Investigations into the fate and functionality of apoptotic vesicles

apoptosis;vesicles;microvesicles;chemotherapy;cancer

Black, Lane Vincent Charles ; McLellan, Alexander

University of Otago

关键词: apoptosis;    vesicles;    microvesicles;    chemotherapy;    cancer;   

Others  :  https://ourarchive.otago.ac.nz/bitstream/10523/4923/3/BlackLane%202014PhD.pdf

美国|英语

来源: Otago University Research Archive

【 摘 要 】

AbstractIn a variety of cell types, apoptosis leads to the fragmentation and packaging of cellular contents into discrete blebs. Blebs may be released from the plasma membrane as ;;apoptotic vesicles’ and ingested by a range of cells with phagocytic function including neutrophils, macrophages and dendritic cells. The clearance of these apoptotic debris by phagocytes is important in maintaining immune function; the failure of which can lead to an inflammatory response against self-antigens and can result in autoimmunity Our laboratory has discovered that exosomes (cell-derived nanoparticles) are trapped in the sub-capsular sinus (SCS) of the lymph nodes and in the MZ (MZ) of the spleen by a population of macrophages. This capture was discovered to be mediated by the sialoadhesin CD169 (sialic acid receptor). The objective of this study was to determine if vesicles derived from apoptotic cells (Apo-V) are captured in a similar manner, to investigate the effect of captured Apo-V on the immune system, and the relationship between Apo-V, CD169, and immune function. Previous studies had shown both tolerogenic and immunostimulatory effects of apoptotic microvesicles slthough this study is the first to show a relationship between apoptotic microvesicles, CD169, and immune function. Utilising a combination of intravenously and subcutaneously administered Apo-V, a frozen section binding assay, CD169 deficient (CD169-/-) mice, and flow cytometry this study confirmed that the capture of T lymphocyte (EL4)derived Apo-V is mediated through the sialoadhesin molecule CD169 expressed on macrophages found within the MZ of the spleen and the SCS of the lymph nodes. This pattern of capture is identical to that displayed previously in this laboratory with B lymphocyte-derived exosomes. Further in vivo studies using adoptively transferred ovalbumin-specific T lymphocytes, showed that subcutaneously administered Apo-V stimulated CD8+ T lymphocyte proliferation, whereas intravenously administered Apo-V did not. Additionally, this study utilised flow cytometry to investigate the immune response to Apo-V. In vivo cytotoxicity assays revealed that co-administration of Apo-V with dendritic cells significantly reduced the cytotoxic response of the dendritic cells to the target cells. However, there was no significant difference between the wild-type and CD169-/- groups indicating that this was independent of CD169. This investigation also revealed that intravenous administration of Apo-V co-cultured with ovalbumin protein significantly increased the hosts’ cytotoxic response to target cells when compared to the PBS control. Interestingly this response was significantly enhanced in the CD169-/-.Investigations into the possible immunological cell subset involved in this process revealed subtle changes in immune cells subsets in response to Apo-V; however further investigation is required in order to fully understand the contributions of these changes to immune function.In toto these results demonstrate that CD169+ macrophages are involved in the capture of Apo-V, and that these vesicles play a role in both the enhancement and attenuation of cytotoxic responses of the adaptive immune response.