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目的探讨细胞外调节蛋白激酶和核因子-κB活化对重症急性胰腺炎(severe acute pancreatitis,SAP)大鼠中性粒细胞凋亡延迟的调控作用。方法 60只雄性SD大鼠随机分为对照组、SAP组、吡咯烷二硫代氨基甲酸盐(pyrrolidine thiocarbamate,PDTC)组(CPTC组)和细胞外调节蛋白激酶特异性抑制剂PD98059组各15只,对照组大鼠仅作开腹再缝合,SAP组、PDTC组和PD98059组大鼠采用胆胰管逆行注射质量分数4%牛磺胆酸钠建立SAP模型,PDTC组和PD98059组大鼠造模前分别注射PDTC和PD98059,对照组和SAP组仅注射等量生理盐水。24h后于大鼠下腔静脉取血分离中性粒细胞,体外培养16h后采用流式细胞仪检测中性粒细胞凋亡率和Bcl-xl基因表达情况。结果体外培养16h后,SAP组中性粒细胞凋亡率明显低于对照组、PDTC组和PD98059组(P<0.01),Bcl-xl基因表达水平明显高于对照组、PDTC组和PD98059组(P<0.01);PDTC组和PD98059组中性粒细胞凋亡率低于对照组(P<0.01);PD98059组Bcl-xl基因表达水平高于对照组(P<0.01),PDTC组与对照组比较差异无统计学意义(P>0.05)。结论 SAP时大鼠中性粒细胞中Bcl-xl表达明显增加,导致其自发凋亡率减低,PD98059或PDTC可明显抑制Bcl-xl表达,恢复其自发凋亡率。
Objective To investigate the regulatory effect of extracellular regulated protein kinase and nuclear factor-κB activation on the apoptosis of neutrophils in rats with severe acute pancreatitis (SAP). Methods Sixty male Sprague Dawley rats were randomly divided into control group, SAP group, PDTC group (CPTC group) and PD98059 group (15 mg / kg) Only the control group rats were only used for laparotomy and resuscitation. SAP group, PDTC group and PD98059 group were injected with 4% sodium taurocholate into the biliary and pancreatic ducts to establish SAP model. PDTC group and PD98059 group PDTC and PD98059 were injected before model and normal saline was injected into control and SAP groups respectively. Neutrophils were isolated from vena cava of inferior vena cava 24 h later and cultured for 16 h. Flow cytometry was used to detect neutrophil apoptosis rate and Bcl-xl gene expression. Results The apoptosis rate of neutrophils in SAP group was significantly lower than that in control group, PDTC group and PD98059 group (P <0.01), and the expression of Bcl-xl gene was significantly higher than that in control group, PDTC group and PD98059 group P <0.01). The apoptosis rate of neutrophils in PDTC group and PD98059 group was lower than that in control group (P <0.01). The expression of Bcl-xl gene in PD98059 group was higher than that in control group (P <0.01) The difference was not statistically significant (P> 0.05). Conclusions The expression of Bcl-xl in rat neutrophils increased significantly during SAP, which led to the decrease of spontaneous apoptosis rate. PD98059 or PDTC could significantly inhibit the expression of Bcl-xl and restore its spontaneous apoptosis rate.