Avaliação do tratamento preventivo com o citral sobre o eixo tecido adiposo-hipotálamo em camundongos obesos induzidos por dieta hiperlipídica
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Data
2023-07-31
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Universidade Estadual Paulista (Unesp)
Resumo
Em 2022, a obesidade já atingiu mais de 41 milhões de brasileiros. O alto consumo de nutrientes e o baixo gasto energético provocam injúria do tecido adiposo (TA), desencadeando um processo de inflamação crônica de baixo grau com o predomínio na liberação de adipocinas pró-inflamatórias e diminuição de mediadores anti-inflamatórios. Ocorre também um aumento de espécies reativas de oxigênio (EROs) devido ao aumento de infiltração de células do sistema imunológico e amplificação da inflamação na região do hipotálamo. Diante do desbalanço sistêmico, ocorre um desequilíbrio da homeostase energética. Contudo, são poucos os estudos que investigam as vias de sinalização envolvidos na interação do eixo TA-hipotálamo como potenciais alvos terapêuticos para o tratamento da obesidade. O citral é um monoterpeno, de origem vegetal, que possui atividade antioxidante e anti-inflamatória já comprovada na literatura. Assim, nosso objetivo foi avaliar a ação preventiva do citral na inflamação ocasionada pela obesidade em camundongos C57BL/6J, investigando os mecanismos anti-inflamatórios e antioxidantes do eixo TA-hipotálamo. Foram utilizados camundongos machos C57BL/6J com 6 semanas de idade (n=10), e a indução de obesidade com dieta hiperlipídica (HFD; 60% calorías provenientes de lipídios) durante 17 semanas (CEUA nº 3547180722). Os animais foram separados em cinco grupos: alimentados com dieta padrão (DP) e os que receberam HFD mais o veículo e os tratamentos com o citral por via oral (25, 100 ou 300 mg/kg). Ao final do período de indução, foram realizados os testes de tolerância à glicose e insulina (TOTG e TTI). Os TA epididimal e hipotalâmico foram coletados para quantificação de mediadores inflamatórios interleucina (IL)-4 e IL-10 pelo ensaio Luminex. Para a análise de expressão gênica dos mediadores inflamatórios IL-6 e IL-10, assim como o receptor de adiponectina no hipotálamo pela técnica de RT-qPCR. Foi realizada a determinação de mecanismos antioxidantes no tecido hipotalâmico, através quantificação da atividade da enzima superóxido dismutase (SOD) e quantificação da enzima catalase (CAT). Por fim, foi executado a quantificação de leptina e adiponectina através do imunoensaio ELISA sanduíche, no TA e soro. Foi efetuado o teste de normalidade e em seguida, os dados foram submetidos ao teste t de Student não pareado para comparação entre dois grupos e análise de variância (ANOVA) de uma via ou duas vias para comparação entre três grupos ou mais, seguido do teste de Tukey (p < 0,05). Observamos o aumento significativo da massa corporal, índice de adiposidade e hiperglicemia dos animais que receberam HFD em comparação ao grupo DP. O tratamento dos animais HFD com citral 300 mg/kg foi capaz de promover a redução da massa corporal dos animais em relação ao grupo HFD sem tratamento. O grupo de animais tratados com citral 300 mg/kg apresentou redução da glicemia e diminuição da ingestão calórica. Tais resultados podem ser explicados pelo aumento da IL-4 no tecido hipotalâmico, aumento da IL-10 e IL-6 no TA com os animais HFD previamente tratados com citral na dose de 300 mg/kg em relação aos animais HFD que receberam somente o veículo. Também, o citral foi capaz de promover a redução dos níveis séricos de leptina e aumento de adiponectina. Em síntese, a administração de citral na dose de 300 mg/kg promoveu uma ação anti-inflamatória na resolução de alterações metabólicas decorrentes da obesidade. Diante disso, os novos desafios serão caracterizar mecanismos neurais da atuação do citral sobre neurônios hipotalâmicos.
In 2022, obesity affected more than 41 million Brazilians. The high consumption of nutrients and low energy expenditure causes injury to the adipose tissue (AT), triggering a process of chronic low-grade inflammation with a predominance of pro- inflammatory adipokines and a decrease in anti-inflammatory mediators, fundamental for the control of an inflammatory response. There is also an increase in reactive oxygen species (ROS) due to increased infiltration of immune system cells and amplification of inflammation in the hypothalamus region. Together with systemic imbalance, there is an imbalance in energy homeostasis. However, there have been few studies that investigate the signaling pathways involved in the interaction of the TA-hypothalamus axis as potential therapeutic targets for the treatment of obesity. Citral is a monoterpene, of plant origin, which has antioxidant and anti-inflammatory activity already proved in the literature. Thus, the aim of this study was to evaluate the preventive action of citral in low-grade chronic inflammation caused by obesity, investigating the anti-inflammatory and antioxidant mechanisms of the TA-hypothalamus axis, in C57BL/6J mice. Male C57BL/6J mice (n=10) were used, and obesity was induced with high-fat diet (HFD; 60% calories from lipids) for 17 weeks (CEUA nº 3547180722). The animals were divided into five groups: those fed with a standard diet (SD) and those that received HFD plus oral citral treatment (25, 100 or 300 mg/kg). At the end of the induction period, glucose and insulin tolerance tests (TOTG and TTI, respectively) were performed. Epididymal AT and hypothalamic tissue were collected for quantification of interleukin (IL)-4 and IL-10 inflammatory mediators by Luminex. The gene expression of the inflammatory mediators IL-6 and IL-10, as well as the adiponectin receptor in the hypothalamic tissue, were performed by the RT-qPCR technique. The determination of antioxidant mechanisms was performed, through activity quantification of the enzyme superoxide dismutase (SOD) and quantification of the enzyme catalase (CAT). Lastly, the quantification of adipokines, leptin and adiponectin were performed through the sandwich ELISA immunoassay. A normality test was performed to analyze whether the data are parametric or non-parametric. Data were subjected to unpaired Student's t-test for comparison between two groups and one-way or two-way analysis of variance (ANOVA) for comparison between three groups or more, followed by Tukey's test, p < 0,05. We observed an increase in body weight, adiposity index and hyperglycemia of the animals that received HFD compared to the SD-fed animals. Citral 300 mg/kg was able to promote the reduction of the animal's body weight in relation to the HFD group. Citral 300 mg/kg-treated mice showed a reduction in glycemia compared to eutrophic mice and a decrease in caloric intake. This can be explained by the IL-4 increase in the hypothalamic tissue and the IL-10 increase in AT with the citral preventive treatment at a dose of 300 mg/kg. In addition, this dose promotes a leptin levels reduction and an adiponectin increase in serum. In conclusion, the administration of citral at a dose of 300 mg/kg possibly promoted an anti-inflammatory action in the resolution of metabolic alterations such as: insulin resistance, caloric intake and body mass gain. Therefore, we will characterize the neural control of citral action on anorexigenic and orexigenic neurons through immunohistochemical techniques.
In 2022, obesity affected more than 41 million Brazilians. The high consumption of nutrients and low energy expenditure causes injury to the adipose tissue (AT), triggering a process of chronic low-grade inflammation with a predominance of pro- inflammatory adipokines and a decrease in anti-inflammatory mediators, fundamental for the control of an inflammatory response. There is also an increase in reactive oxygen species (ROS) due to increased infiltration of immune system cells and amplification of inflammation in the hypothalamus region. Together with systemic imbalance, there is an imbalance in energy homeostasis. However, there have been few studies that investigate the signaling pathways involved in the interaction of the TA-hypothalamus axis as potential therapeutic targets for the treatment of obesity. Citral is a monoterpene, of plant origin, which has antioxidant and anti-inflammatory activity already proved in the literature. Thus, the aim of this study was to evaluate the preventive action of citral in low-grade chronic inflammation caused by obesity, investigating the anti-inflammatory and antioxidant mechanisms of the TA-hypothalamus axis, in C57BL/6J mice. Male C57BL/6J mice (n=10) were used, and obesity was induced with high-fat diet (HFD; 60% calories from lipids) for 17 weeks (CEUA nº 3547180722). The animals were divided into five groups: those fed with a standard diet (SD) and those that received HFD plus oral citral treatment (25, 100 or 300 mg/kg). At the end of the induction period, glucose and insulin tolerance tests (TOTG and TTI, respectively) were performed. Epididymal AT and hypothalamic tissue were collected for quantification of interleukin (IL)-4 and IL-10 inflammatory mediators by Luminex. The gene expression of the inflammatory mediators IL-6 and IL-10, as well as the adiponectin receptor in the hypothalamic tissue, were performed by the RT-qPCR technique. The determination of antioxidant mechanisms was performed, through activity quantification of the enzyme superoxide dismutase (SOD) and quantification of the enzyme catalase (CAT). Lastly, the quantification of adipokines, leptin and adiponectin were performed through the sandwich ELISA immunoassay. A normality test was performed to analyze whether the data are parametric or non-parametric. Data were subjected to unpaired Student's t-test for comparison between two groups and one-way or two-way analysis of variance (ANOVA) for comparison between three groups or more, followed by Tukey's test, p < 0,05. We observed an increase in body weight, adiposity index and hyperglycemia of the animals that received HFD compared to the SD-fed animals. Citral 300 mg/kg was able to promote the reduction of the animal's body weight in relation to the HFD group. Citral 300 mg/kg-treated mice showed a reduction in glycemia compared to eutrophic mice and a decrease in caloric intake. This can be explained by the IL-4 increase in the hypothalamic tissue and the IL-10 increase in AT with the citral preventive treatment at a dose of 300 mg/kg. In addition, this dose promotes a leptin levels reduction and an adiponectin increase in serum. In conclusion, the administration of citral at a dose of 300 mg/kg possibly promoted an anti-inflammatory action in the resolution of metabolic alterations such as: insulin resistance, caloric intake and body mass gain. Therefore, we will characterize the neural control of citral action on anorexigenic and orexigenic neurons through immunohistochemical techniques.
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Palavras-chave
Tecido adiposo, Tecido hipotalâmico, Inflamação hipotalâmica, Obesidade, Citral, Atividade anti-inflamatória, Adiposetissue, Hypothalamictissue, Hypothalamicinflammation, Obesity, Anti-inflammatoryactivity