NFκB信号通路中泛素蛋白酶体系统在牙周病的发病机制和防治中的作用

Periodontal disease is a polymicrobial infectious disease and it is characterized by loss of tooth-supportitve tissue including periodontal ligament and alveolar bone. Periodontal tissue comprises multicopartmental groups of interacting cells and matrices that provide continuous support, attachment, proprioception, and physical protection for the teeth. So once lost the attachment by the periodontal ligament, it is difficult for the regeneration of periodontal ligament. It reported by World health organization that the periodontal disease is the most important cause of tooth loose in adult. Besides, periodontal disease is related to kinds of systemic diseases, such as cardiovascular disease...Current theory predicts the existence of at least four basic components of an inflammatory response: inducers of inflammation，their sensors，inflammatory mediators secreted by sensors after stimulation by inducers，and effectors influencing the tissues affected by inflammation. On the one hand, dental plaque is considered as the main cause of periodontal diseases. And nuclear factor-kappaB ligand (NF-kB) path-way is the one which mediates the pathological role of the dental plaque on the periodontal tissue injury. Dental plaque could activate the NF-kB path-way will promote the expression of various inflammatory mediators such as interleukin (IL)-6, IL-8 and tumor necrosis factor-alpha (TNF-a). And the inflammatory mediators will amplify the inflammation and cause periodontal tissue injury. On the other hand, beta-TRCP, an ubiquitin E3 ligase and cylindromatosis (CYLD),a de-ubiquitin enzyme, are the positive and negative regulator of NF-kB, respectively. IkBa is the most important inhibitor of NF-kB, and beta-TRCP triggers the degradation of IkBa. It is discovered that all the IkBs were degraded by the mediation of beta-TRCP. And under the stimulation of TNFα and other inflammation mediator,CYLD blocked the activation of NF-kB path-way by de-ubiquitin TRAF2 and TRAF6...As it is shown in our pre-experiment results that proteasome inhibitor, beta-TRCP and CYLD all played an important role in the osteoclastgenesis. In this study, via cell, biochemical and animal experiment, we will reveal the effect of Ubiquitin Ptroteasome System and de-ubiquitin enzyme CYLD on the pathological mechanisms prevention and treatment of periodontal diseases. And, we will investigate the effect of proteasome inhibitor on the treatment of periodontal disease. All of these basic evidences will offer the clinical application of proteasome inhibitor.

据世界卫生组织统计牙周病为口腔的第二大常见病。牙周病与多种疾病相关联，是关乎广大人民群众切身利益急需攻克的重要课题。牙菌斑是牙周炎的主要致病因素， NF-kB信号通路在介导菌斑引起的牙周组织损伤中起到重要作用。泛素蛋白酶系统，特别是泛素连接酶beta-TRCP和去泛素化酶CYLD对NF-kB起到重要的调控作用，通过NF-kB途径调控组织炎症和骨组织代谢。同时蛋白酶体抑制剂在组织炎症和骨组织代谢中起到重要的调控作用。课题组前期工作表明：泛素连接酶Beta-TRCP, 去泛素化酶CYLD, 蛋白酶体抑制剂通过NF-KB信号通路对破骨细胞分化起着重要的调控作用。本课题揭示泛素蛋白酶体系统和去泛素化酶CYLD在牙周病中的致病机理和牙周组织修复中的重要作用，通过多角度的研究为蛋白酶体抑制剂治疗牙周病的科研成果转化，从分子水平、细胞水平和动物体水平提供依据，具有重要的学术理论意义和临床实用价值。

背景：牙周病为口腔常见病是成人牙齿脱落主原，与多种疾病关联是关乎广大人民群众健康急需攻克的重要课题。NF-kB 信号通路在介导菌斑引起的牙周组织损伤中起到重要作用。泛素蛋白酶系统中β-TRCP 等通过 NF-κB 途径调控组织炎症和骨组织代谢。蛋白酶体抑制剂在组织炎症和骨组织代谢中起到重要的调控作用。.主要研究内容：从基因，分子，蛋白，细胞，动物等层面，通过PCR、细胞免疫荧光，荧光探针设计、WB、流式细胞仪，大鼠牙周病模型，micro CT，基因敲除老鼠，免疫组织化学等方法阐明了NFκB信号通路中泛素蛋白酶体系统（UPS）在牙周病发病机制和防治中的作用，泛素蛋白酶体抑制剂Bortezomib（BTZ）的治疗机制和作用。设计、制备、研究炎症代谢产物探针。根据国际牙周病概念外延包含种植体周围炎，延伸研究植入颗粒的炎症反应。.重要成果和关键数据：1成功培养人牙周膜细胞（hPDLCs）和牙周膜干细胞（hPDLSCs）并鉴定。2发现UPS在牙周炎中的致病机制和再生修复中的作用。LPS引起hPDLCs中IκBa降解和NF-κB/p65核移激活NF-κB通路，引起TNF-a、IL-1β、IL-6和IL-8等表达，导致下游CyclinD1，Caspase3等异常表达。 而hPDLCs的凋亡、NF-κB核转移可被BTZ抑制。3 UPS对炎症刺激下hPDLSCs干性变化调控：LPS引起hPDLSCs衰老和凋亡。敲除NF-κB通路β-TRCP可阻断细胞衰老、凋亡等，并可被BTZ抑制。4 BTZ抑制IκBa的降解和NF-κB/p65的核移抑制NF-κB活性。同时证明1nM以内的BTZ在hPDLCs、成骨细胞，破骨细胞中的药物安全性。5发现BTZ通过抑制MKP1的降解等，抑制MAPKs/AP-1信号通路活性，从而抑制牙周炎进展。6 BTZ对大鼠牙周炎模型作用和机制。7种植体周围炎的内涵外延，研究发现UPS对颗粒引起的炎症和无菌性骨吸收的调控作用。8制备β-TRCP等基因敲除鼠进一步研究UPS的作用。9制备炎症代谢产物次磺酸检测探针。.科学意义：为UPS在牙周病防治中的作用提供新的策略和依据，为BTZ的应用提供了坚实的理论基础。在美国牙周病协会会刊JOP等顶级SCI及其它外文杂志共发表9篇（2篇SCI），中文核心3篇，另有1篇SCI 审稿中，获批发明专利1项、在审2项，培养硕士博士共9名。