Single cell RNA-sequencing of lineage-negative fraction of stromal cells isolated from inguinal white adipose tissue in mouse

ABSTRACT: A subset of adipocytes residing within the inguinal white adipose tissue (ingWAT) of mice exhibit thermogenic activity in response to various external stimuli, including cold exposure. The inducible nature of this thermogenic response, coupled with its robust energy-depleting capacity have prompted investigation into the adipose precursor cells (APCs) from which thermogenic adipocytes derive. To this end, we performed single-cell transcriptomics on cells derived from ingWAT, interscapular brown adipose tissue (iBAT) and epididymal WAT. A subset of single cells collected from ingWAT and epiWAT of mice were chronically (4 days) treated with CL-316,243 (dose at 1 mg kg -1). Tissues of n=28, 10 week-old mice were digested and stromal cells were subsequently purified via differential centrifugation. Single-cell RNA extraction and mRNA amplification were performed on the C1™ Single-Cell Auto Prep Integrated Fluidic Circuit (IFC) following the protocol (PN 100-7168, http://www.fluidigm.com/). Following centrifugation and removal of the medium, cells were resuspended at a concentration of 150–500 cells/μL. This cell suspension was mixed with C1 Cell Suspension Reagent (Fluidigm, Cat # 634833) at the recommended ratio of 3:2 immediately before loading 5 μL of this final mix on the C1 IFC. We obtained, on average, 2.5 million mapped reads per one single cell and successfully reconstructed single-cell expression of ~9,000 genes. Our results revealed a unique cluster of cells that exhibited enriched expression of canonical thermogenic and adipogenic gene markers. Notably, we identified tetraspanin CD81 as a discretely expressed membrane-bound protein conserved specifically within this population. All experiments were performed at our facility at the University of California, San Francisco.

INSTRUMENT(S): Illumina HiSeq 4000

ORGANISM(S): Mus musculus

SUBMITTER: Rachana Pradhan 

PROVIDER: E-MTAB-8464 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress