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Surface expression of Tim-3 inhibitory molecule on antigen-specific CD8+ T cells expanded in vitro using dendritic cells for cell-based cancer immunotherapy
Svobodová, Hana ; Smrž, Daniel (advisor) ; Funda, David (referee)
Cancer is the second most common cause of death in the world, and the number of people with the disease increases each year. The therapy of the disease currently stands on four pillars; surgery, chemotherapy, radiotherapy, and immunotherapy. Through the past few years, immunotherapy has become the fastest developing treatment modality. However, despite its unprecedented efficacy in some patients, the majority of patients still does not respond to the therapy. Therefore, there is a need to investigate the mechanisms that make immunotherapy inefficient. Cell-based cancer immunotherapy is the treatment modality which uses live ex vivo-produced tumor-targeting immune cells to treat cancer. One of the mechanisms that may compromise its therapeutic efficacy is the expression of inhibitory molecules on the surface of the produced immune cells. Tim-3 is the inhibitory molecule which attracts attention in recent years. Tim-3 expression in the tumor cells and the tumor-infiltrating immune cells is often associated with worse prognosis and more aggressive forms of the disease. However, its role in the in vitro or ex vivo-produced immune cells is difficult to predict. In this work, an in vitro study model which is based on in vitro-produced antigen-specific CD8+ T cells with high expression of Tim-3 has been...
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Phosphatidylserine and phospholipid scramblase in mast cell signaling
Smrž, Daniel ; Dráber, Petr (advisor) ; Bezouška, Karel (referee) ; Šebo, Peter (referee)
6. CONCLUSIONS 1. We found that mast cell stimulation can induce PS externalization in the absence of secretory response. 2. We identified GPI-APs as molecules whose engagement can induce sustained and reversible non-apoptotic PS externalization. 3. GPI-AP-induced PS externalization was determined as non- apoptotic and distinct from the FceRl-induced PS externalization. 4. The effect of multiple triggering on PS externalization was additive and dependent on a tlpe of stimulus and cells engaged. 5. We identifred PLSCR1 as a molecule that becomes tyrosine phosphorylated in mast cells stimulated through GPI-APs. 6. We found that the PLSCR1 tyrosine phosphorylation is not associated with mast cell secretory response' and with GPI-AP- or FceRl-induced non-apoptotic PS externalization. 7. Using confocal microscopy and electron microscopy visualization of PLSCR1 in the course of mast cell activation we found that PLSCR1: (1) is not co-localized with extemalized PS, (2) is not co- localized with aggregated Thy-l.l or FceR[, and (3) does not form self-aggregates. 8. We děřelóped a modifred one-tube semi-nested PCR-ELISA. The modified assay showed higher sensitivity and specificity than the conventional hybridization-based and a modified semi-nested- based PCR-ELISA. Due to its versatility and robustness, the...
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