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A method for demonstrating shortened platelet survival utilizing recovery from aspirin effect
350
March 1974 The Journal o f P E D I A T R I C S
A methodfor demonstrating shortened platelet survival utilizing recoveryfrom aspirin effect `4 method for demonstrating shortened platelet survival was devised by utilizing the effect of aspirin on platelet aggregation. Twenty control subjects had complete recovery of platelet aggregation four to seven days following ingestion o f aspirin,"seven subjects with previous histories o f thrombocytopenia thought to be due to rapid platelet turnover had complete recovery from drug effect within two to three days. The aggregation recovery time following drug ingestion appears to be a simple method o f demonstrating the presence o f a compensated thrombocytolytic state using small amounts o f blood without exposing the patient to radioactive materials. In addition it may permit differentiation o f thrombocytopenia due to inCreasedplatelet destruction from thrombocytopenia secondary to platelet pooling within the spleen.
A l l e n D . S c h w a r t z , M . D . , Chicago, Ill.
THE INGESTION of acetylsalicylic acid results in a permanent defect of the platelet-release mechanism, resulting in a measurable abnormality of platelet aggregationJ -3 This defect is not corrected until a small number of platelets which have not been exposed to the drug enter into the circulation from the bone marrow. 2 A method of demonstrating shortened platelet survival and the presence of a compensated thrombocytolytic state was devised by measuring the recovery of platelet aggregation from this drug effect. MATERIALS
AND METHODS
Twenty normal control subjects were instructed to take no medication, including aspirin, for at least one week prior to being studied. Each subject then ingested 10 grains of aspirin after his platelet aggregation was demonstrated to be normal. Platelet aggregation was From the Department of Pediatrics, Northwestern University, and the Children's Memorial Hospital. Supported in part by United States Public Health Service Grant R R 05475 and a grant from the Otho S. ,4. Spragg Institute. A number o f the patients were studied in the Children's Clinical Research Center, supported by United States Public Health Service Clinical Research Center Grant FR -199. Reprint address:Children"sMemoriaIHospital, 2300 Children's Plaza, Chicago, IlL 60614.
Vol. 84, No. 3. pp. 350-354
measured two to four hours after aspirin ingestion and at 24 hour intervals until it had returned to normal. The subjects were told to take no medication during the study period. Eight patients with a history of thrombocytopenia probably due to shortened platelet survival were studied in the same manner. At the time of study four had normal platelet counts, one had thromhocytosis, and three had platelet counts below normal levels. Five had evidence of autoimmune thrombocytopenia, two had thrombocytopenia believed to be secondary to hypersplenism, and one had a history of thrombocytopenia believed to be the result of cardiac valvular disease. Abbreviations used PRP: platelet-rich plasma PPP: platetet-poor plasma ITP: immune thrombocytopenic purpura Blood was collected in a plastic syringe through a No. 20 needle and 4.5 ml. were added to a plastic tube containing 0.5 ml. of 3.2 per cent buffered citrate anticoagulant. The blood was centrifuged at 500 r.p.m, for 15 minutes at room temperature. A 1 ml. amount of the supernate platelet-rich plasma (PRP) was removed and the remaining sample centrifuged at 3,400 r.p.m, for I0 minutes to prepare platelet-poor plasma (PPP). Platelet
Volume 84 Number 3
aggregation was measured by a modification of the turbidimetric method of Born and Cross 4 with a Chrono-log platelet aggregometer Model 300. The aggregating agent was prepared by adding 1 ml. of epinephrine U.S.P. to 4.5 ml. of isotonic saline solution. PRP was constantly stirred at 1,200 r.p.m, by a Tefloncoated magnetic stir bar with the temperature maintained at 37 ~ C. To 0.5 ml. of PRP was added 0.025 ml. of the aggregating agent, resulting in a final concentration of 4• epinephrine. Increase in light transmission occurring during platelet aggregation was recorded on a Heath Model EU-20 V strip chart recorder after PRP was set as 0 per cent transmission and PPP, with 0.025 ml. of epinephrine added, set as 100 per'cent transmission. All aggregation studies were performed within one hour of the time of blood collection. Three patients with a history of autoimmune thrombocytopenia and one control subject had platelet survival determined from the disappearance of 51Cr-labeled autologous platelets based on a modification of previously described methods, s, 6 The 5~Cr-tabeled ptatelets were infused into the subjects immediately prior to their ingestion of aspirin, and the recovery of platelet aggregation from aspirin effect was measured simultaneously with the 51Cr platelet survival time. The aspirin was given after the infusion of the radioactively labeled platelets to insure that transfused platelets were also affected by the drug. Platelet counts were performed in duplicate using the method of Brecher and Cronkite. 7 Daily counts were obtained on the eight patients on the days on which aggregation studies were performed. RESULTS Control subjects. All of the control subjects had 75 to 100 per cent aggregation prior to aspirin ingestion, with aggregation being completed within three to four minutes after the addition of epinephrine. The second wave of aggregation was abolished after aspirin ingestion and did not revert to a normal pattern for four to seven days (Fig. 1). This finding is similar to the results reported by others. 8, 9 A n example of the return of platelet aggregation to normal is demonstrated in Fig. 2. It should be noted that although platelet aggregation was complete by day 3 in this subject, the rate of aggregation was abnormally retarded. A normal pattern was not achieved until day 4. The one control subject who had a 51Cr platelet survival time during his aggregation studies had a half-life of 5.2 days (normal T 89 = 3.3 to 5.3 days). Platelet aggregation returned to normal four days after aspirin ingestion in this individual. This observation is in agree-
Measurement o f platelet survival
351
10 9 7 kL~ uJ 6 ~
5
A II1 ~ a
D
z
E G 1
2
4
3
5 6
7
DAYS FOLLOWING ASPIRIN INGESTION
Fig. 1. Complete recovery of platelet aggregation following aspirin ingestion. Shaded boxes represent control subjects and lettered boxes represent patients listed in Table I. 100-
9080-
PRE-ASPIRIN Y S
~
70~
60-
oo. /// ,o.
30- ~
///
/
1DA,
3HOURS
2010" TIME(Minutes) Fig, 2. Representative tracings of platelet-aggregation studies in a control subject following aspirin ingestion demonstrating eventual return to the preaspirin aggregation pattern. merit with previous reports that aspirin does not appear to affect the 51Cr platelet survival time in the normal subject. 1~ Patients. Results of the 51Cr platelet survival times and aggregation studies are recorded in Table I. Patients A, B, and C, each with a history of chronic immune thrombocytopenic purpura (ITP), had shortened 51Cr platelet survival times and rapid recovery of aggregation from aspirin effect. Typical aggregation patterns from Patient A are shown in Fig. 3. Patients D and F, with a
35 2
Schwartz
The Journal o f Pediatrics March 1974
T a b l e I. Laboratory data and summaries of histories o f patients studied
Patient
Platelet count (normal = 150,000400,000/cu. ram.)
R eco very from A SA (normal = 4- 7 days)
Sl Cr platelet survival (normal T 89= 3.35.3 days)
A
450,000
2
2.6
B
203,000
3
1.5
C
106,000
3
1.8
D
150,000
2
E
190,000
2
F
900,000
3
G
90,~0
2
H
140,000
5
100-
9
PRE-ASPIRIN
908070-
//
Z
o_
60-
o~ O)
s
=E 5 0 U~ Z ,r n,- 4 0 I-p30_J
20-
Chronic ITP and Coombs' + hemolytic anemia (Evans' syndrome); postsplenectom y Chronic ITP; taking azathioprine; postsplenectomy Chronic ITP; postsplenectomy Chronic ITP; postsplenectom y Rheumatic cardiac disease with aortic stenosis; postoperative for mitral valve commissurotomy; history of platelet counts less than 100,000/cu. mm. Systemic lupus erythematosus; Coomhs' + hemolytic anemia and ITP; postsplenectomy Thalassemia major with massive splenomegaly Sickle-cell anemia with splenomegaly
history o f immune thrombocytopenia, also had a rapid recovery of platelet aggregation. Patient E had a history o f platelet counts below 100,000 per cubic millimeter for a period of time following cardiac surgery, but had a normal platelet count at the time aggregation studies were performed. This patient's platelets showed rapid recovery from the effects of aspirin. Of the two patients with thrombocytopenia believed to be secondary to hypersplenism, patient G had a rapid recovery of aggregation and Patient H recovered after 5 days, a value comparable with that of the normal control subjects. No patient had a significant change in platelet number during the study period. DISCUSSION
10-
o
Remarks
;
~
~
~
k
TIME (Minutes)
Fig. 3. Tracings of platelet-aggregation studies in a patient with Evans' syndrome (Patient A) following aspirin ingestion demonstrating a rapid return to the preaspirin aggregation pattern.
The addition of epinephrine to platelet-rich plasma results in a primary wave o f platelet aggregation. Endogenous adenosine diphosphate and other constituents are then released from normal platelets, leading to a secondary wave of irreversible platelet aggregation. A number of drugs, including aspirin, inhibit the plateletrelease reaction preventing the second wave of aggrega-
Volume 84 Number 3
tion. Once induced, this platelet defect is permanent and is corrected only when new platelets that have not been exposed to the drug have entered into the circulation. The present method of estimating platelet survival was devised by taking advantage of this effect of aspirin.on platelet function. All of the control subjects studied did not have return of normal platelet aggregation until four to seven days following ingestion of 10 grains of aspirin. This period of time required for complete return of function is the same as that reported by others. 9 The eight patients studied were suspected of having shortened platelet survival times because of their previous medical histories. Four had a history of idiopathic thrombocytopenic purpura, one had thrombocytopenia associated with systemic lupus erythematosus, and two had thrombocytopenia believed to be secondary to splenomegaly. The remaining patient had severe cardiac valvular disease which has been shown to result in platelet destruction and rapid platelet turnover, l~ 11 She had documented thrombocytopenia in the past. Seven of the eight patients had rapid recovery of platelet aggregation following aspirin ingestion, suggesting that there was a more rapid return into the circulation of undamaged platelets in those individuals. The one patient with sickle-cell anemia, mild thrombocytopenia, and a large spleen (Patient H) had complete return of function in five days, suggesting splenic platelet pooling rather than increased platelet destruction to be the etiology of his thrombocytopenia. This mechanism of producing thrombocytopenia has been well documented.12 There are two conditions which theoretically could have invalidated the present study. The first is the possibility that aspirin might interfere with the mechanisms of platelet destruction in the patients, thus tending to correct the abnormally shortened platelet survival time. It has already been demonstrated that aspirin does not appear to affect platelet survival in the normal individual) ~ The shortened 51Cr survival times in the patients taking aspirin in the present study, in addition to the lack of rise of platelet number in the thrombocytopenic patients, indicate that aspirin in the dosage used does not appear to correct an abnormally shortened platelet survival in the disease states studied. Previous studies have also demonstrated an inability of aspirin to correct shortened platelet survival in patients with prosthetic heart valves, unless this drug is used in combination with dipyridamole, it The second possible problem encountered would have been abnormal platelet aggregation secondary to the disease state itself. One study of patients with chronic idiopathic thrombocytopenic purpura d e m o n s t r a t e d
Measurement o f platelet survival
353
qualitative platelet abnormalities, including impaired epinephrine-induced aggregation, x3 This finding could not be demonstrated in the patients in this report, all of whom had normal aggregation prior to aspirin ingestion. This apparent discrepancy could be due to the marked difference between the concentrations of the aggregating agents used in the two studies. Only a small percentage of platelets is required to correct the aspirin-induced aggregation defect. It has been shown that the addition of only 10 per cent undamaged platelets to 90 per cent aspirin-treated platelets will completely correct aggregation? As little as five per cent undamaged platelets added to aspirin-treated platelets resulted in normal aggregation curves in this laboratory. Because normal platelet survival is about seven to 10 days, approximately 10 to 15 per cent newly produced platelets should enter the circulation every day. The fact that it requires four to seven days rather than one day for aggregation to recover completely in the normal person suggests that the megakaryocyte pool is also damaged by aspirin and abnormal platelets are being released into the circulation for a period of time. The production of platelets is believed to be regulated by a humoral stimulator which increases the size and number of megakaryocytes in the bone marrow.laThus a larger megakaryocyte pool may be susceptible to aspirin damage in states of compensated thrombocytopenia and create the apparent paradox of nearly normal aggregation recovery time after aspirin ingestion despite the markedly shortened 51Cr platelet survival times. This may explain the results obtained in Patients A, B, and C. Patients B and C had shorter 51Cr platelet survival times than Patient A, yet Patient A required only two days to recover from aspirin while Patients B and C required three days. Thus this method does not appear to be able to accurately quantitate platelet survival time. It does appear to be useful in demonstrating the presence of a compensated thrombocytolytic state, and of differentiating the thrombocytopenia of splenic platelet pooling from the thrombocytopenia due to increased platelet destruction. REFERENCES
1. Evans, G., Packham, M. A., Nishizawa, E. E., Mustard, J. F., and Murphy, E. A.: The effect of acetylsalicylic acid on platelet function, J. Exp. Med. 128: 877, 1968. 2. O'Brien, J. R.: Effects of salicylates on human platelets, Lancet 1: 779, 1968. 3. Zucker, M. B., and Peterson, J.: Inhibition of adenosine diphosphate-induced secondary aggregation and other platelet functions by acetylsalicylic acid ingestion, Proc. Soc. Exp. Biol. Med. 127: 547, 1968. 4. Born, G. V. R., and Cross, M. J.: The aggregation of blood platelets, J. Physiol. 168: 178, 1963.
354
5.
6.
7.
8.
9.
Schwartz
Murphy, S., Oski, F. A., and Gardner, F. H.: Hereditary thrombocytopenia with an intrinsic platelet defect, N. Engl. J. Med. 281: 857, 1969. Abrahamsen, A. F.: A modification of the technique for 51Cr-labelling of blood platelets giving increased circulating platelet radioactivity, Scand. J. Haematol. 5: 53, 1968. Brecher, G., and Cronkite, E. P.: Morphology and enumeration of human blood platelets, J. Appl. Physiol. 3: 365, 1950. Weiss, H. J., and Aledort, L. M.: Impaired platelet/connective tissue reaction in man after aspirin ingestion, Lancet 2: 495, 1967. Weiss, H. J., Aledort, L. M., and Kochwa, S.: The effect of salicylates on the hemostatic properties of platelets in man, J. Clin. Invest. 47: 2169, 1968.
The Journal of Pediatrics March 1974
10. Harker, L. A., and Slichter, S. J.: Platelet and fibrinogen consumption in man, N. Engl. J. Med. 20: 999, 1972. 11. Harker, L. A., and Slichter, S. J.: Studies of platelet and fibrinogen kinetics in patients with prosthetic heart valves, N. Engl. J. Med. 283: 1302, 1970. 12. Aster, R. H.: Pooling of platelets in the spleen: Role in the pathogenesis of"hypersplenic" thrombocytopenia, J. Clin. Invest. 45: 645, 1966. 13. Clancy, R., Jenkins, E., and Firkin, B.: Qualitative platelet abnormalities in idiopathic thrombocytopenic purpura, N. Engl. J. Med. 286: 622, 1972. 14. Harker, L. A.: Current concepts: Platelet production, N. Engl. J. Med. 282: 492, 1970.
March 1974 The Journal o f P E D I A T R I C S
A methodfor demonstrating shortened platelet survival utilizing recoveryfrom aspirin effect `4 method for demonstrating shortened platelet survival was devised by utilizing the effect of aspirin on platelet aggregation. Twenty control subjects had complete recovery of platelet aggregation four to seven days following ingestion o f aspirin,"seven subjects with previous histories o f thrombocytopenia thought to be due to rapid platelet turnover had complete recovery from drug effect within two to three days. The aggregation recovery time following drug ingestion appears to be a simple method o f demonstrating the presence o f a compensated thrombocytolytic state using small amounts o f blood without exposing the patient to radioactive materials. In addition it may permit differentiation o f thrombocytopenia due to inCreasedplatelet destruction from thrombocytopenia secondary to platelet pooling within the spleen.
A l l e n D . S c h w a r t z , M . D . , Chicago, Ill.
THE INGESTION of acetylsalicylic acid results in a permanent defect of the platelet-release mechanism, resulting in a measurable abnormality of platelet aggregationJ -3 This defect is not corrected until a small number of platelets which have not been exposed to the drug enter into the circulation from the bone marrow. 2 A method of demonstrating shortened platelet survival and the presence of a compensated thrombocytolytic state was devised by measuring the recovery of platelet aggregation from this drug effect. MATERIALS
AND METHODS
Twenty normal control subjects were instructed to take no medication, including aspirin, for at least one week prior to being studied. Each subject then ingested 10 grains of aspirin after his platelet aggregation was demonstrated to be normal. Platelet aggregation was From the Department of Pediatrics, Northwestern University, and the Children's Memorial Hospital. Supported in part by United States Public Health Service Grant R R 05475 and a grant from the Otho S. ,4. Spragg Institute. A number o f the patients were studied in the Children's Clinical Research Center, supported by United States Public Health Service Clinical Research Center Grant FR -199. Reprint address:Children"sMemoriaIHospital, 2300 Children's Plaza, Chicago, IlL 60614.
Vol. 84, No. 3. pp. 350-354
measured two to four hours after aspirin ingestion and at 24 hour intervals until it had returned to normal. The subjects were told to take no medication during the study period. Eight patients with a history of thrombocytopenia probably due to shortened platelet survival were studied in the same manner. At the time of study four had normal platelet counts, one had thromhocytosis, and three had platelet counts below normal levels. Five had evidence of autoimmune thrombocytopenia, two had thrombocytopenia believed to be secondary to hypersplenism, and one had a history of thrombocytopenia believed to be the result of cardiac valvular disease. Abbreviations used PRP: platelet-rich plasma PPP: platetet-poor plasma ITP: immune thrombocytopenic purpura Blood was collected in a plastic syringe through a No. 20 needle and 4.5 ml. were added to a plastic tube containing 0.5 ml. of 3.2 per cent buffered citrate anticoagulant. The blood was centrifuged at 500 r.p.m, for 15 minutes at room temperature. A 1 ml. amount of the supernate platelet-rich plasma (PRP) was removed and the remaining sample centrifuged at 3,400 r.p.m, for I0 minutes to prepare platelet-poor plasma (PPP). Platelet
Volume 84 Number 3
aggregation was measured by a modification of the turbidimetric method of Born and Cross 4 with a Chrono-log platelet aggregometer Model 300. The aggregating agent was prepared by adding 1 ml. of epinephrine U.S.P. to 4.5 ml. of isotonic saline solution. PRP was constantly stirred at 1,200 r.p.m, by a Tefloncoated magnetic stir bar with the temperature maintained at 37 ~ C. To 0.5 ml. of PRP was added 0.025 ml. of the aggregating agent, resulting in a final concentration of 4• epinephrine. Increase in light transmission occurring during platelet aggregation was recorded on a Heath Model EU-20 V strip chart recorder after PRP was set as 0 per cent transmission and PPP, with 0.025 ml. of epinephrine added, set as 100 per'cent transmission. All aggregation studies were performed within one hour of the time of blood collection. Three patients with a history of autoimmune thrombocytopenia and one control subject had platelet survival determined from the disappearance of 51Cr-labeled autologous platelets based on a modification of previously described methods, s, 6 The 5~Cr-tabeled ptatelets were infused into the subjects immediately prior to their ingestion of aspirin, and the recovery of platelet aggregation from aspirin effect was measured simultaneously with the 51Cr platelet survival time. The aspirin was given after the infusion of the radioactively labeled platelets to insure that transfused platelets were also affected by the drug. Platelet counts were performed in duplicate using the method of Brecher and Cronkite. 7 Daily counts were obtained on the eight patients on the days on which aggregation studies were performed. RESULTS Control subjects. All of the control subjects had 75 to 100 per cent aggregation prior to aspirin ingestion, with aggregation being completed within three to four minutes after the addition of epinephrine. The second wave of aggregation was abolished after aspirin ingestion and did not revert to a normal pattern for four to seven days (Fig. 1). This finding is similar to the results reported by others. 8, 9 A n example of the return of platelet aggregation to normal is demonstrated in Fig. 2. It should be noted that although platelet aggregation was complete by day 3 in this subject, the rate of aggregation was abnormally retarded. A normal pattern was not achieved until day 4. The one control subject who had a 51Cr platelet survival time during his aggregation studies had a half-life of 5.2 days (normal T 89 = 3.3 to 5.3 days). Platelet aggregation returned to normal four days after aspirin ingestion in this individual. This observation is in agree-
Measurement o f platelet survival
351
10 9 7 kL~ uJ 6 ~
5
A II1 ~ a
D
z
E G 1
2
4
3
5 6
7
DAYS FOLLOWING ASPIRIN INGESTION
Fig. 1. Complete recovery of platelet aggregation following aspirin ingestion. Shaded boxes represent control subjects and lettered boxes represent patients listed in Table I. 100-
9080-
PRE-ASPIRIN Y S
~
70~
60-
oo. /// ,o.
30- ~
///
/
1DA,
3HOURS
2010" TIME(Minutes) Fig, 2. Representative tracings of platelet-aggregation studies in a control subject following aspirin ingestion demonstrating eventual return to the preaspirin aggregation pattern. merit with previous reports that aspirin does not appear to affect the 51Cr platelet survival time in the normal subject. 1~ Patients. Results of the 51Cr platelet survival times and aggregation studies are recorded in Table I. Patients A, B, and C, each with a history of chronic immune thrombocytopenic purpura (ITP), had shortened 51Cr platelet survival times and rapid recovery of aggregation from aspirin effect. Typical aggregation patterns from Patient A are shown in Fig. 3. Patients D and F, with a
35 2
Schwartz
The Journal o f Pediatrics March 1974
T a b l e I. Laboratory data and summaries of histories o f patients studied
Patient
Platelet count (normal = 150,000400,000/cu. ram.)
R eco very from A SA (normal = 4- 7 days)
Sl Cr platelet survival (normal T 89= 3.35.3 days)
A
450,000
2
2.6
B
203,000
3
1.5
C
106,000
3
1.8
D
150,000
2
E
190,000
2
F
900,000
3
G
90,~0
2
H
140,000
5
100-
9
PRE-ASPIRIN
908070-
//
Z
o_
60-
o~ O)
s
=E 5 0 U~ Z ,r n,- 4 0 I-p30_J
20-
Chronic ITP and Coombs' + hemolytic anemia (Evans' syndrome); postsplenectom y Chronic ITP; taking azathioprine; postsplenectomy Chronic ITP; postsplenectomy Chronic ITP; postsplenectom y Rheumatic cardiac disease with aortic stenosis; postoperative for mitral valve commissurotomy; history of platelet counts less than 100,000/cu. mm. Systemic lupus erythematosus; Coomhs' + hemolytic anemia and ITP; postsplenectomy Thalassemia major with massive splenomegaly Sickle-cell anemia with splenomegaly
history o f immune thrombocytopenia, also had a rapid recovery of platelet aggregation. Patient E had a history o f platelet counts below 100,000 per cubic millimeter for a period of time following cardiac surgery, but had a normal platelet count at the time aggregation studies were performed. This patient's platelets showed rapid recovery from the effects of aspirin. Of the two patients with thrombocytopenia believed to be secondary to hypersplenism, patient G had a rapid recovery of aggregation and Patient H recovered after 5 days, a value comparable with that of the normal control subjects. No patient had a significant change in platelet number during the study period. DISCUSSION
10-
o
Remarks
;
~
~
~
k
TIME (Minutes)
Fig. 3. Tracings of platelet-aggregation studies in a patient with Evans' syndrome (Patient A) following aspirin ingestion demonstrating a rapid return to the preaspirin aggregation pattern.
The addition of epinephrine to platelet-rich plasma results in a primary wave o f platelet aggregation. Endogenous adenosine diphosphate and other constituents are then released from normal platelets, leading to a secondary wave of irreversible platelet aggregation. A number of drugs, including aspirin, inhibit the plateletrelease reaction preventing the second wave of aggrega-
Volume 84 Number 3
tion. Once induced, this platelet defect is permanent and is corrected only when new platelets that have not been exposed to the drug have entered into the circulation. The present method of estimating platelet survival was devised by taking advantage of this effect of aspirin.on platelet function. All of the control subjects studied did not have return of normal platelet aggregation until four to seven days following ingestion of 10 grains of aspirin. This period of time required for complete return of function is the same as that reported by others. 9 The eight patients studied were suspected of having shortened platelet survival times because of their previous medical histories. Four had a history of idiopathic thrombocytopenic purpura, one had thrombocytopenia associated with systemic lupus erythematosus, and two had thrombocytopenia believed to be secondary to splenomegaly. The remaining patient had severe cardiac valvular disease which has been shown to result in platelet destruction and rapid platelet turnover, l~ 11 She had documented thrombocytopenia in the past. Seven of the eight patients had rapid recovery of platelet aggregation following aspirin ingestion, suggesting that there was a more rapid return into the circulation of undamaged platelets in those individuals. The one patient with sickle-cell anemia, mild thrombocytopenia, and a large spleen (Patient H) had complete return of function in five days, suggesting splenic platelet pooling rather than increased platelet destruction to be the etiology of his thrombocytopenia. This mechanism of producing thrombocytopenia has been well documented.12 There are two conditions which theoretically could have invalidated the present study. The first is the possibility that aspirin might interfere with the mechanisms of platelet destruction in the patients, thus tending to correct the abnormally shortened platelet survival time. It has already been demonstrated that aspirin does not appear to affect platelet survival in the normal individual) ~ The shortened 51Cr survival times in the patients taking aspirin in the present study, in addition to the lack of rise of platelet number in the thrombocytopenic patients, indicate that aspirin in the dosage used does not appear to correct an abnormally shortened platelet survival in the disease states studied. Previous studies have also demonstrated an inability of aspirin to correct shortened platelet survival in patients with prosthetic heart valves, unless this drug is used in combination with dipyridamole, it The second possible problem encountered would have been abnormal platelet aggregation secondary to the disease state itself. One study of patients with chronic idiopathic thrombocytopenic purpura d e m o n s t r a t e d
Measurement o f platelet survival
353
qualitative platelet abnormalities, including impaired epinephrine-induced aggregation, x3 This finding could not be demonstrated in the patients in this report, all of whom had normal aggregation prior to aspirin ingestion. This apparent discrepancy could be due to the marked difference between the concentrations of the aggregating agents used in the two studies. Only a small percentage of platelets is required to correct the aspirin-induced aggregation defect. It has been shown that the addition of only 10 per cent undamaged platelets to 90 per cent aspirin-treated platelets will completely correct aggregation? As little as five per cent undamaged platelets added to aspirin-treated platelets resulted in normal aggregation curves in this laboratory. Because normal platelet survival is about seven to 10 days, approximately 10 to 15 per cent newly produced platelets should enter the circulation every day. The fact that it requires four to seven days rather than one day for aggregation to recover completely in the normal person suggests that the megakaryocyte pool is also damaged by aspirin and abnormal platelets are being released into the circulation for a period of time. The production of platelets is believed to be regulated by a humoral stimulator which increases the size and number of megakaryocytes in the bone marrow.laThus a larger megakaryocyte pool may be susceptible to aspirin damage in states of compensated thrombocytopenia and create the apparent paradox of nearly normal aggregation recovery time after aspirin ingestion despite the markedly shortened 51Cr platelet survival times. This may explain the results obtained in Patients A, B, and C. Patients B and C had shorter 51Cr platelet survival times than Patient A, yet Patient A required only two days to recover from aspirin while Patients B and C required three days. Thus this method does not appear to be able to accurately quantitate platelet survival time. It does appear to be useful in demonstrating the presence of a compensated thrombocytolytic state, and of differentiating the thrombocytopenia of splenic platelet pooling from the thrombocytopenia due to increased platelet destruction. REFERENCES
1. Evans, G., Packham, M. A., Nishizawa, E. E., Mustard, J. F., and Murphy, E. A.: The effect of acetylsalicylic acid on platelet function, J. Exp. Med. 128: 877, 1968. 2. O'Brien, J. R.: Effects of salicylates on human platelets, Lancet 1: 779, 1968. 3. Zucker, M. B., and Peterson, J.: Inhibition of adenosine diphosphate-induced secondary aggregation and other platelet functions by acetylsalicylic acid ingestion, Proc. Soc. Exp. Biol. Med. 127: 547, 1968. 4. Born, G. V. R., and Cross, M. J.: The aggregation of blood platelets, J. Physiol. 168: 178, 1963.
354
5.
6.
7.
8.
9.
Schwartz
Murphy, S., Oski, F. A., and Gardner, F. H.: Hereditary thrombocytopenia with an intrinsic platelet defect, N. Engl. J. Med. 281: 857, 1969. Abrahamsen, A. F.: A modification of the technique for 51Cr-labelling of blood platelets giving increased circulating platelet radioactivity, Scand. J. Haematol. 5: 53, 1968. Brecher, G., and Cronkite, E. P.: Morphology and enumeration of human blood platelets, J. Appl. Physiol. 3: 365, 1950. Weiss, H. J., and Aledort, L. M.: Impaired platelet/connective tissue reaction in man after aspirin ingestion, Lancet 2: 495, 1967. Weiss, H. J., Aledort, L. M., and Kochwa, S.: The effect of salicylates on the hemostatic properties of platelets in man, J. Clin. Invest. 47: 2169, 1968.
The Journal of Pediatrics March 1974
10. Harker, L. A., and Slichter, S. J.: Platelet and fibrinogen consumption in man, N. Engl. J. Med. 20: 999, 1972. 11. Harker, L. A., and Slichter, S. J.: Studies of platelet and fibrinogen kinetics in patients with prosthetic heart valves, N. Engl. J. Med. 283: 1302, 1970. 12. Aster, R. H.: Pooling of platelets in the spleen: Role in the pathogenesis of"hypersplenic" thrombocytopenia, J. Clin. Invest. 45: 645, 1966. 13. Clancy, R., Jenkins, E., and Firkin, B.: Qualitative platelet abnormalities in idiopathic thrombocytopenic purpura, N. Engl. J. Med. 286: 622, 1972. 14. Harker, L. A.: Current concepts: Platelet production, N. Engl. J. Med. 282: 492, 1970.