Bioreactor produced wharton's jelly mesenchymal stem cell- derived exosomes prevent lung injury in a pre-clinical model of bronchopulmonary dysplasia.

Background & Aim Mesenchymal stem cells (MSCs) have been safely and effectively used to treat lung and heart diseases. However, cell product variability and limited survival post transplantation has led to varying degrees of clinical success. Recent evidence suggest that exosomes rather than stem cell engraftment and differentiation mediate MSC regenerative effects. Exosomes are nanosized extracellular vesicles that a play a key role in intercellular and interorgan communications. Therefore, exosome therapy has emerged as a new therapeutic alternative to MSCs. One of the challenges for the transition of exosomes from the bench to the bedside is the generation of high quantity GMP-compliant products. Thus, we aimed to develop a production methodology of Wharton's Jelly MSC (WJMSC) derived exosomes in a bioreactor and test the hypothesis that the exosome product would significantly reduce neonatal lung injury in a preclinical model of Bronchopulmonary Dysplasia. Methods, Results & Conclusion WJMSCs were isolated from Wharton's Jelly and expanded to passage 1. After harvest, 30 × 106 cells were loaded into the quantum bioreactor (Terumo BCT) and expanded for 1 week to achieve 4.05 × 108 cells. Serum-free aMEM conditioned media was collected every 24 hours for 96 hours for exosome precipitation by ultracentrifugation. NanoSight analysis of the final products revealed a production of 15.0 × 1012± 0.32 × 1012 total particles with an average mode size of 111±4.78 nm. Furthermore, protein analysis revealed positive expression of exosome markers CD9, CD63, and CD81. To test the regenerative potential of this product, we treated newborn Sprague-Dawley rats (n=9/group) exposed to normoxia or hyperoxia (85% O2) from postnatal day 1-14 at day 3 with 1 × 1010 total particles (EXO) or placebo (PL). The degree of lung vascular remodeling, alveolarization, and pulmonary hypertension was then assessed. Results confirmed a significant lung protective effect of the EXO treatment in the hyperoxia groups by the demonstration of decreased percentage of muscularized vessels (67 ± 9% in PL vs 14 ± 7% in EXO, P