Metabolic Profiling of hASCs

OC and ECAR were measured using an XF96 oxymeter (Seahorse Biosciences, North Billerica, MA, USA). hASCs of donors not related to the RNA-Seq analysis were seeded and differentiated in 96-well XF96 assay plates. Baseline, dibutyril-cAMP (Sigma-Aldrich cat#D0627) stimulated, β-guanidinopropionic acid (Sigma-Aldrich cat#G6878), and oligomycin (Enzo Life Sciences, Farmingdale, NY, USA cat#ALX-380-037) inhibited OC and ECAR were recorded. As the last step, cells received a single bolus dose of Antimycin A (Sigma-Aldrich cat# U8674) at 10 μM final concentration for baseline correction. The oxygen consumption rate (OCR) and ECAR were normalized to protein content and normalized readings were shown. For statistical analysis, relative OC and ECAR levels were determined to compare basal, cAMP stimulated and oligomycin inhibited (both in unstimulated and stimulated cells) OCRs/ECARs of each sample to the basal OCR/ECAR of untreated SC white adipocytes [6 (link),7 (link),26 (link)].

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Tóth B.B., Arianti R., Shaw A., Vámos A., Veréb Z., Póliska S., Győry F., Bacso Z., Fésüs L, & Kristóf E. (2020). FTO Intronic SNP Strongly Influences Human Neck Adipocyte Browning Determined by Tissue and PPARγ Specific Regulation: A Transcriptome Analysis. Cells, 9(4), 987.

Publication 2020

XF96 oxymeter dibutyril-cAMP oligomycin Antimycin A

Antimycin a Assay Cells Donors Guanidinopropionic acid Oligomycin Oxygen consumption Protein Rna seq Seahorse White adipocytes

Corresponding Organization : University of Debrecen

Other organizations : University of Szeged

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Variable analysis

independent variables

Dibutyril-cAMP (Sigma-Aldrich cat#D0627) stimulation
β-guanidinopropionic acid (Sigma-Aldrich cat#G6878) treatment
Oligomycin (Enzo Life Sciences, Farmingdale, NY, USA cat#ALX-380-037) inhibition

dependent variables

Oxygen consumption rate (OCR)
Extracellular acidification rate (ECAR)

control variables

HASCs of donors not related to the RNA-Seq analysis were seeded and differentiated in 96-well XF96 assay plates
Antimycin A (Sigma-Aldrich cat# U8674) at 10 μM final concentration for baseline correction
Normalized OCR and ECAR readings to protein content

positive controls

Basal OCR/ECAR of untreated SC white adipocytes

negative controls

Not explicitly mentioned

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