组织特异性细胞外基质构建三维涎腺类器官及其作用机制的研究

Salivary gland hypofunction(i.e. hyposalvation) is a common oral disease. Certain clinical conditions, such as X-irradiation therapy for the patients with head and neck cancers, , Sjögren’s Syndrome, and congenital absence , could cause salivary gland hypofunction. This often leads to dry mouth syndrome, subsequent dental caries, and swallowing difficulty, which seriously affects the patient's quality of life. Currently, effective treatments are not readily available in spite of various attempts of therapies to recover salivary gland function. Bioengineered salivary glands and organoids culture provide a promising solution for the patients with hyposalivation. Based on our recent studies and reports from other groups, tissue specific extracellular matrix (ECM) provides the optimal microenvironment for the cells originated from the same type of tissue to adhere, proliferate and differentiate. We hypothesize that salivary gland tissue specific ECM with hydrogel renders a three dimensional milieu for salivary gland progenitor cells to form functional organoids of salivary glands. The goals of this study are to determine the biological effect of tissue specific ECM on phonotype and function of salivary gland cells in vitro, and also explore the molecular mechanisms of how salivary gland organoids within 3D ECM gel regenerate in vivo. Therefore, we will assess the cytokines and biological components within the ECM, cell growth, mRNA and protein makers for saliva secretion functions of the salivary gland organoids after culturing salivary gland progenitor cells on the ECM using hanging drop cell culture, as well as autophagy function, anti-apoptosis, antioxidation in vivo experiment. We anticipate that tissue specific ECM will promote the formation of functional salivary gland organoids, which provides a promising new treatment option for patients with salivary gland hypofunction.

头颈部恶性肿瘤的放射治疗、舍格伦综合征或其他原因导致的涎腺分泌功能障碍，可引起口干、继发性龋、吞咽困难等，严重影响患者生活质量。目前尚无有效治疗方法。组织工程涎腺和类器官培养为治疗涎腺功能障碍带来了希望。我们的前期研究显示：组织特异性细胞外基质(ECM)为细胞三维生长和分化提供最佳的的微环境。因此本研究提出以下科学假设：涎腺组织ECM为涎腺前体细胞提供三维生长的微环境，促进其形成具有分泌功能的涎腺类器官。为验证此假设，明确涎腺类器官构建过程中所必要的生物活性因子/蛋白，将涎腺前体细胞与组织特异性ECM水凝胶进行悬滴/动态培养并植入动物体内；评价涎腺类器官的结构、功能以及植入放射性涎腺损伤动物模型后类器官中的细胞表型、自噬功能、抗凋亡和抗氧化应激等。本研究从组织特异性ECM构建功能性三维类器官这个新视点，揭示涎腺类器官的生物学特点及其修复涎腺损伤的分子机制，为类器官治疗这类疾病提供新思路。

头颈部恶性肿瘤的放射治疗、舍格伦综合征、发育异常均可导致涎腺分泌功能降低或丧失，引起口干、继发性龋、吞咽困难，严重影响患者的生活质量和身体健康，目前尚无有效的治疗方法。三维涎腺类器官构建可能为治疗涎腺功能障碍带来希望。本研究利用SD大鼠下颌下腺及肝组织分别获取其脱细胞基质并制备可以进行体外三维培养用的肝素-透明质酸-肝（颌下腺）细胞外组织特异性水凝胶细胞外基质（HP-HA-肝（颌下腺）ECM水凝胶），通过对ECM中与细胞生长相关生物因子的缓释情况进行了初步评估，发现下颌下腺脱细胞ECM中含有EGF、VEGF、bFGF、IGF等生长因子，并能平稳缓慢释放达7天以上，具有缓释细胞因子的作用，同时细胞在特异性水凝胶细胞外基质中相容性良好。将涎腺干细胞与HP-HA-肝（颌下腺）ECM水凝胶复合培养，第 10 天时细胞表达 AMY-1、AQP5、CK7 及 CK14，并观察到有导管样结构形成，初步获得了能表达唾液腺细胞功能的类器官。现在正在将体外培养的类器官植入SD大鼠放射性损伤的颌下腺组织中进行相关功能修复的实验研究。本研究揭示涎腺类器官可能为治疗涎腺功能障碍性疾病提供有效的新方法思路。