Novel anti-inflammatory mesothelial signaling to the spleen

项目摘要/摘要 脓毒症是一种复杂的致命性疾病，其定义为“由器官调节失调引起的危及生命的器官功能障碍”。 宿主对感染的反应“。在美国，脓毒症与每年约100万次住院有关，而发病率 脓毒症的发病率继续上升。脓毒症患者的死亡率仍然高得令人无法接受。新奇数据 从我们的实验室最近证明，简单地移动脾到中线，然后再回到它的 原位置导致脾包膜纤维化和免疫细胞的促炎性转移 脾的表型。虽然在老鼠身上也有类似的发现，但目前还不清楚这些 手法促进了促炎表型的形成。我们的新数据表明，特化间皮细胞 形成与脾被膜的连接可能通过以下途径向脾传递补益抗炎信号 通过α7烟碱受体传递胆碱能信号。除了胆碱能信号的标志物，这些 间皮细胞强烈表达神经元标志物，胞浆内可见神经元样细胞器。 由于连接本身是由胶原和间皮细胞组成的，所以脾被膜纤维化 在这些连接中断之后可能代表着试图重新建立 间皮细胞对细胞的信号转导。这项建议的主要目标是确定胆碱能信号 通过特殊的间皮细胞确实将强健的抗炎信号传递到脾胶囊 以及该信号的中断是否具有功能性反应，从而加剧全身炎症 对内毒素的反应。因此，Aim 1将测试外科手术是否破坏了间皮细胞向脾发出的信号 胶囊具有持久的抗炎作用，并加剧了对内毒素的炎症反应。 我们将通过侧腹切开进入大鼠的脾，并清除脾中的间皮连接，而不是 干扰脾。基线免疫谱(流式细胞仪/细胞因子分析)和生理学 麻醉破坏和假手术大鼠对实验性炎性休克(内毒素)的反应 在外科手术切断间皮连接后24小时、21天和3个月确定 脾目标2将测试间皮细胞之间的非神经性胆碱能信号是否介导强直性 在脾的抗炎反应‘。α7nAChR在小鼠体内的全局缺失导致促炎反应 然而，调节这一效应的细胞类型(S)仍不清楚。我们将使用Wilms Tumor 1 同源基因启动子Cre和chrna 7tm1使小鼠在间皮细胞中特异性地“敲除”α7nAChR。 我们预测，α7nAChR在间皮细胞中的特异性缺失将促进促炎表型 在脾里。如果我们的假设是正确的，通过间皮细胞在远端部位之间的神经元样信号 代表了一种以前未被认识到的生物交流途径，很可能有一个主要的 不仅影响我们对脓毒症中调节全身免疫反应的因素的理解，还影响到 特发性纤维化累及间皮的其他器官。

Project Summary/Abstract Sepsis is a complex lethal condition defined as “life- threatening organ dysfunction caused by a dysregulated host response to infection”. In the U.S, sepsis is related to ~1 million hospitalizations annually, and the incidence of sepsis continues to rise. The mortality rate from in patients with sepsis remains unacceptably high. Novel data from our laboratory recently demonstrated that simply moving the spleen to midline and then returning it to its original position causes development of splenic capsular fibrosis and a pro-inflammatory shift in the immune cell phenotype of the spleen. While similar findings have been reported in mice, it remains unclear how these maneuvers promote a pro-inflammatory phenotype. Our novel data indicate that specialized mesothelial cells that form connections to the splenic capsule may transmit tonic anti-inflammatory signals to spleen via cholinergic signaling through alpha 7 nicotinic receptors. In addition to markers of cholinergic signaling, these mesothelial cells strongly express neuronal markers and display neuronal-like organelles within their cytoplasm. As the connections themselves are made of collagen lined with mesothelial cells, splenic capsular fibrosis following disruption of these connections may represent a physiological response to try to re-establish mesothelial cell to cell signaling. The primary goal of this proposal is to determine whether cholinergic signaling through specialized mesothelial cells does indeed transmit tonic anti-inflammatory signals to the splenic capsule and whether disruption of this signaling has a functional response, exacerbating the systemic inflammatory response to endotoxin. As such, Aim 1 will test whether `Surgical disruption of mesothelial signaling to the splenic capsule results in a prolonged anti-inflammatory profile and exacerbated inflammatory response to endotoxin'. We will access the spleen in rats via a flank incision and clear the spleen of mesothelial connections without interfering with the spleen. Baseline immune profiles (flow cytometry/cytokine analysis) and the physiological response to experimental inflammatory shock (endotoxin) in anesthetized disrupted and sham surgical rats will be determined 24 hours, 21 days and 3 months following surgical disruption of mesothelial connections to the spleen Aim 2 will test whether `Non-neuronal cholinergic signaling between mesothelial cells mediates a tonic anti-inflammatory response in the spleen'. Global deletion of α7nAChR in mice results in a pro-inflammatory splenic phenotype however, the cell type(s) mediating this effect remain unclear. We will use Wilms tumor 1 homolog gene promoter Cre and Chrna7tm1 floxed mice to `knockout' α7nAChR specifically in mesothelial cells. We predict that loss of the α7nAChR specifically in mesothelial cells will promote a pro-inflammatory phenotype in the spleen. If our hypothesis is correct, neuronal like signaling between distal sites through mesothelial cells would represent a previously unrecognized pathway of biological communication and would likely have a major impact not only on our understanding of the factors that regulate systemic immune responses in sepsis, but also idiopathic fibrosis of other organs involving the mesothelium.