Role of sphingosine-1-phosphate receptor 2 in osteoblastogenesis and bone regeneration in periodontitis

1-磷酸鞘氨醇受体2在牙周炎成骨细胞生成和骨再生中的作用

• 批准号: 10270131
• 负责人: Hong Yu
• 金额: $ 22.58万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-06 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10270131
• 关键词: Actinobacillus actinomycetemcomitans; Adult; Affect; Alkaline Phosphatase; Alveolar Bone Loss; BMP2 gene; BMP7 gene; BMPR2 gene; Bacteria; Bone Marrow; Bone Morphogenetic Proteins; Bone Regeneration; Bone Resorption; Bone Tissue; C57BL/6 Mouse; Cell Adhesion; Cell fusion; Cells; Chronic; Couples; Cultured Cells; Data; Dimethyl Sulfoxide; Disease; Excision; Food; Foundations; G-Protein-Coupled Receptors; GTP-Binding Protein alpha Subunits, Gs; Generations; Genes; Gingiva; Goals; In Vitro; Infection; Inflammatory; Interleukin-1 beta; Interleukin-6; Knowledge; Ligature; Lipopolysaccharides; MAP Kinase Gene; Mammalian Cell; Maxilla; Membrane; Messenger RNA; Microbial Biofilms; Modality; Mononuclear; Mus; Natural regeneration; Oral; Osteoblasts; Osteocalcin; Osteoclasts; Patients; Periodontitis; Pharmacology; Play; Powder dose form; Process; Production; Protein Family; Protein Kinase; Proteins; Role; Signal Pathway; Signal Transduction; Silk; Smad Proteins; Sphingosine-1-Phosphate Receptor; Stromal Cells; TNF gene; TNFSF11 gene; Testing; Tissues; Tooth Loss; Tooth structure; Topical application; United States; alveolar bone; bone; bone morphogenetic protein receptors; cytokine; dysbiosis; inflammatory bone loss; inhibitor/antagonist; knock-down; macrophage; mesenchymal stromal cell; monocyte; novel; novel therapeutic intervention; novel therapeutics; oral pathogen; osteoblast differentiation; osteoblast proliferation; osteoclastogenesis; osteogenic; osteogenic protein; receptor; recruit; response; small hairpin RNA; transcription factor

Periodontitis is a bacteria-driven inflammatory bone loss disease affecting 47% of adults in the United States. Oral pathogens induce the generation of inflammatory cytokines (including IL-1β, IL-6, TNF-α, and RANKL), which attract monocytes to gingival tissues. Under RANKL stimulation, these mononuclear cells can further differentiate and fuse to form multinucleated osteoclasts, resulting in alveolar bone loss. Meanwhile, inflammatory cytokines associated with periodontitis inhibit osteoblast differentiation and bone regeneration. With current limited treatment modalities, periodontitis is still the major cause of alveolar bone loss and tooth loss in adults. Our long-term goal is to develop effective new therapies to treat the disease. We were the first to demonstrate that sphingosine-1-phosphate receptor 2 (S1PR2, a G-protein-coupled receptor) plays a key role in regulating the inflammatory bone loss response in periodontitis. Knockdown of S1PR2 by a S1PR2-directed shRNA or inhibition of S1PR2 by its specific inhibitor (JTE013) reduced PI3K, NF-κB, and MAPKs protein kinases induced by the oral pathogen Aggregatibacter actinomycetemcomitans (Aa), and decreased IL-1β, IL- 6, TNF-α levels induced by Aa. Moreover, treatment with the S1PR2 shRNA or JTE013 decreased cells adhesion units (podosome components) induced by RANKL, inhibited osteoclastogenesis and bone resorption induced by RANKL. Oral topical administration of JTE013 alleviated inflammatory bone loss in C57BL/6 mice with periodontitis induced by ligature placement. However, there is a knowledge gap on how S1PR2 regulates osteoblastogenesis and bone regeneration. Thus, the goal of this application is to determine the role of S1PR2 in controlling osteoblastogenesis and bone regeneration. Our preliminary data show that inhibition of S1PR2 by JTE013 in bone marrow-derived stromal cells (BMSCs) cultured in osteogenic media increases osteoblastogenesis compared with vehicle treatment. Treatment with JTE013 increased the mRNA levels of osteogenic genes, including alkaline phosphatase, Runt-related transcription factor 2 (Runx2), osteocalcin, osterix, and bone morphogenetic protein (BMP) 2 compared with vehicle treatment. Additionally, treatment with JTE013 increased the levels of osteogenic proteins, including BMP2, BMP7, BMP receptor II, BMP receptor 1A, and p-Smad 1/5/9 protein compared with control vehicle treatment. Thus, we hypothesize that inhibition of S1PR2 in pre-osteoblast cells promotes osteoblastogenesis and bone regeneration. Our specific aims will determine 1) if shRNA knockdown or inhibition of S1PR2 in vitro will increase osteoblastogenesis via BMPs/Smad signaling pathway and/or other signaling pathways; and 2) whether pharmacological inhibition of S1PR2 by JTE013 in mice with periodontitis promotes alveolar bone regeneration following inflammatory bone loss. This application defines novel signaling pathways regulated by S1PR2 in promoting osteoblastogenesis. It will lay the foundation to develop a novel therapeutic approach for patients with periodontitis to alleviate inflammatory bone loss, while promoting bone regeneration.

牙周炎是一种细菌驱动的炎症性骨质疾病，影响美国47％的成年人。 口腔病原体诱导炎性细胞因子（包括IL-1β，IL-6，TNF-α和RANKL）的产生， 在RANKL刺激下，这些单核细胞可以进一步 分化和融合形成多核破骨细胞，从而导致肺泡骨质流失。同时， 与牙周炎有关的炎性细胞因子抑制成骨细胞分化和骨再生。 由于目前的治疗方式有限，牙周炎仍然是肺泡骨质流失和牙齿的主要原因 成人损失。我们的长期目标是开发有效的新疗法来治疗该疾病。我们是第一个 证明鞘氨醇-1-磷酸受体2（S1PR2，G蛋白偶联受体）起关键作用 在调节牙周炎的炎症性骨质损失反应时。由S1PR2导向击倒S1PR2 SHRNA或通过其特异性抑制剂对S1PR2的抑制（JTE013）降低了PI3K，NF-κB和MAPKS蛋白 由口腔病原体聚集型放线菌（AA）诱导的激酶，并改善了IL-1β，IL- 6，由AA诱导的TNF-α水平。此外，用S1PR2 shRNA或JTE013的处理减少了细胞 由RANKL诱导的粘附单元（足组成分），抑制破骨细胞生成和骨骼分辨率 由Rankl诱导。 JTE013的口服局部给药可缓解C57BL/6小鼠的炎症性骨质流失 牙周炎是由结扎的放置诱导的。但是，关于S1PR2如何调节的知识差距 成骨细胞生成和骨再生。这是该应用程序的目的是确定S1PR2的作用 在控制成骨细胞生成和骨再生时。我们的初步数据表明，抑制S1PR2 在成骨培养基中培养的骨髓衍生基质细胞（BMSC）中的JTE013增加 与车辆处理相比，成骨细胞生成。用JTE013的治疗增加了mRNA水平 成骨基因，包括酒精磷酸酶，Runt相关转录因子2（Runx2），骨钙素，骨钙素， 与媒介物处理相比，Osterix和骨形态发生蛋白（BMP）2。另外，处理 JTE013增加了成骨蛋白的水平，包括BMP2，BMP7，BMP受体II，BMP受体 与对照媒介物处理相比，1A和P-SMAD 1/5/9蛋白。那我们假设抑制作用 S1PR2在骨细胞前细胞中的促进骨细胞生成和骨再生。我们的具体目标 将确定1）SHRNA敲低或体外S1PR2的抑制是否会通过 BMP/SMAD信号通路和/或其他信号通路； 2）药物抑制是否 JTE013在患有牙周炎的小鼠中的S1PR2促进炎症骨后肺泡再生 损失。该应用定义了由S1PR2调节在促进成骨细胞生成中的新型信号通路。 它将为牙周炎患者开发一种新型热方法以减轻的基础 炎症性骨质流失，同时促进骨骼再生。