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使用3D悬浮和甲基纤维素基培养系统研究成人胰腺导管祖细胞样细胞的方案。

Protocol to study ductal progenitor-like cells from the adult human pancreas using 3D suspension and methylcellulose-based culture systems.

作者信息

Zook Heather N, Quijano Janine C, Ortiz Jose A, Donohue Cecile, Erdem Neslihan, Ku Hsun Teresa

机构信息

Department of Translational Research and Cellular Therapeutics, Arthur Riggs Diabetes and Metabolism Research Institute, City of Hope, Duarte, CA 91010, USA.

Department of Translational Research and Cellular Therapeutics, Arthur Riggs Diabetes and Metabolism Research Institute, City of Hope, Duarte, CA 91010, USA.

出版信息

STAR Protoc. 2025 Jun 20;6(2):103847. doi: 10.1016/j.xpro.2025.103847. Epub 2025 May 26.

DOI:10.1016/j.xpro.2025.103847
PMID:40424137
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC12152497/
Abstract

Primary human ductal progenitor-like cells derived from donated pancreas have the potential to serve as a source of therapeutic insulin-producing beta cells for the treatment of diabetes. Here, we present a protocol for studying ductal progenitor-like cells using a good manufacturing practice (GMP)-compatible 3D suspension culture system and a methylcellulose- and Matrigel-based 3D colony assay. We describe steps for dispersing and cryopreserving human pancreatic cells and initiating and maintaining cultures. We then detail how to prepare ductal spheroids or colonies for downstream applications. For complete details on the use and execution of this protocol, please refer to Zook et al. and Quijano et al..

摘要

源自捐赠胰腺的原代人导管样祖细胞有潜力成为治疗糖尿病的胰岛素分泌β细胞的治疗来源。在此，我们展示了一种使用符合药品生产质量管理规范（GMP）的3D悬浮培养系统以及基于甲基纤维素和基质胶的3D集落测定法来研究导管样祖细胞的方案。我们描述了分散和冷冻保存人胰腺细胞以及启动和维持培养的步骤。然后，我们详细说明了如何为下游应用制备导管球体或集落。有关本方案使用和执行的完整详细信息，请参考佐克等人以及基哈诺等人的研究。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/26bc656bf179/gr8.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/d0c890a407c1/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/cf872d53ec08/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/c1ac5414a2ec/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/a8e30b9b87ad/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/bb9b7a77f9cc/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/b84a518a18b1/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/6f9ba930b8cc/gr6.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/a60cd3396ebe/gr7.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/26bc656bf179/gr8.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/d0c890a407c1/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/cf872d53ec08/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/c1ac5414a2ec/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/a8e30b9b87ad/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/bb9b7a77f9cc/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/b84a518a18b1/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/6f9ba930b8cc/gr6.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/a60cd3396ebe/gr7.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/fa80/12152497/26bc656bf179/gr8.jpg

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本文引用的文献

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Activation of ductal progenitor-like cells from adult human pancreas requires extracellular matrix protein signaling.从成人胰腺中激活导管祖细胞样细胞需要细胞外基质蛋白信号传导。
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A matrigel-free method for culture of pancreatic endocrine-like cells in defined protein-based hydrogels.一种在特定蛋白基水凝胶中培养胰腺内分泌样细胞的无基质胶方法。
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Methylcellulose colony assay and single-cell micro-manipulation reveal progenitor-like cells in adult human pancreatic ducts.
甲基纤维素集落测定和单细胞微操作揭示成人胰腺导管中的祖细胞样细胞。
Stem Cell Reports. 2023 Mar 14;18(3):618-635. doi: 10.1016/j.stemcr.2023.02.001. Epub 2023 Mar 2.
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A 3D system to model human pancreas development and its reference single-cell transcriptome atlas identify signaling pathways required for progenitor expansion.一个用于模拟人类胰腺发育的 3D 系统及其参考单细胞转录组图谱确定了祖细胞扩增所需的信号通路。
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