Therapeutic Potential of Placenta-derived Mesenchymal Stem/stromal Cells and Derived Extracellular Vesicles Cultured in Human Platelet Lysate for Acquired Spinal Cord Injury

Introduction: Spinal Cord Injury (SCI) is a life-altering event that can result in irreversible disabilities. Placenta-derived mesenchymal stem/stromal cells (PMSCs) cultured in fetal bovine serum have been successfully used as a therapy for myelomeningocele, a congenital form of SCI. However, to limit the risk of any zoonotic transmissions, we propose serum-free based expansion of PMSCs and extracellular vesicles (EVs) using Human Platelet Lysate (HPL) for the treatment of acquired SCIs.

Methods: PMSCs were isolated from discarded 2nd-trimester human placenta via an established explant culture method in HPL supplemented medium and EVs were isolated using ultracentrifugation. PMSCs were evaluated for growth kinetics, trilineage differentiation potential, MSC phenotype by flow cytometry, and basal concentrations of brain-derived neurotrophic factor (BDNF), vascular endothelial growth factor (VEGF), and hepatocyte growth factor (HGF) by enzyme-linked immunosorbent assay (ELISA). EV phenotype and size were assessed via Western blot and nanoparticle tracking analysis respectively. The neuroprotective capability of both PMSCs and derived EVs was studied using a staurosporine-induced apoptotic human neuroblastoma cell line. The immunomodulatory effect of the PMSCs was observed via a leukocyte suppression assay (LSA).

Results: HPL-supplemented medium increased PMSC proliferation as evaluated by their doubling time. PMSCs expressed the typical MSC surface markers and maintained their differentiation potential into adipogenic, chondrogenic and osteogenic lineages. The secretory profiles of the cells contained substantial amounts of BDNF, VEGF, and HGF. Average size of the EVs was 125 nm and Western blotting confirmed the presence of typical EV markers. PMSCs and EVs demonstrated robust neuroprotective capabilities based on increased neurite outgrowth. HPL PMSCs also inhibigited T-cell proliferation in LSAs from differing lymphocyte donors.

Conclusion : These results demonstrated that PMSCs cultured in HPL containing medium and derived EVs maintain their therapeutic potential. This paves the way for therapies that could improve motor function after SCI, clinically translating into better prognosis.