单向滤膜减压重建炎症牙髓修复及再生适宜微环境的机制及其转化研究

课题组前期工作建立了单向滤膜减压保髓新技术，有效隔离口腔微生物，缓解髓腔高压，在小型猪模型上实现了不可复性牙髓炎的牙髓保存治疗；临床亦成功用于部分病例。该技术，有助于牙髓炎症消退，但其对牙髓微环境的调控及如何提高牙髓保存质量，避免过度矿化，仍有待解决。本研究将探索该技术重建炎症牙髓修复再生适宜微环境的机制。应用小型猪牙髓炎模型，观察单向滤膜减压前后，缺氧诱导因子HIF-1α及各种炎症因子动态表达谱，鉴定发挥关键作用的因子；体外低氧诱导联合靶向敲除HIF-1α，鉴定各种因子的细胞来源，体内应用HIF-1α抑制剂和激活剂，验证HIF-1α对各种因子的影响以及它对牙髓组织矿化的影响；应用细胞归巢方法，序列促进牙髓组织及牙本质再生评估筛选关键生长因子对牙髓再生修复的影响。本研究有望从牙髓炎症组织水平、细胞水平及分子水平揭示单向滤膜减压重建牙髓修复再生适宜微环境的机制，推动临床转化。

Dental pulp preservation is vital in maintaining the physiological function and lifespan of teeth. Our previous studies have established a new decompressed filling technique that enables us to obtain hypertension decompression, bacteria isolation and pulp preservation. The decompressed filling technique allowed us to successfully achieve vital pulp therapy on the irreversible pulpitis with local abscess using miniature swine models. In clinic, we also demonstrated that patients with irreversible pulpitis also had good results via this method. The patented decompressed filling membrane is our core technology that has been approved for clinical research. Though the decompressed filling innovation is effective to subside pulp inflammation, the mechanism that regulates the dental pulp microenvironment still remains incompletely understood, and the strategy to improve the pulp preservation has yet been fully explored. This study will explore the mechanism of decompressed filling technique in improving the microenvironment of inflammatory pulp and promoting pulp regeneration. The study includes the application of miniature swine pulpitis model to observe the correlation between hypoxic-induced factor HIF-1α, cytokines, chemokines, growth factor expression and neutrophils infiltration before and after decompressed filling treatments. Meanwhile, the in vitro primary culture of dental pulp tissue source neutrophils and fibroblasts, hypoxic induction and Crispr-cas9 technology targeted knockout HIF-1α, to examine the role of HIF-1α in the process of inflammatory hypoxia. Moreover, HIF-1α promoter and inhibitor will be used in the miniature swine model to confirm the functions of HIF-1α.After inflammation subsided, cell homing (with screened key growth factors ) method will be used to promote the regeneration of pulp soft tissue and dental pulp-dentin complex sequentially. This study is expected to reveal the mechanism of decompressed filling technique reconstruction of suitable microenvironment from pulp inflammatory tissue level, cell level and molecular level, thus laying out a solid foundation for the clinical translation of decompressed filling technique.