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Endocrine Abstracts (2018) 56 GP254 | DOI: 10.1530/endoabs.56.GP254

ECE2018 Guided Posters Thyroid non cancer (9 abstracts)

Proteome analysis of 3,5-T2-treated primary mouse hepatocytes

Janine Golchert 1 , Julika Lietzow 2 , Uwe Völker 1 , Georg Homuth 1 & Josef Köhrle 2


1Department of Functional Genomics, Interfaculty Institute for Genetics and Functional Genomics, University Medicine Greifswald, Greifswald, Germany; 2Institute of Experimental Endocrinology, Charité – Universitätsmedizin Berlin, Berlin, Germany.


Introduction: Besides the classical thyroid hormones T4 and T3 the endogenous thyroid hormone metabolite 3,5-diiodothyronine (3,5-T2) attracted attention during the last years due to its metabolic activity. 3,5-T2 exerts thyromimetic action on the hypothalamus-pituitary-thyroid axis and the heart as well as on energy and lipid metabolism. In previous liver transcriptome analyses of 3,5-T2-treated lean and diet-induced obese male mice, effects on hepatic lipid, steroid, xenobiotic, and thyroid hormone metabolism were observed.

Objectives: Applying an untargeted mass spectrometry based proteome analysis approach for 3,5-T2-treated primary mouse hepatocytes, we intended to confirm previous in vivo transcriptome data and to identify novel protein targets and pathways of hepatic 3,5-T2 action.

Materials & methods: Primary hepatocytes isolated from male mouse liver were cultivated under FCS-free conditions and treated with 100 nM 3,5-T2 for either 6, 24, 48 or 72 h (n=3 per group). Proteins were prepared and digested using Trizol and FASP (filter-aided sample preparation), respectively. Proteomes were analyzed using LC-MS/MS measurement and data analysis was performed via MaxQuant and GeneData Analyst software. Pathway analysis of proteins exhibiting significantly altered amounts was carried out using the Ingenuity Pathway Analysis (IPA) software.

Results: The proteomics approach revealed 852 proteins exhibiting significantly altered amounts in 3,5-T2-treated primary hepatocytes compared to their time matched controls (P ≤ 0.05, fold change ≥ |1.5|). Notably, the highest number of proteins with significantly changed amounts was observed after 48 h of treatment, while after 6 and 24 h 4.5-times less proteins were altered. The greatest overlap of 92 proteins was observed between the time points 24 and 48 h of 3,5-T2 treatment. As identified by IPA, proteins showing significant alterations mainly belong to the pathways ‘EIF2 signaling’, ‘regulation of eIF4 and p70S6K signaling’, ‘mTOR signaling’, various degradation pathways for intermediary metabolites, as well as proteins involved in steroid hormone metabolism.

Conclusion: The untargeted mass spectrometry approach identified a variety of proteins and pathways altered by 3,5-T2 treatment in primary mouse hepatocytes as well as the confirmation of previously indicated 3,5-T2 effects on mouse liver transcriptomes and functional readouts. To further extend our knowledge of 3,5-T2 action, a transcriptome analysis of the current study is planned to create a complementary dataset.

Volume 56

20th European Congress of Endocrinology

Barcelona, Spain
19 May 2018 - 22 May 2018

European Society of Endocrinology 

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