Exosomes from osteoarthritic synovial fibroblasts promote angiogenesis

Abstract

Purpose: Exosomes containing proteins, microRNAs and mRNAs are found in extracellular spaces such as blood and other body fluids and function as messengers in cell-cell communication through transfer of their molecular content.Wehypothesize that exosomes containingmiRNAs function in anovel communication mechanism among joint cells in osteoarthritis (OA) pathogenesis.Wepreviouslyobservedthat thebalanceofanabolicandcatabolicgene expression in chondrocytes was impaired by exosomes derived from OAsynovial fibroblasts (SFB). Angiogenesis in various joint tissues, including synovium, ligaments, menisci and the osteochondral junction, is a recently recognized important factor inOApathogenesis. The purpose of this studywas to investigate potential angiogenic function of exosomes from OA-SFB. Methods: Human SFB were obtained from a normal, OA and rheumatoid arthritis (RA) knee joints, and cultured with or without interleukin-1b (IL1b). Exosomes were isolated by ExoQuick from IL-1b stimulated medium and control medium. Isolated exosomes fraction was measured for protein content using BCA protein assay kit. To examine the angiogenic functions of exosomes derived from SFB, we examined whether migration and tube formation in human umbilical vein endothelial cells (HUVECs)were affected by normal-, OA-, RA-SFB derived exosomes. Migration assay was evaluated using HUVECs in a modified Boyden chamber. In the tube formation assay, HUVECs were seeded into plates coated with growth-factor-reduced Matrigel and tube length was measured. miRNA profiles in the exosome preparations were established using 3D-Gene miRNA microarray. Results:Migration and tube formation activity were significantly higher in endothelial cells treated with exosomes from OA and RA SFB as compared to exosomes from normal SFB. IL-1b stimulation of SFB enhanced the angiogenic activity in their exosomes. However, migration and tube formation were not induced in endothelial cells by cytokine IL-1b alone. miRNA array data showed that 349 miRNAs were changed in OA SFB exosomes as compared to normal SFB expsomes. OAand RA-SFB exosomes shared 29miRNAs that were up-regulated and 35miRNAs that were down-regulated compared normal SFB exosomes. This includes several differentially expressed angiogenesis-related miRNAs. Conclusions: Exosomes from OA SFB accelerate angiogenic activity in HUVECs, and might be involved in OA development by promoting angiogenesis throughout the joint.miRNAarraydata suggest anOA-relatedmiRNA profile in exosomes that may serve as a novel biomarker. These results supportourhypothesis that exosomes containingmiRNAs function in anovel regulatory network that contributes to various aspects of OA pathogenesis.