Dissertation (Ph.D.).--University of Texas Graduate School of Biomedical Sciences at San Antonio, 2007. / Vita. Includes bibliographical references.

Thesis (Ph.D.). / Written for the Dept. of Neurology and Neurosurgery. Title from title page of PDF (viewed 2009/06/09). Includes bibliographical references.

Orientador: Fabíola Freitas de Paula Lopes / Resumo: A função do espermatozoide pode ser comprometida por condições ambientais adversas. Estudos já demonstraram que a exposição in vivo e in vitro de espermatozoides bovinos a temperatura elevada induz a morte celular, reduz a motilidade espermática e o potencial fertilizante do espermatozoide. No entanto, as alterações celulares induzidas pelo choque térmico em espermatozoides bovinos ainda são controversas. Portanto, o objetivo deste estudo foi determinar o efeito do choque térmico em espermatozoides de touros holandeses na motilidade espermática, produção de espécies reativas de oxigênio (EROs), atividade mitocondrial, atividade de caspase, potencial de fertilização, cinética e desenvolvimento embrionário pré-implantacional. As amostras de sêmen foram descongeladas e submetidas ao gradiente de Percoll. Na amostra controle (sem incubação) os espermatozoides foram avaliados imediatamente após o gradiente de Percoll. Posteriormente, as amostras foram incubadas à 35 °C (Controle da temperatura testicular), 38,5 °C (temperatura corporal) e 41 °C (choque térmico) por 4 horas. O choque térmico de 41 °C reduziu a motilidade espermática após 2 h de incubação em comparação com 35 e 38,5 °C. A exposição de espermatozoides a diferentes temperaturas aumentou a produção de EROs, sendo este efeito mais acentuado no grupo 41°C em relação aos demais tratamentos. O aumento das EROs em espermatozoides submetidos ao choque térmico foi seguido da redução na atividade mitocondrial espermática e au... (Resumo completo, clicar acesso eletrônico abaixo) / Doutor

Effector caspase activation is a critical regulatory step in apoptosis, as well as an essential inductive cut in numerous non-death related processes that occur within complex cell systems. Here we report two novel studies detailing mechanisms by which effector caspase activation advances muscle cell differentiation. In the first study, we demonstrate that caspase 3 triggered DNA damage leads to rapid formation of XRCC1 repair foci within differentiating myonuclei, which dissipates as the maturation program proceeds. Skeletal myoblast deletion of XRCC1 does not impact cell growth, yet leads to perinatal lethality, with sustained DNA damage and impaired myofiber development. These observations demonstrate that the temporal deployment of the XRCC1-related DNA repair mechanisms are effector caspase mediated, and essential for muscle cell differentiation. In the second study, we sought to investigate whether effector caspase enzymes altered chromatin structure to promote the early differentiation of muscle progenitor cells. Past research has shown that Matrix Attachment region proteins known as Special AT-rich binding proteins are expressed abundantly in stem and progenitor cells, showing rapid decrease in expression as the cell advances into its mature phenotype. Here we demonstrate that effector caspase-7 is responsible for cleavage of Satb2, rather than caspase 3. Satb2 degradation alters the expressed genetic profile leading to acceleration of the muscle differentiation program. Our cumulative work adds novel roles in which effector caspases are vital in the development of cells.

One of the key features of cancer cells is the ability to escape programmed cell death (apoptosis). As a mechanism of apoptosis inactivation in cancer cells, somatic mutations of pro-apoptotic genes have been reported in many cancers. Caspase 10 is an initiator caspase whose physiological function remains poorly understood. Also the ability of caspase 10 to substitute for caspase 8 in the death receptors apoptotic pathway is still controversial. However, the fact that some of the mutations found in CASP10 gene was associated with apoptosis defects (79, 81) suggest that caspase 10 could be also important in apoptosis initiation. In our lab, there was found a heterozygous mutation in CASP10 gene of Jurkat (human T-acute lymphoblastic leukemia) clone resistant to TRAIL (J-TR1). This mutation influence the amino acid composition close to the active site of the enzyme. The aim of this thesis was to confirm the mutation by ARMS-PCR and to determine if an overexpression of normal (unmutated) or mutated caspase 10 D in TRAIL sensitive and/or TRAIL resistant Jurkat cells (J-WT and/or J-TR1) will influence TRAIL induced apoptosis. Mutation was confirmed. We created J-WT and J-TR1 stable clones transfected by vector with unmutated or mutated CASP10 D (CASP10 D WT or CASP10 D MUT). CASP10 D MUT overexpression in J-WT...

Glucocorticoids are widely used as anti-inflammatory and chemotherapeutic agents. However, prolonged use of glucocorticoids leads to osteoporosis. This study was designed to examine the mechanism of dexamethasone (DEX)-induced apoptosis in murine osteoblastic MC3T3-E1 cells. Total RNA was extracted from MC3T3-E1 cells treated with 10-7 M DEX for 6 h. DEX exerted a variety of effects on apoptotic gene expression in osteoblasts. Ribonuclease protection assays (RPA) revealed that DEX upregulated mRNA levels of caspases-1, -3, -6, -8, -11, -12, and bcl-XL. Western blot analysis showed enhanced processing of these caspases, with the appearance of their activated enzymes 8 h after DEX treatment. In addition, DEX also induced the activation of caspase-9. DEX elevated the levels of cleaved poly(ADP-ribose) polymerase and lamin A, a caspase-3 and a caspase-6 substrate, respectively. Expression of bcl-XL protein level was upregulated by DEX. Cytochrome c release was detected in the cytosol of DEX-treated cells. Furthermore, caspase-3 enzyme activity was elevated by 2-fold after DEX treatment for 7 h. Finally, early apoptotic cells were detected in cells treated with DEX for 3 h. Our results demonstrate that DEX-induced apoptosis involves gene activation of a number of caspases.

Previously we found that 3,4-(methylenedioxy)-1-(2′,3′- epoxypropyl)-benzene (safrole oxide) induced a typical apoptosis in A549 human lung cancer cells. In this study, we further investigated which caspases were activated by safrole oxide during the apoptosis. The data showed that the activity of caspase-3, -8, and -9 was significantly enhanced by the compound, which suggested that safrole oxide might be used as a caspase promoter to initiate lung cancer cell apoptosis.

京都大学 / 0048 / 新制・課程博士 / 博士(情報学) / 甲第21393号 / 情博第679号 / 新制||情||117(附属図書館) / 京都大学大学院情報学研究科知能情報学専攻 / (主査)教授 阿久津 達也, 教授 山本 章博, 教授 鹿島 久嗣 / 学位規則第4条第1項該当 / Doctor of Informatics / Kyoto University / DFAM

No description available.

The caspase family of proteins is critical to biological understanding, because they serve as the final arbiters of life and death, being the initiators and executioners of cell death. Specifically, caspase-7 plays a key role in apoptosis, however its full complement of targets within the cell has not yet been elucidated, nor has its function been targeted by drug design efforts. These factors stem from the lack of fundamental understanding of the structural dynamics of the protein, including the mobile loops that constitute the active site binding groove of caspase-7, and their ability to modulate the function of the protein. In this work, we describe the importance of the entire loop bundle for catalysis, demonstrate a novel approach for allosteric control using loop movement, develop computational methods to engineer a new binding site for an allosteric effector and discover a hereunto unseen native disulfide within caspase-7 that may contribute to specificity and catalysis. The information obtained within this study is applicable for not only the study of caspase-7, but also the greater field of apoptosis research.