Orientador: Eliana Aguiar Petri Nahas / Resumo: Objetivo: Avaliar os fatores de risco cardiovascular em mulheres na pós-menopausa tratadas de câncer de mama comparadas a mulheres na pós-menopausa sem câncer de mama e a associação das heat shock proteins (HSP) 60 e 70 com o câncer de mama em mulheres na pós-menopausa. Métodos: Realizou-se estudo clínico de corte transversal com 96 mulheres tratadas de câncer de mama comparadas a 192 mulheres na pós-menopausa (controle), com idades entre 45 e 75 anos. Foram incluídas no grupo principal mulheres com amenorréia > 12 meses e idade ≥ 45 anos, com diagnóstico histológico de câncer de mama, sem doença metastática e sem doença cardiovascular (DCV) estabelecida. O grupo controle foi constituído por mulheres com amenorréia > 12 meses, idade ≥ 45 anos, sem câncer de mama e DCV. Os grupos foram pareados por idade, tempo de menopausa e índice de massa corpórea (IMC) na proporção 1 caso para 2 controles, conforme cálculo amostral, com o mínimo de 92 pacientes tratadas de câncer de mama. Dados clínicos e antropométricos (IMC e circunferência da cintura) foram coletados por meio de entrevista e exame físico. Para análise bioquímica foram solicitados colesterol total, HLD, LDL, triglicerídeos, glicose e insulina. Foram consideradas com síndrome metabólica (SM), as mulheres que apresentaram três ou mais critérios diagnósticos: circunferência da cintura (CC) > 88 cm; TG ³ 150 mg/dL; HDL colesterol < 50 mg/dL; pressão arterial ³ 130/85 mmHg; glicose ³ 100 mg/dL. Para determinação das concentra... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: To evaluate cardiovascular risk factors in postmenopausal breast cancer survivors as compared to postmenopausal women without breast cancer and the association of heat shock proteins (HSP) 60 and 70 with breast cancer in postmenopausal women. Methods: In this cross-sectional study, 96 postmenopausal breast cancer survivors were compared with 192 postmenopausal women (controls), aged 45 to 75 years. The principal group included women with amenorrhea > 12 months and age ≥45 years, with histological diagnosis of breast cancer, without metastatic disease and without established cardiovascular disease (CVD). The control group consisted of women with amenorrhea > 12 months, age ≥45 years, without breast cancer and CVD. The groups were matched by age, time since menopause, and body mass index (BMI) in the proportion 1 case for 2 controls, according to the sample calculation, with a minimum of 92 breast cancer survivors. Clinical and anthropometric data (BMI and waist circumference) were collected by interview and physical examination. For biochemical analysis, total cholesterol, HLD, LDL, triglycerides, glucose and insulin levels were measured. Women who presented three or more of the following criteria were diagnosed as having metabolic syndrome (MetS): waist circumference (WC)> 88 cm; TG≥ 150 mg/dL; HDL cholesterol <50 mg/dL; blood pressure ≥ 130/85 mmHg; glucose ≥ 100 mg/dL. For measuring plasma concentrations of HSP 60 and 70, immunoassays by ELISA technique were used. Atheroscl... (Complete abstract click electronic access below) / Doutor

Heat shock protein 90 (HSP90), a conserved molecular chaperone, has become a potential molecular target for cancer therapeutics. HSP90 inhibition (HSP90i) causes inhibition of several oncogenic pathways simultaneously and leads to anti-cancer activities in multiple cancers including in triple-negative breast cancer (TNBC). TNBC is a subtype of breast cancer with poor prognosis and lack of approved targeted therapies. Although HSP90i has shown promising initial clinical data, resistance to HSP90i can still arise in TNBC patients and its resistance mechanisms are not yet understood. In this study, using an in vitro system, we report for the first time the isolation of TNBC cells with acquired resistance to HSP90i. Proteome and whole transcriptome profiling, and a bioactive small molecule screen were performed to identify the molecular basis of resistance to HSP90i and potential therapeutic approaches to overcome acquired resistance to HSP90i in TNBC cells. Two independent HSP90i-resistant clones were acquired through prolonged exposure of a TNBC cell line (Hs578T) to HSP90i. The clones showed significant resistance to HSP90i, notably to resorcinol-based HSP90i. The HSP90i-resistant clones also shared genomic sequence variants, suggesting a pre-existing population of resistant cells within the parental cells. We demonstrate that upregulated expression of UGT1A9, possibly due to an increased intrinsic oxidative stress, is associated with acquired resistance to resorcinol-based HSP90i in TNBC cells, and sensitivity to HSP90i can be restored with a competitive inhibitor of UGT1A9. The HSP90i-resistant clones also exhibited slower growth and upregulated IL6- mediated JAK2-STAT3 survival signalling pathway, which might contribute to the crossresistance to chemotherapeutics and other targeted therapies seen in the clones. Finally, we demonstrate that inhibition of JAK2-STAT3 signalling pathway is able to increase the cytotoxic effects of HSP90i to TNBC cells. We conclude that by using in vitro assays, we are able to identify potential mechanisms of acquired resistance to HSP90i in TNBC cells. We propose that expression of UGT1A9 or STAT3 might be a potential biomarker of sensitivity to HSP90i in TNBC cells. A combined inhibition of HSP90 and JAK2 might be a potential therapeutic approach for the development of effective targeted therapies in TNBC patients.

Heat shock proteins (HSPs) play important roles in the process of maintaining proteostasis in a cell. HSP72, the inducible form of the HSP70 family, is expressed in response to stress on the cell or tissue, including those stresses caused by tumor growth. Increasing evidence suggests that HSP72 is necessary for a cancerous cell to survive under the stresses of a tumor microenvironment. This has naturally raised interest in identifying an inhibitor selective for HSP72. The Haystead Laboratory at Duke University identified such a small-molecule inhibitor, referred to as HS-72, and proposed the scaffold as an ideal starting point to develop a family of therapeutic agents targeting HSP72. This work examines the potency and effectiveness of HS-72 and a number of its analogs developed by the Haystead Laboratory. These results suggest that HS-159 is a more effective inhibitor of HSP72 on a range of human tumor cell lines than HS-72. Further studies are needed to quantify how much more potent HS-159 is than HS-72 and potentially identify even more potent compounds.

Alcanivorax species are gram negative bacteria that usually require aliphatic hydrocarbon as the sole carbon source for growth. The ability to use petroleum in polluted environments as energy source makes Alcanivorax species biotechnologically relevant in bioremediation. Universal stress proteins confer ability to respond to unfavourable environments, thus the present study was done to analyse the genomic context of genes for universal stress proteins in Alcanivorax genomes. A combination of bioinformatics and visual analytics approaches were used to analyze genome-enabled data including sequences and gene expression datasets. On the basis of transcription unit and adjacent genes, two types of Alcanivorax USP genes observed were (i) adjacent to cyclic nucleotide-binding and oxygen sensing functions; and (ii) adjacent to sulfate transporter function. Both types of genes encode two universal stress protein domains (pfam00582) also referred to as tandem-type universal stress proteins. The sequence and structural characteristics of each of the four USP domains in Alcanivorax needs to be further investigated. This dissertation research evaluated data from Alcanivorax borkumensis cells (grown on either pyruvate or hexadecane as carbon source) that were stressed with 1-octanol and data collected at 15 min, 30 min, 60 min and 90 min after 1-octanol addition. The two genes for Alcanivorax borkumensis SK2 universal stress proteins, ABO_1340 and ABO_1511, had the same direction of expression for adjacent genes. A limitation of this research was that findings based on bioinformatics and visual analytics methods may need confirmation by molecular methods. The differences observed may also reflect the quality of the annotations provided for genes. The sequence and structural characteristics of each of the four USP domains in Alcanivorax needs to be further investigated. Further research is needed on the relationship between number, length and order of genes in operons that include genes for universal stress proteins. Additionally, in vitro studies to confirm the functional prediction made from the genomic context of the universal stress protein in Alcanivorax genome. The knowledge discovered from this genome context analytics research could contribute to improving the performance of Alcanivorax species in bioremediation of environments polluted with petroleum / College of Agriculture and Environmental Sciences / M. Sc. (Environmental Science)

xi, 52 p. ; ill. (some col.) A print copy of this title is available through the UO Libraries under the call number: SCIENCE QH445.2 .B35 2008 / Activation of Heat shock proteins (Hsps) is critical to adaptation to low oxygen levels (hypoxia) and enduring the oxidative stress of reoxygenation. Hsps are known to be regulated by Heat shock factor (Hsf), but my results demonstrate an unexpected regulatory link between the oxygen sensing and heat shock pathways. Hsf transcription is upregulated during hypoxia due to direct binding by Hypoxia-inducible Factor-1 (HIF-1) to HIF-1 response elements in an Hsf intron. This increase in Hsf transcripts is necessary for full Hsp induction during hypoxia and reoxygenation. The HIF-1-dependent increase in Hsps has a functional impact, as reduced production of Hsps decreases viability of adult flies exposed to hypoxia and reoxygenation. Thus, HIF-1 control of Hsf transcriptional levels is a regulatory mechanism for sensitizing heat shock pathway activity in order to maximize production of protective Hsps. This cross-regulation represents a mechanism by which the low oxygen response pathway has assimilated complex new functions by regulating the heat shock pathway's key transcriptional activator. Beyond studying the regulation of specific genes. I have also developed a method to identify small, yet important, changes within entire genomes. Genetic variation is the foundation of phenotypic traits, as well as many disease states. Variation can be caused by inversions, insertions, deletions, duplications, or single nucleotide polymorphisms (SNPs) within a genome. However, identifying a genetic change that is the cause of a specific phenotype or disease has been a difficult and laborious task for researchers. I developed a technique to quickly and accurately map genetic changes due to natural phenotypic variation or produced by genetic screens. I utilized massively parallel, high-throughput sequencing and restriction site associated DNA (RAD) markers, which are short tags of DNA adjacent to the restriction sites. These RAD markers generate a genome-wide signature of fragments for any restriction enzyme. Taken together with the fact that the vast majority of organisms have SNPs that disrupt restriction site sequences, the differences in the restriction fragment profiles between individuals can be compared. In addition, by using bulk segregant analysis, RAD tags can be used as high-density genetic markers to identify a genetic region that corresponds to a trait of interest. This dissertation includes both previously published and unpublished co-authored materials. / Adviser: Eric Johnson

The aim of this work was to examine the expression of certain Hsp genes upon heat exposure in spruce bark beetle Ips typographus. We determined the level in unstressed and heat-exposed animals, and attempted to assess the importance of Hsp proteins for animals' survival upon heat treatment. We used RNA interference to knock down the expression of these genes, and analyzed the influence on animals exposed to elevated temperatures.

Submitted by Daniel De Araujo Brito Buttros (danielbuttros@hotmail.com) on 2018-06-14T10:28:06Z No. of bitstreams: 1 DOUTORADO DANIEL BUTTROS.pdf: 8920662 bytes, checksum: 8d3d25caa8f04b9c86ef89d60f2cab83 (MD5) / Approved for entry into archive by Sulamita Selma C Colnago null (sulamita@btu.unesp.br) on 2018-06-14T17:51:10Z (GMT) No. of bitstreams: 1 buttros_dab_dr_bot.pdf: 8920662 bytes, checksum: 8d3d25caa8f04b9c86ef89d60f2cab83 (MD5) / Made available in DSpace on 2018-06-14T17:51:10Z (GMT). No. of bitstreams: 1 buttros_dab_dr_bot.pdf: 8920662 bytes, checksum: 8d3d25caa8f04b9c86ef89d60f2cab83 (MD5) Previous issue date: 2018-04-19 / Objetivo: Avaliar os fatores de risco cardiovascular em mulheres na pós-menopausa tratadas de câncer de mama comparadas a mulheres na pós-menopausa sem câncer de mama e a associação das heat shock proteins (HSP) 60 e 70 com o câncer de mama em mulheres na pós-menopausa. Métodos: Realizou-se estudo clínico de corte transversal com 96 mulheres tratadas de câncer de mama comparadas a 192 mulheres na pós-menopausa (controle), com idades entre 45 e 75 anos. Foram incluídas no grupo principal mulheres com amenorréia > 12 meses e idade ≥ 45 anos, com diagnóstico histológico de câncer de mama, sem doença metastática e sem doença cardiovascular (DCV) estabelecida. O grupo controle foi constituído por mulheres com amenorréia > 12 meses, idade ≥ 45 anos, sem câncer de mama e DCV. Os grupos foram pareados por idade, tempo de menopausa e índice de massa corpórea (IMC) na proporção 1 caso para 2 controles, conforme cálculo amostral, com o mínimo de 92 pacientes tratadas de câncer de mama. Dados clínicos e antropométricos (IMC e circunferência da cintura) foram coletados por meio de entrevista e exame físico. Para análise bioquímica foram solicitados colesterol total, HLD, LDL, triglicerídeos, glicose e insulina. Foram consideradas com síndrome metabólica (SM), as mulheres que apresentaram três ou mais critérios diagnósticos: circunferência da cintura (CC) > 88 cm; TG ³ 150 mg/dL; HDL colesterol < 50 mg/dL; pressão arterial ³ 130/85 mmHg; glicose ³ 100 mg/dL. Para determinação das concentrações plasmáticas de HSP 60 e 70 foram empregados imunoensaios pela técnica de ELISA. A doença aterosclerótica foi determinada pela espessura do complexo médio-intimal (CMI > 1mm) das artérias carótidas e/ou pela presença de placa ateromatosa, avaliadas pela ultrassonografia carotídea (scanner 14 duplex). Para análise estatística foram empregados: Teste t-student, Distribuição Gama (variáveis assimétricas), Teste do Qui-Quadrado e Regressão Logística (odds ratio-OR). Resultados: A média de idade das pacientes tratadas de câncer de mama foi de 59,8 ± 9,0 anos com tempo médio de seguimento de 4,2 ± 2,0 anos. Na comparação entre os grupos, mulheres tratadas de câncer de mama apresentaram valores médios elevados da pressão sistólica e diastólica (p<0.001) e valores médios de triglicerídeos e glicose, acima dos valores desejáveis (p<0.05). As pacientes do grupo principal apresentaram valores mais altos de HSP 60 e mais baixos de HSP 70 quando comparados ao controle (p<0.05). Foi observada maior ocorrência de diabetes, SM e placa ateromatosa entre as mulheres tratadas de câncer mama quando comparadas ao controle (19,8% vs 6,8%; 54,2% vs 30,7%; 19.8% vs 9.4%, respectivamente) (p<0.05). Na análise de risco ajustado para idade, tempo de menopausa e IMC, as mulheres tratadas de câncer de mama apresentaram risco significativamente aumentado para SM (OR=4.21; IC 95% 2.28-7.76), presença de placa ateromatosa (OR=2.61; IC 95% 1.19-5.72), diabetes (OR=4.42; IC 95% 1.86-10.49), hipertrigliceridemia (OR=2.32; IC 95% 1.33-4.0) e circunferência de cintura aumentada (OR=11.22; IC 95% 4.0 – 31.65), quando comparadas as mulheres sem câncer de mama. Conclusão: Mulheres tratadas de câncer de mama apresentaram maior risco para síndrome metabólica, diabetes, doença aterosclerótica, hipertrigliceridemia e obesidade abdominal, importantes fatores de risco para doença cardiovascular, quando comparadas as mulheres na pós-menopausa sem câncer de mama. Assim como, mulheres tratadas de câncer de mama apresentaram valores elevados de HSP 60 e diminuídos de HSP 70 quando comparadas às mulheres sem câncer. / To evaluate cardiovascular risk factors in postmenopausal breast cancer survivors as compared to postmenopausal women without breast cancer and the association of heat shock proteins (HSP) 60 and 70 with breast cancer in postmenopausal women. Methods: In this cross-sectional study, 96 postmenopausal breast cancer survivors were compared with 192 postmenopausal women (controls), aged 45 to 75 years. The principal group included women with amenorrhea > 12 months and age ≥45 years, with histological diagnosis of breast cancer, without metastatic disease and without established cardiovascular disease (CVD). The control group consisted of women with amenorrhea > 12 months, age ≥45 years, without breast cancer and CVD. The groups were matched by age, time since menopause, and body mass index (BMI) in the proportion 1 case for 2 controls, according to the sample calculation, with a minimum of 92 breast cancer survivors. Clinical and anthropometric data (BMI and waist circumference) were collected by interview and physical examination. For biochemical analysis, total cholesterol, HLD, LDL, triglycerides, glucose and insulin levels were measured. Women who presented three or more of the following criteria were diagnosed as having metabolic syndrome (MetS): waist circumference (WC)> 88 cm; TG≥ 150 mg/dL; HDL cholesterol <50 mg/dL; blood pressure ≥ 130/85 mmHg; glucose ≥ 100 mg/dL. For measuring plasma concentrations of HSP 60 and 70, immunoassays by ELISA technique were used. Atherosclerotic disease was determined by the intima-media thickness (IMT> 1mm) of the carotid arteries and / or by the presence of atheromatous plaque, assessed by carotid ultrasound (scanner duplex). For statistical nalysis, Student's t-test, Gamma Distribution (asymmetric variables), Chi-Square Test, and Logistic Regression (odds ratio-OR) were used. Results: The mean (± SD) age of breast cancer survivors was 59.8 ± 9.0 years, with a mean (± SD) follow-up of 4.2 ± 2.0 years. The breast cancer survivors presented high mean values of systolic and distal pressure (p <0.001), and mean values of triglycerides and glucose, above the desirable values (p <0.05). Patients with breast cancer had higher levels of HSP 60 and lower HSP 70 when compared to control (p <0.05). There was a higher occurrence of diabetes, MetS, and atheromatous plaque among the breast cancer survivors when compared to the control group (19.8% vs 6.8%, 54.2% vs. 30.7%, 19.8% vs. 9.4% respectively) (p <0.05). In the risk analysis adjusted for age, time since menopause and BMI, women treated for breast cancer had a significantly increased risk for MetS (OR = 4.21, 95% CI 2.28-7.76), presence of atheromatous plaque (OR = 2.61, CI 95% CI 1.19-5.72), diabetes (OR=4.42; IC 95% 1.86-10.49), hypertriglyceridemia (OR = 2.32, 95% CI 1.33-4.0) and large waist circumference (OR = 11.22, 95% CI 4.0 - 31.65 ) when compared to women without breast cancer. Conclusion: Postmenopausal breast cancer survivors had a higher risk for metabolic syndrome, diabetes, atherosclerotic disease, hypertriglyceridemia and abdominal obesity, important risk factors for cardiovascular disease when compared to postmenopausal women without breast cancer. Similarly, women treated for breast cancer presented high levels of HSP 60 and decreased HSP 70 when compared to women without cancer.

Os genes de choque térmico groES e groEL de Caulobacter crescentus foram isolados utilizando-se os genes homólogos de E.coli como sonda e por sequenciamento demonstrou-se que estes genes estão organizados na forma de um operon em um fragmento de DNA de aproximadamente 2,5 kb, contendo também sua região regulatória. \"Northern blots\" de RNA total de células crescidas a 300C ou submetidas a choque térmico mostraram a presença de um único RNA de tamanho aproximado de 2,3kb, altamente induzido por choque térmico, permanecendo em altos níveis mesmo após longos períodos de choque térmico. Amostras de RNA total de células sincronizadas, de diferentes estágios do ciclo celular de Caulobacter, foram também analisadas mostrando que os níveis do mRNA groESL variam durante o ciclo, apresentando um máximo na célula prédivisional. Análises através de \"Western blot\" mostraram uma pequena variação nos níveis da proteína GroEL ao longo do ciclo celular, sendo os tempos 60 e 120 minutos, respectivamente, os pontos de mínimo e máximo acúmulo da proteína concordando com os resultados obtidos em \"N orthern blots\". O mesmo tipo de análise foi feito com extratos totais obtidos a partir uma população mista de células crescidas a 300C e submetidas a choque térmico, observando-se o acúmulo da proteína até 60 minutos depois do choque térmico, com aumento da ordem de 5 vezes nos níveis de GroEL, níveis estes que diminuem lentamente a partir deste ponto. Os inícios de transcrição foram determinados em experimentos de \"primer extension\" utilizando-se RNA total de células incubadas 300C e de células submetidas a diferentes condições de choque térmico. Dois possíveis sítios de início de transcrição foram determinados nas posições -119 e -88 do ATG da metionina iniciadora de groES, sendo as regiões -10 e -35 dos promotores correspondentes (P 1 e P2) identificadas. Somente a transcrição iniciando a partir de P2, que apresenta características de um promotor transcrito pelo &#963;32, aumenta durante o choque térmico. Fusões de transcrição com o vetor repórte placZ/290 e a região 5\' regulatória do operon groESL foram construídas para identificar as sequências responsáveis pelo controle por choque térmico e pelo controle temporal. Fusões de transcrição contendo deleções na região 5\' do operon mostraram que sequências a montante do promotor P2 não são necessárias para a indução por choque térmico ou para o controle temporal. Fusões de transcrição contendo mutações sítio-dirigidas na repetição invertida, encontrada a 3\' do promotor P2, antes do gene groES, revelaram que este elemento, conhecido como CIRCE, regula negativamente a expressão de groESL a 300C e mutações neste elemento levam à perda do controle temporal deste operon. / The heat shock genes groES and groEL of Caulobacter crescentus were isolated using the homologous genes of E.colí as a probe. DNA sequence analysis has shown that these genes are organized as an operan in a fragment of about 2.5kb, which includes the 5\' regulatory region. Northern blot analysis of total RNA from cells grown at 300C or heat shocked treated has shown the presence of a single mRNA species for groESL, of approximately 2.3kb in size, which presented increased leveis even after long periods of heat shock. Samples of total RNA from synchronized cells, corresponding to different stages of the Caulobaaer cell cycle, were also analysed, showing that the amount of groESL mRNA varies during the cycle, with maximum leveis in predivisional cells. Western blot analysis of GroEL leveis in Caulobaaer has shown that the amount of the protein decreases during the first 60 minutes of C.crescentus cell cycle and then starts to increase again. These results corroborate the data obtained with Northern blot analysis. A similar experiment was performed after exposing a mixed population of C.crescentus cells to different times of heat shock at 400C. Western blot of extracts of these cells showed a fivefold increase in the leveis of GroEL after 60 minutes of heat shock, which then begins to decrease. Primer extension experiments were performed using total RNA from cells incubated at normal growth temperature or after heat shock treatment. Two possible transcription start sites were determined at positions -119 and -88 from the ATG of the groES initiator methionine and the -10 and -35 regions of the corresponding promoters (P 1 and P2) were identified. Only trancription initiating from the P2 promoter, which has caracteristics of a &#963;32 promoter, I ncreases during heat shock .Transcription fusions with the reporter vector placZ/290 and the 5\' regulatory region of the groESL operan were contructed in order to identify the sequences responsible for heat shock and cell cycle contral. Deletion analysis in the 5\' region of the operon showed that no sequences upstream of the P2 promoter are necessary for heat shock induction or for temporal contral. Site-directed mutagenesis in the inverted repeat found 3\' of the P2 promoter, in front of the groES gene, revealed that this element, also known as CIRCE, negatively regulates groESL expression at 300C and mutations in it lead to loss of temporal control of this heat shock operon.

Made available in DSpace on 2014-10-09T12:35:32Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T14:04:01Z (GMT). No. of bitstreams: 0 / Tese (Doutoramento) / IPEN/T / Instituto de Pesquisas Energeticas e Nucleares - IPEN-CNEN/SP

Made available in DSpace on 2014-10-09T12:35:32Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T14:04:01Z (GMT). No. of bitstreams: 0 / Moluscos têm sido empregados como bioindicadores em estudos de contaminação ambiental. Nesse contexto, o caramujo de água doce Biomphalaria glabrata tem sido avaliado como um bom modelo laboratorial, e estudos prévios apontaram sua aplicação na pesquisa ambiental. A proteína HSP70 é uma molécula de 70 kDa, pertencente a uma família de proteínas com papel na manutenção da homeostase dos seres vivos: as proteínas de choque térmico (HSPs); e vem sendo estudada como potencial biomarcador de dano ambiental, indicando estresse e protegendo os organismos dos danos às proteínas. Neste trabalho, foi caracterizada a proteína HSP70 de B. glabrata pelo Western blot, com o objetivo de seu emprego em aplicações ambientais futuras. Para isso, caramujos de 5-6 meses de idade, com diâmetro de concha de 14,4 (±1,7) mm, foram expostos ao calor e ao cloreto de cádmio (CdCl2) a fim de se verificar a resposta desta proteína frente a esses estresses. Os animais foram dissecados para investigação da indução da HSP70. As proteínas foram extraídas dos tecidos com tampão RIPA, separadas em eletroforese desnaturante em gel de poliacrilamida, transferidas para uma membrana de nitrocelulose e detectadas com anticorpo específico para HSP70. A CL50/96h foi determinada como sendo 0,34 (0,30-0,37) ppm para o CdCl2 e serviu de referência para os experimentos de indução da proteína. Foi observado que a exposição a temperaturas subletais aumentou a resistência dos caramujos à temperatura letal de 42 °C. Exposições prévias ao calor de 33 °C e ao CdCl2 a 0,22 ppm aumentou a sobrevivência dos caramujos B. glabrata à concentração letal de CdCl2 (0,7 ppm) e à temperatura letal (42 °C), respectivamente. Os achados do Western blot apontaram para um possível papel da HSP70 nesse processo. Os resultados mostraram relação entre a proteína HSP70 e o aumento na sobrevivência aos estímulos letais após prévia exposição a estresses moderados. O Western blot mostrou uma indução da HSP70 nos grupos pré-expostos, se comparados aos grupos controles. A glândula digestiva foi o tecido mais responsivo, no que concerne à indução da proteína HSP70, comparando com tecidos de cabeça/pé e ovoteste. Foi encontrado o pico de indução da HSP70 nos caramujos B. glabrata após 48 horas de exposição ao calor de 33 °C, e após 96 horas de exposição ao CdCl2 a 0,22 ppm. Apesar do bem conhecido papel da HSP70 na termotolerância e tolerância a outros agentes estressores nos organismos vivos, esta foi a primeira vez que isto foi demonstrado no B. glabrata, oferecendo subsídios para a sua aplicação em estudos de monitoramento ambiental. Os resultados apresentados aqui abrem o caminho para estudos futuros dessa proteína no molusco, e fornecem mais bases para o conhecimento do B. glabrata. / Tese (Doutoramento) / IPEN/T / Instituto de Pesquisas Energeticas e Nucleares - IPEN-CNEN/SP