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Exp Clin Endocrinol Diabetes 2020; 128(10): 672-680
DOI: 10.1055/a-1105-6332
Article

Steroidogenesis in the NCI-H295 Cell Line Model is Strongly Affected By Culture Conditions and Substrain

Max Kurlbaum
1   Department of Internal Medicine I, Division of Endocrinology and Diabetes, University of Würzburg, University Hospital Würzburg, Würzburg, Germany
2   University Hospital Würzburg, Central Laboratory, Würzburg, Germany
,
Silviu Sbiera
1   Department of Internal Medicine I, Division of Endocrinology and Diabetes, University of Würzburg, University Hospital Würzburg, Würzburg, Germany
,
Sabine Kendl
1   Department of Internal Medicine I, Division of Endocrinology and Diabetes, University of Würzburg, University Hospital Würzburg, Würzburg, Germany
2   University Hospital Würzburg, Central Laboratory, Würzburg, Germany
,
M. Martin Fassnacht
1   Department of Internal Medicine I, Division of Endocrinology and Diabetes, University of Würzburg, University Hospital Würzburg, Würzburg, Germany
2   University Hospital Würzburg, Central Laboratory, Würzburg, Germany
3   Comprehensive Cancer Center Mainfranken, University of Würzburg, Würzburg, Germany
,
Matthias Kroiss
1   Department of Internal Medicine I, Division of Endocrinology and Diabetes, University of Würzburg, University Hospital Würzburg, Würzburg, Germany
3   Comprehensive Cancer Center Mainfranken, University of Würzburg, Würzburg, Germany
› Author Affiliations Funding: This work was supported in part by the Deutsche Forschungsgemeinschaft (DFG) within the CRC/TRR 205 “The adrenal, central relay in health and disease”, project B16 to MKr and MF, the DFG grants KR4371/1–2 and FA 466/4–2 to MKr and MF, in addition by a short term fellowship of the European Society of Endocrinology to MKu.
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Correction: Steroidogenesis in the NCI-H295 Cell Line Model is Strongly Affected By Culture Conditions and SubstrainExp Clin Endocrinol Diabetes 2020; 128(10): e3-e3
DOI: 10.1055/a-1169-5951

Abstract

Context NCI-H295 cells are the most widely used model for adrenal steroidogenesis and adrenocortical carcinoma and have been used for decades in laboratories worldwide. However, reported steroidogenic properties differ considerably.

Objective To evaluate heterogeneity of steroidogenesis among NCI-H295 cell strains, clarify the influence of culture media and test response to inhibitors of steroidogenesis by using liquid chromatography tandem mass spectrometry (LC-MS/MS).

Methods NCI-H295 cells were obtained from two cell banks and cultivated in different media. An LC-MS/MS-based panel analysis of thirteen steroids was adapted for cell culture supernatant. Cells were treated with metyrapone, abiraterone and mitotane.

Results Mineralocorticoid synthesis was strongly affected by passaging as reflected by reduction of aldosterone secretion from 0.158±0.006 to 0.017±0.001 µg/106 cells (p<0.05). Relevant differences were also found for cells from two vendors in terms of aldosterone secretion (0.180±0.001 vs. 0.09±0.002 µg/106 cells, p<0.05). Selection of medium strongly impacted on cortisol secretion with>4-fold difference (40.6±5.5 vs. 182.1±23 µg/106 cells) and reflected differential activation of the glucocorticoid pathway. Exposure to abiraterone, metyrapone and mitotane resulted in characteristic steroidogenic profiles consistent with known mechanism of drug action with considerable differences in metabolites upstream of the blocked enzyme.

Conclusion We demonstrate that steroid hormone secretion in NCI-H295 cells is strongly affected by the individual strain, passage and growing conditions. These factors should be taken into account in the evaluation of experiments analyzing steroid parameters directly or as surrogate parameters of cell viability.

Key words

Steroid hormones - glucocorticoids - androgens - adrenocortical carcinoma - cell culture

Supplementary Material



Publication History

Received: 16 January 2020
Received: 21 August 2019

Accepted: 23 January 2020

Article published online:
29 April 2020

© 2020. Thieme. All rights reserved.

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