Salivary gland ionocyte organization and function during homeostasis, repair, and disease

稳态、修复和疾病期间唾液腺离子细胞的组织和功能

• 批准号: 10628001
• 负责人: Helen P. Makarenkova
• 金额: $ 59.64万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-18 至 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10628001
• 关键词: Acinar Cell; Acinus organ component; Adult; Affect; Anions; Biology; Cell Separation; Cells; Characteristics; Chronic; Clinical Research; Collaborations; Complex; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Data; Disease; Disease Progression; Duct (organ) structure; Ductal Epithelial Cell; Ductal Epithelium; Electrolytes; Enzymes; Epithelial Cells; Epithelium; Exocrine Glands; FGF10 gene; Fibroblast Growth Factor; Fluids and Secretions; Fluorescence-Activated Cell Sorting; Functional disorder; Future; Gene Expression; Genes; Gland; Heterogeneity; Homeostasis; Immunoglobulins; Inflammation; Injury; Intercalated Duct; Ion Transport; Ions; Knockout Mice; Maintenance; Mesenchymal; Mesenchyme; Modeling; Mucins; Mus; Myoepithelial cell; Natural regeneration; Parotid Gland; Play; Population; Process; Proliferating; Proteins; Proton-Translocating ATPases; RNA analysis; Regulation; Research; Role; Saliva; Salivary; Salivary Glands; Salivary duct structure; Signal Transduction; Sjogren' s Syndrome; Sorting; Tamoxifen; Testing; Tetanus Helper Peptide; Thrombospondin 1; Tissues; Tomatoes; Transplantation; UXT gene; biomarker identification; cell type; experimental study; fibroblast growth factor receptor 2b; functional improvement; gland development; human disease; human tissue; injured; mouse model; novel; oral tissue; overexpression; postnatal; postnatal development; repaired; single cell analysis; single-cell RNA sequencing; stem cell growth; transcriptome sequencing; vacuolar H+-ATPase

The salivary glands are exocrine glands that produce saliva. An adequate supply of saliva is critical to the maintenance of oral tissue. Saliva is composed of a variety of electrolytes and also contains immunoglobulins, proteins, enzymes and mucins, suggesting that saliva has a complex function. An important function of salivary gland ducts is to regulate ion homeostasis[1]. Dysfunction of epithelial ion homeostasis can lead to several human diseases[2, 3]. Our lineage tracing experiments using a tamoxifen inducible FGF10CreERT2:R26- Tomato mouse identified new epithelial populations of the Fgf10 expressing cells in the striated (SD), granular (GCT) and excretory ducts (ED) of submandibular (SMG) and parotid (PG) glands. Importantly Fgf10 lineage tracing shows that until postnatal day 7 (P7) only mesenchymal Fgf10 expressing cells are present, however after P10, Fgf10 expression in the mesenchyme shuts down, while a subset of ductal epithelial cells starts to express Fgf10. Using Fluorescence Activated Cell Sorting (FACS) we isolated both mesenchymal and epithelial populations of Fgf10+ cells and performed RNA sequencing (RNA-seq). Analysis of this RNA- seq data combined with analysis of single cell RNA-Seq (scRNAseq) data from P1 and adult SMGs showed that epithelial Fgf10+ populations overlap with the known duct population marked by Ascl3. Moreover, these epithelial Fgf10+ cells expressed many gene characteristics for an ancient type of ionocyte cell: Bsnd, Foxi1, Foxi2, Asgr1, Stap1, several subunits of the H+- ATPase (V-ATPase) and markers of SLC12A1/2 and Slc9a4 transporters. Most importantly Fgf10+ cells expressed cystic fibrosis transmembrane conductance regulator (Cftr) that plays a key role in exocrine secretion, including in salivary glands. We propose that these epithelial Fgf10+/Ascl3+/Cftr+ cells are specialized duct salivary gland ionocytes. Moreover, using two models of Sjogrens syndrome: the NOD.B10Sn-H2b/J and the thrombospondin-1 null (TSP1-/-) mice, we showed a significant decrease in Fgf10+ expression and the number of ionocytes with disease progression. We also showed that epithelial Fgf10+ cells isolated from the SMG are able to maintain proliferation and growth of progenitor and myoepithelial cells, suggesting an additional role for ionocytes in ductal maintenance. In this application we will determine the role of Fgf10 signaling in SMG regeneration and ionocyte function, define factors that control ionocyte differentiation and determine how chronic inflammation affects ionocyte function. In addition, in collaboration with Dr. Hoffman's group using bulk RNA-seq and scRNA-seq data we will investigate whether salivary glands have one type or several types of ionocytes. Our proposed study will provide a starting platform for future studies of ionocytes in basic biology and clinical research.

唾液腺是产生唾液的外分泌腺。足够的供应 唾液对于维持口腔组织至关重要。唾液由多种电解质组成 还含有免疫球蛋白，蛋白质，酶和粘蛋白，表明唾液具有 复杂函数。唾液腺管道的重要功能是调节离子 稳态[1]。上皮离子稳态的功能障碍会导致几个人类 疾病[2，3]。我们使用他莫昔芬诱导FGF10CREERT2：R26-的谱系追踪实验 番茄小鼠鉴定了纹状体中FGF10表达细胞的新上皮种群 （SD），颗粒（GCT）和排泄管（ED）（SMG）和腮腺（PG） 腺体。重要的是，FGF10谱系跟踪表明，直到产后第7天（P7） 间充质FGF10表达细胞存在，但是在P10之后，FGF10表达在 间质关闭，而导管上皮细胞的一部分开始表达FGF10。使用 荧光活化的细胞分选（FACS）我们分离了间质和上皮 FGF10+细胞的种群和进行RNA测序（RNA-Seq）。该RNA-的分析 SEQ数据与P1和成人的单细胞RNA-Seq（Scrnaseq）数据的分析结合 SMG表明上皮FGF10+种群与已知的导管种群重叠 由ASCL3。此外，这些上皮FGF10+细胞表达了许多基因特征 古代离子细胞细胞：BSND，FOXI1，FOXI2，ASGR1，STAP1，H+ - 几个亚基 ATPase（V-ATPase）和SLC12A1/2和SLC9A4转运蛋白的标记。最重要的是 FGF10+细胞表达囊性纤维化跨膜电导调节剂（CFTR） 在外分泌分泌中的关键作用，包括唾液腺。我们建议这些上皮 FGF10+/ASCL3+/CFTR+细胞是专门的唾液腺离子细胞。而且，使用两个 Sjogrens综合征的模型：NOD.B10SN-H2B/J和drombospondin-1 null（TSP1 - / - ） 小鼠，我们显示出FGF10+表达的显着降低，并且具有与 疾病进展。我们还表明，从SMG分离的上皮FGF10+细胞是 能够维持祖细胞和肌上皮细胞的增殖和生长，这表明 离子细胞在导管维持中的其他作用。在此应用程序中，我们将确定角色 SMG再生和离子细胞功能中FGF10信号传导的定义因素 离子细胞分化，并确定慢性炎症如何影响离子细胞功能。在 此外，使用散装RNA-Seq和Scrna-Seq数据与Hoffman博士的小组合作，我们 将研究唾液腺是否具有一种或几种类型的离子细胞。我们的 拟议的研究将为将来研究基本生物学的离子细胞研究提供一个启动平台 和临床研究。