AIF介导的Caspase非依赖的细胞凋亡信号通路对毛囊周期的影响及其作用机制研究

Hair follicle is a self-renewal organ with definitive cycles. Apoptosis plays an important role in hair cycles. Previous studies show caspase dependent apoptosis signaling pathway is actively involved in regulation of hair follicle cycles. Abnormal regulation of caspase dependent apoptosis on hair cycles often leads to hair loss. Under the support from completed China National Natural Science Foundation, for the first time we find that hair growth can be achieved by retardation of the catagen arrivals through prevention of the expression and translocation of apoptosis-inducing factor (AIF) into the nucleus in hair follicles during transition from anagen to catagen［Jin Yu Li : Fundam Clin Pharmacol. 2015 Jan 25］. It is well-known AIF mediated apoptosis is independence of caspase, which plays a unique role in maintenance the homeostasis in tissues with highly metabolism and active self-renewal, such as hair follicles. As such we speculate that AIF-mediated apoptosis signaling pathway might be a key signaling pathway involved in regulating hair follicle cycles. To this end the role of AIF mediated apoptosis in regulating hair cycles will be investigated and compared with caspase dependent apoptosis using the techniques associated with molecular biology, cell biology and histology. The related mechanism underlying apoptosis and hair cycles will be dissected, which may lay basis for extending hair follicle biology and provide new idea for treatment of hair loss.

毛囊周期性自我更新形成毛囊周期。凋亡在毛囊周期性自我更新中发挥重要作用。以往研究发现Caspase依赖的细胞凋亡参与毛囊周期的调控。该通路异常会导致脱发的发生。新近我们在已结题的国家自然科学基金项目资助下，国际上首次研究发现阻止凋亡诱导因子(AIF)在退行期毛囊中的表达和转位入核，抑制毛囊凋亡的发生，延迟毛囊退行期的到来，促进生发［Jin Yu Li (刘晋宇) : Fundam Clin Pharmacol. 2015 Jan 25］。已知AIF介导的细胞凋亡是一种全新的、不依赖于Caspase的细胞凋亡。该凋亡通路在维持代谢旺盛或自我更新活跃组织(如毛囊) 的内环境稳定方面发挥独特而重要作用。因此本课题拟在上述研究已取得重要进展基础上，探究AIF介导的细胞凋亡对毛囊周期影响的分子机制，并与Caspase介导的细胞凋亡比较，解析其中的关系，建立相应的信号传导通路，为毛囊再生提供理论依据。

我们在国家自然科学基金面上项目资助下，对出生后第11天（毛囊生长期）、16天（毛囊退行期早期）、18天（毛囊退行期晚期）和21天（毛囊静止期）C57小鼠的毛囊组织结构进行了动态检测。HE染色发现C57小鼠背部皮肤毛囊由生长期进入退行期和静止期时，毛囊逐渐萎缩，密度降低。免疫荧光、Western blot 及生物化学检测发现毛囊由生长期进入退行期时，CytoC、Caspase3(17KD) 表达下调，ROS、Tunnel阳性细胞比例增加，细胞核AIF、PARP1表达上调，伴有PAR蓄积，NAD+及NADH表达下调。毛囊由退行期进入静止期时，CytoC、Caspase3(17KD) 表达上调，细胞核AIF、PARP1和PAR表达下调。为了探讨AIF-PARP1和CytoC-Caspase3 信号通路在毛囊周期中的作用。我们分别利用PARP1抑制剂（BMN）和Caspase抑制剂（Ac-DEVD-CHO）处理小鼠。HE染色发现，与对照组相比，Caspase 3 抑制剂组后小鼠皮肤增厚，毛囊数量增多，毛球直径增粗；PARP 1抑制剂组小鼠脂肪增加，毛囊数量减少，毛球显著萎缩，可见脂肪组织显著增生；联合给药组小鼠毛囊形态与对照组小鼠无明显区别，毛球直径无差异，但毛囊数量增多。Western blot检测发现：Caspase 3抑制剂组与对照组相比，Caspase 3(17KD)表达下调, 细胞核AIF表达上调，同时伴有PAR蓄积；PARP 1 抑制剂组与对照组相比，Cyto C 表达显著上调，AIF(57KD)和PAR表达明显降低；双抑制剂组与对照组相比，除了Cyto C表达显著上调，AIF(57KD)表达下调外，PARP1、PAR、Caspase 3(17KD)表达与对照组比统计学上差异。上述结果提示：CytoC-Caspase3和AIF-PARP1信号通路之间存在负向调控作用，共同维持毛囊组织自我更新。