Abstract Body (Do not enter title and authors here): Abstract:
Background: Critical limb ischemia is a severe stage of peripheral artery disease. It causes claudication, ischemic pain, and ulceration. It is a serious condition that increases risks of limb amputation and death. While extracellular vesicles (EVs) secreted naturally from endothelial cells (ECs) or mesenchymal stem cells (MSCs) have shown promise for the treatment of ischemic limb diseases in mice, the clinical translation of EV therapy into patients has been limited by low yields from cultured cells. In this study, we evaluated the potential of xeno-transplanted nanovesicles (NVs) manufactured from human induced-pluripotent stem cells (hiPSCs) for the treatment of ischemic limb diseases in mice without administration of immunosuppressive drugs.
Methods and Results: A hypo-immunogenic hiPSC line with β2-microglobulin knockout (^B2MKOhiPSCs) was used to manufacture NVs (^B2MKOhiPSC-NVs). NV size and concentration were measured using an Nanosight, NV morphology was imaged using an transmission electron microscope, and NV zeta potential was measured using the ZETASIZER Nano series. In vitro, the cytoprotective and proliferative effects of ^B2MKOhiPSC-NVs on human umbilical vein endothelial cells (HUVECs) were determined. In vivo, the therapeutic potential of ^B2MKOhiPSC-NVs was tested in a mouse model of hind limb ischemia, without administration of immunosuppressive drugs. Over 9,500 NVs could be manufactured from one hiPS cell. The zeta potential of ^B2MKOhiPSC-NVs was -16.7 mV, with a mean diameter of 115.9 ± 43.5 nm and bilayer lipid membranes. In vitro, ^B2MKOhiPSC-NVs protected HUVECs from hypoxic injury and promoted their proliferation. In vivo, ^B2MKOhiPSC-NV administration significantly improved blood perfusion, which was accompanied by significantly increased mouse EC proliferation and stimulated neovascularization in ischemic limbs, compared to control mice 14 days after treatment.
Conclusions: ^B2MKOhiPSC-NVs hold significant potential for the treatment of ischemic limb diseases.