Estudo transgeracional dos efeitos da restrição alimentar materna sob a atividade funcional de macrófagos alveolares da prole adulta
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Data
2024-04-30
Autores
Azevedo, Gabriela Araujo de [UNIFESP]
Orientadores
Landgraf, Richardt Gama [UNIFESP]
Tipo
Tese de doutorado
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A baixa oferta de nutrientes durante a gestação pode causar adaptações morfológicas e metabólicas duradouras no indivíduo, algumas também afetar as gerações subsequentes. Embora essas adaptações sejam benéficas a curto prazo, pois acontecem com o propósito de sobrevivência, elas podem levar o indivíduo a maior propensão ao desenvolvimento de doenças na vida adulta, como obesidade, diabetes e hipertensão. Além disso, com o estresse causado pela restrição de alimentos durante a fase de embriogênese, alguns órgãos podem ser diretamente afetados, principalmente o pulmão que possui uma formação complexa, já que seu amadurecimento completo só ocorre após o nascimento. Estudos anteriores do grupo observaram, em modelo animal, que ratos Wistar que sofreram restrição alimentar intrauterinamente apresentaram redução da resposta inflamatória aguda induzida por LPS e da resposta inflamatória pulmonar alérgica. Uma das células que estão ligadas diretamente com a resposta inflamatória e por garantir a homeostase no organismo são os macrófagos. O objetivo do trabalho consiste em avaliar, na prole de ratos Wistar, a influência da programação fetal ocasionada pela restrição alimentar durante toda a prenhez, em animais da 1ª e 2ª geração, avaliando tamanho, atividade dos macrófagos e produção de citocinas na 1ª e 2ª geração. Com esse propósito, realizamos o acasalamento, acompanhamos a prenhez das ratas e separamos em dois grupos, o grupo controle que recebeu ração ad libitum e grupo restrito que recebeu apenas 50% de ração durante todo período gestacional. Após o nascimento da prole (1ª geração) realizamos o acompanhamento de peso e tamanho dos animais. Para obter a geração F2, animais que sofreram restrição intrauterina foram acasalados, pai e/ou mãe. Estes animais não tiveram nenhuma restrição alimentar ou oferta de água reduzida durante toda a gestação e foram divididos em três diferentes grupos: Grupo P-F2, onde somente o pai é proveniente das ratas que sofreram restrição alimentar na F1; Grupo M-F2, onde somente a mãe é proveniente das ratas que sofreram restrição alimentar na F1 e Grupo PM-F2 onde o pai e a mãe são provenientes das ratas que sofreram restrição alimentar na F1. Na 12ª semana de vida obtivemos as células de macrófagos alveolares, para avaliar o a capacidade fagocítica e microbicida desses macrófagos e analisar as citocinas no sobrenadante da cultura de macrófagos. Os resultados obtidos indicam que a restrição intrauterina provoca diminuição no ganho de massa corporal das ratas prenhas, no tamanho e no peso corporal das proles, e que o baixo peso ao nascer e o menor tamanho são características transgeracionais. No entanto, não verificamos diferença entre os grupos da segunda geração. Também observamos que os macrófagos alveolares do grupo restrito apresentam uma capacidade fagocítica e microbicida inferior àquela observada no grupo controle, sendo que a atividade microbicida pouco efetiva se manifesta também na geração F2 em todos os grupos. Avaliando a produção de citocinas pró inflamatórias IL-1β e IL-6, utilizamos como estímulo as leveduras Saccharomyces cerevisiae durante 4 horas e observamos que a expressão de IL-1β e IL-6 está aumentada unicamente no grupo controle + leveduras quando comparado aos demais grupos estimulados (restrito, P2-F2, M2-F2 e PM-F2). No caso da IL-10, uma citocina anti-inflamatória, a expressão está elevada apenas no grupo restrito sem levedura. Logo, a conclusão do trabalho é a de que existe uma interação entre a restrição alimentar intrauterina e a atividade dos macrófagos. Além de alterações de tamanho e massa corporal, alterações de atividade dos macrófagos são transgeracionais, independente se só pai, ou mãe ou ambos sofreram restrição alimentar intrauterina. A restrição alimentar durante a gestação pode estar causando alterações na linha germinativa, que afetam mecanismos epigenéticos do indivíduo, podendo levar a mudanças transgeracionais transmitidos por linhas maternas ou paternas. Porém uma maior compreensão das mudanças causadas na resposta imunológicas e das modificações epigenéticas nesses animais se faz necessária para confirmar se a restrição alimentar durante o período gestacional poderia aumentar a predisposição de doenças nas próximas gerações, mesmo que nestas proles não ocorra redução da oferta de nutrientes e então podemos realizar as intervenções precoces dirigidas à saúde das mulheres grávidas garantirão a saúde da população a seguir.
The low supply of nutrients during pregnancy can cause lasting morphological and metabolic adaptations in the individual, some of which can also affect subsequent generations. Although these adaptations are beneficial in the short term, as they occur for the purpose of survival, they can make the individual more prone to developing diseases in adulthood, such as obesity, diabetes and hypertension. Furthermore, with the stress caused by food restriction during the embryogenesis phase, some organs can be directly affected, especially the lung, which has a complex formation, as its complete maturation only occurs after birth. Previous studies by the group observed, in an animal model, that Wistar rats that suffered intrauterine food restriction showed a reduction in the acute inflammatory response induced by LPS and the allergic pulmonary inflammatory response. One of the cells that are directly linked to the inflammatory response and ensures homeostasis in the body are macrophages. The objective of the work is to evaluate, in the offspring of Wistar rats, the influence of fetal programming caused by food restriction throughout pregnancy, in 1st and 2nd generation animals, evaluating size, macrophage activity and cytokine production in the 1st and 2nd generation. For this purpose, we performed mating, monitored the pregnancy of the rats and separated them into two groups, the control group that received food ad libitum and the restricted group that received only 50% of food throughout the gestational period. After the birth of the offspring (1st generation), we monitor the weight and size of the animals. To obtain the F2 generation, animals that suffered intrauterine restriction were mated, father and/or mother. These animals did not have any food restrictions or reduced water supply throughout pregnancy and were divided into three different groups: Group P-F2, where only the father comes from rats that suffered food restrictions in F1; Group M-F2, where only the mother comes from rats that suffered food restriction in F1 and Group PM-F2 where the father and mother come from rats that suffered food restriction in F1. In the 12th week of life, we obtained alveolar macrophage cells to evaluate the phagocytic and microbicidal capacity of these macrophages and analyze the cytokines in the macrophage culture supernatant. The results obtained indicate that intrauterine restriction causes a decrease in body mass gain in pregnant rats, in the size and body weight of their offspring, and that low birth weight and smaller size are transgenerational characteristics. However, we found no difference between the second generation groups. We also observed that the alveolar macrophages in the restricted group have a lower phagocytic and microbicidal capacity than that observed in the control group, with ineffective microbicidal activity also manifested in the F2 generation in all groups. Evaluating the production of pro-inflammatory cytokines IL-1β and IL-6, we used Saccharomyces cerevisiae yeast as a stimulus for 4 hours and observed that the expression of IL-1β and IL-6 is increased only in the control + yeast group when compared to the others. stimulated groups (restricted, P2-F2, M2-F2 and PM-F2). In the case of IL-10, an anti-inflammatory cytokine, expression is elevated only in the restricted group without yeast. Therefore, the conclusion of the work is that there is an interaction between intrauterine dietary restriction and macrophage activity. In addition to changes in body size and mass, changes in macrophage activity are transgenerational, regardless of whether only the father, mother or both suffered intrauterine food restriction. Food restriction during pregnancy may be causing changes in the germline, which affect the individual's epigenetic mechanisms, which may lead to transgenerational changes transmitted through maternal or paternal lines. However, a greater understanding of the changes caused in the immunological response and epigenetic modifications in these animals is necessary to confirm whether dietary restriction during the gestational period could increase the predisposition to diseases in the next generations, even if these offspring do not reduce the supply of nutrients. and then we can carry out early interventions aimed at the health of pregnant women will ensure the health of the population thereafter.
The low supply of nutrients during pregnancy can cause lasting morphological and metabolic adaptations in the individual, some of which can also affect subsequent generations. Although these adaptations are beneficial in the short term, as they occur for the purpose of survival, they can make the individual more prone to developing diseases in adulthood, such as obesity, diabetes and hypertension. Furthermore, with the stress caused by food restriction during the embryogenesis phase, some organs can be directly affected, especially the lung, which has a complex formation, as its complete maturation only occurs after birth. Previous studies by the group observed, in an animal model, that Wistar rats that suffered intrauterine food restriction showed a reduction in the acute inflammatory response induced by LPS and the allergic pulmonary inflammatory response. One of the cells that are directly linked to the inflammatory response and ensures homeostasis in the body are macrophages. The objective of the work is to evaluate, in the offspring of Wistar rats, the influence of fetal programming caused by food restriction throughout pregnancy, in 1st and 2nd generation animals, evaluating size, macrophage activity and cytokine production in the 1st and 2nd generation. For this purpose, we performed mating, monitored the pregnancy of the rats and separated them into two groups, the control group that received food ad libitum and the restricted group that received only 50% of food throughout the gestational period. After the birth of the offspring (1st generation), we monitor the weight and size of the animals. To obtain the F2 generation, animals that suffered intrauterine restriction were mated, father and/or mother. These animals did not have any food restrictions or reduced water supply throughout pregnancy and were divided into three different groups: Group P-F2, where only the father comes from rats that suffered food restrictions in F1; Group M-F2, where only the mother comes from rats that suffered food restriction in F1 and Group PM-F2 where the father and mother come from rats that suffered food restriction in F1. In the 12th week of life, we obtained alveolar macrophage cells to evaluate the phagocytic and microbicidal capacity of these macrophages and analyze the cytokines in the macrophage culture supernatant. The results obtained indicate that intrauterine restriction causes a decrease in body mass gain in pregnant rats, in the size and body weight of their offspring, and that low birth weight and smaller size are transgenerational characteristics. However, we found no difference between the second generation groups. We also observed that the alveolar macrophages in the restricted group have a lower phagocytic and microbicidal capacity than that observed in the control group, with ineffective microbicidal activity also manifested in the F2 generation in all groups. Evaluating the production of pro-inflammatory cytokines IL-1β and IL-6, we used Saccharomyces cerevisiae yeast as a stimulus for 4 hours and observed that the expression of IL-1β and IL-6 is increased only in the control + yeast group when compared to the others. stimulated groups (restricted, P2-F2, M2-F2 and PM-F2). In the case of IL-10, an anti-inflammatory cytokine, expression is elevated only in the restricted group without yeast. Therefore, the conclusion of the work is that there is an interaction between intrauterine dietary restriction and macrophage activity. In addition to changes in body size and mass, changes in macrophage activity are transgenerational, regardless of whether only the father, mother or both suffered intrauterine food restriction. Food restriction during pregnancy may be causing changes in the germline, which affect the individual's epigenetic mechanisms, which may lead to transgenerational changes transmitted through maternal or paternal lines. However, a greater understanding of the changes caused in the immunological response and epigenetic modifications in these animals is necessary to confirm whether dietary restriction during the gestational period could increase the predisposition to diseases in the next generations, even if these offspring do not reduce the supply of nutrients. and then we can carry out early interventions aimed at the health of pregnant women will ensure the health of the population thereafter.
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AZEVEDO, Gabriela Araujo de. Estudo transgeracional dos efeitos da restrição alimentar materna sob a atividade funcional de macrófagos alveolares da prole adulta. 2024. 84 f. Tese (Doutorado em Medicina Translacional) - Escola Paulista de Medicina, Universidade Federal de São Paulo (UNIFESP). São Paulo, 2024.