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胰成人™类器官培养基（人）

人胰腺类器官起始、生长和建立的培养基
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产品号 #（选择产品）

产品号 #100-0820_C

人胰腺类器官起始、生长和建立的培养基

产品优势

  • 在不同的供体组织中启动和扩展正常和肿瘤来源的类器官
  • 通过去除EGF从kras激活的癌细胞中消耗正常细胞
  • 使用无血清维持培养基减少药物筛选的干扰和可变性
  • 使用灵活的培养格式进行高通量药物筛选

产品组分包括

  • 胰成人™类器官起始培养基（人）（目录#100-0820）
    • 胰成人™类器官基础培养基（人），95 mL
    • 胰成人™类器官生长补充剂（人），5ml
    • 类器官补充剂，50毫升
  • 胰成人™类器官生长培养基（人）（目录#100-0781）
    • 胰成人™类器官基础培养基（人），95 mL
    • 胰成人™类器官生长补充剂（人），5ml
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Products for Your Protocol
To see all required products for your protocol, please consult the Protocols and Documentation.
  1. Prostaglandin E2
    Prostaglandin E2

    Prostanoid pathway activator; Activates prostaglandin receptors EP1, EP2, EP3 and EP4

  2. Y-27632 (Dihydrochloride)
    Y-27632 (Dihydrochloride)

    RHO/ROCK pathway inhibitor; Inhibits ROCK1 and ROCK2

  3. Human Recombinant EGF
    Human Recombinant EGF

    Epidermal growth factor

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实验数据
产品说明书及文档
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概述

可靠地培养胰管类器官，以研究胰管上皮的功能，模拟胰腺疾病，如癌症或囊性纤维化，并进行靶向药物筛选或毒性试验。使用胰管™（人）培养基系统，您有一个完整而稳健的工作流程来建立、扩展和维持胰管类器官。

胰腺类器官可以在胰成人™类器官起始培养基（OIM）中从新鲜或冷冻分离的组织中建立，该培养基经过优化，可提供更高的组织类器官形成率。然后，类器官可以在胰成人™类器官生长培养基（OGM）中培养，进行长期维持、传代、无血清药物筛选或冷冻保存，以备将来的实验。从培养基中选择性去除EGF支持kras激活的癌症类器官培养中正常细胞的消耗。

使用胰成人™（人）培养基系统培养的类器官可以适应各种培养方案，包括二维Transwell®单层培养，稀释康宁®Matrigel®悬浮培养和高通量测定兼容的培养格式。了解如何设置人类胰腺类器官以高通量格式进行药物测试最新的协议 .

如果您打算将此产品用于商业用途，请通过www.huborganoids.nl获取商业用途许可证或HUB许可的相关说明。

Subtype
Specialized Media
 
Cell Type
Pancreatic Cells
 
Species
Human
 
Application
Cell Culture, Expansion, Maintenance, Organoid Culture
 
Brand
PancreaCult
 
Area of Interest
Drug Discovery and Toxicity Testing, Epithelial Cell Biology, Organoids, Stem Cell Biology
 
Formulation Category
Serum-Free
 

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Data Figures

Human Pancreatic Duct Organoids Grown using PancreaCult™ (Human)

Figure 1. Human Pancreatic Duct Organoids

PancreaCult™ Organoid Initiation Medium (OIM; Human) and Organoid Growth Medium (OGM; Human) support the initiation and expansion of human pancreatic duct organoids from pancreatic tissue or previously established organoid cultures. Shown are organoids grown using PancreaCult™ OIM and OGM and imaged on day 7 of passage 3.

Workflow for Generating Pancreatic Ductal Organoids Using PancreaCult™ (Human)

Figure 2. PancreaCult™ Human Enables Initiation and Expansion of Pancreatic Ductal Organoids

Human pancreatic ductal organoids are initiated in PancreaCult™ OIM for the first 3 days of culture, then switched to PancreaCult™ OGM for the remainder of culture. Alternatively, organoids can be grown in PancreaCult™ OGM for a completely serum-free protocol, however, proliferative pancreatic ductal cells are better supported in PancreaCult™ OIM during the first 3 days of culture. Cultures should be passaged after 7 days with further medium changes every 2-3 days. Pancreatic ductal organoids are suitable for experimentation or banking after one full passage in PancreaCult™ OGM. For full culture instructions please refer to the product manual (Document #10000011617)

PancreaCult™ (Human) Provides Robust Expansion of Pancreatic Duct Organoids

Figure 3. PancreaCult™ (Human) Provides Robust Expansion of Pancreatic Duct Organoids

(A) Organoid expansion in PancreaCult™ OGM provides robust expansion of organoids across different donors. (B) Organoids may be initiated in PancreaCult™ OGM for a completely serum-free workflow, however, initiation and maintenance of damaged tissue is better supported by initiating cultures in PancreaCult™ OIM. (C) Comparison of PancreaCult™ Human to two different DIY formulations showed more robust expansion of organoids in PancreaCult™ Human. Shown is the cumulative fold expansion of organoid fragments as counted at the end of each passage.

Pancreatic Duct Organoids Display Features of the Pancreatic Ductal Epithelium by ICC Staining

Figure 4. Pancreatic Duct Organoids Display Features of the Pancreatic Ductal Epithelium

Organoids grown using the PancreaCult™ (Human) display marker expression consistent with the pancreatic ductal epithelium when imaged using immunocytochemistry. Shown are organoids grown in PancreaCult™ OGM and stained for (A) pancreatic ductal marker CK19, (B) pancreatic ductal marker SOX9, (C) epithelial marker EPCAM, (D) proliferation marker KI67, (E) apical pancreatic duct marker MUC1, and (F) pancreatic ductal marker CA2. Organoids were imaged on passage 2 (A), passage 3 (B, C) or passage 10 (D-F).

Pancreatic Duct Organoids Cultured With PancreaCult™ Human Show Pancreatic Marker Expression Levels Similar to Exocrine Tissue

Figure 5. Pancreatic Duct Organoids Cultured With PancreaCult™ Human Show Pancreatic Marker Expression Levels Similar to Exocrine Tissue

Pancreatic duct organoids grown using the PancreaCult™ Human show marker expression levels similar to those observed in exocrine tissue. Analysis by qPCR showed pancreatic duct organoids were enriched for (A) PDX1 (C) CK19 and (F) LGR5 as compared to total pancreatic tissue, demonstrating enrichment of proliferative duct organoids. Comparable expression of (B) SOX9, (D) cystic fibrosis transmembrane receptor (CFTR), and (E) CA2 was observed in pancreatic duct organoids. Expression levels are normalized to TBP and UBC housekeeping genes (ΔCT) and total pancreas for relative expression levels (ΔΔCT).

PancreaCult™ (Human) Efficiently Maintains the Long-Term Growth and Genetic Profile of PDAC (Pancreatic Ductal Adenocarcinoma) Organoids

Figure 6. PancreaCult™ (Human) Efficiently Maintains the Long-Term Growth and Genetic Profile of PDAC (Pancreatic Ductal Adenocarcinoma) Organoids

(A) Pre-established PDAC organoid lines can be maintained and expanded in PancreaCult™ OGM for at least 10 passages (n = 4) and demonstrate comparable growth with PDAC organoid lines maintained in DIY media. *DIY medium culture for this line was terminated after 10 passages. (B) Pre-established PDAC organoids cultured in PancreaCult™ OGM for 5 passages retain somatic SNPs and indels in oncogenic driver and tumor suppressor genes from the parent PDAC organoid line. Whole-exome sequencing data was collected before (teal) and after 5 passages of PDAC organoid cultures in PancreaCult™ OGM (orange) or the DIY media (maroon). Grey and white boxes indicate the overall presence or absence of SNPs or indels in the indicated genes compared to the parent PDAC organoid line pre-established in DIY media. #TP53 reads were undetectable, indicating gene deletion.

PDAC Organoid Line-Specific Differences in GATA6 Expression, Morphology, and Chemotherapy Drug Response

Figure 7. PDAC Organoid Line-Specific Differences in GATA6 Expression, Morphology, and Chemotherapy Drug Response

PDAC organoid lines (PDAC1-3) grown using PancreaCult™ (Human) demonstrate line-specific differences in (A) GATA6 (GATA-binding factor 6) gene expression and (B) morphology. Expression levels are normalized to housekeeping genes (ΔCT) and primary pancreatic tissue for relative expression levels (ΔΔCT). PDAC1 and PDAC2 lines were expanded in PancreaCult™ OGM from pre-established lines while normal organoids and the PDAC3 line were established with PancreaCult™ OIM from dissociated primary tissue and maintained in PancreaCult™ OGM. PDAC3 expansion was facilitated by reduced oxygen culture (5%) and maintained in the absence of EGF. Scale bar = 500 μm. (C) PDAC organoid lines were cultured in 96-well plates within Matrigel® domes or layer cultures and treated with irinotecan (left) or 5-fluorouracil (right) and compared to 0.1% DMSO vehicle control. Full-medium changes with fresh compounds were performed each day and analyses were performed 24 hours after the final fresh compound addition (Day 3). Cell viability for treated, vehicle-treated, and untreated PDAC organoid lines was assessed using the CellTiter-Glo® 3D Cell Viability Assay (Promega Catalog #G9681). Irrespective of layer or dome culture format, PDAC organoid lines demonstrate line-specific differences in chemotherapy drug response. Error bars = SD.

Removing Epidermal Growth Factor (EGF) Abrogates Normal Pancreatic Duct Organoid Growth but Does Not Affect PDAC Organoids Cultured in PancreaCult™ (Human)

Figure 8. Removing Epidermal Growth Factor (EGF) Abrogates Normal Pancreatic Duct Organoid Growth but Does Not Affect PDAC Organoids Cultured in PancreaCult™ (Human)

(A) EGF removal efficiently suppresses normal pancreatic duct organoid growth within the first passage. (B) PDAC organoids were established in PancreaCult™ OIM and maintained in PancreaCult™ OGM for 8 passages in low-oxygen culture (5%) before removing EGF. EGF-depleted PDAC organoids maintained growth for at least 2 passages, indicating the presence of KRAS-activated tumor cells. Scale bar = 500 μm.

Organoids Can Be Established from Cryopreserved PDAC Cells Using PancreaCult™ (Human)

Figure 9. Organoids Can Be Established from Cryopreserved PDAC Cells Using PancreaCult™ (Human)

Mutational profile of organoids established and expanded in PancreaCult™ (Human) (Low O2, no EGF). Orange areas indicate areas of gains (top row) and grey areas indicate areas of losses (middle row) in the indicated chromosomes. There is a lack of high certainty loss of heterozygosity (LOH, bottom row). Microarray analysis was performed with Illumina iScan on the Global Diversity Array with Cytogenetics-8 (v1.0). CNVs (gene copy number variations) were called using the MoChA software tool for mosaic chromosomal alterations detection and analysis, MoChA, with human genome hg38 used as a reference.

Protocols and Documentation

Find supporting information and directions for use in the Product Information Sheet or explore additional protocols below.

Document Type
Product Name
Catalog #
Lot #
Language
Document Type
Product Information Sheet
Product Name
PancreaCult™ Organoid Initiation Medium (Human)
Catalog #
100-0820
Lot #
All
Language
English
Document Type
Product Information Sheet
Product Name
PancreaCult™ Organoid Growth Medium (Human)
Catalog #
100-0781
Lot #
All
Language
English
Document Type
Technical Manual
Product Name
PancreaCult™ Organoid Media (Human)
Catalog #
100-0820
Lot #
All
Language
English
Document Type
Technical Manual
Product Name
PancreaCult™ Organoid Growth Medium (Human)
Catalog #
100-0781
Lot #
All
Language
English
Document Type
Safety Data Sheet 1
Product Name
PancreaCult™ Organoid Initiation Medium (Human)
Catalog #
100-0820
Lot #
All
Language
English
Document Type
Safety Data Sheet 2
Product Name
PancreaCult™ Organoid Initiation Medium (Human)
Catalog #
100-0820
Lot #
All
Language
English
Document Type
Safety Data Sheet 3
Product Name
PancreaCult™ Organoid Initiation Medium (Human)
Catalog #
100-0820
Lot #
All
Language
English
Document Type
Safety Data Sheet 1
Product Name
PancreaCult™ Organoid Growth Medium (Human)
Catalog #
100-0781
Lot #
All
Language
English
Document Type
Safety Data Sheet 2
Product Name
PancreaCult™ Organoid Growth Medium (Human)
Catalog #
100-0781
Lot #
All
Language
English

Applications

This product is designed for use in the following research area(s) as part of the highlighted workflow stage(s). Explore these workflows to learn more about the other products we offer to support each research area.

Research Area
Workflow Stages
Workflow Stages for Organoids
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Resources and Publications

Educational Materials (7)

How to Process Epithelial Organoids and Organoid-Derived Epithelial Monolayers for RNA Isolation
Protocol
How to Process Epithelial Organoids and Organoid-Derived Epithelial Monolayers for RNA Isolation
Drug Screening with Human Pancreatic Organoid Cultures
Protocol
Drug Screening with Human Pancreatic Organoid Cultures
Tools to Establish and Maintain Pancreatic Organoid Cultures
11:22
Webinar
Tools to Establish and Maintain Pancreatic Organoid Cultures
Generating Pancreatic Organoids from Healthy Murine and Human Pancreatic Ducts
15:48
Webinar
Generating Pancreatic Organoids from Healthy Murine and Human Pancreatic Ducts
A Bench-to-Bedside Approach to Overcome Recurrence in PDAC
20:51
Webinar
A Bench-to-Bedside Approach to Overcome Recurrence in PDAC
Robust Establishment and Expansion of Human Pancreatic Duct Organoids
Scientific Poster
Robust Establishment and Expansion of Human Pancreatic Duct Organoids
Dr. Bas Trietsch
Interview
Dr. Bas Trietsch

Publications (1)

Oncogenic KRAS-driven metabolic reprogramming in pancreatic cancer cells utilizes cytokines from the tumor microenvironment P. Dey et al. Cancer Discovery 2020 4

Abstract

A hallmark of pancreatic ductal adenocarcinoma (PDAC) is an exuberant stroma comprised of diverse cell types that enable or suppress tumor progression. Here, we explored the role of oncogenic KRAS in protumorigenic signaling interactions between cancer cells and host cells. We show that KRAS mutation (KRAS) drives cell-autonomous expression of type I cytokine receptor complexes (IL2r?–IL4r? and IL2r?–IL13r?1) in cancer cells that in turn are capable of receiving cytokine growth signals (IL4 or IL13) provided by invading Th2 cells in the microenvironment. Early neoplastic lesions show close proximity of cancer cells harboring KRAS and Th2 cells producing IL4 and IL13. Activated IL2r?–IL4r? and IL2r?–IL13r?1 receptors signal primarily via JAK1-STAT6. Integrated transcriptomic, chromatin occupancy, and metabolomic studies identified MYC as a direct target of activated STAT6 and that MYC drives glycolysis. Thus, paracrine signaling in the tumor microenvironment plays a key role in the KRAS-driven metabolic reprogramming of PDAC. SIGNIFICANCE: Type II cytokines, secreted by Th2 cells in the tumor microenvironment, can stimulate cancer cell-intrinsic MYC transcriptional upregulation to drive glycolysis. This KRAS-driven heterotypic signaling circuit in the early and advanced tumor microenvironment enables cooperative protumorigenic interactions, providing candidate therapeutic targets in the KRAS pathway for this intractable disease.
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Species Human
Formulation Category Serum-Free
Legal Statement: This product was developed under a license to intellectual property owned by Hubrecht Organoid Technology (HUB). This product is sold for research use only. Purchase of this product does not include the right to use it for drug screening aiming for commercial gain, equipment validation, biobanking, or for other commercial purposes. Purchasers wishing to use the product for purposes other than basic research use should contact HUB at www.huborganoids.nl to obtain a further license. Purchasers may apply for a License from HUB, which will not be unreasonably withheld by HUB. PRODUCTS ARE FOR RESEARCH USE ONLY AND NOT INTENDED FOR HUMAN OR ANIMAL DIAGNOSTIC OR THERAPEUTIC USES UNLESS OTHERWISE STATED. FOR ADDITIONAL INFORMATION ON QUALITY AT STEMCELL, REFER TO WWW.STEMCELL.COM/COMPLIANCE.
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