RESUMO
Cell death plays a central role in normal physiology and in disease. Common to apoptotic and necrotic cell death is the eventual loss of plasma membrane integrity. We have produced a small organoarsenical compound, 4-(N-(S-glutathionylacetyl)amino)phenylarsonous acid, that rapidly accumulates in the cytosol of dying cells coincident with loss of plasma membrane integrity. The compound is retained in the cytosol predominantly by covalent reaction with the 90 kDa heat shock protein (Hsp90), the most abundant molecular chaperone of the eukaryotic cytoplasm. The organoarsenical was tagged with either optical or radioisotope reporting groups to image cell death in cultured cells and in murine tumors ex vivo and in situ. Tumor cell death in mice was noninvasively imaged by SPECT/CT using an (111)In-tagged compound. This versatile compound should enable the imaging of cell death in most experimental settings.
Assuntos
Arsenicais/química , Carbocianinas , Proteínas de Choque Térmico HSP90/metabolismo , Neoplasias/diagnóstico , Ácido Pentético , Peptídeos , Animais , Arsenicais/metabolismo , Carbocianinas/química , Carcinoma Pulmonar de Lewis/diagnóstico , Morte Celular , Neoplasias Colorretais/diagnóstico , Humanos , Células Jurkat , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Ácido Pentético/química , Peptídeos/química , Peptídeos/metabolismo , Ligação Proteica , Radioisótopos/químicaRESUMO
We describe the evaluation of three live, attenuated deltaphoP/Q Salmonella enteric serovar Typhimurium strains expressing PEB1 minus its signal sequence (PEB1-ss) from three different plasmids: a pBR-based asd plasmid, an arabinose-based runaway plasmid, which each expressed PEB1-ss in the bacterial cytosol, and a PEB1::HlyA fusion plasmid that directs secretion of PEB1-ss into the extracellular milieu. Serum IgG responses specific for PEB1-ss were induced by pBR-derived and runaway plasmids, with 100 and 90% seroconversion, respectively, at a 1:500 dilution of anti-sera as measured by Western blot analysis, while the PEB1-ss::HlyA fusion plasmid induced serum IgG in only 20% of the mice. Although significant levels of anti-PEB serum IgG were induced, no protection against oral Campylobacter jejuni challenge was observed.
Assuntos
Antígenos de Bactérias/imunologia , Vacinas Bacterianas/imunologia , Infecções por Campylobacter/prevenção & controle , Campylobacter jejuni/imunologia , Salmonella typhimurium/imunologia , Administração Oral , Animais , Anticorpos Antibacterianos/sangue , Antígenos de Bactérias/genética , Vacinas Bacterianas/administração & dosagem , Vacinas Bacterianas/genética , Western Blotting , Infecções por Campylobacter/patologia , Infecções por Campylobacter/fisiopatologia , Contagem de Colônia Microbiana , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Fezes/microbiologia , Feminino , Vetores Genéticos , Proteínas Hemolisinas/genética , Proteínas Hemolisinas/imunologia , Imunoglobulina G/sangue , Camundongos , Camundongos Endogâmicos BALB C , Plasmídeos , Sinais Direcionadores de Proteínas/genética , Salmonella typhimurium/genética , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/imunologiaRESUMO
The Exposure to Violence Screening Measure (EVSM) was evaluated as a structured interview for assessing violence exposure among inner-city youth. Psychometric analysis of the measure on a sample of 352 inner-city teenagers indicated adequate internal consistency. The validity of the EVSM was supported by significant correlations between the total score and measures of life stress and behavioral problems.