Abstract 4369361: Extracellular Vesicles Enhance Cardiovascular and Protein Adaptations in Mice With Pulmonary Hypertension

Introduction: Extracellular vesicles (EV) mediate intercellular communication and may have effects on disease processes. Research question: Can intravenous (IV) treatment with human bone-marrow mesenchymal stem-cell-derived (hBM-MSC) EV improve monocrotaline (MCT)-induced pulmonary hypertension (PH) in mice? Methods: WT Mice (male/female, 6 weeks-old) were randomly divided into four groups: Control (n= 8), MCT (n= 10), Control+EV (n= 7) and MCT+EV (n= 10). MCT and MCT+EV groups received weekly SC injections of MCT (60 mg/kg, 100 µl, Sigma, St. Louis, USA) diluted in 1x DPBS over 4 weeks, while Control and Control+EV received vehicle. Twenty-four hours after each SC injection, Control+EV and MCT+EV received IV injection (100 µl) of 3 x 10 6 hBM-MSC-derived EV diluted in 1x DPBS, while Control and MCT received vehicle. PH was characterized by morphometry, echocardiography, and lung histology. Blood samples (100 µl) were collected from the submandibular vein, centrifuged to purify EV in the serum by Total Exosome Isolation Kit (Invitrogen™), and analyzed by proteomics. Results: There was an attenuation of pulmonary vascular remodeling in MCT+EV mice (decreased by 41.2%, p= 0.0005) compared to the MCT group. MCT+EV mice also presented improved right ventricular (RV) function measured by Tricuspid Annular Plane Systolic Excursion (TAPSE), which increased by 51.7% in the MCT+EV vs the MCT group (p= 0.0004). MCT+EV mice also presented an attenuation of RV hypertrophy (decreased by 19.4%, p= 0.0115) along with an increase in expression of calsequestrin (increased by 33.5%, p= 0.0401) and calreticulin (increased by 29.8%, p= 0.0125) in RV compared to the MCT group. A KEGG pathway analysis of purified EV from mouse blood showed upregulation of proteasome and autophagy in MCT as compared with the Control group after 21 days. The same pathways were downregulated in MCT+EV in comparison with MCT. Further analysis revealed 7 overlapping proteins upregulated in MCT vs control and downregulated with MCT+EV vs MCT. Of the proteins downregulated in MCT vs control and upregulated with MCT+EV vs MCT, 9 overlapping proteins were found. Conclusion: EV treatment attenuated pulmonary vascular remodeling and RV hypertrophy, and improved parameters of RV cardiac function in mice with PH. This research identified 16 proteins in circulating EV that may be involved in the attenuation of PH. Future research investigating them may reveal new biomarkers or targets for treatment of PH.