重症急性胰腺炎与中性粒细胞凋亡蛋白质的相关性

Relationship between development and progression of severe acute pancreatitis and neutrophil apoptosis-related proteins in rats

目的:探讨重症急性胰腺炎(severe acute pancreatitis,SAP)的发生发展与中性粒细胞(polymorpho nuclear neutrophil,PMN)凋亡相关蛋白质是否相关.方法:SD大鼠60只,随机分为2组,每组30只,建立急性坏死性胰腺炎(acute necrotizing pancreatitis,ANP)组和假手术(sham-operated,SO)组模型,分别在制模后3、6、12h分批处死大鼠,抽取下腔静脉血密度梯度离心分离PMN,用流式细胞仪测定PMN凋亡比率,对胰腺组织进行病理评分.并对大鼠制模后12h的PMN裂解,应用Labelfree技术对两组PMN进行蛋白定量差异分析,进一步鉴定出与凋亡有关的差异蛋白质.同时收集我院2008-06/2012-06收治的28例SAP患者,随机分为治疗组(14例)和对照组(14例).对照组采用常规综合治疗,治疗组患者在常规综合治疗的基础上加用生长抑素(somatostatin,SST)持续静脉滴注.比较两组患者的平均腹痛持续时间、淀粉酶恢复时间、平均住院天数以及并发症发生率、转手术率、死亡率.分析凋亡相关蛋白质的抗凋亡机制及生长抑素在治疗SAP中的诱导凋亡作用,推断SAP的发生发展是否与凋亡相关蛋白质有关.结果:ANP组PMN凋亡延迟,与SO组各个时间点比较差异有统计学意义(均P<0.01);鉴定出与PMN凋亡有关的4种蛋白质:78kDa葡萄糖调节蛋白、RhoGTPases、L-乳酸脱氢酶A链和血红蛋白α2链(ANP组与SO组的比值分别为1.953614、3.526625、1.766764、0.609825),两组差异有统计学意义(P<0.05).治疗组患者的平均腹痛持续时间、淀粉酶恢复时间及平均住院天数明显短于对照组患者,两组比较差异有统计学意义(P值分别为0.041、0.001、0.000).治疗组患者的并发症发生率、转手术率和死亡率明显低于对照组患者,两组比较差异有统计学意义(P=0.022,0.029,0.029).结论:ANP时,PMN凋亡延迟,与凋亡有关的差异蛋白质可能参与了ANP外周血PMN凋亡延迟;应用生长抑素治疗SAP,能明显缩短患者的临床症状缓解时间,减少并发症,降低患者的死亡率;SAP的发生发展可能与凋亡相关蛋白质有关.

AIM：To investigate the relationship between the development and progression of severe acute pancreatitis（SAP） and apoptosis-related proteins in rats.METHODS：Sixty SD rats were randomly divided into two groups：acute necrotizing pancreatitis（ANP） group and sham-operated（SO） group（n=30 for each）.At 3,6,and 12 h after induction of ANP,the rats were sacrificed and blood samples were collected from the inferior vena cava.Density gradient centrifugation was conducted to separate polymorpho nuclear neutrophils（PMNs）,and PMN apoptosis was determined by flow cytometry.PMNs collected at 12 h were lysed,and label-free technology was used to identify apoptosis-related proteins.Twenty-eight SAP patients treated at our hospital from June 2008 to June 2012 were randomly divided into a treatment group and a control group（n=14 for each）.The control group underwent conventional treatment,while the treatment group was treated with conventional treatment plus continuous infusion of somatostatin.The mean duration of abdominal pain,amylase recovery time,length of hospital stay,and the incidence of complications,rate of conversion to surgery,and mortality were compared between the two groups.RESULTS：PMN apoptosis was significantly delayed in the ANP group compared to the SO group at all time points（all P〈0.01）.Four PMN apoptosis-related proteins were identified：78 KDa glucose-regulated protein,RhoGTPase,L-lactic acid dehydrogenase A chain,and hemoglobin α2 chain（ANP/SO ratios：1.953614,3.526625,1.766764,0.609825;all P〈0.05）.The mean duration of abdominal pain,amylase recovery time and length of stay were significantly shorter（P = 0.041,0.001,0.000）,and the incidence of complications,rate of conversion to surgery,and mortality were significantly lower in the treatment group than in the control group（P = 0.022,0.029,0.029）.CONCLUSION：PMN apoptosis delay in ANP may be mediated by apoptosis-related proteins.Somatostatin therapy can significantly shorten the duration of patient＇s clinical symptoms and reduce complications and mortality.