Research on mechanism of apoptosis and organ damage after hemorrhagic shock and resuscitation

雄性BALB/c小鼠失血至平均动脉压40+/-5毫米汞柱90min。复苏时给予7.5%氯化钠4ml/kg(高渗盐水；HS)并放血(SB)，或两倍失血量(2LR+SB)+SB或假手术(仅导尿管)或对照(不治疗)。采用末端脱氧核苷酸转移介导的5-三磷酸脱氧尿苷缺口末端标记法和DNA梯状末端标记法检测小肠细胞凋亡率。Western blotting检测caspase-3、HSP70、HSP40表达的激活情况。失血性休克复苏后2小时2LR+SB组大鼠出现细胞凋亡，6h时2LR+SB组大鼠肠道损伤明显高于其他组(P<0.0 1)，HS+SB组2小时的细胞凋亡指数明显低于2LR+SB组(P<0.0 1)，HS+SB组6小时的肠损伤明显高于其他两组(P<0.0 1)。与…组比较，HS+SB组的肠损伤明显减轻(P<0.0 1)。与2LR+Sb相比，6h时降低幅度更大(p&lt；0.01)。HS+SB组与2LR+SB组相比，Hs+SB组2 h时caspase-3活性降低。HS+SB组HSP70、HSP40的表达在2小时显著高于2LR+SB组，肝、胸腺、脾细胞凋亡率在HS+SB组也明显低于2LR+SB组(p<0.05)。HS+SB组的Fas和墨汁在复苏后2小时略低于2LR+SB组。结论：高渗盐水复苏可减轻失血性休克复苏后肠组织的细胞凋亡和肠损伤。因此，高渗盐水复苏对细胞凋亡的影响可能在失血性休克和复苏中起重要作用。HS复苏可能会干扰小肠中caspase-3的激活。HS复苏可防止HSP70和HSP40的丢失，这可能与HS对小肠的有益作用有关。Fas和INK可能参与了失血性休克复苏后肝脏细胞的凋亡。较少

Male BALB/c mice were hemorrhaged to the mean arterial pressure of 40+/-5 mmHg for 90 min. Resuscitation was made by 4ml/Kg of 7.5% NaCl (hypertonic saline; HS) and shed blood (SB) or two times the volume of shed blood (2LR+SB) +SB or sham (catheter only) or control (no treatment).(Small intestine) Intestinal injury was devised and based on the extent of the vacuolation at basal area of the intestinal villi. Apoptosis score of the small intestines were examined by terminal deoxynucleotidyl transfer-mediated deoxyuridine 5-triphosphate nick end labeling (TUNEL) methods and DNA ladder assay. Activation of caspase-3, HSP 70, HSP 40 expression were assessed by Western blotting. Hemorrhagic shock and resuscitation induced the apoptosis in 2LR+SB at 2h and intestinal damage in 2LR+SB at 6h compared to other groups significantly (p<0.01) Apoptosis index of HS+SB at 2h group was significantly reduced compared to 2LR+SB at 2h (p<0.01) and also intestinal damage of HS+SB at 6h group was signiflc … More antly reduced compared to 2LR+SB at 6h (p<0.01). Activation of caspase-3 of HS+SB at 2h group was reduced compared to 2LR+SB at 2h group. On the contrary HSP 70, HSP 40 expression in HS+SB at 2h were higher than 2LR+SB at 2h.(Liver, Thymus, Spleen) Apoptosis of liver, thymus and spleen were also reduced in HS+SB at 2h compared to 2LR+SB at 2h (p<0.05). FAS and INK in HS+SB at 2h were slightly reduced than 2LR+SB at 2h.Conclusions : Hypertonic saline resuscitation reduced the apoptosis and intestinal damage after hemorrhagic shock and resuscitation. Thus, the effect of hypertonic saline resuscitation toward apoptosis may have important role in hemorrhagic shock and resuscitation. HS resuscitation may interfere with caspase-3 activation in the small intestine. HS resuscitation may prevent the loss of HSP 70 and HSP 40, which might be involved in the beneficial effect for HS in the small intestine. FAS and INK may be involved in the apoptosis of the liver after hemorrhagic shock and resuscitation. Less