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Clonal analyses and gene profiling identify genetic biomarkers of the thermogenic potential of human brown and white preadipocytes

Abstract

Targeting brown adipose tissue (BAT) content or activity has therapeutic potential for treating obesity and the metabolic syndrome by increasing energy expenditure. However, both inter- and intra-individual differences contribute to heterogeneity in human BAT and potentially to differential thermogenic capacity in human populations. Here we generated clones of brown and white preadipocytes from human neck fat and characterized their adipogenic and thermogenic differentiation. We combined an uncoupling protein 1 (UCP1) reporter system and expression profiling to define novel sets of gene signatures in human preadipocytes that could predict the thermogenic potential of the cells once they were maturated. Knocking out the positive UCP1 regulators, PREX1 and EDNRB, in brown preadipocytes using CRISPR-Cas9 markedly abolished the high level of UCP1 in brown adipocytes differentiated from the preadipocytes. Finally, we were able to prospectively isolate adipose progenitors with great thermogenic potential using the cell surface marker CD29. These data provide new insights into the cellular heterogeneity in human fat and offer potential biomarkers for identifying thermogenically competent preadipocytes.

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Figure 1: Generation and characterization of immortalized human brown and white fat progenitors.
Figure 2: Use of a UCP1 reporter system for in vitro and in vivo monitoring of UCP1 expression.
Figure 3: Clonal analysis of human brown and white fat progenitors.
Figure 4: Gene expression profiles in adipose progenitors predict the thermogenic capacity of mature adipocytes.
Figure 5: PREX1 and EDNRB are required for determining thermogenic competency.
Figure 6: Isolation of progenitors possessing thermogenic potential using a cell surface marker.

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Acknowledgements

This work was supported in part by US National Institutes of Health (NIH) grants R01DK077097 (to Y.-H.T.), R01DK099511 (to L.J.G.), K23DK081604 (to A.M.C.) and P30DK036836 (to Joslin Diabetes Center's Diabetes Research Center, DRC) from the National Institute of Diabetes and Digestive and Kidney Diseases, a sponsored research grant from Chugai Pharmaceutical Co. (to Y.-H.T. and A.M.C.), a research grant from the American Diabetes Association (ADA 7-12-BS-191, to Y.-H.T.), the Intramural Research Program of the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), and by funding from the Harvard Stem Cell Institute (to Y.-H.T.). M.D.L. was supported by NIH fellowships (T32DK007260 and F32DK102320). We thank M.-E. Patti and K. Hughes of the Advanced Genomics and Genetics Core of Joslin's DRC for advice and expert technical assistance. The authors thank Stryker Regenerative Medicine (Hopkinton, Massachusetts) for the generous gift of recombinant BMP7.

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The study was designed by Y.-H.T., R.X., M.D.L. and A.M.C. The manuscript was written by Y.-H.T., M.D.L., R.X. and J.M.D. R.X. performed the majority of the experiments. M.D.L. did the time-lapse imaging, IVIS scanning and FACS. J.M.D. analyzed microarray data. F.S. performed bioenergetics analyses in knockout cells. T.J.S. and H.Z. established the method of isolation, immortalization and differentiation of human fat progenitors. T.L.H. did the human cell implantation and gene expression microarrays. K.L.T. provided assistance with the Seahorse bioanalyzer. Y.L. provided research assistance. H.T. and L.J.G. helped with fuel utilization experiments. A.M.C., L.S.W. and A.P.W. collected human fat samples. M.S.L. and L.L.R. helped with the time-lapse imaging. All authors contributed to editing the manuscript.

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Correspondence to Yu-Hua Tseng.

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Competing interests

A.M.C. and Y.-H.T. are recipients of a sponsored research grant and licensing payments from Chugai Pharmaceutical Co., Ltd through Joslin Diabetes Center.

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Supplementary Figures 1–9 & Supplementary Tables 1–5 (PDF 1480 kb)

Time lapse imaging of hBAT-SVF differentiation (MOV 31877 kb)

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Xue, R., Lynes, M., Dreyfuss, J. et al. Clonal analyses and gene profiling identify genetic biomarkers of the thermogenic potential of human brown and white preadipocytes. Nat Med 21, 760–768 (2015). https://doi.org/10.1038/nm.3881

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