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NEUROSURGICAL ANESTHESIA

The Effect of Graded Hypothermia (36°C–32°C) on Hemostasis in Anesthetized Patients Without Surgical Trauma

S. C. Kettner, MD^*,, C. Sitzwohl, MD^*, M. Zimpfer, MD MBA^*,, S. A. Kozek, MD^*, A. Holzer, MD^*, C. K. Spiss, MD^*, and U. M. Illievich, MD^*

*Department of Anesthesiology and General Intensive Care, University of Vienna, General Hospital Vienna, Vienna, Austria; and Ludwig Boltzmann Institute of Clinical Anesthesiology and Intensive Care, Vienna, Austria

Address correspondence and reprint requests to Stephan C. Kettner, MD, Department of Anesthesiology and General Intensive Care, General Hospital Vienna, 18-20 Währinger Gürtel, A-1090 Vienna, Austria. Address e-mail to stephan.kettner{at}akh-wien.ac.at

	Abstract

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The isolated effects of hypothermia on hemostasis have not been investigated in healthy humans. We cooled 16 anesthetized patients scheduled for elective intracranial surgery to 32°C body core temperature and assessed prothrombin time (PT), activated partial thromboplastin time, thrombelastogram (TEG®), closure time, and platelet count at 36°C, 34°C, and 32°C body core temperature after the induction of anesthesia but before surgical intervention. Activated partial thromboplastin time, hematocrit, and closure time did not change, whereas PT and platelet count decreased during cooling. Platelet count decreased without a decrease in hematocrit; hence, a dilution by administered fluids seemed unlikely. The small decrease of platelet count is probably clinically irrelevant in patients with normal platelet count and function. The small decrease in PT indicates an alteration of the extrinsic pathway of coagulation. TEG® measurements showed a delay of clot formation in temperature-adjusted measurements but showed no change if the test temperature was 37°C. This indicates that hypothermia reduces plasmatic coagulation and platelet reactivity. However, the clot strength is not altered by hypothermia. All coagulation variables remained within the normal ranges. Our results may indicate that moderate short-term (4-h) hypothermia has only minor adverse effects in healthy humans. We can make no statement about the effects of hypothermia of longer duration.

IMPLICATIONS: This study investigated the isolated effects of hypothermia in healthy anesthetized humans. We found only minor effects of body temperature reduction to 32°C on assessed coagulation variables, indicating only minor effects in otherwise healthy humans.

	Introduction

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Hemostasis is the result of a complex interaction between the endothelium, platelets, and plasmatic coagulation factors. The effects of hypothermia on different parts of the hemostatic system have been investigated in animal studies, in in vitro studies, and in studies investigating trauma patients or patients undergoing cardiac procedures (1–17). Because trauma and massive transfusion or cardiopulmonary bypass affects hemostasis, the isolated effects of hypothermia on the hemostatic system remain unclear.

The function of the hemostatic system is assessed by different coagulation tests. The activity of coagulation factors is usually assessed by plasmatic coagulation tests, such as prothrombin time (PT) and activated partial thromboplastin time (APTT). Plasmatic coagulation can also be assessed by thrombelastogram (TEG®; Haemoscope Corp., Skokie, IL), an easy and reliable coagulation monitor that can be performed at different temperatures. TEG® is a global assessment of the hemostatic system, including interactions between plasmatic coagulation and platelets (18,19).

Another global test to assess platelet function is the bleeding time. Bleeding times are widely used to screen patients, although their accuracy is questioned by many investigators (20). An automated, computerized in vitro form of the bleeding time is the closure time (21). The closure time correlates with platelet function and is sensitive for changes in primary hemostasis (22).

The aim of this study was to investigate the isolated effects of hypothermia on the hemostatic system in healthy anesthetized humans without trauma or surgical intervention. Accordingly, we cooled patients scheduled for elective intracranial surgery to 32°C body temperature (BT) after the induction of anesthesia, but before surgical intervention. At 36°C, 34°C, and 32°C (BT), we assessed PT, APTT, TEG®, closure time, and platelet count.

	Methods

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Sixteen consecutive qualifying patients scheduled for elective intracranial surgery were included after written, informed consent in this IRB-approved study. Patients were anesthetized with propofol and fentanyl, orotracheally intubated after neuromuscular blockade with vecuronium, and controlled-ventilated with a fraction of inspired oxygen of 0.3. Anesthesia was maintained by continuous infusion of 8–12 mg · kg^-1 · h^-1 of propofol, 1.5–4 µg · kg^-1 · h^-1 of fentanyl, and 0.1–0.2 mg · kg^-1 · h^-1 of vecuronium. To maintain normovolemia, 2–4 mL · kg^-1 · h^-1 of normal saline was infused during the study period. Normocapnia was monitored by analysis of arterial blood gases with the alpha-stat regimen. One heparin-free arterial catheter was inserted into a radial artery for invasive blood pressure monitoring and drawing of blood samples. One peripheral venous line was inserted into a forearm vein to maintain normovolemia by infusion of 0.9% saline. BT was measured by a temperature probe inserted into the distal esophagus. The correct position of the probe was verified by a chest radiograph. Patients were cooled to 32°C by surface cooling with a forced-air cooling system (Polar Air^TM; Augustine Medical Inc., Eden Prairie, MN) and a cooling blanket (Blanketroll II^TM; Sub-Zero, Cincinnati, OH) before the start of the surgical intervention. Arterial blood samples for all measurements were obtained at 36°C, 34°C, and 32°C BT. Blood samples were collected in silicone-coated tubes containing 0.129 M buffered sodium citrate. Platelet and red cell count were measured by a Coulter counter. For the plasmatic coagulation test, citrated plasma was frozen and stored at -80°C. PT and APTT were measured at the end of the collection period by using a photometric coagulation analyzer. Factor VII was determined in plasma samples with a one-stage clotting assay by using factor-deficient plasma from Behringwerke (Marburg, Germany) and Thromborel® as the PT reagent on a STA analyzer (normal range for Factor VII, 75%–130%)

Four temperature-adaptable TEGs® were used in this investigation. One was adjusted to 37°C and served as the control; the others were adjusted to 36°C, 34°C, and 32°C. The accuracy of temperature was verified by measuring the temperature of the thrombelastograph cuvettes filled with normal saline and was within a deviation of ±0.2°C. Measurements were performed with disposable plastic pins and cups (Haemoscope), which were inserted at least 20 min before measurements to confirm the exact temperature of the cup. Immediately after sample collection, TEG® was performed with 300 µL of citrated blood and 40 µL of 0.645% CaCl₂. The temperature-adjusted and normothermic measurements were performed from the same blood sample and started simultaneously.

The TEG® reaction time (r) is the time from the start of measurement until initial fibrin formation. The clot formation time (k) measures the time necessary to reach 20 mm of clot strength. Values for r and k are expressed in millimeters; because the chart speed is 2 mm/min, the time in minutes is equal to the distance in millimeters divided by 2 (normal ranges: r, 10–19 mm; k, 4–10 mm). The angle measures the speed of fibrin buildup and cross-linking, which resembles the speed of clot strengthening (normal range, 44°–56°). The maximum amplitude (MA) measures the maximal clot strength, which is dependent on platelet function and, to a lesser extent, on fibrinogen level (normal MA range, 50–64 mm).

A Thrombostat 4000^TM was used to measure the closure time. This closure time reflects variables of primary hemostasis in vitro according to the system of Kratzer and Born (21). A Teflon capillary, serving as an artificial vessel, is perfused with whole blood under a constant pressure of -40 mm Hg at 37°C. An aperture of a cellulose-acetate filter with a 150-µm diameter reflects an injured part of a cut arteriole where platelets adhere and aggregate, forming a plug. The cellulose-acetate filter is soaked with 40 µL of adenosine 5'-diphosphate (20 mM) 2 min before the start of the measurement. As the microthrombus grows and gradually occludes the aperture, blood flow diminishes. Blood flow passing the capillary is measured over time, and the result is expressed in terms of closure time.

The samples of 800 µL of citrated blood were incubated for 2 min before measurement in the Thrombostat to reach a temperature of 37°C. All measurements of closure time were performed at 37°C.

After testing for normal distribution of the data (Kolmogorov-Smirnov test), the Mann-Whitney U-test, followed by Bonferroni’s correction, was performed to test for differences between the different temperature groups. Significance was considered for P 0.05; data are expressed as mean ± SD, except where otherwise indicated.

	Results

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We investigated 16 patients (7 men and 9 women) with a median age of 38 yr (range, 20–55 yr) and a median body weight of 68 kg (range, 45–91 kg). All patients were scheduled for temporal lobe epilepsy surgery, and no patient was taking medication except anticonvulsants. Active cooling to 32°C BT required 182 ± 53 min. During this time, patients received 1020 ± 270 mL, or 5–7 mL · kg^-1 · h^-1, of normal saline and had a urine output of 400 ± 220 mL, or 2–3 mL · kg^-1 · h^-1.

APTT, hematocrit, and closure times did not change during the study period and were within the normal ranges (Table 1). PT and platelet count decreased with temperature reduction but stayed within the normal ranges (Table 1).

	Discussion

Top Abstract Introduction Methods Results Discussion References

 
In this study, the effects of hypothermia on coagulation in healthy anesthetized humans were determined by preoperative assessment of PT, APTT, TEG®, closure time, and platelet count during active cooling to 32°C BT. Our data showed no changes in APTT or closure time in patients during active cooling, whereas PT and platelet count decreased.

The platelet count decreased without a decrease in hematocrit; hence, a dilution of platelet count by the administered fluids seems unlikely. A decrease of platelet count during active cooling has been described in dogs (14,16), where a BT reduction to 32°C caused an approximately 70% decrease in platelet count. This decrease was caused by pooling of platelets in the spleen. No study has shown a decrease of platelet count by hypothermia in humans. Our data show a minor decrease of platelet count of approximately 5% (P < 0.05) at 32°C BT. Although statistically significant, this 5% decrease in platelet count is probably clinically irrelevant in patients with normal platelet counts. Furthermore, this decrease of platelet count did not lead to a decrease in the hemostatic function of platelets, as assessed by closure time or TEG®.

The decrease in PT with BT reduction indicates an alteration of the extrinsic pathway of coagulation, whereas the APTT, which assesses the intrinsic pathway of coagulation, remained unchanged. The extrinsic pathway includes only two reactions: the binding of Factor VII or VIIa to tissue factor and the subsequent activation of Factor X by the tissue factor/Factor VIIa complex (23). Unlike the other coagulation factors, tissue factor does not circulate in plasma, and for the assessment of the PT, tissue factor is added to the assay. Therefore, a likely cause for the decrease of PT would have been a decrease in the plasma levels of Factor VII, which is the coagulation factor with the shortest half-life. However, we measured the plasma levels of Factor VII in the frozen plasma samples and found no change in the plasma levels of Factor VII during active cooling to 32°C. Hence, we have no explanation for the decrease in PT with BT reduction.

TEG® measurements at 37°C did not change during cooling. In contrast, r, k, and changed in temperature-adjusted TEG® measurements (32°C). Similar results have been shown in hypothermic patients during liver transplantation and in patients undergoing cardiopulmonary bypass (11,13). The r represents the time necessary for building the first fibrin strands, and k represents the time for reaching a certain clot strength. The angle represents the formation rate of the clot. The r is mainly dependent on coagulation factor activity. The k and angle depend on plasmatic coagulation and the interaction of platelets with fibrin, increasing the stability of the clot. The change of these variables seems to indicate a reduction of activity in both coagulation factors and platelet function with decreasing temperature. This is consistent with the literature: plasmatic coagulation tests and r are sensitive to the temperature at which the test is performed (3–5). Test temperatures of 29°C or 28°C prolong PT and APTT by approximately 50% (4,5). Temperature reduction to 32°C increased clotting time by approximately 50% in our study. This high sensitivity of TEG® to temperature change might be explained by a cumulative slowing of the enzymatic reactions in the steps of the coagulation cascade, because TEG® includes many more steps than plasmatic coagulation tests such as PT or APTT. In contrast to plasmatic coagulation tests (e.g., PT and APTT), TEG® is a global assessment of hemostatic function. TEG® assesses the enzymatic coagulation cascade from the time of the initial fibrin forming, through platelet aggregation, to clot strengthening and fibrin cross-linkage (18). Our measurements revealed differences in normothermic and temperature-adjusted k and angle , indicating a reversibly reduced platelet function, as described in animal models and in humans (2,14,16).

Another variable for the platelet function is the MA of TEG®, which reflects the absolute clot strength, but not a coagulation time, such as r and k. The MA is mainly dependent on the platelet function and, to a lesser extent, on plasma levels of fibrinogen. It was not affected by temperature reduction to 32°C, whether measurements were performed with the temperature adjusted or at 37°C. This finding, together with increased r and k and smaller angle , indicates a decrease in the speed of clot formation by temperature reduction, but not a reduction of clot quality, which is represented by the MA.

In our opinion, performing the TEG® at 37°C in hypothermic patients might overestimate the speed of clot formation. However, temperature-adjusted TEG® measurements could lead to unnecessary therapeutic interventions, when prolongation of r and coagulation time are caused by hypothermia and treated with the administration of coagulation factors.

Another described side effect of hypothermia is the release of a heparin-like substance during profound hypothermia in dogs (24). Neither APTT nor TEG® measurements, which are sensitive for heparin, changed during the study when measured at 37°C. This indicates that the release of a heparin-like substance is unlikely in humans during mild to moderate hypothermia.

A limitation of this study is that we investigated only the effects of short-term hypothermia, because we investigated the effects of BT reduction to 32°C within 4 hours. We can make no statement about the effects of hypothermia of longer duration.

We conclude that hypothermia has only minor effects on the coagulation system in anesthetized healthy patients during active cooling to 32°C BT. Coagulation times as assessed by temperature-adjusted TEG® measurements are prolonged, indicating a slowing of both the enzymatic reactions of the coagulation cascade and the speed of the interaction between the coagulation cascade and platelets. However, the resulting clot strength, as indicated by the MA of TEG®, was not altered by hypothermia. All other coagulation variables remained within the normal ranges. This may indicate that short-term (four-hour) mild to moderate hypothermia has only minor effects in anesthetized healthy humans but might amplify preexisting coagulopathies.

	Acknowledgments

 
Supported by a grant from the Medical Fund of the Mayor of Vienna (Project No. 1601).

	References

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Anesthesia & Analgesia® is published for the International Anesthesia Research Society® by Lippincott Williams & Wilkins with the assistance of Stanford University Libraries' HighWire Press®. Copyright 2006 by the International Anesthesia Research Society. Online ISSN: 1526-7598   Print ISSN: 0003-2999