Mestrado em Biotecnologia

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http://repositorio.ufes.br/handle/10/17567

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    O papel da L-arginina sobre o sistema oxidativo em células mda-mb-231 de câncer de mama triplo negativo
    (Universidade Federal do Espírito Santo, 2025-03-18) João Augusto Diniz Moura; Gouvea, Sonia Alves; https://orcid.org/0000-0001-5180-471X; http://lattes.cnpq.br/7268228122543743; Meira, Débora Dummer; Sartório, Carmem Luíza
    L-Arginine is a semi-essential amino acid whose role in cancer has been under investigation, showing beneficial results due to its involvement in various physiological systems. Therefore, it is necessary to investigate which pathways involving this amino acid directly contribute to tumor regression, such as those related to oxidative stress. In this context, the use of the MDA-MB-231 triple-negative breast cancer cell line, known for its high aggressiveness and limited treatment options due to the lack of therapeutic targets, highlighted the effects of different concentrations of L-ARG on cell viability and the oxidative system. The treatment involved exposure to L-ARG at concentrations of 800 µg/mL, 1600 µg/mL, and 3200 µg/mL for 48 hours. A dihydroethidium (DHE) staining assay was performed to detect the increased presence of superoxide anion (O₂⁻). The AlamarBlue assay was employed to measure cell proliferation and assess cell viability. AOPP values were determined according to the protocol described by Witko-Sarsat et al. (1996) to evaluate the production of advanced oxidation protein products. Western blotting was used to determine the influence of L-arginine treatment on SOD1 protein expression in MDA-MB-231 cells across the NT, 800 µg/mL, 1600 µg/mL, and 3200 µg/mL groups. Data were analyzed using GraphPad Prism® version 8.0.2 and corrected by Tukey’s test. The main findings of the study were: (I) Reduced cell viability; (II) Increased production of superoxide anion radicals (O₂⁻) at all concentrations; (III) Elevated levels of advanced oxidation protein products; and (IV) Reduced SOD1 expression. Therefore, through the significant results that contributed to elucidating the effects of L-arginine on the oxidative system in MDA-MB-231 cells, this study opens new perspectives with potential translational implications.
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    Identificação de potenciais biomarcadores genéticos da rede de inflamação e seus riscos de desenvolver dispneia pós-COVID-19
    (Universidade Federal do Espírito Santo, 2024-02-24) Ventorim, Vinicius do Prado; Meira, Débora Dummer; https://orcid.org/0000-0002-6092-2459; http://lattes.cnpq.br/7199119599752978; Louro, Iúri Drumond ; https://orcid.org/0000-0001-5160-9615; http://lattes.cnpq.br/3817361438227180; https://orcid.org/0009-0001-9958-5023; http://lattes.cnpq.br/2427358142499457; Paula, Flávia; https://orcid.org/0000-0001-8679-2982; http://lattes.cnpq.br/7913201450663683; Carvalho, Elizeu Fagundes; https://orcid.org/0000-0003-4620-7253; http://lattes.cnpq.br/2742420738858309
    The study aimed to identify genetic biomarkers associated with the progression of dyspnea in individuals post-COVID-19. Following approval by the Research Ethics Committee (CEP), a case-control study was conducted involving 277 individuals. Participants were selected based on clinical and epidemiological criteria, allowing a representative sample of the study population. For genetic analysis, whole-exome sequencing was performed, the identification of genetic variants potentially associated with the clinical outcome. Rigorous statistical methodologies were applied to select the most relevant variants, focusing on determining the allelic risk for the progression of dyspnea. The prevalence of dyspnea among participants was 15.16%, underscoring the importance of investigating genetic factors that may influence its manifestation. Eleven genetic variants with p-values lower than 0.05 were identified, suggesting a statistically significant association with the risk of dyspnea progression. The identified variants are located in ten genes that play key roles in the immune system, in the regulation of vascular hemostasis, and in the response to epithelial damage. These findings indicate that genetic predisposition may contribute to the persistence of dyspnea in individuals who have overcome the acute phase of COVID 19. Gene analysis revealed a strong correlation with critical biological processes in the inflammatory response and tissue recovery. The immune system plays a central role in regulating the persistent inflammation observed in some post-COVID-19 individuals, and may influence the maintenance of chronic cardiorespiratory symptoms. Vascular hemostasis, on the other hand, may be associated with coagulation disorders previously described in patients with COVID-19, impacting pulmonary perfusion and contributing to dyspnea. Genes related to epithelial damage are possibly involved in lung tissue repair, being decisive for the functional recovery of the organ after infection. These results provide valuable directions for future clinical and epidemiological research in the State of Espírito Santo (Brazil). The identification of genetic biomarkers can help in the risk stratification of patients, allowing for more personalized and targeted therapeutic approaches. Moreover, the findings reinforce the need for further investigations to understand the mechanisms by which these genetic variants influence the evolution of post-COVID-19 dyspnea. Furthermore, this study contributes to the growing body of knowledge regarding sequelae of COVID-19 and its genetic bases, highlighting the importance of the integration between genomics and epidemiology for the development of public health strategies. Based on these findings, it is expected that further research can validate the identified biomarkers and deepen the understanding of the genetic factors involved in the recovery of post COVID-19 patients. This approach may, ultimately, lead to the development of more effective interventions to mitigate the impacts of persistent dyspnea on the quality of life of affected individuals
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    Análise de variantes nos genes ACE1 e LRP1 e sua associação com a doença de Alzheimer na Grande Vitória/ES : potenciais biomarcadores genéticos para diagnóstico complementar
    (Universidade Federal do Espírito Santo, 2025-02-26) Lopes, Victor Alves; Paula, Flávia de ; https://orcid.org/0000-0001-8679-2982; http://lattes.cnpq.br/7913201450663683; https://orcid.org/0009-0006-1332-6079; http://lattes.cnpq.br/1133525392804189; Errera, Flavia Imbroisi Valle; https://orcid.org/0000-0002-8069-6372; http://lattes.cnpq.br/9337327437538048; Maranduba, Carlos Magno da Costa ; https://orcid.org/0000-0001-7327-1934; http://lattes.cnpq.br/4763153859701731
    The accelerated aging of the Brazilian population has contributed to the increasing prevalence of chronic diseases such as Alzheimer’s disease (AD), a multifactorial condition involving complex interactions between genetic and environmental factors. This study aimed to analyze genetic variants in the ACE1 (INDEL rs4646994) and LRP1 genes and their association with AD in a population from Greater Vitória, Espírito Santo, to identify potential genetic biomarkers for complementary diagnosis. A cross-sectional, analytical, and observational study was conducted with 246 participants clinically diagnosed with sporadic AD, matched by sex, age, and ethnicity. Blood samples were collected for genomic DNA extraction, followed by PCR and agarose gel electrophoresis for genotyping. The results showed no significant association between the INDEL variant of the ACE1 gene and AD or the APOE ε4 allele status, suggesting a limited contribution of this variant in the studied population. Conversely, the LRP1 gene showed a significant association, with the D allele (DD+ID) displaying a potentially protective role against AD. These findings reinforce the multifactorial nature of AD and highlight the influence of regional genetic characteristics, considering the historical diversity and genetic admixture of the Espírito Santo population. This study contributes to understanding genetic differences between populations and supports the development of personalized medicine strategies for the diagnosis and treatment of AD
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    Bioprospecção do fucoidan e ouabaína: efeitos na motilidade e vitalidade espermática humana in vitro
    (Universidade Federal do Espírito Santo, 2025-03-18) Silva, Paula Gabriella Pedras; Nogueira, Breno Valentim; Meira, Débora Dummer; https://orcid.org/0000-0002-6092-2459; Paula, Flavia de; Maranduba, Carlos Magno da Costa
    Oral hormonal contraceptives (OC) are widely used in family planning but pose significant health risks to women. Although effective, they can cause severe adverse effects, such as cardiovascular alterations that increase the risk of pulmonary embolism, venous and arterial thrombosis, and metabolic diseases such as diabetes and dyslipidemia. Cardiovascular diseases (CVD) are the leading cause of death in Brazil and globally, and hormonal contraceptive methods are associated with an increased risk of cerebrovascular accidents and other CVDs. Plant-based spermicides are alternatives to chemical spermicides like Nonoxynol-9 (N-9), which can cause vaginal mucosa damage and increase the risk of infections. Alternatives to N-9, such as Sodium Lauryl Sulfate (SLS), also present adverse effects, including skin and mucosal irritation. Fucoidan, a sulfated polysaccharide found in brown seaweeds, has beneficial biological activities and is investigated as a drug carrier due to its low toxicity. Ouabain, extracted from the plant Strophanthus gratus, acts on sperm capacitation and inhibits Na+/K±ATPase, playing a significant role in various physiological and pathological processes. This study aimed to analyze the spermicidal activity of fucoidan, extracted from the seaweed Fucus vesiculosus, and ouabain on human sperm in vitro. The methodology involved collecting semen samples from volunteers aged 18 to 45 years, after approval from the Ethics and Research Committee under the Ethical Appreciation Presentation Certificate (CAAE) 79783124.5.0000.5060, which were subjected to treatments with negative control, positive control (SLS), fucoidan, ouabain, and a combination of both. Sperm motility and vitality were assessed at two different time points (0 and 15 minutes) using seminal analysis techniques standardized by the World Health Organization. The treatments significantly reduced sperm motility and vitality compared to the positive control (SLS). The combination of fucoidan and ouabain showed the greatest reduction in sperm motility, with approximately a 28.3% decrease in progressive motility and a 28.1% decrease in vitality, suggesting a synergistic effect between the compounds. Statistical analysis using non-parametric tests of Kruskal-Wallis and Wilcoxon post hoc confirmed the significance of these reductions at times T0 and T15. Moreover, the interaction of the compounds with reactive oxygen species (ROS) suggests a mechanism of action that compromises cellular integrity and sperm function. These findings highlight the potential of fucoidan and ouabain as effective spermicidal agents, contributing to the development of safer and more efficient contraceptive methods. However, it is necessary to consider the costs associated with each treatment and conduct further research to confirm these hypotheses and explore their future biotechnological applications
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    Classificação de gravidade e identificação de biomarcadores na Covid-19: análise do exoma de pacientes através de máquinas de vetores de suporte com kernel linear (SVM)
    (Universidade Federal do Espírito Santo, 2025-02-24) Zetum, Aléxia Stefani Siqueira; Meira, Débora Dummer; https://orcid.org/0000-0002-6092-2459; http://lattes.cnpq.br/7199119599752978; Louro, Iuri Drumond; https://orcid.org/0000-0001-5160-9615; http://lattes.cnpq.br/3817361438227180; https://orcid.org/0000-0002-5086-411X; Paula, Flávia de; https://orcid.org/0000-0001-8679-2982; Carvalho, Elizeu Fagundes de; https://orcid.org/0000-0003-4620-7253
    Introduction: SARS-CoV-2 infection presents a wide spectrum of clinical manifestations. Genetic variations may influence the host's response to the virus. The use of Machine Learning (ML) has shown promise in identifying genetic biomarkers and individuals who may develop severe forms of the disease. Objective: To develop an ML model using exome data to predict clinical outcomes in COVID-19 patients and identify genes potentially associated with disease severity. Methodology: The study involved data from 239 COVID-19 patients ("Non-severe" and "Severe"). DNA sequencing was performed, and ancestry analysis was conducted. A Support Vector Machine (SVM) model with a linear kernel was developed to predict COVID-19 severity, utilizing Recursive Feature Elimination (RFE) to select the most influential variants. Metrics such as Area Under the Curve-Receiver Operating Characteristic (AUC-ROC), accuracy, F1 score, sensitivity, and specificity were used. Subsequently, logistic regression (LR) analysis was performed with the variants selected by SVM-RFE and confounding variables. Results and Discussion: The SVM model with a linear kernel achieved an AUC-ROC of 0,81, accuracy of 83%, and an F1 score of 0,78, indicating a good capacity to discriminate between "Severe" and "Non-severe" cases of COVID-19. Fifteen variants were selected by the model, of which seven were significantly associated with disease severity in the LR analysis. Risk variants include WSCD1 (rs2302837 "A/A" or "A/G," 95% CI: 1,32–7,24, OR: 3,09, P < 0,01), PTPRS (rs1143700 "A/A" or "A/G," 95% CI: 1,54–7,07, OR: 3,30, P < 0,01), ARVCF (rs2073744 "A/A" or "A/G," 95% CI: 1,31–6,30, OR: 2,88, P < 0,01), and LVRN (rs10078759 "G/G" or "G/C," 95% CI: 1,07–4,31, OR: 2,08, P = 0,04). Conversely, protective variants include ALDH4A1 (rs6426813 "G/G" or "G/A," 95% CI: 0,23–0,93, OR: 0,48, P = 0,02), ARHGAP22 (rs10776601 "C/C" or "C/T," 95% CI: 0,09–0,56, OR: 0,23, P < 0,01), and C3 (rs423490 "A/A" or "A/G," 95% CI: 0,14–0,70, OR: 0,32, P < 0,01). The results demonstrated that the SVM with a linear kernel is effective in predicting COVID-19 severity using exome data. The protein-protein interaction (PPI) network analysis identified biological pathways associated with the immune system, inflammatory response, and blood coagulation. Genes such as C3, PTPRS, and LVRN stood out in functions related to immune response regulation and inflammation modulation, suggesting these pathways are directly linked to adverse COVID-19 outcomes. The network also revealed the interconnection between cellular signaling processes and stress response mechanisms, which may explain the variability in clinical responses observed among patients. Conclusion: The SVM with a linear kernel using our data proved effective in predicting COVID-19 severity. This study highlights the importance of integrative approaches to better understanding the disease. Identifying genetic biomarkers can aid in treatment and management of future pandemics