Long-term in vitro 2D-culture of SDHB and SDHD-related human paragangliomas and pheochromocytomas.
Adrenal Gland Neoplasms
/ pathology
Chromogranin A
/ metabolism
Culture Media, Serum-Free
Germ-Line Mutation
Humans
Lactates
Paraganglioma
/ genetics
Pheochromocytoma
/ pathology
Phosphopyruvate Hydratase
/ metabolism
Succinate Dehydrogenase
/ genetics
Synaptophysin
/ metabolism
Tyrosine 3-Monooxygenase
/ metabolism
Journal
PloS one
ISSN: 1932-6203
Titre abrégé: PLoS One
Pays: United States
ID NLM: 101285081
Informations de publication
Date de publication:
2022
2022
Historique:
received:
02
06
2022
accepted:
29
08
2022
entrez:
30
9
2022
pubmed:
1
10
2022
medline:
5
10
2022
Statut:
epublish
Résumé
The neuroendocrine tumours paraganglioma and pheochromocytoma (PPGLs) are commonly associated with succinate dehydrogenase (SDH) gene variants, but no human SDH-related PPGL-derived cell line has been developed to date. The aim of this study was to systematically explore practical issues related to the classical 2D-culture of SDH-related human paragangliomas and pheochromocytomas, with the ultimate goal of identifying a viable tumour-derived cell line. PPGL tumour tissue/cells (chromaffin cells) were cultured in a variety of media formulations and supplements. Tumour explants and dissociated primary tumour cells were cultured and stained with a range of antibodies to identify markers suitable for use in human PPGL culture. We cultured 62 PPGLs, including tumours with confirmed SDHB, SDHC and SDHD variants, as well as several metastatic tumours. Testing a wide range of basic cell culture media and supplements, we noted a marked decline in chromaffin cell numbers over a 4-8 week period but the persistence of small numbers of synaptophysin/tyrosine hydroxylase-positive chromaffin cells for up to 99 weeks. In cell culture, immunohistochemical staining for chromogranin A and neuron-specific enolase was generally negative in chromaffin cells, while staining for synaptophysin and tyrosine hydroxylase was generally positive. GFAP showed the most consistent staining of type II sustentacular cells. Of the media tested, low serum or serum-free media best sustained relative chromaffin cell numbers, while lactate enhanced the survival of synaptophysin-positive cells. Synaptophysin-positive PPGL tumour cells persist in culture for long periods but show little evidence of proliferation. Synaptophysin was the most consistent cell marker for chromaffin cells and GFAP the best marker for sustentacular cells in human PPGL cultures.
Identifiants
pubmed: 36178902
doi: 10.1371/journal.pone.0274478
pii: PONE-D-22-14862
pmc: PMC9524698
doi:
Substances chimiques
Chromogranin A
0
Culture Media, Serum-Free
0
Lactates
0
SDHD protein, human
0
Synaptophysin
0
Tyrosine 3-Monooxygenase
EC 1.14.16.2
SDHB protein, human
EC 1.3.5.1
Succinate Dehydrogenase
EC 1.3.99.1
Phosphopyruvate Hydratase
EC 4.2.1.11
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
e0274478Déclaration de conflit d'intérêts
The authors have declared that no competing interests exist.
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