The transcription factor AP-1 as a molecular target in sepsis therapy

采用盲肠结扎穿刺法（CLP）诱导BALB/c小鼠（8-12周龄）多微生物脓毒症。电泳迁移率转移测定显示，在clp诱导的脓毒症发作后，小鼠肺组织中AP-1的DNA结合活性呈时间依赖性增加。这种增加可通过体内转染AP-1寡核苷酸诱饵（odn）有效消除。脓毒症诱导显著增加脓毒症后肺和主动脉组织中TNF-R1、Fas、DR4、DR5等死亡受体的表面表达。此外，在脓毒症诱导后，FADD的基因和蛋白水平升高，FADD将procaspase-8招募到诱导死亡的信号复合体中。这些败血症引起的改变可通过全身应用AP-1诱骗odn来消除。TUNEL实验显示，AP-1诱骗ODNs系统治疗可防止脓毒症小鼠肺和主动脉组织切片中出现明显的细胞凋亡。静脉注射10mg/kg脂多糖（LPS）诱导小鼠内毒素休克时，炎症分子IL-1R、IL-6R、HMGB-1、gp130的表达水平显著升高，AP-1诱饵ODN处理可显著抑制炎症分子表达水平。所有接受LPS治疗的动物均在48小时内死亡，LPS治疗后给予AP-1诱饵odn治疗的动物存活率显著提高。我们的研究结果表明，AP-1诱骗ODN治疗是治疗败血症的有效策略。此外，系统给药靶向FADD的siRNA（被发现被AP-1反激活）可以阻止CLP小鼠急性肺损伤的发生，并提高其生存率。这些发现表明FADD siRNA在脓毒症基因治疗中的潜在作用。

Polymicrobial sepsis was induced by cecal ligation and puncture (CLP) in BALB/c mice (8-12 wk of age). The DNA binding activity of AP-1, as assessed by electrophoretic mobility shift assay, was time-dependently increased in lung tissues from mice after the onset of CLP-induced sepsis. This increase was effectively eliminated by in vivo transfection of AP-1 decoy oligonucleotides (ODNs). Sepsis induction significantly increased surface expression of death receptors, such as TNF-R1, Fas, DR4, and DR5, in lung and aortic tissues after sepsis. Furthermore, the gene and protein levels of FADD, which recruits procaspase-8 into the death-inducing signaling complex, were increased after sepsis induction. These sepsis-induced changes were eliminated by systemic application of AP-1 decoy ODNs. TUNEL assays revealed that the significant appearance of cell apoptosis in lung and aortic tissue sections from septic mice was prevented by systemic treatment with AP-1 decoy ODNs. When endotoxic shock was induced by an intravenous injection of 10mg/kg lipopolysaccharide (LPS) in mice, expression levels of inflammatory molecules, including IL-1R, IL-6R, HMGB-1, and gp130, were highly increased, which was significantly inhibited by AP-1 decoy ODN treatment. All animals which received LPS died within 48h, and the animals that were treated with AP-1 decoy ODNs after LPS exhibited a striking improvement of survival. Our results suggest that AP-1 decoy ODN therapy represent an effective strategy in the treatment of sepsis. In addition, systemic administration of siRNA targeting FADD, which was found to be transactivated by AP-1, prevented the development of acute lung injury in CLP mice, and improved their survival. These findings indicate the potential usefulness of FADD siRNA for gene therapy of the septic syndrome.