气管干细胞增殖分化过程中Shh信号通路的动态观察

目的 探讨大鼠气管上皮损伤后，气管干细胞增殖分化过程中Shh信号通路的动态变化。方法 大鼠麻醉后，暴露气管，于平胸骨上缘处离断，与肺相连一端行气管插管维持呼吸，近端应用5-氟尿嘧啶（5-FU）诱发气管上皮损伤，动态观察修复过程；同时在观察的各个时间点取气管上皮标本通过IKT—PCIK检测Shh的表达情况。结果 5-Fu作用30min后大鼠气管上皮细胞绝大部分脱落，仅残留少量裸核样G0期细胞位于基底膜上，之后这些细胞数目逐渐增多，逐渐经过扁平、立方形态，向纤毛细胞、粘液细胞、基底细胞等分化，于去除5-FU后48h基本恢复假复层纤毛柱状上皮；正常组织及5-FU作用30min时的气管上皮均无Shh表达，去除5-Fu恢复3h后可见Shh轻度表达，于恢复12h表达达高峰，之后表达减弱，48h时Shh表达已极微弱。结论 shh参与了气管干细胞的增殖分化过程。

Dynamic observation of Shh signaling pathway during process of tracheal stem cell proliferation and differentiation

[Objective] To explore the dynamic change of Shh in the process of tracheal epithelial stem cell proliferation and differentiation after tracheal injury. [Methods] The trachea of rat was exposed and cut at the level of suprasternal notch after the rat was anesthetized. The distal part which connected with the lung was intubated to keep breathing and the proximal part was injured by 5-FU. The injury-repair process was dynamically observed un- der LM. The tracheal epithelia were sampled to detect the expression of Shh by RT-PCR at different time as well. [Results] Most epithelial cells exfoliated 30 minutes after being treated by 5-FU. A few nude-nucleus-like cells in Go phase were randomly distributed on the basement membrane. After 5-FU being removed, the number of the cells on the basal membrane increased gradually. The cells which undergoing the shape of fiat and cuboid differentiated into the ciliated, mucosal and basal cells. Finally, 48 hours later the pseudostratified ciliated clumnar epithelium was restored the similar as original mode. There was no expression of Shh in normal epithelia and the epithelia which were treated by 5-FU for 30 min. Slight expression of Shh emerged in the epithelia 3 hours after 5-FU being removed. The expression of Shh increased 6 hours later and reached its peak 12 hours later. After that time, the expression of Shh became decreased and was very faint 48 hours later. [Conclusion] Shh took part in the process of proliferation and differentiation of tracheal epithelial stem cells.