低温和酸中毒对人全血凝血功能损害的协同作用

背景低温和酸中毒对不同临床状态下凝血疾患的影响已有报道。本研究中，我们对全血的凝血功能进行评估，以探讨低温和酸中毒对凝血的影响。方法采自10名健康志愿者（2女，8男）的全血样本（3．000μl），加入40μl摩尔浓度递加的盐酸，使血液酸化至pH（α-pH固定计）介于7．0和7．37之间。用内源性（InTEM^TM）和外源性（ExTEM^TM）活化方法孵化30分钟后，再用旋转血栓弹性测定法分析凝血功能。为了评估凝血功能对温度的依赖性，所有试验均在血液／血栓弹性测定仪的温度30、33、36和39℃下分别进行。此外，通过添加细胞松弛素D进行了另一项外源性激活方法，以此来检测在无血小板作用时，血凝块的形成情况。结果正常pH下低温导致了凝血时间延长[ExTEM：65秒±3．6（36℃）vs85±4（30℃），P〈0．001；InTEM：181秒±10（36℃）vs226±9，P〈0．001]，血凝块形成时间延长[ExTEM：105秒±5（36℃）vs 187±6（30℃），P〈0．001；InTEM：101秒±5（36℃）vs175±7，P〈0．001]，并且伴有α角的缩小，[ExTEM：65．6±1．8（36℃）vs 58±1．1，P〈0．01；InTEM：70．5±1．8（36℃）vs 60．2±1．5，P〈0．001]。最大血栓弹力仅在InTEM测定法中显著受损[56．9mm±0．9（36℃）vs 52．7±0．9，P〈0．05]。相反，正常体温下酸中毒本身无明显作用。酸中毒放大了低温的作用，并在内源性和外源性激活实验中，两者对凝血时间、d角、最大血栓弹力的影响有协同作用。用细胞松弛素D消除血小板功能后进行的纤维蛋白凝块形成试验显示，凝血功能并没有受损。血凝块溶解在低温和（或）酸中毒环境下减少，而在体温升高的情况下增加。结论在此体外研究中，血栓弹性测定法显示：酸中毒使低温引起的凝血功能改变恶化，而不伴低温的酸中毒对凝血功能无显著影响。这种情况是由于凝血因子和血小板功能受到抑制引起的。因此，在37℃下进行的血栓弹性测定法高估了低温尤其伴酸中毒时凝血功能的完整性。

BACKGROUND： Hypothermia and acidosis were reported to influence coagulopathy in different clinical settings. We evaluated whole blood coagulation to determine the effects of hypothermia and/or acidosis on hemostasis. METHODS： Whole blood samples （3.000 μl） from 10 healthy volunteers （2 female, 8 male） were addified by adding 40 μl of hydrochloric acid of increasing molarity to achieve a blood pH （α-stat） between 7. 0 and 7. 37, and coagulation was analyzed by rotational thromboelastometry after an incubation period of 30 min using both intrinsically （InTEMTM） and extrinsically （ExTEMTM） activated assays. To assess temperature-dependent effects, all tests were performed at blood/thromboelastometer temperatures of 30, 33, 36, and 39℃, respectively. An additional extrinsically activated test with addition of cytochalasin D was performed to examine dot formation without platelet contribution. RESULTS： Hypothermia at a normal pH prodhced an increased coagulation time [ ExTEM： 65 s ±3.6 （36℃） vs 85±4 （30℃）, P 〈0. 001; coagulation time, InTEM： 181 s ± 10 （36℃） vs 226±9, P 〈0. 0011 and dot formation time [ ExTEM： 105 s ± 5 （36℃） vs 187 ± 6 （30℃）, P 〈0. 001 ] ; clot formation time [ InTEM： 101 s ±5 （36℃） vs 175 ± 7, P 〈0. 001 ], as well as decreased α angle [ ExTEM： 65.6 ± 1.8 （36℃） vs 58 ± 1.1, P 〈0. 01, P 〈 0. 01 ; InTEM： 70. 5 ± 1.8 （36℃） vs 60. 2 ±1.5, P 〈 0. 001 ]. Maximum dot firmness was significantly impaired only in InTEM assays [ 56.9 mm± 0.9 （36℃） vs 52.7 ± 0. 9, P 〈 0.05]. In contrast, acidosis per se had no significant effects during normothermia. Acidosis amplified the effects of hypothermia, and synergistically impaired dotting times, α angle, and decreased maximum dot firmness, again in both extrinsically and intrinsically activated assays. Formation of a fibrin dot tested after abolition of platelet function by cytochalasin D was not impaired. Clot lysis decreased under hypothermic and/ or addotic conditions, but increased with hyperthermia. CONCLUSIONS： In this in vitro study, hypothermia produced coagulation changes that were worsened by acidosis whereas acidosis without hypothermia has no significant effect on coagulation, as studied by thromboelastometry. This effect was mediated by the inhibition of coagulation factors and platelet function. Thus, thromboelastometry performed at 37℃ overestimated integrity of coagulation during hypothermia in particular in combination with acidosis.