Shh信号通路对肺CD45-/CD31-/Sca-1+细胞的调控及其在COPD发病中作用的研究

Lung epithelial stem/progenitor cells play critical role in maintenance and repair of lung epithelium. Sonic hedgehog (Shh) signaling pathway is essential for stem cell renewal and maintenance in many tissues and organs in adult. In our preliminary study, we have isolated and identified lung CD45-/CD31-/Sca-1+ cells, which comprise a heterogeneous population exhibiting some progenitor properties. And we've also detected that mild expansion of lung CD45-/CD31-/Sca-1+ cells in mice is induced by cigarette smoking, but it is inadequate to recover the injured lung epithelium. To date, the underlying mechanisms that Shh signaling pathway mediates the functions of lung CD45-/CD31-/Sca-1+ cells to affect the balance between injury and repair of epithelium and to participate in the pathogenesis of chronic obstructive pulmonary disease (COPD) has not been elucidated. In this study, we will use recombinant Shh and Shh inhibitor to treat cigarette smoke extract (CSE) stimulated lung CD45-/CD31-/Sca-1+ cells and cigarette smoke (CS) induced COPD mouse model, and revaluate the results by flow cytometry, magnetic activated cell sorting and immunochemistry. Hopefully, through this research, we would give an insight into the pathogenesis of COPD and explore a new therapeutic target for it.

肺上皮干/祖细胞直接参与受损肺上皮的修复，Sonic hedgehog（Shh）信号通路参与多种组织器官内干/祖细胞的维持。我们的前期研究发现肺CD45-/CD31-/Sca-1+细胞是一个异质肺上皮干/祖细胞群，香烟烟雾能够刺激该细胞群扩增，但其扩增程度不能完全再生受损的肺上皮。Shh信号通路是否调控肺CD45-/CD31-/Sca-1+细胞，香烟烟雾是否通过Shh信号通路影响肺CD45-/CD31-/Sca-1+细胞功能，致肺上皮损伤/修复失衡，从而参与COPD的发病目前尚无报道。本研究拟通过干预肺CD45-/CD31-/Sca-1+细胞和吸烟COPD小鼠模型中的Shh信号通路，采用流式细胞术和免疫磁珠细胞分选等技术首次探讨香烟烟雾通过抑制Shh信号通路，导致肺CD45-/CD31-/Sca-1+细胞功能障碍，影响肺上皮的修复再生，为阐明COPD发病机制及探索其治疗新途径提供理论依据。

研究表明吸烟可能通过肺上皮干/祖细胞的增殖/修复失衡参与COPD疾病的发生与发展，但具体机制尚不明确，而肺CD45-/CD31-/Sca-1+是一个异质肺上皮干/祖细胞群。本研究的体内实验表明，与健康组相比，肺气肿小鼠模型组肺CD45-/CD31-/Sca-1+数目明显减少，Shh、Ptch1和Gli1的 mRNA及蛋白相对表达量均较PBS组降低，免疫组化结果显示Shh信号通路的三个关键组分Shh、Ptch1和Gli1和在小鼠肺组织中的主要表达于肺泡以上气道的上皮细胞。体外实验结果表明，肺CD45-/CD31-/Sca-1+细胞具有自我增殖能力，CSE可使肺CD45-/CD31-/Sca-1+细胞Shh 、Ptch1和Gli1的 mRNA及蛋白相对表达量均较表达下调。通过上述研究，我们首次提出香烟可抑制肺干/祖细胞Shh信号通路，干扰肺上皮干/祖细胞的增殖与修复，使肺干/祖细胞增殖能力减弱，参与COPD的发病过程。