A Paracrine Mechanism Accelerating Expansion of Human Induced Pluripotent Stem Cell-Derived Hepatic Progenitor-Like Cells

Tsuruya, Kota; Chikada, Hiromi; Ida, Kinuyo; Anzai, Kazuya; Kagawa, Tatehiro; Inagaki, Yutaka; Mine, Tetsuya; Kamiya, Akihide

doi:10.1089/scd.2014.0479

Cited by 11 publications

(5 citation statements)

References 60 publications

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“…Prior studies support the concept that soluble factors from mesenchymal and endothelial cells induce hepatic differentiation of fetal hepatic progenitors (Tsuruya et al, 2015). Among the paracrine Fig.…”

Section: Discussionmentioning

confidence: 86%

Paracrine signals regulate human liver organoid maturation from iPSC

et al. 2017

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A self-organizing organoid model provides a new approach to study the mechanism of human liver organogenesis. Previous animal models documented that simultaneous paracrine signaling and cell-to-cell surface contact regulate hepatocyte differentiation. To dissect the relative contributions of the paracrine effects, we first established a liver organoid using human induced pluripotent stem cells (iPSCs), mesenchymal stem cells (MSCs) and human umbilical vein endothelial cells (HUVECs) as previously reported. Time-lapse imaging showed that hepatic-specified endoderm iPSCs (HE-iPSCs) self-assembled into three-dimensional organoids, resulting in hepatic gene induction. Progressive differentiation was demonstrated by hepatic protein production after organoid transplantation. To assess the paracrine contributions, we employed a Transwell system in which HE-iPSCs were separately co-cultured with MSCs and/or HUVECs. Although the three-dimensional structure did not form, their soluble factors induced a hepatocyte-like phenotype in HE-iPSCs, resulting in the expression of bile salt export pump. In conclusion, the mesoderm-derived paracrine signals promote hepatocyte maturation in liver organoids, but organoid self-organization requires cell-to-cell surface contact. Our model demonstrates a novel approach to identify developmental paracrine signals regulating the differentiation of human hepatocytes.

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Section: Discussionmentioning

confidence: 86%

Paracrine signals regulate human liver organoid maturation from iPSC

et al. 2017

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“…However, these studies relied mainly only on biomarker analyses to evaluate the bipotency and lacked many of the functional analyses and confirmation through animal transplantation was limited. In addition, some of them included undefined components such as fetal bovine serum or feeder cells leading to safety concerns for clinical applications [15, 17–19].…”

Section: Introductionmentioning

confidence: 99%

Synergistic modulation of signaling pathways to expand and maintain the bipotency of human hepatoblasts

et al. 2019

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BackgroundThe limited proliferative ability of hepatocytes is a major limitation to meet their demand for cell-based therapy, bio-artificial liver device, and drug tests. One strategy is to amplify cells at the hepatoblast (HB) stage. However, expansion of HBs with their bipotency preserved is challenging. Most HB expansion methods hardly maintain the bipotency and also lack functional confirmation.MethodsOn the basis of analyzing and manipulating related signaling pathways during HB (derived from human induced pluripotent stem cells, iPSCs) differentiation and proliferation, we established a specific chemically defined cocktails to synergistically regulate the related signaling pathways that optimize the balance of HB proliferation ability and stemness maintenance, to expand the HBs and investigate their capacity for injured liver repopulation in immune-deficient mice.ResultsWe found that the proliferative ability progressively declines during HB differentiation process. Small molecule activation of Wnt or inhibition of TGF-β pathways promoted HB proliferation but diminished their bipotency, whereas activation of hedgehog (HH) signaling stimulated proliferation and sustained HB phenotypes. A cocktail synergistically regulating the BMP/WNT/TGF-β/HH pathways created a fine balance for expansion and maintenance of the bipotency of HBs. After purification, colony formation, and expansion for 20 passages, HBs retained their RNA profile integrity, normal karyotype, and ability to differentiate into mature hepatocytes and cholangiocytes. Moreover, upon transplantation into liver injured mice, the expanded HBs could engraft and differentiate into mature human hepatocytes and repopulate liver tissue with restoring hepatocyte mass.ConclusionOur data contribute to the understanding of some signaling pathways for human HB proliferation in vitro. Simultaneous BMP/HGF induction, activation of Wnt and HH, and inhibition of TGF-β pathways created a reliable method for long-term stable large-scale expansion of HBs to obtain mature hepatocytes that may have substantial clinical applications.Graphical abstract

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“…Recent studies have shown that human pluripotent stem cells (hPSCs) can differentiate into cholangiocytes and hepatocytes by various differentiation protocols (Tsuruya et al 2015;Takayama et al 2017;Asumda et al 2018;Dianat et al 2014;Ogawa et al 2015). However, these differentiation processes are complex and require a long time.…”

Section: Discussionmentioning

confidence: 99%

Direct and indirect coculture of mouse hepatic progenitor cells with mouse embryonic fibroblasts for the generation of hepatocytes and cholangiocytes

et al. 2019

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The widespread use of hepatocytes and cholangiocytes for regenerative medicine and tissue engineering is restricted by the limited number of hepatocytes and cholangiocytes; a simple and effective method for the expansion and differentiation of the hepatic progenitor cells (HPCs) is required. Recent studies demonstrated that mouse embryonic fibroblasts (MEFs) play an important role in supporting the proliferation of the mouse hepatic progenitor cells (mHPCs). However, the effect of direct and indirect coculture of MEFs with mHPCs on the differentiation of mHPCs is poorly studied. Herein, we show that mHPCs rapidly proliferate and form colonies in direct or indirect contact coculture with MEFs in the serumfree medium. Importantly, after direct contact coculture of the mHPCs with MEFs for 6 days, mHPCs expressed the hepatic marker albumin (ALB) and did not express the cholangiocyte marker CK19, indicating their differentiation into hepatocytes. In contrast, after indirect contact coculture of the mHPCs with MEFs for 6 days, mHPCs expressed the cholangiocyte marker CK19 and did not express the hepatic marker ALB, indicating their differentiation into cholangiocytes. These results indicate that direct and indirect contact cocultures of the mHPCs with MEFs are useful for rapidly producing hepatocytes and cholangiocytes.

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A Paracrine Mechanism Accelerating Expansion of Human Induced Pluripotent Stem Cell-Derived Hepatic Progenitor-Like Cells

Cited by 11 publications

References 60 publications

Paracrine signals regulate human liver organoid maturation from iPSC

Paracrine signals regulate human liver organoid maturation from iPSC

Synergistic modulation of signaling pathways to expand and maintain the bipotency of human hepatoblasts

Direct and indirect coculture of mouse hepatic progenitor cells with mouse embryonic fibroblasts for the generation of hepatocytes and cholangiocytes

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