高压电烧伤大鼠血浆FIB、D-二聚体、血清FN的变化及乌司他丁的干预作用

发布时间：2018-08-17 18:32

【摘要】：目的：高压电烧伤是一种特殊原因的烧伤，可引起一系列复杂的病理学变化，微循环障碍则是其中重要的病理变化之一。凝血机制异常在机体微循环障碍发生和发展过程中起重要作用。烧伤后由于血管内皮受损、血小板活化、炎症介质释放、凝血系统功能增强及继发性纤溶系统功能相对增强，导致血液高凝状态，甚至引起弥漫性血管内凝血（disseminatedintravascular coagulation，DIC）。血浆纤维蛋白原（fibrinogen，FIB）、D-二聚体是血液发生血栓及血栓前状态的凝血及纤溶系统活性改变的分子标志物，可反映体内凝血和纤溶过程的变化；纤维连接蛋白（fibronectin，FN）在血小板聚集和黏附过程中起重要作用，可反映烧伤后血液的黏滞性。因此，我们设计本实验，旨在研究高压电烧伤早期大鼠FIB、D-二聚体、FN变化，并通过乌司他丁（ulinastain，UTI）进行干预，研究烧伤后凝血功能和纤溶活性的分子机制，探讨UTI治疗高压电烧伤的可行性。方法： 1.实验动物分组：健康成年SD大鼠（由河北医科大学动物实验中心提供，合格证编号为1211057）180只，按随机数字表法分成三组，即假高压电烧伤组（简称对照组）、高压电烧伤组（简称电伤组）、高压电烧伤UTI治疗组（简称治疗组），每组各60只。每组又按观察时间分为电击前15min、电击后5min、电击后1h、2h、4h、8h六个时相，即T0、T1、T2、T3、T4、T5，每时相10只。 2.实验前准备：将大鼠编号、称重，左上肢、右下肢及前胸脱毛。将实验药品按实验需要配成所需浓度。 3.高压电烧伤模型制作：首先连接好实验变压器和调压器电线。用1%戊巴比妥钠腹腔注射麻醉大鼠，按40mg/kg给药，麻醉成功后，将大鼠仰卧于专用电击实验台上，固定四肢，将两个1cm×1cm电极片分别固定于大鼠的左上肢（电流入口）、右下肢（电流出口）脱毛区。接通电源后，调整调压器使升压器输出电压至2kV，连接升压器，使高压电流通过大鼠，电击3s。对照组制作假电伤模型，不合闸通过电流，其余步骤与电伤组一致。电击伤后即刻，治疗组腹腔注射5×104u/kg UTI，对照组及电伤组腹腔内注射等量的生理盐水。 4.标本采集与保存：将高压电烧伤模型复制成功的大鼠开胸暴露心脏，直视下心脏抽血6mL，其中3mL置于一次性使用真空采血管（红）中，3mL置于一次性使用真空采血管（蓝）中，轻轻颠倒数次后静置30min，上离心机，以3000转/min离心10min，红管中取上清液置于Eppendorf管中在-70℃条件下保存。 5．指标检测：蓝管离心分离血浆后使用全自动血凝仪上机自动检测每组大鼠六个时相组FIB及D-二聚体含量。置于冰箱的血清采用ELISA双抗体夹心法，检测FN含量。 6．实验数据处理：采用SPSS13.0统计软件，行两因素析因设计的方差分析，多重比较采用LSD法t检验。以P＜0.05为有统计学差异。结果： 1．大鼠血浆FIB含量变化电伤组FIB含量总体高于对照组（P＜0.01），且FIB含量受伤后受时间因素的影响（P＜0.01）,至伤后8h达到最高值。治疗组FIB含量总体低于电伤组（P＜0.01），且FIB含量治疗后受时间因素的影响（P＜0.01），，但只有治疗组伤后5min高于本组伤前值（P＜0.01），伤后1h、2h、4h、8h与伤前无明显差异（P＞0.05）。 2．大鼠血浆D-二聚体含量变化电伤组D-二聚体含量总体高于对照组（P＜0.01），且D-二聚体含量受伤后受时间因素的影响（P＜0.01），呈增高趋势。治疗组D-二聚体含量总体低于电伤组（P＜0.01），且D-二聚体含量治疗后受时间因素的影响（P＜0.01），治疗组伤后5min～8h各时相均高于本组伤前值（P＜0.001） 3．大鼠血清FN含量变化电伤组FN含量总体低于对照组（P＜0.01），且FN含量受伤后受时间因素的影响（P＜0.05），伤后各时相均低于本组伤前值（P＜0.05）。治疗组FN含量总体高于电伤组（P＜0.05），但FN含量治疗后不受时间因素的影响（P＞0.05）。结论： 1．高压电烧伤早期大鼠血浆FIB、D-二聚体表达水平均有不同程度升高，血清FN表达水平下降，说明高压电烧伤后存在高凝状态、纤溶系统功能异常及微循环障碍。 2．高压电烧伤早期UTI治疗后FIB、D-二聚体表达水平降低，FN表达水平升高，说明UTI治疗可以抑制凝血亢进与继发性纤溶，减轻高凝状态，从而改善微循环，防止血栓形成。
[Abstract]:Objective: High voltage electrical burn is a special cause of burns, which can cause a series of complex pathological changes. Microcirculation disorder is one of the important pathological changes. Abnormal coagulation mechanism plays an important role in the occurrence and development of microcirculation disorder. Release, enhancement of coagulation system and secondary fibrinolysis system lead to hypercoagulability, even diffuse intravascular coagulation (DIC). Plasma fibrinogen (FIB) and D-dimer are coagulation and fibrinolysis system activities in the thrombosis and pre-thrombotic state of the blood. The changes of molecular markers can reflect the changes of coagulation and fibrinolysis in vivo. Fibronectin (FN) plays an important role in the process of platelet aggregation and adhesion, and can reflect the blood viscosity after burns. Ulinastatin (UTI) was used to study the molecular mechanism of coagulation function and fibrinolytic activity after burns, and to explore the feasibility of UTI in the treatment of high voltage electrical burn.
Method:
1. Experimental animal groups: 180 healthy adult SD rats (provided by the Animal Experimental Center of Hebei Medical University, with the certificate number of 121,1057) were randomly divided into three groups: sham high-voltage electrical burn group (control group), high-voltage electrical burn group (electrical burn group), high-voltage electrical burn UTI treatment group (treatment group), 60 rats in each group. Group A was divided into six phases, namely, T0, T1, T2, T3, T4, T5, 5 minutes before and 1, 2, 4 and 8 hours after electric shock.
2. Pre-experiment preparation: Rats were numbered, weighed, left upper extremity, right lower extremity and chest depilation.
3. Making high voltage electric burn model: Firstly, the experimental transformer and the voltage regulator wire were connected. Anesthetized rats were injected intraperitoneally with 1% sodium pentobarbital. After anesthesia, the rats were given 40 mg/kg. After successful anesthesia, the rats were lying on their backs on a special electric shock test bench, fixed their limbs, and fixed two 1 cm *1 cm electrode plates on the left upper limbs (current inlet) and right lower limbs (current inlet). After switching on the power supply, adjust the voltage regulator to make the output voltage of the booster to 2 kV, and connect the booster to make the high voltage current pass through the rats and shock them for 3 s. The control group made the model of false electric injury, which was not suitable for the switch through the current, and the other steps were the same as that of the electric injury group. The same amount of saline was injected into the abdominal cavity.
4. Sample Collection and Preservation: The heart was exposed after thoracotomy in rats with high-voltage electrical burn. Blood was drawn from the heart under direct vision for 6 mL. Among them, 3 mL was placed in a disposable vacuum (red), 3 mL was placed in a disposable vacuum (blue), and 30 minutes was left after several times of inversion. The heart was centrifuged for 10 minutes at 3000 rpm. The supernatant was placed in the Eppendorf tube and stored at -70 C.
5. Indicators: After centrifugation, the plasma was separated by blue tube and the contents of FIB and D-dimer were detected automatically by automatic coagulator. The serum in refrigerator was detected by ELISA double antibody sandwich method.
6. Data processing: SPSS13.0 statistical software was used to analyze the variance of two-factor factorial design, and LSD t test was used for multiple comparisons.
Result:
Changes of plasma FIB content in 1. rats
The content of FIB in the treatment group was lower than that in the electric injury group (P < 0.01), but only 5 minutes after injury was higher than that in the control group (P < 0.01). There was no significant difference in 1H, 2h, 4H and 8h after injury (P > 0.05).
Changes of plasma D- two dimer content in 2. rats
The content of D-dimer in the treatment group was lower than that in the electric injury group (P < 0.01), and the content of D-dimer in the treatment group was higher than that in the control group (P The pre injury values were (P < 0.001).
Changes of serum FN level in 3. rats
The FN content in the treatment group was higher than that in the electric injury group (P < 0.05), but the FN content was not affected by the time factor (P > 0.05).
Conclusion:
1. The levels of FIB and D-dimer in plasma and FN in serum of rats with high-voltage electrical burn increased to different degrees in the early stage, indicating that there was hypercoagulable state, abnormal fibrinolytic system function and microcirculation disturbance after high-voltage electrical burn.
2. After early UTI treatment, the expression of FIB, D-dimer and FN decreased, indicating that UTI treatment can inhibit hypercoagulation and secondary fibrinolysis, reduce hypercoagulability, improve microcirculation and prevent thrombosis.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R647

【参考文献】