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dc.contributor.authorDe Leon Rodriguez, Carlos Miguel
dc.identifier.citationSource: Dissertation Abstracts International, Volume: 78-09(E), Section: B.;Advisors: Arturo Casadevall.
dc.description.abstractCryptococcus neoformans (Cn) is a facultative intracellular pathogen that is the causative agent of cryptococcal meningoencephalitis, which occurs primarily in immunocompromised patients. Cn colonizes lung tissue after its spores are inhaled from the environment. In the lungs, Cn is phagocytosed by alveolar macrophages. Cn does not block phagosomal maturation, which is completed without major alterations. Cn and macrophage interaction results in one of the following outcomes (a) killing of Cn by the macrophage, (b) non-lytic exocytosis, or (c) lytic exocytosis. It is unknown what factors determine the outcome of the macrophage infection. Phagolysosomal membrane permeabilization (PMP) can be induced by exogenous or endogenous stimulus that results in partial or complete damage of the phagolysosomal membrane. PMP results in the release of phagolysosomal content into the cytosol, causing the activation of different macrophage cell death pathways. The lysosomal proteases cathepsins (CTS) are known to activate apoptosis after being released into the cytosol. Our laboratory and others have shown that caspases, proteases involved in programmed cell death, are activated after Cn phagocytosis. Activation of caspases occurs through different cell death pathways initiated by PMP. Previous studies from our laboratory and others have shown that Cn induces PMP. However, its effect in macrophage viability remains unknown. Recent studies have revealed that PMP is required for Cn intracellular replication and that interferon-y treatment protected the phagolysosomal membrane. The Cn virulence factor extracellular phospholipase B 1 (PLB 1) has been proposed to mediate its effect by degrading the phagolysosomal membrane, but its contribution to PMP remains unknown. We hypothesize that phagolysosomal membrane stability plays a role determining the outcome of Cn macrophage interaction.;Cn-infected macrophages that showed PMP were also apoptotic, whereas macrophage infected with heat-killed Cn showed no PMP. Cn-induction of PMP increases over time and with the number of intracellular Cn. Interestingly, cathepsin D, cathepsin H and cathpesin S protein expression increases over time in Cn-infected macrophage. Treatment with inhibitors of cysteine and aspartic cathepsins, or with caspases inhibitors did not block apoptosis. Strikingly, we showed that PLB 1 contributes to Cn-induction of PMP by activating apoptosis, a finding that provides a mechanism of action for this virulence factor. Previous studies have shown discrepancies on the pH of Cn-containing phagolysosomes. We found that Cn-infected macrophages that become apoptotic are not able to maintain the same phagolysosomal pH in comparison to non-apoptotic Cn-infected macrophages, which showed a more acidic pH. Finally, chemically induced PMP increases non-lytic exocytosis in Cn-infected macrophages. Together these data imply that the phagolysosome plays an important role determining the outcome of the Cn-macrophage interplay and, by extension, the host-pathogen interaction.
dc.publisherProQuest Dissertations & Theses
dc.titleCryptococcus neoformans and macrophage interaction is determine by the phagolysosomal membrane integrity

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