Brief Report
Effect of Prostaglandin E-1 Upon Microaggregate and Fibrin Formation in Stored Blood J . R. HARP, B. E. MARSHALL, H . A. WURZEL,AND A. S. MILLER From the University of Pennsylvania. Philadelphia, Pennsylvania
RESPIRATORY F A I L U R E has been identified be a desirable approach, since aspirin irrea s a common life-threatening complication of versibly inhibits platelet function.34 Platelets t r a ~ m a . Massive ~ . ~ ~ ~blood ~ ~ transfusion (> have been shown to be essential for microaggrega t e form a tion in stored blood, 17*23*33and 10 units) may be a significant contributing f a ~ t o r . ~ , ' ~ ,S t' o~r e. d~ blood ~ . ~ ~contains ~ ~ ~ modification of platelet behavior to prevent microaggregates consisting of platelets, leumicroaggregate formation seems logical. Prostaglandin E-l (PGE,) is a potent incocytes, fibrin strands, and o t h e r debris hibitor of platelet aggregationI4*l5and has which may be small enough ( < 170 microns) been used to facilitate preparation of platelet to pass standard transfusion filters and enter t h e circulation. I I . 17.18.22.27.32.33 R emoval of rich p l a ~ m a . ~Evidence * ~ * ~ ~ for inhibition of microaggregate formation by 20 mg P G E , t h e s e m i c r o e m b o l i by filtration in t h e pulmonary circulation may lead to per ml whole blood h a s been found by pulmonary vascular disturbance and measurement of screen filtration rate (SFR), capillary endothelial damage.7.'2'13.25.28.30 a modification of screen filtration pressure (SFP). S F R was markedly lower in control A variety of micropore filters have been designed to remove microaggregate material blood after five days storage ( I .75 ml on day from blood prior to i n f u ~ i o n . ~ - ' ~ . ~ ~1 .vs. ~ ~0.25 . ~ ~ml~ on ~ ~ day 5 ) , but only moderately lower in P G E , treated blood (1.80 ml on day These filters a r e now in widespread use, and have been reported in prospective studies to 1 vs. I .45 ml on day 5).30 be effective in reducing the incidence of T h e present study examines the effect respiratory failure following massive transfuupon S F P , platelet count and screen debris morphology of adding P G E , (20 mg per ml) sion in man.3o However, all of the micropore to whole human blood collected in acid filters remove significant amounts of cellular citrate dextrose (ACD) solution during 0 to elements, particularly platelets and leuc0cytes.8.20.32.33 21 days storage a t 4 C . An alternate approach to prevention of Respiratory Distress Syndrome ( R D S ) Methods following massive transfusion may be desirBlood Collection able, particularly since a second common After obtaining informed consent, young complication, bleeding diathesis, is now healthy male volunteers were subjected to a stanknown to be due to dilutional thrombocytodard medical examination, including medical history, blood pressure, hemoglobin, urinalysis and penia,le.21a condition which could be exacerwhite blood cell count. Those selected were bated by micropore filtration. One earlier instructed to refrain from all drug ingestion for study described an a t t e m p t to prevent one week and from smoking for 12 hours. Blood microaggregate formation in stored blood by was collected into 67.5 ml acid-citrate-dextrose addition of aspirin.' This does not appear to (ACD) solution, U S P formula A. A n equal volume, as determined by weight, was transfered to a second bag, and the connecting tubing was sealed and divided. PGE, was dissolved in absolute Received for publication July 4, 1975; accepted August 16, 1975. ethanol, 10 mg in 1.0 ml, and then wasdiluted with
277 Transfusion May-June 1976
Volume 16 Number3
278
Transfusion May-June 1976
HARP, ET A L Table 1. Effect of PGEl on SFP (torr) in Stored Blood
0
Storage Days Control units P G E l treated units
38 26
f f
10" 2"
7
14
21
391 ?: 117 141 f 30
406 f 116 21 1 f 52
644 f 84 304 66
Significance of difference of means P G E l vs. control: day 7. p "SFP (mean k standard error of mean).
sterile veronal buffer, pH 6.55, to a concentration of 5 p g per ml. The medication injection site was used to add 5 wg PGE, to one aliquot of blood to give a concentration of approximately 20 mg per ml. The same volume of buffer without PGE, was added to the second aliquot of blood. After mixing, samples were withdrawn using sterile precautions through a 170 p filter. Both the control and the PGE, treated aliquots were then divided equally, as determined by weight, into three separate bags. The tubing used to transfer the blood was sealed and divided. The blood was stored at 4 C without agitation. Samples were obtained through a 170 p filter on day 7, 14, or 21 after the appropriate unit had been warmed to room temperature and had been mixed by inversion. Units which were sampled on day 7 were resampled on day 10. All other fractional units were sampled one time only. Screen Filtration Pressure
Screen filtration pressure (SFP) was measured in triplicate using a modification of the Swank technique. Blood was forced through a 2.54 sq mm area of nickel screen (pore size 20 x 20 M)* a t a rate of 2.0 ml per sec by means of a heavy duty Harvard Apparatus infusion pump. Pressure to force blood through the screen was recorded on a Texas Galvanometer via a Statham Microdot Transducer calibrated to 1,250 torr. S F P was recorded as the pressure maximum at the end of ten seconds. Reported values represent the mean of triplicate measurements. Platelet Count
Platelet counts were performed in triplicate using a standard phase h e m ~ c y t o m e t r y . ~ Screen Debris Morphology
Screens used to determine S F P were washed with saline and then fixed in 3 per cent gluteraldehyde. The sample was dehydrated by passage through graded alcohols and amyl acetate and then subjected to critical point drying. T h e *Buckbee Mears Co., St. Paul, Minn
*
< 0.05; day 21, p < 0.01.
specimen was then layered with gold and photographed at 60 and 1800x magnification with a JEOL JSM 50-A scanning electron microscope. Analysis of Results
Determination of significance of difference o f means for S F P and platelet count was by onetailed paired T test.
Results Screen Filtration Pressure
Table 1 presents mean S F P for treated and untreated units at day 0, 7, 14, and 21. On day 7 and day 21, the mean S F P of PGE, treated units was significantly (P < 0.05 and P < 0.01) lower than that o f the control units. S F P was higher in the PGE, treated than in its untreated control only once (donor 5 on day 14). Changes in S F P resulting from storage of the control units were consistent with earlier studies done in from this laboratory. l . I 7 . In Units used for day 7 samples were stored at 4 C and were resampled on day 10. In these units, there was a marked increase in SFP. On day 10, the S F P was 744 + 184 torr in untreated samples and 486 * 130 torr in PGE, treated samples. These results were higher than those obtained on undisturbed units stored for 14 days (not reported here). Removal of blood from refrigerated storage, warming to room temperature, and mixing results in an increase in SFP. Platelet Counr
Table 2 represents changes in platelet count during storage. The platelet count was significantly (P < 0.02 and P < 0.01) higher in PGE, treated samples than in the control samples on day I4 and 21 o f storage. Screen Debris Morphology
Typical scanning electron photomicrographs of screens from treated and untreated samples after 7 and 21 days storage are shown in Figures 1-4. Fibrillar material identified as fibrin
Volume 16 Number 3
279
EFFECT O F PROSTAGLANDIN Table 2. Effect of PGEl on Platelet Count in Stored Blood Storage Days
0
7
14
21
Control units PGEl treated units
140 i loc 144 * 12”
114+ 12 128+ 12
44 i 9 8 0 + 16
58 + 6 88 i 7
*Platelet count X 10-3 + standard error of mean. Significance of difference of means PGEl vs. control: day 14,p
in earlier studiesli.lsis present in both treated and untreated fractional units. I t is not possible to quantitate the rnorphologic findings. However, there does appear to be less fibrin on screens from PGE, treated blood.
< 0.05;day 21,p < 0.01.
Signijicance o f S F P S F P is apparently linearly related to microaggregate content in human blood stored in ACD.li*lsThis is not the case when human blood is stored in C P D ” nor when blood from sheep or horses is stored in ACD.I8 I n each of these latter circumstances, large amounts of microaggregate material, consisting primarily of clumped
platelets, can be identified while S F P is not significantly elevated. Screen debris morphology studies as well as flourescine-labeled rabbit anti-fibrin antibody assay indicate that S F P elevation occurs in stored blood when fibrin is present in addition to microaggregate material.” I n earlier studies, we have cited convincing evidence that pulmonary damage follows infusion of microaggregates plus fibrin, and that infusion of microaggregates alone does not produce lung damage.20This is important for interpretation of the present findings, since mitigation of S F P elevation during blood storage by PGE, must indicate reduction of the rate of fibrin formation. This, in t u r n , should reduce t h e probability of
FIG. I . Scanning electron microgram (18OOx)of debris on screens after filtration of PEG, treated blood at 7 days, S F P I80Torr.
FIG.2. Scanning electron microgram (1800x)of debris on screens after filtration of control blood at 7 days, S F P 420 Torr.
Discussion
280
HARP, ET AL.
Transfusion
May-June 1976
FIG.3. Scanning electron microgram (1800x)ofdebris on screens after filtration of PEG, treated blood of 21 days, S F P 261 Torr.
FIG.4. Scanning electron microgram (1800x)of debris on screens after filtration of control blood at 21 dyas, S F P 433 Torr.
respiratory distress syndrome (RDS) following massive transfusion.
Clinical Irnplicarions The present observations comprise a pilot study. Critical questions remain to be answered regarding duration of survival of functional platelets during blood storage at 4 C . An encouraging partial answer is found in Becker's3 observation that single units of platelets harvested from blood treated with PGE,, 10 mg per ml and stored for six days in A C D preservative a t 4 c significantly shortened the duration of the bleeding time abnormality produced by aspirin ingestion. Approximately one-third of transfused blood is storaged six days or less" and blood in this age range is therefore readily available for use in massive transfusion. The clinical significance of the reduction in microaggregate and fibrin formation noted in the present study is unclear. Both microaggregates and fibrin are present after seven days storage. For this reason, microfiltration might still be indicated for massive transfusion of PGE, treated blood. The possibility remains, however, that PGE, treatment may
Platelet Count While the platelet count in the present material was consistently higher in PGE, treated blood, lack of data reflecting platelet function impose constraint upon interpretation. Storage at 4 C might be expected to prolong survival of cellular elements through reduction of metabolic requirements. However, it seems unlikely that functional platelets could remain throughout 21 days. Beck e r3 noted mo rp holog i c a bno r m al i t i es after six days in platelets stored at 4 C in ACD in PGE, 20 mg per ml. We interpret the consistently higher platelet count in PGE, treated blood to indicate that clumping a n d / o r aggregation were inhibited throughout the period of platelet viability, possibly five to seven days. However, both in vivo and in vitro tests of platelet function during the entire period of blood storage with and without PGE, appear to be indicated.
Volume 16 Number 3
EFFECT O F PROSTAGLANDIN
reduce microaggregate and fibrin formation sufficiently to remove the need for micropore filtration. While PGE, is rapidly removed from the circulation by the lung, and levels of 0.1 mg per kg per minute can be tolerated by healthy volunteers," it is not known what effect shock, or anesthesia may have upon tolerance to PGE, infusion. The present results do indicate that it is possible to reduce microaggregate and fibrin formation throughout 21 days of blood storage. Possible prolongation of platelet survival is also suggested. Microaggregate embolism may contribute to respiratory failure following massive t r a n ~ f u s i o n ,and ~~ dilutional thrombocytopeniaZ1 appears to be the most common cause of coagulation abnormalities following massive transfusion. PGE, treatment of blood at time of collection may alleviate both of these problems.
References I.
2.
3.
4.
5.
6.
7.
8.
9.
Arrington, P. J., and J. J. McNamara: Effect of aspirin on microaggregate formation in banked blood. Surgery 76:295, 1974. Becker, G. A,, M. K. Chalos, M. Tuccelli, and R. H. Aster: Prostaglandin E, in preparation and s t o r a g e of platelet concentrates. Science 175538, 1972. -, T . Kunicki, and R. H. Aster: Effect of Prostaglandin E, on harvesting of platelets from refrigerated whole blood. J . Lab. Clin. Med. 83:304, 1974. Blaisdell, F. W., and R. M. Schlobohm: The respiratory distress syndrome: a review. Surgery 74:251, 1973. Brecher, G., M. Schneiderman, and E. P. Cronkite: The reproductibility and constancy of the platelet count. Am. J. Clin. Pathol. 23:15, 1953. Carlson, L. A,, L. Ekelund, and L. Oro: Clinical and metabolic effects of different doses of Prostaglandin E, i n man. A c t a Med. S c a n d . 183:423, 1968. Connell, R. S., and R. L. Swank: Pulmonary microembolism after blood transfusions. Ann. Surg. 177:40, 1973. Cullen, D. J., and L. Ferrara: Comparative evaluation of blood filters. Anesthesiology 41:568, 1974. Gervin, A. S., T. J . Limbird, C. L. Puckett, and D. Silver: Ultrapore hemofiltration: the effects on the coagulation and fibrinolytic mechanisms i n
10.
I I.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
23.
24. 25.
28 1
fresh and stored blood. Arch. Surg. 106:333, 1973. Goldiner, P. L., W. S. Howland, and C. Ray: Filter for prevention of microembolism during massive transfusions. Anesth. Analg. 51:717, 1972. Harp, J. R., M. Q. Wyche, B. E. Marshall, and H. A. Wurzel: Some factors determining rate of microaggregate formation in stored blood. Anesthesiology 40:398, 1974. Hissen, W., and R. L. Swank: Screen filtration pressure and pulmonary hypertension. Am. J. Physiol. 209:715, 1965. Jenevein, E. P., and D. L. Weiss: Platelet microemboli associated with massive blood transfusion. Am. J. Pathol. 45:313, 1964. Kinlough-Rathbone, R. L., M. A . Packham, and J. F. Mustard: The effect of Prostaglandin E, on platelet function in vitro and in viva Br. J. Haematol. 19559, 1970. Kloeze, J.: Influence of Prostaglandins on platelet adhesiveness and platelet aggregation. In Proceedings of the 2nd Nobel Symposium, Stockholm, June 1966. S. Bergstrom, and B. Samuelsson, Eds. New York, Interscience, 1967, p.241. Lim, R. C., Jr. C. Olcott IV, A . J . Robinson, and F. W. Blaisdell: Platelet response and coagulation changes following massive blood replacement. J. Trauma 13577, 1973. Marshall, R. E., N. Ellison, H. A . Wurzel, et a/.: Microaggregate formation in stored blood: 11. Influence of anticoagulants and blood components. Circ. Shock 2:185, 1975. -, L. R. Soma, J. R. Harp, e / a/.: Microaggregate formation in stored blood. I. Species differences and storage time. Circ. Shock 2:175, 1975. -, L. R. Soma, J. R. Harp, G. R. Neufeld, H. A. Wurzel, and D. C. Dodd: Pulmonary function after exchange transfusion of stored blood in dogs. Ann. Surg. 179:46, 1974. -, H. A . Wurzel, N. Ellison, et a/.: Microaggregate formation in stored blood: 111. Comparison of Bentley, Fenwal, Pall, and Swank micropore filtration. Circ. Shock 2:249, 1975. Miller, R. D., T . 0. Robbins, J. M. Tong, and S. L. Barton: Coagulation defects associated with massive blood transfusions. Ann Surg. 174:794, 1971. McNamara, J. J., D. Boatright, E. L. Burran, el a/.: Changes in some physical properties of stored blood. Ann. Surg. 17458, 1971. -, E. L. Burran, E. Larson, G. Omiya, G. Suehiro, and H. Yamase: Effect of debris in stored blood on pulmonary microvasculature. Ann. Thor. Surg. 14:133, 1972. -, E. L. Burran, and G. Suehiro: Effective filtration of banked blood. Surg. 71594, 1972. -, M. D. Molot, and J . F. Stremple: Screen filtration pressure in combat casualities. Ann. Surg. 172334, 1970
282 26. 27.
28.
29.
30.
31.
32.
33.
HARP, ET A L . Moore, F. D.: Terminal mechanisms in human injury. Am. J. Surg. 110:317, 1965. Moseley, R. V., and D. B. Doty: Changes in the filtration characteristics of stored blood. Ann. Surg. 171:329, 1970. -, and D. B. Doty: Death associated with multiple pulmonary emboli soon after battle injury. Ann. Surg. 171:336, 1970. Pontoppidan, H., B. Geffin, and E. Lowenstein: Acute respiratory failure in the adult. N. Engl. J . Med. 287:690, 1972. Reul, G. J., Jr., A . C . Beall, and S . D. Greenberg: Protection of the pulmonary microvasculature by fine screen filtration. Chest 66:4, 1974. Shio, H., and P. W. Ramwell: Prostaglandin E, in platelet harvesting: an in vitro study. Science 175536, 1972. Solis, R. T., and M. B. Gibbs: Filtration of the microaggregates in stored blood. Transfusion 12:245, 1972. Swank, R. L.: Alteration of blood on storage:
34.
Transfusion May-June 1976
m e a s u r e m e n t of adhesiveness of “aging” platelets and leucocytes and their removal by filtration. N. Engl. J . Med. 265:728, 1961. Weiss, H. J., L . M. Aledort, and S. Kochwa: The effect of salicylates on the hemostatic properties of platelets in man. J. Clin. Invest. 47:2169, 1968.
J . R . H a r p , M.D., Professor & C h a i r m a n of Anesthesiology, Temple University Health Science Center, 3400 N. Broad Street, Philadelphia, Pa. 19140. B. E. Marshall, M.D., Professor of Anesthesiology, Hospital of the University of Pennsylvania, 3400 Spruce Street, Philadelphia, Pa. 19104. H.A . Wurzel, M.D., Associate Professor of Medicine and Pathology, Hospital of the Univeristy of Pennsylvania. A. S. Miller, B.S., Medical Technologist, Hospital of the University of Pennsylvania.
Effect of Prostaglandin E-1 Upon Microaggregate and Fibrin Formation in Stored Blood J . R. HARP, B. E. MARSHALL, H . A. WURZEL,AND A. S. MILLER From the University of Pennsylvania. Philadelphia, Pennsylvania
RESPIRATORY F A I L U R E has been identified be a desirable approach, since aspirin irrea s a common life-threatening complication of versibly inhibits platelet function.34 Platelets t r a ~ m a . Massive ~ . ~ ~ ~blood ~ ~ transfusion (> have been shown to be essential for microaggrega t e form a tion in stored blood, 17*23*33and 10 units) may be a significant contributing f a ~ t o r . ~ , ' ~ ,S t' o~r e. d~ blood ~ . ~ ~contains ~ ~ ~ modification of platelet behavior to prevent microaggregates consisting of platelets, leumicroaggregate formation seems logical. Prostaglandin E-l (PGE,) is a potent incocytes, fibrin strands, and o t h e r debris hibitor of platelet aggregationI4*l5and has which may be small enough ( < 170 microns) been used to facilitate preparation of platelet to pass standard transfusion filters and enter t h e circulation. I I . 17.18.22.27.32.33 R emoval of rich p l a ~ m a . ~Evidence * ~ * ~ ~ for inhibition of microaggregate formation by 20 mg P G E , t h e s e m i c r o e m b o l i by filtration in t h e pulmonary circulation may lead to per ml whole blood h a s been found by pulmonary vascular disturbance and measurement of screen filtration rate (SFR), capillary endothelial damage.7.'2'13.25.28.30 a modification of screen filtration pressure (SFP). S F R was markedly lower in control A variety of micropore filters have been designed to remove microaggregate material blood after five days storage ( I .75 ml on day from blood prior to i n f u ~ i o n . ~ - ' ~ . ~ ~1 .vs. ~ ~0.25 . ~ ~ml~ on ~ ~ day 5 ) , but only moderately lower in P G E , treated blood (1.80 ml on day These filters a r e now in widespread use, and have been reported in prospective studies to 1 vs. I .45 ml on day 5).30 be effective in reducing the incidence of T h e present study examines the effect respiratory failure following massive transfuupon S F P , platelet count and screen debris morphology of adding P G E , (20 mg per ml) sion in man.3o However, all of the micropore to whole human blood collected in acid filters remove significant amounts of cellular citrate dextrose (ACD) solution during 0 to elements, particularly platelets and leuc0cytes.8.20.32.33 21 days storage a t 4 C . An alternate approach to prevention of Respiratory Distress Syndrome ( R D S ) Methods following massive transfusion may be desirBlood Collection able, particularly since a second common After obtaining informed consent, young complication, bleeding diathesis, is now healthy male volunteers were subjected to a stanknown to be due to dilutional thrombocytodard medical examination, including medical history, blood pressure, hemoglobin, urinalysis and penia,le.21a condition which could be exacerwhite blood cell count. Those selected were bated by micropore filtration. One earlier instructed to refrain from all drug ingestion for study described an a t t e m p t to prevent one week and from smoking for 12 hours. Blood microaggregate formation in stored blood by was collected into 67.5 ml acid-citrate-dextrose addition of aspirin.' This does not appear to (ACD) solution, U S P formula A. A n equal volume, as determined by weight, was transfered to a second bag, and the connecting tubing was sealed and divided. PGE, was dissolved in absolute Received for publication July 4, 1975; accepted August 16, 1975. ethanol, 10 mg in 1.0 ml, and then wasdiluted with
277 Transfusion May-June 1976
Volume 16 Number3
278
Transfusion May-June 1976
HARP, ET A L Table 1. Effect of PGEl on SFP (torr) in Stored Blood
0
Storage Days Control units P G E l treated units
38 26
f f
10" 2"
7
14
21
391 ?: 117 141 f 30
406 f 116 21 1 f 52
644 f 84 304 66
Significance of difference of means P G E l vs. control: day 7. p "SFP (mean k standard error of mean).
sterile veronal buffer, pH 6.55, to a concentration of 5 p g per ml. The medication injection site was used to add 5 wg PGE, to one aliquot of blood to give a concentration of approximately 20 mg per ml. The same volume of buffer without PGE, was added to the second aliquot of blood. After mixing, samples were withdrawn using sterile precautions through a 170 p filter. Both the control and the PGE, treated aliquots were then divided equally, as determined by weight, into three separate bags. The tubing used to transfer the blood was sealed and divided. The blood was stored at 4 C without agitation. Samples were obtained through a 170 p filter on day 7, 14, or 21 after the appropriate unit had been warmed to room temperature and had been mixed by inversion. Units which were sampled on day 7 were resampled on day 10. All other fractional units were sampled one time only. Screen Filtration Pressure
Screen filtration pressure (SFP) was measured in triplicate using a modification of the Swank technique. Blood was forced through a 2.54 sq mm area of nickel screen (pore size 20 x 20 M)* a t a rate of 2.0 ml per sec by means of a heavy duty Harvard Apparatus infusion pump. Pressure to force blood through the screen was recorded on a Texas Galvanometer via a Statham Microdot Transducer calibrated to 1,250 torr. S F P was recorded as the pressure maximum at the end of ten seconds. Reported values represent the mean of triplicate measurements. Platelet Count
Platelet counts were performed in triplicate using a standard phase h e m ~ c y t o m e t r y . ~ Screen Debris Morphology
Screens used to determine S F P were washed with saline and then fixed in 3 per cent gluteraldehyde. The sample was dehydrated by passage through graded alcohols and amyl acetate and then subjected to critical point drying. T h e *Buckbee Mears Co., St. Paul, Minn
*
< 0.05; day 21, p < 0.01.
specimen was then layered with gold and photographed at 60 and 1800x magnification with a JEOL JSM 50-A scanning electron microscope. Analysis of Results
Determination of significance of difference o f means for S F P and platelet count was by onetailed paired T test.
Results Screen Filtration Pressure
Table 1 presents mean S F P for treated and untreated units at day 0, 7, 14, and 21. On day 7 and day 21, the mean S F P of PGE, treated units was significantly (P < 0.05 and P < 0.01) lower than that o f the control units. S F P was higher in the PGE, treated than in its untreated control only once (donor 5 on day 14). Changes in S F P resulting from storage of the control units were consistent with earlier studies done in from this laboratory. l . I 7 . In Units used for day 7 samples were stored at 4 C and were resampled on day 10. In these units, there was a marked increase in SFP. On day 10, the S F P was 744 + 184 torr in untreated samples and 486 * 130 torr in PGE, treated samples. These results were higher than those obtained on undisturbed units stored for 14 days (not reported here). Removal of blood from refrigerated storage, warming to room temperature, and mixing results in an increase in SFP. Platelet Counr
Table 2 represents changes in platelet count during storage. The platelet count was significantly (P < 0.02 and P < 0.01) higher in PGE, treated samples than in the control samples on day I4 and 21 o f storage. Screen Debris Morphology
Typical scanning electron photomicrographs of screens from treated and untreated samples after 7 and 21 days storage are shown in Figures 1-4. Fibrillar material identified as fibrin
Volume 16 Number 3
279
EFFECT O F PROSTAGLANDIN Table 2. Effect of PGEl on Platelet Count in Stored Blood Storage Days
0
7
14
21
Control units PGEl treated units
140 i loc 144 * 12”
114+ 12 128+ 12
44 i 9 8 0 + 16
58 + 6 88 i 7
*Platelet count X 10-3 + standard error of mean. Significance of difference of means PGEl vs. control: day 14,p
in earlier studiesli.lsis present in both treated and untreated fractional units. I t is not possible to quantitate the rnorphologic findings. However, there does appear to be less fibrin on screens from PGE, treated blood.
< 0.05;day 21,p < 0.01.
Signijicance o f S F P S F P is apparently linearly related to microaggregate content in human blood stored in ACD.li*lsThis is not the case when human blood is stored in C P D ” nor when blood from sheep or horses is stored in ACD.I8 I n each of these latter circumstances, large amounts of microaggregate material, consisting primarily of clumped
platelets, can be identified while S F P is not significantly elevated. Screen debris morphology studies as well as flourescine-labeled rabbit anti-fibrin antibody assay indicate that S F P elevation occurs in stored blood when fibrin is present in addition to microaggregate material.” I n earlier studies, we have cited convincing evidence that pulmonary damage follows infusion of microaggregates plus fibrin, and that infusion of microaggregates alone does not produce lung damage.20This is important for interpretation of the present findings, since mitigation of S F P elevation during blood storage by PGE, must indicate reduction of the rate of fibrin formation. This, in t u r n , should reduce t h e probability of
FIG. I . Scanning electron microgram (18OOx)of debris on screens after filtration of PEG, treated blood at 7 days, S F P I80Torr.
FIG.2. Scanning electron microgram (1800x)of debris on screens after filtration of control blood at 7 days, S F P 420 Torr.
Discussion
280
HARP, ET AL.
Transfusion
May-June 1976
FIG.3. Scanning electron microgram (1800x)ofdebris on screens after filtration of PEG, treated blood of 21 days, S F P 261 Torr.
FIG.4. Scanning electron microgram (1800x)of debris on screens after filtration of control blood at 21 dyas, S F P 433 Torr.
respiratory distress syndrome (RDS) following massive transfusion.
Clinical Irnplicarions The present observations comprise a pilot study. Critical questions remain to be answered regarding duration of survival of functional platelets during blood storage at 4 C . An encouraging partial answer is found in Becker's3 observation that single units of platelets harvested from blood treated with PGE,, 10 mg per ml and stored for six days in A C D preservative a t 4 c significantly shortened the duration of the bleeding time abnormality produced by aspirin ingestion. Approximately one-third of transfused blood is storaged six days or less" and blood in this age range is therefore readily available for use in massive transfusion. The clinical significance of the reduction in microaggregate and fibrin formation noted in the present study is unclear. Both microaggregates and fibrin are present after seven days storage. For this reason, microfiltration might still be indicated for massive transfusion of PGE, treated blood. The possibility remains, however, that PGE, treatment may
Platelet Count While the platelet count in the present material was consistently higher in PGE, treated blood, lack of data reflecting platelet function impose constraint upon interpretation. Storage at 4 C might be expected to prolong survival of cellular elements through reduction of metabolic requirements. However, it seems unlikely that functional platelets could remain throughout 21 days. Beck e r3 noted mo rp holog i c a bno r m al i t i es after six days in platelets stored at 4 C in ACD in PGE, 20 mg per ml. We interpret the consistently higher platelet count in PGE, treated blood to indicate that clumping a n d / o r aggregation were inhibited throughout the period of platelet viability, possibly five to seven days. However, both in vivo and in vitro tests of platelet function during the entire period of blood storage with and without PGE, appear to be indicated.
Volume 16 Number 3
EFFECT O F PROSTAGLANDIN
reduce microaggregate and fibrin formation sufficiently to remove the need for micropore filtration. While PGE, is rapidly removed from the circulation by the lung, and levels of 0.1 mg per kg per minute can be tolerated by healthy volunteers," it is not known what effect shock, or anesthesia may have upon tolerance to PGE, infusion. The present results do indicate that it is possible to reduce microaggregate and fibrin formation throughout 21 days of blood storage. Possible prolongation of platelet survival is also suggested. Microaggregate embolism may contribute to respiratory failure following massive t r a n ~ f u s i o n ,and ~~ dilutional thrombocytopeniaZ1 appears to be the most common cause of coagulation abnormalities following massive transfusion. PGE, treatment of blood at time of collection may alleviate both of these problems.
References I.
2.
3.
4.
5.
6.
7.
8.
9.
Arrington, P. J., and J. J. McNamara: Effect of aspirin on microaggregate formation in banked blood. Surgery 76:295, 1974. Becker, G. A,, M. K. Chalos, M. Tuccelli, and R. H. Aster: Prostaglandin E, in preparation and s t o r a g e of platelet concentrates. Science 175538, 1972. -, T . Kunicki, and R. H. Aster: Effect of Prostaglandin E, on harvesting of platelets from refrigerated whole blood. J . Lab. Clin. Med. 83:304, 1974. Blaisdell, F. W., and R. M. Schlobohm: The respiratory distress syndrome: a review. Surgery 74:251, 1973. Brecher, G., M. Schneiderman, and E. P. Cronkite: The reproductibility and constancy of the platelet count. Am. J. Clin. Pathol. 23:15, 1953. Carlson, L. A,, L. Ekelund, and L. Oro: Clinical and metabolic effects of different doses of Prostaglandin E, i n man. A c t a Med. S c a n d . 183:423, 1968. Connell, R. S., and R. L. Swank: Pulmonary microembolism after blood transfusions. Ann. Surg. 177:40, 1973. Cullen, D. J., and L. Ferrara: Comparative evaluation of blood filters. Anesthesiology 41:568, 1974. Gervin, A. S., T. J . Limbird, C. L. Puckett, and D. Silver: Ultrapore hemofiltration: the effects on the coagulation and fibrinolytic mechanisms i n
10.
I I.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
23.
24. 25.
28 1
fresh and stored blood. Arch. Surg. 106:333, 1973. Goldiner, P. L., W. S. Howland, and C. Ray: Filter for prevention of microembolism during massive transfusions. Anesth. Analg. 51:717, 1972. Harp, J. R., M. Q. Wyche, B. E. Marshall, and H. A. Wurzel: Some factors determining rate of microaggregate formation in stored blood. Anesthesiology 40:398, 1974. Hissen, W., and R. L. Swank: Screen filtration pressure and pulmonary hypertension. Am. J. Physiol. 209:715, 1965. Jenevein, E. P., and D. L. Weiss: Platelet microemboli associated with massive blood transfusion. Am. J. Pathol. 45:313, 1964. Kinlough-Rathbone, R. L., M. A . Packham, and J. F. Mustard: The effect of Prostaglandin E, on platelet function in vitro and in viva Br. J. Haematol. 19559, 1970. Kloeze, J.: Influence of Prostaglandins on platelet adhesiveness and platelet aggregation. In Proceedings of the 2nd Nobel Symposium, Stockholm, June 1966. S. Bergstrom, and B. Samuelsson, Eds. New York, Interscience, 1967, p.241. Lim, R. C., Jr. C. Olcott IV, A . J . Robinson, and F. W. Blaisdell: Platelet response and coagulation changes following massive blood replacement. J. Trauma 13577, 1973. Marshall, R. E., N. Ellison, H. A . Wurzel, et a/.: Microaggregate formation in stored blood: 11. Influence of anticoagulants and blood components. Circ. Shock 2:185, 1975. -, L. R. Soma, J. R. Harp, e / a/.: Microaggregate formation in stored blood. I. Species differences and storage time. Circ. Shock 2:175, 1975. -, L. R. Soma, J. R. Harp, G. R. Neufeld, H. A. Wurzel, and D. C. Dodd: Pulmonary function after exchange transfusion of stored blood in dogs. Ann. Surg. 179:46, 1974. -, H. A . Wurzel, N. Ellison, et a/.: Microaggregate formation in stored blood: 111. Comparison of Bentley, Fenwal, Pall, and Swank micropore filtration. Circ. Shock 2:249, 1975. Miller, R. D., T . 0. Robbins, J. M. Tong, and S. L. Barton: Coagulation defects associated with massive blood transfusions. Ann Surg. 174:794, 1971. McNamara, J. J., D. Boatright, E. L. Burran, el a/.: Changes in some physical properties of stored blood. Ann. Surg. 17458, 1971. -, E. L. Burran, E. Larson, G. Omiya, G. Suehiro, and H. Yamase: Effect of debris in stored blood on pulmonary microvasculature. Ann. Thor. Surg. 14:133, 1972. -, E. L. Burran, and G. Suehiro: Effective filtration of banked blood. Surg. 71594, 1972. -, M. D. Molot, and J . F. Stremple: Screen filtration pressure in combat casualities. Ann. Surg. 172334, 1970
282 26. 27.
28.
29.
30.
31.
32.
33.
HARP, ET A L . Moore, F. D.: Terminal mechanisms in human injury. Am. J. Surg. 110:317, 1965. Moseley, R. V., and D. B. Doty: Changes in the filtration characteristics of stored blood. Ann. Surg. 171:329, 1970. -, and D. B. Doty: Death associated with multiple pulmonary emboli soon after battle injury. Ann. Surg. 171:336, 1970. Pontoppidan, H., B. Geffin, and E. Lowenstein: Acute respiratory failure in the adult. N. Engl. J . Med. 287:690, 1972. Reul, G. J., Jr., A . C . Beall, and S . D. Greenberg: Protection of the pulmonary microvasculature by fine screen filtration. Chest 66:4, 1974. Shio, H., and P. W. Ramwell: Prostaglandin E, in platelet harvesting: an in vitro study. Science 175536, 1972. Solis, R. T., and M. B. Gibbs: Filtration of the microaggregates in stored blood. Transfusion 12:245, 1972. Swank, R. L.: Alteration of blood on storage:
34.
Transfusion May-June 1976
m e a s u r e m e n t of adhesiveness of “aging” platelets and leucocytes and their removal by filtration. N. Engl. J . Med. 265:728, 1961. Weiss, H. J., L . M. Aledort, and S. Kochwa: The effect of salicylates on the hemostatic properties of platelets in man. J. Clin. Invest. 47:2169, 1968.
J . R . H a r p , M.D., Professor & C h a i r m a n of Anesthesiology, Temple University Health Science Center, 3400 N. Broad Street, Philadelphia, Pa. 19140. B. E. Marshall, M.D., Professor of Anesthesiology, Hospital of the University of Pennsylvania, 3400 Spruce Street, Philadelphia, Pa. 19104. H.A . Wurzel, M.D., Associate Professor of Medicine and Pathology, Hospital of the Univeristy of Pennsylvania. A. S. Miller, B.S., Medical Technologist, Hospital of the University of Pennsylvania.
