258
Injury, 7, 258-262
Lactic dehydrogenase isoenzymes following head injury D. G. T. Thomas and T. D. Rowan Departments of Neurosurgery and Biochemistry, Southern General Hospital, Glasgow Summary Serial levels of lactic dehydrogenase (LDH) isoenzymes have been measured in 92 neurosurgical patients with damaged brains, including 77 suffering from head injury. Serum LDH isoenzymes 1 and 2 rise in the injured group and generally reflect the degree of parenchymal brain damage. The method has possible diagnostic applications, as well as some prognostic use.
INTRODUCTION TRE D[A6NOSLS and prognosis of head-injured patients shortly after injury is difficult because the extent of the damage to the brain is not evident initially: patients with different degrees of injury, as judged by their final condition, may at first show similar or identical clinical pictures (Editorial, Lancet, 1973). An index of total brain damage is therefore highly desirable, and over the past decade a number of biochemical assays have been applied to this purpose. Serum and cerebrospinal fluid levels of LDH, creatine kinase and the transaminase enzymes have been studied in patients with damaged brains and in experimental animals (Rabow et al., 1971; Hausd6rfer et al., 1973). This study examines both the outcome and the underlying pathology of a series of such patients in relation to serial L D H isoenzyme levels up to one month after
injury. L D H is a cytoplasmic enzyme present in the brain in relatively high concentration (Lindblom et al., 1967). It appears in the soluble fraction of disintegrated brain, and it is understandable that it should pass into the serum of patients with damaged brains. L D H is also present in organs
other than the brain. The heart, red cells and kidneys contain LDH-1-3, while LDH-4-5 are predominant in skeletal muscle and liver (Lindblom et al., 1967). However, the activities of isoenzymes LDH-1-3 are particularly high in the brain.
METHODS Total serum LDH was assayed by the optimized method of McQueen (1972) using an LKB 8600 reaction rate analyser at 37~ (normal range 240-525 i.u./l). In all cases the serum was removed from the cells as soon as possible and stored at room temperature until the analyses were performed, which was within 4 days. Serum LDH isoenzymes were separated by cellulose acetate electrophoresis in tris-barbitone buffer (pH 8.3) at 100 V for 18 minutes. After treatment of the strips at 37~ with lactate-coenzyme substrate in the presence of nitroblue tetrazolium and phenazonium methosulphate, the purple formazons formed were scanned in a Millipore Phoroscope at 580 nm. Alternative methods relying on the head stability of the faster moving isoenzymes were found to be less reliable. Initial blood samples were taken from the patients as soon as possible after admission, and before operation. Further samples were taken, so that levels of isoenzymes were obtained in most cases for the periods 0-24 hours, 2-3 days, 4-6 days, 7-10 days, 11-14 days after injury, and in some cases also at 15-22 days and 28-35 days.
PATIENTS The patients studied were 92 cases admitted to the Institute of Neurological Sciences, Glasgow, for
Thomas and Rowan : Lactic Dehydrogenase Isoenzymes
259
neurosurgical management; 77 had suffered head injury and 15 cerebral damage from other causes (Table I). Sera from 22 normal people were examined to establish reference levels for this laboratory (Table H).
disabled, and 21 (27 per cent) moderately disabled. Twenty-six out of 77 had a good result (34 per cent). The control patients with their brains damaged otherwise than by injury were also severely ill, 4 of 15 dying (27 per cent), 5
Table I. Diagnosis and outcome of 92 patients with brain damage Patients
Died
Extracerebral haematoma (9) Extracerebral haematoma with intracerebral damage (24) Severe intracerebral damage (23) Moderate intracerebral damage (21) Control brain-damaged patients (15)
Outcome Severely Moderately disabled disabled
1
Total
Good
1
7
8 7 1 4
6 5 2 5
8 9 3 2
2 2 15 4
21
18
23
30
Table H. Reference levels of LDH isoenzymes in 22 normal people expressed as i.u./I
Mean s.d. +
LDH- 1
LDH-2
LDH-3
LDH-4
LDH-5
125 19
184 33
46 19
30 9
16 8
The injured patients were classified as having extracerebral haematoma, mixed intracerebral damage and extracerebral haematoma, severe intracerebral damage without extracerebral haematoma, and moderate intracerebral damage (Table I). The evidence for the pathological diagnosis was obtained from investigation and operative or post-mortem findings. Several of the patients had broken limbs or other injury, but in all cases it was concluded that brain damage was the major injury. The outcome, reviewed up to 3 months from injury, is specified as dead, severely disabled, moderately disabled or good, as described by Jennett and Bond (1975). The severely disabled are not able to lead an independent existence, and have persisting neurological deficits. The moderately disabled are out of institutional care and generally have some persisting neurological deficit, for example hemiparesis or ataxia. Those termed good are apparently free of symptoms and signs attributable to their cer~ebral damage. The head injuries were fairly severe, with 17 out of 77 (22 per cent) dying, 13 (17 per cent) severely
(33 per cent) severely disabled, 2 (13 per cent) moderately disabled and 4 (27 per cent) with a good result.
RESULTS The levels of LDH-1 and, to a lesser extent, of LDH-2 and-3 did rise following head injury. The extent and time course of the rise in enzyme levels varied according to the outcome for different groups of patients. The pattern of serial L D H isoenzyme levels for those 17 patients who died following head injury is shown in Fig. 1. In the patients who died, the mean serum LDH-1 level shows a biphasic curve with peaks at 2-3 days and again at 11-14 days. LDH-2 also follows this pattern, whereas LDH-3 tended to rise until day 2 or 3 and then not to show a further peak. LDH-1 appears to show the highest rise compared with the normal range, and in the other groups of injured patients it also appears to be the most labile isoenzyme. LDH-4 and LDH-5 did not rise in the injured patients, whatever the outcome. The pattern of LDH-1 levels following injury for patients placed into four categories by
260
Injury: the British Journal of Accident Surgery Vol. 7/No. 4
LDH ISOENZYMES1-5 ( i.u.lL ) 600
500.
DEAD
z,O0 9 300 9
~
i
200
100
o
;
2'-3
4'-6
7-'~ 11"14
TIME AFTER INJURY (
days
is
~s
)
_FIE. l. Mean levels of isoenzymes ]-5 of L D H (i.u./l) in the serum of patients who died following head injury. severity of outcome is shown in Fig. 2. The 13 severely disabled show a slightly later peak compared with those who died, at 4-6 days, and their levels tend to remain up rather than follow a biphasic pattern. The 21 moderately disabled injured patients show a definite but less marked rise in serum levels, while those with a good survival have mean levels just at the upper limit of the normal range.
the mean serum LDH-1 levels for the injured patients are shown divided into four categories by pathological diagnosis. The cases with extracerebral haematoma show a rise to a peak at 7-10 days, and subsequent gradual decline, but these are not statistically significantly different from brain-damaged control patients (Student's t test P>0.05). The 24 cases with mixed extracerebral clot and disruption of brain, however, showed increased LDH-1 levels earlier, within the first 2-3 days, which persisted. The early rise from day 1 to days 4-6 in this group is significant (P~s
z-'3
4'-6
7"1o 11"14 A5
~s
Fig. 3. Mean level of isoenzyme LDH-1 (i.u./l) in the serum of patients following head injury, grouped according to type of brain damage; and in control patients with brain damage due to other causes. * Indicates levels significantly different from control group (Student's t test P15
>28
0
;
TIME AFTER INJURY ( doys ) n = t =
18 1"35
28 1.z~9
25 109
24 1'86
13 0"85
12 0'50
n =
6 0"77
in the serum of patients moderately disabled and in those with good survival following head injury. LDH l ( i , / u . / I ) 600 -
* P
Injury, 7, 258-262
Lactic dehydrogenase isoenzymes following head injury D. G. T. Thomas and T. D. Rowan Departments of Neurosurgery and Biochemistry, Southern General Hospital, Glasgow Summary Serial levels of lactic dehydrogenase (LDH) isoenzymes have been measured in 92 neurosurgical patients with damaged brains, including 77 suffering from head injury. Serum LDH isoenzymes 1 and 2 rise in the injured group and generally reflect the degree of parenchymal brain damage. The method has possible diagnostic applications, as well as some prognostic use.
INTRODUCTION TRE D[A6NOSLS and prognosis of head-injured patients shortly after injury is difficult because the extent of the damage to the brain is not evident initially: patients with different degrees of injury, as judged by their final condition, may at first show similar or identical clinical pictures (Editorial, Lancet, 1973). An index of total brain damage is therefore highly desirable, and over the past decade a number of biochemical assays have been applied to this purpose. Serum and cerebrospinal fluid levels of LDH, creatine kinase and the transaminase enzymes have been studied in patients with damaged brains and in experimental animals (Rabow et al., 1971; Hausd6rfer et al., 1973). This study examines both the outcome and the underlying pathology of a series of such patients in relation to serial L D H isoenzyme levels up to one month after
injury. L D H is a cytoplasmic enzyme present in the brain in relatively high concentration (Lindblom et al., 1967). It appears in the soluble fraction of disintegrated brain, and it is understandable that it should pass into the serum of patients with damaged brains. L D H is also present in organs
other than the brain. The heart, red cells and kidneys contain LDH-1-3, while LDH-4-5 are predominant in skeletal muscle and liver (Lindblom et al., 1967). However, the activities of isoenzymes LDH-1-3 are particularly high in the brain.
METHODS Total serum LDH was assayed by the optimized method of McQueen (1972) using an LKB 8600 reaction rate analyser at 37~ (normal range 240-525 i.u./l). In all cases the serum was removed from the cells as soon as possible and stored at room temperature until the analyses were performed, which was within 4 days. Serum LDH isoenzymes were separated by cellulose acetate electrophoresis in tris-barbitone buffer (pH 8.3) at 100 V for 18 minutes. After treatment of the strips at 37~ with lactate-coenzyme substrate in the presence of nitroblue tetrazolium and phenazonium methosulphate, the purple formazons formed were scanned in a Millipore Phoroscope at 580 nm. Alternative methods relying on the head stability of the faster moving isoenzymes were found to be less reliable. Initial blood samples were taken from the patients as soon as possible after admission, and before operation. Further samples were taken, so that levels of isoenzymes were obtained in most cases for the periods 0-24 hours, 2-3 days, 4-6 days, 7-10 days, 11-14 days after injury, and in some cases also at 15-22 days and 28-35 days.
PATIENTS The patients studied were 92 cases admitted to the Institute of Neurological Sciences, Glasgow, for
Thomas and Rowan : Lactic Dehydrogenase Isoenzymes
259
neurosurgical management; 77 had suffered head injury and 15 cerebral damage from other causes (Table I). Sera from 22 normal people were examined to establish reference levels for this laboratory (Table H).
disabled, and 21 (27 per cent) moderately disabled. Twenty-six out of 77 had a good result (34 per cent). The control patients with their brains damaged otherwise than by injury were also severely ill, 4 of 15 dying (27 per cent), 5
Table I. Diagnosis and outcome of 92 patients with brain damage Patients
Died
Extracerebral haematoma (9) Extracerebral haematoma with intracerebral damage (24) Severe intracerebral damage (23) Moderate intracerebral damage (21) Control brain-damaged patients (15)
Outcome Severely Moderately disabled disabled
1
Total
Good
1
7
8 7 1 4
6 5 2 5
8 9 3 2
2 2 15 4
21
18
23
30
Table H. Reference levels of LDH isoenzymes in 22 normal people expressed as i.u./I
Mean s.d. +
LDH- 1
LDH-2
LDH-3
LDH-4
LDH-5
125 19
184 33
46 19
30 9
16 8
The injured patients were classified as having extracerebral haematoma, mixed intracerebral damage and extracerebral haematoma, severe intracerebral damage without extracerebral haematoma, and moderate intracerebral damage (Table I). The evidence for the pathological diagnosis was obtained from investigation and operative or post-mortem findings. Several of the patients had broken limbs or other injury, but in all cases it was concluded that brain damage was the major injury. The outcome, reviewed up to 3 months from injury, is specified as dead, severely disabled, moderately disabled or good, as described by Jennett and Bond (1975). The severely disabled are not able to lead an independent existence, and have persisting neurological deficits. The moderately disabled are out of institutional care and generally have some persisting neurological deficit, for example hemiparesis or ataxia. Those termed good are apparently free of symptoms and signs attributable to their cer~ebral damage. The head injuries were fairly severe, with 17 out of 77 (22 per cent) dying, 13 (17 per cent) severely
(33 per cent) severely disabled, 2 (13 per cent) moderately disabled and 4 (27 per cent) with a good result.
RESULTS The levels of LDH-1 and, to a lesser extent, of LDH-2 and-3 did rise following head injury. The extent and time course of the rise in enzyme levels varied according to the outcome for different groups of patients. The pattern of serial L D H isoenzyme levels for those 17 patients who died following head injury is shown in Fig. 1. In the patients who died, the mean serum LDH-1 level shows a biphasic curve with peaks at 2-3 days and again at 11-14 days. LDH-2 also follows this pattern, whereas LDH-3 tended to rise until day 2 or 3 and then not to show a further peak. LDH-1 appears to show the highest rise compared with the normal range, and in the other groups of injured patients it also appears to be the most labile isoenzyme. LDH-4 and LDH-5 did not rise in the injured patients, whatever the outcome. The pattern of LDH-1 levels following injury for patients placed into four categories by
260
Injury: the British Journal of Accident Surgery Vol. 7/No. 4
LDH ISOENZYMES1-5 ( i.u.lL ) 600
500.
DEAD
z,O0 9 300 9
~
i
200
100
o
;
2'-3
4'-6
7-'~ 11"14
TIME AFTER INJURY (
days
is
~s
)
_FIE. l. Mean levels of isoenzymes ]-5 of L D H (i.u./l) in the serum of patients who died following head injury. severity of outcome is shown in Fig. 2. The 13 severely disabled show a slightly later peak compared with those who died, at 4-6 days, and their levels tend to remain up rather than follow a biphasic pattern. The 21 moderately disabled injured patients show a definite but less marked rise in serum levels, while those with a good survival have mean levels just at the upper limit of the normal range.
the mean serum LDH-1 levels for the injured patients are shown divided into four categories by pathological diagnosis. The cases with extracerebral haematoma show a rise to a peak at 7-10 days, and subsequent gradual decline, but these are not statistically significantly different from brain-damaged control patients (Student's t test P>0.05). The 24 cases with mixed extracerebral clot and disruption of brain, however, showed increased LDH-1 levels earlier, within the first 2-3 days, which persisted. The early rise from day 1 to days 4-6 in this group is significant (P~s
z-'3
4'-6
7"1o 11"14 A5
~s
Fig. 3. Mean level of isoenzyme LDH-1 (i.u./l) in the serum of patients following head injury, grouped according to type of brain damage; and in control patients with brain damage due to other causes. * Indicates levels significantly different from control group (Student's t test P15
>28
0
;
TIME AFTER INJURY ( doys ) n = t =
18 1"35
28 1.z~9
25 109
24 1'86
13 0"85
12 0'50
n =
6 0"77
in the serum of patients moderately disabled and in those with good survival following head injury. LDH l ( i , / u . / I ) 600 -
* P
