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Browsing by Subject "Cell interaction."

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    In vitro and in vivo immunomodulatory effects of extracellular vesicles
    (2013) Şahin, Mehmet
    The major theme of this thesis was to characterize and understand the immunomodulatory potential of extracellular vesicles (EVs) isolated from different mouse and human cell lines on immune cells. To this end, we first purified and characterized the particle nature of exosomes and microparticles (MPs) and exosomes by AFM and DLS analyses. Next, we documented their in vitro or in vivo differential uptake/internalization kinetics by immune cells. Finally, we tested the potential application of EVs as a drug delivery system. The physicochemical characterization studies confirmed that EVs were in vesicular form and upon reconstitution followed by lyophilization they retained their original sizes (i.e. size ranges were 100-150 nm for exosomes, and 250-500 nm for MPs, respectively). MPs derived from mouse cell lines (macrophage, T-cell and fibroblast) were stained and administered in vitro and in vivo to track their internalization by immune cells. When incubated in culture, origins of MPs greatly affected uptake rate and ratio by different immune cells. This differential internalization pattern of distinct MPs was reproduced when they were injected i.p. to mice. When incubated in culture B-cells took up the most MPs from fibroblast origin cells. Macrophages of peritoneal exudate cells took up RAW264.7 derived MPs at the highest level upon 24h post-ip injection. We tested whether MPs have a role in transportation or presentation of bacterial products during an ongoing infection by adhering circulating pathogenic by-products and carrying them to distant immune cells. Our data suggest that MPs could contribute to the severity of ongoing infection in vivo, since MPs from different cells can adhere distinct ligands such as LPS or DNA or even RNA and transmit these ligands to naive innate immune cells augmenting the immunostimulatory response raised against these ligands. Lastly, the immunotherapeutic potential of EVs harboring immunosuppressive synthetic oligodeoxynucleotide sequence namely, A151 was tested following dehydrationrehydration method on mouse splenocytes. EV-associated A151 displayed improved inhibition of immune activation triggered by TLR7/8 and TLR9 ligands on spleen cells. When taken together, this study established that various types of EVs derived from different cells induces plethora of activities on immune cells. Furthermore, EVs are potential drug carrier systems when loaded externally with suitable agents and can be harnessed in immunotherapy of diseases.

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