抑制Ape1/Ref-1调控老化细胞SASP对放射性肺损伤的防护作用及机制研究

Radiation-induced lung injury (RILI) is the common and serious complication of thoracic tumor radiotherapy, and the mechanisms of which has not yet been fully elucidated. Recent studies suggested that the cellular senescence of alveolar type II epithelial cells in the radiation field is closed related to RILI, and the senescence-associated secretory phenotype (SASP) plays a key suppressive role to RILI. Telomeric DNA damage initiates cellular senescence and repair of which possibly only processed through base excision repair (BER). Our preliminary data showed that knocking down of the core BER enzyme APE1 rendered apoptosis in telomerase positive tumor cells and cellular senescence in telomerase negative primary fibroblasts. We then hypothesized that the telomeric DNA lesion level is the key determinative factor for cell fate caused by APE1 deficiency. Our current project will focus on the impact of APE1 deficiency on cellular senescence and SASP in suppressing RILI by taking advantage of APE1 conditional knockout mice model. In addition, we will explore the mechanisms of APE1 in telomeric DNA repair by utilizing the primary alveolar epithelial cells and cell model with different telomere length/telomerase status. Our project will provide experimental support for APE1-based targeted therapy.

放射性肺损伤是胸部肿瘤放射治疗的常见严重并发症，其发生机制尚未完全阐明。新近研究表明射野内肺泡Ⅱ型上皮细胞老化与放射性肺损伤密切相关，老化细胞衰老相关分泌表型(SASP)为抑制放射性肺纤维化的关键因素。细胞老化自端粒DNA损伤始动，而端粒DNA损伤可能仅对碱基切除修复(BER)途径开放。我们前期研究证实敲低BER途径核心限速酶脱嘌呤脱嘧啶核酸内切酶(APE1)可促进电离辐射所诱导，端粒酶阳性的肿瘤细胞凋亡、端粒酶阴性的原代成纤维细胞老化。据此我们首先提出端粒DNA损伤程度可能是APE1缺陷导致不同细胞命运的决定因素。本项目利用APE1条件敲除小鼠构建放射性肺损伤模型，研究抑制APE1调控老化细胞SASP对放射性肺损伤的影响；并通过复制原代培养肺泡上皮细胞和不同端粒长度/端粒酶表达状态差异组合细胞模型，重点探讨APE1参与端粒DNA损伤与修复的分子机制，为基于APE1的靶向治疗提供实验依据。

放射性肺损伤是胸部肿瘤放射治疗的常见严重并发症，其发生机制尚未完全阐明。新近研究表明射野内肺泡Ⅱ型上皮细胞老化与放射性肺损伤密切相关，老化细胞衰老相关分泌表型(SASP)为抑制放射性肺纤维化的关键因素。细胞老化自端粒DNA损伤始动，而端粒DNA损伤可能仅对碱基切除修复(BER)途径开放。我们前期研究证实敲低BER途径核心限速酶脱嘌呤脱嘧啶核酸内切酶(APE1)可促进电离辐射所诱导，端粒酶阳性的肿瘤细胞凋亡、端粒酶阴性的原代成纤维细胞老化。据此我们首先提出端粒DNA损伤程度可能是APE1缺陷导致不同细胞命运的决定因素。本项目利用APE1条件敲除小鼠构建放射性肺损伤模型，研究抑制APE1调控老化细胞SASP对放射性肺损伤的影响；并通过复制原代培养肺泡上皮细胞和不同端粒长度/端粒酶表达状态差异组合细胞模型，重点探讨APE1参与端粒DNA损伤与修复的分子机制。本项目研究发现APE1在体内模型中可以防止DNA损伤并阻止细胞老化，并且APE1可对端粒区的DNA损伤具有特殊的保护作用。APE1敲除型小鼠放射性肺损伤严重程度高于APE1野生型小鼠, Ape1敲除小鼠组织中的老化细胞数量明显增多, 增殖能力明显减弱。放射治疗后，APE1-KO小鼠出现老化、肺纤维化现象更明显、SASP的mRNA水平高于野生型小鼠、SASP相关通路激活。这为后期通过抑制APE1/ Ref-1调控SASP来预防胸部肿瘤放疗所引起的放射性肺损伤的转化研究提供试验基础。