Platelet Aggregation in Blood Donors

A. D. SCHWARTZ

From the Yale University School of Medicine, Department of Pediatrics, New Haven, Connecticut

One hundrcd eighteen consecutive volunteer donors to a Red Cross Mobile Collecting Center were questioned about ingestion of medication prior to blood donation. Blood samples were col- lected from each donor and platelet aggregation measured. Fortythree (36%) gave a history of having taken a drug known to affect platelet func- tion, the most common of which was acetylsalicylic acid. Of these, 88 per cent had abnormal platelet aggregation. A total of 59 per cent of the blood donors were found to have abnormal ag- gregation. When 50 control subjects were asked one week prior to testing not to take any medica- tion, only one had abnormal aggregation. This study indicates that in our area the majority of blood donors have abnormal platelet aggregation due to ingestion of medication.

DURING the past year, active investiga- tion of the role of the platelet in hemo- stasis and the effect of pharmacologic agents upon platelet function have been pursued in this laboratory. Considerable difficulty was encountered in obtaining control subjects with entirely normal plate- let function. The platelet abnormalities were due in most instances to previous in- gestion of salicylate-containing drugs. This observation prompicd a study of platelet aggregation in a population of healthy blood donors. The results of this study in- dicate that platelets with abnormal hemo- static function are often donated for clini- cal use.

Materials and Methods

One hundred eigli teen consecutive volunteer donors to a Red Cross Mobile Collecting Cen- ter were asked to respond to a short question- naire. Blood donors were accepted in accord-

Received for publication November 26, 1971; ac- cepted May 7, 1972.

ance witli the rccotnmendations ol the N;itional Ked Cross. Each donor was asked whether he h;td ingested any medication, including aspirin, during the week prior to blood donation and was requested to list the name of each drug. Finally, he was asked if he felt that he bruised or bled easily.

Following the witldrawal of one unit of blood from each donor, an extra 4.5 ml of blood were taken from the intravenous tubing and addcd to a plastic tube cont:tining 0.5 ml of 3.2 per cent buffered citrate anticoagulant. The blood was centrifuged at 500 rpm for 15 minutes. One milliliter of the supernate plate- let-rich plasma (PKP) was removed and the remaining sample centrifuged a t 3,400 rpm for ten minutes to preparc platelct-poor plasma (PPP). Platelet aggregation was nicasured by a modification of the turbiclimetric method of Born and Cross1 with a Chrono-log platelet aggregometer RIodel 300.

The aggregating agent was prepared by adding 1 ml of epineplirine U.S.P. to 4.5 ml of physiologic saline solution. PRP was con- stantly stirred at 1,200 rpm by a teflon-coated magnetic stir bar with the temperature main- tained at 37 C. T o 0.5 nil of PKP was added 0.025 ml of the aggregating ageiit, resulting ui a final concentration of 4 X 10-5 A3 epinephrine. Increase i l l light transmissioii occurring during platelet aggregation Wiis recorded on a Heath Model EU-2OV strip chart recorder after PRP was set as 0 per cent tr;insmission and PPP with 0.025 ml of epinephrine added set as 100 per cent transmission.

Fifty control subjects were rcquested in ad- vance to refrain from taking any medication for one week prior to testing. Twenty of the controls were children between the ages of 4 and 12 years. Blood was collected in a plastic syringe through a No. 20 needle, and prepared and studied in the same manner as samples obtained from blood donors. All aggregation studies were performed within two hours of the time of blood collection.

Transfusion Scpt.-Oct. 1972

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Volume 1 2 Number 5

PLATELET AGGREGATION IN BLOOD DONORS 345

T A n L E 1. Results of Platelet Aggregntion Studies in Blood Donors and Control Subjects

Blood Donors

Controls Females Males Total

History of drug ingestion 0 41% (29/66) 27% (14/52) 36% (43/118)

Abnormal aggregation with positive drug history 0 SG% (25/29) 93% (13/14) 88% (38/43)

No history of drug ingcstion 100% (50/50) 56% (37/66) 73% (38/52) 64% (75/118) Abnormal aggregation with

negative drug history 2% (1/50) 41% (15/37) 45% (17/38) 43% (32/75)

Total with abnormal aggregation 2% (1/50) 61% (40/66) 58% (30/52) 59% (70/118)

Results

All but one of tlie control subjects had 75-100 per cent aggregation, with aggregation being completed between 3 to 4 minutes after addition of epinephrine. This pattern repre- sents a normal result in this laboratory. Platelet aggregation was considered abnormal if there was less than 60 per cent aggregation 5 minutes following addition of epinephrine to PRP.

Results of the aggregation studies of tlie blood donors are recorded in Table 1. Of the 70 donors with abnormal function, 60 had no secondary wave of aggregation. T h e remaining ten had a markedly delayed secondary wave which seldom reached 50 per cent aggregation. A few samples had a rate of aggregation slightly slower than that of the controls, but were con- sidered norma1.

A positive history was defined by ingestion of drugs known to affect platelet function and was found in 36 per cent of the donor popula- tion. Of tlie 43 individrials who gave a positive drug history, 40 Itad ingested salicylate-contain- ing compounds and three liad taken antihista- mines. Eighty-eiglit per cent of individuals with ;I positive history had abnormal platelet aggre- gation. Altliougli 75 persons denied drug ingestion, 32 of them (43%) had abnormal aggregation. Men denied drug ingestion more commonly than women, but there appeared to be no difference in the results of aggregation studies between the sexes. A total of 70 of the 118 blood donors (59%) hat1 abnormal p1;ltelet aggregation.

Fourteen blood donors believed that they bruised or bled more easily than normal. The one male with such a history denied taking medication. but liad abnormal platelet aggre- gation. T h e remaining 13 were women, a total of 20 per cent of the female blood donor population.

A comparison of women with and without a history suggesting a hemorrhagic problem is made in Table 2. \*\'omen who believed tliem- selves to bruise or bleed more easily th;in nor- mal had a 54 per cent incidence of drug inges- tion and a 77 per cent incidence of abnormal platelet aggregation. I n comparison, those who denied easy bruising or bleeding had a 9 per cent incidence of drug ingestion and 36 per cent incidence of abnormal aggregation. T h e differences between these two groups a s deter- mined by the Chi square test is statistically significant with a P value of less than 0.01.

Discussion

The addition of epinephrine to PKP re- sults in a primary wave of platelet aggrega- tion. Under this stimulus, endogenous ade- nosine diphosphate (ADP) is released from normal platelets leading to a secondary wave of irreversible platelet aggregation. A number of drugs, including aspirin,2- 7, 11

inhibit the release of endogenous ADP, thus preventing the secondary wave of ag- gregation.

Difficulty in obtaining normal control subjects for platelet function studies led to the present study. Frequently a subject initially denied drug ingestion, but after abnormal aggregation was demonstrated, upon requestioning, they recalled ingestion of a headache tablet or salicylate-contain- ing powder for indigestion. Some indi- viduals stated that they often took such drugs, but were unsure whether they had taken medication in the past few days. Unfortunately, it was not possible to take

346 SCHIVARTZ Transfusion Sept.-Uct. 1972

TARLE 2. Relntionshilj of Bleeding History t o Plntelet A g g q n / i o n nnd History of Drug Ingeslion

Abnormal Aggregation

History of Drug Ingestion

Wornen wilh positive history

Women with negative history of bruising or bleeding 77% (10/13) 54% (7/13)

of bruising or bleeding 36% (19/53) 9% (5/53)

a second history from the blood donors described here.

A series of normal control subjects was finally obtained by asking individuals one week in advance to avoid taking any medi- cation. Aggregation studies were normal in all but one of 50 persons tested, clearly im- plicating drug ingestion as the cause of abnormal aggregation.

Experience with young children was dif- ferent from adults. Control subjects with normally functioning platelets were more easily obtained in the younger age group. The rare child with abnormal platelet ag- gregation usually had a history of drug ingestion which was associated with an ill- ness and such medication was remembered by a parent. T h e adult control subject often took the drug for minor indications.

McCann et al.6 have previously reported an 11 per cent incidence of positive serum salicylate levels in a random sampling of blood donors, all of whom denied drug ingestion. The higher incidence in the present series may reflect the fact that aspirin is rapidly cleared from the blood while the platelet abnormality caused by aspirin persists much longer and is not completely corrected for four to seven days.10 Therefore, once induced, the plate- let defect is corrected only when new plate- lets that have not been exposed to the drug have been released from the bone marrow. As little as 5 grains of aspirin can produce the platelet defect.10- 11

Under usual circumstances the effect of drugs upon platelets probably has little

clinical significance. However, if a patient recently has undergone a surgical proce- dure or has another underlying defect, drug ingestion may cause significant bleed- ing. An association of aspirin with post- tonsillectomy bleeding has been recognized for over 20 years.3- 9 Furthermore, aspirin has been shown to aggravate the bleeding tendencies of patients with congenital4~ 8

and acquired5 coagulation disorders. Most hematologists have had the experi-

ence of seeing patients for evaluation of a mild hemorrhagic disorder in whom no coagulation abnormality can be found. It is likely that many of these cases represent unrecognized drug-induced bleeding. The observation that 20 per cent of women in the group of blood donors gave a history of easy bruising or bleeding, and that 77 per cent of these individuals had abnormal platelet aggregation and 54 per cent a his- tory of drug ingestion, leiids weight to this speculation.

The present study indicated that in our area the majority of bloocl donors have platelets that function abnormally and that the abnormality is due to ingestion of mecli- cation. Blood collected from such donors is usually separated into various com- ponents, and the platelets are given to leukemic or other thrombocytopenic pa- tients. Although hemostatic effectiveness of these functionally abnormal platelets has not yet been determined, experience with postoperative and hemophilic patients sug- gest that optimal hemostatic support may not be provided by such damaged platelets.

Volume 12 PLATELET AGGREGATION IN R1.00D DONORS Number 5

T h e incidence of platelet dysfunction in a panel of individuals could he decreased by asking subjects i n advance to avoid medication. This suggests that the same results could be obtained in a population of informed blood donors.

Acknowledgment

T h e technical assistance of Mrs. Carol Johnson and Mrs. Billie Jean Norman is gratefully acknowl- edged.

References

I . Born, G . V. R.. and M. J. Cross: The aggrega- tion of blood platelets. J. Physiol. 168: 178, 1963.

2. Evans, G., M. A. Packham, E. E. Nishizawa, J. F. Mustard, and E. A. Murphy: The effect of acetylsalicylic acid on platelet function. J. Exper. Med. 128: 877, 1968.

3. Fox, S. L., and G. B. West: Vitainin K and late tonsillar hemorrhage. Laryngoscopc 57: 564, 1947.

4. Kaneshiro. M. M., C. H. Mielke, C. K. Kasper, and S. I. Rapaport: Bleeding time after

347

aspirin in disorders of intrinsic clotting. Ncw Eng. J. Med. 281: 1039, 1969.

5. Koch-Weser, V. A,. and E. M. Sellcrs: Drug intcractioiis with couinarin anticoagulants. New Eng. J. Med. 285:547, 1971.

6. McCann, W. P., E. I. McGowan, 0. L. Ihirnctt, and 1’. A. I’alinisano: Seriini salicylatc levcls in blood donors. J.A.M.A. 214: 553, 1970.

7. O’Brien, J. R.: Effects of salicylatcs on human platelets. Lancct 1: 799, 1968.

8. Quick, A. J.: Salicylatcs and bleeding: T h e aspirin tolerance test. Aiiier. J. Med. Sci. 252: 265, 1966.

9. Singer, R.: Acetylsalicylic acid, a probable cause €or secondary post-tonsillectomy hemor- rhage. Arch. Otolaryng. 42: 19, 1945.

10. Weiss, H. J., and L. M. Aledort: Imparicd platelet/connective tissue reaction in mail after aspirin ingestion. Lancet 2: 495, 1967.

11. Zucker, M. B., and J. Peterson: Inhibition of adenosine disphophate-induced secondary ag- gregation and other platelet functions by acetylsalicylic acid ingestion. Proc. SOC. Exper. Biol. 127: 547, 1968.

Allen D. Schwartz, M.D., Children’s Memorial Hospital, Division of Hematology, 2300 Children’s Plaza, Chicago, Illinois 60614.