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从人多能干细胞规模化生产类组织血管化的肝类器官。

Scalable production of tissue-like vascularized liver organoids from human PSCs.

机构信息

Hybrid Technology Hub-Centre of Excellence, Institute of Basic Medical Sciences, University of Oslo, Oslo, Norway.

Department of Pediatric Research, Oslo University Hospital, Oslo, Norway.

出版信息

Exp Mol Med. 2023 Sep;55(9):2005-2024. doi: 10.1038/s12276-023-01074-1. Epub 2023 Sep 1.


DOI:10.1038/s12276-023-01074-1
PMID:37653039
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC10545717/
Abstract

The lack of physiological parity between 2D cell culture and in vivo culture has led to the development of more organotypic models, such as organoids. Organoid models have been developed for a number of tissues, including the liver. Current organoid protocols are characterized by a reliance on extracellular matrices (ECMs), patterning in 2D culture, costly growth factors and a lack of cellular diversity, structure, and organization. Current hepatic organoid models are generally simplistic and composed of hepatocytes or cholangiocytes, rendering them less physiologically relevant compared to native tissue. We have developed an approach that does not require 2D patterning, is ECM independent, and employs small molecules to mimic embryonic liver development that produces large quantities of liver-like organoids. Using single-cell RNA sequencing and immunofluorescence, we demonstrate a liver-like cellular repertoire, a higher order cellular complexity, presenting with vascular luminal structures, and a population of resident macrophages: Kupffer cells. The organoids exhibit key liver functions, including drug metabolism, serum protein production, urea synthesis and coagulation factor production, with preserved post-translational modifications such as N-glycosylation and functionality. The organoids can be transplanted and maintained long term in mice producing human albumin. The organoids exhibit a complex cellular repertoire reflective of the organ and have de novo vascularization and liver-like function. These characteristics are a prerequisite for many applications from cellular therapy, tissue engineering, drug toxicity assessment, and disease modeling to basic developmental biology.

摘要

二维细胞培养与体内培养之间缺乏生理一致性,这导致了更类器官型模型的发展。已经为许多组织开发了类器官模型,包括肝脏。目前的类器官方案的特点是依赖于细胞外基质 (ECM)、二维培养中的图案形成、昂贵的生长因子以及缺乏细胞多样性、结构和组织。目前的肝类器官模型通常比较简单,由肝细胞或胆管细胞组成,与原组织相比,它们的生理相关性较低。我们开发了一种不需要二维图案形成、不依赖 ECM 且使用小分子模拟胚胎肝脏发育的方法,从而产生大量类肝器官。通过单细胞 RNA 测序和免疫荧光,我们证明了具有类肝细胞组成、更高阶的细胞复杂性、具有血管腔结构和驻留巨噬细胞(库普弗细胞)的群体。类器官表现出关键的肝脏功能,包括药物代谢、血清蛋白产生、尿素合成和凝血因子产生,同时保留了翻译后修饰,如 N-糖基化和功能。类器官可以在小鼠中移植并长期维持,产生人白蛋白。类器官表现出反映器官的复杂细胞组成,并具有新形成的血管化和类肝功能。这些特征是从细胞治疗、组织工程、药物毒性评估和疾病建模到基础发育生物学的许多应用的先决条件。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/71b5ab4815ed/12276_2023_1074_Fig8_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/46a961965d54/12276_2023_1074_Fig1_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/b85591460a48/12276_2023_1074_Fig2_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/857fcd2cc0f6/12276_2023_1074_Fig3_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/81001d2c1929/12276_2023_1074_Fig4_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/3fb8169ff190/12276_2023_1074_Fig5_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/decab2fe45f9/12276_2023_1074_Fig6_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/ffc7a49199ce/12276_2023_1074_Fig7_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/71b5ab4815ed/12276_2023_1074_Fig8_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/46a961965d54/12276_2023_1074_Fig1_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/b85591460a48/12276_2023_1074_Fig2_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/857fcd2cc0f6/12276_2023_1074_Fig3_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/81001d2c1929/12276_2023_1074_Fig4_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/3fb8169ff190/12276_2023_1074_Fig5_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/decab2fe45f9/12276_2023_1074_Fig6_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/ffc7a49199ce/12276_2023_1074_Fig7_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/d30b/10545717/71b5ab4815ed/12276_2023_1074_Fig8_HTML.jpg

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本文引用的文献

[1]
Temporal analyses of postnatal liver development and maturation by single-cell transcriptomics.

Dev Cell. 2022-2-7

[2]
N-Glycosylation as a Tool to Study Antithrombin Secretion, Conformation, and Function.

Int J Mol Sci. 2021-1-6

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Front Physiol. 2020-7-21

[4]
Generation of Functional Liver Sinusoidal Endothelial Cells from Human Pluripotent Stem-Cell-Derived Venous Angioblasts.

Cell Stem Cell. 2020-8-6

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Large-Scale Production of LGR5-Positive Bipotential Human Liver Stem Cells.

Hepatology. 2020-7

[6]
Blastula stage specification of avian neural crest.

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Continuous WNT Control Enables Advanced hPSC Cardiac Processing and Prognostic Surface Marker Identification in Chemically Defined Suspension Culture.

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