To Investigate the Effects of Anthrax Lethal Toxin in Caspase-Independent Macrophage Cell Death
| Autor: | Wang, Leng-Lin, 王稜琳 |
|---|---|
| Rok vydání: | 2015 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 103 Anthrax lethal toxin (LeTx) is secreted by Bacillus anthracis as two subunits, protective antigen (PA) and lethal factor (LF). LF enters mammalian cells through PA-receptor binding mediated endocytosis, then cleaves mitogen-activated protein kinase kinases (MAPKK) and leads to cell death. Because macrophages build a first line of defense in the innate immune system, the targeting of macrophages by LF may incapacitate the host immune response and initiate the infection. However, the molecular basis of LeTx-mediated cell death is still not clear. In our previous study, differentiated THP-1 cells undergoing LeTx-mediated death show not typical features of apoptosis, they are neither protected by the pan-caspase inhibitor ZVAD-FMK nor appear active forms of caspase-3. Thus, in this study, we would investigate how LeTx kills differential THP-1 cells in a caspase-independent manner and what the mechanisms regulate the fate of macrophage underlying the effects of LeTx. Based on current hypothesis, LeTx could enhance ROS production dramatically and lead to cytotoxicity through a caspase-independent pathway in differential THP-1 cells. Therefore, we propose two objectives to study LeTx-induced caspase-independent cell death by using THP-1 cells treated with LeTx and the pan-caspase inhibitor Z-VAD-FMK simultaneously. First of all, we found that caspase 3, an apoptosis marker, is not activated with LeTx treatment. And LeTx could lead to cell death with or without Z-VAD-FMK. It is proved that LeTx mediates cell death without caspase 3 activation. Secondly, the caspase-independent cell death could classify into two categories, autophagic cell death and apoptosis-like cell death. Autophagy is the fundamental catabolic mechanism that involves cell degradation of unnecessary or dysfunctional cellular components through the actions of lysosomes. The breakdowns of cellular components ensure cellular survival during starvation and stress by maintaining cellular energy levels. Recent studies have shown that autophagy is related with cell death closely. But the certain route of causing autophagic cell death is still not clear. We found that autophagy is inhibited in protein level and the expressions of some autophagy-related genes are altered in RNA level. In addition to autophagy, alternative route, apoptosis-like cell death correlates with mitochondrial dysfunction, apoptosis-inducing factor (AIF) translocation, poly (ADP-ribose) polymerase (PARP) activation and cathepsin B release. And we confirmed that mitochondria lost the function via LeTx treated. Furthermore, we will examine whether LeTx-mediated cytotoxicity involves these characteristics of apoptosis-like cell death in the future. In conclusion, this study will provide novel insight into the mechanism of LeTx-induced caspase-independent cell death in macrophage, and serve as the basis for in vivo studies to further investigate the effects of LeTx on the first line of innate immunity. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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