亚低温对脓毒症大鼠肝肺组织的保护作用

The protective effects of mild hypothermia on liver and lung tissues in septic rats

目的测定亚低温(MHT)情况下脓毒症大鼠肝组织能量代谢和肺组织肿瘤坏死因子-α(TNF-α)mRNA表达的变化,并结合肝细胞和肺组织超微结构的改变,初步探讨MHT能否减轻脓毒症大鼠组织损伤的程度。方法24只Wistar大鼠随机分为正常对照组(NC组)、内毒素组(LPS组)和MHT组,应用高效液相色谱法及生化法分别测定各组肝组织腺嘌呤核苷酸及乳酸水平,应用RT-PCR方法分别测定各组肺组织TNF-α mRNA表达变化,并在透射电镜下观察肝细胞、肺组织超微结构的变化。结果LPS组大鼠肝组织(湿重)腺苷酸水平为ATP(4.093±0.424)μmol·g-1,ADP(1.331±0.136)μmol·g-1,AMP(1.331±0.312)μmol·g-1;MHT组大鼠肝组织(湿重)腺苷酸水平为ATP(4.519±0.028)μmol·g-1,ADP(1.483±0.108)μmol·g-1,AMP(1.544±0.301)μmol·g-1;LPS组和MHT组均较NC组腺苷酸水平ATP(4.990±0.455)μmol·g-1,ADP(1.632±0.181)μmol·g-1,AMP(1.737±0.407)μmol·g-1降低,尤以LPS组明显。MHT组肝组织腺苷酸水平较LPS组明显升高;MHT组肝乳酸含量为每克蛋白(1.424±0.213)mmol,明显低于LPS组的每克蛋白(1.676±0.267)mmol(P<0.05),而与NC组差异无统计学意义[vs每克蛋白(1.379±0.314)mmol,P>0.05]。LPS组大鼠肺组织TNF-α mRNA1、2和3h表达分别为(1.029±0.055)、(1.132±0.068)和(1.107±0.044);MHT组大鼠肺组织相应时点TNF-α mRNA表达分别为(0.835±0.041)、(1.056±0.037)和(1.056±0.032),均较NC组肺组织相应各时点TNF-α mRNA表达(0.625±0.009,0.631±0.007,0.612±0.011)明显增加(P<0.05),尤以LPS组明显。MHT组肺组织TNF-α mRNA较LPS组表达明显减少(P<0.05)。透射电镜下可见MHT组肝细胞及肺组织超微结构损伤明显轻于LPS组。结论MHT可减缓脓毒症大鼠肝组织能量的消耗,减轻肝组织损伤,还可减少脓毒症大鼠肺组织TNF-α mRNA的表达,减轻肺组织炎性损伤,MHT可作为防治脓毒症的一种新方法。

Objective To investigate the effects of mild hypothermia on septic rats, including the energy metabolization of liver and the levels of tumor necrotic factor-or mRNA combined with the ultrastructure changes of liver and lung and to discuss preliminarily whether mild hypothermia treatment（MHT） can decrease the degrees of septic injury . Methods Twenty four Wistar rats were divided randomly into normal control （NC） group, lipopolysaccharide （LPS） group , mild hypothermia theatment （MHT） group . The models of septic rats were prepared. Livers were conservated in liquefacient Nitrogen to determine the levels of Adenine nucleotides, and lactate and lungs were frozen in Eppendorf tubes at - 80℃ to determine the levels of TNF-αmRNA. The levels of adenine nucleotides and lactate were determined by high performance liquid chromatography （HPLC） and biochemistry method. The levels of TNF-α mRNA were determined by RT-PCR method. The ultrastructure changes of liver and lung were investigated using electronic microscope. Finally the data were dealed with SAS statistical software. Results Compared with the NC group rats [ATP （4.990 ±0.455）, ADP （ 1.632 ±0. 181 ） ,AMP （ 1. 737 ±0.407）μmol·g^-1 （wet） ] , the levels of adenine nucleotides in liver tissue of LPS group rats [ ATP （4.093± 0. 424）, ADP （ 1. 331 ± 0. 136）, AMP （ 1.331± 0.312） μmol·g^-1 （wet） ] and MHT group rats [ ATP （4.519 ± 0.028 ）, ADP （ 1. 483 ± 0. 108 ）, AMP （ 1. 544 ± 0. 301 ）μmol·g^-1 （wet） ] were decreased.The levels of adenine nueleotides were increased significantly in MHT group rats than that in LPS group rats; The lactate levels of MHTgroup [（1. 424 ± 0. 213） mmol ·g^-1（prot）] were lower than that of LPS group [（1. 676 ± 0. 267） mmol·g^-1 （prot）,P〈0.05] , and no significant difference was found comparing with NC group[ （1. 379 ± 0. 314）mmol ·g^-1 （prot） ,P 〉 0.05 ] ;Compared with the NC group rats （0. 625 ± 0. 009,0. 631 ± 0. 007, 0. 612 ±0. 011 ） , the levels of TNF-α mRNA in lung tissue of LPS group rats （ 1. 029 ± 0. 055,1. 132 ± 0. 068,1. 107 ± 0. 044） and MHT group rats （0. 835 ± 0. 041, 1. 056 ±0. 037,1. 056 ±0. 032）were increased （P 〈 0.05 ）. The levels of lung TNF-α mRNA were lower significantly in MHT group rats than that in LPS group rats（ P 〈 0.05 ） ; The liver and lung in MHT group rats showed less severe uhrastructure changes than that in LPS group rats electronic-microscopically. Conclusions Mild hypothermia could slow the energy expenditure in the liver tissue of septic rats, decrease TNF-α mRNA expression in the lung tissue of septic rats, and decrease the severity of liver and lung injury to some extent.