摘要
目的通过检测抑／促凋亡调控基因Bcl-xL／Bax表达变化，研究大黄素（EMD）影响脓毒症患者中性粒细胞（PMN）凋亡的机制。方法采集5例健康志愿者及16例脓毒症患者PMN进行体外培养。健康志愿者的血标本作为对照组（HC组）；脓毒症患者分为脓毒症组（sE组，8例）、EMD干预脓毒症组（EMD-SE组）。采用反转录-聚合酶链反应（RT-PCR）及免疫细胞化学技术检测PMN抑凋亡基因Bcl-xL或促凋亡基因Bax的mRNA及蛋白表达水平的差异。结果sE组抑凋亡基因Bcl-xLmRNA[吸光度（4值）]表达和蛋白阳性率均较Hc组显著增加【Bcl-xLmRNA：0．98±0．12比0．32±0．07，Bcl-xL蛋白：87．5％（7／8）比20．0％（1／5），均P〈0．05]，而促凋亡基因Bax表达与HC组比较差异无统计学意义[BaxmRNA（A值）：1．13±0．08比1．28±0．13，Bax蛋白：62．5％（5／8）比80．0％（4／5），均P〉0．05]；EMD-SE组Bcl-xL表达均较sE组明显减少[Bcl-xLmRNA（A值）：0．39±0．06比0．98±0．12，Bcl-xL蛋白：25．0％（2／8）比87．5％（7／8），均P〈0．05]，EMD-SE组与sE组Bax表达差异均无统计学意义[BaxmRNA（A值）：1．19±0．11比1．13±0．08，Bax蛋白：87．5％（7／8）比62．5％（5／8），均P〉0．05]。SE组Bcl-xL／BaxmRNA明显高于HE组（0．87±0．07比0．25±0．04．P〈0．05），EMD-SE组Bcl-xL／BaxmRNA明显低于SE组（0．32±0．03比0．87±0．07，P〈0．05）。结论EMD可使脓毒症时增高的Bcl-xL／Bax比值下调，提示在EMD促PMN凋亡中抑／促凋亡调控基因Bcl-xL／Bax表达变化可能是其分子机制之一。
Objective By detecting expression changes of apoptotic gene Bcl-xL/Bax to investigate the molecular mechanisms of emodin （EMD） effect on apoptosis of polymorphonuclear neutrophil （PMN） in patients with sepsis of abdominal cavity infection. Methods Peripheral venous blood was collected from healthy volunteers and patients with sepsis. PMN was separated from peripheral blood and cultivated in vitro. The blood samples of healthy volunteers were recognized as control group （HC group）, and the patients with sepsis were divided into sepsis group （SE group） and EMD intervention sepsis group （EMD-SE group）. The changes of PMN mRNA and protein expressions of anti-/pro-apoptotic gene Bcl-xL/Bax were detected separately by using reverse transcription-polymerase chain reaction （RT-PCR） and immunocytochemistry. Results Compared with HC group, the mRNA [absorbance （A value）] and protein positive expression rate of anti-apoptotic gene Bcl-xL were significantly higher in SE group [Bcl-xL mRNA： 0.98 ± 0.12 vs. 0.32 ± 0.07, Bcl-xL protein positive expression rate： 87.5% （7/8） vs. 20.0% （1/5）, P 〈 0.05]; however, the mRNA and protein positive expression rate of Bax of the pro-apoptotic gene were not obviously different in SE group from those in the HC group [mRNA （A value）： 1.13 ± 0.08 vs. 1.28 ± 0.13, protein positive expression rate： 62.5% （5/8） vs. 80.0% （4/5）, both P 〉 0.05]. Compared with SE group, either the expression of Bcl-xL mRNA or protein positive expression rate in EMD-SE group was significantly decreased [Bcl-xL mRNA （A value）： 0.39 ± 0.06 vs. 0.98± 0.12, Bcl-xL protein positive expression rate： 25.0% （2/8） vs. 87.5% （7/8）, all P 〈 0.05]; however, the expression of either Bax mRNA or protein was not markedly different in EMD-SE group from either one of them in control group [Bax mRNA （A value）： 1.19± 0.11 vs. 1.13 ± 0.08, Bax protein positive expression rate： 87.5% （7/8） vs. 62.5% （5/8）, both P 〉 0.05]. The ratio of Bcl-xL/Bax mRNA in SE group was significantly higher than that in HC group, being 0.87 ± 0.07 vs. 0.25 ± 0.04; the ratio of Bcl-xL/Bax mRNA in EMD-SE group was obviously lower than that in SE group （0.32 ± 0.03 vs. 0.87 ± 0.07, both P 〈 0.05）. Conclusion EMD can down-regulate the elevated ratio of Bcl-xL/Bax in sepsis, suggesting that in the effect of EMD in pro-apoptosis of PMN, the expression changes of anti-/pro-apoptosis regulatory gene Bcl-xL/Bax might be possibly one of the molecular mechanisms.
出处
《中国中西医结合急救杂志》
CAS
北大核心
2016年第2期129-132,共4页
Chinese Journal of Integrated Traditional and Western Medicine in Intensive and Critical Care
基金
国家自然科学基金资助项目（2013A007）
