放射治疗损伤颌下腺线粒体稳态调节机制及策略

Radiotherapy for head and neck cancer commonly leads to damage to normal tissues located in the radiation field, especially the salivary glands. As a result, patients can develop radiation sialadenitis, mucositis, radiation caries, and xerostomia, all of which significantly affect their quality of life. Thus, it is of great importance to find therapeutic approaches that can decrease the injury to the salivary glands induced by radiation. It has previously been found that the destruction of the crests and intra–mitochondrial vacuolization is the main pathophysiologic characteristic in irradiated salivary glands. Nevertheless, its relevant molecular mechanisms remain unknown. .Nicotinamide phosphoribosyltransferase (NAMPT) is a key enzyme for the nicotinamide adenine dinucleotide (NAD) salvage pathway, but also a cytokine participating in cell survival, metabolism, and longevity, with a broad effect on cellular functions in physiology and pathology. However, the regulatory events of NAMPT in response to the radiation in salivary glands are not well understood. In our recent studies, NAMPT was mainly distributed in the cytoplasm of granular convoluted tubule cells and ductal cells in normal submandibular gland. Both the mRNA and the protein expressions of NAMPT were downregulated after radiation but upregulated with phenylephrine pretreatment both in vivo and in vitro. Moreover, the protein expression of phosphorylated-Akt was decreased in irradiated glands, and phenylephrine pretreatment restored the expression of p-Akt. Since the NAD salvage pathway plays a vital role in the energy metabolism of mitochondria in cells, and PI3K/Akt signal pathway has been well known in regulation of the biosynthesis metabolism of mitochondria in condition of oxidation stress, our findings suggested that NAMPT upregulation might be involved in the regulatory mechanisms of mitochondrial homeostasis in irradiated salivary gland. .Therefore, in the present study, we hypothesize that the disturbance of mitochondrial homeostasis is the key regulatory mechanisms induced by ionizing radiation, and NAMPT may play an important role in regulating signal pathways of mitochondrial homeostasis. In order to understand the regulatory mechanisms of mitochondrial homeostasis in irradiated salivary gland, we propose to study the characteristics of this process, including the mitochondrial energy metabolisms, the mitochondrial biogenesis and degradation, and the NAMPT-mediated signal pathways with cellular and animal models of irradiated submandibular gland of rat. Also, we will investigate the roles of phenylephrine and nicotinamide in the potential restoration of mitochondrial homeostasis. The results obtained from the proposed research will elucidate the molecular mechanisms of radiation damage in salivary gland, and provide targets and strategies for potential interventions for radiation sialadenitis.

放射治疗是头颈癌等恶性肿瘤术后重要治疗措施，但不可避免地造成唾液腺放射性损伤，是世界范围内临床尚未攻克的难题。线粒体变性是唾液腺电离辐射损伤的重要病理特征，但分子机制不清楚。我们前期研究发现，电离辐射下调了颌下腺烟酰胺磷酸核糖转移酶（NAMPT）的表达，而苯肾上腺素（PE）能够上调NAMPT及其上游分子PI3K/Akt的表达，从而促进辐射损伤颌下腺结构和功能的修复。由于NAMPT是线粒体能量代谢途径中关键限速酶，PI3K/Akt是调控线粒体合成重要分子，所以我们推测线粒体稳态调节紊乱是唾液腺电离辐射损伤的关键机制。故本课题拟从线粒体能量代谢和生成降解代谢途径研究电离辐射致颌下腺损伤的分子机制，系统阐述NAMPT介导的多条分子途径在线粒体稳态调节中的关键靶点，针对性地利用PE或其它能够改善线粒体功能的分子来有效恢复线粒体稳态，为临床头颈癌等恶性肿瘤病人提供防治唾液腺电离辐射损伤的新策略。

（1）项目的背景：放射性唾液腺炎是严重影响病人生存质量的临床常见疾病，又是世界范围内尚无有效治疗方法的难治性疾病。辐射所致唾液腺损伤的重要病理特征是线粒体损伤，导致组织细胞产生过量的活性氧，进一步引起线粒体稳态破坏，但目前以线粒体为靶点预防辐射损伤的治疗方法尚罕见。文献报道苯肾上腺素（phenylephrine, PE）、烟酰胺（nicotinamide, NAM）、甘草酸（glycyrrhizic acid, GA）和红景天苷（salidroside, Sal）在多种组织和细胞具有抗氧化应激的作用，但在辐射损伤唾液腺中的作用尚未见报道。因此，本项目针对性地研究PE、NAM、GA和Sal对电离辐射损伤的颌下腺细胞线粒体稳态的影响，为头颈癌等恶性肿瘤病人防治唾液腺辐射损伤提供新的高效安全的防治策略。.（2）主要研究内容：在动物（大鼠）唾液腺组织及细胞水平，研究了PE、NAM、GA和Sal对电离辐射损伤唾液腺细胞的线粒体的保护作用。揭示了其分子机制可能为降低辐射后腺体细胞线粒体的活性氧水平、促进线粒体能量代谢及生物合成，最终减轻辐射所致细胞凋亡，从而维持唾液腺较正常的分泌功能。.（3）重要结果：1）PE经α1-AR活化减轻内辐射所致的腺体线粒体数量、结构和功能的损伤，其分子机制与NAMPT介导的线粒体能量代谢途径相关，亦与激活α1-AR后PI3K/AKT途径而促进线粒体合成增加相关；2）NAM可通过影响线粒体的能量合成，减轻辐射损伤细胞的凋亡；3）GA和Sal亦可通过降低辐射后线粒体的活性氧水平，维持线粒体能量代谢及生物合成，减轻辐射所致的细胞凋亡，从而维持唾液腺较正常的分泌功能。.（4）关键数据：GA和Sal可以显著降低辐射后颌下腺细胞内线粒体ROS水平，其效果与线粒体ROS靶向清除剂MitoTempo效率相当，提示补气中药作为线粒体营养素在抗唾液腺辐射损伤中的重要作用。.（5）科学意义：本研究提示PE、NAM、GA和Sal均可作为线粒体营养素，靶向减轻辐射后唾液腺细胞线粒体的氧化损伤，对于唾液腺辐射损伤的防治具有良好的潜在应用前景，将极大提高头颈癌患者的生存质量，具有极显著的社会效益。