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Increase in superoxide dismutase after cerebrovascular accident
Life Sciences, VoW.54, No. 11, pp. 711-713, 1994 Copyright © 1994 Elsevier Science l~d Printed in the USA. All rights reserved 0024-3205/94 $6.0O + .00
Pergamon
INCREASE IN SUPEROXIDE DISMUTASE AFTER CEREBROVASCULAR ACCIDENT N. Gruener, B. Gross" O. Gozlan and M. Barak Departments of Biochemistry and "Neurology Carmel Hospital Haifa 34362, Israel
(Received in final form December 23, 1993) Summ~arv Superoxide dismutase (SOD), neuron specific enolase (NSE) and lactic dehydrogenase (LDH) were measured in the serum and cerebrospinal fluid (CSF) of ischemic cerebrovascular patients, other neurological patients and in age-matched healthy controls (serum only). The levels of SOD in the CSF or serum of the ischemic patients in the first 24 hrs after stroke were similar to the control groups. However, SOD levels in the ischemic patients increased after two days, reaching their peak values after one week (2-3 fold of the initial values). NSE showed a similar kinetics while LDH showed no change. These results suggest that oxygen radicals are formed in the ischemic patients and the increased synthesis of SOD may protect the patients from the potential damage of such radicals.
It has been hypothesized that oxygen free radicals can be formed in the cell after ischemiareperfusion injury (I). The origins of the oxygen free radicals are not clear and several mechanisms have been proposed. Apparently, the first step in the formation of such radicals is the reduction of oxygen to superoxide radicals which can lead to the appearance of other radicals by a variety of chemical and biological reactions (2,3). Of special importance is the accumulation of lipid peroxidation products which may cause structural and functional changes in the cell (4). The enzyme superoxide dismutase catalytic activity is directed toward the destruction of superoxide anion, thus preventing the radical "cascade" (2). The role of oxygen free radicals in the pathogenesis of cerebral ischemia has been intensively investigated (5-7). We present here data that show that superoxide dismutase levels in the cerebrospina[ fluid and plasma of patients who suffer a stroke increase during the first two weeks and then return to the initial values.
Materials And Methods Cerebrospinal fluid (CSF) samples were collected for clinical purposes from I_12 patients who suffered various neurological problems. Eighty seven of them had an acute stroke. The other 25 patients participated in the control group. Plasma samples were collected from the same patients and from an additional twenty three healthy controls of the same age profile (mean age 72, range 45-90, male patients 52%). CuZn superoxide dismutase concentration was measured by competitive EL]SA using antibodies against recombinant superoxide dismutase raised in rabbits. Neuron specific enolase was measured with a sandwich enzyme immunoassay, which contains monoclonal and polyclona] antibodies to NSE.
712
SOD Levels in CSF After Stroke
Voi. 54, No. 11, 1994
Results And Discussion Figure ] presents the kinetics of the superoxide dismutase levels in CSF and plasma of patients after stroke• Both show continuous increase after a lag of 24 hrs, which peak after one week in the CSF and two weeks in the plasma. At three weeks they level off to the initial concentrations. ~,9/ml
~,9/ml
)4.
•
p<05
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..............................................
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,
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i Control
1 day
2 dayS
3.7 dayS
8.14 days 15.21 dayS
Control
lday
Days after admission
• 2dayS
"
.
3-7dayS
"
..
.
'
8-14days 1 5 2 1 d a y S
Days after admLssion
Fig. 1 Superoxide dismutase concentrations in CSF (a) and plasma (b) after cerebrovascular accident•
Neuron specific enolase (NSE), a marker of neuronal pathology, was also measured along period. Figure 2 depicts the results of NSE and shows a kinetic profile similar to the one found superoxide dismutase. We could not find a change in the level of lactic dehydrogenase (data presented) which suggests that the increase in the synthesis of superoxide dismutase and NSE are a result of lysis of neurons. ng/ml 50
this for not not
r¢o~/~ .o
~.p<05 <0005
I
• p<05 "" p < 0,005
a
40
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.
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¢~r.....
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-
O. Control
1 day
2 dayS
3-7
dayS 8-14 days 15.21 dayS
Daysafteradmi~slon
Control
I day
2 days
15-21da~
Daysafteradm~s,on
Fig. 2 Neuron-specific enolase concentrations in CSF (a) and plasma (b) after cerebrovascular accident.
Vol. 54, No. 11, 1994
SOD Levels in CSF After Stroke
713
Injury to the brain due to ischemia is caused by lack of oxygenation. Evidence is now accumulating that injury can occur also during the reperfusion phase, and that the injury is due to oxygen free radicals. Superoxide dismutase is a specific scavenger of superoxide radicals. These radicals are the starting species of the "radical chain", which lead to the formation of several kinds of radicals. The presented findings suggest that increased levels of superoxide radicals after an ischemia-reperfusion cycle induce the synthesis of superoxide dismutase. The parallel kinetics of superoxide dismutase and NSE suggest that the increase of the enzyme activity originates in the neurons. We suggest that the increase in superoxide dismutase activity is a protection mechanism against the risk from radical damage. These results support the suggestion that superoxide dismutase (8, 9) or other superoxide scavengers (9) can be used in the treatment of stroke patients (10).
Acknowledqment We wish to thank Genera] Biotechno]ogy (Rehovot, Israel) for the donation of recombinant human superoxide dismutase.
References I. 2. 3. 4.
M.D. WEISFELDT, Clin. Res. 35 13-20 (1987). I. FRIDOVICH, Science 201 875-880 (1978). JM. MCCORD, N. Eng. J. Med. 312 159-163 (1985). B.C. WHITE, G.S. KRAUSE, S.D. AUST and G.E EYSTER, Ann. Emerg. Med. 14 8 0 4 - 8 0 9 (1985). 5. ES. FLAMM, H B . DEMOPOULOS, M.L. SELIGMAN, R.G. ROSER and J. RANSOHOFF, Stroke 9 445-447 (1978). 6. P.H. CHAN, J.W. SCHMIDLEY, R.A. FISHMAN and S.M. I_ANGER, Neurology 34 315-320 (1984). 7. H. KINOCHI, C.J. EPSTEIN, T. MIZUI, CARLSON, S.F. CHEN and PH. CHAN, Proc. Nat. Acad. Sci. USA 88 I I 1 5 6 - I I 1 6 2 (1991). 8. P.M. CHAN, S. LONGAR and R.A. FISHMAN, Ann. Neurol. 21 540-547 (1987). 9. M. TAGAYA, M. MATSUMOTO, K. KITAGAWA, M. NIINOBE, T. OHTSUKI, R HATA, S. OGAWA, N. HANDA, K. MIKOSHIBA and T. KAMADA, Life Sci. 51 253-259 (1992). I0. D. MARTZ, G. RAYOS, G.P SCHIELKE and A.U BETZ, Stroke 2 0 488-494 (1989).
Pergamon
INCREASE IN SUPEROXIDE DISMUTASE AFTER CEREBROVASCULAR ACCIDENT N. Gruener, B. Gross" O. Gozlan and M. Barak Departments of Biochemistry and "Neurology Carmel Hospital Haifa 34362, Israel
(Received in final form December 23, 1993) Summ~arv Superoxide dismutase (SOD), neuron specific enolase (NSE) and lactic dehydrogenase (LDH) were measured in the serum and cerebrospinal fluid (CSF) of ischemic cerebrovascular patients, other neurological patients and in age-matched healthy controls (serum only). The levels of SOD in the CSF or serum of the ischemic patients in the first 24 hrs after stroke were similar to the control groups. However, SOD levels in the ischemic patients increased after two days, reaching their peak values after one week (2-3 fold of the initial values). NSE showed a similar kinetics while LDH showed no change. These results suggest that oxygen radicals are formed in the ischemic patients and the increased synthesis of SOD may protect the patients from the potential damage of such radicals.
It has been hypothesized that oxygen free radicals can be formed in the cell after ischemiareperfusion injury (I). The origins of the oxygen free radicals are not clear and several mechanisms have been proposed. Apparently, the first step in the formation of such radicals is the reduction of oxygen to superoxide radicals which can lead to the appearance of other radicals by a variety of chemical and biological reactions (2,3). Of special importance is the accumulation of lipid peroxidation products which may cause structural and functional changes in the cell (4). The enzyme superoxide dismutase catalytic activity is directed toward the destruction of superoxide anion, thus preventing the radical "cascade" (2). The role of oxygen free radicals in the pathogenesis of cerebral ischemia has been intensively investigated (5-7). We present here data that show that superoxide dismutase levels in the cerebrospina[ fluid and plasma of patients who suffer a stroke increase during the first two weeks and then return to the initial values.
Materials And Methods Cerebrospinal fluid (CSF) samples were collected for clinical purposes from I_12 patients who suffered various neurological problems. Eighty seven of them had an acute stroke. The other 25 patients participated in the control group. Plasma samples were collected from the same patients and from an additional twenty three healthy controls of the same age profile (mean age 72, range 45-90, male patients 52%). CuZn superoxide dismutase concentration was measured by competitive EL]SA using antibodies against recombinant superoxide dismutase raised in rabbits. Neuron specific enolase was measured with a sandwich enzyme immunoassay, which contains monoclonal and polyclona] antibodies to NSE.
712
SOD Levels in CSF After Stroke
Voi. 54, No. 11, 1994
Results And Discussion Figure ] presents the kinetics of the superoxide dismutase levels in CSF and plasma of patients after stroke• Both show continuous increase after a lag of 24 hrs, which peak after one week in the CSF and two weeks in the plasma. At three weeks they level off to the initial concentrations. ~,9/ml
~,9/ml
)4.
•
p<05
p<05 "" p < 0 0 0 5
"" p <
b
0 005
..............................................
............................................... 08
°°
............
i
12.
q~.....................................
i
..... 10.
•. d
~
l
.
"...........
.
°o 06 0
8.
/
i
I
'
I
"
..
"
..
,
•
i Control
1 day
2 dayS
3.7 dayS
8.14 days 15.21 dayS
Control
lday
Days after admission
• 2dayS
"
.
3-7dayS
"
..
.
'
8-14days 1 5 2 1 d a y S
Days after admLssion
Fig. 1 Superoxide dismutase concentrations in CSF (a) and plasma (b) after cerebrovascular accident•
Neuron specific enolase (NSE), a marker of neuronal pathology, was also measured along period. Figure 2 depicts the results of NSE and shows a kinetic profile similar to the one found superoxide dismutase. We could not find a change in the level of lactic dehydrogenase (data presented) which suggests that the increase in the synthesis of superoxide dismutase and NSE are a result of lysis of neurons. ng/ml 50
this for not not
r¢o~/~ .o
~.p<05 <0005
I
• p<05 "" p < 0,005
a
40
~a
30
Z0'
i
i i
• ..
-
.!
.............
10,
.:.,.
~,~i~i. ,.:-:
'..:::i
.
'.*.?"'.2
¢~r.....
3-7da~
8-14d~
-
O. Control
1 day
2 dayS
3-7
dayS 8-14 days 15.21 dayS
Daysafteradmi~slon
Control
I day
2 days
15-21da~
Daysafteradm~s,on
Fig. 2 Neuron-specific enolase concentrations in CSF (a) and plasma (b) after cerebrovascular accident.
Vol. 54, No. 11, 1994
SOD Levels in CSF After Stroke
713
Injury to the brain due to ischemia is caused by lack of oxygenation. Evidence is now accumulating that injury can occur also during the reperfusion phase, and that the injury is due to oxygen free radicals. Superoxide dismutase is a specific scavenger of superoxide radicals. These radicals are the starting species of the "radical chain", which lead to the formation of several kinds of radicals. The presented findings suggest that increased levels of superoxide radicals after an ischemia-reperfusion cycle induce the synthesis of superoxide dismutase. The parallel kinetics of superoxide dismutase and NSE suggest that the increase of the enzyme activity originates in the neurons. We suggest that the increase in superoxide dismutase activity is a protection mechanism against the risk from radical damage. These results support the suggestion that superoxide dismutase (8, 9) or other superoxide scavengers (9) can be used in the treatment of stroke patients (10).
Acknowledqment We wish to thank Genera] Biotechno]ogy (Rehovot, Israel) for the donation of recombinant human superoxide dismutase.
References I. 2. 3. 4.
M.D. WEISFELDT, Clin. Res. 35 13-20 (1987). I. FRIDOVICH, Science 201 875-880 (1978). JM. MCCORD, N. Eng. J. Med. 312 159-163 (1985). B.C. WHITE, G.S. KRAUSE, S.D. AUST and G.E EYSTER, Ann. Emerg. Med. 14 8 0 4 - 8 0 9 (1985). 5. ES. FLAMM, H B . DEMOPOULOS, M.L. SELIGMAN, R.G. ROSER and J. RANSOHOFF, Stroke 9 445-447 (1978). 6. P.H. CHAN, J.W. SCHMIDLEY, R.A. FISHMAN and S.M. I_ANGER, Neurology 34 315-320 (1984). 7. H. KINOCHI, C.J. EPSTEIN, T. MIZUI, CARLSON, S.F. CHEN and PH. CHAN, Proc. Nat. Acad. Sci. USA 88 I I 1 5 6 - I I 1 6 2 (1991). 8. P.M. CHAN, S. LONGAR and R.A. FISHMAN, Ann. Neurol. 21 540-547 (1987). 9. M. TAGAYA, M. MATSUMOTO, K. KITAGAWA, M. NIINOBE, T. OHTSUKI, R HATA, S. OGAWA, N. HANDA, K. MIKOSHIBA and T. KAMADA, Life Sci. 51 253-259 (1992). I0. D. MARTZ, G. RAYOS, G.P SCHIELKE and A.U BETZ, Stroke 2 0 488-494 (1989).