LIVER-LUNG INTERACTIONS IN ACUTE LUNG INJURY

我们已经证明，环氧合酶(COX)基因在肺中的高表达可以阻止内毒素对实验动物肺的直接影响。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，对肝脏的影响对于内毒素诱导的肺损伤的充分表达具有重要意义。需要更彻底地了解内毒素反应中与肝-肺相互作用相关的生化、分子和病理生理事件，以便确定可能的干预措施，以防止这两个器官的反应。我们提出以下假设：1)内毒素通过直接和间接作用对肺造成损害。直接影响主要是局部产生的血栓烷引起的血管收缩。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因子α(TNFpha)所介导的间接影响。2)内皮源性前列腺素E_2(PGE_2)可调节产生前列腺素E_2的器官的内毒素反应，与循环血液中的浓度无关(“旁分泌”效应)。3)利用质粒-阳离子脂质体递送系统，可以将COX转基因靶向主要在肺内皮细胞(静脉递送)或主要在肝脏(经动脉递送)的内皮细胞表达。由此产生的局部PGE2浓度的增加将主要缓解内毒素对转基因器官的影响。我们已经开发出一种猪制剂，它允许肺和肝脏分开或共同灌流。我们将利用这项准备来验证上述假设，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、组织和灌流液中前列腺素和肿瘤坏死因子α的浓度、支气管肺泡灌洗液中细胞总数和分类计数、前列腺素和肿瘤坏死因子α的浓度、肺(和肝)中肿瘤坏死因子α、环氧合酶-1和环氧合酶-2基因的表达以及核因子-kappaB(NFkappaB)激活的影响。2)由于PGE2在通过肺或肝脏的一次转运中从血液中完全清除，我们将通过将PGE2注入传入血液供应来增加每个器官中PGE2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转基因基因在与阳离子脂质体形成复合体的载体中传递后，确定其表达的器官分布。在记录了器官特异性转基因表达后，我们将确定先前通过每条途径与COX基因体内转导对内毒素反应的影响。这些研究将进一步阐明内毒素性肺损伤中重要的肝肺相互作用机制，为新的治疗干预措施提供基础。

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.