PMN integrins, apoptosis and inflammation

描述(由申请人提供)： 中性粒细胞(PMN)参与炎症反应提供了抵御入侵生物的第一道防线。然而，炎症被称为“双刃剑”，因为PMN会对周围组织造成损害，而持续的炎症反应会导致各种病理情况。循环中的中性粒细胞重新聚集到炎症部位需要刺激中性粒细胞与内皮细胞的黏附，然后是它们的外渗和趋化作用。β2整合素、α-Lβ-2和α-Mβ-2介导了中性粒细胞与随后的渗出所需的内皮细胞的紧密黏附。尽管Beta2整合素已经得到了广泛的研究，但焦点一直集中在识别黏附所需的构象变化上，而负责调节Beta2介导的PMN黏附的细胞信号通路仍然知之甚少。我们已经确定了一条与PMN凋亡相关的信号通路，该通路可以减弱β2介导的PMN与肺微血管内皮细胞的黏附。刺激PMN Fas“Death”受体可减少PMN与肺微血管内皮细胞的黏附。初步数据表明，黏附的丧失是Fas/Cerama/Protein Kinase CDelta(PKCDelta)通路中的一个非常早期的事件，参与了PMN的凋亡。具体地说，Fas的激活增加了PMN神经酰胺的浓度，并导致PKC Delta的胞浆定位。我们认为，PKC Delta的胞浆定位可以减少黏附，因为它反对PKC Delta的膜定位，而PKC Delta是Beta2介导的黏附所必需的。目的1检测中性粒细胞表面介导内皮细胞黏附的两种β2整合素受体，α-Lβ2和αMβ2，以确定哪些β2受体受Fas激活的影响。目的2确定Fas激活对PKC Delta的影响，并操纵PKC Delta的细胞定位，以确定其对β2整合素和黏附的影响。目的3检测Fas对神经酰胺的影响，以证实神经酰胺可增加Fas对PKC Delta、β2整合素和黏附的影响。目的4确定通过操纵Fas/神经酰胺/PKC Delta途径减少黏附是否也能减轻体外大鼠肺缺血/再灌流所致的损伤。对这一途径的了解将为控制炎症提供新的靶点。

DESCRIPTION (provided by applicant): The participation of Polymorphonuclear neutrophils (PMN) in the inflammatory response provides a first line of defense against invading organisms. However, inflammation has been called "double edged sword" as PMN can cause damage to surrounding tissues and a prolonged inflammatory response can contribute to a variety of pathological conditions. The recruitment of circulating PMN to inflammatory sites requires the stimulated adhesion of PMN to the endothelium followed by their extravasation and chemotaxis. The beta2 integrins, alpha-L beta-2 and alpha-M beta-2 mediate the tight adhesion of PMN to the endothelial cells required for subsequent extravasation. Although the beta2 integrins have been widely studied, the focus has been on identifying conformational changes necessary for adhesion while the cellular signaling pathways responsible for regulating beta2 mediated adhesion of PMN remain poorly understood. We have identified a signaling pathway associated with PMN apoptosis that attenuates the beta2 mediated adhesion of PMN to pulmonary microvascular endothelial cells. Stimulation of the PMN Fas "death" receptor results in a reduction of PMN adhesion to pulmonary microvascular endothelial cells. Preliminary data indicates that the loss of adhesion is a very early event in the Fas/Ceramide/Protein Kinase Cdelta (PKCdelta) pathway that contributes to PMN apoptosis. Specifically, Fas activation increases PMN ceramide concentration and causes the cytosolic localization of PKCdelta. We propose that the cytosolic localization of PKCdelta could reduce adhesion because it opposes the membrane localization of PKCdelta that is necessary for beta2 mediated adhesion. Aim 1 examines the two beta2 integrin receptors on PMN, alpha-L beta-2 and alphaM beta2, that mediate endothelial cell adhesion to determine which of the beta2 receptors are affected by Fas activation. Aim 2 determines the effect of Fas activation on PKCdelta and manipulates the cellular localization of PKCdelta to determine its effects on the beta2 integrins and adhesion. Aim 3 examines the effects of Fas on ceramide to confirm that ceramide increases mediate Fas effects on PKCdelta, beta2 integrins, and adhesion. Aim 4 determines if reducing adhesion by manipulating the Fas/ceramide/PKCdelta pathway can also reduce ischemia/reperfusion-induced injury in an ex-vivo rat lung. An understanding of this pathway would provide novel targets for the control of inflammation.