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Inducible UCP1 silencing: A lentiviral RNA-interference approach to quantify the contribution of beige fat to energy homeostasis

Abstract

Energy consuming, heat-producing beige adipocytes, located in classic white adipose tissue (WAT), hold promise for the treatment of obesity. Few reports have quantitatively assessed the contribution of browned 'WAT' to energy expenditure. There is a need for methods to examine beige-fat thermogenesis, independently of classical brown fat. The aim of this study is to optimize an inducible lentiviral shRNA to conditionally knock-down Ucp1 and assess the effects on 'browned' WAT. Primary adipocytes from mouse inguinal WAT converted into thermogenic adipocytes when stimulated with beta-adrenergic agonist and thiazolidinedione. There was increased UCP1 protein and importantly increases in various indicators of mitochondrial bioenergetics. Next, we determined optimal transfection conditions for the UCP1-shRNA lentiviral system and subsequently applied this to 'browned' WAT. UCP1 knockdown decreased the brown/beige-fat gene profile and decreased mitochondrial respiration. In summary, this study optimizes lentiviral UCP1-shRNA technology in vitro. This technique could be applied to inguinal fat depots in vivo. This would allow investigation of contribution of depots to whole-body metabolism to help elucidate the physiological relevance of beige fat.

Type Journal
ISBN 1932-6203 (Electronic) 1932-6203 (Linking)
Authors Khor, N. W. M.; Swarbrick, M. M.; Gunton, J. E.
Responsible Garvan Author Prof Jenny Gunton
Publisher Name PLoS One
Published Date 2019-11-21
Published Volume 14
Published Issue 11
Published Pages e0223987
Status Always Electronic
DOI 10.1371/journal.pone.0223987
URL link to publisher's version https://www.ncbi.nlm.nih.gov/pubmed/31751350