Project description:From subcutaneous adipsoe tissue from 2 particpants we performed scRNA-seq on the stromal vascular fraction, snRNA-seq on isolated adipocytes and snRNA-seq on frozen adipose tissue. We performed clustering analysis, differential gene expression analysis and over-representation analysis to decipher the different cell populations detectable with each technique. The SVF fraction was able to show more diverse immune cell populations. Whilst the snRNA-seq frozen adipose tissue showed diverse clusters of adipocytes that was not observable from a snRNA-Seq fraction
Project description:Adipose tissue mass and adiposity change throughout the lifespan. During aging, while visceral adipose tissue (VAT) tends to increase, peripheral subcutaneous adipose tissue (SAT) decreases significantly. Unlike VAT, which is linked to metabolic diseases, SAT has beneficial effects. However, the molecular details behind aging-associated loss of SAT remain unclear. Here we compare scRNA-seq of total SVF of SAT from young and aging mice to identify a novel Aging-dependent Regulatory Cell (ARC) that emerges in SAT of aged mice. Inguinal white adipose tissue (iWAT) was used as a representative SAT; iWAT pads of 2 mice from each age group were subjected to collagenase digestion and treated with a hypotonic buffer to remove red blood cells before subjection to scRNA-seq by 10X Genomics Chromium Single Cell Kit. The findings showed that ARCs express adipogenic markers but lack adipogenic capacity and inhibit differentiation of neighboring adipose precursors.
Project description:Using our robust snRNA-seq protocol effectively enhancing nucleus integrity and RNA quality, we comprehensivly characterize the dynamics of cell populations within mouse epididymal and inguinal adipose tissues during obesity.
Project description:Single nucleus RNA sequencing (snRNA-seq), an alternative to single cell RNA sequencing (scRNA-seq), encounters technical challenges in obtaining high-quality nuclei and RNA, persistently hindering its applications. Here, we present a robust technique for isolating nuclei across various tissue types, remarkably enhancing snRNA-seq data quality. Employing this approach, we comprehensively characterize the depot-dependent cellular dynamics of various cell types underlying adipose tissue remodeling during obesity. By integrating nuclear RNA-seq data from adipocyte nuclei of varying sizes, we identify distinct adipocyte subpopulations categorized by size and functionality. Specifically, we characterize dysfunctional hypertrophic adipocytes prevalent in visceral adipose tissues during obesity, exhibiting cellular stress, inflammation and impaired metabolic gene expression. Obesity-induced changes in gene expression profiles of adipocyte subpopulations reveal their distinct contributions to adipose tissue pathophysiology. Our study establishes a robust snRNA-seq method, providing novel insights into the mechanisms orchestrating adipose tissue remodeling during obesity, with broader applicability across diverse biological systems.