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

在BALB/c小鼠（8 - 12周龄）中通过盲肠结扎和穿孔（CLP）诱导多微生物脓毒症。AP-1的DNA结合活性，通过电泳迁移率变动分析评估，在CLP诱导的脓毒症发作后的小鼠肺组织中呈时间依赖性增加。通过体内转染AP-1诱饵寡核苷酸（ODN）有效地消除了这种增加。脓毒症诱导显著增加脓毒症后肺和主动脉组织中死亡受体如TNF-R1、Fas、DR4和DR5的表面表达。此外，在脓毒症诱导后，FADD的基因和蛋白水平增加，FADD将半胱氨酸天冬氨酸蛋白酶原-8募集到死亡诱导信号复合物中。这些脓毒症诱导的变化被消除全身应用AP-1诱饵ODNs。TUNEL分析显示，在脓毒症小鼠的肺和主动脉组织切片中细胞凋亡的显着外观被阻止与AP-1诱饵ODNs的全身治疗。当静脉注射10mg/kg脂多糖（LPS）诱导小鼠内毒素休克时，IL-1R、IL-6R、HMGB-1和gp130等炎症分子的表达水平显著升高，而AP-1 decoy ODN可显著抑制这些炎症分子的表达。所有接受LPS的动物在48小时内死亡，并且在LPS后用AP-1诱饵ODNs处理的动物表现出显著的存活改善。我们的研究结果表明，AP-1诱饵ODN治疗是一种有效的策略，在脓毒症的治疗。此外，全身施用靶向FADD的siRNA，发现FADD被AP-1反式激活，防止了CLP小鼠中急性肺损伤的发展，并改善了它们的存活率。这些发现表明FADD siRNA对于脓毒症基因治疗的潜在用途。

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.