Vilatoba Mario, Eckstein Christopher, Bilbao Guadalupe, Smyth Cheryl A, Jenkins Stacie, Thompson J Anthony, Eckhoff Devin E, Contreras Juan L
Division of Transplantation and Transplant Center, Department of Surgery, University of Alabama at Birmingham, USA.
Surgery. 2005 Aug;138(2):342-51. doi: 10.1016/j.surg.2005.04.019.
Evidence is emerging that the endoplasmic reticulum (ER) participates in initiation of apoptosis induced by the unfolded protein response and by aberrant Ca(++) signaling during cellular stress such as ischemia/reperfusion injury (I/R injury). ER-induced apoptosis involves the activation of caspase-12 and C/EBP homologous protein (CHOP), and the shutdown of translation initiated by phosphorylation of eIF2alpha. Sodium 4-phenylbutyrate (PBA) is a low molecular weight fatty acid that acts as a chemical chaperone reducing the load of mutant or unfolded proteins retained in the ER during cellular stress and also exerting anti-inflammatory activity. It has been used successfully for treatment of urea cycle disorders and sickle cell disease. Thus, we hypothesized that PBA may reduce ER-induced apoptosis triggered by I/R injury to the liver.
Groups of male C57BL/6 mice were subjected to warm ischemia (70% of the liver mass, 45 minutes). Serum aspartate aminotransferase was assessed 6 hours after reperfusion; apoptosis was evaluated by enzyme-linked immunosorbent assays of caspase-12 and plasma tumor necrosis factor alpha, Western blot analyses of eIF2alpha, and reverse transcriptase-polymerase chain reaction of CHOP expression.
A dose-dependent decrease in aspartate aminotransferase was demonstrated in mice given intraperitoneal PBA (1 hour before and 12 hours after reperfusion), compared with vehicle-treated controls; this effect was associated with reduced pyknosis, parenchymal hemorrhages, and neutrophil infiltrates in PBA-treated mice, compared with controls. In a lethal model of total liver I/R injury, all vehicle-treated controls died within 3 days after reperfusion. In contrast, 50% survival (>30 days) was observed in animals given PBA. The beneficial effects of PBA were associated with a greater than 45% reduction in apoptosis, decreased ER-mediated apoptosis characterized by significant reduction in caspase-12 activation, and reduced levels of both phosphorylated eIF2alpha and CHOP. Significant reductions in plasma levels of tumor necrosis factor alpha and liver myeloperoxidase content were demonstrated after PBA treatment.
Reduction in ER stress-induced hepatocellular injury was achieved by the administration of PBA. Targeting the ER-associated cell death pathway might offer a novel approach to reduce I/R injury to the liver.
越来越多的证据表明，内质网（ER）在细胞应激（如缺血/再灌注损伤，I/R损伤）期间，参与由未折叠蛋白反应和异常Ca(++)信号诱导的细胞凋亡的起始过程。内质网诱导的细胞凋亡涉及半胱天冬酶-12（caspase-12）和C/EBP同源蛋白（CHOP）的激活，以及由真核生物翻译起始因子2α（eIF2α）磷酸化引发的翻译终止。4-苯丁酸钠（PBA）是一种低分子量脂肪酸，作为一种化学伴侣，可减少细胞应激期间内质网中滞留的突变或未折叠蛋白的负荷，并具有抗炎活性。它已成功用于治疗尿素循环障碍和镰状细胞病。因此，我们推测PBA可能减少由肝脏I/R损伤引发的内质网诱导的细胞凋亡。
将雄性C57BL/6小鼠分组，进行温缺血（肝脏质量的70%，45分钟）。再灌注6小时后评估血清天冬氨酸转氨酶；通过半胱天冬酶-12的酶联免疫吸附测定、血浆肿瘤坏死因子α、eIF2α的蛋白质印迹分析以及CHOP表达的逆转录聚合酶链反应来评估细胞凋亡。
与给予赋形剂处理的对照组相比，腹腔注射PBA（再灌注前1小时和再灌注后12小时）的小鼠中天冬氨酸转氨酶呈剂量依赖性降低；与对照组相比，这种作用与PBA处理的小鼠中核固缩、实质出血和中性粒细胞浸润减少有关。在全肝I/R损伤的致死模型中，所有给予赋形剂处理的对照组在再灌注后3天内死亡。相比之下，给予PBA的动物中观察到50%的存活率（>30天）。PBA的有益作用与细胞凋亡减少超过45%、内质网介导的细胞凋亡减少（以半胱天冬酶-12激活显著降低为特征）以及磷酸化eIF2α和CHOP水平降低有关。PBA处理后，血浆肿瘤坏死因子α水平和肝脏髓过氧化物酶含量显著降低。
给予PBA可减轻内质网应激诱导的肝细胞损伤。针对内质网相关的细胞死亡途径可能为减轻肝脏I/R损伤提供一种新方法。
