Intestinal epithelial apoptosis initiates gut mucosal injury during extracorporeal membrane oxygenation in the newborn piglet.
Intestinal epithelial apoptosis initiates gut mucosal injury during extracorporeal membrane oxygenation in the newborn piglet.
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DOI：
10.1038/labinvest.2013.149
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发表时间：
2014-02
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在体外膜氧合（ECMO）和心肺转流（CPB）期间暴露于体外循环的新生儿和幼儿有发生全身炎症反应综合征（SIRS）伴多器官功能障碍的风险。我们使用ECMO的仔猪模型来研究上皮细胞凋亡是ECMO相关肠损伤期间绒毛损伤之前的早期事件的假设。健康的3周龄仔猪接受ECMO长达8小时。通过组织病理学分析、核成像和末端脱氧核苷酸转移酶dUTP缺口末端标记来测量上皮细胞凋亡。采用酶免疫法测定血浆脂肪酸结合蛋白（I-FABP）水平。通过荧光辅助细胞分选分离肠肥大细胞。通过免疫组化、Western印迹和逆转录-定量聚合酶链反应研究了切割型caspase-8、caspase-9、磷酸化p38 MAPK和Fas配体的表达。仔猪ECMO与肠上皮细胞凋亡增加相关。ECMO 2 h后，绒毛尖端和散在的隐窝细胞中观察到广泛的凋亡变化。8h后，绒毛被剥脱，大多数隐窝细胞出现明显的凋亡变化。循环中I-FABP水平的增加是肠上皮损伤的标志，表明ECMO期间发生了上皮损伤。我们在上皮细胞中检测到增加的裂解半胱天冬酶-8，但没有裂解半胱天冬酶-9，表明外源性凋亡途径是活跃的。ECMO与肠道肥大细胞中Fas配体表达增加相关，这是通过激活p38丝裂原活化蛋白激酶诱导的。在ECMO的仔猪模型中，上皮细胞凋亡是启动肠粘膜损伤的早期事件。
Neonates and young infants exposed to extracorporeal circulation during extracorporeal membrane oxygenation (ECMO) and cardiopulmonary bypass (CPB) are at risk of developing a systemic inflammatory response syndrome (SIRS) with multi-organ dysfunction. We used a piglet model of ECMO to investigate the hypothesis that epithelial apoptosis is an early event that precedes villous damage during ECMO-related bowel injury. Healthy 3-week-old piglets were subjected to ECMO for up to 8h. Epithelial apoptosis was measured in histopathological analysis, nuclear imaging, and terminal deoxynucleotidyl transferase dUTP nick end labeling. Plasma intestinal-fatty acid-binding protein (I-FABP) levels were measured by enzyme immunoassay. Intestinal mast cells were isolated by fluorescence-assisted cell sorting. Cleaved caspase-8, caspase-9, phospho-p38 MAPK, and fas ligand expression was investigated by immunohistochemistry, Western blots, and reverse transcriptase-quantitative polymerase chain reaction. Piglet ECMO was associated with increased gut epithelial apoptosis. Extensive apoptotic changes were noted on villus tips and in scattered crypt cells after 2h of ECMO. After 8h, the villi were denuded and apoptotic changes were evident in a majority of crypt cells. Increased circulating I-FABP levels, a marker of gut epithelial injury, showed that epithelial injury occurred during ECMO. We detected increased cleaved caspase-8, but not cleaved caspase-9, in epithelial cells indicating that the extrinsic apoptotic pathway was active. ECMO was associated with increased fas ligand expression in intestinal mast cells, which was induced through activation of the p38 mitogen-activated protein kinase. Epithelial apoptosis is an early event that initiates gut mucosal injury in a piglet model of ECMO.