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丢失疾病，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， 其将单核细胞吸引到牙龈组织。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，导致牙槽骨丢失。同时， 与牙周炎相关的炎性细胞因子抑制成骨细胞分化和骨再生。 在目前有限的治疗方式下，牙周炎仍然是导致牙槽骨丢失和牙齿脱落的主要原因。 成年人的损失。我们的长期目标是开发有效的新疗法来治疗这种疾病。我们是第一个 表明鞘氨醇-1-磷酸受体2（S1 PR 2，一种G蛋白偶联受体）在 在调节牙周炎炎症性骨丢失反应中的作用。通过S1 PR 2定向的 shRNA或通过其特异性抑制剂（JTE 013）抑制S1 PR 2可降低PI 3 K、NF-κB和MAPK s蛋白 口腔病原菌伴放线菌聚集杆菌（Aa）诱导的IL-1β、IL-10、IL-11、IL-12、IL-13、IL-14、IL-15、IL-16、IL-17、IL-18、IL-19、IL 6、Aa诱导的TNF-α水平。此外，用S1 PR 2 shRNA或JTE 013处理减少了细胞数量。 RANKL诱导的粘附单位（podosome组分），抑制破骨细胞生成和骨吸收 由RANKL诱导。口服局部给药JTE 013减轻C57 BL/6小鼠的炎性骨丢失 结扎引起的牙周炎然而，关于S1 PR 2如何调节 成骨细胞生成和骨再生。因此，本申请的目的是确定S1 PR 2的作用 在控制成骨细胞生成和骨再生中的作用。我们的初步数据表明，S1 PR 2的抑制， 在成骨培养基中培养的骨髓源性基质细胞（BMSCs）中的JTE 013增加 成骨细胞生成与赋形剂治疗相比。用JTE 013处理增加了 成骨基因，包括碱性磷酸酶，Runt相关转录因子2（Runx 2），骨钙素， osterix和骨形态发生蛋白（BMP）2与赋形剂治疗相比。此外，治疗 JTE 013增加成骨蛋白的水平，包括BMP 2、BMP 7、BMP受体II、BMP受体 1A和p-Smad 1/5/9蛋白与对照媒介物处理相比。因此，我们假设抑制 前成骨细胞中S1 PR 2的表达促进成骨细胞生成和骨再生。我们的具体目标 将确定1）体外shRNA敲低或抑制S1 PR 2是否会通过以下途径增加成骨细胞生成： BMP/Smad信号通路和/或其他信号通路的药理学抑制;和2）BMP/Smad信号通路和/或其他信号通路的药理学抑制是否 JTE 013诱导的S1 PR 2促进牙周炎小鼠炎性骨再生 损失本申请定义了由S1 PR 2调节的促进成骨细胞生成的新信号通路。 这将为开发一种新的治疗牙周炎的方法，以减轻牙周炎患者的痛苦奠定基础 炎性骨丢失，同时促进骨再生。