Unraveling the Developmental Roadmap toward Human Brown Adipose Tissue

Abstract

Summary Increasing brown adipose tissue (BAT) mass and activation is a therapeutic strategy to treat obesity and complications. Obese and diabetic patients possess low amounts of BAT, so an efficient way to expand their mass is necessary. There is limited knowledge about how human BAT develops, differentiates, and is optimally activated. Accessing human BAT is challenging, given its low volume and anatomical dispersion. These constraints make detailed BAT-related developmental and functional mechanistic studies in humans virtually impossible. We have developed and characterized functionally and molecularly a new chemically defined protocol for the differentiation of human pluripotent stem cells (hPSCs) into brown adipocytes (BAs) that overcomes current limitations. This protocol recapitulates step by step the physiological developmental path of human BAT. The BAs obtained express BA and thermogenic markers, are insulin sensitive, and responsive to β-adrenergic stimuli. This new protocol is scalable, enabling the study of human BAs at early stages of development.

Reconstructing human Brown Fat developmental trajectory in vitro

“…This also provided us with a benchmark to which the differentiating hPSCs could be compared. Our comparison of the differentiated human adipocytes generated in vitro in our protocol with those generated by two other recently published protocols (Carobbio et al, 2021; Zhang et al, 2020) suggest that the transcriptional signature of the adipocytes generated in our conditions is closer to endogenous fetal brown adipocytes. In humans, BAT activity shows an inverse correlation with body mass index and percentage of total body fat (Cypess et al, 2009; Saito et al, 2009).…”

“… M – Heatmap comparing transcriptomic profiles of iPSC derived brown adipocytes (iPSC-BA), human fetal brown adipose tissue (fBAT), human embryonic stem cell derived brown adipocytes in Zhang et al 2020 (H9-d50) and iPSC derived brown adipocytes in Carobbio et al 2021 (KOLF2-C1-d25).…”

“…When analyzed using the ProFat database (Cheng et al, 2018), the browning probability of iPSC-BAs was comparable to fetal brown adipocytes (Figure 5L). Recently, two studies have described the directed differentiation of brown adipocytes from hPSCs (Carobbio et al, 2021; Zhang et al, 2020). Comparison of our iPSC-BAs with the terminally differentiated adipocytes from the two studies – 50 day old adipocytes from Zhang et al (H9-d50) and 25 day old adipocytes from Carobbio et al (KOLF2-C1-d25) showed that cells differentiated according to our protocol exhibit a higher degree of similarity to the human fetal BAT and brown adipocytes gene expression profile (Figure 5M, Supplementary Figure 5E).…”

“…Other reported methods rely on serum-based spontaneous differentiation or treatment of hPSCs with a cytokine cocktail (Guenantin et al, 2017; Hafner et al, 2016; Nishio et al, 2012). Recently, protocols allowing the generation of brown adipocytes from hPSCs by recapitulating developmental cues have also been reported (Carobbio et al, 2021; Zhang et al, 2020). However, to date, a well characterized roadmap of brown fat lineage development is still missing.…”

FTO m6A Demethylase in Obesity and Cancer: Implications and Underlying Molecular Mechanisms

“…Human-induced pluripotent stem cells (iPSCs) represent an attractive ideal cellular model that is promising in delineating mechanistic details of FTO SNPs on adipocyte differentiation, including production of thermogenic adipocytes at higher frequencies [ 127 , 128 ]. Several reports published directed differentiation protocols to produce thermogenic adipocytes using iPS cells without gene transfer methods [ 129 , 130 , 131 , 132 ]. Using iPS cells to produce thermogenic adipocytes and to investigate FTO’s role as an m6A demethylase throughout the adipogenesis process has not been investigated yet.…”