Generation and Culture of Lingual Organoids Derived from Adult Mouse Taste Stem Cells.
Generation and Culture of Lingual Organoids Derived from Adult Mouse Taste Stem Cells.
复制标题
DOI：
10.3791/62300
复制
发表时间：
2021-04-05
期刊：
影响因子：
--
通讯作者：
Barlow LA
中科院分区：
文献类型：
--
作者：
Shechtman LA;Piarowski CM;Scott JK;Golden EJ;Gaillard D;Barlow LA
味觉是由舌头上的味蕾调节的，味蕾由快速更新的味觉感受器细胞(TrCs)组成。这种持续的更替是由当地的祖细胞提供动力的，使得味觉功能容易受到多种医疗治疗的干扰，这反过来又严重影响生活质量。因此，在药物治疗的背景下研究这一过程对于了解味觉前体功能和TRC的产生是否以及如何受到影响至关重要。考虑到伦理问题和人类味觉组织的有限可获得性，老鼠模型通常被使用，它的味觉系统与人类相似。与耗时、昂贵、不适合高通量研究的体内方法相比，小鼠语言类器官可以使实验快速运行，重复次数多，小鼠数量少。在这里，修改了以前发表的方案，并提出了一种标准化的方法，从成年小鼠的环状乳头(CVP)中分离出味觉前体细胞来产生味觉有机体。CVP中的味觉祖细胞表达LGR5，并可通过EGFP荧光激活细胞分选(FACS)从携带Lgr5EGFP-IRES-CreERT2等位基因的小鼠中分离出来。将分离的细胞接种到基于基质凝胶的3D培养系统上，培养12天。在培养的前6天，类有机物通过增殖扩张，然后进入分化阶段，在此期间它们产生所有三种味觉细胞类型以及非味觉上皮细胞。成熟后的第12天或生长过程中的任何时间都可以收获类有机物质，用于RNA表达和免疫组织化学分析。从成人干细胞生产舌类器官的标准化培养方法将提高重复性，并推动舌类器官作为一种强大的药物筛选工具来帮助经历味觉障碍的患者。
The sense of taste is mediated by taste buds on the tongue, which are composed of rapidly renewing taste receptor cells (TRCs). This continual turnover is powered by local progenitor cells and renders taste function prone to disruption by a multitude of medical treatments, which in turn severely impacts the quality of life. Thus, studying this process in the context of drug treatment is vital to understanding if and how taste progenitor function and TRC production are affected. Given the ethical concerns and limited availability of human taste tissue, mouse models, which have a taste system similar to humans, are commonly used. Compared to in vivo methods, which are time-consuming, expensive, and not amenable to high throughput studies, murine lingual organoids can enable experiments to be run rapidly with many replicates and fewer mice. Here, previously published protocols have been adapted and a standardized method for generating taste organoids from taste progenitor cells isolated from the circumvallate papilla (CVP) of adult mice is presented. Taste progenitor cells in the CVP express LGR5 and can be isolated via EGFP fluorescence-activated cell sorting (FACS) from mice carrying an Lgr5EGFP-IRES-CreERT2 allele. Sorted cells are plated onto a matrix gel-based 3D culture system and cultured for 12 days. Organoids expand for the first 6 days of the culture period via proliferation and then enter a differentiation phase, during which they generate all three taste cell types along with non-taste epithelial cells. Organoids can be harvested upon maturation at day 12 or at any time during the growth process for RNA expression and immunohistochemical analysis. Standardizing culture methods for production of lingual organoids from adult stem cells will improve reproducibility and advance lingual organoids as a powerful drug screening tool in the fight to help patients experiencing taste dysfunction.
登录
查看更多内容
人类多能干细胞的肾单位祖细胞和肾脏器官的产生。
影响因子：
14.8
作者：
Morizane R;Bonventre JV
通讯作者：
Bonventre JV
味觉的细胞生物学。
影响因子：
7.8
作者：
Chaudhari, Nirupa;Roper, Stephen D.
通讯作者：
Roper, Stephen D.
影响因子：
14.8
作者：
Dekkers, Johanna F.;Alieva, Maria;Rios, Anne C.
通讯作者：
Rios, Anne C.
DOI：
10.1073/pnas.2001833118
发表时间：
2021-01-12
影响因子：
11.1
作者：
Lin, Xiaoli;Lu, Chanyi;Jiang, Peihua
通讯作者：
Jiang, Peihua
DOI：
10.1073/pnas.1409064111
发表时间：
2014-11-18
影响因子：
11.1
作者：
Ren, Wenwen;Lewandowski, Brian C.;Jiang, Peihua
通讯作者：
Jiang, Peihua