Lyso-PCs activate PMNs and endothelium via PKC

脂质在血液的常规储存过程中积累，并刺激多种中性粒细胞（PMN）功能，以及引起人肺微血管内皮细胞（HMVEC）的激活。流行病学研究表明，老年红细胞压积与急性肺损伤和多器官功能衰竭的发生独立相关。此外，我们已经将这些脂质与人输血相关的急性肺损伤（TRALI）联系起来，并提出了这种危及生命的疾病的两个事件模型，类似于急性呼吸窘迫综合征的病理生理学。在这两个事件模型中，患者的临床状态是第一个事件，输注生物活性脂质 是第二个事件。TRALI的动物模型已经证实，来自储存而非新鲜血液和血液成分的血浆和脂质会导致TRALI，并且体外模型已经证明了这种两次事件肺模型所需的大部分细胞生理学。这些事件包括以下要求：1）内皮活化（粘附分子和趋化因子释放的增加），导致PMN引发和粘附，随后2）这些粘附的“高反应性”PMN的活化，最终导致肺损伤。 溶血磷脂酰胆碱（lyso-PC）是在细胞血液组分中积累的主要生物活性物质，并且这些化合物通过细胞溶质钙的快速增加和蛋白激酶C（PKC）的活化来刺激PMN和HMVEC。我们假设储存血液中的脂质通过激活特异性的 PKC亚型刺激内皮床和/或激活隔离的中性粒细胞，导致急性肺损伤。这一假设将通过以下具体目标的完成来检验：1）确定溶血PC从其受体到其效应激酶的信号通路：2）证实PKC γ是溶血PC介导的信号通路中的效应激酶; 3）确定PKC激活在PMN和HMVEC中的作用，和4）使用临床测试的PKC抑制剂研究在充分描述的TRALI动物模型中PKC抑制的作用。这些特定目标的完成将可能导致体外和可能的临床干预的目标，以开发抑制或减弱这些脂质和脂质代谢产物的作用的方法。 最终使输血更安全。

Lipids accumulate during the routine storage of blood and stimulate multiple neutrophil (PMN) functions as well as causing activation of human pulmonary microvascular endothelial cells (HMVECs). Epidemiologic studies have demonstrated that older packed red blood cells are independently associated with the development of acute lung iinjury and multiple organ failure in injured aptients. Moreover, we have implicated these lipids in human transfusion related acute lung injury (TRALI) and have proposed a two event model for this life threatening illness, similar to the pathophysiology of the acute respiratory distress syndrome. In this two event model the clinical status of the patient is the first event and the infusion of biologically active lipids in stored blood components is the second event. An animal model of TRALI has verified that the plasma and lipids from stored, but not fresh, blood and blood components cause TRALI and an in vitro model has demonstrated much of the cellular physiology required for this two event lung model. These events include the requirements of 1) endothelial activation (increase in adhesion molecules and chemokine release) resulting in PMN priming and adherence followed by 2) activation of these adherent "hyperresponsive" PMNs, culminating in lung injury. Lysophosphatidylcholines (lyso-PCs) are the major biologically active species that accumulate in cellular blood components, and these compounds stimulate both PMNs and HMVECs through rapid increases in cytosolic calcium and activation of protein kinase C (PKC). We hypothesize that lipids from stored blood stimulate human PMNs and HMVECs through activation of specific PKC isoforms that stimulate endothelial beds and/or activate sequestered PMNs resulting in acute lung injury. This hypothesis will be tested through completion of the following specific aims: 1) to determine the signaling pathways of lyso-PCs from its receptor to its effector kinases; 2) to confirm the that PKC gamma is the effector kinase in lyso-PC mediated signaling; 3) to determine the role of PKC activation in PMNs and HMVECs, and 4) to investigate the effects of PKC inhibition in a well-described animal model of TRALI using a clinically tested PKC inhibitor. Completion of these specific aims will likely result in targets for in vitro, and possibly clinical, intervention to develop methods to inhibit or to attenuate the effects of these lipids and ultimately make transfusion safer.