Loss of platelet fibrinogen receptors during clinical cardiopulmonary bypass

In 10 patients, cardiopulmonary bypass decreased the number of fibrinogen binding sites from 31,730 +/- 12,802 per platelet to 18,590 +/- 9,644 per platelet. Bypass also decreased the amount of the platelet membrane glycoprotein IIIa, which is part of the fibrinogen receptor complex, from 17.1 +/- 3.6 ng/10(9) platelets to 12.9 +/- 4.7. The fibrinogen binding constant did not change. Platelet sensitivity to adenosine diphosphate did not change; however, template bleeding times increased from 5.2 +/- 1.5 minutes before bypass to 8.5 +/- 2.3 minutes after bypass. Analysis of detergent washings from the perfusion circuit after bypass in five patients indicated that platelet material remains attached to the surface as membrane fragments and degranulated platelets. These data further elucidate the mechanism of platelet loss and dysfunction during cardiopulmonary bypass and highlight the importance of platelet membrane fibrinogen receptors and surface adsorbed fibrinogen in this process.     

The platelet in cardiopulmonary bypass

Platelets are the smallest of the blood cells and are known to be activated during cardiopulmonary bypass. They play a role in many associated complications. Both quantitative and qualitative platelet defects have been demonstrated, resulting in microvascular hemorrhage and thromboembolism. As their interactions with endothelium and other blood cells are unraveled, the important contribution they make toward the systemic inflammatory response to operation seen in cardiopulmonary bypass is increasingly evident. In this review, we consider platelet activation during cardiopulmonary bypass, the resultant clinical effects, and potential approaches to therapy and prevention.     

体外循环期间血小板膜糖蛋白的定量研究及临床意义

目的　探讨体外循环 (CPB)对血小板膜糖蛋白的影响。方法　用流式细胞术对 40例先天性心脏病房、室间隔缺损患者在 CPB下行心内修补术时的血小板膜糖蛋白 CD41 a、CD42 b、CD6 2 p和CD6 3的含量免疫荧光定量测定 ,同时与本组患者和健康供血者试管内血小板膜糖蛋白进行比较。结果　在 CPB中血小板总数明显下降 ,从术前值的 (310± 91)× 10 3/ m l降至术后值的 (181± 44 )× 10 3/ml,CD41 a无明显变化 ,CD42 b明显降低 (P<0 .0 5 ) ,CD6 2 p和 CD6 3明显增加 (P<0 .0 5 )。本组和健康供血者的试管内血小板膜糖蛋白 CD41 a、CD42 b、CD6 2 p和 CD6 3均呈平行变化。结论　 CPB期间既有粘附凝集功能下降 ,也有粘附凝集功能增强的血小板膜糖蛋白 ,而凝血机制缺陷的原因可能与血小板总数下降、低温及肝素的后续作用有关。     

Inhibition of platelet receptors involved in neutrophil-platelet interaction in model cardiopulmonary bypass

We investigated the interactions between neutrophils, platelets, and artificial surfaces, and whether blocking of relevant receptors on platelets reduced unwanted activation responses in model cardiopulmonary bypass. Isolated neutrophils and platelets resuspended in heparin-anticoagulated plasma were recirculated with and without blocking antibodies to CD62P, CD42b, or junctional adhesion molecule C (JAM-C) in polyvinyl chloride tubing using a roller pump. Platelet adhesion to the tubing was inhibited by anti-CD42b and anti-CD62P, and adhesion of neutrophils by anti-JAM-C. Formation of platelet-neutrophil and platelet aggregates was reduced by anti-CD62P. Anti-JAM-C decreased platelet-neutrophil aggregation at low concentrations and platelet macroaggregates at high concentrations. Anti-CD62P increased neutrophil CD11b expression but not degranulation. Anti-JAM-C substantially increased neutrophil degranulation and slightly increased CD11b expression. Platelet activation increased when CD62P was blocked and decreased with anti-CD42b antibody. High-dose anti-JAM-C reduced platelet activation. In conclusion, inhibiting platelet and neutrophil-platelet interactions had useful effects but no single blocking antibody seemed capable of inducing only beneficial effects.     

Hemofiltration during cardiopulmonary bypass.

Several factors combine to facilitate the evolution towards heart and multi-organ failure following cardiac surgery. Some of these factors are related to pure cardiac aspects like the existence of a preoperative heart disease, the use of aortic cross clamping or performance of cardiotomy. Cardiopulmonary bypass (CPB) also plays an important role in the occurrence of postoperative organ dysfunctions by two principal means: firstly by inducing a profound hemodilution, which impairs oxygen transport through tissues. This phenomenon is pointed out in the postoperative period by the existence of increased transpulmonary O2 gradients, extravascular lung water volume and subsequent impairments of O2 transport. Secondly CPB is deleterious by triggering an important inflammatory reaction. This reaction is largely related to the ratio of the circuit area to the patient's body surface area and is therefore maximal in children. It has been widely demonstrated that the very early paths of this reaction imply several humoral factors including kinins, coagulation factor-XII and complement fragments. The activation of these factors is self-amplified and triggers both expression and release of numerous mediators by endothelial cells and leukocytes. Finally, these mediators are responsible for the well described "post-bypass syndrome" which is, from a clinical viewpoint, very close to hyperkinetic septic shocks. Several methods have been proposed to reduce the deleterious effects of both cardiac surgery and CPB. The older one is hypothermia that considerably reduces the triggering of the inflammatory mediators network. Heparin-coated circuits may also reduce this reaction to some extent. Hemofiltration has been introduced in the 90's in CPB management. Because of its very high tolerance in patients with compromised circulatory status this technique was already used in the postoperative period to treat patients with acute renal failure. Initially hemofiltration was intended to correct the accumulation of extravascular water during or immediately following the surgical procedure. Nevertheless several of its "side-effects" appeared to be useful like reduction of postoperative blood loss and immediate hemodynamics improvement. Several studies attempted to point out the mechanism of action of hemofiltration and although removal of inflammatory mediator occurs, there is currently no proofs that this removal is the actual mechanism by which this technique acts. At the early beginning of the use of its utilization hemofiltration during cardiac surgery aimed either to concentrate blood at the end of the procedure or to rapidly restore a normal fluid and electrolytes balance. Today some new implementations of this technique are proposed either to reduce the triggering of the inflammatory reaction to CPB or to reduce the immediate postoperative drug support.     

Leukocyte activation with increased expression of CR3 receptors during cardiopulmonary bypass.

The effects of cardiopulmonary bypass (CPB) on the expression of leukocyte adhesive receptors, ie, complement receptor type 3 (CR3), were studied in 16 patients. The CR3 expression on leukocytes was determined by time-resolved fluoroimmunoassay on a standardized number of cells isolated from blood samples taken during various times during CPB. The results demonstrated that CR3 expression on leukocytes increased immediately after the start of CPB (p less than 0.05), concomitant with an early sharp increase of plasma concentrations of C3a (p less than 0.01). After release of the aortic cross-clamp, a second peak of leukocyte CR3 expression was induced (p less than 0.05), paralleled by a significant increase of leukotriene B4 (p less than 0.05) and elastase (p less than 0.05) levels in the late period of CPB. In vitro studies with leukocytes isolated from healthy donors (n = 5) showed a dose-dependent increase of CR3 expression stimulated by zymosan-activated plasma, indicating that the rapid CR3 expression on leukocytes is likely mediated by complement activation. However, the mechanisms for the second peak of leukocyte CR3 expression during CPB remain to be further elucidated. In conclusion, CR3 expression on leukocytes increased immediately after the start of CPB and was followed by a second peak of expression in the late phase of CPB. Pharmacological blockage of these adhesive receptors might reduce the leukocyte-mediated deleterious effects of CPB.     

肝素涂层管道对体外循环中血小板的保护作用

目的：探讨常规剂量肝素体外循环（CPB）下使用肝素涂层CPB管道（HCC）对血小板的影响。方法：机械瓣膜置换术患者随机分为HCC组（n=8）和对照组（n=15）。分别在CPB前、CPB60min、肝素中和后30、60min及手术后12h测定血小板计数（PLT），血浆α颗粒膜蛋白－140（GMP－140）的浓度及手术后12h出血量。利用电子显微镜观察两组转流后微栓过滤网上沉积物的附着情况。结果；对照组PLC在转流中、后显著低于HCC组，差异有显著性（P＜0．05），且血浆GMP－140浓度在转流60min、转流中和后30、60min显著高于HCC组，差异有显著性（P＜0．05），但手术后12h差异无显著性（P＞0．05）；HCC组手术后12h出血量较对照组少，差异有显著性（P＜0．05）；HCC组动脉过滤网表面光洁，网眼边缘清晰，偶见白细胞和血小板附着；对照组动脉过滤网表面可见纤维蛋白沉积，网眼边缘模糊，有大量白细胞及少数活化血小板粘附。结论：HCC在CPB中对血小板具有保护作用，较好地改善CPB装置的血液相容性。     

体外循环中止血芳酸对血小板的保护作用

目的：探讨止血芳酸在体外循环中对血小板的保护作用及其临床意义。方法：测定止血芳酸组和对照组血小板数量、出血量和输血量,及体外循环前后血浆α－颗粒膜蛋白（ＧＭＰ－１４０）,血栓烷Ｂ２（ＴＸＢ２）和６－酮－前列腺素Ｆ１a(6-k-PGF1a）的浓度变化。结果,体外循环术后血小板计数组明显高于对照组,出血量明显少于对照组。ＧＭＰ－１４０、ＴＸＢ２上升幅度小于对照组,二者相比差异显著（Ｐ＜０．０５）。结论：止血芳酸在体外循环术中可以起到保护血小板,减少术后出血的作用。     

Heparin-induced platelet dysfunction and cardiopulmonary bypass

Background. Cardiopulmonary bypass is associated with impaired platelet macroaggregation. Heparin contributes to platelet dysfunction before extracorporeal circulation. In vitro heparinization of whole blood does not impair macroaggregation. Heparin releases several endothelial proteins; thus heparin may inhibit macroaggregation indirectly.

Methods. Patients undergoing operations using cardiopulmonary bypass and ABO blood group compatible volunteers were studied. Whole blood impedance aggregometry assessed macroaggregation in response to collagen (0.6 μg ml⁻¹) in blood diluted either with normal saline or with platelet poor plasma, obtained from patients at different stages of cardiopulmonary bypass.

Results. Before heparinization, blood diluted with its own platelet poor plasma recorded an impedance change of 13.0 (4.7 to 15.6) Ohms. Platelet poor plasma obtained after heparinization or during extracorporeal circulation reduced this response to 3.7 (1.1 to 8.4) and 2.0 (1.1 to 3.3) Ohms, respectively (both p < 0.0001 versus pre-heparin; n = 13). Macroaggregation in blood from volunteers was similarly inhibited by patients’ platelet poor plasma (n = 30). The macroaggregatory response in blood sampled after heparinization for cardiopulmonary bypass, decreased gradually from 11.4 (8.2 to 15.9) Ohms immediately after sampling to 1.7 (1.4 to 4.1) Ohms 2 hours later (p < 0.0001; n = 11).

Conclusions. In vivo heparinization induces plasma changes that inhibit platelet macroaggregation. This is an indirect, delayed inhibition that is transferable in vitro to normal platelets.     

Differential expression of neutrophil adhesion molecules during coronary artery surgery with cardiopulmonary bypass.

BACKGROUND: Activation of neutrophil adhesion molecules and subsequent neutrophil adhesion to vascular endothelium are key events initiating inflammatory organ dysfunction after cardiopulmonary bypass and ischemic reperfusion. OBJECTIVES: We sought to characterize neutrophil integrin CD11b and L-selectin activation associated with coronary artery bypass graft surgery and to determine whether neutrophil activation contributes to their sequestration on postbypass reperfusion. METHODS: Twenty patients undergoing routine coronary artery bypass were studied. Heparinized whole blood was simultaneously sampled from a central venous line, aorta, coronary sinus, and right and left atrium before, during, and up to 20 minutes after cardiopulmonary bypass. Neutrophil counts were obtained, and neutrophil CD11b and L-selectin expression was determined by flow cytometric analysis in whole blood. RESULTS: CD11b expression on circulating neutrophils increased during cardiopulmonary bypass, peaking at 145% of baseline level after release of the aortic clamp and then declined by 20 minutes after bypass (analysis of variance, P =.003). No change in neutrophil L-selectin expression was observed during cardiopulmonary bypass. Neutrophils responded to ex vivo stimulation by C5a and leukotriene B(4) during cardiopulmonary bypass but not at 24 hours after the operation. After reperfusion, neutrophil loss, but not local activation, was demonstrated in the coronary and pulmonary circulations. CONCLUSIONS: Upregulated CD11b expression on neutrophils is likely to contribute to neutrophil sequestration in the heart and lungs after bypass, but neutrophil activation may be limited by their reduced responsiveness to agonist stimulation. CD11b represents a potential therapeutic target for diminishing inflammation after cardiac operations.     

Expression of chemokine receptors CXCR1 and CXCR2 during cardiopulmonary bypass.

OBJECTIVE: This study investigated the effects of cardiopulmonary bypass on neutrophil expression of chemokine receptors, CXCR1 and CXCR2, and the beta2 integrin CD11b. METHODS: Ten patients undergoing coronary artery grafting with cardiopulmonary bypass were studied. Blood samples were collected preoperatively, before bypass, at termination of bypass, and 12 to 18 hours postoperatively. In vitro studies were performed on control subjects to determine changes in the surface expression of CXCR1, CXCR2, and CD11b on stimulation with interleukin 8. Receptor expression was measured by flow cytometry. Plasma levels of interleukin 8 from the patients were determined by enzyme-linked immunoassay. RESULTS: After bypass, CXCR2 expression fell by 66% (P <.0001) and remained low postoperatively (P <.0001). CXCR1 expression persisted at preoperative levels. CD11b expression increased significantly after bypass (P <.0001), returning to prebypass levels postoperatively. In vitro studies showed a dose-related fall of both CXCR1 (P <.0001) and CXCR2 expression (P <.0001) and a significant rise in CD11b expression (P <.0001). Plasma interleukin 8 increased significantly after bypass (P <.0001), remaining elevated 12 to 18 hours postoperatively (P =.02). Correlations between interleukin 8 levels and CXCR2 expression (P <.0001) and CD11b expression (P <.03) were demonstrated. CONCLUSIONS: CXCR2 expression is significantly down-regulated after bypass; in contrast, CXCR1 expression remains unchanged. In addition, whereas interleukin 8 is an important determinant of both CXCR1 and CXCR2 expression in vitro, it only correlates with CXCR2 and CD11b expression in vivo. This has implications in the search for antagonists against CXC chemokines and their receptors.     

Nitric oxide: platelet protectant properties during cardiopulmonary bypass/ECMO

Postoperative bleeding is a major complication in patients who have been placed on extracorporeal circulatory support for various cardiac procedures (1). The increase in hemorrhage is well documented and is associated with various factors, which include, high-dose systemic heparinization, thrombocytopenia, and impaired platelet function. Platelets activate when exposed to the large foreign surface of the extracorpeal circuit, with the largest area being the oxygenator. Despite adequate heparinization, platelet levels continue to decrease. This aggregation phenomenon has also been extensively studied, and it cannot be attributed to the use of aminocarproic acid, aprotinin, propofol, or amicar (2). Other factors found to be unrelated include, the brand or type of oxygenator, the use of heparin coatings (3), activated clotting time (ACT) levels while on bypass, the operative procedure, preoperative medications, or the types of anesthetic agents used (2). Therefore, it may be beneficial to add nitric oxide to the sweep gas to decrease platelet loss, platelet damage, postoperative bleeding, and lessening the need for post-operative blood transfusions.     

异丙酚对儿童体外循环血小板功能的影响

目的 探讨异丙酚对儿童体外循环(CPB)期间血小板(Plt)功能的影响。方法将40例CPB下行心脏修补术的先心病儿童随机分为对照组(C组)和异丙酚组(P组)。两组均以芬太尼5μg·kg-1·h-1辅助吸入0.7％异氟醚维持麻醉。P组自诱导至术毕持续泵注异丙酚3 mg·kg-1·h-1持续泵注。两组病儿分别于麻醉前、转流30min、手术结束时、术后24 h取中心静脉血测定血小板a-颗粒膜蛋白(GMP-140)和Plt血栓烷B2(TXB2)浓度,及全血激活凝固时间(ACT)、凝集速率、Plt功能(PF)、快速Plt功能(qPF)值,同时测定各时段的Plt计数。结果 两组GMP-140、TXB2在转流期间显著性升高(P<0.01);Plt计数在转流期间显著性下降(P<0.01);ACT转流期间显著性延长(P<0.01),术后恢复基础水平。PF、qPF转流期间测不出,术后恢复至基础水平。C组和P组之间Plt各观测值相比差异无显著性。结论 持续泵注异丙酚3 mg·k-1·h-1对儿童CPB中的血小板功能无明显影响。