LIVER-LUNG INTERACTIONS IN ACUTE LUNG INJURY

我们已经证明，肺中环氧合酶（考克斯）基因的过表达可防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均表明，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。 需要更深入地了解与内毒素反应中的肝-肺相互作用相关的生化、分子和病理生理学事件，以确定潜在的干预措施，从而防止两个器官的反应。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伤。 直接作用主要是局部产生血栓素引起的血管收缩。 显著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（TNF α）介导的间接效应。 2)内皮衍生的前列腺素类，前列腺素E2（PGE 2），用于调节其产生的器官中的内毒素反应，而与循环血液中达到的浓度无关（“旁分泌”效应）。 3)使用质粒-阳离子脂质体递送系统，可以将考克斯转基因的表达靶向主要在肺中（静脉内递送）或主要在肝中（动脉内递送）的内皮细胞。 由此产生的PGE 2局部浓度增加将主要缓和内毒素对转染器官的影响。 我们已经开发了一种猪制剂，它允许肺和肝的单独或共同灌注。我们将利用这一准备工作来检验上述假设，具体目标如下：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、前列腺素类和TNF α的组织和灌注液浓度、支气管肺泡灌洗液（BAL）总细胞计数和分类细胞计数、前列腺素类和TNF α的BAL浓度、TNF α的表达的影响，考克斯-1和考克斯-2基因以及肺（和肝）中核因子κ B（NF κ B）的激活，在灌注回路中包括和不包括肝。 2)由于PGE 2在通过肺或肝的单次运输中完全从血液中清除，我们将通过将PGE 2注入传入血液供应来增加每个器官中PGE 2选择性的浓度，并确定对内毒素反应的影响。3)我们将确定考克斯转基因在与阳离子脂质体复合的质粒中通过静脉内、主动脉根或两种途径递送后表达的器官分布。 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种途径用考克斯基因体内转染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，这是重要的内毒素诱导的肺损伤，提供新的治疗干预措施的基础。

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.