Molecular and Cellular Endocrinology

MicroRNA-126 upregulation, induced by training, plays a role in controlling microcirculation in dexamethasone treated rats Publication date: 5 April 2020 Source: Molecular and Cellular Endocrinology, Volume 505 Author(s): Naiara A. Herrera, Francine Duchatsch, Lidieli P. Tardelli, Thiago J. Dionísio, Andre L. Shinohara, Carlos F. Santos, Sandra Lia Amaral Abstract Microcirculation maintenance is associated with microRNAs. Nevertheless, the role of microRNAs induced by training in preventing dexamethasone (DEX)-induced microvascular rarefaction remains unknown. The study aim was to investigate if training-induced microRNAs are able to improve microcirculation proteins and prevent DEX-induced microvascular rarefaction. Rats underwent training for 8 weeks and then were treated with DEX (50 μg/kg per day, s.c.) for 14 days. Arterial pressure was measured and tibialis anterior (TA) muscle was collected for analyses. DEX induced hypertension concomitantly with capillary density loss (CD, −23.9%) and decrease of VEGF (−43.0%), p-AKT/AKT (−39.6%) and Bcl-2 (−23.0%) and an increase in caspase-3-cleaved protein level (+34.0%) in TA muscle. Training upregulated microRNA-126 expression (+13.1%), prevented VEGF (+61.4%), p-AKT/AKT (+37.7%), Bcl-2 (+7.7%) decrease and caspase-3-cleaved (−23.1%) increase associated with CD (+54.7%) reduction and hypertension prevention. MiRNA-126 upregulation, induced by training, plays a role in controlling microcirculation, which may be a potential target against DEX-induced microvascular rarefaction.

Hyposialylated angiopoietin-like-4 induces apoptosis of podocytes via β1 Integrin/FAK signaling in diabetic nephropathy Publication date: 5 April 2020 Source: Molecular and Cellular Endocrinology, Volume 505 Author(s): Kaifeng Guo, Pan Pan, Mian Wu, Yiwen Ma, Junxi Lu, Haibing Chen Abstract Angiopoietin-like-4 (ANGPTL4) is reported to mediate proteinuria in some types of glomerulonephropathy. However, the mechanism underlying the effect on podocytes of ANGPTL4 under pathologic conditions in diabetic nephropathy (DN) is unclear. We investigated the role of ANGPTL4 in the pathogenesis of DN. In DN rats, elevated ANGPTL4 expression was associated with increased proteinuria, glomerular hypertrophy, and ultrastructural changes in podocytes. In vitro, hyperglycemia induced the upregulation of ANGPTL4, which led to activation of integrin-β1/FAK signaling with increased apoptosis of podocytes and actin cytoskeleton derangement. These pathological changes were reversed by transfection with a lentivirus expressing short hairpin RNA against integrin-β1 or an ANGPTL4-neutralizing antibody in vitro. Furthermore, supplementation with the sialic acid precursor ManNAc reversed these pathological changes and conferred renoprotection in a mouse model of DN. Our findings suggest that ANGPTL4 mediates high glucose-induced loss of podocytes by modulating their detachment and apoptosis in vivo and in vitro. This study deepens our understanding of the mechanisms of podocyte loss in DN and shows targeting ANGPTL4-related signaling has therapeutic potential for DN.

Transcriptional regulation of retinol binding protein 4 by Interleukin-6 via peroxisome proliferator-activated receptor α and CCAAT/Enhancer binding proteins Publication date: 5 April 2020 Source: Molecular and Cellular Endocrinology, Volume 505 Author(s): Muzaida Aminah Mohd, Nur Adelina Ahmad Norudin, Tengku Sifzizul Tengku Muhammad Abstract Interleukin-6 (IL-6) is a major mediator of the acute phase response (APR) that regulates the transcription of acute phase proteins (APPs) in the liver. During APR, the plasma levels of negative APPs including retinol binding protein 4 (RBP4) are reduced. Activation of the IL-6 receptor and subsequent signaling pathways leads to the activation of transcription factors, including peroxisome proliferator-activated receptor alpha (PPARα) and CCAAT/enhancer binding protein (C/EBP), which then modulate APP gene expression. The transcriptional regulation of RBP4 by IL-6 is not fully understood. Therefore, this study aimed to elucidate the molecular mechanisms of PPARα and C/EBP isoforms in mediating IL-6 regulation of RBP4 gene expression. IL-6 was shown to reduce the transcriptional activity of RBP4, and functional dissection of the RBP4 promoter further identified the cis-acting regulatory elements that are responsible in mediating the inhibitory effect of IL-6. The binding sites for PPARα and C/EBP present in the RBP4 promoter were predicted at −1079 bp to −1057 bp and −1460 bp to −1439 bp, respectively. The binding of PPARα and C/EBPs to their respective cis-acting elements may lead to antagonistic interactions that modulate the IL-6 regulation of RBP4 promoter activity. Therefore, this study proposed a new mechanism of interaction involving PPARα and different C/EBP isoforms. This interaction is necessary for the regulation of RBP4 gene expression in response to external stimuli, particularly IL-6, during physiological changes.

Growth differentiation factor 15 (GDF-15) is a novel orexigen in fish Publication date: 5 April 2020 Source: Molecular and Cellular Endocrinology, Volume 505 Author(s): Ayelén Melisa Blanco, Juan Ignacio Bertucci, Cristina Velasco, Suraj Unniappan Abstract Growth differentiation factor 15 (GDF-15), an anti-inflammatory and anti-tumorigenic cytokine, has been emerging as a regulator of appetite and energy homeostasis in mammals. In fish, the physiological role of this peptide remains to be elucidated. This research aimed to determine the possible role of GDF-15 on food intake in goldfish (Carassius auratus). To achieve our objectives, we first obtained a 595 bp gdf-15 cDNA sequence from goldfish tissues, and examined the tissue expression profile of mRNAs encoding both GDF-15 and its receptor (GFRAL). Both mRNAs were detected in several goldfish tissues, including the hypothalamus, foregut and liver (main tissues regulating appetite and energy balance). Food deprivation for 3 and 7 days significantly upregulated gdf-15 mRNAs in the foregut, but downregulated them in the liver. Our in vivo study using diets with varying amounts of carbohydrates, proteins and fats, and our in vitro study exposing goldfish tissues to different macronutrients revealed that gdf-15 mRNAs are importantly modulated by macronutrients. In general terms, we found an increase in gdf-15 mRNA levels in the goldfish foregut and liver in response to all macronutrients tested. Finally, our in vivo study testing the effects of GDF-15 on appetite levels demonstrated an important dose-dependent orexigenic role for this peptide in goldfish. Results from this study described GDF-15 as a novel regulator of appetite in fish, importantly modulated by food availability and diet composition.

Improvement of human pancreatic islet quality after co-culture with human adipose-derived stem cells Publication date: 5 April 2020 Source: Molecular and Cellular Endocrinology, Volume 505 Author(s): Bianca M. de Souza, Michelle Rodrigues, Fernanda S. de Oliveira, Liana P.A. da Silva, Ana P. Bouças, Ciro P. Portinho, Bruno P. dos Santos, Melissa Camassola, Dagoberto Rocha, Simone Lysakowski, Juliano Martini, Cristiane B. Leitão, Nance B. Nardi, Andrea C. Bauer, Daisy Crispim Abstract The aim of this study was to investigate whether co-culture of human islets with adipose-derived stem cells (ASCs) can improve islet quality and to evaluate which factors play a role in the protective effect of ASCs against islet dysfunction. Islets and ASCs were cultured in three experimental groups for 24 h, 48 h, and 72 h: 1) indirect co-culture of islets with ASC monolayer (Islets/ASCs); 2) islets alone; and 3) ASCs alone. Co-culture with ASCs improved islet viability and function in all culture time-points analyzed. VEGFA, HGF, IL6, IL8, IL10, CCL2, IL1B, and TNF protein levels were increased in supernatants of islet/ASC group compared to islets alone, mainly after 24 h. Moreover, VEGFA, IL6, CCL2, HIF1A, XIAP, CHOP, and NFKBIA genes were differentially expressed in islets from the co-culture condition compared to islets alone. In conclusion, co-culture of islets with ASCs promotes improvements in islet quality.

p.L571P in the linker domain of rat thyroglobulin causes intracellular retention Publication date: 5 April 2020 Source: Molecular and Cellular Endocrinology, Volume 505 Author(s): Cintia E. Citterio, Sofia Siffo, Christian M. Moya, Mauricio Gomes Pio, Maricel F. Molina, Karen G. Scheps, Osvaldo A. Rey, Peter Arvan, Carina M. Rivolta, Héctor M. Targovnik Abstract Thyroglobulin (TG), a large glycosylated protein secreted by thyrocytes into the thyroid follicular lumen, plays an essential role in thyroid hormone biosynthesis. Rattus norvegicus TG (rTG) is encoded by a large single copy gene, 186-kb long, located on chromosome 7 composed of 48 exons encoding a 8461-kb mRNA. Although the TG gene displays sequence variability, many missense mutations do not impose any adverse effect on the TG protein, whereas other nucleotide substitutions may affect its TG stability and/or TG intracellular trafficking. In order to gain a further understanding of the protein domains regulating its intracellular fate, we cloned a full-length cDNA from rTG into the pcDNA6/V5-His B expression vector. However, transient expression of the cDNA in HEK293T cells showed that the encoded protein was not a wild-type molecule, as it was unable to be secreted in the culture supernatant. Sequencing analyses revealed three random mutations, which accidentally emerged during the course of cloning: c.1712T>C [p.L571P] in the linker domain (amino acid positions 360 to 604), c.2027A>G [p.Q676R] in TG type 1–6 repeat and c.2720A>G [p.Q907R] in the TG type 1–7 repeat. Expression of cDNAs encoding a combination of two mutations [p.Q676R-p.Q907R], [p.L571P-p.Q907R] or [p.L571P-p.Q676R] indicated that any TG bearing the p.L571P substitution was trapped intracellularly. Indeed, we expressed the single point mutant p.L571P and confirmed that this point mutation was sufficient to cause intracellular retention of mutant TG in HEK293T cells. Endo H analysis showed that the p.L571P mutant is completely sensitive to the enzyme, whereas the will-type TG acquires full N-glycan modifications in Golgi apparatus. This data suggest that the p.L571P mutant contains the mannose-type N-glycan, that was added at the first stage of glycosylation. Complex-type N-glycan formation in the Golgi apparatus does not occur, consistent with defective endoplasmic reticulum exit of the mutant TG. Moreover, predictive analysis of the 3D linker domain showed that the p.L571P mutation would result in a significant protein conformational change. In conclusion, our studies identified a novel amino acid residue within the linker domain of TG associated with its conformational maturation and intracellular trafficking.

ALK4-SMAD3/4 mediates the effects of activin A on the upregulation of PAI-1 in human granulosa lutein cells Publication date: 5 April 2020 Source: Molecular and Cellular Endocrinology, Volume 505 Author(s): Beili Chen, Hsun-Ming Chang, Zhiguo Zhang, Yunxia Cao, Peter C.K. Leung Abstract In the mammalian ovary, the proteolysis of the extracellular matrix is dynamically regulated by plasminogen activator and plasminogen activator inhibitor (PAI), and it is a critical event that influences various physiological and pathological processes. Activin A is a member of the transforming growth factor-β superfamily and is expressed at a high level in human luteal cells that play an essential role in the regulation of the luteal function. At present, it is not known whether activin A can regulate the expression and production of PAI in human granulosa lutein (hGL) cells. The present study aimed to examine the effects of activin A on the expression and production of intraovarian PAI-1 and the underlying molecular mechanisms. Using primary and immortalized hGL cells as the cell model, we demonstrated that activin A upregulated the expression of PAI-1 and increased the production of PAI-1 in an autocrine/paracrine manner. Additionally, using a dual inhibition approach (molecular inhibitors and siRNA-mediated knockdown), we showed that this biological function is mediated by the ALK4-mediated SMAD3-SMAD4-dependent signaling pathway. Our findings suggest that activin A may be involved in the regulation of luteal function via the induction of PAI-1 expression and an increase in PAI-1 production.

Hormones and human developmental plasticity Publication date: 5 April 2020 Source: Molecular and Cellular Endocrinology, Volume 505 Author(s): Davide Ponzi, Mark V. Flinn, Michael P. Muehlenbein, Pablo A. Nepomnaschy Abstract Natural selection favors the evolution of mechanisms that optimize the allocation of resources and time among competing traits. Hormones mediate developmental plasticity, the changes in the phenotype that occur during ontogeny. Despite their highly conserved functions, the flexibilities of human hormonal systems suggest a strong history of adaptation to variable environments. Physiological research on developmental plasticity has focused on the early programming effects of stress, the hypothalamus-pituitary-adrenal axis (HPAA) and the hypothalamus-pituitary-gonadal axis (HPGA) during critical periods, when the hormones produced have the strongest influence on the developing brain. Often this research emphasizes the maladaptive effects of early stressful experiences. Here we posit that the HPAA and HPAG systems in human developmental plasticity have evolved to be responsive to complex and dynamic problems associated with human sociality. The lengthy period of human offspring dependency, and its associated brain development and risks, is linked to the uniquely human combination of stable breeding bonds, extensive paternal effort in a multi-male group, extended bilateral kin recognition, grandparenting, and controlled exchange of mates among kin groups. We evaluate an evolutionary framework that integrates proximate physiological explanations with ontogeny, phylogeny, adaptive function, and comparative life history data.

The tissue-specific effects of different 17β-estradiol doses reveal the key sensitizing role of AF1 domain in ERα activity Publication date: 5 April 2020 Source: Molecular and Cellular Endocrinology, Volume 505 Author(s): Coralie Fontaine, Melissa Buscato, Alexia Vinel, Frank Giton, Isabelle Raymond-Letron, Sung Hoon Kim, Benita S. Katzenellenbogen, John A. Katzenellenbogen, Pierre Gourdy, Alain Milon, Gilles Flouriot, Claes Ohlsson, Françoise Lenfant, Jean-François Arnal Abstract 17β-Estradiol (E2) action can be mediated by the full-length estrogen receptor alpha (ERα66), and also by the AF1 domain-deficient ERα (ERα46) isoform, but their respective sensitivity to E2 is essentially unknown. We first performed a dose response study using subcutaneous home-made pellets mimicking either metestrus, proestrus or a pharmacological doses of E2, which resulted in plasma concentrations around 3, 30 and 600 pM, respectively. Analysis of the uterus, vagina and bone after chronic exposure to E2 demonstrated dose-dependent effects, with a maximal response reached at the proestrus-dose in wild type mice expressing mainly ERα66. In contrast, in transgenic mice harbouring only an ERα deleted in AF1, these effects of E2 were either strongly shifted rightward (10–100-fold) and/or attenuated, depending on the tissue studied. Finally, experiments in different cell lines transfected with ERα66 or ERα46 also delineated varying profiles of ERα AF1 sensitivity to E2. Altogether, this work emphasizes the importance of dose in the tissue-specific actions of E2 and demonstrates the key sensitizing role of AF1 in ERα activity.

The START-domain proteins in intracellular lipid transport and beyond Publication date: 15 March 2020 Source: Molecular and Cellular Endocrinology, Volume 504 Author(s): Barbara J. Clark Abstract The Steroidogenic Acute Regulatory Protein-related Lipid Transfer (START) domain is a ~210 amino acid sequence that folds into an α/β helix-grip structure forming a hydrophobic pocket for lipid binding. The helix-grip fold structure defines a large superfamily of proteins, and this review focuses on the mammalian START domain family members that include single START domain proteins with identified ligands, and larger multi-domain proteins that may have novel roles in metabolism. Much of our understanding of the mammalian START domain proteins in lipid transport and changes in metabolism has advanced through studies using knockout mouse models, although for some of these proteins the identity and/or physiological role of ligand binding remains unknown. The findings that helped define START domain lipid-binding specificity, lipid transport, and changes in metabolism are presented to highlight that fundamental questions remain regarding the biological function(s) for START domain-containing proteins.