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• W3096958298 abstract "Background Ischemic cardiovascular diseases are the major causes of death in the world. Recently, mesenchymal or tissue progenitor cell derived exosomes are being investigated as an emerging therapeutic agent for treating cardiovascular diseases due to their potentials in restoring the damaged vasculature via promoting angiogenesis. Exosome therapy has the advantages in low immunogenicity and stability features comparing to cells 1. Tissue and organ derived exosomes could have a broader functionality than those derived from a homogeneous cell population as they contain multiple cell types. Placenta is a large and highly vascularized organ and placenta exosomes are known to play essential roles during pregnancy in supporting fetus development 2. Placenta is abundant with endothelial cells, epithelial cells, cytotrophoblast, syncytiotrophoblasts and stem cells including both hematopoietic and non-hematopoietic stem cells. It is perceivable that exosomes isolated from a placenta possess the functions of the cells in a placenta. Materials and Methods Full-term human placentas were obtained under the full consent of healthy donors. Placenta tissues were cultured in serum-free DMEM medium supplemented with antibiotics. After culturing 8 to 16 hours, supernatant was harvested, medium was changed every 8 to 12 hours for up to two days. Placenta derived exosomes (pExo) were isolated through sequential centrifugation of the supernatant of the placenta culture and final preparation were suspended in PBS and stored at -80oC. Quantification of pExo was performed with Bicinchoninic Acid protein assay kit. pExo size was determined using NanoSight. MILLIPLEX-MAP human cytokine/chemokine-PX41 was used to analyze the cytokine composition of pExo. In vitro cell migration assays were performed with seeding 1x10e5/mL human umbilical vessel endothelial cells (HUVEC) in basal media (BM) on the top chamber of an 8-um transwell on a 24-well plate with 500uL BM with or without pExo in the bottom chamber. After 6-24 hours of culture, transwells were observed under an inverted microscope after staining. In vitro cell proliferation assay was performed by seeding HUVEC at 2-4x10e3 cells/96-well with a water-soluble tetrazolium salt-based cell viability assay. To evaluate the pro-angiogenic activity of pExo in vivo, a hind limb ischemia (HLI) model in db/db diabetic mice was used. After surgical induction of HLI on Day 0, pExo from two different donors or vehicle (n=15 each group) was injected via i.v. at 100ug/mice or 100uL on Days 1, 6 and 11. The blood flow through hind paws was measured by Speckle Doppler. Results Utilizing the established cultivation and isolation methods, we achieved the average yield of pExo at 327±91 mg per placenta (n=10). pExo has an average size of 118±15 nm (n=10) as determined with nano-particle tracking analysis, consistent with consensus exosome size ranging 50nm to 200nm 1. Flow cytometry and Western blot analyses confirmed that pExo exhibits characteristic markers known for exosomes including CD9, CD63 and CD81. pExo contained abundant IL-8, HGF, FGF2, PDGF-BB, RANTES, MCP-1 and GM-CSF which are known to play roles in supporting angiogenesis and chemotaxis. In co-culture assays, pExo promoted the proliferation of HUVECs by 149±13% (n=5) comparing to basal medium controls. pExo also demonstated chemotactic activities comparable to that of complete HUVEC growth medium in stimulating the migration of HUVECs across membrane of transwell. Chlorpromazine and Pitstop2 abolished pExo-induced transwell-migration of HUVECs, suggesting that HUVECs may uptake pExo through endocytosis. In the HLI study, pExo treated mice showed significantly higher (50% to 250%) blood flow on Days 14, 28 and 35 comparing to the vehicle group. Histology analysis showed pExo treated mice had 40% to 140% higher density of CD34+ capillary on Day 35, suggesting a significant increase of angiogenesis in the pExo treated groups comparing to the vehicle group. Conclusion: In summary, pExo contains proangiogenic cytokines and chemokines and demonstrates the promising pro-angiogenic activities in vitro and in HLI mice model. These results support further development of pExo as a potential therapeutic agent for the treatment of cardiosvascular diseases. References: Phinney and Pittenger (2017). Stem Cells. 35:851 Sarker et al., (2014). Journal of Translational Medicine. 12:204 Disclosures Ye: Celularity Inc.: Current Employment, Patents & Royalties. He:Celularity Inc.: Current Employment, Current equity holder in private company. Gleason:Celularity Inc.: Current Employment, Current equity holder in private company. Stout:Celularity: Current Employment, Current equity holder in private company. Shah:Celularity: Current Employment. Somanchi:Celularity Inc.: Current Employment, Current equity holder in private company. Zhang:Celularity Inc.: Current Employment, Current equity holder in private company. Hariri:Celularity Inc.: Current Employment, Current equity holder in private company." @default.
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• W3096958298 date "2020-11-05" @default.
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• W3096958298 title "Human Placenta Derived Exosomes Enhance Ischemia Repair in a Mouse Model of Hind-Limb Ischemia" @default.
• W3096958298 doi "https://doi.org/10.1182/blood-2020-137452" @default.
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