Effect of heparin reversal and fresh platelet transfusion on platelet emboli post-cardiopulmonary bypass in a pig model.

Heparin reversal by protamine and fresh platelet transfusion may decrease bleeding complications post-cardiopulmonary bypass (CPB) and may increase the level of organ trapped platelet emboli. Platelet emboli were quantified in two groups of 12 Yorkshire pigs (30-35 kg), where 111indium labeled autologous platelets (INPLT: 850-1,200 microCi) were injected intravenously before and after CPB (BCPB, ACPB), and the platelet emboli level in intact organs and their samples (brain, heart, kidneys, lung, liver, and spleen) was quantified with an ion chamber and a gamma counter, respectively. All pigs were systemically heparinized (ACT > 400 sec). CPB was carried out at 2.5-3.5 L/min at 28 degrees C using a centrifugal pump, an oxygenator (OX:Bentley Univox 1.8 m2), an arterial filter (AF:0.25 m2), and a cardiotomy reservoir (CR: BMR 250) for 90 min. Heparin was reversed with an equivalent dose of protamine. The percent of INPLT dose (ID%, mean +/- SD) in organs of BCPB and ACPB pigs was calculated. The sequence of platelet emboli on a unit weight basis (ID%/g) had the following order: Spleen > Liver > Lung > Kidneys > Heart > Brain. The presence of significantly higher levels of emboli in brain, heart, and kidneys in the ACPB than the BCPB group suggest that platelet transfusion after heparin reversal with protamine may increase the risk of platelet emboli. However, it is an acceptable risk for patients having bleeding complications post-CPB.

In vitro and in vivo studies of PEO-grafted blood-contacting cardiovascular prostheses.

The initial step of thrombus formation on blood-contacting biomaterials is known to be adsorption of blood proteins followed by platelet adhesion. Poly(ethylene oxide) (PEO) has been frequently used to modify biomaterial surfaces to minimize or prevent protein adsorption and cell adhesion. PEO was grafted onto a number of biomaterials in our laboratory. Nitinol stents and glass tubes were grafted with PEO by priming the metal surface with trichlorovinylsilane (TCVS) followed by adsorption of Pluronic and y-irradiation. Nitinol stents were also coated with Carbothane for PEO grafting. Chemically inert polymeric biomaterials, such as Carbothane, polyethylene, silicone rubber, and expanded polytetrafluoroethylene (e-PTFE), were first adsorbed with PEO-polybutadiene-PEO (PEO-PB-PEO) triblock copolymers and then exposed to gamma-irradiation for covalent grafting. For PEO grafting to Dacron (polyethylene terephthalate), the surface was sequentially treated with PEO-PB-PEO and Pluronics followed by gamma-irradiation. In vitro studies showed substantial reduction in fibrinogen adsorption and platelet adhesion to the PEO-grafted surfaces compared with control surfaces. Fibrinogen adsorption was reduced by 70-95% by PEO grafting on all surfaces, except for e-PTFE. The platelet adhesion corresponded to the fibrinogen adsorption. When the PEO-grafted surfaces were tested ex vivo/in vivo, however, the expected beneficial effect of PEO grafting was inconsistent. The beneficial effect of the PEO grafting was most pronounced on the PEO-grafted nitinol stents. Thrombus formation was reduced by more than 85% by PEO grafting on metallic stents. Only moderate improvement (i.e. 35% decrease in platelet deposition) was observed with PEO-grafted tubes of polyethylene, silicone rubber, and glass. For PEO-grafted heart valves made of Dacron, however, no effect of PEO grafting was observed at all. It appears that the extent of thrombus formation on PEO-grafted biomaterials was directly related to the extent of tissue damage during implantation surgery. Platelets can be activated and form aggregates in the bulk blood, and the formed platelet aggregates may be able to deposit on the PEO monolayer overcoming its repulsive property. Our studies indicate that the testing of in vitro platelet adhesion should include adhesion of large platelet aggregates, in addition to adhesion of individual platelets. Furthermore, the surface modification methods should be improved over the current monolayer grafting concept so that the repulsive force by the grafted PEO layers is large enough to prevent adhesion of platelet aggregates formed in the bulk blood before arriving at the biomaterial surface.

Thrombogenicity of polyethylene oxide-bonded Dacron sewing ring in a mechanical heart valve.

BACKGROUND AND AIM OF THE STUDY: The aim of the study was to evaluate the effect of binding hydrophilic polyethylene oxide (PEO) onto Dacron fibers in the sewing ring of a mechanical heart valve (MHV), in terms of thrombogenicity of the prosthesis. METHODS: The study was performed in blinded fashion. Six Yorkshire-cross pigs (bodyweight 35-45 kg) were implanted with MHVs, in the mitral annulus, with the PEO-treated sewing ring. An additional five pigs implanted with identical MHVs, but with untreated sewing rings, served as controls. PEO of chain-length 10,000 Da was grafted to Dacron fibers using gamma irradiation. PEO-bonded Dacron fibers (diameter 100 microns) were used to weave the sewing ring, which was then assembled on a titanium stent (OD 25 mm). Autologous platelets were labeled with 111In-tropolone and injected intravenously (850-1250 microCi per injection) into the pigs on removal from cardiopulmonary bypass (CPB). At 20-24 h after surgery, platelet thrombi adherent to MHV components, and shed emboli trapped in the brain, lung, heart, kidneys and other organs/connective tissues were imaged using a gamma camera. The animals were killed and the amounts of thrombi adherent to MHV components and organ-trapped emboli quantified using an ionization chamber and gamma counter. RESULTS: There was no statistically significant difference in the adhesion of 111In-labeled platelets to either control sewing rings (0.08 +/- 0.06% dose) or PEO-treated rings (0.19 +/- 0.21% dose). The thrombogenicity of MHV components in both animal groups was in the ascending order: Dacron ring > Teflon pledgets > polypropylene sutures > titanium housing > pyrolytic carbon. The number of platelet-emboli trapped in the organs was not significantly different between the two groups. CONCLUSIONS: Simple modifications may not reduce platelet thrombosis or wound-healing of the sewing ring in the acute phase, at which time several complex processes are activating and inactivating platelets and coagulant factors during CPB and implantation of MHVs.

Quantification of device adherent, circulating, and organ pool of thrombin and fibrinogen after cardiopulmonary bypass in a pig model.

The pool of thrombin and fibrinogen in circulation, in organs, and on cardiopulmonary bypass devices was quantified during and after cardiopulmonary bypass in four groups of 24 Yorkshire pigs (weight, 30-35 kg); two groups of 10 unoperated pigs were used as controls. Thrombin-alpha and fibrinogen were iodinated with 125iodide using an iodogen transfer technique; 250-300 microCi of these tracers were injected intravenously 1 hr before cardiopulmonary bypass. All pigs were systematically heparinized (activated clotting time > 400 sec); cardiopulmonary bypass was performed at 2.5-3.5 L/min at 28 degrees C using a centrifugal pump, oxygenator (Bentley Univox 1.8 m2; Bentley Inc., Irvine, CA), arterial filter (0.25 m2), and cardiotomy reservoir (BMR 3500) for 90 min, followed by a 90 min reperfusion and 180 min of cardiopulmonary bypass. Iodinated thrombin-alpha and fibrinogen in intact organs and samples of blood, organs, tissues, and oxygenator-arterial filter-cardiotomy reservoir were quantified with an ion chamber and a gamma counter, respectively. The percent of injected iodinated thrombin-alpha and fibrinogen dose (mean +/- SD) in organs and cardiopulmonary bypass devices of all groups of cardiopulmonary bypass pigs was calculated. Thrombin generated at the small area of surgical wounds (0.016-0.038 m2), and fibrin deposited on surfaces of cardiopulmonary bypass devices (2.59 m2), initiate and propagate thrombus formation and embolization. The protein level reached saturation values on all cardiopulmonary bypass devices at 180 min. High levels of thrombin and fibrinogen-fibrin circulate in blood and organs, and are adsorbed on cardiopulmonary bypass devices; this large blood pool of pro-coagulants in the cardiac cradle, tissues, and perfused organs may account for thrombi and emboli during and after cardiopulmonary bypass.

Quantitation of adsorption and conjugation of plasma proteins by residual glutaraldehyde in fixed collagenous tissue with radioiodinated plasma proteins.

Residual glutaraldehyde (GA) in collagenous cardiovascular tissue prostheses after multiple saline rinses remains in the prostheses and accounts for adsorption and conjugation of a variety of plasma proteins. This may account for later beneficial or adverse effects. Human serum albumin (SA), gamma globulin (GG), and fibrinogen (FB) were iodinated with 125I using the iodogen-transfer technique. Bovine pericardium (PC) was fixed with 0.5% GA for 24 hr and rinsed to remove excess GA. Fresh and GA-fixed PC (FRPC, GAPC: 1 x 1 cm2), in triplicate, were incubated with 0.5-1.0 microCi of tracers in human, porcine, or bovine blood (2 ml) for a period of 0.5, 1, 2, and 3 hr and washed (5x) with saline. Maximum adsorbed proteins per unit weight of collagen (pmol/mg of PC, mean +/- SD) at 3 hr on FRPC and GAPC were quantified with a gamma counter. Fixed PC absorbed significantly more plasma proteins from blood than fresh PC. These conjugated plasma proteins are tightly bound to fixed PC. The adsorbed and conjugated plasma proteins for GAPC and FRPC have the same sequence: SA > GG > FB vs SA > GG > FB. Protein conjugation may affect the remodeling of collagenous cardiovascular tissue prostheses post implantation.

Posttraumatic cerebral ischemia after fluid percussion brain injury: an autoradiographic and histopathological study in rats.

OBJECTIVES: Mild-to-moderate reductions in local cerebral blood flow (ICBF) have been reported to occur in rats after moderate (1.7-2.2 atm) fluid percussion brain injury. The purpose of this study was to determine whether evidence for severe ischemia (i.e., mean ICBF < 0.25 ml/g/min) could be demonstrated after severe brain injury. In addition, patterns of indium-labeled platelet accumulation and histopathological outcome were correlated with the hemodynamic alterations. METHODS: Sprague-Dawley rats (n = 23), anesthetized with halothane and maintained on a 70:30 mixture of nitrous oxide:oxygen and 0.5% halothane, underwent normothermic (37 degrees C) parasagittal fluid percussion brain injury (2.4-2.6 atm). Indium-111-tropolone-labeled platelets were injected 30 minutes before traumatic brain injury (TBI), while 14C-iodoantipyrine was infused 30 minutes after trauma for ICBF determination. Sham-operated animals (n = 8) underwent similar surgical procedures but were not injured. For histopathological analysis, traumatized rats (n = 5) were perfusion-fixed 3 days after TBI. RESULTS: In autoradiographic images of indium-labeled platelets, abnormal platelet accumulation that was most pronounced overlying the pial surface was commonly associated with severe reductions in ICBF within underlying cortical regions 30 minutes after TBI. For example, within the lateral parietal cortex, ICBF was significantly reduced from 1.67 +/- 0.11 ml/g per minute (mean +/- standard error of the mean) in sham-operated animals to 0.23 +/- 0.03 ml/g per minute within the traumatized group. In addition to focal severe ischemia, moderate reductions in ICBF were detected throughout the traumatized hemisphere, including the frontal and occipital cortices, hippocampus, thalamus, and striatum. Mild decreases in ICBF were also observed throughout the contralateral cerebral cortex. At 3 days after severe TBI, histopathology demonstrated intracerebral and subarachnoid hemorrhage associated with cerebral contusion and selective neuronal necrosis. CONCLUSION: These data indicate that multiple cerebrovascular abnormalities, including subarachnoid hemorrhage, focal platelet accumulation, and severe ischemia, are important early events in the pathogenesis of cortical contusion formation after TBI. Injury severity is expected to be a critical factor in determining what therapeutic strategies are attempted in the clinical setting.

Tumor necrosis factor-alpha in plasma during cardiopulmonary bypass in a pig model: correlation with marginated neutrophils and cerebral edema by magnetic resonance imaging.

Tumor necrosis factor (TNFalpha) initiates the cytokine cascade via the nuclear factor (NFkappaB), increasing vascular permeability, inflammation, and edema during and after cardiopulmonary bypass (CPB). Cerebral edema before and after thoracotomy and CPB was evaluated by magnetic resonance imaging. TNF-alpha in plasma was measured in 12 Yorkshire pigs with an enzyme linked immunosorbent assay technique using a monoclonal antibody made against porcine TNFalpha. Blood samples were taken 30 min before and 1, 30, 60, 90, 120, 150, and 180 min after initiation of CPB. Plasma was separated from packed cells after centrifugation, and frozen at -80 degrees C. The level of TNFalpha, measured by color intensity, was read with a 96 well microtiter plate reader. Normal values in plasma were low (range, 0-17 pg/ml; mean, 4.7 pg/ml). In sham control pigs undergoing sternotomy, and pigs that had undergone CBP, TNFalpha reached a peak value at 120-150 min, and subsequently declined. TNFalpha in plasma increased in pigs that underwent thoracotomy and CPB. The increase was significantly higher in pigs that underwent CPB. Neutrophil mapping in the pig brain with 111In-labeled autologous neutrophils 180 min after CPB, suggested almost uniform distribution and accounted for the 0.02% of total circulating neutrophils in the brain, corresponding to a total of 8 x 10(5) neutrophils. Magnetic resonance imaging of the pig brain indicated the presence of cerebral edema after CPB, as depicted by the loss of structural details of the sulci, gyri, and ventricles. Activated neutrophils, monocytes, and other inflammatory cells may induce multi-organ edema and injury via TNFalpha and other regional cytokines.

The effect of nitric oxide synthase inhibition on acute platelet accumulation and hemodynamic depression in a rat model of thromboembolic stroke.

The relative importance of hemodynamic factors in the pathogenesis of thrombotic or embolic stroke is unclear. Of particular therapeutic interest are those substances that facilitate vasodilation and the clearance of platelet aggregates in the compromised microvasculature. A likely contributor to these functions is nitric oxide because it is known to inhibit platelet aggregability and promote vascular relaxation. To investigate the involvement of nitric oxide in the hemodynamic changes after experimental ischemia, photochemically induced nonocclusive common carotid artery thrombosis (CCAT) was studied. CCAT is a rat model of unilateral carotid artery stenosis and platelet embolization to the brain. This study characterized the acute hemodynamic consequences of CCAT and the resultant pattern of platelet deposits with and without nitric oxide synthase inhibition by nitro-L-arginine methyl ester (L-NAME). In addition, the subacute local cerebral blood flow changes were studied at 24 hours. Right CCAT was produced in 30 male Wistar rats injected with (111)In-labeled platelets. Between 5 and 15 minutes after thrombosis, rats were treated with either 15 mg/kg of L-NAME (intravenously) or saline vehicle. Hemodynamic changes were studied 30 to 45 minutes after thrombosis using [14C]iodoantipyrine autoradiography. Eight coronal levels were analyzed, and cortical and subcortical regions of interest were defined. Significant increases were observed in total platelets in the ipsilateral hemisphere after L-NAME treatment, and in the distribution of platelets in the anterior frontal and occipital cortices with nitric oxide synthase inhibition, encompassing the anterior and posterior border zone areas of the ipsilateral cortex. Otherwise, foci of labeled platelets were detected throughout the ipsilateral and contralateral hemispheres. Mean local cerebral blood flow images (n = 5) revealed a moderate bilateral global reduction in flow acutely, which normalized in the untreated thrombosed group by 24 hours. In contrast, the L-NAME-treated groups (sham and experimental) had lasting, widespread reductions in flow of approximately 25%. Pairwise comparisons between groups showed that CCAT/L-NAME was significantly different from shams in the corpus callosum and different from L-NAME shams in the internal capsule (P < 0.05) These hemodynamic and platelet accumulation changes may partially account for the aggravation of cognitive and sensorimotor deficits previously reported in this model of thromboembolic stroke.

Successful prosthetic mitral valve implantation in pigs.

Clotting mechanisms, the coagulation cascade, platelet function, and platelet-leukocyte-endothelial cell interactions are all very similar in humans and pigs. Because of these similarities, the authors concluded that the pig would be an ideal model for the study of thromboembolism resulting from prosthetic heart valves. To date, they have successfully recovered a total of 11 pigs (52.9 +/- 8.1 kg), 3 with bioprosthetic valves and 8 with mechanical valves, all in the mitral position (25 mm od). The normal presence of high numbers of pulmonary endothelial macrophages and other unique aspects of porcine cardiovascular and pulmonary function dictate somewhat different surgical protocols than those normally used for human patients and ruminant species. Some of these special procedures include 1) crystalloid prime without the use of plasma volume expanders, especially those with a starch base; 2) pharmacologic protection against arrhythmias (lidocaine, 4 mg/kg); 3) special attention to adequate hypothermic cardioprotection during the time of cross-clamp; 4) the use of shock doses of corticosteroid (prednisolone sodium succinate, 0.5 mg/kg) before removal of the aortic cross-clamp; and 5) positive inotropic support (dopamine, 0.008 mg/kg) while weaning from cardiopulmonary bypass. Gamma camera images of 111In tagged autologous platelets 24 hours after surgery show most thrombi located on the sewing ring with fewer on the pledgets and anchor sutures. The latter observations were confirmed by quantification of platelet deposition using a gamma counter.

A new method for quantitation of platelet microthrombi and microemboli from cardiopulmonary bypass in organs using 111In labeled platelets.

During cardiopulmonary bypass (CPB), showers of microemboli (ME) distribute among the organs and connective tissues according to regional blood flow. Post CPB, ME were quantified by subtracting residual platelets (RP) in the organs of a group of unoperated control Yorkshire pigs (n = 6) from those of operated pigs. The RP level was minimized by heparinization (300 IU/kg) before death and exsanguination. The number of adherent microthrombi (MT) and ME from the oxygenator (OX), arterial filter (AF), and thoracotomy site were determined using 111In labeled autologous platelets (INPLT) (525-585 microCi administered 24 hr before CPB) in two CPB groups (ACT > 400 sec) of 12 pigs (30-35 kg). CPB was carried out at a flow of 2.5-3.5 L/min at 28 degrees C with a roller or a centrifugal pump, OX (Bentley Univox 1.8 m2), AF (0.25 m2), and cardiotomy reservoir (CR) (Bentley BR: 3,500), for 90 (n = 6) and 180 (CPB 180, n = 6) min. Six pigs underwent thoracotomy without CPB. L-Arginine was infused at a dose of 2 mg/ kg/min during CPB (n = 6). Flow cytometry was used to estimate the circulating ME in blood. MT and organ trapped ME were imaged with a gamma camera and measured with an ion chamber and a gamma counter. ME values (percent of injected INPLT dose) in six organs and four connective tissues were calculated for all five groups. INPLT distribution indicated a uniform distribution of low level platelet MT in the CR and AF. Circulating ME amounted to 2.5% of total platelets. In the CPB circuit, ME generation in AF was the rate-limiting step (n = 4 x 10(5)). Similar studies in organs and tissues suggested the presence of a uniform distribution of the total events of ME (n = 500 x 10(6)). ME increase in brain, lung, liver, and skeletal muscle following thoracotomy and CPB was significant. The low level of ME in ischemia sensitive organs also indicated the presence of a thrombolytic threshold for cumulative ME. ME disaggregation was activated at an early stage to prevent ischemic damage, specifically in the brain. Measurement of trapped ME provided a novel, reliable, and one step method of evaluation of thrombogenicity of a CPB device and drugs.

Fluid mechanics of arterial stenosis: relationship to the development of mural thrombus.

In this study, we analyzed blood flow through a model stenosis with Reynolds numbers ranging from 300 to 3,600 using both experimental and numerical methods. The jet produced at the throat was turbulent, leading to an axisymmetric region of slowly recirculating flow. For higher Reynolds numbers, this region became more disturbed and its length was reduced. The numerical predictions were confirmed by digital particle image velocimetry and used to describe the fluid dynamics mechanisms relevant to prior measurements of platelet deposition in canine blood flow (R.T. Schoephoerster et al., Atherosclerosis and Thrombosis 12:1806-1813, 1993). Actual deposition onto the wall was dependent on the wall shear stress distribution along the stenosis, increasing in areas of flow recirculation and reattachment. Platelet activation potential was analyzed under laminar and turbulent flow conditions in terms of the cumulative effect of the varying shear and elongational stresses, and the duration platelets are exposed to them along individual platelet paths. The cumulative product of shear rate and exposure time along a platelet path reached a value of 500, half the value needed for platelet activation under constant shear (J.M.. Ramstack et al., Journal of Biomechanics 12: 113-125, 1979).

Emboli from an extraluminal blood flow hollow fiber oxygenator with and without an arterial filter during cardiopulmonary bypass in a pig model.

The effect of an arterial filter on visceral emboli was quantified with autologous indium-111 labeled platelets (INPLT) during cardiopulmonary bypass (CPB) in Yorkshire pigs. Biodistribution of INPLT was determined in 12 control pigs (30-35 kg, unoperated control [n = 6] and sham operated control [n = 6]). CPB was carried out with (n = 6) and without (n = 6) an arterial filter in 12 pigs at a flow rate of 2.5-3.5 L/min. Platelets labeled with In-111 tropolone (650-780 microCi) were injected intravenously 24 hr before CPB. All pigs were systemically heparinized (activated coagulation time > 400 sec); CPB was instituted with a roller pump, an extraluminal blood flow oxygenator (Bentley Univox, 1.8 m2), and an arterial filter (0.25 m2) and continued for 3 hr. Platelet kinetics, pooling, and counts were monitored by a Geiger probe and a Coulter counter. The thrombi in the oxygenator and arterial filter and emboli in viscera and brain were imaged with a gamma camera and measured with an ion chamber and gamma counter. Percentage of INPLT (mean +/- SD) in organs, tissues, and components of the circuit in four groups of pigs was calculated. Flow cytometry with antibodies to CD61 (GPIIIa) and CD62P (GMP-140: control) of porcine platelets was carried out with blood samples taken before, during, and after CPB for estimation of circulating platelet aggregates and platelet microparticles. Pulmonary, renal, cardiac, and cerebral emboli in pigs undergoing CPB with and without a filter were similar (p < 0.1). The amount of filter adherent thrombi was small (0.04 +/- 0.01%); oxygenator adherent thrombus in both groups was similar (p < 0.1). Emboli were found in the cerebral medulla, hippocampus, and posterior cerebral cortex in both groups. During CPB, the arterial filter functioned minimally as a trap for platelet thrombi detached from the oxygenator and circulating emboli. Flow cytometry of blood demonstrated the shift of equilibria from single platelets to platelet aggregates and microparticles during CPB and their gradual reversal to single platelets after CPB; the loosely adherent emboli disaggregated and further shifted these equilibria to single platelets and smaller aggregates, probably through the action of endogenous nitric oxide and prostacyclin. The emboli were trapped in organs and tissues and microparticles were sequestered by the reticuloendothelial system.

Reduction of neutrophil margination by L-arginine during hypothermic cardiopulmonary bypass in a pig model.

Nitric oxide generation by L-arginine (2 mg/kg/min) infusion during cardiopulmonary bypass (CPB) increases blood flow to all organs and reduces cytokine induced organ damage by reducing the level of marginating neutrophils (Ns). The N-trapping in the oxygenator (OX), arterial filter (AF), cardiotomy reservoir (CR), and N-margination were quantified with indium 111 labeled autologous neutrophils (INN) in nine groups of 40 Yorkshire pigs (30-35 kg). Cardiopulmonary bypass (180 min or 90 min CPB, 90 min reperfusion) was carried out at 2.5-3.5 L/min and at two temperatures (18 degrees C, 28 degrees C). The INN (650-780 microCi) was administered intravenously 15 mins before CPB. All pigs received heparin systemically (activated coagulation time > 400 secs); CPB was instituted with a roller pump, OX (Univox 1.8 m2), AF (0.25 m2), and CR (BCR-3500, Bentley Lab, Irvine, CA). The INN distribution in the device (OX, AF, CR) and organs was imaged with a gamma camera and measured with an ion chamber and a gamma counter. The LA infusion decreased N-trapping, estimated as the percent of injected INN (mean +/- standard deviation), in OX from control (2.7 +/- 2.02)% to (0.94 +/- 0.29)%, and margination in lung from control (48 +/- 4)% to minimal levels (23 +/- 2)% (p < 0.01). In the CPB reperfusion group, a beneficial effect was observed at LA low dose and toxicity of higher N-margination at 15 mg/ kg/min. Neither CPB temperature nor Leumedin affected N-margination significantly.

Steady flow in an aneurysm model: correlation between fluid dynamics and blood platelet deposition.

Laminar and turbulent numerical simulations of steady flow in an aneurysm model were carried out over Reynolds numbers ranging from 300 to 3600. The numerical simulations are validated with Digital particle Image Velocimetry (DPIV) measurements, and used to study the fluid dynamic mechanisms that characterize aneurysm deterioration, by correlating them to in vitro blood platelet deposition results. It is shown that the recirculation zone formed inside the aneurysm cavity creates conditions that promote thrombus formation and the viability of rupture. Wall shear stress values in the recirculation zone are around one order of magnitude less than in the entrance zone. The point of reattachment at the distal end of the aneurysm is characterized by a pronounced wall shear stress peak. As the Reynolds number increases in laminar flow, the center of the recirculation region migrates toward the distal end of the aneurysm, increasing the pressure at the reattachment point. Under fully turbulent flow conditions (Re = 3600) the recirculation zone inside the aneurysm shrinks considerably. The wall shear stress values are almost one order of magnitude larger than those for the laminar cases. The fluid dynamics mechanisms inferred from the numerical simulation were correlated with measurements of blood platelet deposition, offering useful explanations for the different morphologies of the platelet deposition curves.

Widespread hemodynamic depression and focal platelet accumulation after fluid percussion brain injury: a double-label autoradiographic study in rats.

Cerebrovascular damage leading to subsequent reductions in local cerebral blood flow (lCBF) may represent an important secondary injury mechanism following traumatic brain injury (TBI). We determined whether patterns of 111-indium-labeled platelet accumulation were spatially related to alterations in lCBF determined autoradiographically 30 min after TBI. Sprague-Dawley rats (n = 8), anesthetized with halothane and maintained on a 70:30 (vol/vol) mixture of nitrous oxide/oxygen and 0.5% halothane, underwent parasagittal fluid percussion brain injury (1.7-2.2 atm). 111-Indium-tropolone-labeled platelets were injected 30 min prior to TBI while [14C]-iodoantipyrine was infused 30 min after trauma. Sham-operated animals (n = 7) underwent similar surgical procedures but were not injured. In autoradiographic images of the indium-labeled platelets, focal sites of platelet accumulation within the traumatized hemisphere were restricted to the pial surface (five of eight rats), the external capsule underlying the lateral parietal cortex (five of eight rats), and within cerebrospinal fluid (CSF) compartments (six of eight rats). In contrast, mild-to-moderate reductions in lCBF, not restricted to sites of platelet accumulation, were seen throughout the traumatized hemisphere. Flow reductions were most severe in coronal sections underlying the impact site. For example, within the lateral parietal cortex and hippocampus, lCBF was significantly reduced [p <0.01; analysis of variance (ANOVA)] from 1.71 +/- 0.34 (mean +/- SD) and 0.78 +/- 0.12 ml/g/min, respectively, versus 0.72 +/- 0.17 and 0.41 +/- 0.06 ml/g/min within the traumatized hemisphere. Significant flow reductions were also seen in remote cortical and subcortical areas, including the right frontal cortex and striatum. These results indicate that focal platelet accumulation and widespread hemodynamic depression are both early consequences of TBI. Therapeutic strategies directed at these early microvascular consequences of TBI may be neuroprotective by attenuating secondary ischemic processes.

Acadesine reduces indium-labeled platelet deposition after photothrombosis of the common carotid artery in rats.

BACKGROUND AND PURPOSE: The adenosine-regulating agent acadesine has been shown to reduce the incidence of myocardial infarction and stroke after cardiopulmonary bypass surgery. The present study examined the effect of acadesine on the accumulation of indium-labeled platelet emboli and infarct size after photothrombosis of the common carotid artery. METHODS: Rats were anesthetized with halothane and preloaded with 111In-tropolone-labeled platelets (50 to 80 microCi) 30 minutes before nonocclusive common carotid artery thrombosis induced by a rose bengal-mediated photochemical insult. Intravenous infusion of acadesine (0.5, 1, or 2 mg/kg per minute) or vehicle was begun 30 minutes before right common carotid artery thrombosis and continued for an additional 15 minutes. Rats were then killed and brains processed for the autoradiographic quantitation of labeled platelet aggregates. In a separate group of rats, infarct areas and volumes were determined in treated (acadesine 1 mg/kg per minute) (n = 9) and nontreated (n = 9) rats 7 days after thrombosis. RESULTS: Although the ratio of right-to-left common carotid artery radioactivity was not affected by treatment, acadesine at 1 and 2 mg/kg per minute significantly decreased (P < .01) platelet deposition within the right cerebral cortex, hippocampus, and striatum. For example, within the frontoparietal cortex, numbers of platelet aggregates were 11.8 +/- 1.8 (mean +/- SEM), 6.1 +/- 1.4, 2.3 +/- 0.6, and 3.2 +/- 0.8 in rats infused with vehicle, 0.5, 1, and 2 mg/kg per minute acadesine, respectively. In addition, infarct volume was reduced by 48% in acadesine-treated (1 mg/kg per minute) rats, with a significant reduction in infarct area at the coronal level 3.7 mm anterior to bregma (P < .01). CONCLUSIONS: These results support a prophylactic role for acadesine in reducing the accumulation of platelet emboli during vascular thrombosis and subsequent brain infarction. Acadesine treatment in patients at risk for embolic stroke could potentially lead to cerebral protection.

Neutrophil dynamics and retention in lung, oxygenator, and arterial filter during cardiopulmonary bypass in a pig model.

Interactions of neutrophils with adsorbed proteins in components of the cardiopulmonary bypass (CPB) circuit and expression of leukocyte adhesion molecules on activated neutrophils affect neutrophil kinetics and margination. Lung and skeletal muscle along with oxygenator (OX) and arterial filter (AF) in the extracorporeal circuit provide the major areas of neutrophil (N) interaction. The dynamics of N-interaction and N-retention during 3 hr CPB was quantified with autologous In-111 labeled neutrophils (INN) in 4 groups of 20 Yorkshire pigs (28-35 kg, 5 sham; 5 CPB, 1 hr; 5 CPB, 3 hr and 5 CPB with heparinized circuit, 3 hr); anesthetized pigs were injected with INN (500-650 microCi), 30 min before CPB and heparinized, and underwent CPB with a roller pump, a hollow fiber OX (Bentley CM 50, 5.0 m2) and AF (Bentley AF 025, 0.25 m2) at 2.5-3.6 l/min for 3 hr. N-dynamics on OX and AF was monitored by a calibrated Geiger probe. Neutrophil deposition, like that of plasma proteins on OX, reached a steady state almost instantly, but increased on filter with CPB time. INN distribution was viewed with a gamma camera; total INN was measured with an ion chamber and INN in samples of fibers and tissues was quantified with a gamma counter. INN in lung did not change significantly during CPB and increased in liver. The percentage of injected INN in lung, liver, and brain changed with CPB time and showed significant increase over sham-operated animals. Heparin coating of components decreased INN retention. INN/meter2 of lung, OX, and AF at 3 hr were 0.26 +/- 0.07%, 0.06 +/- 0.02%, and 6.17 +/- 3.94%, and significantly lower on a heparin coated filter (2.14 +/- 1.30%). Capillary surface areas of viscera and connective tissues (lung, 100; liver, 134; spleen, 20; heart, 7; skeletal muscle, 92; fat, 12; bone, 3; bone marrow, 5; brain, 0.1 meter2) were estimated from distribution of activated INN in pigs. Lung INN retention was much higher than that of the polymer surfaces of OX/AF, indicating the role of cell adhesion molecules on INN retention on endothelial cells of lung and viscera. By direct continuous monitoring and quantitation of INN at the end of CPB, a sensitive technique for quantitation of neutrophil kinetics, margination, and retention during CPB was developed.