Melatonin Prevents Immune Stimulation of Pituitary ACTH production in Rats Fed Sucrose Rich Diets

CALANNI, JS; SANCHEZ PUCH, S; MERCAU, MARÍA E.; REPETTO, EM; ARANDA, M; CYMERYNG, CORA B.

Chicago

Congreso; ENDO 2018; 2018

Endocrine Society

Excessiveconsumption of simple carbohydrates has been associated with an increasedincidence of insulin resistance and obesity. Several reports have shown aconcomitant dysfunction of the hypothalamus­ pituitary­ adrenal (HPA) axis inthese patients. We have previously reported that rats fed a sucrose­ rich diet(SRD, 30% sucrose in the drinking water) for 7 weeks show a significantdecrease in insulin sensitivity. Analysis of HPA axis activity 3 weeks afterthe initiation of the dietary modification, when changes in insulin sensitivitywere still not evident, showed augmented levels of circulating ACTH and ahigher pituitary expression of POMC that was accompanied by an increase inoxidative stress parameters. As a chronic low-grade inflammatory state has beenassociated with the development of insulin resistance and type 2 diabetes andseveral inflammatory cytokines have been shown to activate the HPA axis, we designedpresent experiments to analyze the involvement of inflammation on the observedpituitary dysfunction. Animals were randomly distributed in 4 groups: 1)Control, 2) SRD, 3) Mel and 4) SRD + Mel. In groups 3 and 4, subcutaneousmelatonin implants were surgically implanted and changed every ten days.Animals were sacrificed after 3 weeks and pituitary glands were dissected andprocessed to obtain protein and RNA. Plasma ACTH levels were assessed by achemiluminiscent assay and corticosteronemia was determined by RIA. Expressionof specific proteins and mRNAs were evaluated by western blot and RT-qPCRrespectively. AtT20 (murine corticotrophs) and J774 (murine macrophages) cellswere used in in vitro experiments.Compared to the corresponding controls our results showed an increase in themRNA levels of TNFα (p<0.001), IL-1 beta (p<0.01) andinflammasome components (ASC, NALP3, p<0.01) in pituitary tissues obtainedfrom the SRD group. This increase in markers of inflammation was prevented bymelatonin treatment. An augmented expression of POMC, Iba-1 and F4-80,determined by immunohistochemistry and immunoblot in the SRD-group, was alsoattenuated by melatonin treatment.Incubation of the corticotroph AtT20 cell line with conditioned media obtainedfrom control (CMC) or activated J774 cells (with 100ng/ml LPS and 7.5 mMglucose for 6 h and further incubated for 24 h with fresh medium, CMLG) showedan increase in the reporter activity of POMC-LUC by CMLG that was alsoprevented by the addition of 50 nM melatonin to macrophage J774 cells(p<0.001).Insummary, our results suggest that the hyperactivation of the synthesis andsecretion of ACTH detected in SRD-treated rats, prior to the development ofinsulin resistance, could involve inflammation-related pathways activated inaddition to or as a consequence of the generation of oxidative stress, as wepreviously demonstrated in pituitary tissues from SRD-treated rats.