Exosomes and other extracellular vesicles (EVs) have emerged as an important mechanism of cell-to-cell communication. However, previous studies either did not fully resolve what genetic materials were shuttled by exosomes or only focused on a specific set of miRNAs and mRNAs. A more systematic method is required to identify the genetic materials that are potentially transferred during cell-to-cell communication through EVs in an unbiased manner.
In this work, researchers from the Icahn School of Medicine at Mount Sinai present a novel next generation of sequencing (NGS) based approach to identify EV mediated mRNA exchanges between co-cultured adipocyte and macrophage cells. They performed molecular and genomic profiling and jointly considered data from RNA sequencing (RNA-seq) and genotyping to track the “sequence varying mRNAs” transferred between cells. They identified 8 mRNAs being transferred from macrophages to adipocytes and 21 mRNAs being transferred in the opposite direction. These mRNAs represented biological functions including extracellular matrix, cell adhesion, glycoprotein, and signal peptides.
The experimental workflow used to identify mRNA transfers between adipocytes and macrophages in a co-culture system
(a) A schema illustrating the experimental design for cell culture and sample collection including (a) adipocyte cells cultured alone, from which two cell pellet samples ADaloneN1 and ADaloneN2 were retrieved for RNA-seq, adipocyte B1 and B2 were technical duplicates of genotyping using Illumina Omni2.5 SNP array, and an exosome extraction ADexosome was prepared for RNA-seq; (b) same as (a) but for macrophage cells; (c) co-culture of two cell lines, from which three adipocyte cell pellet samples ADcoN1-N3, three macrophage cell pellet samples MOcoN1-N3, and an exosome sample ADMOexosome were profiled by RNA-seq. b The analytical pipeline including the pre-processing of (a) genotype data and (b) RNA-seq data, (c) the Bayesian model to call mRNA transfers between two cell lines, and (d) the final output of this pipeline
This study sheds new light on EV mediated RNA communications between adipocyte and macrophage cells, which may play a significant role in developing insulin resistance in diabetic patients. This work establishes a new method that is applicable to examining genetic material exchanges in many cellular systems and has the potential to be extended to in vivo studies as well.