LIVER-LUNG INTERACTIONS IN ACUTE LUNG INJURY

我们已经证明，高表达的环加氧酶（COX）基因在肺阻止内毒素对肺的直接影响的实验动物。然而，内毒素的直接影响只是故事的一部分。临床经验和动物研究都表明，内毒素对肝脏的影响对于内毒素诱导的肺损伤的充分表达至关重要。我们需要对内毒素反应中与肝-肺相互作用相关的生化、分子和病理生理事件有更深入的了解，以便确定可能阻止这两个器官反应的干预措施。我们提出以下假设：1)内毒素对肺有直接和间接的损伤作用。直接作用主要是局部生成血栓素导致血管收缩。明显的肺部炎症和肺血管通透性增加是由肝脏产生肿瘤坏死α (TNFalpha)介导的间接效应。2)内皮来源的类前列腺素，前列腺素E2 (PGE2)，作用于调节其产生器官的内毒素反应，而不依赖于循环血液中的浓度（“旁分泌”效应）。3)使用质粒阳离子脂质体给药系统，可以将COX转基因靶向表达到内皮细胞，主要是肺（静脉给药）或肝脏（动脉给药）。由此产生的PGE2局部浓度的增加将主要调节内毒素对转染器官的影响。我们已经开发了一种猪制剂，允许肺和肝脏分别或共同灌注。我们将利用这一准备来检验上述假设，具体目的如下：1)我们将确定内毒素血症对肺血管阻力、肺血管通透性、肺含水量、前列腺素和TNFalpha的组织和灌注浓度、支气管肺泡灌洗（BAL）总细胞和分化细胞计数、前列腺素和TNFalpha的BAL浓度、TNFalpha、COX-1和COX-2基因的表达以及肺（和肝脏）中核因子κ B （NFkappaB）的激活的影响，无论是否将肝脏纳入灌注回路。2)由于PGE2在通过肺部或肝脏的单次运输中完全从血液中清除，我们将通过将PGE2输注到传入血液供应中来增加PGE2在每个器官中的选择性浓度，并确定对内毒素反应的影响。3)我们将通过静脉、主动脉根部或两种途径将COX转基因基因通过与阳离子脂质体结合的质粒传递给主动脉根部，从而确定COX转基因基因在器官中的表达分布。在记录了器官特异性转基因表达后，我们将确定通过每种途径预先在体内转染COX基因对内毒素反应的影响。这些研究将进一步阐明肝-肺相互作用的机制，这对内毒素诱导的肺损伤具有重要意义，为新的治疗干预措施提供依据。

We have shown that hyperexpression of the cyclooxygenase (COX) gene in the lungs prevents the direct effects of endotoxin on the lungs of experimental animals. However, direct effects of endotoxin are only part of the story. Both clinical experience and animal studies implicate effects on the liver as important for the full expression of endotoxin induced lung injury. A more thorough understanding of the biochemical, molecular and pathophysiological events related to liver-lung interactions in the endotoxin response is needed in order to identify potential interventions which would prevent the response in both organs. We propose the following hypotheses: 1) Endotoxin injures the lungs by both direct and indirect effects. The direct effects are mainly vasoconstriction resulting from local generation of thromboxane. Marked lung inflammation and increased pulmonary vascular permeability are indirect effects mediated by production of tumor necrosis alpha (TNFalpha) by the liver. 2) The endothelial derived prostanoid, prostaglandin E2 (PGE2), acts to modulate endotoxin responses in the organ where it is produced, independent of concentrations achieved in circulating blood (a "paracrine" effect). 3) Using a plasmid-cationic liposome delivery system, it is possible to target expression of a COX transgene to endothelial cells mainly in the lungs (intravenous delivery) or mainly in the liver (intra-arterial delivery). The resulting increased local concentrations of PGE2 will moderate principally the endotoxin effects on the transfected organ. We have developed a swine preparation which permits separate or common perfusion of the lungs and the liver. We will use this preparation to test the above hypotheses with the following specific aims: 1) We will determine effects of endotoxemia on pulmonary vascular resistance, lung vascular permeability, lung water content, tissue and perfusate concentrations of prostanoids and TNFalpha, bronchoalveolar lavage (BAL) total and differential cell counts, BAL concentrations of prostanoids and TNFalpha, expression of TNFalpha, COX-1 and COX-2 genes and activation of nuclear factor kappa B (NFkappaB) in the lungs (and liver) with and without inclusion of the liver in the perfusion circuit. 2) Since PGE2 is completely cleared from the blood in a single transit through the lungs or liver, we will increase concentrations of PGE2 selectivity in each organ by infusing PGE2 into the afferent blood supply and determine effects on the endotoxin response. 3) We will determine organ distribution of expression of a COX transgene following delivery of the gene in a plasmid complexed to cationic liposomes either intravenously, into the aortic root or by both routes. Having documented organ- specific transgene expression, we will determine effects of prior in vivo transfection by each route with the COX gene the endotoxin response. These studies will further clarify mechanisms of liver-lung interactions which are important to endotoxin induced lung injury, providing a basis for new therapeutic interventions.