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，我们在当前拨款方面的工作已取得以下结果：a）单个小鼠腮腺细胞 发现形成三维 (3D) 细胞簇，显示 TJ 和激动剂诱导的分泌反应 当在生长因子减少的基质胶 (GFR-MG) 与纤维蛋白水凝胶 1 (FH) 组合上生长时，b) GRF-MG 和 FH 的比例得到优化，最终目标是创建功能性且临床安全的 支架1，c）我们的唾液细胞分离方法得到改进，以维持分泌颗粒，形成TJ和 促进细胞存活，d) 小鼠颌下腺 (mSMG) 细胞，而不是腮腺或舌下腺， 被发现是创建显示有组织形态的唾液细胞簇的最佳选择，e) 关键 GFR-MG 中发现了增强唾液腺 (SG) 的成分（即 EGF 和 IGF-1） 聚合为 FH 时的分化，f) EGF 和 IGF-1 被发现无法独立分化 产生有组织的细胞簇；然而，这个目标是使用 Laminin-1 (L1), g) 肽实现的 （对应于促进完整 SG 形成的四个 L1 区域）已合成并与 FH、h 缀合 特定的 L1 肽诱导大鼠 protid 细胞系 Par-C10 中腺泡球的形成（与 细胞仅在 FH 上生长），i) 添加来自人毛囊间充质干细胞的条件培养基 (hHF-MSC CM) 改善 mSMG 细胞的唾液细胞簇组织和管腔形成，j) 唾液细胞 簇维持腺泡、导管和肌上皮细胞，同时响应分泌激动剂，k) 腔 hHF-MSC CM 中 FGF10 的阻断阻碍了 FGF10 的形成，l) 在 hHF-MSC CM 中检测到大量 FGF2 hHF-MSC CM，其中生长有完整的唾液细胞簇，m) L1 肽与化学缀合 与单独的 FH 相比，将荧光 FH 应用于体内受伤的 mSMG 以形成新的唾液组织 或没有支架，并且治疗后小鼠是健康的，以上研究表明我们。 在这笔赠款的前三年内，我们已经实现了大部分最初的目标，并且发现可行 针对遇到的困难的替代方案（例如，使用L1肽来增强FH并改善唾液细胞 簇的形成，如上所述）在接下来的资助期内，我们将完善我们的设计以实现临床安全。 环境如下：a) 产生用 L1 肽和生长因子修饰的 FH，b) 生长分化 c) 使用改良的 FH 支架在体内形成新的功能性组织 （目的是稍后将这些发现应用于因干燥综合征以及 头颈部 γ 射线治疗）。