Journal article
Evaluation of different media compositions promoting hepatocyte differentiation in the canine liver organoid model
Heliyon, Vol.10(7), e28420
04/15/2024
DOI: 10.1016/j.heliyon.2024.e28420
PMCID: PMC10999936
PMID: 38590903
Abstract
Organoids are 3-dimensional (3D) self-assembled structures capable of replicating the microanatomy and physiology of the epithelial components of their organ of origin. Adult stem cell (ASC) derived organoids from the liver have previously been shown to differentiate into primarily mature cholangiocytes, and their partial differentiation into functional hepatocytes can be promoted using specific media compositions. While full morphological differentiation of mature hepatocytes from ASCs has not yet been reported for any species, the functional differentiation can be approximated using various media compositions.
Six differentiation media formulations from published studies on hepatic organoids were used for the differentiation protocol. Target species for these protocols were humans, mice, cats, and dogs, and encompassed various combinations and concentrations of four major hepatocyte media components: Bone morphogenetic protein 7 (BMP7), Fibroblast Growth Factor 19 (FGF19), Dexamethasone (Dex), and Gamma-Secretase Inhibitor IX (DAPT). Additionally, removing R-spondin from basic organoid media has previously been shown to drive the differentiation of ASC into mature hepatocytes. Differentiation media (N = 20) were designed to encompass combinations of the four major hepatocyte media components. The preferred differentiation of ASC-derived organoids from liver tissue into mature hepatocytes over cholangiocytes was confirmed by albumin production in the culture supernatant.
Out of the twenty media compositions tested, six media resulted in the production of the highest amounts of albumin in the supernatant of the organoids. The cell lines cultured using these six media were further characterized via histological staining, transmission electron microscopy, RNA in situ hybridization, analysis of gene expression patterns, immunofluorescence, and label-free proteomics. The results indicate that preferential hepatocyte maturation from canine ADC-derived organoids from liver tissue is mainly driven by Dexamethasone and DAPT components. FGF19 did not enhance organoid differentiation but improved cell culture survival. Furthermore, we confirm that removing R-spondin from the media is crucial for establishing mature hepatic organoid cultures.
Details
- Title: Subtitle
- Evaluation of different media compositions promoting hepatocyte differentiation in the canine liver organoid model
- Creators
- Vojtech Gabriel - Iowa State UniversityAddison Lincoln - Iowa State UniversityChristopher Zdyrski - Iowa State UniversityAbigail Ralston - 3D Health Solutions Inc., Ames, IA, USAHannah Wickham - Iowa State UniversitySydney Honold - Iowa State UniversityBasant H. Ahmed - Iowa State UniversityKarel Paukner - Institute of Clinical and Experimental MedicineRyan Feauto - Iowa State UniversityMaria M. Merodio - Iowa State UniversityPablo Piñeyro - Iowa State UniversityDavid Meyerholz - University of IowaKarin Allenspach - Iowa State UniversityJonathan P. Mochel - Solutions Inc.
- Resource Type
- Journal article
- Publication Details
- Heliyon, Vol.10(7), e28420
- Publisher
- Elsevier Ltd
- DOI
- 10.1016/j.heliyon.2024.e28420
- PMID
- 38590903
- PMCID
- PMC10999936
- ISSN
- 2405-8440
- eISSN
- 2405-8440
- Language
- English
- Date published
- 04/15/2024
- Academic Unit
- Pathology
- Record Identifier
- 9984577113802771
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