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Advisor(s)
Abstract(s)
Introdução: As β-hemoglobinopatias são uma das doenças monogénicas mais comuns em todo o mundo. Atualmente, um dos tratamentos disponíveis passa pela indução farmacológica da hemoglobina fetal (HbF) através de compostos, tais como, a hidroxiureia (HU), contudo o seu perfil de segurança e o elevado custo em países subdesenvolvidos limitam a sua utilização. Nesse contexto, é essencial estudar novos compostos indutores de HbF que tenham efeitos adversos escassos e que possam estar amplamente disponíveis, como é o caso dos compostos naturais. Objetivo: Avaliar o efeito do extrato metanólico das folhas de Carica papaya (EMFCP) e da Quercetina, em células linfoblásticas em suspensão (células K562), na expressão dos genes HBG1 e HBG2. Métodos: Células K562 foram expostas durante 24 horas ao EMFCP (0,5; 50 e 100 μg/mL), à Quercetina (0,2 e 20 μM) e à HU (25 μg/mL). Após a exposição ao EMFCP e à Quercetina, os efeitos na expressão génica foram quantificados a partir do RNA total através do PCR quantitativo em Tempo Real. Os genes estudados foram HBA, HBB, HBG1 e HBG2, genes reguladores da γ-globina (MYB, KLF1, BCL11A e BGLT3), assim como moduladores epigenéticos desta globina (DNMT1, DNMT3a, DNMT3b e HDAC classe I), tendo como gene de referência o GAPDH. Resultados: Enquanto a proliferação celular é afetada pela Quercetina a 20 μM, a viabilidade não é afetada por nenhum dos compostos nas concentrações avaliadas. A análise transcricional demonstrou que o EMFCP e a Quercetina têm efeitos opostos nos padrões de expressão dos genes HBB, MYB e KLF1, o que sugere diferentes mecanismos de ação. Quanto à análise do potencial epigenético, o EMFCP parece aumentar a transcrição da maioria dos moduladores epigenéticos, enquanto a Quercetina parece ter um efeito inibitório nos genes HDAC2 e HDAC3. Conclusão: No geral, este estudo preliminar sugere que o EMFCP e a Quercetina podem modular a expressão da γ-globina, dos genes reguladores e dos moduladores epigenéticos da mesma, constituindo assim uma abordagem potencialmente eficaz para o tratamento de β-hemoglobinopatias.
ABSTRACT - Background: β-Hemoglobinopathies are one of the most common human monogenic disorders in the world. Fetal hemoglobin (HbF) pharmacological induction, through compounds such as hydroxyurea (HU), is one of the available treatments, however, their safety concerns and the expensive cost in low- and middle-Income countries limit their use. In this context, it is essential to study novel HbF-inducing compounds that have scarcer adverse effects and can be widely available, such as natural compounds. Objective: Here, we evaluate the effect of Carica papaya methanolic leaf extract (CPMLE) and Quercetin, in suspension lymphoblastic cells (K562 cells), in the expression of HBG1 and HBG2 genes. Methods: Cells were exposed for 24 hours to CPMLE (0,5; 50 and 100 μg/mL), Quercetin (0,2 and 20 μM) and HU (25 μg/mL). Gene expression differential effects after CPMLE and Quercetin exposure were quantified using total RNA through quantitative Real-Time PCR. The studied genes were HBA, HBB, HBG1, and HBG2, γ-globin expression regulators (MYB, KLF1, BCL11A, and BGLT3), as well as epigenetic modulators of this globin (DNMT1, DNMT3a, DNMT3b and HDAC class I), and GAPDH was used as a reference gene. Results: While cell proliferation is affected by 20 μM of Quercetin, viability is not affected by any of the compounds at the assessed concentrations. The transcriptional analysis demonstrated that CPMLE and Quercetin have divergent effects on HBB, MYB, and KLF1 expression levels, which suggests different mechanisms of action. As for the epigenetic potential analysis, CPMLE appears to increase the transcription of most epigenetic modulators, while Quercetin appears to have an inhibitory effect on genes HDAC2 and HDAC3. Conclusion: Overall, this preliminary study suggests that CPMLE and Quercetin can modulate the expression of γ-globin, regulator genes, and epigenetic modulators of this globin, thus potentially constituting an effective approach for the treatment of β-hemoglobinopathies.
ABSTRACT - Background: β-Hemoglobinopathies are one of the most common human monogenic disorders in the world. Fetal hemoglobin (HbF) pharmacological induction, through compounds such as hydroxyurea (HU), is one of the available treatments, however, their safety concerns and the expensive cost in low- and middle-Income countries limit their use. In this context, it is essential to study novel HbF-inducing compounds that have scarcer adverse effects and can be widely available, such as natural compounds. Objective: Here, we evaluate the effect of Carica papaya methanolic leaf extract (CPMLE) and Quercetin, in suspension lymphoblastic cells (K562 cells), in the expression of HBG1 and HBG2 genes. Methods: Cells were exposed for 24 hours to CPMLE (0,5; 50 and 100 μg/mL), Quercetin (0,2 and 20 μM) and HU (25 μg/mL). Gene expression differential effects after CPMLE and Quercetin exposure were quantified using total RNA through quantitative Real-Time PCR. The studied genes were HBA, HBB, HBG1, and HBG2, γ-globin expression regulators (MYB, KLF1, BCL11A, and BGLT3), as well as epigenetic modulators of this globin (DNMT1, DNMT3a, DNMT3b and HDAC class I), and GAPDH was used as a reference gene. Results: While cell proliferation is affected by 20 μM of Quercetin, viability is not affected by any of the compounds at the assessed concentrations. The transcriptional analysis demonstrated that CPMLE and Quercetin have divergent effects on HBB, MYB, and KLF1 expression levels, which suggests different mechanisms of action. As for the epigenetic potential analysis, CPMLE appears to increase the transcription of most epigenetic modulators, while Quercetin appears to have an inhibitory effect on genes HDAC2 and HDAC3. Conclusion: Overall, this preliminary study suggests that CPMLE and Quercetin can modulate the expression of γ-globin, regulator genes, and epigenetic modulators of this globin, thus potentially constituting an effective approach for the treatment of β-hemoglobinopathies.
Description
Mestrado em Tecnologias Moleculares em Saúde
Keywords
β-hemoglobinopatias Carica papaya Reativação da hemoglobina fetal Expressão génica Quercetina β-hemoglobinopathies Fetal hemoglobin reactivation Gene expression Quercetin
Citation
Mendes MI. Efeitos de extratos de Carica papaya nos padrões de expressão de genes reguladores da hemoglobina fetal [dissertation]. Lisboa: Escola Superior de Tecnologia da Saúde de Lisboa/Instituto Politécnico de Lisboa; 2022.
Publisher
Instituto Politécnico de Lisboa, Escola Superior de Tecnologia da Saúde de Lisboa