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Cited by 78Pregnancy and preeclampsia studiesReproductive System and PregnancyGestational Trophoblastic Disease Studies

Isolation, purification and in vitro differentiation of cytotrophoblast cells from human term placenta

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Li, Liping, Schust, Danny J·2015·Reproductive Biology and Endocrinology
DOI 10.1186/s12958-015-0070-8PMC PMC4497497
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Abstract

BACKGROUND: The syncytialization of cytotrophoblast cells to syncytiotrophoblast is central to human placental transport and hormone production. Many techniques for in vitro study of this process have been proposed and new investigators to the field may find the literature in the field daunting. Here, we present a straightforward and reliable method to establish this important model using modern but readily available tools and reagents. METHODS: Villous cytotrophoblast cells are obtained from term placenta using mild enzymatic degradation, Percoll gradient centrifugation, negative magnetic cell sorting using an antibody against classical major histocompatibility complex molecules and in vitro culture on a matrix-coated growth surface. RESULTS: The purity of isolated cytotrophoblast cells exceeds 98 % as assessed by cytokeratin-7 expression using flow cytometry. Contamination by mesenchymal cells, extravillous trophoblast cells, leukocytes, Hofbauer and endothelial cells is minimized (less than 2 % when analyzed for vimentin, HLA-G, CD45, CD163 and CD31 using flow cytometry). Isolated cytotrophoblast cells began to aggregate into monolayers of mononucleated cells within about 12 h of plating. By 72 h in culture, most cytotrophoblast cells have differentiated into syncytiotrophoblast as demonstrated by a loss of intercellular E-cadherin expression upon fusion into multinucleated syncytia. After 72 h in culture, nearly every cultured cell expresses syncytiotrophoblast markers, including cytokeratin-7, human chorionic gonadotropin-β (β-hCG) and the fusion-related proteins glial cell missing-1 (GCM-1) and syncytin. CONCLUSIONS: We present an efficient and reliable method for isolating of cytotrophoblast cells with high purity and complete differentiation into syncytiotrophoblast in vitro.

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  • Culture of isolated primary term cytotrophoblasts, which differentiate in culture to model syncytiotrophoblasts, can be employed to overcome the incomplete syncytiotrophoblast barrier issue.

    “Culture of isolated primary term cytotrophoblasts, which differentiate in culture to model syncytiotrophoblasts can be employed to overcome this issue (138).”

    From Factors Affecting the FcRn-Mediated Transplacental Transfer of Antibodies and Implications for Vaccination in Pregnancy (2017) · cites this paper as [138]

    The Results section reports that primary cytotrophoblast cells isolated from human term placentas fused into multinucleated syncytia by 72 hours in culture, supporting their use as an in vitro syncytiotrophoblast model. The paper does not establish that this culture overcomes an incomplete syncytiotrophoblast barrier issue. (LLM, conf=0.84)

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Cited by

Sorted by citations

Authors
Li, L., & Schust, D. J.
Affiliations
  • First Affiliated Hospital of Guangzhou Medical University (CN)
  • University of Missouri Health System (US)
Corresponding author
Schust, Danny J
ORCID
Venue
Reproductive Biology and Endocrinology
Publisher
Springer Science and Business Media LLC
Published
2015-07-09
Volume
13
Issue
1
Pages
71-71
ISSN
1477-7827 (ISSN-L)
DOI
10.1186/s12958-015-0070-8
PMC
PMC4497497
OpenAlex
W1927898783
Cited by
78
References
0
Citations by year
2026: 4 · 2025: 4 · 2024: 11 · 2023: 8 · 2022: 6 · 2021: 13 · 2020: 11 · 2019: 8 · 2018: 3 · 2017: 5 · 2016: 4 · 2015: 1
FWCI
4.93
Type
journal article
Language
en
Open access
Licence
Locations
  • Reproductive Biology and Endocrinology (publishedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
Funders
  • Natural Science Foundation of Guangzhou City
  • National Natural Science Foundation of China
  • National Institutes of Health (NIH-R01)
  • Guangzhou Municipal Science and Technology Project
  • Eunice Kennedy Shriver National Institute of Child Health and Human Development (R01HD077108)
Topics
  • Health Sciences › Medicine › Obstetrics and Gynecology › Pregnancy and preeclampsia studies
  • Life Sciences › Immunology and Microbiology › Immunology › Reproductive System and Pregnancy
  • Health Sciences › Medicine › Public Health, Environmental and Occupational Health › Gestational Trophoblastic Disease Studies
Keywords
cytotrophoblast, syncytiotrophoblast, syncytialization, placenta, cell isolation, cell purification, Percoll density gradient centrifugation, magnetic cell separation, HLA-G, CK7, E-cadherin, syncytiotrophoblast differentiation, human chorionic gonadotropin, term placenta
MeSH terms
Cell Differentiation, Cell Separation, Cells, Cultured, DNA-Binding Proteins, Female, Flow Cytometry, Chorionic Gonadotropin, Humans, Nuclear Proteins, Placenta, Pregnancy, Transcription Factors, Trophoblasts, Giant Cells, Cadherins, Keratin-7, In Vitro Techniques

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