Mimicking and Manipulating Pancreatic Acinar-to-Ductal Metaplasia in 3-dimensional Cell Culture
Mimicking and Manipulating Pancreatic Acinar-to-Ductal Metaplasia in 3-dimensional Cell Culture
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DOI：
10.3791/59096
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发表时间：
2019-02-01
影响因子：
1.2
通讯作者：
Storz, Peter
中科院分区：
文献类型：
--
作者：
Martinez, Alicia K. Fleming;Storz, Peter
在胰腺炎和胰腺癌早期发展过程中，腺泡细胞向导管细胞的分化是一个需要进一步研究的关键过程。为了了解腺泡到导管化生（ADM）的调节机制，体外3D培养和原代腺泡细胞分化为导管细胞比其他系统有许多优势。利用本研究的技术，蛋白表达的调节简单而快速，仅需一天的时间即可分离、刺激或病毒感染，并开始培养原代腺泡细胞以研究ADM过程。与使用基底膜基质相比，在I型胶原细胞外基质中播种腺泡细胞簇，可使腺泡细胞在操作前保持其腺泡特性。在测试各种成分对诱导adv的贡献时，这是至关重要的。通过该技术不仅可以测试细胞因子或其他异位给药因素的影响，而且可以通过使用腺病毒或慢病毒载体对原代腺泡细胞进行病毒感染来测试常见突变、蛋白质表达增加或蛋白质表达抑制的贡献。此外，可以在终点从胶原或基底膜基质中重新分离细胞并分析蛋白质表达。
The differentiation of acinar cells to ductal cells during pancreatitis and in the early development of pancreatic cancer is a key process that requires further study. To understand the mechanisms regulating acinar-to-ductal metaplasia (ADM), ex vivo 3D culture and differentiation of primary acinar cells to ductal cells offers many advantages over other systems. With the technique herein, modulation of protein expression is simple and quick, requiring only one day to isolate, stimulate or virally infect, and begin culturing primary acinar cells to investigate the ADM process. In contrast to using basement membrane matrix, the seeding of acinar cell clusters in collagen I extracellular matrix, allows acinar cells to retain their acinar identity before manipulation. This is vital when testing the contribution of various components to the induction of ADM. Not only are the effects of cytokines or other ectopically administered factors testable through this technique, but the contribution of common mutations, increased protein expression, or knockdown of protein expression is testable via viral infection of primary acinar cells, using adenoviral or lentiviral vectors. Moreover, cells can be re-isolated from collagen or basement membrane matrix at the endpoint and analyzed for protein expression.