The role of syndecan-4 in the regulation of JNK activation in human neutrophils

描述（由申请人提供）：单核细胞募集到急性炎症区域是炎症过程的后期。我们之前已经证明，在lps诱导的急性炎症模型中，p38或c-Jun nh2末端激酶（JNK）的全身抑制限制了中性粒细胞向肺的募集。然而，正如我们将展示的，抑制JNK，而不是p38，限制了单核细胞的肺部募集，这是通过单核细胞化学吸引蛋白1 （MCP-1）和中性粒细胞依赖途径发生的。因此，了解导致中性粒细胞中JNK活化的机制将是理解单核细胞募集的基础。虽然中性粒细胞中p38的激活发生在悬浮条件下，但JNK的激活需要中性粒细胞在贴壁条件下培养，尽管对调节细胞粘附的细胞表面蛋白以及JNK的激活知之甚少。其中一个家族，syndecans，与细胞外基质（ECM）结合，诱导细胞粘附。此外，syndecan-4已被证明与CXCR4共同作用，我们将在这里展示与中性粒细胞中的IL-8受体CXCR2共同作用。我们拟通过检测syndecan-4是否通过上调PKC活性来增强JNK的激活，进而检测syndecan-4在调节il -8诱导的中性粒细胞JNK激活中的作用，从而检测MCP-1的表达，检测syndecan-4与CXCR2共关联的机制，以及这种共关联增强il -8诱导的JNK激活的必要性。我们将使用多种技术，包括化学抑制剂、阻断抗体、小干扰rna和中性粒细胞和PLB-985细胞系的蛋白质转导，这些细胞系可以分化为中性粒细胞样细胞。此外，为了证实中性粒细胞和PLB-985细胞的发现，我们将利用来自syndecan-4、CXCR2和PKC（缺陷）小鼠的中性粒细胞。利用上述技术，我们将研究我们假设调节syndecan-4诱导的PKC(和JNK激活的成分，包括PKC-（和PP2A），或调节syndecan-4与CXCR2的共关联，包括接头蛋白CASK和syntenin。这些研究将为中性粒细胞中JNK活化的调节机制提供重要的见解，并将为肺部炎症，特别是肺单核细胞募集的调节提供新的途径。

DESCRIPTION (provided by applicant): The recruitment of monocytes to areas of acute inflammation is a later stage in the inflammatory process. We have previously shown that systemic inhibition of either p38 or c-Jun NH2-terminal kinase (JNK) limits neutrophil recruitment to the lung in an LPS-induced model of acute inflammation. As we will show, however, inhibition of JNK, but not p38, limits the pulmonary recruitment of monocytes, which occurs through monocyte chemoattractant protein 1 (MCP-1) and neutrophil dependent pathways. Understanding mechanisms that lead to JNK activation in neutrophils will, therefore, be fundamental to the understanding of monocyte recruitment. While activation of p38 in neutrophils occurs in suspended conditions, activation of JNK requires neutrophils to be cultured in adherent conditions, although the cell surface proteins that regulate cell adhesion, and thereby JNK activation, are poorly understood. One such family, the syndecans, bind to the extracellular matrix (ECM) inducing cell adhesion. In addition, syndecan-4 has been shown to co-associate with CXCR4, and we will show here with the IL-8 receptor CXCR2 in neutrophils. We propose to examine the role of syndecan-4 in regulating IL-8-induced JNK activation in neutrophils, and thereby expression of MCP-1, by examining if syndecan-4 enhances JNK activation via an upregulation of protein kinase C alpha (PKC() activity, examine mechanisms for the co-association of syndecan-4 with CXCR2, and the necessity of the co-association for the enhancement of IL-8-induced JNK activation. We will use several techniques including chemical inhibitors, blocking antibodies, small interfering RNAs, and protein transduction of neutrophils and the PLB-985 cell line, which can be differentiated into neutrophil-like cells. In addition, to confirm our findings from neutrophils and PLB-985 cells, we will utilize neutrophils derived from syndecan-4, CXCR2, and PKC( deficient mice. Utilizing the above techniques we will examine components that we hypothesize to regulate syndecan-4-induced PKC( and JNK activation, including PKC-( and PP2A, or to regulate the co-association of syndecan-4 with CXCR2, including the adaptor proteins CASK and syntenin. These studies will provide vital insights into mechanisms that regulate JNK activation in neutrophils and will suggest roles for novel pathways in the regulation of lung inflammation, and in particular pulmonary monocyte recruitment.