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.

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 Platelet Thrombi and Emboli by L-Arginine during Cardiopulmonary Bypass in a Pig Model.

We wanted to test the hypothesis that NO generation by L-arginine (LA) infusion will be beneficial in increasing blood flow to all organs to counteract the process of global ischemia during cardiopulmonary bypass (CPB) and to reduce platelet emboli by platelet inhibition. The effect of LA infusion on NO formation, vasodilation, and reduction of thromboembolic burden in organs and tissues after CPB was quantified with In-111-labeled autologous platelets in two major groups: 180 minutes CPB (CPB) and 90 minutes CPB plus 90 minutes reperfusion (RP). Platelets labeled with In-111 tropolone (650-780 µCi) were administered 24 hours before CPB and LA infusion (bolus, 10 mg/kg and infusion at 2 mg/kg/min, 21 pigs for 180 minutes CPB) in 8 groups of 30 Yorkshire pigs (30-35 kg, 6 pigs; LA 2 mg/kg/min, 3 pigs; sham-thoracotomy control, 6 pigs; unoperated control, 6 pigs). Two groups of 9 pigs (control CPB, 6 pigs; LA 2 mg/kg/min, 3 pigs) underwent 90 minutes of CPB and 90 minutes of reperfusion. All pigs were heparinized (ACT > 400 seconds); CPB was instituted with a roller pump, an oxygenator (OX: Bentley Univox, 1.8 m2), and an arterial filter (AF: 0.25 m2, Bentley) at a blood flow of 2.5-3.5 l/min. Radioactive thrombi in OX and AF and emboli in viscera, brain, and connective tissues were imaged with a gamma camera and were finally measured with an ion chamber and a gamma counter. The percent of injected platelets (mean +/- SD) in the organs and tissues of all pigs was calculated. Cerebral emboli were mapped in 25 regions of both hemispheres of pig brain. 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. Coronary bypass with LA infusion decreased the amount of adherent thrombi in OX and AF (p < 0.07). The embolic burden in brain and lung also decreased. Regional cerebral mapping of In-111 platelets showed reduced emboli in almost all regions, including the medulla, hip pocampus, and posterior cerebral cortex in both LA-treated groups. Flow cytometry of blood samples demonstrated the shift of equilibria from single platelet to platelet-aggregate-microparticle during CPB and steady-state level after the first 5-10 minutes of initiation of CPB. The L-arginine infusion reduced thrombi and emboli during CPB in the pig model.

Effect of Thoracotomy and Cardiopulmonary Bypass on Activated Platelet and Neutrophil Dynamics and Platelet Emboli in a Pig Model.

The effects of thoracotomy and components of extracorporeal circuits on dynamics of platelets and neutrophils were quantified with autologous In-111-labeled platelets (INPLT) and neutrophils (INN) during cardiopulmonary bypass (CPB) operations in Yorkshire pigs. Cardiopulmonary bypass was carried out with a hollow-fiber oxygenator and an arterial filter in 48 pigs (30-35 kg; 12 unoperated controls for platelets and neutrophils; 12 sham operated controls; 12 with 180 minutes of CPB with platelets and neutrophils; 12 with 90 minutes of CPB and 90 minutes of reperfusion at 2.5-3.5 one/min. Platelets and neutrophils were labeled with In-111 tropolone and were injected intravenously: platelets at 24 hours and neutrophils at 15 minutes before CPB. All pigs were systemically heparinized [activated coagulation time (ACT) > 400 seconds]; CPB was instituted with a roller pump, oxygenator (OX; Bentley Univox, 1.8 m2), and arterial filter (AF; 0.025 m2) for durations of 180 minutes and 90 minutes of bypass, followed by 90 minutes of reperfusion. The kinetics and pooling of platelets and neutrophils were monitored by a Geiger probe. The adherent thrombi and neutrophils in the OX, AF, viscera, and brain were imaged with a gamma camera and were measured with an ion chamber and a gamma counter. The percentile distribution of labeled platelets and neutrophils expressed as the mean +/- standard deviation of injected dose in eight groups was calculated and statistical analyses were performed (ANOVA and paired t-test). Sham operation alone increased platelet retention in the lung, heart, and brain significantly (p < 0.001) over that of unoperated pigs. Neutrophil margination to lung immediately after injection was high; CPB and reperfusion altered the distribution in blood, viscera, and connective tissues. During CPB, an equilibrium among single platelets, platelet thrombi, and emboli was reached in the blood, oxygenator, arterial filter, perfused organs, and tissues. After CPB, the pulmonary neutrophil retention increased significantly (p < 0.001). Reperfusion of 90 minutes following 90 minutes of CPB decreased the level of neutrophils and increased the level of platelets in the lung. Only a small amount of platelets and neutrophils was retained in the oxygenator and arterial filter. Neutrophil retention in the OX and AF was higher than that of platelets. The small amount of retained neutrophils in the heart, kidneys, and brain suggested that cytokines, rather than marginated neutrophils alone, may play a major role in inflammatory insult to these organs during and after CPB. OX thrombi increased with the time of CPB; AF thrombus in both groups was almost similar. During CPB, AF functioned minimally as a thrombus trap with a small percent of retained thrombi; reperfusion post-CPB did not change the amount. Thoracotomy alone has a significant effect on platelet and neutrophil kinetics, and on the subsequent effect of thrombus formation, embolization, and neutrophil margination in organs during the CPB procedure.

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.

Noninvasive imaging of c-myc oncogene messenger RNA with indium-111-antisense probes in a mammary tumor-bearing mouse model.

UNLABELLED: The c-myc oncogene is amplified in leukemia and solid tumors, thus making the c-myc messenger RNA (mRNA) a suitable target for following the progression of malignancy by noninvasive imaging with radiolabeled antisense pharmaceuticals or radiolabeled antisense oligodeoxynucleotide (RASON) probes. Considering the higher stability of phosphorothioate over phosphodiester, the probe stability and tumor localization was compared with both derivatives. METHODS: The 15-mer oligonucleotide sequence was synthesized, aminolinked [sense and antisense phosphodiester (O) and monothioester (S)] and coupled with diethylenetriamine pentaacetate (DTPA)-isothiocyanate and aliquots were lyophilized to make a DTPAAHON kit. The radionuclide 111In was chelated to DTPAAHON derivatives, and free 111In was separated by gel filtration. The radiolabeled antisense and sense probes were injected intravenously in mammary tumor-bearing BALB/c mice (1 x 10(6) cells, 8 days postinoculation). RESULTS: The highest uptake was observed at 2 hr with both thio and oxo derivatives of RASON probes, and small tumors could be imaged noninvasively. Tumor uptake and tumor/blood and tumor/muscle ratios for the sense probe (control) were significantly lower (p < 0.001) than those of the antisense probe. CONCLUSION: The radiolabeled antisense probe may provide a new sensitive tool for noninvasive imaging of c-myc oncogene mRNA for a variety of malignant tumors at an earlier stage.

Kinetics of hybridization of mRNA of c-myc oncogene with 111In-labeled antisense oligodeoxynucleotide probes by high-pressure liquid chromatography.

Antisense oligodeoxynucleotides (ASON) were labeled with gamma-emitting 123I, 99mTc and 111In radionuclides. The hybridization kinetics of 111In-labeled ASON probes [phosphorothioate (S) and phosphodiester (O)] with intact leukemic cells (P388) and purified mRNA was studied by gel filtration technique. The 15-mer oligodeoxynucleotide (ON) sequence was synthesized, amino linked and coupled to diethylenetriaminepentaacetic acid (DTPA)-isothiocyanate, and aliquots were lyophilized to make a kit for convenient preparation. 111In radionuclide was chelated to DTPAASON derivatives and free 111In was separated by gel filtration. The probe was incubated with P388 cells and mRNA extract of P388 cells. Hybridization kinetics was studied by measuring the free and mRNA-bound probe separated by the HPLC technique. The distribution of radioactivity associated with proteins, DNA and mRNAs was directly measured with a gamma camera and further quantified with an ionization chamber. The kinetics of direct and indirect hybridization of 111In-labeled antisense probes with mRNA and intact cells was similar.

Comparative thrombogenicity of pacemaker leads.

To evaluate the thrombogenicity of transvenous silicone and polyurethane pacemaker leads, 9 of 12 anesthetized Yorkshire pigs (27-32 kg) were implanted with silicone (n = 5) or polyurethane (n = 4) pacemaker leads via a femoral vein. The remaining three pigs served as controls. All 12 pigs were injected with autologous indium-111 labeled platelets (300-420 muCi) 24 hours before anesthesia induction. The pigs were monitored for 3 hours under a gamma camera. Radioactivity in blood and lead segments was measured with a gamma counter. Platelet deposits were denser on silicone leads (441.58 +/- 915.0 to 2.19 +/- 2.07) than on polyurethane leads (1.21 +/- 1.33 to 0.27 +/- 0.14) (P > 0.05). Denser platelet deposits were detected at the tip of all leads. Density of platelet deposits declined from tip to distal segments in silicone leads. The percentage of injected platelet radioactivity in the lungs of pigs with either silastic leads (12.9 +/- 2.3%) of polyurethane leads (10.1 +/- 2.2%) was higher than in the controls (4.6 +/- 0.5%) (P < 0.05). This difference indicates thrombus formation and embolization in the lungs early after lead implantation. Thrombogenicity of polyurethane leads may be lower than that of silicone leads.

In vitro and in vivo evaluation of the comparative thrombogenicity of cellulose acetate hemodialyzers with radiolabeled platelets.

Platelet consumption and platelet kinetics during hemodialysis were quantified in Yorkshire pigs with In-111 labeled platelets. Six anesthetized pigs (20-25 kg) were hemodialyzed at 150 ml/min for 3 hr. All pigs were injected with autologous In-111 labeled platelets (300-420 microCi) 24 hr before dialysis and were systemically heparinized (ACT > 400 sec) before cannulation. Hemodialysis was instituted with a Drake-Willock hemodialysis machine and a hollow fiber dialyzer (Cobe4, 0.6 m2). In vitro sham dialysis was carried out at 150 ml/min for 3 hr with six more dialyzers in a flow-loop with the blood reservoir maintained at 37 degrees C. In vitro thrombogenicity over-estimates (10-fold) in vivo values. In both systems, platelet deposition on dialyzers reached a steady state, suggesting a constant rate of thrombus formation and embolization in the hollow fiber system. The relative thrombus distribution after 3 hr of dialysis was similar in both systems, with adherent thrombi in the entry and exit ports and highest numbers in the midsection of the hemodialyzer. Biodistribution after 3 hr of dialysis indicated that thrombosis of the hemodialyzer and arterial and venous traps as well as embolization reduced the platelet pool in the blood and increased platelet emboli in lung, brain, kidneys, and skeletal muscle, as measured by the In-111 labeled platelets.

Effects of local geometry and fluid dynamics on regional platelet deposition on artificial surfaces.

An important aspect of blood-material interactions is the activation, adhesion, and subsequent aggregation of blood platelets on the artificial surface, all of which are directly affected by local fluid dynamics. The objective of this work was to directly correlate changing local fluid dynamic conditions produced by various vessel geometries, including stenosis, aneurysm, and separate contraction and expansion geometries, with quantitative in vitro measurements of regional platelet deposition. We directly measured platelet deposition as a function of axial position along four Lexan flow chambers with axisymmetric models of these geometries using 111In-labeled platelets. Platelet deposition was maximum in observed areas of flow recirculation and reattachment and minimum in locations of high shear and separation. For the stenosis geometry, two distinct regions of increased platelet deposition were apparent, one proximal to and one distal to the stenosis throat. An approximately linear increase in platelet densities was produced in the aneurysm region, increasing in the direction of flow. Through a comparison of platelet deposition with local fluid streamline orientation, we have shown that platelet deposition is increased in certain areas due to the enhanced convective transport of platelets and blood cells to the vessel wall along locally curved streamlines with velocity components perpendicular to the vessel wall.

A higher blood flow window of reduced thrombogenicity and acceptable fragmentation in a hollow fiber hemodialyzer.

The effect of pulsatile blood flow on platelet thrombogenicity and platelet fragmentation (PF) in a hollow fiber hemodialyzer (HFD) was quantified with 111In labeled platelets and 125I labeled fibrinogen; 150 ml of blood was collected from Beagle dogs, Yorkshire pigs, and a human volunteer (non-smoker). Platelets were labeled with 111In tropolone (300 microCi) and fibrinogen was labeled with 125I. Sham dialysis (SHD) was performed with 120 HFDs (0.9 meter2) at 37 degrees C, with flow-rates of 150, 250, 500, and 950 ml/min.; after SHD, the washed HD radioactivity was measured with an ionization chamber. PF was measured by flow cytometry with GP IIb-IIIa murine monoclonal antibody. Platelet deposition decreased significantly for 3 species at higher flow; fibrinogen deposition (10-12%, 55-65 mg/m2), was not affected by flow. Adherent platelet thrombus decreased from (8.2 +/- 3.4) to (3.1 +/- 1.0) with human blood as flow rate increased from 150 to 950 ml/min; platelet thrombus level also decreased significantly (p < 0.005) from (20.3 +/- 6.2) to (4.5 +/- 1.9) with canine blood. Higher values were obtained for canine than human and porcine platelets. Platelet fragmentation, on the other hand, increased from 2.1-2.2% to 10.2-11.3% with increase of flow. Like platelets, deposition of canine fibrinogen was slightly higher than that of pig and human. The studies of adherent thrombus and platelet fragmentation identified an important flow-window of reduced thrombogenicity and acceptable fragmentation, encouraging extracorporeal circulation at higher blood flow.

Quantitation of thrombogenicity of hemodialyzer with technetium-99m and indium-111 labeled platelets.

The platelet thrombogenicity of a hemodialyzer was quantified with 99mTc- and 111In-labeled platelets. The platelets collected from blood of Beagle dogs, Yorkshire pigs and human volunteers were labeled with 111In-tropolone (detergent-free) and 99mTc-HMPAO. Hemodialysis was performed with a hollow-fiber dialyzer (HFD) in a flow-loop, the temperature of which was maintained at 37 degrees C, with flow-rates of 7, 150 and 270 mL/min; after dialysis, the HFD radioactivity was measured with an ionization chamber and imaged with a gamma-camera. The radioactivity of samples of hollow-fibers taken from the top, middle and bottom of the dialyzer was determined with a gamma-counter. The mean values of hemodialyzer-adherent platelet radioactivity were calculated for both radionuclides. The canine platelets were found to be more thrombogenic than porcine and human platelets. The adhesivity of porcine platelets to the biomaterial (cellulose-acetate) of the dialyzer approximated that of human platelets. The 99mTc label underestimated the thrombus formation (P < 0.01). The dynamic processes of thrombosis and embolization from the hemodialyzer resulted in the large standard deviations around the mean values of the adherent thrombus. In spite of this limitation of the dynamic pathology, the quantitation of comparative thrombogenicity with 111In- and 99mTc-labeled platelets suggests that both radionuclides could be used for measurement of device-induced thrombogenicity and may provide an estimation of prosthesis-induced thrombogenicity of human platelets from animal studies.

Rate constants of embolization and quantitation of emboli from the hollow-fiber oxygenator and arterial filter during cardiopulmonary bypass.

A direct technique was developed to estimate parameters related to half life (T1/2) and rate constants of embolization (RCE) and to quantitate emboli of three sizes (small, medium, and large) shed from the oxygenator and arterial filter during cardiopulmonary bypass (CPB). Cardiopulmonary bypass was performed in 16 Yorkshire pigs divided as follows: systemic heparin group (SHG:6), systemic heparin/heparinized circuit group (SH/HCG:5), and Iloprost (Bentley) (2 ng/kg/min)/heparinized circuit group (IHCG:5) with In-111 labeled autologous platelets. The anesthetized pigs (20-25 kg) underwent CPB at 2.5-3.0 L/min for 3 hours. Pigs were injected with In-111 platelets (300-420 microCi) 24 hours before CPB. Cardiopulmonary bypass was instituted with a roller pump and hollow-fiber oxygenator, and thrombosis and embolization on the oxygenator and filter were monitored by a calibrated Geiger probe (WMB Johnson Associates, Montvale, NJ). The radioactivity in the oxygenator and filter reached peak values at 25-45 min after CPB; the radioactivity then declined in the oxygenator but remained at a steady state in the filter, suggesting continuous embolization at the same rate of trapping. The curve stripping technique of the normalized radioactivity time curve of the oxygenator was used for RCE estimation of different sized emboli shed from the oxygenator; 42% of thrombus embolized from the oxygenator in SHG with three rate constants, with T1/2s of 12 min, 42 min, and 13 hr; the SH/HCG embolized 35% with T1/2 of 78 min, and the IHCG embolized 30%, with a T1/2 of 22 min. This indicates that there is less embolization in the IHCG.(ABSTRACT TRUNCATED AT 250 WORDS)

Quantitation of comparative thrombogenicity of dog, pig, and human platelets in a hemodialyzer.

Comparative platelet thrombogenicity was quantified with In111 labeled platelets. The platelets collected from the blood of Beagle dogs, Yorkshire pigs, and human volunteers were labeled with detergent free In111 tropolone, and sham hemodialysis (SHD) was performed with a hollow fiber dialyzer in a flow loop at 37 degrees C, with flow rates of 7, 150, and 270 ml/min. After SHD, hemodialyzer radioactivity was measured with an ionization chamber, gamma counter, and it was imaged with a gamma camera. The mean values of hemodialyzer-adherent platelet radioactivity were calculated. Canine platelets are more thrombogenic than porcine and human platelets. Quantitation of comparative thrombogenicity with In111 platelets may provide an estimation of prosthesis induced thrombogenicity of human platelets from animal studies.

Platelet preservation during cardiopulmonary bypass with iloprost and Duraflo-II heparin-coated surfaces.

To test the hypothesis that temporary platelet inhibition during cardiopulmonary bypass (CPB) with surface heparinized systems may result in platelet preservation, nine Yorkshire pigs were placed on CPB for 3 hours. Platelet labeling was done in all pigs with Indium-111 tropolone. CPB was instituted with a roller pump, a hollow fiber membrane oxygenator (Bentley CM-50 [Baxter-Bentley Laboratories, Irvine, CA]), and an arterial filter. The extracorporeal perfusion systems were surface-coated with the Duraflo-II heparin complex. Group A pigs (n = 5) were systemically heparinized (activated coagulation time longer than 400 sec). Group B pigs (n = 4) were placed on CPB without systematic heparinization, but have received the stable prostacyclin-analog Iloprost (ZK36374) at 1 ng/kg/min i.v. from 30 min before and during CPB. Platelet counts declined in group A pigs at 5 min, 1 hr, 2 hr, and 3 hr of CPB to 79.8% (mean), 66.5%, 71.3%, and 69.0% of pre-CPB values, respectively (p less than 0.05). In group B pigs, mean platelet count during CPB was higher than 90% of control value. Percentage of injected radioactivity detected in the oxygenator was 2.82% in group A pigs versus 0.73% in group B pigs (p = 0.0541). Surface heparinization with the Duraflo II heparin coating complex in combination with Iloprost-induced temporary platelet inhibition resulted in platelet count preservation during CPB in the pig model.

Cardiopulmonary bypass with a surface-heparinized extracorporeal perfusion system.

To evaluate the effect of surface heparinization on platelet consumption during cardiopulmonary bypass (CPB) ten pigs were placed on CPB for 3 hours. All pigs were injected with autologous Indium-111 labeled platelets (300-420 uCi) 24 hours prior to CPB and were systemically heparinized prior to cannulation for CPB. CPB was established with a roller pump, a hollow fiber membrane oxygenator (HFMO, Bentley CM-50) and an arterial filter (AF, Bentley 1025). In six pigs the extracorporeal system was untreated whereas in four pigs it was surface heparinized with the Duraflo-II method. Cardiotomy suction was not used. Percent of injected radiation dose in HFMO and AF at 3 hours of CPB in the nontreated systems was 1.53 +/- 1.12 and 0.88 +/- 0.63%, whereas in the surface heparinized systems was 2.45 +/- 1.71 and 0.49 +/- 0.39% respectively (NS). (Values are mean +/- SD). Blood loss during (CPB) was 225 +/- 179 ml in the nontreated systems, and 263 +/- 103 ml in the surface heparinized systems (NS). Platelet counts were reduced by 12% or 21.8% at 3 hours of CPB in the two groups of pigs respectively (NS). No difference was observed in platelet consumption (in HFMO and in AF) or in platelet count reduction between the two groups of pigs. Surface heparinization did not improve platelet preservation in systemically heparinized pigs at 3 hours of CPB.

Increase of intraplatelet free calcium ion during extracorporeal circulation with a hollow fiber oxygenator: arterial filter and dynamics of platelet thrombosis on oxygenator and filter in a pig model.

Intraplatelet free calcium (IPFC) ions provide a common pathway for platelet activation leading to thrombosis and embolization. IPFC levels were determined by chlorotetracycline fluorometry during extracorporeal circulation (ECC) with systemic heparin in eight Yorkshire pigs (weighing 30-40 kg; 3 control and 5 ECC); the ratio of slow phase organelle calcium sequestration to fast phase platelet-membrane binding is an index of free calcium. During 3 hr of ECC with a hollow fiber oxygenator (HFO) (Bentley CM-50) and AF (Bentley 1025), seven blood samples were collected 5 min before and during ECC. The platelet deposition (CPM/microCi) on HFO (PDHFO) was simultaneously measured with In-111-labeled autologous platelets (300-400 microCi) and a Geiger probe detector at -5, 0, 5, 30, 45, 60, 120, and 180 min. During ECC, IPFC and HFO thrombus increase significantly (p less than 0.05) at 45 min with respect to control IPFC values of 0.4 +/- 0.1, suggesting direct participation of calcium activated platelets in thrombosis on HFO. The decline of IPFC is due to extrusion and sequestration by dense granules, and decline in HFO thrombus is due to embolization. On the other hand, the embolus in the arterial filter was trapped in a linear fashion, with a consistent increase with time of ECC.

The dynamics of platelet thrombus formation rate, thrombus retention time, and rate of embolization on a control and heparin bonded polyurethane angio-catheter.

The dynamics of platelet deposition and embolization from control and heparin bonded polyurethane catheters (CPC and HBPC) was evaluated with In-111 labeled autologous platelets (IN-PLT) and a computerized gamma camera (CGC). Ten non-heparinized dogs (18-25 kg) were catheterized in both femoral arteries with 10 cm of CPC and HBPC (5 Fr., Cordis) 24 hr after injection of 300-420 microCi of In-PLT, and imaged for 3 hr with the computerized gamma camera. The regional platelet deposition curves (RPDC) indicated multiple peaks and valleys; the curves were analyzed for early rate of thrombus formation (upswing), thrombus retention time (full width at half maxima of the RPDC-peak), and rate of embolization (downswing) on both catheters. The four parameters (mean +/- SD) of thrombosis on catheters and integral of the radioactivity time curve for the 3 hr duration of imaging were calculated from normalized counts/sec. The rate of thrombus formation and rate of embolization are higher for the control than HBPC, suggesting that heparin-bonding decreases the early rate of thrombosis and embolization. The thrombus adhesivity and retention time appear shorter for the control catheter, indicating that the control thrombogenic catheter forms multiple thrombi and emboli than HBPC. The integral appears larger for the control catheter than HBPC. In vivo (dynamic) studies, in vitro studies, and critical analyses of the radioactivity time curve were essential for complete evaluation of thrombogenicity of catheters and other cardiovascular prostheses.