Anti-Inflammatory Effects of Exosomes Derived from Human Umbilical Cord Mesenchymal Stem Cells on Neuroglia

Abstract

Wharton’s jelly mesenchymal stem cells (WJ-MSC) have the capacity to reduce neuroinflammation and induce tissue regeneration in perinatal brain damage despite of their low long-term survival in host tissue. The therapeutic function of WJ- MSC is mainly ascribed to their paracrine secretion involving the shedding of cell-derived exosomes. The aim of this study is to evaluate the anti-inflammatory effects of WJ-MSC-derived exosomes on neuroglia in vitro. WJ-MSC derived exosomes were isolated from cell culture supernatants using a protocol consisting of several steps of successive centrifugations and ultra-centrifugations. The isolated exosomes were characterized by their expression of endosomal markers and their size using a membrane-based antibody array and electron microscopy. In vitro models involving oxygen glucose deprivation and reoxygenation (OGD-R) and lipopolysaccharide (LPS) stimulation were used to test the anti-inflammatory effects of the exosomes on activated primary astrocytes and immortalized microglia cells. After the co-culture with WJ-MSC derived exosomes, glia cells were evaluated for their expression of activation markers and production of pro-inflammatory cytokines by real-time PCR, enzyme-linked immunosorbent assay (ELISA) and Western blot. WJ-MSC-derived exosomes were positive for endosomal markers, including TSG101 and ALIX, and had a mean diameter of 34 nm. In co-culturing experiments, WJ-MSC-derived exosomes prevented the upregulation of the astrocyte activation marker glial fibrillary acidic protein (Gfap) in response to 6h of OGD and 48h of reoxygenation. WJ-MSC-derived exosomes further tend to suppress the upregulation of pro-inflammatory cytokines such as interleukin 1 beta (IL-1b), tumor necrosis factor alpha (TNF-α) and inducible isoform of nitric oxide synthase (iNOS) in response to 24h LPS stimulation. In conclusion, we demonstrate that WJ-MSC-derived exosomes are potent modulators of neuroglia activation in hypoxia/ ischemia and inflammation. Hence not only WJ-MSC, but also WJ-MSC-derived exosomes are able to support tissue regeneration by reducing inflammation. As a result, WJ-MSC-derived exosomes might represent a novel cell-free approach to treat perinatal brain damage