Choudhry Mashkoor A, Ren Xiangping, Romero Adriana, Kovacs Elizabeth J, Gamelli Richard L, Sayeed Mohammed M
Center for Surgical Research and Department of Surgery, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
J Surg Res. 2004 Sep;121(1):62-8. doi: 10.1016/j.jss.2004.02.013.
Recent studies from our laboratory have suggested that acute alcohol ingestion prior to burn injury enhances gut bacterial translocation by suppressing T cell-mediated intestinal immune defense. To determine the mechanism responsible for suppressed T cell function, we examined the activation of mitogen-activated protein kinases (MAPK) members p-38 and ERK-1/2. Both p-38 and ERK-1/2 are known to play a significant role in the T cell proliferation and their cytokine production. Rats were gavaged with ethanol to achieve a blood alcohol level of approximately 100 mg/dl, before they were subjected to a 25% total body surface area burn injury. Two days after injury, rats were sacrificed and mesenteric lymph node (MLN) T cells were isolated and their ability to proliferate in response to anti-CD3 was determined. For p-38 and ERK-1/2 determination, T cells were divided into two groups. Cells in one group were stimulated with anti-CD3 for 3 min and lysed. The cells in the second group were cultured for approximately 18 h in the presence of anti-CD3 and lysed. MAPK status in 18-h cultured cells allowed us to determine whether or not the changes in p-38 and ERK-1/2 are transient or persist in the proliferating cells. Two days after injury, anti-CD3-mediated MLN T cell proliferation was more suppressed in rats gavaged with alcohol prior to burn injury compared to rats receiving either burn injury alone or sham-injured rats regardless of their exposure. Western blot analyses showed significant inhibition of ERK-1/2 phosphorylation in both freshly isolated and 18-h cultured T cells from alcohol and burn-injured rats compared to the sham rat T cells. The inhibition of p-38 phosphorylation in T cells derived from alcohol and burn-injured rats was found to be transient as no significant difference in p-38 phosphorylation was noted between the 18 h incubated MLN T cells of sham and alcohol and burn-injured rats. Taken together, our findings suggest that low levels of ERK-1/2 activation is likely to play a significant role in MLN T cell proliferative suppression in alcohol and burn-injured rats.
我们实验室最近的研究表明，烧伤前急性摄入酒精会通过抑制T细胞介导的肠道免疫防御增强肠道细菌易位。为了确定T细胞功能受抑制的机制，我们检测了丝裂原活化蛋白激酶（MAPK）成员p-38和ERK-1/2的激活情况。已知p-38和ERK-1/2在T细胞增殖及其细胞因子产生中起重要作用。在大鼠遭受25%体表面积烧伤之前，用乙醇灌胃使其血液酒精水平达到约100mg/dl。受伤两天后，处死大鼠，分离肠系膜淋巴结（MLN）T细胞，并测定其对抗CD3刺激的增殖能力。为了检测p-38和ERK-1/2，将T细胞分为两组。一组细胞用抗CD3刺激3分钟后裂解。第二组细胞在抗CD3存在的情况下培养约18小时后裂解。18小时培养细胞中的MAPK状态使我们能够确定p-38和ERK-1/2的变化是短暂的还是在增殖细胞中持续存在。受伤两天后，与仅接受烧伤或假手术的大鼠相比，烧伤前用酒精灌胃的大鼠中抗CD3介导的MLN T细胞增殖受到更明显的抑制，无论它们是否暴露于酒精。蛋白质印迹分析显示，与假手术大鼠的T细胞相比，来自酒精和烧伤大鼠的新鲜分离及18小时培养的T细胞中ERK-1/2磷酸化均受到显著抑制。发现来自酒精和烧伤大鼠的T细胞中p-38磷酸化的抑制是短暂的，因为在假手术组与酒精和烧伤大鼠组培养18小时的MLN T细胞之间，p-38磷酸化没有显著差异。综上所述，我们的研究结果表明，低水平的ERK-1/2激活可能在酒精和烧伤大鼠的MLN T细胞增殖抑制中起重要作用。
