Exosomes for Wound Healing: Purification Optimization and Identification of Bioactive Components

Abstract

Human mesenchymal stem cell exosomes have been shown to promote cutaneous wound healing. Their bioactivity is most often attributed to their protein and nucleic acid components, while the function of exosomal lipids remains comparatively unexplored. This work specifically assesses the involvement of lipids and the transmembrane enzyme CD73 in the exosomes' biological activity in stimulating the cutaneous wound healing process. Since exosome preparation processes are not harmonized yet, certain production and purification parameters are first systematically investigated, enabling the optimization of a standardized protocol delivering high exosome integrity, yield, and purity. An in situ enzymatic assay is introduced to specifically assess the vesicle functionality, and quantitative proteomics is employed to establish the cell culture conditions yielding a stable exosome protein profile. Using a combination of in vitro approaches, CD73 and constitutional lipids of HPV-16 E6/E7 transformed human bone marrow stromal cell-derived exosomes are identified as key bioactive components promoting the exosome-driven acceleration of processes required for wound repair. A pilot wound healing study in mice indeed suggests a role of lipids in the exosomes' biological activity. Strikingly, the extent of the bioactivity of these exosomal components is found to be dependent on the target cell type.

Differential Therapeutic Effect of Extracellular Vesicles Derived by Bone Marrow and Adipose Mesenchymal Stem Cells on Wound Healing of Diabetic Ulcers and Correlation to Their Cargoes

“…Our data demonstrated that both BMSC and ADSC-EVs promoted fibroblast and keratinocyte migration in vitro ( Figure 5 A). The ability of EVs to drive keratinocyte and fibroblast migration during wound healing was correlated to their CD73 content [ 52 ] and previous reports demonstrated that BMSC and ADSC express similar level of CD73 [ 7 ]. Here, we demonstrated that BMSC and ADSC-EVs express similar levels of CD73 by FACS analysis ( Figure 1 C), possibly explaining their similar effect on keratinocyte and fibroblast migration in diabetic wound healing process.…”

The Unique Immunomodulatory Properties of MSC-Derived Exosomes in Organ Transplantation

“…Thus, MSC-derived exosomes have broad prospects as a therapeutic agent. It is worth noting that although MSC exosomes avoid the risks of immunogenicity and tumorigenicity that are associated with cell transplantation, there are still many problems associated with application in the clinical setting, including production standards (e.g., culture separation, cell phenotype, quantification, and positioning tracking after infusion) and the monitoring of efficacy ( 131 ). The determination of clinical indications for different diseases, especially the production of standard dosage in the field of organ transplantation, is a problem that needs to be solved urgently.…”

Exosomes, PD-L1 and aGvHD: Perspectives for WJMSCmediated Therapy

“…WJSMC exosomes were defined as positive for CD9, CD63, and CD73 [33]. Cytokine TNF-α stimulation not only increased the amount of exosome secreted from gingivaderived MSCs but also enhanced the exosomal expression of CD73 [34].…”