal range and both groups subsequently showed similar evolutionary patterns during the course of therapy, i.e. subsequent return towards the lower limit of the normal range during the following 10-12 days. Surprisingly, this tendency toward normalization was more marked in the LC patients. Finally, the mean nitrogen balance was calculated for the two groups for each day of TPN (Fig. 8). Except for an initial positive peak in the LUC group, both groups showed
ENZYMES
mu/ml
IN TOTAL
PARENTERAL
191
NUTRITION
_
L.U.C. GROUP
.----.
L.C.
GROUP
300-
25O-
Y
T
N.V.
50.
1
Fig. 4. TPN.
mu/ml
Changes
5
10
in the mean alkaline
15
phosphatase
activity
1 25 TPN
20
in the two groups
DAYS
of selected patients
_
t..U.C.
.-e-B.
L.C.
during
GROUP GROUP
200.
150.
100.
50.
l-2
Fig. 5. Changes during TPN.
5-6
in the gamma-glutamyl
9-10
transpeptidase
13-14
plasma
17-16
activity
21-22
in the same groups
TPN
DAYS
of patients
a protracted period of negative balance, which lasted approximately 18 days, with subsequent tendency toward positive balance, reached earlier by LUC group. Discussion Several findings have prompted us to consider the alterations in the enzyme activity during TPN from a functional point of view. First, the patients selected for
192
M. ZANELLO AND OTHERS
mu/ml
700
900
1
500. 400. 300.
8
‘\
‘\,
200.
.... -
i&c.
oooo -----
Lc.
GROUP GROUP
0 ‘\
‘ii
iFi 50. 40. 30. N.V.
20
10
1* 0’
l-2
9-10
5-9
17-19
13-u
21-22
TPN DAYS
Fig. 6. Spread of individual values and mean changes of creating phosphokinase plasma activity in the ‘laboratory uncomplicated’ and ‘laboratory complicated’ groups during TPN.
mu/ml
_
L.U.C.
c--d
L.C.
9J
GROUP GROUP
J
f
I
N.V.
I 1
Fig. 7.
3
5
7
Mean plasma pseudocholinesterase
9
11
13
15
17
q9
21 TPN DAYS
activity in the same two groups of patients during TPN.
ENZYMES IN TOTAL PARENTERAL NUTRITION
9
_
L.u.c.
.-m-d
L.C.
193
GROUP
GROUP
N/day
1
Fig. 8.
5
10
, 15
20
25 TPN
DAYS
Changes in mean nitrogen balance for each TPN day in the two groups of patients.
this study did not have liver disease. Secondly, although both groups of patients showed alterations in plasma enzyme activities, they did not reach the levels generally associated with severe organic lesions or necrosis in either group. Thirdly, notwithstanding the difference in the enzyme patterns noted between the two groups, the mortality and recovery rates were similar. Generally alterations in plasma enzyme activity can be considered to result from four conditions: decrease in synthesis; increase in postinductive synthesis; increase in loss or leakage from cells; defective excretion (Zimmerman & Seeff, 1970; Schmidt & Schmidt, 1971; Schepartz, 1973). Some of these mechanisms come into play, for various functional reasons, in critically ill patients undergoing TPN. One of the principal disturbances in cellular function, resulting from altered energetic post-traumatic metabolism, is abnormal membrane permeability (Flear, 1970, 1974). The subsequent loss or leakage of small molecules (ions, amino acids, substrates etc.) and also larger molecules (such as small proteins) can lead to variations in the normal plasma levels of these substances (McFarlane, 1969; Courtice, Adams, Shannon & Bishop, 1974). Further variations in the plasma activities of various hepatic and muscular enzymes have been described, which reflect the response to changes in nitrogen content of the diet; these include thereonine-serine dehydrogenase (Harper, 1965); some enzymes of the urea cycle (Schimke, 1962; Das & Waterlow, 1974) and several aminotransferases including glutamic acid-oxaloacetate aminotransferase (Steigink & Baker, 1971). Probably an abundance of amino acids entering the liver causes an increase in the amount of transforming enzymes: the enzyme induction seems to be a very fundamental biological reaction. The complicated interaction, also, of multiple
194
M. ZANELLO
AND OTHERS
transaminases in cells and mitochondria has been extensively reviewed by Munro (1964, 1970). Theoretically, the elevated content in amino acids of the TPN results in a condition which is similar to that described above. Further, the increased endogenous turnover of amino acids and protein catabolism (primarily muscular), which characterize the poststress phase in critically ill patients (Cuthberson & Tiltson, 1969; Kinney, Duke, Long & Gump, 1970; Moore & Brennan, 1975) are further inductive factors in this nutritional context. The concentration of amino acids in the blood is also regulated by certain hormones such as the catecholamines, insulin, glucagon, growth hormone and glucocorticosteroids, whose circulation concentrations are also increased in the acute post-traumatic phase (Wilmore, Long, Skreen, Mason & Pruitt, 1974; Johnston, 1974; Moore & Brennan, 1975; Blackburn, Maini & Pearce, 1977). It is also known that some of these hormones have a direct action of enzyme induction (Wannemacher, 1975). Further, it has been demonstrated that there is a reduction in the levels of certain enzymes whose synthesis is depressed in the condition of malnutrition; various hepatic enzymes (which are labile proteins) (Munro, 1964), amylase and alkaline phosphatase (Srikantia, Jacob & Reddy, 1964) and pseudocholinesterase (Waterlow & Stephen, 1970). The serial measurement of plasma activity of certain enzymes in patients undergoinng TPN during the acute phase of recovery in the intensive care unit show two general evolutionary patterns. Some enzymes, e.g., GOT, GPT, GlDH, whose activity was elevated in the plasma in the initial post-acute period or in relation to a worsening clinical course, usually show a global tendency towards return to normal of values with improvement of the clinical and nutritional condition of the patient. Other enzymes, such as AP and gamma-GT, on other hand, tend to show a continuous increase in relation to the duration of TPN. These two patterns of evolution of the enzymes examined appear more pronounced in the LC group than in the LUC group as defined earlier, although both groups were treated similarly with respect to TPN. The TPN itself did not seem to have a direct effect on these enzymatic alterations, since in most cases in the LC group enzyme activities were usually significantly elevated even before the onset of TPN. The increase in plasma activity of those enzymes connected with gluconeogenesis (GOT, GPT), urea cycle (GlDH) and general energetic metabolism (LDH, HBDH, CPK) therefore seems to be related to the particular posttraumatic metabolic response of critically ill patients. The final effect of the post-traumatic catabolic reaction seems to be the production of small molecules, intermediate substrates of glucose metabolism which can be inserted into the multiple anabolic pathways of gluconeogenesis, protein and lipid synthesis or degradative pathways to yield energy (Baruh, Sherman, Kolodny & Singh, 1973; Wannemacher, 1975; Owen, Pate1 & Block, 1976), processes that necessitate various enzyme systems, including those followed in this study. The increase of cellular enzymes in the blood as a result of altered membrane permeability from unbalanced energetic metabolism due to toxic or hypoxic conditions (Flear, 1970; 1974) is supported by the finding that the level of these enzymes remained lower and returned to normal levels more quickly in the patients in which TPN was initiated sooner, possibly because this reduced the degree of enzyme induction or decreased celiular leakage by improving the energetic state of the cells.
ENZYMES
IN TOTAL PARENTERAL
NUTRITION
195
In this context TPN exerts an effect on metabolic homeostasis, which, at least theoretically, modulates the plastic and energetic cellular requirements, and therefore, also changes the orientation of cellular metabolism by reducing those conditions which promote catabolism. The alteration in transaminases activity (GOT, GPT, GlDH) is considered to reflect the extent of protein catabolism, the amount of transformation and transport of NH:, and the rate of gluconeogenesis. It seems that the increase in transaminase activity is the result of enzyme induction in response to endogenous, and perhaps exogenous, increased substrate levels, the latter due to amino acids during total parenteral nutrition. The most elevated levels of enzyme activity observed in the LC group (clearly more catabolic, as demonstrated by the considerable negative nitrogen balance: Fig. 8) seems to confirm this supposition, even though we have not measured the specific substrates of the reactions. The degradation of branched-chain amino acids resulting in the formation of glutamine (Felig, Wahren, Karl, Cerasi, Luft & Kipnis, 1973) and the liberation of alanine (Felig, Pozefsky, Marliss & Cahill, 1970; Marliss, Aoki, Pozetsky, Rost & Cahill, 1971) on the part of muscle tissue seems to be the fundamental mechanisms for the endogenous production of some of those substrates. Various amino acids, but particularly alanine, must first be converted into pyruvate in order to enter the hepatic gluconeogenetic pathway (Wannemacher, 1975), which can explain the obligatory rise in the converting enzyme, GPT (alanine to pyruvate), as noted in the more catabolic patients of the LC group. The increased activity of the converting enzyme GOT (aspartate to oxaloacetate), which was also observed in these patients, must be considered as the result of a more non-specific increase in the degradative process involving many amino acids, but which serves as a fundamental step in subsequent gluconeogenesis, and acts as the metabolic key for glucose metabolism and the Krebs cycle. The increased production of NH: by transamination with secondary acceleration of the urea cycle activity, probably induces synthesis of GlDH, which participates in the transport of ammonium radicles from the muscle to the liver, and catalyses the amination of many amino acids, as demonstrated during starvation (Das & Waterlow, 1974). In the post-traumatic state, in addition to accelerated activities of the glucosealanine cycle (Feling et al., 1970) and the glutamine cycle (Wannemacher, 1975), one notes also the induction of the Cori cycle (glucose-lactate-glucose) (Bassler, 1976). The increases in LDH activity in both groups of patients seems to indicate either an induction of alternative energetic pathways or the permanence of conditions of anaerobic metabolism. These same mechanisms may be similarly active in the elevation of the HBDH activity, the enzyme which catalyses the reaction acetoacetate-betahydroxybutyrate, making ketone bodies as an alternative energy source in post-stress metabolism (Barton, 1970). The elevated levels of CPK observed more markedly in the LC patients because of the magnitude of the values reached, the particular evolutionary patterns observed and the simultaneous presence of negative nitrogen balance (as expression of energy deficit), seem to be more related to muscular trauma than to enzyme induction and accumulation of high-energy substrates as described in other clinical conditions (Clark, 1971; Nevins, Madhukar, Bright & Lyon, 1973; Sklar & Wigand, 1973).
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The tendency towards increased activities of other enzymes, such as AP and gammaGT, in relation to the duration of TPN therapy, seems to signify an increased need for amino acid transport in the gamma-glutamyl cycle, or some functional hepatic changes, probably cholestatic in nature, even though hyperbilirubinaemia is an inconstant finding. One cannot exclude the protracted infusion of hypertonic glucose solutions or the eventual deficit or imbalance of amino acid mixtures employed in TPN as probable causes of these cholestatic-like enzyme patterns (Dubrick et al., 1972; Touloukian & Downing, 1973). However, it is important to emphasize that critically ill patients often suffer from clinical conditions which can lead to hepatic damage, cholestasis or changes in the entero-hepatic circulation of bile salts (e.g., peritonitis, septicaemia, intestinal occlusion, prolonged and massive drug treatment etc.), which cannot be neglected as possible aetiological agents for some of these enzyme alterations. The constant observation of depressed plasma ChE values is interpreted as an expression of the general reduced synthesis of this rapidly turning over enzyme probably due to effective malnutrition. This view is supported by the finding of progressively increasing levels of this enzyme related to improvement in nitrogen balance. The relatively more elevated mean values of the LC group are not easily explained. It is possible that the more frequent blood and plasma transfusions given to this group of patients served as an exogenous source of enzyme. On the other hand, the difference between the mean values for the two groups may be only apparent as it was not found to be statistically significant. Finally, comparing the individual enzyme patterns of the two groups of patients with their respective nitrogen balance curves, one notes a definitive relationship between improvement in nitrogen balance and normalization of plasma enzyme levels. The delay in the nitrogen balance becoming positive in the LC group with respect to the LUC group could be dependent on the delay in initiation of TPN in this group, and the lack of positive initial values can be considered as an expression of nitrogen deficit accumulated before initiation of nutritional parenteral support. In view of what has been discussed so far, it is now possible to answer the four questions which were posed in the Introduction to this paper. On the basis of our data, the answers are: (i) In spite of the great variability of enzyme levels between individual patients, each group (LC and LUC) showed a characteristic evolutionary pattern for some enzymes and, above all, of the transaminases. (ii) The altered levels of plasma enzymes observed in these patients were probably due to functional disturbances of the cellular activity. However, at the same time, TPN, as exogenous source of amino acids, could induce specific enzyme synthesis in relation to the metabolic tendency of the post-stress condition. (iii) The patients whose clinical course was more severe seem to have more markedly altered metabolic indices. In addition, there was usually a general improvement indicated by these laboratory findings, with improvement of the clinical conditions. (iv) Early initiation of TPN improves the pattern of laboratory findings in critically ill patients probably by several mechanisms. It is clear that patients receiving early TPN have much more improved nitrogen and energy balances. That probably leads to a decreased leakage of intracellular enzymes even if the absolute enzyme activity has been increased because of induction. According to this point of view, TPN seems to be an effective therapeutic means of control of the metabolic response to injury.
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