SOC通道在循环纤维细胞参与哮喘气道重塑中的作用研究

Fibrocytes are bone marrow-derived mesenchymal cell precursors, which have been shown to be increased in peripheral blood and accumulated beneath the epithelial basement membrane of asthma patients, implicating that these cells play an important role in airway remodelig of chronic asthma. Store-operated calcium (SOC) channels are the major calcium influx pathway of blood cells including monocytes,macrophages and dendritic cells.We have previously demonstrated that SOC channels regualtes the function of monocytes-derived dendritic cells and macrophages, whether SOC channels have a similarly effect on the function of monicytes-derived fibrocytes is still unknown. Our previous study on airway smooth muscel cells showed that Th2 cytokine Il-13 promotes Ca2+ entry through SOC channels.Since Th2 cytokeines IL-4 and IL-13 promote, whereas Th1 cytokein INF-gamma inhibits the differentiation of fibrocytes，we hypothesized that SOC channels are involved in the functional regulation of circulating fibrocytes, and thus contribute to the subepithelial fibrosis in chronic asthma..In the present study,we will firstly illustrate the possible role of SOC channels in airway remodeling and the accumulation of fibrocytes beneath the epithelial basement membrane by using SOC blockers on chronic sensitized mice models. We will also compare the expression and activity of SOC channels in fibrocytes from normal controls and asthmatics; in addition, we will observe the effect of STIM1 and Orai1 knockdown,two essential component and regulating molecules of SOC channels, on the differentiation, proliferation, migration and secretion of extracellular matrix of fibrocytes.These expriments will elucidate the importance of SOC chanenls to biological processes of fibrocytes. Finally, we will figure out the modulating effect of Th1,Th2 and Th17 cytokines on SOC channels; the regulatory effect of serum amyloid P (SAP),a fibrocyte-inhibiting factor in plasma, on SOC channels will also be investigated .By these experiments, we will clarify the importance of SOC channels to fibrocytes and the subepithelial fibrosis of chronic asthma.

循环纤维细胞是慢性哮喘气道上皮下纤维化过程中成纤维细胞的重要来源。我们前期研究证实，钙库操纵的钙通道（SOC）是单核细胞来源的巨噬细胞和树突状细胞的重要钙信号机制，Th2细胞因子如IL-13对SOC通道功能具有调控作用；纤维细胞同样由单核细胞分化而来，IL-13也可以促进纤维细胞分化和胶原分泌；据此我们假设，SOC通道是纤维细胞分化、迁移和分泌等功能的重要调控机制，在纤维细胞募集至气道上皮下参与哮喘气道重塑中具有重要作用。本研究将首先建立慢性哮喘小鼠模型，从整体水平研究SOC通道抑制对气道气道上皮下纤维化和纤维细胞募集的作用；其次，从细胞水平证实SOC通道对纤维细胞分化、增殖、迁移以及胶原和细胞因子分泌等功能的调控作用；最后，探讨Th1、Th2相关细胞因子和纤维细胞抑制因子SAP对纤维细胞SOC通道的调控作用。这些研究将明确慢性气道炎症诱导纤维细胞的分化和迁移，促进气道重塑的离子通道基础。

循环纤维细胞（circulating fibrocytes， CF）参与了慢性哮喘患者的上皮下纤维化过程，是慢性哮喘气道炎症导致不可逆气流受限和气道重塑的重要机制，但CF如何参与这一过程尚未阐明。 我们的流式细胞术研究结果显示，哮喘患者外周血单个核细胞中CF比例明显高于正常对照患者，提示CF参与了哮喘发病过程；进一步研究发现，ATP可以诱导CF的钙释放和外钙内流，且这一作用可以被SOC通道阻断剂SKF和BTP-2抑制，提示CF功能性表达SOC通道； 体外培养的CF的分化受SOC通道阻断剂的影响，即非选择性的阻断剂SKF抑制CF的分化，而SOC相对选择性的阻断剂BTP-2可以部分促进CF的分化，提示这两种抑制剂存在功能差异；血清淀粉样蛋白P（SAP）是循环中天然存在的CF抑制因子，我们研究发现，SAP可以抑制SOC通道活性和CF的分化。在慢性哮喘小鼠模型研究发现，气道粘膜下CR45+ColI+ 循环纤维细胞比例高于正常对照，而孟鲁斯特处理可以明显减少哮喘小鼠的气道粘膜下纤维细胞比例。这些结果初步证实了循环纤维细胞参与了慢性哮喘的气道重塑过程，其机制可能与SOC通道介导的钙信号相关，而SAP可能用于抑制慢性气道重塑。