THE USE OF FIBRIN HYDROGELS TO PROMOTE SALIVARY GLAND REGENERATION

使用纤维蛋白水凝胶促进唾液腺再生

• 批准号: 9428224
• 负责人: Stelios Theoharis Andreadis
• 金额: $ 9.16万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-16 至 2018-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9428224
• 关键词: Acinar Cell; Aftercare; Agonist; Autoimmune Diseases; Award; Biomedical Engineering; Blood Vessels; C10; Cancer Etiology; Cell Line; Cell Polarity; Cell Separation; Cell Survival; Cell physiology; Cells; Clinical; Conditioned Culture Media; Dimensions; Dryness; Ductal Epithelial Cell; EGF gene; Ectodermal Dysplasia; Endothelial Cells; Engineering; Environment; Extracellular Matrix; FGF10 gene; FGF2 gene; Fibrin; Functional disorder; Funding; Funding Opportunities; Gland; Goals; Grant; Growth; Growth Factor; Hair follicle structure; Head and Neck Cancer; Head and neck structure; Hereditary Disease; Human; Hydrogels; Immobilization; In Vitro; Insulin-Like Growth Factor I; Lead; Link; Mechanical Stimulation; Mesenchymal Stem Cells; Methods; Morphology; Mus; Myoepithelial cell; Natural regeneration; Oral cavity; Oral health; Parotid Gland; Peptides; Polymers; Rattus; Research; Saliva; Salivary; Salivary Gland Tissue; Salivary Glands; Secretory Vesicles; Sjogren' s Syndrome; Sublingual Gland; Submandibular gland; Surface; Testing; Tissues; Work; chemical conjugate; design; improved; improved functioning; in vivo; irradiation; laminin-1; matrigel; mechanical properties; mouse model; parotid cell; repaired; response; salivary acinar cell; salivary cell; scaffold; translation to humans; tumorigenesis; tyrosyl-isoleucyl-glycyl-seryl-arginine

ABSTRACT Our work to date on the current grant has yielded the following results: a) single mouse parotid cells were found to form three-dimensional (3D) cell clusters displaying TJ and agonist-induced secretory responses when grown on Growth Factor-Reduced Matrigel (GFR-MG) in combination with Fibrin Hydrogel1 (FH), b) the ratio of GRF-MG and FH was optimized with the ultimate goal of creating a functional and clinically safe scaffold1, c) our salivary cell isolation method was improved to maintain secretory granules, form TJ and facilitate cell survival, d) mouse submandibular gland (mSMG) cells, rather than parotid or sublingual glands, were found to be the best choice for creating salivary cell clusters displaying organized morphology, e) critical components were identified within GFR-MG (i.e., EGF and IGF-1) that enhance salivary gland (SG) differentiation when polymerized to FH, f) EGF and IGF-1 were found to be incapable of independently producing organized cell clusters; however, this goal was achieved using Laminin-1 (L1), g) peptides (corresponding to four L1 regions that promote intact SG formation) were synthesized and conjugated to FH, h) specific L1 peptides induced formation of acinar spheres in the rat parotid cell line Par-C10 (as compared to cells grown on FH alone), i) addition of a conditioned medium from human hair follicle mesenchymal stem cells (hHF-MSC CM) improved salivary cell cluster organization and lumen formation in mSMG cells, j) salivary cell clusters maintained acinar, ductal and myoepithelial cells while responding to secretory agonists, k) lumen formation was impeded with the blocking of FGF10 in hHF-MSC CM, l) high amounts of FGF2 were detected in a hHF-MSC CM in which intact salivary cell clusters were grown, m) L1 peptides chemically conjugated to fluorescent FH were applied to wounded mSMG in vivo to form new salivary tissue, as compared to FH alone or no scaffold, and mice were healthy after the treatment. Taken as a whole, the above studies indicate we have accomplished the majority of our initial goals during the first three years of this grant and found viable alternatives for the difficulties encountered (e.g., use of L1 peptides to enhance FH and improve salivary cell clusters formation, as noted above). In the coming grant period, we will refine our design for a clinically safe environment as follows: a) produce a FH modified with L1 peptides and growth factors, b) grow differentiated salivary cell clusters in these matrices and c) use modified FH scaffold to form new and functional tissue in vivo (with aim of later applying these findings to humans with SG dysfunction due to Sjögren's syndrome as well as head and neck γ-irradiation treatments).

抽象的 迄今为止，我们在当前赠款上的工作产生了以下结果：a）单小鼠腮腺细胞是 发现形成三维（3D）细胞簇，显示TJ和激动剂引起的秘密响应 在生长因子还原因子降低Matrigel（GFR-MG）与纤维蛋白水凝胶1（FH）结合时生长时，b） GRF-MG和FH的比率是优化的，其最终目标是创建功能和临床安全 脚手架1，c）改进了我们的唾液细胞分离方法以维持分泌颗粒，形式TJ和 促进细胞存活，d）小鼠下颌腺（MSMG）细胞，而不是腮腺或舌下腺体 被发现是创建唾液细胞簇显示有组织形态的最佳选择，e）关键 在GFR-MG（即EGF和IGF-1）中鉴定出成分，以增强唾液腺（SG） 分化聚合到FH时，发现F）EGF和IGF-1无能力独立 产生有组织的细胞簇；但是，使用层粘连蛋白1（L1），g）肽实现了这一目标 （对应于促进完整SG形成的四个L1区）合成并连接到FH，H） 特异性L1肽在大鼠腮腺细胞系PAR-C10中诱导的腺泡球的形成（与 单独生长在FH上），i）从人毛囊间充质干细胞中添加条件培养基 （HHF-MSC CM）改善MSMG细胞中唾液细胞簇组织和管腔形成，J）唾液细胞 群集在响应秘密激动剂时保持腺泡，导管和肌上皮细胞 在HHF-MSC CM中FGF10的阻塞阻碍了形成，在HHF-MSC CM中，在 HHF-MSC CM，其中完整的唾液细胞簇生长，M）L1肽化学连接到 与单独的FH相比 或没有脚手架，治疗后小鼠很健康。从总体上讲，上述研究表明我们 在这笔赠款的头三年中，已经实现了我们最初的目标，并发现可行 遇到困难的替代方法（例如，使用L1 Pepperides来增强FH并改善唾液细胞 簇形成，如上所述）。在未来的赠款期间，我们将精心设计临床安全的设计 环境如下：a）产生用L1辣椒和生长因子修饰的FH，b）成长分化 这些物质中的唾液细胞簇和c）使用改良的FH支架在体内形成新的和功能性组织 （以后将这些发现应用于Sjögren综合征以及SG功能障碍的人类的目的 头部和颈γ-辐照处理）。