A Novel UPLC-MS Metabolomic Analysis-Based Strategy to Monitor the Course and Extent of iPSC Differentiation to Hepatocytes

Marta Moreno-Torres; Manoj Kumar; Guillem García-Llorens; Guillermo Quintás; Tine Tricot; Ruben Boon; Laia Tolosa; Burak Toprakhisar; Francois Chesnais; Catherine Verfaillie; José V. Castell

doi:10.1021/acs.jproteome.1c00779

A Novel UPLC-MS Metabolomic Analysis-Based Strategy to Monitor the Course and Extent of iPSC Differentiation to Hepatocytes

Marta Moreno-Torres, Manoj Kumar, Guillem García-Llorens, Guillermo Quintás, Tine Tricot, Ruben Boon, Laia Tolosa, Burak Toprakhisar, Francois Chesnais, Catherine Verfaillie, José V. Castell

Unidad de Hepatología Experimental, Health Research Institute La Fe, Valencia 46026, Spain
KU Leuven
Unidad de Hepatología Experimental, Health Research Institute La Fe, Valencia 46026, Spain,Departamento de Bioquímica y Biología Molecular, Universidad de Valencia, Valencia 46010, Spain
Health and Biomedicine, LEITAT Technological Center, Valencia 46026, Spain,Analytical Unit, Health Research Institute La Fe, Valencia 46026, Spain
Department of Development and Regeneration, Stem Cell Institute, KU Leuven, Leuven 3000, Belgium,Massachusetts General Hospital Cancer Center, Harvard Medical School, Boston 02114, Massachusetts, United States,The Broad Institute, Harvard & MIT, Cambridge 02142, Massachusetts, United States
Department of Development and Regeneration, Stem Cell Institute, KU Leuven, Leuven 3000, Belgium,Academic Center of Reconstructive Science, King’s College London, London SE1 9RT, U.K.
Unidad de Hepatología Experimental, Health Research Institute La Fe, Valencia 46026, Spain,Analytical Unit, Health Research Institute La Fe, Valencia 46026, Spain,Departamento de Bioquímica y Biología Molecular, Universidad de Valencia, Valencia 46010, Spain,Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas, Instituto de Salud Carlos III, Madrid 28029, Spain

Research output: Contribution to journal › Article › peer-review

5 Citations (Scopus)

Abstract

Typical protocols to differentiate induced pluripotent stem cells (iPSCs) from hepatocyte-like cells (HLCs) imply complex strategies that include transfection with key hepatic transcription factors and the addition to culture media of nutrients, growth factors, and cytokines. A main constraint to evaluate the hepatic phenotype achieved arises from the way the grade of differentiation is determined. Currently, it relies on the assessment of the expression of a limited number of hepatic gene transcripts, less frequently by assessing certain hepatic metabolic functions, and rarely by the global metabolic performance of differentiated cells. We envisaged a new strategy to assess the extent of differentiation achieved, based on the analysis of the cellular metabolome along the differentiation process and its quantitative comparison with that of primary human hepatocytes (PHHs). To validate our approach, we examined the changes in the metabolome of three iPSC progenies (transfected with/without key transcription factors), cultured in three differentiation media, and compared them to PHHs. Results revealed consistent metabolome changes along differentiation and evidenced the factors that more strongly promote changes in the metabolome. The integrated dissimilarities between the PHHs and HLCs retrieved metabolomes were used as a numerical reference for quantifying the degree of iPSCs differentiation. This newly developed metabolome-analysis approach evidenced its utility in assisting us to select a cell's source, culture conditions, and differentiation media, to achieve better-differentiated HLCs.

Original language	English
Pages (from-to)	702-712
Number of pages	11
Journal	JOURNAL OF PROTEOME RESEARCH
Volume	21
Issue number	3
Early online date	4 Jan 2022
DOIs	https://doi.org/10.1021/acs.jproteome.1c00779
Publication status	Published - 4 Mar 2022

Keywords

General Chemistry
Biochemistry

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10.1021/acs.jproteome.1c00779

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Moreno-Torres, M., Kumar, M., García-Llorens, G., Quintás, G., Tricot, T., Boon, R., Tolosa, L., Toprakhisar, B., Chesnais, F., Verfaillie, C., & Castell, J. V. (2022). A Novel UPLC-MS Metabolomic Analysis-Based Strategy to Monitor the Course and Extent of iPSC Differentiation to Hepatocytes. JOURNAL OF PROTEOME RESEARCH, 21(3), 702-712. https://doi.org/10.1021/acs.jproteome.1c00779

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title = "A Novel UPLC-MS Metabolomic Analysis-Based Strategy to Monitor the Course and Extent of iPSC Differentiation to Hepatocytes",

abstract = "Typical protocols to differentiate induced pluripotent stem cells (iPSCs) from hepatocyte-like cells (HLCs) imply complex strategies that include transfection with key hepatic transcription factors and the addition to culture media of nutrients, growth factors, and cytokines. A main constraint to evaluate the hepatic phenotype achieved arises from the way the grade of differentiation is determined. Currently, it relies on the assessment of the expression of a limited number of hepatic gene transcripts, less frequently by assessing certain hepatic metabolic functions, and rarely by the global metabolic performance of differentiated cells. We envisaged a new strategy to assess the extent of differentiation achieved, based on the analysis of the cellular metabolome along the differentiation process and its quantitative comparison with that of primary human hepatocytes (PHHs). To validate our approach, we examined the changes in the metabolome of three iPSC progenies (transfected with/without key transcription factors), cultured in three differentiation media, and compared them to PHHs. Results revealed consistent metabolome changes along differentiation and evidenced the factors that more strongly promote changes in the metabolome. The integrated dissimilarities between the PHHs and HLCs retrieved metabolomes were used as a numerical reference for quantifying the degree of iPSCs differentiation. This newly developed metabolome-analysis approach evidenced its utility in assisting us to select a cell's source, culture conditions, and differentiation media, to achieve better-differentiated HLCs.",

keywords = "General Chemistry, Biochemistry",

author = "Marta Moreno-Torres and Manoj Kumar and Guillem Garc{\'i}a-Llorens and Guillermo Quint{\'a}s and Tine Tricot and Ruben Boon and Laia Tolosa and Burak Toprakhisar and Francois Chesnais and Catherine Verfaillie and Castell, {Jos{\'e} V.}",

note = "Funding Information: This work has received funding from the European Union{\textquoteright}s Horizon 2020 Research and Innovation Program under Grant No. 681002 (EU-ToxRisk). M.M.T. acknowledges support from the European Consortium EU-ToxRisk (EU Grant No. 681002) and the Ministerio de Ciencia e Innovaci{\'o}n with Grant No. IJC2018-036209-I. L.T. was supported by the Institute of Health Carlos III through Grant No. CP16/00097. G.G.L. acknowledges the Generalitat Valenciana and the European Social Fund for hiring predoctoral research staff with grant (ACIF/2019/145). C.V. received funding from KU Leuven C14/17/111-3D-MuSYC, C32/17/053, IWT-HILIM-3D, FWO-SBO-QPG-359638-iPSC-LIMIC, and FWO G0D9917N. C.V. and M.K. received funding KU Leuven C3/20/107. M.K. also received funding from the European Union{\textquoteright}s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie-IF Grant No. 657701. T.T. was funded by FWO (1185918N). R.B. was funded by IWT fellowship SB-121393. Publisher Copyright: {\textcopyright} 2022 The Authors. Published by American Chemical Society",

year = "2022",

month = mar,

day = "4",

doi = "10.1021/acs.jproteome.1c00779",

language = "English",

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pages = "702--712",

journal = "JOURNAL OF PROTEOME RESEARCH",

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Moreno-Torres, M, Kumar, M, García-Llorens, G, Quintás, G, Tricot, T, Boon, R, Tolosa, L, Toprakhisar, B, Chesnais, F, Verfaillie, C & Castell, JV 2022, 'A Novel UPLC-MS Metabolomic Analysis-Based Strategy to Monitor the Course and Extent of iPSC Differentiation to Hepatocytes', JOURNAL OF PROTEOME RESEARCH, vol. 21, no. 3, pp. 702-712. https://doi.org/10.1021/acs.jproteome.1c00779

TY - JOUR

T1 - A Novel UPLC-MS Metabolomic Analysis-Based Strategy to Monitor the Course and Extent of iPSC Differentiation to Hepatocytes

AU - Moreno-Torres, Marta

AU - Kumar, Manoj

AU - García-Llorens, Guillem

AU - Quintás, Guillermo

AU - Tricot, Tine

AU - Boon, Ruben

AU - Tolosa, Laia

AU - Toprakhisar, Burak

AU - Chesnais, Francois

AU - Verfaillie, Catherine

AU - Castell, José V.

N1 - Funding Information: This work has received funding from the European Union’s Horizon 2020 Research and Innovation Program under Grant No. 681002 (EU-ToxRisk). M.M.T. acknowledges support from the European Consortium EU-ToxRisk (EU Grant No. 681002) and the Ministerio de Ciencia e Innovación with Grant No. IJC2018-036209-I. L.T. was supported by the Institute of Health Carlos III through Grant No. CP16/00097. G.G.L. acknowledges the Generalitat Valenciana and the European Social Fund for hiring predoctoral research staff with grant (ACIF/2019/145). C.V. received funding from KU Leuven C14/17/111-3D-MuSYC, C32/17/053, IWT-HILIM-3D, FWO-SBO-QPG-359638-iPSC-LIMIC, and FWO G0D9917N. C.V. and M.K. received funding KU Leuven C3/20/107. M.K. also received funding from the European Union’s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie-IF Grant No. 657701. T.T. was funded by FWO (1185918N). R.B. was funded by IWT fellowship SB-121393. Publisher Copyright: © 2022 The Authors. Published by American Chemical Society

PY - 2022/3/4

Y1 - 2022/3/4

N2 - Typical protocols to differentiate induced pluripotent stem cells (iPSCs) from hepatocyte-like cells (HLCs) imply complex strategies that include transfection with key hepatic transcription factors and the addition to culture media of nutrients, growth factors, and cytokines. A main constraint to evaluate the hepatic phenotype achieved arises from the way the grade of differentiation is determined. Currently, it relies on the assessment of the expression of a limited number of hepatic gene transcripts, less frequently by assessing certain hepatic metabolic functions, and rarely by the global metabolic performance of differentiated cells. We envisaged a new strategy to assess the extent of differentiation achieved, based on the analysis of the cellular metabolome along the differentiation process and its quantitative comparison with that of primary human hepatocytes (PHHs). To validate our approach, we examined the changes in the metabolome of three iPSC progenies (transfected with/without key transcription factors), cultured in three differentiation media, and compared them to PHHs. Results revealed consistent metabolome changes along differentiation and evidenced the factors that more strongly promote changes in the metabolome. The integrated dissimilarities between the PHHs and HLCs retrieved metabolomes were used as a numerical reference for quantifying the degree of iPSCs differentiation. This newly developed metabolome-analysis approach evidenced its utility in assisting us to select a cell's source, culture conditions, and differentiation media, to achieve better-differentiated HLCs.

AB - Typical protocols to differentiate induced pluripotent stem cells (iPSCs) from hepatocyte-like cells (HLCs) imply complex strategies that include transfection with key hepatic transcription factors and the addition to culture media of nutrients, growth factors, and cytokines. A main constraint to evaluate the hepatic phenotype achieved arises from the way the grade of differentiation is determined. Currently, it relies on the assessment of the expression of a limited number of hepatic gene transcripts, less frequently by assessing certain hepatic metabolic functions, and rarely by the global metabolic performance of differentiated cells. We envisaged a new strategy to assess the extent of differentiation achieved, based on the analysis of the cellular metabolome along the differentiation process and its quantitative comparison with that of primary human hepatocytes (PHHs). To validate our approach, we examined the changes in the metabolome of three iPSC progenies (transfected with/without key transcription factors), cultured in three differentiation media, and compared them to PHHs. Results revealed consistent metabolome changes along differentiation and evidenced the factors that more strongly promote changes in the metabolome. The integrated dissimilarities between the PHHs and HLCs retrieved metabolomes were used as a numerical reference for quantifying the degree of iPSCs differentiation. This newly developed metabolome-analysis approach evidenced its utility in assisting us to select a cell's source, culture conditions, and differentiation media, to achieve better-differentiated HLCs.

KW - General Chemistry

KW - Biochemistry

UR - http://www.scopus.com/inward/record.url?scp=85122668211&partnerID=8YFLogxK

U2 - 10.1021/acs.jproteome.1c00779

DO - 10.1021/acs.jproteome.1c00779

M3 - Article

SN - 1535-3893

VL - 21

SP - 702

EP - 712

JO - JOURNAL OF PROTEOME RESEARCH

JF - JOURNAL OF PROTEOME RESEARCH

IS - 3

ER -

A Novel UPLC-MS Metabolomic Analysis-Based Strategy to Monitor the Course and Extent of iPSC Differentiation to Hepatocytes

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