巨噬细胞移动抑制因子在感染性休克大鼠心血管机能障碍中的作用

Role of macrophage migration inhibitory factor in septic shock-induced cardiovascular dysfunction:experiment with rats

目的观察应用巨噬细胞移动抑制因子(MIF)阻断剂及糖皮质激素(GC)后感染性休克大鼠心血管机能和 MIF 表达的变化。方法 56只 SD 大鼠250～300 g,随机分7组(n_1=8):空白组(Sham 组);CLP 组(盲肠结扎穿刺);ISO-1组(MIF 拓仆酶抑制剂);MIF-Ab 组(MIF 多克隆抗体);DEX-1、DEX-5和 DEX-20组(地塞米松分别1 mg/kg、5 mg/kg、和20 mg/kg)。CLP 后6 h 超声心动仪观察每组大鼠左室舒张末和收缩末直径(LVEDD、LVESD)、左室短轴缩短率(FS%)、心输出量(CO)及肺动脉血灌流量(PP)。继后监测 MAP 和去氧肾上腺素(PE,0.5、1.0、2.0和2.5μg/kg)引起的MAP 增长百分比(ΔMAP%)。活体实验结束后,取大鼠胸主动脉环做离体张力实验,建立 PE 的剂量一反应曲线并计算 E_(max)、EC_(50)值和检测心肌和主动脉中 MIF 蛋白质表达。第二部分大鼠(n_2=10)实验干预后观察其72 h 存活率。结果心脏 LVEDD、LVESD 在 CLP 后6 h 分别下降56%和54%,应用ISO-1、MIF-Ab 和 DEX-20后显著升高(P<0.05)。FS%、CO 及 PP 也相应地得到提高(P<0.001)。而应用 PE 后,ΔMAP%由 CLP 组的6%分别升高至 ISO-1 12.1%、MIF-Ab 10.4%和 DEX-20 9.3%(均 P<0.05)。PE 引起的动脉环最大张力在 ISO-1和 MIF-抗体组较 CLP 组的0.87 g 升高至1.25 g和1.18 g(均 P<0.05),大剂量 GC 组在 PE 1.0×10^(-7)～1.0×10^(-5)mol/L 之间对动脉环张力的影响较 CLP 有改善(P<0.05),而大于1.0×10^(-5)mol/L 时差异无统计学意义。ISO-1、MIF-Ab 和 DEX-20的应用使心肌和主动脉内MIF的表达明显较 CLP 下降(P<0.05);然而,DEX-1和 DEX-5引起 MIF表达较 DEX-20增加(P<0.05)。应用 ISO-1、MIF-Ab 后大鼠72 h 生存率由 CLP 的0%分别提高到70%和60%(P<0.05);而 DEX 的应用均不能使大鼠生存率得到改善。结论 MIF 在感染性休克循环功能障碍中起着重要的中介作用,对 MIF 的拮抗和阻断明显改善休克时血流动力学、血管反应性及预后;大、小剂量 GC 对感染性休克大鼠的血流动力学及血管反应性的影响截然相反,但均不能改善其预后。

objective To investigate the role of macrophage migration inhibitory factor （MIF） in septic shock-induced cardiovascular dysfunction. Methods 56 SD rats were randomly divided into 7 equal groups ： CLP group （ undergoing cecal ligation and puncture so as to cause septic shock）, CLP ＋ ISO-1 group （ ISO-1, was injected before and after CLP）, CLP ＋ MIF antibody group （ MIF-Ab was injected before and after CLP）, CLP ＋ dexamethasone （ DEX）-1, 5, and 20 groups [ I, 5, or 20 mg/kg was injected 1 h after CLP）, and sham operation group. Echocardiography was performed 6 h after CLP to measure the LVEDD, LVESD, FS%, CO, and PP. Catheters were inserted into the femoral artery and vein to measure the mean arterial pressure （MAP） and to be used as the route of drug administration. Phenylephrine （PE） of the concentrations of 0. 5, 1, 2, and 2. 5 μg/kg was injected intravenously and then the MAP increase percentage （ A MAP% ） was calculated. Then the rats were euthanized with their hearts and aortas taken out. The aortas were cut into rings, and bathed in Krebs solution. PE of the concentrations of 1 mol/L to 30 μmol/L was added into the solution cumulatively to produce the dose-reaction curve of PE. The maximum energy （Emax） and median effective concentration （EC50） of PE were calculated. Western blotting was used to examine the protein expression of MIF in the myocardium and aorta. Another 70 SD rats were divided into 7 groups as mentioned above to observe the cumulative survival rates within 72 h. Results The LVEDD and LVESD of the CLP group decreased by 56% and 54% respectively 6 h after CLP, and the LVEDD and LVESD of the ISO-1, MOF-Ab, and DEX-20 groups were all significantly higher than that of the CLP group （ all P 〈 0. 05 ） The FS% of the CLP group was significantly lower than that of the Sham groups, and the FS% of the ISO-1, MOF-Ab, and DEX-20 groups were all significantly higher than that of the CLP group （ all P 〈 0. 05 ）. The PP value of the ISO-1, MIF-Ab, and DEX-20 groups were all significantly higher than that of the CLP group （ all P 〈 0. 06 ）. The CO of the CLP group was significantly lower than that of the Sham group, and those of the ISO-1, MIF-Ab, and DEX-20 groups were all significantly higher than that of the CLP group （all P 〈 0. 001 ）. The A MAP% of different group all increased after the addition of PE dosedependently, however, the A MAP% was significantly lower in the CLP group than in the Sham group （P 〈 0. 05 ）, and t significantly higher in the ISO-1, MIF-Ab, and DEX-20 groups than in the CLP group （ all P 〈 0. 05 ） . The values of PE-induced maximum aorta tension of the SO-1 and MIF-Ab groups were both significantly higher than that of the CLP group （ both P 〈 0. 05 ）. The values of PE-induced maximum aorta tension of the DEX-20 group were all higher than those of the CLP group when the PE concentration was between 1.0×10^-6 ～ 1.0×10^-5 mol/L （all P 〈0. 05）, however, were not significantly different those of the CLP group when the PE concentration was over 1.0×10^-5 mol/L. The values of Emax were significantly lower in the 6 experimental groups than in the Sham group （ all P 〈 0. 05 ）, however, were all significantly higher in the ISO-1, MIF-Ab, and DEX-20 groups than in the CLP group （ all P 〈 0. 05 ）. The values of EC50 were significantly higher in the 6 experimental groups than in the Sham group （ all P 〈 0. 05 ）, however, were significantly lower in the ISO-1, MIF-Ab, and DEX-20 groups than in the CLP group （all P 〈 0. 05）. The protein expression levels of MIF in the heart and aorta were significantly higher in the 6 experimental groups than in the Sham group （ all P 〈 0. 05 ）, however, the DEX-1 and DEX-5 groups showed significantly higher MIF expression than DEX-20 group （ both P 〈 0. 05 ）. The 72h survival rates of the ISO- 1 and MIF-Ab groups were both significantly higher than that of the CLP group （0%, both P 〈0. 05 ）. DEX of different dose failed to increase the survival rate. Conclusion MIF plays a pivotal role in the circulation dysfunction in septic ambience. Antagonism and blockade of MIF improve corresponding hemodynamics, vascular responsiveness, and prognosis. Glycocorticoid of high and low dose are poles apart in effects on septic hemodynamics and vaso-reactivity, however, fails to improve the prognosis of sepsis no matter how high is the dose.