Modulation of therapeutic cells exosome content by autophagy

Information

  • Research Project
  • 10054819
  • ApplicationId
    10054819
  • Core Project Number
    R15GM128189
  • Full Project Number
    3R15GM128189-01S1
  • Serial Number
    128189
  • FOA Number
    PA-18-586
  • Sub Project Id
  • Project Start Date
    7/1/2018 - 5 years ago
  • Project End Date
    6/30/2021 - 2 years ago
  • Program Officer Name
    MAAS, STEFAN
  • Budget Start Date
    7/1/2018 - 5 years ago
  • Budget End Date
    6/30/2021 - 2 years ago
  • Fiscal Year
    2020
  • Support Year
    01
  • Suffix
    S1
  • Award Notice Date
    4/13/2020 - 4 years ago
Organizations

Modulation of therapeutic cells exosome content by autophagy

Project Summary We are currently investigating the use of human mesenchymal stromal cells (MSCs) for tissue repair by injecting MSCs into the damaged organ. Recent discoveries indicate that many of the therapeutic benefits of MSCs can be attributed to secretion of various biomolecules which can be secreted via exosomes, small membrane vesicles of endocytic origin. Most cell types secrete exosomes which contain proteins, DNA, mRNA, and microRNA, all of which which are thought to play a role in cell-cell communication. While previous research efforts largely focused on the characterization of proteins found in exosomes, our current research focuses on exosomal RNAs. Increasing evidence suggests that exosome formation and release are regulated by the autophagy pathway, a homeostatic quality control pathway that recycles proteins and organelles via recognition, sequestration, and lysosomal degradation. Conditions that stimulate autophagy pathway can inhibit exosome release, but at the same time pharmacological inhibitors of autophagy enhance the release of exosomes. For our study, we propose utilizing a subtype of MSCs called ?marrow-isolated adult multilineage inducible? (MIAMI) cells due to their ease of isolation from bone marrow, differentiation capacity, their immunomodulatory and tissue repair capacities, and ability to secrete various chemokines/growth factors. We hypothesize that autophagy mediates release of exosomes from MIAMI cells, regulates their RNA content and their immunomodulatory capacity. To stimulate MIAMI cells, we will expose them to inflammatory response stimulator, IFN?, while simultaneously applying pharmacologic stimulators or inhibitors of autophagy. Subsequently, we will isolate and characterize MIAMI cell-derived exosomes by using NanoSight, electron microscopy and immunoblotting to characterize and compare exosomes size distributions, exosome yield and markers (CD9, CD63 and CD81) (Aim 1). We will then determine how modulation of autophagy regulates exosomal RNA content by identifying and validating long (more than 200 nucleotides) and short (less than 200 nucleotides) RNAs in MIAMI cells-derived exosomes (Aim 2). Lastly, we will evaluate immunoregulatory properties of MIAMI cells upon modulation of autophagy using MIAMI cell-T cell co-cultures and subsequent flow cytometry analysis to assess T cell markers of proliferation and cytokine secretion (Aim 3). Results of this mechanistic study will increase our understanding of the role that autophagy plays in regulating RNA content in exosomes and it will also reveal whether targeting autophagy could be used to manipulate RNA content and subsequently immunomodulation. Ultimately, such knowledge is anticipated to foster further development of cell therapies for tissue regeneration. 1

IC Name
NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES
  • Activity
    R15
  • Administering IC
    GM
  • Application Type
    3
  • Direct Cost Amount
    47404
  • Indirect Cost Amount
    24650
  • Total Cost
    72054
  • Sub Project Total Cost
  • ARRA Funded
    False
  • CFDA Code
    859
  • Ed Inst. Type
    SCH ALLIED HEALTH PROFESSIONS
  • Funding ICs
    NIGMS:72054\
  • Funding Mechanism
    Non-SBIR/STTR RPGs
  • Study Section
  • Study Section Name
  • Organization Name
    NOVA SOUTHEASTERN UNIVERSITY
  • Organization Department
    OTHER HEALTH PROFESSIONS
  • Organization DUNS
    002971240
  • Organization City
    Fort Lauderdale
  • Organization State
    FL
  • Organization Country
    UNITED STATES
  • Organization Zip Code
    333147796
  • Organization District
    UNITED STATES