炎症微环境通过PI3K/AKT/FOXO信号通路促进牙周膜干细胞衰老的机制研究

The functional decline of PDLSCs directly influence the regenerative capacity of periapical lesions. PI3K/Akt/FOXO signaling pathway regulating stem cells senescence is associated with impaired regeneration of various tissues. In preliminary experiment we indicated the senescence of hDPSLCs derived from periapical lesions. Based on the above, we proposes a new hypothesis that PI3K/AKT/FOXO signal pathway modulate the senescence of PDLSCs in periapical inflammatory microenvironment. We isolated PDLSCs from human periapical lesion and establish a mouse model of apical periodontitis. Then we investigate the correlations of environmental factors, cell senescence and dynamic expression of PI3K/AKT/FOXO during different stages of apical periodontitis. Next we mimic inflammatory microenvironment in vitro and construct Akt and FOXO knockdown PDLSCs strain and further discuss the effect mechanism of PI3K/AKT/FOXO axis on PDLSCs senescence. Therefore, the present project may deepen molecular understanding of cell senescence under inflammatory microenvironment that may provide theoretical basis for tissue regeneration partly through adapting biological properties of stem cells.

研究证实PDLSCs功能受损是抑制根尖周病变组织修复的重要原因。近年来研究注意到PI3K/Akt/FOXO信号通路调控的干细胞衰老与组织再生能力下降密切相关。前期研究提示根尖周炎症来源的PDLSCs呈现衰老状态，据此我们提出在慢性根尖周炎病理过程中，炎症微环境通过调控PI3K/AKT/FOXO信号通路促进PDLSCs衰老的假设。本项目采集人病变组织及建立根尖周动物模型，探讨根尖周病变不同阶段影响PDLSCs衰老的微环境因素，检测PDLSCs衰老与PI3K/AKT/FOX活化水平的相关性；体外模拟炎症微环境，通过构建Akt及FOXO基因沉默细胞模型，探讨PI3K/AKT/FOXO信号轴调控炎症环境下PDLSCs衰老的分子机制。本项目研究有助于阐明炎症微环境下PDLSCs功能变化的调节机理，为通过干细胞调控促进组织再生修复提供理论依据。

临床上经久不愈的根尖周炎病损是困扰牙体牙髓科医生的一大难题，在这种情况下患者最终不得不接受手术治疗甚至拔牙术，因此如何促进根尖周病损区的修复引起学者们密切关注。最近研究发现微环境变化导致的干细胞衰老是抑制组织再生的重要因素，本课题拟从炎症环境下细胞衰老的角度探讨根尖周炎发生发展的机制。在根尖周炎模型鼠中，检测到慢性炎症期根尖周组织细胞出现衰老，且磷酸化的AKT和FOXO蛋白表达上调，当采用PI3K/AKT通路抑制剂LY294002注射后根尖周炎症病损区范围明显缩小，炎症因子表达量亦显著下降。接下来通过采集和分析临床根尖周囊肿及肉芽肿标本，发现与根尖周肉芽肿相比，囊肿的炎症因子表达量较低，但是细胞衰老的发生率显著提高，磷酸化的AKT和FOXO蛋白表达明显上调；相关性分析显示AKT和FOXO1阳性细胞数和PTRF表达呈正相关。最后体外实验进一步分析得出双氧水诱导能显著上调牙周膜干细胞的衰老相关基因和蛋白的表达，且该表达水平能被LY294002所抑制。本项目阐明了PI3K/AKT/FOXO信号轴参与了牙周膜干细胞的衰老和大鼠根尖周炎发展过程，该衰老过程可能与根尖周囊肿的形成机制密切相关。