Long-Term Culture of Primary Human Hepatocytes with Preservation of Proliferative Capacity and Differentiated Functions

Katsura, Nagato; Ikai, Iwao; Mitaka, Toshihiro; Shiotani, Tomohiro; Yamanokuchi, Satoshi; Sugimoto, Shinichi; Kanazawa, Akiyoshi; Terajima, Hiroaki; Mochizuki, Yohichi; Yamaoka, Yoshio

doi:10.1006/jsre.2002.6446

Cited by 47 publications

(34 citation statements)

References 26 publications

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“…In addition, the clinical use of isolated hepatocytes for transplantation, either as single cells or attached to microcarriers, and for extracorporal liver support systems, has been extensively studied and acknowledged during the last decade [1][2][3][4]. Moreover, because of the strategic anatomical location of the liver and the role of liver parenchymal cells in maintaining body homeostasis and detoxifying xenobiotics, isolated hepatocytes and their cultures are subject of extensive research in the field of pharmaco-toxicology of xenobiotics.…”

Section: Introductionmentioning

confidence: 99%

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Molecular Mechanisms Underlying the Dedifferentiation Process of Isolated Hepatocytes and Their Cultures

Elaut

Henkens²,

Papeleu³

et al. 2006

CDM

257

213

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Primary hepatocytes and their cultures are a simple but versatile, well-controlled, and relatively easy to handle in vitro system that is well-accepted for investigating xenobiotic biotransformation, enzyme induction and inhibition, and (biotransformation-mediated) hepatotoxicity. In addition, hepatocyte cultures have proven to be valuable tools in the study of liver physiology, viral hepatitis, and liver regeneration and are proposed as an alternative to orthotopic liver transplantation. It has been observed, however, that a number of liver-specific functions are progressively lost with time when hepatocytes are isolated and cultivated. These phenotypic changes are primarily the result of fundamental changes in gene expression concomitant with a diminished transcription of the relevant liver-specific genes, and can be interpreted as a 'dedifferentiation' of the isolated hepatocytes. Ischemia-reperfusion stress induced during the isolation process, disruption of the normal tissue architecture, as well as an adaptation to the in vitro environment are underlying factors and will be extensively discussed. A detailed description of the regulation of the hepatocyte phenotype in vivo in the first section of this review will help to understand the effect of these factors on hepatocyte gene expression. Although different approaches, mainly mimicking the in vivo hepatocyte environment, have been succesfully used to prevent or slow down the dedifferentiation of primary hepatocytes in monolayer culture, the ideal hepatocyte-based culture model, characterized by a long-term expression of hepatocyte-specific functions comparable to the in vivo level, does not exist at the moment. Consequently, alternative strategies should focus on the isolation procedure, during which dedifferentiation is already initiated. In addition, identification of the conditions needed for the full in vitro maturation of hepatic progenitor cells to quiescent, functional hepatocyte-like cells opens promising perspectives.

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Section: Introductionmentioning

confidence: 99%

“…They are particularly useful for investigating xenobiotic biotransformation, enzyme induction and inhibition, and hepatotoxicity which may or may not be caused by biotransformation. Typically, in vitro methods can answer questions at the cellular or molecular level (mechanistic toxicology) [4][5][6][7].…”

Section: Introductionmentioning

confidence: 99%

Molecular Mechanisms Underlying the Dedifferentiation Process of Isolated Hepatocytes and Their Cultures

Elaut

Henkens²,

Papeleu³

et al. 2006

CDM

257

213

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“…7B). From our previous experiment, urea synthesis rate of rat mature hepatocytes was about 32-37 g/10 6 cells/day [29]. So it is postulated that the differentiated UC-MSCs synthesize relatively low quantity of urea when compared to human mature hepatocytes (about 36 g/10 6 cells/day) [29].…”

Section: Hepatic Differentiation Of Uc-mscmentioning

confidence: 98%

In vitro hepatic differentiation of umbilical cord-derived mesenchymal stem cell

Yoon¹,

Jung²,

Seo³

et al. 2010

Process Biochemistry

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“…The isolation procedure was previously described. 18 Briefly, the liver tissue was examined for small vessels and Hanks' balanced salt solution (HBSS; GIBCO-BRL Invitrogen, Grand Island, NY) supplemented with EGTA (Dojindo Chemical Laboratories, Kumamoto, Japan) was poured into the vessels. After the washout of blood, HBSS containing 0.05% collagenase (Wako Pure Chemical, Osaka, Japan) and 0.5% dispase (Godo Shyusei, Tokyo, Japan) was poured into the vessels.…”

Section: Isolation Of Human Hepatocytesmentioning

confidence: 99%

Hepatic Organoid Formation in Collagen Sponge of Cells Isolated from Human Liver Tissues

et al. 2005

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We examined whether small hepatocytes (SHs), which are hepatic progenitor cells, could be isolated from a normal human liver and whether human hepatic cells could form hepatic organoids in a collagen sponge. Normal liver tissues were obtained from resected specimens from nine patients who underwent hepatic resection. Isolated hepatic cells were plated on dishes and a collagen sponge. More than 1 month later, SH-like cells appeared and proliferated on the dishes, whereas cell aggregates were formed in the sponge and showed characteristic tissue architecture: columnar and/or cuboidal epithelial cells lined the surface of the sponge. Clusters of epithelial cells with a large cytoplasm and ductular structures were observed under the lining cells. The lining and ductular cells were positive for cytokeratins 7 and 19, which indicated they were biliary epithelial cells (BECs), and the epithelial cells forming clusters were positive for the anti-human hepatocyte antibody, identifying them as hepatocytes. Some lining cells were positive for both the hepatic marker and the BEC markers. The cells in the collagen sponge actively proliferated and the hepatocytes excreted albumin into the medium. Thus, hepatic organoids could be reconstructed in a collagen sponge by normal human liver cells.

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Long-Term Culture of Primary Human Hepatocytes with Preservation of Proliferative Capacity and Differentiated Functions

Abstract: This hepatocyte culture method facilitates the prolonged culture of primary human hepatocytes with preservation of hepatocyte differentiation, function, and proliferative capacity.

Cited by 47 publications

References 26 publications

Molecular Mechanisms Underlying the Dedifferentiation Process of Isolated Hepatocytes and Their Cultures

Molecular Mechanisms Underlying the Dedifferentiation Process of Isolated Hepatocytes and Their Cultures

In vitro hepatic differentiation of umbilical cord-derived mesenchymal stem cell

Hepatic Organoid Formation in Collagen Sponge of Cells Isolated from Human Liver Tissues

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