(PO07-56-22) Genetically Altered Bovine Milk Exosomes (BMEs) Evade Elimination by Murine Bone Marrow-Derived Macrophages (BMDMs)

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Presenting Author(s)

AN

Alice K. Ngu, Bachelor
– Graduate Student, University of Nebraska-Lincoln, Lincoln, Nebraska, United States

Co-Author(s)

Janos Zempleni, Ph.D.
– Professor, University of Nebraska-Lincoln, Lincoln, Nebraska, United States

Disclosure(s):

Alice K. Ngu, Bachelor: No relevant financial relationship(s) with ineligible companies to disclose.

Objectives: To develop BMEs that evade elimination by BMDMs.

Methods: Bovine mammary alveolar MAC-T cells secrete MEs and were used to engineer MEs that express proprietary protein features suspected to decrease elimination by BMDMs, denoted UNL1 and UNL2. MAC-T cells were transformed by using lentiviral vectors. BMEs featuring UNL1 or UNL2 were isolated from MAC-T cell culture media supernatant using polyethylene glycol (PEG) precipitation after removing cell debris by using a 0.22-µm filter. The BMEs were labeled using a carbonyl-reactive fluorescent dye and purified by ultracentrifugation. Primary BMDMs was isolated from mouse hind legs (C57BL/6J, aged 8-10 weeks) and seeded in 96-well plates for assessing BME uptake at a physiological concentration (1010 BMEs/mL). Uptake was compared to unmodified BMEs and normalized for BMDM density. Time points were compared pairwise by using t-test, and p < 0.05 was considered significant.

Results: The uptake of BME UNL1 by BMDMs was reduced by 37%, 42%, 48% and 47% as compared to unmodified BMEs after 12 h, 24 h, 36 h and 48h, respectively in culture dishes (p <.05; n = 5). Data are preliminary (n = 3), yet encouraging, for BME UNL2: The uptake of BME UNL2 was reduced by 41%, 44%, 46% and 46% compared to unmodified BMEs after 12 h, 24 h, 36 h and 48 h, respectively.

Conclusions: The elimination of BMEs UNL1 and UNL2 is significantly reduced compared unmodified BMEs in BMDM cultures. This is of great importance when using BMEs for delivering therapeutics.

Funding Sources: NIH P20GM104320, NIFA2016-67001-25301 and 2022-67021-36407), USDA Hatch and W-4002, and the SynGAP Research Fund (all to J. Z.). J.Z. serves as consultant for PureTech Health, Inc.