Knocking down CDKN2A in 3D hiPSC-derived brown adipose progenitors potentiates differentiation, oxydative metabolism and browning process

Human induced pluripotent stem cells (hiPSCs) have the potential to be differentiated into any cell type, making these cells a relevant tool for therapeutic purposes such as cell-based therapies. In particular, they show great promise for obesity treatment as they represent an unlimited source of brown/brite adipose progenitors (hiPSC-BAPs), although the low brown/brite adipocyte differen-tiation potential in 2D culture represents a strong limitation for clinical use. In adipose tissue, be-sides to its cell cycle regulator functions, the cyclin-dependent kinase inhibitor 2A (CDKN2A) lo-cus modulates the commitment of stem cells to the brown-like type fate, mature adipocyte energy metabolism and browning of adipose tissue. Here, using a new method of hiPSC-BAPs 3D culture, via the formation of an organoid-like structure, we silenced CDKN2A expression during hiPSC-BAP adipogenic differentiation and observed that knocking down CDKN2A potentiates adipogenesis, oxidative metabolism and browning process, resulting in brown-like adipocytes by promoting UCP1 expression and beiging markers. Our results suggest that modulating CDKN2A levels could be relevant for hiPSC-BAPs cell-based therapies. Overall design: Comparative gene expression profiling analysis of RNA-seq data for triplicats sample of hiPSC-BAP 3D at two time points during differentiation and effect of CDKN2A deficiency (siCtrl D0, siCtrl D10, siCDKN2A D0,siCDKN2A D10).