强化胰岛素对严重烧伤大鼠心肌细胞TNF-α表达的抑制作用

Inhibitory effects of intensive insulin on myocardium expression of tumor necrosis factor alpha in severely burnt rats

目的探讨强化胰岛素对严重烧伤大鼠心肌细胞TNF-α表达的抑制作用及其可能机制。方法健康雄性SD大鼠18只,右颈静脉及左心室插管,随机均分为3组:假伤组,大鼠给以假烫;烧伤组,制作30%TBSAⅢ度烫伤后,常规给以液体复苏;强化组,烫伤后给以强化胰岛素治疗,液体总量同烫伤组。测定伤后0、1、2、3、4、5、6h的血糖及左室收缩压(LVSP)、左室舒张末压(LV-EDP)。伤后6h处死动物,留取左心室组织,分别用ELISA和定量PCR法测定TNF-α蛋白及其mRNA水平,Westernblot观察p-IκBα和IκBα变化。同时用烧伤血清培养新生鼠心肌细胞,观察胰岛素及其活化抑制剂LY294002对p-IκBα水平的影响。结果与假伤组比较,烧伤组动物伤后出现应激性高血糖反应,LVSP降低,LVEDP升高,左心室组织TNF-α蛋白及mRNA水平升高,p-IκBα蛋白表达增高,IκBα蛋白表达降低(P<0.05或0.01)。烧伤大鼠经强化胰岛素治疗后,血糖维持在4.5～5.2mmol/L之间,左心室功能得到明显改善,左心室组织TNF-α蛋白及mRNA水平显著降低,κBα的磷酸化和降解明显减轻(P<0.05或0.01)。LY294002能够消除胰岛素对烧伤血清培养心肌细胞κBα磷酸化的抑制作用。结论强化胰岛素可能通过减少IκBα的磷酸化降解而降低NF-κB活化,进而抑制心肌细胞TNF-α的转录与表达,发挥改善心肌功能的作用。

Objective To investigate the inhibitory effects of intensive insulin on myoeardiurn expression of tumor necrosis factor alpha （TNF-α） in severely burnt rats and its correlative mechanism. Methods In eighteen SD rats the right jugular vein and the left ventricle were cathetetized, and were divided into three groups randomly： （1） the sham bum group, the rats were treated by sham bum; （2） the bum group, the rats were inflicted with 30% TBSA Ⅲ degree bum, and then given fluid resuscitation routinely; （3） the intensive insulin group, the rats received intensive insulin treatment after bum, and the liquid volume given was equal to the burn group. The level of plasma glucose contents, the left ventricular systolic pressure （LVSP） and left ventricular end-diastolic pressure （LVEDP） were recorded at 1, 2, 3, 4, 5, 6h after bum, and animals were sacrificed at 6 hours after burn. The level of TNF-α in myocardium tissue was assayed by ELISA, and mRNA expression was determined by real-time PCR. Western blot was used to detect KBa phospborylation and degradation in left ventricle tissue. Meanwhile, bum serum-challenged cardiomyocytes were also used to detect the levels of insulin and LY294002, a specific insulin activation inhibitor, on the phosphorylation of IkBα. Results The animals showed stress hyperglycemia after bums, the function of heart was damaged （LVSP degraded and LVEDP increased）, the level of TNF and the expression of mRNA in the myocardium increased apparently, IkBα phosphorylation and degradation increased in left ventricle tissue and bum serum-challenged cardiomyocytes. Given the intensive insulin treatment to keep the blood glucose level between 4. 5 to 5. 2 mmol/L, the function of heart was significantly improved, the level of TNF-α was apparently lowered, and the IkBα phosphorylation and degradation were significantly decreased. LY294002 could abolish the inhibitory effect of insulin, and it acted on the IkBα phosphorylation on bum serum-challenged cardiomyocytes model. Conclusion Intensive insulin may decrease the activation of NF-kB through diminishing the IkBa phosphorylation and degradation, which inhibit the transcription and expression of TNF-α in myocardial cell, and improve the heart function accordingly.