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Lactylation in HEPG2 toxicity response
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RESUMO: A regeneração de tecidos permite aos organismos restaurar estruturas danificadas através de vias de sinalização complexas e coordenadas que promovem a proliferação e diferenciação celular. No entanto, esta capacidade regenerativa varia entre espécies. Enquanto alguns animais conseguem regenerar totalmente os tecidos, os mamíferos — incluindo os humanos — dependem principalmente da cicatrização, um processo que frequentemente falha em restaurar plenamente a função original do tecido, podendo resultar em consequências clínicas prolongadas. Para ultrapassar estas limitações, é fundamental compreender os mecanismos que regulam as respostas celulares ao dano e à reparação. O metabolismo celular desempenha um papel central na adaptação ao stress e na regeneração, sendo comum as células alterarem o seu perfil metabólico para a glicólise aeróbica, favorecendo a sobrevivência e o restabelecimento do tecido. Esta reprogramação metabólica leva ao aumento da produção de lactato, que atua como molécula sinalizadora e reguladora epigenética. Estudos prévios do nosso laboratório em peixe-zebra demonstram que a atividade glicolítica aumentada durante a regeneração da barbatana caudal promove a formação do blastema e a regeneração eficaz do osso. Adicionalmente, a intensificação da glicólise foi associada a níveis elevados de lactilação de histonas em osteoblastos, com regiões de cromatina lactiladas enriquecidas junto a genes regulados por Oct4. Estes resultados sugerem que a remodelação da cromatina mediada por lactato pode modular programas de transcrição relevantes para a reparação de tecidos. No entanto, esta relação permanece inexplorada em modelos mamíferos. Neste estudo, investigámos se o dano celular induzido por toxicidade em células hepáticas humanas promove a acumulação de lactato e a lactilação de histonas, facilitando potencialmente a remodelação da cromatina nos locais de ligação de Oct4 e ativando uma resposta transcricional. Para testar a dependência da lactilação face ao metabolismo glicolítico, manipulámos a produção de lactato utilizando 2-desoxi-D-glucose, galoflavina, oxamato de sódio e lactato exógeno. Em seguida, avaliámos a acumulação de lactato extracelular e os níveis de lactilação após exposição a três insultos tóxicos distintos: acetaminofeno, peróxido de hidrogénio e cloreto de cobalto. Estes compostos foram selecionados por recriarem danos celulares fisiologicamente relevantes que caracterizam ambientes danificados. Por fim, explorámos se estas condições afetavam a expressão de OCT4 em função do estado de lactilação. Os nossos resultados demonstram que a lactilação é regulada pela atividade glicolítica. A acumulação de lactato e a lactilação induzidas por toxicidade dependeram do tipo de insulto. Apesar da clara modulação da lactilação, a expressão de OCT4 manteve-se inalterada, sugerindo que a lactilação não regula diretamente a transcrição deste gene. Ao esclarecer a interligação entre metabolismo, modificação das histonas e regulação transcricional durante o dano celular, este estudo fornece novos contributos para a compreensão da biologia regenerativa humana. O conhecimento destes mecanismos poderá apoiar o desenvolvimento de estratégias terapêuticas baseadas na modulação metabólica para promover a reparação de tecidos, especialmente em órgãos com capacidade regenerativa limitada.
ABSTRACT: Tissue regeneration enables organisms to restore damaged structures through coordinated and complex signalling pathways that drive cell proliferation and differentiation. However, this regenerative capacity varies across species. While some animals can fully regenerate tissues, mammals — including humans — primarily rely on wound healing, a process that often fails to restore full functionality and may lead to long-term health consequences. To overcome these limitations, it is essential to understand the mechanisms driving cellular responses to damage and repair. Cellular metabolism plays a pivotal role in stress adaptation and tissue regeneration, as cells shift toward aerobic glycolysis to promote survival and support tissue restoration. This metabolic reprogramming process increases lactate production, which can act as a signalling molecule and epigenetic regulator. Prior work from our lab in zebrafish demonstrated that enhanced glycolytic activity during caudal fin regeneration supports blastema formation and effective bone regeneration. Moreover, elevated glycolysis was associated with increased histone lactylation in osteoblasts, with lactylated chromatin regions enriched near genes regulated by Oct4. These findings suggest that lactate-driven chromatin remodelling may modulate transcriptional programs relevant to tissue repair. However, this relationship remains unexplored in mammalian systems. In this study, we investigated whether toxicity-induced cellular damage in human hepatic cells promotes lactate accumulation and histone lactylation, potentially facilitating chromatin remodelling at Oct4 binding sites and activating a transcriptional response. To test the dependence of lactylation on glycolytic metabolism, we manipulated lactate production using 2-deoxy-D-glucose, galloflavin, sodium oxamate, and exogenous lactate. We then assessed extracellular lactate production and lactylation following exposure to three distinct toxic insults: acetaminophen, hydrogen peroxide, and cobalt chloride. These compounds were selected to model physiologically relevant damaging conditions that characterize early injured environments. Finally, we explored whether these conditions influenced OCT4 expression in relation to lactylation status. Our results demonstrate that lactylation is tightly regulated by glycolytic activity. Toxicity-induced lactate accumulation and lactylation were dependent on the type of insult, highlighting insult-specific metabolic responses. Despite clear modulation of lactylation, OCT4 expression remained unchanged, suggesting that lactylation does not directly regulate OCT4 transcription but may instead enhance its chromatin accessibility and influence downstream transcriptional responses. By elucidating the interplay between metabolism, histone modifications, and transcriptional regulation during cellular damage, this study offers new insights into human regenerative biology. Understanding these processes may support the development of metabolic-based therapeutic strategies to enhance tissue repair, particularly in organs with limited regenerative capacity.
ABSTRACT: Tissue regeneration enables organisms to restore damaged structures through coordinated and complex signalling pathways that drive cell proliferation and differentiation. However, this regenerative capacity varies across species. While some animals can fully regenerate tissues, mammals — including humans — primarily rely on wound healing, a process that often fails to restore full functionality and may lead to long-term health consequences. To overcome these limitations, it is essential to understand the mechanisms driving cellular responses to damage and repair. Cellular metabolism plays a pivotal role in stress adaptation and tissue regeneration, as cells shift toward aerobic glycolysis to promote survival and support tissue restoration. This metabolic reprogramming process increases lactate production, which can act as a signalling molecule and epigenetic regulator. Prior work from our lab in zebrafish demonstrated that enhanced glycolytic activity during caudal fin regeneration supports blastema formation and effective bone regeneration. Moreover, elevated glycolysis was associated with increased histone lactylation in osteoblasts, with lactylated chromatin regions enriched near genes regulated by Oct4. These findings suggest that lactate-driven chromatin remodelling may modulate transcriptional programs relevant to tissue repair. However, this relationship remains unexplored in mammalian systems. In this study, we investigated whether toxicity-induced cellular damage in human hepatic cells promotes lactate accumulation and histone lactylation, potentially facilitating chromatin remodelling at Oct4 binding sites and activating a transcriptional response. To test the dependence of lactylation on glycolytic metabolism, we manipulated lactate production using 2-deoxy-D-glucose, galloflavin, sodium oxamate, and exogenous lactate. We then assessed extracellular lactate production and lactylation following exposure to three distinct toxic insults: acetaminophen, hydrogen peroxide, and cobalt chloride. These compounds were selected to model physiologically relevant damaging conditions that characterize early injured environments. Finally, we explored whether these conditions influenced OCT4 expression in relation to lactylation status. Our results demonstrate that lactylation is tightly regulated by glycolytic activity. Toxicity-induced lactate accumulation and lactylation were dependent on the type of insult, highlighting insult-specific metabolic responses. Despite clear modulation of lactylation, OCT4 expression remained unchanged, suggesting that lactylation does not directly regulate OCT4 transcription but may instead enhance its chromatin accessibility and influence downstream transcriptional responses. By elucidating the interplay between metabolism, histone modifications, and transcriptional regulation during cellular damage, this study offers new insights into human regenerative biology. Understanding these processes may support the development of metabolic-based therapeutic strategies to enhance tissue repair, particularly in organs with limited regenerative capacity.
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Palavras-chave
Lactylation Lactate Glycolysis Oct4 Metabolic reprogramming Cellular damage response Regeneration