Membrane particles derived from mesenchymal stromal cells as a novel cell free therapy for immunomodulation and regeneration

Background & Aim Mesenchymal stromal cells (MSC) are widely studied for their immunomodulatory and regenerative properties. However, MSC are large and get trapped in the capillary networks of the lungs after intravenous infusion, where they have a short survival time. Recent work demonstrated that inactivated MSC which lost their capacity to secrete factors maintain their regulatory capacity, suggesting that cell membrane dependent interactions with immune cells are responsible for the regulatory effects. We propose a new cell-free therapy based on the generation of small plasma Membrane Particles (MP) from adipose tissue MSC. The aim of this work is to study the therapeutic potential of MP for immune and degenerative disorders Methods, Results & Conclusion MP from adipose tissue MSC were generated by hypotonic shock and extrusion. Parameters such as size (by nanosight), morphology (by cryo-electron microscopy), and enzyme activity were analysed. The interaction of MP with cells was analysed by confocal microscopy. The therapeutic potential of MP was tested in three in vitro models:1.Immune cells were treated with MP and monocyte apoptosis were performed by flow cytometry.2.Inflammed endothelial cells (EC) model (EC+TNFα). Monocyte transmigration through an EC monolayer, and EC angiogenesis capacity were analyzed by microscopy. Endothelial layer integrity was evaluated by colorimetric assay.3.Human lung fibroblasts from biopsies of Idiopathic pulmonary fibrosis patients were stimulated with TGFb to increase the expression of profibrotic molecules (collagen I, and III, fibronectin, PDL-1, tenascin-c and MMP1). The protein gene expression was analysed by PCR. The results showed that the average size of MP was 120 nm, and they showed a spherical shape. MP exhibited ATPase and nucleotidase activity, indicating they are enzymatically active. MP bound to the cell membranes of monocytes and were internalized by EC and fibroblasts. With respect to the in vitro functional immune studies, MP induced apoptosis of proinflammatory monocytes. On EC, MP decreased the permeability of the EC monolayer reducing the number of transmigrating monocytes. They also stimulated the formation of angiogenic structures in vitro.In the fibrosis model, MP decreased the expression of profibrotic proteins in fibroblasts such as collagen II, tenascin-c, and PDL-1. In conclusion, MP have immunomodulatory and regenerative properties and have potential as a novel cell-free therapy for treatment of immunological and degenerative disorders.