Effects of membrane plasma exchange on the hemostatic system and on thyroid hormones in critically ill patients

The present study examined the effects of repeated plasma exchanges with membrane filtration (Plasmaflux P2 membrane, 4.3 I human albumin solution) on the hemostatic system and on thyroid hormones in critically ill patients. Clotting factors V, VII, VIII, IX, X, XI, XII, XIII, fibrinogen, antithrombin III (ATIII), prothrombin time (PT), activated partial thromboplastin time (PTT), total (TT3) and free tri-iodothyronine (FT3), total (TT4) and free thyroxine (FT4), and thyroxine binding globulin (TBG) were determined before, immediately after, 1, 3, 6 and 24 h after plasma exchange in 6 patients (3 with glomerulonephritis, 2 with IgG myeloma, 1 with chronic polyneuritis) during 18 plasma exchanges. After plasma exchange levels of clotting factors and ATIII were markedly lowered but except for fibrinogen and factor XIII, they were not reduced by repeated exchange procedures. Thyroid hormones and TBG were reconstituted after 24 h. Pre-exchange levels, except for TT4 and TBG, were not lowered by repeated exchange procedures. Risk of bleeding or thrombosis is small and there is no appreciable risk of loss of thyroid hormones during repeated plasma exchanges.     

Pre-instrumental variables in coagulation testing.

A number of variables thought to affect measurement of prothrombin time (PT) and partial thromboplastin time (PTT) were examined in an effort to determine more precisely their effects on these measurements. On the basis of these studies, it is proposed that blood specimens be anticoagulated with one part 3.8% (w./v.) sodium citrate solution to 19 parts whole blood to avoid excessive anticoagulation of blood samples drawn from patients with polycythemia. Because of the smaller amount of plasma in such samples, relatively larger amounts of anticoagulant are used, and spuriously prolonged PT and PTT measurements commonly result. No deleterious effect on anemic specimens is evident when the smaller amount of citrate is used. Studies of the stability of these specimens indicate that unopened, vacuum-drawn specimens do not noticeably deteriorate for as long as 6 hours, even when kept at room temperature. Prothrombin time measurements remain constant for as long as 24 hours. However, a 10--15% lengthening of the partial thromboplastin time is evident after 24 hours of storage.     

Intraindividual comparison of the impact of two selective apheresis methods (DALI and HELP) on the coagulation system

We performed an intraindividual comparison of the effect on the coagulation system of two selective apheresis procedures: Direct Adsorption of Lipoproteins (DALI) and Heparin-induced Lipoprotein Fibrinogen Precipitation (HELP). Six patients suffering from heterozygous familial hypercholesterolemia have been treated with 2 sessions of each procedure. Anticoagulation was carried out according to usual recommendations. Blood samples were taken before, immediately after and on the second day after the sessions. We assessed global coagulation tests (prothrombin time, activated partial thromboplastin time), fibrinogen, prothrombin fragment F 1 + 2 and a variety of factors (Factors II, V, VII, XIII, IX, X, XI, XII, XIIa; von Willebrand Factor; collagen-binding activity, prekallikrein, high-molecular weight kininogen) and antagonists (antithrombin III, protein S activity, free protein S). In fact, all parameters measured have been influenced by the apheresis treatment. Fibrinogen is lowered more by HELP which also has a more definite impact on factors belonging to the prothrombin complex (II, VII, X). In contrast, the major effects of the DALI system have been seen on the intrinsic pathway of the coagulation system (IX, XI, prekallikrein, high-molecular-weight kininogen). With both systems, no increases in activated Factor XII or in prothrombin fragment F 1 + 2 have been observed. These data provide a solid basis for individual adaptations of anticoagulant doses.     

Effects of PEG-PLA-nano artificial cells containing hemoglobin on kidney function and renal histology in rats

This study is to investigate the long-term effects of PEG-PLA nano artificial cells containing hemoglobin (NanoRBC) on renal function and renal histology after 1/3 blood volume top loading in rats. The experimental rats received one of the following infusions: NanoRBC in Ringer lactate, Ringer lactate, stroma-free hemoglobin (SFHB), polyhemoglobin (PolyHb), autologous rat whole blood (rat RBC). Blood samples were taken before infusions and on days 1, 7 and 21 after infusions for biochemistry analysis. Rats were sacrificed on day 21 after infusions and kidneys were excised for histology examination. Infusion of SFHB induced significant decrease in renal function damage evidenced by elevated serum urea, creatinine and uric acid throughout the 21 days. Kidney histology in SFHb infusion group revealed focal tubular necrosis and intraluminal cellular debris in the proximal tubules, whereas the glomeruli were not observed damaged. In all the other groups, NanoRBC, PolyHb, Ringer lactate and rat RBC, there were no abnormalities in renal biochemistry or histology. In conclusion, injection of NanoRBC did not have adverse effects on renal function nor renal histology.     

Response of pulmonary (circulating) megakaryocytes to experimentally induced consumption coagulopathy in rabbits.

The effects of slow temporary infusion of a tissue thromboplastin solution into the superior vena cava on pulmonary as well as circulating megakaryocytes were studied in 40 rabbits (2-48 hours after infusion) and related to 6 noninfused and 7 infused with normal saline. This is a simple and specific method of inducing a fall in blood platelet levels and thereby an activation of thrombocytopoiesis and megakaryocytopoiesis. The induced intravascular coagulation is probably counterbalanced by an activated fibrinolysis allowing the animals to survive the infusion and thereby offering the possibility of studying the long-term effects. An increase to about 300% of the normal values in circulating as well as pulmonary megakaryocytes was found 20 and 24 hours after the onset of the infusions respectively. The number of circulating and pulmonary megakaryocytes, showing great individual variations, however, dropped to normal levels within 28 hours after onset of the infusions, which means that megakaryocytes remain detectable for less than eight hours in the lungs. No increase was found in pulmonary megakaryocytes in the control (saline infused) group. In our opinion the entrance of megakaryocytes from the bone marrow into the blood circulation in an incidental event, the number in the circulation reflecting the activity of megakarycocytopoiesis. This experiment supports our suggestion that intravascular coagulation is one of the major pathophysiological mechanisms leading to an increase in pulmonary megakaryocytes.     

Intravascular circulation and distribution of human 51Cr-DBBF stroma-free hemoglobin, 51Cr-plasma, 51Cr-saline, 59FE-plasma, and 125I-albumin in the mouse

Male B6C3HF1 mice were infused with human 51Cr-labeled DBBF (bis 3,5-dibromosalicyl fumarate) crosslinked stroma-free hemoglobin (SFH). In the first hour following SFH infusion, 11.2% of the infused radioactivity was found in the skin, 11.4% in muscle, 9.1% in the skeleton, and 5% in the liver. Twenty-four hours after infusion, 15.4% of the radioactivity was found in the skin, 10.3%, in the muscle, 16.6% in the skeleton, and 6.7% in the liver. The circulation and distribution of 51Cr-labeled DBBF-SFH were compared with levels of 51Cr labeled plasma, 51Cr in saline, 59Fe labeled plasma, and 125I albumin. The radioactivity in the blood was similar for 51Cr-DBBF-SFH, 51Cr-plasma, and 59Fe-plasma. During the 24-hour post-infusion period, extravascular distribution of the 51Cr-saline, 51Cr-plasma, and 125I albumin within the organs was similar to that of 51Cr-DBBF-SFH, with the highest levels being in skin, muscle, skeleton and liver, and no increase in the levels in the lung or spleen. The distribution of 59Fe compared to that of 51Cr-DBBF, 51Cr-plasma, 51Cr-saline, and 125I albumin can be explained by the fact that 59Fe is utilized in the production of new red blood cells.     

Heparin stability: effects of diluent, heparin activity, container, and pH.

The effects have been studied of diluent, heparin activity after dilution, container, and pH on the stability of heparin solutions stored under conditions resembling those present during heparin infusion by intravenous drip or syringe pump. Heparin activity was measured by activated partial thromboplastin time and thrombin clotting time (and, in one set of studies, also by factor Xa inhibitor assay and protamine sulphate neutralisation). Heparin activity was stable for 6 hours regardless of storage conditions. After 24 hours heparin activity was stable when the drug was diluted in 0.9% saline and stored in plastic, but a small loss of activity was observed in several studies after dilution in 5% dextrose or storage in glass. A more extensive comparison confirmed a 3-5% loss in heparin activity over 24 hours after dilution in 5% dextrose. Changing the pH to 3.5 or 10.0 had little effect on storage stability. We conclude that heparin activity in vitro remains stable during short infusions but recommend dilution in 0.9% saline and a plastic container when a heparin solution is infused over 24 hours.     

The determination of INR in stored whole blood.

AIMS: To examine the reliability of international normalised ratio (INR) determination on samples stored as whole blood for up to two days at room temperature. METHODS: The INR of 40 patients receiving oral anticoagulants was determined on fresh blood and on samples stored for 24 and 48 hours, using five locally calibrated prothrombin time systems. These incorporated Manchester reagent, Recombiplastin, IL PT Fibrinogen HS Plus, Manchester combined capillary prothrombin time reagent, and a freeze dried in-house reference rabbit brain thromboplastin, RBT 1010. In addition, factors II, V, VII, and X were determined on samples obtained from 18 of these patients before and after incubation at room temperature. RESULTS: The INR of the samples changed by differing amounts during storage, depending on which system was employed. Although the mean change after 24 hours storage was relatively small, there were individual samples that changed by > 0.5 INR with all systems. These changes would lead to adjustment in dosage of certain patients. After 48 hours these effects were greater with all systems except that employing Recombiplastin. There were only small reduction in the measured factors by 48 hours. CONCLUSIONS: After storage of samples for only 24 hours, some patients' INR changed sufficiently to affect dosage. In view of these observations, the practice of storing whole blood samples for INR determination cannot be recommended.     

The effects of short term diazinon exposure on blood clotting activity in the rat

Female rats were exposed to a drinking solution containing 157 parts per million (ppm) of the organophosphate insecticide diazinon for 14 days. Plasma samples were assayed for prothrombin time (PT), partial thromboplastin time (APTT), hematocrit (HCT), platelet count (PLT) and clotting times for coagulation factors I (Fibrinogen), II, V, VII, VIII, X and XII. Significant change following ingestion of diazinon were noted for the PT, APTT, Fibrinogen, factors II, VII, VIII, X, XII and hematocrit. The data suggest that in the rat, diazinon ingestion has a direct effect on the clotting activity of various blood coagulation parameters.     

Coagulation changes in baboons during acute experimental hemoglobinemia and dextran infusion.

Evidence of disseminated intravascular coagulation (DIC) was dought in normal baboons infused with autologous hemolyzed whole blood, preceded or followed by infusion of dextran (molecular weight, 70,000). Mean peak plasma hemoglobin following a rapid single injection was 370 mg/100 ml in 2 animals and 1,236 mg/100 ml in 1 animal, while levels during continuous 5 hour infusion in 2 animals averaged 326 and 474 mg/100 ml, respectively. Dextran infusion immediately preceded hemoglobin injection in 2 baboons and followed hemoglobin injection by 1 1/2 and 2 1/2 hours, respectively, in 2 baboons. Coagulation studies showed a moderate although significant fall in platelet count with prolongation of the partial thromboplastin time following hemoglobin infusion, and shortening of the thrombin time after dextran. Fibrin degradation products developed in four of five experiments after hemolysate injection. The induction of acute experimental hemoglobinemia results, therefore, in the development of coagulation changes consistent with milk DIC. Preliminary infusion of dextran (molecular weight, 70,000) may facilitate this response by either initiating the development or impeding the clearance of fibrin degradation products.     

The effects of chronic cadmium toxicity on the hemostatic system

Cadmium, a highly toxic heavy metal, is distributed widely in the general environment. The characteristic clinical manifestations of chronic cadmium intoxication include renal proximal tubular dysfunction, osteomalacia and anemia. Accumulating evidence suggests that cadmium toxicity may also affect various organs such as the liver, lung, testis and hematopoietic system. The aim of this study was to determine the effect of chronic cadmium exposure on the anticoagulant system in rats. Fourty-five adult Wistar albino rats were randomly allocated into 2 groups. While the control group was given tap water, the animals in the cadmium group were treated with 15 ppm CdCl(2) for 4 weeks. Blood cadmium concentration, prothrombin time, activated partial thromboplastin time, plasma protein C and antithrombin activity, and platelet count were determined in the rats. Blood cadmium concentrations increased in the experiment group compared to the control group (p < 0.001). Results also show that cadmium exposure shortened prothrombin time (p < 0.05) and activated partial thromboplastin time (p < 0.01) in rats. Protein C (p < 0.001) and antithrombin (p < 0.001) decreased to statistically significantly lower levels in rat plasma after cadmium exposure when compared to the control group. When the number of thrombocytes was compared between 2 groups, a decrease was observed in the group treated with CdCl(2), which was, however, not statistically significant (p > 0.05). In conclusion, when the parameters of the hemolytic system are considered, the decrease in protein C and antithrombin activities and the shortening of prothrombin time and activated partial thromboplastin time suggests the presence of a hypercoagulable state during chronic cadmium intoxication. Therefore, it may be stated that chronic cadmium toxicity sets the stage for hypercoagulation and hence increases the risk of thrombosis.     

Abnormalities of pathways of fibrin turnover in lung lavage of rats with oleic acid and bleomycin-induced lung injury support alveolar fibrin deposition.

Alveolar fibrin deposition commonly accompanies acute lung injury, but the nature of the local abnormalities of coagulation and fibrinolysis that support pathologic fibrin deposition are not well understood. The trended abnormalities of procoagulant and fibrinolytic activities occurring in lung lavage fluids of Fischer 344 rats after lung injury induced by intravenous oleic acid (OA) or intratracheal bleomycin were studied. After injury by either agent, bronchoalveolar lavage (BAL) contained increased procoagulant activity and decreased fibrinolytic activity. Lavage procoagulant activity was mainly due to an activator of Factor X attributable to the extrinsic coagulation pathway, and fibrinolytic activity was almost completely plasminogen dependent. Major mechanisms of inhibition of fibrinolytic activity involved both the inhibition of the plasminogen activator (PA) and plasmin. These abnormalities were temporally associated with prominent alveolar fibrin deposition in both models. In OA-treated animals, lavage fibrinolytic activity was absent or profoundly decreased, and antiplasmin and procoagulant activities were increased within 4 hours after the induction of acute lung injury. By 24 hours after OA, lavage PA inhibitor (PAI) activity was elevated with sustained antiplasmin activity. By 3 days after OA, these abnormalities had resolved in association with almost complete resolution of alveolar fibrin deposits. Within 3 days after bleomycin-induced lung injury, lavage procoagulant activity was increased and fibrinolytic activity was depressed due to increased antiplasmin and PAI activities. These conditions persisted for 2 weeks, during which time alveolar fibrin deposition was associated with the development of pulmonary fibrosis. These data indicate that a disruption of the normal balance between procoagulant and fibrinolytic activities occurs in alveolar lining fluids of rats with alveolitis induced by either OA or bleomycin, and that concurrent abnormalities of pathways of fibrin turnover that occur in alveolar lining fluids promote the alveolar fibrin deposition associated with these lung injuries.     

Effects of a natural estrogen, estradiol, on hemostasis in guinea pigs

17-beta-estradiol was administered weekly to female guinea-pigs at the dosage of 2-3 mg/kg body weight during 4-6 weeks. Compared to a control population, no significant effect was observed on: recalcification time; thromboelastography; PTT; APTT; Quick one stage prothrombin time; antigenic and active plasma fibrinogen level; antithrombin III level; factor XI level; red and white blood cell count. A significant increase in levels of factor II (+25%), of factor V, of factor XII (+40%) and a decrease in levels of complex VII + X was observed. Values for AT III, factor XII, chronometric and immunologic fibrinogen plasma levels, unknown until now for the guinea-pig, were established.     

Hormone replacement therapy and hemostasis: effects in Brazilian postmenopausal women

OBJECTIVE: To evaluate the impact that administration of transdermal estradiol gel combined with medroxyprogesterone acetate (MPA) has on hemostasis. METHODS: In this open prospective longitudinal study, thirty postmenopausal women received transdermal estradiol gel (1 mg/day) continuously combined with oral MPA (5 mg/day) for 12 days/month. The following parameters were determined: prothrombin time (PT), activated partial thromboplastin time (aPTT), factors VII, X, and XII activity, fibrinogen levels, thrombin-antithrombin complex levels, protein C and S antigen, antithrombin activity, plasminogen activator inhibitor type 1 (PAI-1) antigen, tissue-type plasminogen activator (t-PA) antigen, plasminogen activity and fibrin degradation products (FbDP) antigen. They were evaluated before and after 6 months of treatment. RESULTS: There was a significant decrease in factor VII activity (P=0.001), factor X activity (P=0.016), protein C antigen (P=0.022), antithrombin activity (P=0.025), plasminogen activity (P=0.023), t-PA antigen (P=0.043) and FbDP antigen (P=0.048) compared with baseline values. CONCLUSION: The results suggest that the treatment with transdermal estradiol gel combined with MPA avoids any major activation of coagulation and does not produce any overall effect on fibrinolysis. Therefore, this treatment might provide interesting effects on hemostasis in postmenopausal Brazilian women.     

Clinically applicable laboratory end-points for treating snakebite coagulopathy

AIMS: To determine which coagulation tests best reflect the return of clotting function after snakebite venom induced consumptive coagulopathy (VICC). METHODS: Cases of snake envenoming were prospectively recruited to the Australian Snakebite Project (ASP). This study examined cases with VICC treated with antivenom and monitored with serial measures of clottable fibrinogen, prothrombin time (PT) and activated partial thromboplastin time (aPTT). The main outcome measures were times from antivenom treatment until a moderate recovery in the PT (<24 seconds), a measurable aPTT and detectable fibrinogen. RESULTS: Forty-six cases were examined, including 27 brown snakes with proven complete venom neutralisation by antivenom in 25, 16 tiger snake group and three taipans. The times from initial antivenom dose to recovery were: PT<24 seconds, median 9.2 hours (IQR 6.2-11.3 hours); measurable aPTT, median 5.2 hours (IQR 3.4-8.8 hours); and detectable fibrinogen, median 8.8 hours (IQR 5.4-11.7 hours). In 10 cases where fibrinogen was detectable earlier than recovery of the PT, the mean fibrinogen was 0.25 g/L (SD 0.10) compared with 0.6 g/L (SD 0.28) in the remaining 36 cases (p<0.0001), reflecting differing sensitivities between laboratories. In only three patients (7%) was fibrinogen measurable before the other two outcomes, using highly sensitive fibrinogen assays. CONCLUSION: The combination of the PT and aPTT is an effective, clinically available and cost-effective end-point for treating VICC, and may take longer to return to normal after venom neutralisation than previously believed. The fibrinogen assays that are generally in use do not provide any additional useful information.     

Intracerebroventricular infusions of rat satietin into rats does not produce conditioned taste aversion

In a previous study we found that while human satietin (h-SAT) suppressed the food intake of rats it was also aversive to them. In the present study rat satietin (r-SAT) was tested for aversiveness in rats fitted with chronic third ventricle (ICV) cannulas. The rats were then given access to water for 1-hr/day and food ad lib for ten days. Fluid intake, food intake during fluid access and 24-hr total food consumption were recorded. The rats were then ICV infused with saline and 30 min later half of the animals given access to banana flavored water (Group 1) while the remainder were presented with almond flavored water (Group 2). The next day Group 1 was infused with saline and Group 2 with 100 micrograms/rat of r-SAT. Thirty minutes later the flavors presented to the rats were the reverse of the previous day. Satietin significantly reduced food intake during fluid access and for 24 hours. Thereafter, fluid and food ingestion of the groups was normal and similar. Thus no rebound feeding occurred in the r-SAT treated group. Two days after r-SAT or saline the rats were given a two-bottle choice test. Both groups displaced equal preference for the flavors, therefore r-SAT produced no taste aversion. The r-SAT treated rats lost more body weight than saline treated animals the first day after treatment. This difference increased the next day and remained significant for seven days post infusion, whereas, food consumption did not differ between the groups after the first day. The data indicate the food intake suppression in rats produced by r-SAT is not due to the compound being aversive.     

Proximal tubular reabsorption during renal vasodilation and increased arterial blood pressure in saline loaded rats

Summary Rats were infused with 0.31 ml per min isotonic saline + bicarbonate intravenously. A further infusion of acetylcholine and bradykinin into the renal artery resulted in natriuresis, diuresis, an increase in PAH clearance and a higher content of hemoglobin in the kidney tissue; blood pressure and inulin clearance remained constant. After acetylcholine infusion, a shortening of transit time of Lissamine green through the proximal convolution and a narrowing of tubuli were observed without significant change in reabsorptivet _1/2 (split-oil-droplet). After bradykinin infusion, these changes were much smaller and insignificant. It seems therefore, that a common mechanism of natriuresis after kidney vasodilation does not probably exist.The changes in renal parameters of saline-bicarbonate infused rats after body overheating (natriuresis and diuresis, an increase in PAH clearance and hemoglobin content, shortening of transit time and narrowing of tubuli) were probably due to an elevation of blood pressure. If this increase was prevented by an aortic clamp just below the diaphragm, all values did not differ from those in control rats.     

Hereditary hemorrhagic telangiectasia. A family study

The authors report a comprehensive evaluation of the hemostatic system in eight related patients with hereditary hemorrhagic telangiectasia (HHT). Unlike in previous reports, they could find no evidence for abnormalities in platelet aggregation or for qualitative abnormalities of the Factor VIII complex. The authors did identify a subgroup of the more severely affected patients in whom Factor VIIIc levels were increased, with shortened activated partial thromboplastin times (APTTs) associated with mild elevations of antithrombin III.     

The effects of stroma-free and dextran-conjugated hemoglobin on hemodynamics and carotid blood flow in hemorrhaged guinea pigs

Hemoglobin solutions are potential resuscitative fluids with volume expanding and oxygen delivery abilities developed to reduce the use of blood transfusion. Most hemoglobin solutions in clinical trials increase transiently arterial pressure by inhibiting nitric oxide-dependent vasodilation. Our objective was to compare the effects on central hemodynamics and carotid blood flow of two hemoglobin solutions after resuscitation from hemorrhage in anesthetized guinea pigs. After anesthesia and instrumentation, severe hemorrhage was induced by withdrawing 50% of the blood volume. Resuscitation was performed after 15 min of hypovolemia with 5% albumin, stroma-free hemoglobin, or hemoglobin conjugated to dextran-benzenetetracarboxylate (Dex-BTC-Hb). The mean arterial pressure (MAP), carotid blood flow (CBF), vascular resistance index and heart rate (HR) were monitored for 3 hours after resuscitation. After hemorrhage, MAP and CBF dropped to 57.6 +/- 4.4% and 58.9 +/- 3.7% of control values respectively. Albumin failed to maintain hemodynamics in the decompensatory phase of shock. Both hemoglobin solutions gave rise to a transient increase in MAP (35%); stroma-free hemoglobin increased the CBF (150%) and resistance index (24%) whereas Dex-BTC-Hb had no effect on CBF and vascular resistances. None of the solutions affected the HR. Modified hemoglobin has attenuated effects on CBF and resistance index compared to stroma-free hemoglobin. This may be due to a balance between the stimulation of nitric oxide synthesis by shear-stress and the inhibition of vasodilation by nitric oxide trapping.     

Hemocompatibility of high strength oxide ceramic materials: an in vitro study

High strength oxide ceramic materials like alumina and zirconia are frequently used for artificial joints because of their biocompatibility and high wear resistance. Their suitability as materials for implants and biomedical devices with direct blood contact, such as cardiovascular implants or components for blood pumps and dialyzers, has not been confirmed to date. The objective of this study was to investigate whether oxide ceramics show sufficient hemocompatibility. Dense specimens were made out of alumina, zirconia, titanium oxide, and aluminum titanate. Polyvinylchloride and silicone were additionally tested as reference materials. Interactions of human blood with the surfaces were studied by investigating partial thromboplastin time (PTT), thrombin antithrombin III complex (TAT), free plasma hemoglobin concentration, complete blood count, complement factor 5a, and protein adsorption. The results from the PTT and TAT tests clearly indicated higher blood activation by the ceramic materials when compared to the two polymer materials. However, alumina and zirconia showed lower C5a concentrations and less protein adsorption than the reference materials. Our results revealed that oxide ceramic materials alone cannot be used for implants in direct blood contact without modification of the ceramic surface, for example, by made-to-measure inert nanocoatings.