Platelet dysfunction in acute type A aortic dissection evaluated by the laser light-scattering method

Transcription

1 Platelet dysfunction in acute type A aortic dissection evaluated by the laser light-scattering method Masashi Tanaka, MD Koji Kawahito, MD Hideo Adachi, MD Takashi Ino, MD Objectives: Platelet dysfunction contributes to bleeding tendency in acute type A aortic dissection. Particle counting by new laser light-scattering methods more accurately quantifies changes in the number of different-sized platelet aggregates than do conventional optical density methods. We studied platelet aggregation kinetics and patterns of aggregation deficiency in acute-phase aortic dissection with laser light scattering. Dr Tanaka Methods: Blood from 20 acute type A aortic dissection patients undergoing surgery was sampled during acute ( hours from onset) and chronic (postoperative day 20, control) phases of aortic dissection. Platelet count and aggregability were assessed by optical density and laser light-scattering methods after aggregation was induced (addition of 2.0 g/ml collagen to samples). From the Department of Cardiovascular Surgery, Omiya Medical Center, Jichi Medical School, Saitama, Japan. Received for publication March 5, 2003; revisions requested March 25, 2003; revisions received April 6, 2003; accepted for publication April 15, Address for reprints: Masashi Tanaka, MD, Department of Cardiovascular Surgery, Omiya Medical Center, Jichi Medical School, Amanuma, Saitama , Japan ( masashi@omiya.jichi.ac. jp). J Thorac Cardiovasc Surg 2003;126: Copyright 2003 by The American Association for Thoracic Surgery /2003 $ doi: /s (03) Results: Optical density showed significant reduction in acute-phase platelet aggregation (acute vs chronic: 65 27% vs 77 17%, P.03). Laser light scattering showed significant reduction in medium (25-50 m) and large (50-70 m) but not small aggregate (9-25 m) generation (acute vs chronic: small, vs , NS; medium, vs , P.001; large, vs , P.001). Acute- versus chronic-phase platelet counts were significantly decreased ( / L vs / L, P.001). Conclusions: Platelet aggregation is suppressed in acute-phase aortic dissection. This suppression does not occur in the initial phase of small aggregate formation; rather, it occurs during the conglomeration of small aggregates into larger aggregates.recent reports show improved surgical outcomes in patients with acute type A aortic dissection. However, acute type A dissection continues to be associated with high mortality and morbidity rates, 1-7 and the associated bleeding tendency can be life-threatening. Some reports describe an acute hemorrhagic state typically associated with arterial dissection One of the greatest contributors to this bleeding tendency may be loss of platelet aggregability. Acute massive consumption coagulopathy in the false lumen may affect platelet function, but the mechanism underlying platelet dysfunction has not been fully investigated. Platelet aggregation is conventionally evaluated by measuring optical density (OD) changes 11 or by impedance analysis. 12 However, neither method provides information about temporal changes in the number of platelet aggregates of different sizes after stimulation with an aggregation agent. The newly developed laser light-scattering (LS) method (AG-10 aggregometer; Kowa Co, Ltd, Tokyo, Japan) facilitates selective detection of aggregates of different sizes and is more sensitive than conventional methods. We used this new method to investigate the aggregation The Journal of Thoracic and Cardiovascular Surgery Volume 126, Number 3 837

2 Tanaka et al TABLE 1. Clinical characteristics and operative results of study patients Mean age (years) Age range (years) Sex (male/female) 10/10 Dissection extent Ascending aorta and aortic arch only n 2 (10) To the descending aorta n 18 (90) Time from onset of symptoms until blood collection (hours) Surgery Ascending aortic replacement n 17 (85) Aortic root replacement n 2 (10) Total arch replacement n 1 (5) Hospital mortality n 0 Postoperative complications Reexploration for bleeding n 1 (5) Stroke n 0 Total amount of blood transfusion (units) Data are shown as mean SD unless otherwise indicated, and numbers in parentheses are percentages. kinetics of platelets of different sizes in the acute and chronic phases of aortic dissection and to evaluate the pattern of loss of aggregation in acute type A aortic dissection. Patients and Methods The study subjects comprised 20 patients (10 men and 10 women) who underwent surgery for acute type A aortic dissection at our hospital between April 2000 and February Mean age was years (range, years). Seventeen patients underwent graft replacement of the ascending aorta with supracoronary prosthesis, 2 patients underwent aortic root replacement (modified Bentall method), and 1 patient underwent total arch replacement with antegrade selective cerebral perfusion (Table 1). Informed consent was obtained from each patient before surgery. Although all 20 patients underwent emergency surgery within 48 hours of the onset of symptoms, they were hemodynamically stable at the time of admission and the initial blood draw. An arterial blood sample was obtained during the acute phase of aortic dissection before surgery ( hours from onset) and during the chronic phase on postoperative day 20 (control). In addition, blood samples were taken from healthy volunteers and used as reference values (n 8). Platelet Aggregation Studies The LS method determines variations in particle size and concentration in platelet-rich plasma by detecting the respective intensities of light scattered by particles passing through a laser beam. The principles of the LS method have been described previously. 13,14 Briefly, an He-Ne laser beam with a diameter of 40 m is passed through 300 L of platelet-rich plasma that is being rotated in a small cylindrical glass cuvette. Light scattered by particles in the observation volume is detected by a 4-channel photodiode array; light intensity directly corresponds to particle size. Each of the 4 photodiodes detects the intensity of light scattered by particles within its corresponding observation volume. The amplitude and frequency of alternating current signals were analyzed on a personal computer (PC-98 As2; NEC, Tokyo, Japan) according to the algorithm derived by Lentz from the Mie scattering theory. 15 The signal frequency was recorded at 10-second intervals. To measure platelet aggregation during the acute and chronic phases, 2.0 ml of whole blood was collected, and platelet-rich plasma was obtained by centrifugation at 120 g for 10 minutes at room temperature followed by aspiration of the supernatant. Platelet-rich plasma aggregation was determined by simultaneously evaluating the maximum percentage of decrease in OD and assessing the LS intensity; the AG-10 was used for both methods. With the OD method, output from the aggregometer was adjusted so that the difference in light transmittance between platelet-rich plasma and platelet-poor plasma was 100%. Aggregation was induced by addition of 2.0 g/ml collagen (final concentration; Sigma Chemical Co, St Louis, Mo). After collagen stimulation, small aggregates quickly formed in the first phase of aggregation and larger aggregates formed in the second phase. Data are expressed as the change over time in the number of aggregates of each of 3 sizes (determined by light intensity and expressed in millivolts). Particles with an intensity of 25 to 400 mv were counted as small aggregates (9-25 m), those with an intensity of 400 to 1000 mv were counted as medium aggregates (25-50 m), and those with an intensity of 1000 to 2048 mv were counted as large aggregates (50-70 m). Data were recorded on a 2-dimensional graph showing the change over time of total light intensity expressed as a cumulative summation at 10-second intervals of scattered light intensity and the number of particles of each size. Total intensity was recorded at 10-second intervals for 9 minutes. The number of particles of each size was estimated by quantifying the area under the curve, which represents the sum of measurements of LS intensity. Statistical Analysis All results are expressed as mean SD. Data were compared, and differences between pre- and postoperative values were analyzed by paired t test. P.05 was regarded as statistically significant. All analyses were performed with the StatView v5.0 statistical package (SAS Institute, Cary, NC). Results Operative Results There were no in-hospital deaths or postoperative strokes in this series. Reexploration for bleeding was required in 1 of the 20 patients (5%). The average number of blood transfusions per patient was units (Table 1). Platelet Counts Preoperative platelet counts were significantly lower than postoperative platelet counts (acute vs chronic phase: / L vs / L, P.001; Table 2). Platelet Aggregation Determined by OD Assay Aggregation stimulated by collagen and measured by the conventional OD method was significantly reduced in the 838 The Journal of Thoracic and Cardiovascular Surgery September 2003

3 acute phase (acute vs chronic phase: 65 27% vs 77 17%, P.03; Table 2). Platelet Aggregation Pattern Determined by Laser LS Assay A typical pattern of size-dependent LS intensity after stimulation with collagen in patients with acute type A aortic dissection is shown in Figure 1. Changes in size-dependent aggregate quantity (area under the curve) are shown in Table 2. The number of small aggregates did not differ significantly between the acute and chronic phases of aortic dissection. In contrast, medium and large aggregate formations were significantly inhibited in the acute phase (acute vs chronic phase: small, vs , NS; medium, vs , P.001; large, vs , P.001; Table 2). Discussion Platelet aggregation is a 3-phase response that involves shape change, microaggregation (primary aggregation), and macroaggregation (secondary aggregation). 11 Our results suggest that although the formation of small aggregates is preserved, development of microaggregation into macroaggregation is suppressed in the acute phase of aortic dissection. Previous investigators have shown that macroaggregation, the formation of large stable aggregates, is also impaired by cardiopulmonary bypass, whereas microaggregation, the formation of aggregates containing up to 100 platelets each, is well preserved The suppression of macroaggregation in the context of acute type A dissection indicates that small aggregates do not transform into medium and larger aggregates. Zilla and coworkers 20 reported on the basis of a scanning and transmission electron microscopy study that the morphology of circulating platelets recovers after initial activation despite the maintenance of cardiopulmonary bypass. The initial platelet consumption and dysfunction during cardiopulmonary bypass is caused by reversible primary rather than irreversible secondary aggregation phenomena. The small aggregates of platelets may be activated by additional triggers such as increased shear stress, 21,22 activated leukocytes, 23,24 and activated complement factors, 23 resulting in the development of irreversible macroaggregation, which is a prerequisite for thromboembolic events because it reinforces the platelet plug. Consumption coagulopathy caused by extensive thrombus formation can occur in the uncomplicated arterial aneurysm as well as in acute aortic dissection. 25 Fisher and associates 26 reported that only 3 out of 76 (4%) patients with stable aortic aneurysm showed evidence of clinical consumptive coagulopathy, although laboratory tests TABLE 2. Acute- and chronic-phase platelet aggregation measured by optical density and light-scattering methods Measurement Acute phase Chronic phase Reference value Platelet count ( 10 5 / L) OD (%)* LS ( 10 7 /300 L) Small Medium Large Acute phase is just after onset ( hours from onset), and chronic phase is on postoperative day 20 (control). Blood samples were taken from healthy volunteers and used as reference values. Results are shown as mean SD unless otherwise indicated. *Optical density (OD) is expressed as maximum % reduction in optical density. The number of particles of each size estimated by quantifying the area under the curve representing the sum of measurements of laser light scattering (LS) intensity. P.001 vs acute phase. P.03 vs acute phase. showed altered fibrinogen and fibrin degradation products in 39% of the patients. Some reports regarding the mechanism of coagulopathy in patients with acute aortic dissection have emphasized abnormality of the coagulation and fibrinolysis systems. Ten Cate and associates 9 proposed 3 possible mechanisms leading to coagulation deficiency in patients with acute aortic dissection: activation of factor XII by subendothelial tissues such as collagen and subsequent activation of the intrinsic coagulation system; activation of factor VII by thromboplastic aortic wall material; and triggering of fibrinolysis mediators located in the aortic adventitia. Although coagulopathy associated with acute aortic dissection is well discussed from the perspective of coagulation and fibrinolysis system abnormalities, platelet dysfunction should definitely be considered as a factor in the bleeding tendency. Our results suggest that impaired platelet aggregation also contributes to perioperative hypocoagulability and blood loss. Use of the LS method to enhance coagulation monitoring could provide for preoperative recognition of platelet dysfunction, thereby providing a strategy for clotting factor administration and cellular replenishment that can be adapted to the patients individual requirements. Aortic dissection requires extensive surgery associated with cardiopulmonary bypass and hypothermic circulatory arrest. In the context of acute aortic dissection surgery, multiple interrelated factors, including interference with the vascular integrity, extensive surgical dissection, blood replenishment, hemodilution, hypothermia, ischemia and reperfusion, and activation of systemic inflammatory responses, may cause excessive blood loss. Despite exposure to these nonphysiological conditions, platelet aggregability as determined by the LS method recovers 3 weeks after surgery. The Journal of Thoracic and Cardiovascular Surgery Volume 126, Number 3 839

4 Tanaka et al Figure 1. Typical patterns of platelet aggregation determined by optical density (OD) and laser light-scattering (LS) assays of blood samples obtained from a 77-year-old woman diagnosed with acute type A aortic dissection in the acute phase (A) and chronic phase (B) (agonist: 2.0 g/ml collagen). Small aggregates (S), medium aggregates (M), and large aggregates (L) are measured by LS assay. Platelet aggregation capacity (T) is measured by OD assay. Small aggregates are formed in the first phase of aggregation, and larger aggregates are formed in the second phase. After stimulation, the quantity of small aggregates is increased transiently and the quantity of larger aggregates is increased after that. Arrow indicates the point at which agonist is added. The LS assay scale is depicted on the left; the OD scale is depicted on the right. One possible limitation of the study involves comparison of the medium and large aggregates between the acute and chronic phases. The samples drawn in the acute phase had a significantly smaller total number of platelets than that of samples drawn in the chronic phase. However, the samples drawn in the chronic phase had a disproportionately higher number of medium and large aggregates in comparison with samples drawn in the acute phase, suggesting that the blood in the preoperative samples had not only diminished total platelets but also impaired platelet function. In conclusion, platelet aggregation appears to be suppressed in patients with acute aortic dissection. Although the generation of medium and large aggregates estimated by LS is significantly decreased in patients with acute aortic dissection, there is no significant change in the number of small aggregates. Thus, platelet dysfunction associated with acute aortic dissection may be caused mainly by inhibition of the conglomeration of small aggregates into larger aggregates. References 1. Hagan PG, Nienaber CA, Isselbacher EM, et al. The International Registry of Acute Aortic Dissection (IRAD): new insights into an old disease. JAMA. 2000;283: Pansini S, Gagliardotto PV, Pompei E, et al. Early and late risk factors in surgical treatment of acute type A aortic dissection. Ann Thorac Surg. 1998;66: Bachet J, Goudot B, Dreyfus GD, et al. Surgery for acute type A aortic dissection: the Hopital Foch experience ( ). Ann Thorac Surg. 1999;67: Sabik JF, Lytle BW, Blackstone EH, McCarthy PM, Loop FD, Cosgrove DM. Long-term effectiveness of operations for ascending aortic dissections. J Thorac Cardiovasc Surg. 2000;119: David TE, Armstrong S, Ivanov J, Barnard S. Surgery for acute type A aortic dissection. Ann Thorac Surg. 1999;67: Kawahito K, Adachi H, Yamaguchi A, Ino T. Preoperative risk factors for hospital mortality in acute type A aortic dissection. Ann Thorac Surg. 2001;71: Kawahito K, Adachi H, Yamaguchi A, Ino T. Early and late surgical outcomes of acute type A aortic dissection in patients aged 75 years and older. Ann Thorac Surg. 2000;70: Fine NL, Applebaum J, Elguezabal A, Castleman L. Multiple coagulation defects in association with dissecting aneurysm. Arch Intern Med. 1967;119: ten Cate JW, Timmers H, Becker AE. Coagulopathy in ruptured or dissecting aortic aneurysms. Am J Med. 1975;59: Puxeddu A, Ribacchi R, Scionti L, Gatteschi C, Berrettini M. Disseminated intravascular coagulation in dissecting aortic aneurysm. A case report. Panminerva Med. 1981;23: Born GV. Observations on the change in shape of blood platelets brought about by adenosine diphosphate. J Physiol. 1970;209: Cardinal DC, Flower RJ. The electronic aggregometer: a novel device for assessing platelet behavior in blood. J Pharmacol Methods. 1980; 3: Ozaki Y, Satoh K, Yatomi Y, Yamamoto T, Shirasawa Y, Kume S. Detection of platelet aggregates with a particle counting method using light scattering. Anal Biochem. 1994;218: Tohgi H, Takahashi H, Watanabe K, Kuki H, Shirasawa Y. Development of large platelet aggregates from small aggregates as determined by laser-light scattering: effects of aggregant concentration and antiplatelet medication. Thromb Haemost. 1996;75: Lentz WJ. Generating Bessel function in Mie scattering calculation using continued fractions. Appl Opt. 1976;15: Tanaka M, Kawahito K, Adachi H, Isawa H, Ino T. Platelet damage caused by the centrifugal pump: laser-light scattering analysis of aggregation patterns. Artif Organs. 2001;25: Friedenberg WR, Myers WO, Plotka ED, et al. Platelet dysfunction associated with cardiopulmonary bypass. Ann Thorac Surg. 1978;25: Rinder CS, Bohnert J, Rinder HM, Mitchell J, Ault K, Hillman R. Platelet activation and aggregation during cardiopulmonary bypass. Anesthesiology. 1991;75: The Journal of Thoracic and Cardiovascular Surgery September 2003

5 19. Kestin AS, Valeri CR, Khuri SF, et al. The platelet function defect of cardiopulmonary bypass. Blood. 1993;82: Zilla P, Fasol R, Groscurth P, Klepetko W, Reichenspurner H, Wolner E. Blood platelets in cardiopulmonary bypass operations. Recovery occurs after initial stimulation, rather than continual activation. J Thorac Cardiovasc Surg. 1989;97: O Brien JR. Shear-induced platelet aggregation. Lancet. 1990;335: Tabuchi N, Huet RC, Sturk A, Eijsman L, Wildevuur CR. Hemostatic function of aspirin-treated platelets vulnerable to cardiopulmonary bypass. Altered shear-induced pathway. J Thorac Cardiovasc Surg. 1995;110: Wachtfogel YT, Kucich U, Hack CE, et al. Aprotinin inhibits the contact, neutrophil, and platelet activation systems during simulated extracorporeal perfusion. J Thorac Cardiovasc Surg. 1993;106: Primack C, Walenga JM, Koza MJ, Shankey TV, Pifarre R. Aprotinin modulation of platelet activation in patients undergoing cardiopulmonary bypass operations. Ann Thorac Surg. 1996;61: Bieger R, Vreeken J, Stibbe J, Loeliger EA. Arterial aneurysm as a cause of consumption coagulopathy. N Engl J Med. 1971;285: Fisher DF Jr, Yawn DH, Crawford ES. Preoperative disseminated intravascular coagulation associated with aortic aneurysms. A prospective study of 76 cases. Arch Surg. 1983;118: The Journal of Thoracic and Cardiovascular Surgery Volume 126, Number 3 841