Showing papers in "Journal of Endocrinology in 2020"

SCFAs alleviated steatosis and inflammation in mice with NASH induced by MCD.

Abstract: This study aimed to assess the effects of three major SCFAs (acetate, propionate, and butyrate) on NASH phenotype in mice. C57BL/6 mice were fed a methionine- and choline-deficient (MCD) diet and treated with sodium acetate, sodium propionate, or sodium butyrate during the 6-week feeding period. SCFA treatment significantly reduced serum levels of alanine aminotransferase and aspartate transaminase, the numbers of lipid droplets, and the levels of triglycerides and cholesterols in livers of the mice compared with control treatment. SCFAs also reduced MCD-induced hepatic aggregation of macrophages and proinflammatory responses. Among the three SCFAs, sodium acetate (NaA) revealed the best efficacy at alleviating MCD-induced hepatic steatosis and inflammation. Additionally, NaA increased AMP-activated protein kinase activation in the liver and induced the expression of fatty acid oxidation gene in both the liver and cultured hepatocytes. In vitro, NaA decreased MCD-mimicking media-induced proinflammatory responses in macrophages to a greater extent than in hepatocytes. These results indicated that NaA alleviates steatosis in a manner involving AMPK activation. Also, NaA alleviation of hepatic inflammation appears to be due to, in large part, suppression of macrophage proinflammatory activation. SCFAs may represent as a novel and viable approach for alleviating NASH.

Mechanisms controlling hormone secretion in human gut and its relevance to metabolism

Abstract: The homoeostatic regulation of metabolism is highly complex and involves multiple inputs from both the nervous and endocrine systems. The gut is the largest endocrine organ in our body and synthesises and secretes over 20 different hormones from enteroendocrine cells that are dispersed throughout the gut epithelium. These hormones include GLP-1, PYY, GIP, serotonin, and CCK, each of whom play pivotal roles in maintaining energy balance and glucose homeostasis. Some are now the basis of several clinically used glucose-lowering and weight loss therapies. The environment in which these enteroendocrine cells exist is also complex, as they are exposed to numerous physiological inputs including ingested nutrients, circulating factors and metabolites produced from neighbouring gut microbiome. In this review, we examine the diverse means by which gut-derived hormones carry out their metabolic functions through their interactions with different metabolically important organs including the liver, pancreas, adipose tissue and brain. Furthermore, we discuss how nutrients and microbial metabolites affect gut hormone secretion and the mechanisms underlying these interactions.

Liver-expressed antimicrobial peptide 2 antagonizes the effect of ghrelin in rodents.

Abstract: Ghrelin, a stomach-derived peptide, promotes feeding and growth hormone (GH) secretion. A recent study identified liver-expressed antimicrobial peptide 2 (LEAP2) as an endogenous inhibitor of ghrelin-induced GH secretion, but the effect of LEAP2 in the brain remained unknown. In this study, we showed that intracerebroventricular (i.c.v.) administration of LEAP2 to rats suppressed central ghrelin functions including Fos expression in the hypothalamic nuclei, promotion of food intake, blood glucose elevation, and body temperature reduction. LEAP2 did not inhibit neuropeptide Y (NPY)-induced food intake or des-acyl ghrelin-induced reduction in body temperature, indicating that the inhibitory effects of LEAP2 were specific for GHSR. Plasma LEAP2 levels varied according to feeding status and seemed to be dependent on the hepatic Leap2 expression. Furthermore, ghrelin suppressed the expression of hepatic Leap2 via AMPK activation. Together, these results reveal that LEAP2 inhibits central ghrelin functions and crosstalk between liver and stomach.

Hyperandrogenism and insulin resistance modulate gravid uterine and placental ferroptosis in PCOS-like rats

Abstract: Women with polycystic ovary syndrome (PCOS) have hyperandrogenism and insulin resistance and a high risk of miscarriage during pregnancy Similarly, in rats, maternal exposure to 5α-dihydrotestosterone (DHT) and insulin from gestational day 75 to 135 leads to hyperandrogenism and insulin resistance and subsequently increased fetal loss A variety of hormonal and metabolic stimuli are able to trigger different types of regulated cell death under physiological and pathological conditions These include ferroptosis, apoptosis and necroptosis We hypothesized that, in rats, maternal hyperandrogenism and insulin-resistance-induced fetal loss is mediated, at least in part, by changes in the ferroptosis, apoptosis and necroptosis pathways in the gravid uterus and placenta Compared with controls, we found that co-exposure to DHT and insulin led to decreased levels of glutathione peroxidase 4 (GPX4) and glutathione, increased glutathione + glutathione disulfide and malondialdehyde, aberrant expression of ferroptosis-associated genes (Acsl4, Tfrc, Slc7a11, and Gclc), increased iron deposition and activated ERK/p38/JNK phosphorylation in the gravid uterus In addition, we observed shrunken mitochondria with electron-dense cristae, which are key features of ferroptosis-related mitochondrial morphology, as well as increased expression of Dpp4, a mitochondria-encoded gene responsible for ferroptosis induction in the uteri of rats co-exposed to DHT and insulin However, in the placenta, DHT and insulin exposure only partially altered the expression of ferroptosis-related markers (eg region-dependent GPX4, glutathione + glutathione disulfide, malondialdehyde, Gls2 and Slc7a11 mRNAs, and phosphorylated p38 levels) Moreover, we found decreased expression of Dpp4 mRNA and increased expression of Cisd1 mRNA in placentas of rats co-exposed to DHT and insulin Further, DHT + insulin-exposed pregnant rats exhibited decreased apoptosis in the uterus and increased necroptosis in the placenta Our findings suggest that maternal hyperandrogenism and insulin resistance causes the activation of ferroptosis in the gravid uterus and placenta, although this is mediated via different mechanisms operating at the molecular and cellular levels Our data also suggest that apoptosis and necroptosis may play a role in coordinating or compensating for hyperandrogenism and insulin-resistance-induced ferroptosis when the gravid uterus and placenta are dysfunctional

Endocrine aspects of ACE2 regulation: RAAS, steroid hormones and SARS-CoV-2.

Abstract: Coronavirus disease (COVID-19) is caused by a new strain of coronavirus, the severe acute respiratory syndrome coronavirus 2 or SARS-CoV-2. At the time of writing, SARS-CoV-2 has infected over 5 million people worldwide. A key step in understanding the pathobiology of the SARS-CoV-2 was the identification of -converting enzyme 2 (ACE2) as the receptor for SARS-CoV-2 to gain entry into host cells. ACE2 is an established component of the 'protective arm' of the renin-angiotensin-aldosterone-system (RAAS) that opposes ACE/angiotensin II (ANG II) pressor and tissue remodelling actions. Identification of ACE2 as the entry point for SARS-CoV-2 into cells quickly focused attention on the use of ACE inhibitors (ACEi), angiotensin receptor blockers (ARB) and mineralocorticoid receptor antagonists (MRA) in patients with hypertension and cardiovascular disease given that these pharmacological agents upregulate ACE2 expression in target cells. ACE2 is cleaved from the cells by metalloproteases ADAM10 and ADAM17. Steroid hormone receptors regulate multiple components of the RAAS and may contribute to the observed variation in the incidence of severe COVID-19 between men and women, and in patients with pre-existing endocrine-related disease. Moreover, glucocorticoids play a critical role in the acute and chronic management of inflammatory disease, independent of any effect on RAAS activity. Dexamethasone, a synthetic glucocorticoid, has emerged as a life-saving treatment in severe COVID-19. This review will examine the endocrine mechanisms that control ACE2 and discusses the impact of therapies targeting the RAAS, glucocorticoid and other endocrine systems for their relevance to the impact of SARS-CoV-2 infection and the treatment and recovery from COVID-19-related critical illness.

JAK2/STAT3 regulates estrogen-related senescence of bone marrow stem cells

Abstract: Emerging evidence has indicated that estrogen deficiency contributes to osteoporosis by affecting the level of inflammation. The inflammation microenvironment affects many cellular physiological processes, one of which may be cellular senescence according to previous studies. Senescent cells cannot function normally and secrete inflammatory cytokines and degradative proteins, which are referred to as senescence-associated secretory phenotype (SASP) factors, inducing further senescence and inflammation. Thus, stopping this vicious cycle may be helpful for postmenopausal osteoporosis treatment. Here, we used ovariectomized (OVX) mice as an estrogen-deficient model and confirmed that OVX bone marrow mesenchymal stem cells (BMSCs) displayed a senescent phenotype and upregulated SASP factor secretion both in vitro and in vivo. Furthermore, JAK2/STAT3, an important cytokine secretion-related signalling pathway that is associated with SASP secretion, was activated. Estrogen addition and estrogen receptor blockade confirmed that the JAK2/STAT3 axis participated in OVX BMSC senescence by mediating SASP factors. And JAK inhibition reduced SASP factor expression, alleviated senescence and enhanced osteogenic differentiation. Intraperitoneal injection of a JAK inhibitor, ruxolitinib, prevented bone loss in OVX mice. Collectively, our results revealed that JAK2/STAT3 plays an important role in the inflammation-senescence-SASP feedback loop in OVX BMSCs and that JAK inhibition could be a new method for treating postmenopausal osteoporosis.

PKA functions in metabolism and resistance to obesity: lessons from mouse and human studies

Abstract: Both direct and indirect evidence demonstrate a central role for the cAMP-dependent protein kinase (PKA) signaling pathway in the regulation of energy balance and metabolism across multiple systems. However, the ubiquitous pattern of PKA expression across cell types poses a challenge in pinpointing its tissue-specific regulatory functions and further characterizing its many downstream effects in certain organs or cells. Mouse models of PKA deficiency and over-expression and studies in living cells have helped clarify PKA function in adipose tissue (AT), liver, adrenal, pancreas, and specific brain nuclei, as they pertain to energy balance and metabolic dysregulation. Limited studies in humans suggest differential regulation of PKA in AT of obese compared to lean individuals and an overall dysregulation of PKA signaling in obesity. Despite its complexity, under normal physiologic conditions, the PKA system is tightly regulated by changes in cAMP concentrations upstream via adenylate cyclase and downstream by phosphodiesterase-mediated cAMP degradation to AMP and by changes in PKA holoenzyme stability. Adjustments in the PKA system appear to be important to the development and maintenance of the obese state and its associated metabolic perturbations. In this review we discuss the important role of PKA in obesity and its involvement in resistance to obesity, through studies in humans and in mouse models, with a focus on the regulation of PKA in energy expenditure, intake behavior, and lipid and glucose metabolism.

Androgen signaling pathways driving reproductive and metabolic phenotypes in a PCOS mouse model.

Abstract: As the mechanistic basis of polycystic ovary syndrome (PCOS) remains unknown, current management relies on symptomatic treatment. Hyperandrogenism is a major PCOS characteristic and evidence supports it playing a key role in PCOS pathogenesis. Classically, androgens can act directly through the androgen receptor (AR) or, indirectly, following aromatization, via the estrogen receptor (ER). We investigated the mechanism of androgenic actions driving PCOS by comparing the capacity of non-aromatizable dihydrotestosterone (DHT) and aromatizable testosterone to induce PCOS traits in WT and Ar-knockout (ARKO) mice. DHT and testosterone induced the reproductive PCOS-like features of acyclicity and anovulation in WT females. In ARKO mice, DHT did not cause reproductive dysfunction; however, testosterone treatment induced irregular cycles and ovulatory disruption. These findings indicate that direct AR actions and indirect, likely ER, actions of androgens are important mediators of PCOS reproductive traits. DHT, but not testosterone, induced an increase in body weight, body fat, serum cholesterol and adipocyte hypertrophy in WT mice, but neither androgen induced these metabolic features in ARKO mice. These data infer that direct AR-driven mechanisms are key in driving the development of PCOS metabolic traits. Overall, these findings demonstrate that differing PCOS traits can be mediated via different steroid signaling pathways and indicate that a phenotype-based treatment approach would ensure effective targeting of the underlying mechanisms.

Androgens, oestrogens and endometrium: a fine balance between perfection and pathology

Abstract: The endometrium is a complex multicellular tissue that is exquisitely sensitive to the actions of sex steroids synthesised in the ovary (endocrine system). Recent studies have highlighted a previously under-appreciated role for local (intracrine) metabolism in fine-tuning tissue function in both health and disease. In this review we have focused on the impact of oestrogens and androgens on endometrial function summarising data from studies on normal endometrial physiology and disorders including infertility, endometriosis and cancer. We consider the evidence that expression of enzymes including aromatase, sulphatase and AKR1C3 by endometrial cells plays an important role in tissue function and malfunction and discuss results from studies using drugs targeting intracrine pathways to treat endometrial disorders. We summarise studies exploring the spatial and temporal expression of oestrogen receptors (ERalpha/ESR1, ERbeta/ESR2 and GPER) and their role in mediating the impact of endogenous and synthetic ligands on cross-talk between vascular, immune, epithelial and stromal cells. There is a single androgen receptor gene and androgens play a key role in stromal-epithelial cross-talk, scar-free healing of endometrium during menstruation and regulation of cell proliferation. The development of new receptor-selective drugs (SERMs, SARMs, SARDs) has reinvigorated interest in targeting receptor subtypes in treatment of disorders including endometriosis and endometrial cancer and some show promise as novel therapies. In summary, understanding the mechanisms regulated by sex steroids provides the platform for improved personalised treatment of endometrial disorders as well as novel insights into the impact of steroids on processes such as tissue repair and regeneration.

Liraglutide and sitagliptin counter beta- to alpha-cell transdifferentiation in diabetes

Abstract: Transdifferentiation of beta- to alpha-cells has been implicated in the pathogenesis of diabetes. To investigate the impact of contrasting aetiologies of beta-cell stress, as well as clinically approved incretin therapies on this process, lineage tracing of beta-cells in transgenic Ins1 Cre/+/Rosa26-eYFP mice was investigated. Diabetes-like syndromes were induced by streptozotocin (STZ), high fat feeding (HFF) or hydrocortisone (HC), and effects of treatment with liraglutide or sitagliptin were investigated. Mice developed the characteristic metabolic features associated with beta-cell destruction or development of insulin resistance. Liraglutide was effective in preventing weight gain in HFF mice, with both treatments decreasing energy intake in STZ and HC mice. Treatment intervention also significantly reduced blood glucose levels in STZ and HC mice, as well as increasing either plasma or pancreatic insulin while decreasing circulating or pancreatic glucagon in all models. The recognised changes in pancreatic morphology induced by STZ, HFF or HC were partially, or fully, reversed by liraglutide and sitagliptin, and related to advantageous effects on alpha- and beta-cell growth and survival. More interestingly, induction of diabetes-like phenotype, regardless of pathogenesis, led to increased numbers of beta-cells losing their identity, as well as decreased expression of Pdx1 within beta-cells. Both treatment interventions, and especially liraglutide, countered detrimental islet cell transitioning effects in STZ and HFF mice. Only liraglutide imparted benefits on beta- to alpha-cell transdifferentiation in HC mice. These data demonstrate that beta- to alpha-cell transdifferentiation is a common consequence of beta-cell destruction or insulin resistance and that clinically approved incretin-based drugs effectively limit this.

Igf2 deletion alters mouse placenta endocrine capacity in a sexually dimorphic manner

Abstract: The placenta regulates materno-fetal nutrient transfer and secretes hormones that enable maternal physiological support of the pregnancy. In mice, these functions are performed by the labyrinth (Lz) and junctional (Jz) zones, respectively. Insulin-like growth factor 2 (Igf2) is an imprinted gene expressed by the conceptus that is important for promoting fetal growth and placenta formation. However, the specific role of Igf2 in the Jz in regulating placental endocrine function and fetal development is unknown. This study used a novel model to investigate the effect of conditional loss of Igf2 in the Jz (Jz-Igf2UE) on placental endocrine cell formation and the expression of hormones and IGF signaling components in placentas from female and male fetuses. Jz-Igf2UE altered gross placental structure and expression of key endocrine and signaling genes in a sexually dimorphic manner. The volumes of spongiotrophoblast and glycogen trophoblast in the Jz were decreased in placentas from female but not male fetuses. Expression of insulin receptor was increased and expression the MAPK pathway genes (Mek1, P38α) decreased in the placental Jz of female but not male fetuses. In contrast, expression of the type-1 and -2 IGF receptors and the MAPK pathway genes (H-ras, N-ras, K-ras) was decreased in the placental Jz from male but not female fetuses. Expression of the steroidogenic gene, Cyp17a1, was increased and placental lactogen-2 was decreased in the placenta of both sexes. In summary, we report that Jz-Igf2UE alters the cellular composition, IGF signaling components and hormone expression of the placental Jz in a manner largely dependent on fetal sex.

Fixing the broken clock in adrenal disorders: focus on glucocorticoids and chronotherapy

Abstract: The circadian rhythm derives from the integration of many signals that shape the expression of clock-related genes in a 24-h cycle. Biological tasks, including cell proliferation, differentiation, energy storage, and immune regulation, are preferentially confined to specific periods. A gating system, supervised by the central and peripheral clocks, coordinates the endogenous and exogenous signals and prepares for transition to activities confined to periods of light or darkness. The fluctuations of cortisol and its receptor are crucial in modulating these signals. Glucocorticoids and the autonomous nervous system act as a bridge between the suprachiasmatic master clock and almost all peripheral clocks. Additional peripheral synchronizing mechanisms including metabolic fluxes and cytokines stabilize the network. The pacemaker is amplified by peaks and troughs in cortisol and their response to food, activity, and inflammation. However, when the glucocorticoid exposure pattern becomes chronically flattened at high- (as in Cushing's syndrome) or low (as in adrenal insufficiency) levels, the system fails. While endocrinologists are well aware of cortisol rhythm, too little attention has been given to interventions aimed at restoring physiological cortisol fluctuations in adrenal disorders. However, acting on glucocorticoid levels may not be the only way to restore clock-related activities. First, a counterregulatory mechanism on the glucocorticoid receptor itself controls signal transduction, and second, melatonin and/or metabolically active drugs and nutrients could also be used to modulate the clock. All these aspects are described herein, providing some insights into the emerging role of chronopharmacology, focusing on glucocorticoid excess and deficiency disorders.

microRNA-375 regulates glucose metabolism-related signaling for insulin secretion

Abstract: Enhanced beta cell glycolytic and oxidative metabolism are necessary for glucose-induced insulin secretion. While several microRNAs modulate beta cell homeostasis, miR-375 stands out as it is highly expressed in beta cells where it regulates beta cell function, proliferation and differentiation. As glucose metabolism is central in all aspects of beta cell functioning, we investigated the role of miR-375 in this process using human and rat islets; the latter being an appropriate model for in-depth investigation. We used forced expression and repression of mR-375 in rat and human primary islet cells followed by analysis of insulin secretion and metabolism. Additionally, miR-375 expression and glucose-induced insulin secretion were compared in islets from rats at different developmental ages. We found that overexpressing of miR-375 in rat and human islet cells blunted insulin secretion in response to glucose but not to α-ketoisocaproate or KCl. Further, miR-375 reduced O2 consumption related to glycolysis and pyruvate metabolism, but not in response to α-ketoisocaproate. Concomitantly, lactate production was augmented suggesting that glucose-derived pyruvate is shifted away from mitochondria. Forced miR-375 expression in rat or human islets increased mRNA levels of pyruvate dehydrogenase kinase-4, but decreased those of pyruvate carboxylase and malate dehydrogenase1. Finally, reduced miR-375 expression was associated with maturation of fetal rat beta cells and acquisition of glucose-induced insulin secretion function. Altogether our findings identify miR-375 as an efficacious regulator of beta cell glucose metabolism and of insulin secretion, and could be determinant to functional beta cell developmental maturation.

Loss of Cyp11c1 causes delayed spermatogenesis due to the absence of 11-ketotestosterone.

Abstract: The impacts of androgens and glucocorticoids on spermatogenesis have intrigued scientists for decades. 11β-hydroxylase, encoded by cyp11c1, is the key enzyme involved in the synthesis of 11-ketotestosterone and cortisol, the major androgen and glucocorticoid in fish, respectively. In the present study, a Cyp11c1 antibody was produced. Western blot and immunohistochemistry showed that Cyp11c1 was predominantly expressed in the testicular Leydig cells and head kidney interrenal cells. A mutant line of cyp11c1 was established by CRISPR/Cas9. Homozygous mutation of cyp11c1 caused a sharp decrease of serum cortisol and 11-ketotestosterone, and a delay in spermatogenesis which could be rescued by exogenous 11-ketotestosterone or testosterone, but not cortisol treatment. Intriguingly, this spermatogenesis restored spontaneously, indicating compensatory effects of other androgenic steroids. In addition, loss of Cyp11c1 led to undersized testes with a smaller efferent duct and disordered spermatogenic cysts in adult males. However, a small amount of viable sperm was produced. Taken together, our results demonstrate that cyp11c1 is important for testicular development, especially for the initiation and proper progression of spermatogenesis. 11-ketotestosterone is the most efficient androgen in tilapia.

SGK1 activation exacerbates diet-induced obesity, metabolic syndrome and hypertension.

Abstract: The serum- and glucocorticoid-induced kinase 1 (SGK1) is a transcriptional target of steroid hormones including glucocorticoids or aldosterone in addition to other stimuli such as glucose. SGK1 is activated via phosphoinositide 3-kinase, placing it downstream of insulin signaling. SGK1 participates in the upregulation of kidney Na+ reabsorption by aldosterone and has been linked to obesity-related hypertension in humans. We hypothesized that a systemic increase in SGK1 activity may trigger a multiplicity of mechanisms leading to simultaneous development of the main conditions that characterize the metabolic syndrome (MetS), including hypertension. We used a transgenic mouse model made with a bacterial artificial chromosome containing the whole mouse Sgk1 gene modified to introduce an activating point mutation. Wild type or transgenic 14-week-old male mice were fed with standard chow diet or high-fat diet for up to 18 weeks. Development of the main features of MetS and hepatic steatosis were monitored, and in vitro adipocyte differentiation was studied. Our results show that transgenic animals under high-fat diet rapidly and markedly develop MetS characterized by obesity, glucose intolerance, insulin resistance, dyslipidemia and hypertension. In addition, SGK1 gain-of-function accelerates the development of hepatic steatosis. Our study suggests that inappropriate SGK1 activity represents a risk factor in developing MetS with hypertension and related end-organ damage. Our data support SGK1 as a possible therapeutic target in MetS and related complications and provides a useful gain-of-function model for pre-clinical drug testing.

The therapeutic effect of interleukin-22 in high androgen-induced polycystic ovary syndrome.

Abstract: Polycystic ovary syndrome (PCOS) is a complex syndrome involving both endocrine and metabolic disorders. Gut microbiota and the intestinal immune factor IL-22 play an important role in the pathogenesis of PCOS. However, the therapeutic role of IL-22 in high androgen-induced PCOS mice is not clear. We aimed to determine the therapeutic effects of IL-22 on the DHEA-induced PCOS mouse model and to explore the possible mechanism of IL-22 in regulating hyperandrogenism-associated PCOS. Insulin resistance levels and ovarian functions were investigated in DHEA-induced PCOS mice with or without additional IL-22 treatment. We found that IL-22 could reverse insulin resistance, disturbed estrous cycle, abnormal ovary morphology, and decreased embryo number in DHEA mice. Mechanistically, IL-22 upregulated the browning of white adipose tissue in DHEA mice. This study demonstrated that IL-22-associated browning of white adipose tissue regulated insulin sensitivity and ovarian functions in PCOS, suggesting that IL-22 may be of value for the treatment of PCOS with a hyperandrogenism phenotype.

Glucocorticoid and mineralocorticoid receptor activation modulates postnatal growth

Abstract: During early development, stress or exogenous glucocorticoid (GC) administration reduces body mass in vertebrates, and this is associated with the glucocorticoid receptor (GR) activation. Although GCs also activate the mineralocorticoid receptor (MR), the physiological significance of MR activation on early developmental growth is unknown. We tested the hypothesis that activation of both GR and MR are required for postnatal growth suppression by GCs. Differential regulation of GR and MR activation was achieved by using ubiquitous GR- (GRKO) and MR- (MRKO) knockout zebrafish (Danio rerio) in combination with exogenous cortisol treatment. MR activation increased protein deposition in zebrafish larvae and also upregulated lepa and downregulated lepr transcript abundance. Cortisol treatment reduced body mass and protein content in the WT, and this corresponded with the upregulation of muscle proteolytic markers, including murf1 and redd1 by GR activation. The combined activation of MR and GR by cortisol also upregulated the gh and igf1 transcript abundance, and insulin expression compared to the WT. However, cortisol-mediated reduction in body mass and protein content required the activation of both MR and GR, as activation by GR alone (MRKO + cortisol) did not reduce the larval protein content. Collectively, our results indicate that MR activation favors protein deposition and GR activation stimulates proteolysis, while their combined activation is involved in cortisol-mediated growth suppression. Overall, this work provides insight into the physiological significance of MR activation in regulating protein deposition during early development at a systems level.

Plasticity in medaka gonadotropes via cell proliferation and phenotypic conversion.

Abstract: Follicle-stimulating hormone (Fsh) and luteinizing hormone (Lh) produced by the gonadotropes play a major role in control of reproduction. Contrary to mammals and birds, Lh and Fsh are mostly produced by two separate cell types in teleost. Here, we investigated gonadotrope plasticity, using transgenic lines of medaka (Oryzias latipes) where DsRed2 and hrGfpII are under the control of the fshb and lhb promotors respectively. We found that Fsh cells appear in the pituitary at 8 dpf, while Lh cells were previously shown to appear at 14 dpf. Similar to Lh cells, Fsh cells show hyperplasia from juvenile to adult stages. Hyperplasia is stimulated by estradiol. Both Fsh and Lh cells show hypertrophy during puberty with similar morphology. They also share similar behavior, using their cellular extensions to make networks. We observed bi-hormonal gonadotropes in juveniles and adults but not in larvae where only mono-hormonal cells are observed, suggesting the existence of phenotypic conversion between Fsh and Lh in later stages. This is demonstrated in cell culture, where some Fsh cells start to produce Lhβ, a phenomenon enhanced by gonadotropin-releasing hormone (Gnrh) stimulation. We have previously shown that medaka Fsh cells lack Gnrh receptors, but here we show that with time in culture, some Fsh cells start responding to Gnrh, while fshb mRNA levels are significantly reduced, both suggestive of phenotypic change. All together, these results reveal high plasticity of gonadotropes due to both estradiol-sensitive proliferation and Gnrh promoted phenotypic conversion, and moreover, show that gonadotropes lose part of their identity when kept in cell culture.

Night-time feeding of Bmal1-/- mice restores SCFA rhythms and their effect on ghrelin.

Abstract: The known crosstalk between short-chain fatty acids (SCFAs) and the circadian clock is tightly intertwined with feeding time. We aimed to investigate the role of the core clock gene Bmal1 and feeding time in the diurnal rhythms in plasma and caecal SCFA levels and in their effect on the release of the hunger hormone ghrelin in the stomach and colon. WT, Bmal1-/- (ad libitum fed) and night-time-restricted-fed (RF)-Bmal1-/- littermates were killed at zeitgeber time (ZT) 4 and 16. SCFA concentrations were measured by gas chromatography. To investigate the effect of SCFAs on ghrelin release, stomach and colonic full-thickness strips were incubated with Krebs or a SCFA mix mimicking plasma or caecal concentrations, after which octanoyl ghrelin release was measured by RIA. Diurnal rhythms in caecal and plasma SCFAs oscillated in phase but rhythmic changes were abolished in Bmal1-/- mice. RF of Bmal1-/- mice restored fluctuations in caecal SCFAs. Plasma SCFA concentrations failed to affect gastric ghrelin release. The effect of caecal SCFA concentrations on colonic ghrelin release was rhythmic (inhibition at ZT 4, no effect at ZT 16). In Bmal1-/- mice, the inhibitory effect of SCFAs at ZT 4 was abolished. RF Bmal1-/- mice restored the inhibitory effect and increased colonic Clock expression. To conclude, diurnal fluctuations in caecal SCFAs and the effect of SCFAs on colonic ghrelin release are regulated by feeding time, independent of the core clock gene Bmal1. However, local entrainment of other clock genes might contribute to the observed effects.

The interplay between stress, circadian clocks, and energy metabolism

Abstract: Endogenous circadian clocks adapt an organism's physiology and behavior to predictable changes in the environment as a consequence of the Earth's rotation around its axis. In mammals, circadian rhythms are the output of a ubiquitous network of cellular timers coordinated by a hypothalamic master pacemaker. Circadian clock function is closely connected to the stress response system which has evolved to ensure survival under less predictable situations of danger. Disruptions in both of these functions are highly prevalent in modern society and have been linked to pathologic alterations in metabolic setpoints, promoting overeating, obesity, and type-2 diabetes. This paper describes the different levels of interaction between the circadian clock and acute and chronic stress responses. It summarizes studies assessing clock-stress crosstalk in the context of metabolic homeostasis and outlines options to use this interaction for diagnostic and therapeutic measures targeting metabolic health and well-being in the highly chronodisruptive environment of modern 24-h globalized societies.

Neurosecretory protein GL induces fat accumulation in mice.

Abstract: We recently discovered a novel gene encoding a small secretory protein, neurosecretory protein GL (NPGL), which stimulates feeding behavior in mice following acute administration. These findings suggest that dysregulation of NPGL contributes to obesity and metabolic disease. To explore this possibility, we investigated the impact of prolonged exposure to NPGL through 13 days of chronic intracerebroventricular (i.c.v.) infusion and examined feeding behavior, body composition, expressions of lipid metabolic factors, respiratory metabolism, locomotor activity, and food preference. Under standard chow diet, NPGL increased white adipose tissue (WAT) mass without affecting feeding behavior and body mass. In contrast, when fed a high-calorie diet, NPGL stimulated feeding behavior and increased body mass concomitant with marked fat accumulation. Quantitative reverse transcription polymerase chain reaction (RT-PCR) analysis revealed that mRNA expressions for key enzymes and related factors involved in lipid metabolism were increased in WAT and liver. Likewise, analyses of respiratory metabolism and locomotor activity revealed that energy expenditure and locomotor activity were significantly decreased by NPGL. In contrast, selective feeding of macronutrients did not alter food preference in response to NPGL, although total calorie intake was increased. Immunohistochemical analysis revealed that NPGL-containing cells produce galanin, a neuropeptide that stimulates food intake. Taken together, these results provide further support for NPGL as a novel regulator of fat deposition through changes in energy intake and locomotor activity.

Sucrose consumption alters steroid and dopamine signalling in the female rat brain.

Abstract: Sucrose consumption is associated with type 2 diabetes, cardiovascular disease, and cognitive deficits. Sucrose intake during pregnancy might have particularly prominent effects on metabolic, endocrine, and neural physiology. It remains unclear how consumption of sucrose affects parous females, especially in brain circuits that mediate food consumption and reward processing. Here, we examine whether a human-relevant level of sucrose before, during, and after pregnancy (17-18 weeks total) influences metabolic and neuroendocrine physiology in female rats. Females were fed either a control diet or a macronutrient-matched, isocaloric sucrose diet (25% of kcal from sucrose). Metabolically, sucrose impairs glucose tolerance, increases liver lipids, and increases a marker of adipose inflammation, but has no effect on body weight or overall visceral adiposity. Sucrose also decreases corticosterone levels in serum but not in the brain. Sucrose increases progesterone levels in serum and in the brain and increases the brain:serum ratio of progesterone in the mesocorticolimbic system and hypothalamus. These data suggest a dysregulation of systemic and local steroid signalling. Moreover, sucrose decreases tyrosine hydroxylase (TH), a catecholamine-synthetic enzyme, in the medial prefrontal cortex. Finally, sucrose consumption alters the expression pattern of FOSB, a marker of phasic dopamine signalling, in the nucleus accumbens. Overall, chronic consumption of sucrose at a human-relevant level alters metabolism, steroid levels, and brain dopamine signalling in a female rat model.

Estrogen accelerates heart regeneration by promoting the inflammatory response in zebrafish

Abstract: Sexual differences have been observed in the onset and prognosis of human cardiovascular diseases, but the underlying mechanisms are not clear. Here, we found that zebrafish heart regeneration is faster in females, can be accelerated by estrogen and is suppressed by the estrogen-antagonist tamoxifen. Injuries to the zebrafish heart, but not other tissues, increased plasma estrogen levels and the expression of estrogen receptors, especially esr2a. The resulting endocrine disruption induces the expression of the female-specific protein vitellogenin in male zebrafish. Transcriptomic analyses suggested heart injuries triggered pronounced immune and inflammatory responses in females. These responses, previously shown to elicit heart regeneration, could be enhanced by estrogen treatment in males and reduced by tamoxifen in females. Furthermore, a prior exposure to estrogen preconditioned the zebrafish heart for an accelerated regeneration. Altogether, this study reveals that heart regeneration is modulated by an estrogen-inducible inflammatory response to cardiac injury. These findings elucidate a previously unknown layer of control in zebrafish heart regeneration and provide a new model system for the study of sexual differences in human cardiac repair.

Gut-liver axis modulation in fructose-fed mice: a role for PPAR-alpha and linagliptin

Abstract: Fructose dietary intake affects the composition of the intestinal microbiota and influences the development of hepatic steatosis. Endotoxins produced by gram-negative bacteria alter intestinal permeability and cause bacterial translocation. This study evaluated the effects of gut microbiota modulation by a purified PPAR-alpha agonist (WY14643), a DPP-4 inhibitor (linagliptin), or their association on intestinal barrier integrity, endotoxemia, and hepatic energy metabolism in high-fructose-fed C57BL/6 mice. Fifty mice were divided to receive the control diet (C group) or the high-fructose diet (HFRU) for 12 weeks. Subsequently, the HFRU group was divided to initiate the treatment with PPAR-alpha agonist (3.5 mg/kg/BM) and DPP-4 inhibitor (15 mg/kg/BM). The HFRU group had glucose intolerance, endotoxemia, and dysbiosis (with increased Proteobacteria) without changes in body mass in comparison with the C group. HFRU group showed damaged intestinal ultrastructure, which led to liver inflammation and marked hepatic steatosis in the HFRU group when compared to the C group. PPAR-alpha activation and DPP-4 inhibition countered glucose intolerance, endotoxemia, and dysbiosis, ameliorating the ultrastructure of the intestinal barrier and reducing Tlr4 expression in the liver of treated animals. These beneficial effects suppressed lipogenesis and mitigated hepatic steatosis. In conclusion, the results herein propose a role for PPAR-alpha activation, DPP-4 inhibition, and their association in attenuating hepatic steatosis by gut-liver axis modulation in high-fructose mice model. These observations suggest these treatments as potential targets to treat hepatic steatosis and avoid its progression.

Effects of ERβ and ERα on OVX-induced changes in adiposity and insulin resistance.

Abstract: Loss of ovarian hormones leads to increased adiposity and insulin resistance (IR), increasing the risk for cardiovascular and metabolic diseases. The purpose of this study was to investigate whether the molecular mechanism behind the adverse systemic and adipose tissue-specific metabolic effects of ovariectomy requires loss of signaling through estrogen receptor alpha (ERα) or estrogen receptor β (ERβ). We examined ovariectomized (OVX) and ovary-intactwild-type (WT), ERα-null (αKO), and ERβ-null (βKO) female mice (age ~49 weeks; n = 7-12/group). All mice were fed a phytoestrogen-free diet (<15 mg/kg) and either remained ovary-intact (INT) or were OVX and followed for 12 weeks. Body composition, energy expenditure, glucose tolerance, and adipose tissue gene and protein expression were analyzed. INT αKO were ~25% fatter with reduced energy expenditure compared to age-matched INT WT controls and βKO mice (all P < 0.001). Following OVX, αKO mice did not increase adiposity or experience a further increase in IR, unlike WT and βKO, suggesting that loss of signaling through ERα mediates OVX-induced metabolic dysfunction. In fact, OVX in αKO mice (i.e., signaling through ERβ in the absence of ERα) resulted in reduced adiposity, adipocyte size, and IR (P < 0.05 for all). βKO mice responded adversely to OVX in terms of increased adiposity and development of IR. Together, these findings challenge the paradigm that ERα mediates metabolic protection over ERβ in all settings. These findings lead us to suggest that, following ovarian hormone loss, ERβ may mediate protective metabolic benefits.

Testosterone improves muscle fiber asset and exercise performance in a metabolic syndrome model.

Abstract: Lifestyle modifications, including physical exercise (PhyEx), are well-known treatments for metabolic syndrome (MetS), a cluster of metabolic and cardiovascular risk factors often associated to hypogonadism. Given the trophic role of testosterone on skeletal muscle (SkM), this study was aimed at evaluating the effects of testosterone treatment on SkM metabolism and exercise performance in male rabbits with high-fat diet (HFD)-induced MetS. HFD rabbits, treated or not with testosterone (30 mg/kg/week) for 12 weeks, were compared to regular diet animals (RD). A subset of each group was exercise-trained for 12 weeks. HFD increased type-II (fast, glycolytic) and decreased type-I (slow, oxidative) muscle fibers compared to RD as evaluated by RT-PCR and histochemistry. Testosterone reverted these effects, also inducing the expression of mitochondrial respiration enzymes and normalizing HFD-induced mitochondrial cristae reduction. Moreover, testosterone significantly increased the expression of myogenic/differentiation markers and genes related to glucidic/lipid metabolism. At the end of the PhyEx protocol, when compared to RD, HFD rabbits showed a significant reduction of running distance and running time, while testosterone counteracted this effect, also decreasing lactate production. In the trained groups, muscle histology showed a significant reduction of oxidative fibers in HFD compared to RD and the positive effect of testosterone in maintaining oxidative metabolism, as also demonstrated by analyzing mitochondrial ultrastructure, succinate dehydrogenase activity and ATP production. Our results indicate that testosterone could be useful to promote oxidative muscle metabolism altered by MetS, thus improving exercise performance. Conversely, testosterone administration to otherwise eugonadal rabbits (RD) only increased muscle fiber diameter but not endurance performance.

The P450 side-chain cleavage enzyme Cyp11a2 facilitates steroidogenesis in zebrafish

Abstract: Cytochrome P450 side-chain cleavage enzyme, encoded by the CYP11A1 gene, catalyzes the first and rate-limiting step of steroid hormone biosynthesis. Previous morpholino-knockdown studies in zebrafish suggested cyp11a2 is a functional equivalent of human CYP11A1 and is essential for interrenal steroidogenesis in zebrafish larvae. The role of Cyp11a2 in adult zebrafish, particularly in gonadal steroidogenesis, remains elusive. To explore the role of Cyp11a2 in adults, we developed zebrafish mutant lines by creating deletions in cyp11a2 using the CRISPR/Cas9 genomic engineering approach. Homozygous cyp11a2 mutant zebrafish larvae showed an upregulation of the hypothalamic-pituitary-interrenal axis. Furthermore, these Cyp11a2-deficient zebrafish demonstrated profound glucocorticoid and androgen deficiencies. Cyp11a2 homozygotes only developed into males with feminized secondary sex characteristics. Adult cyp11a2 -/- mutant fish showed a lack of natural breeding behaviors. Histological characterization revealed disorganized testicular structure and significantly decreased numbers of mature spermatozoa. These findings are further supported by the downregulation of the expression of several pro-male genes in the testes of cyp11a2 homozygous zebrafish, including sox9a, dmrt1 and amh. Moreover, the spermatogonia markers nanos2 and piwil1 were upregulated, while the spermatocytes marker sycp3 and spermatids marker odf3b were downregulated in the testes of cyp11a2 homozygous mutants. Our expression analysis is consistent with our histological studies, suggesting that spermatogonia are the predominant cell types in the testes of cyp11a2 homozygous mutants. Our work thus demonstrates the crucial role of Cyp11a2 in interrenal and gonadal steroidogenesis in zebrafish larvae and adults.

MicroRNA-200a improves diabetic endothelial dysfunction by targeting KEAP1/NRF2

Abstract: Over a half of the diabetic individuals develop macrovascular complications that cause high mortality. Oxidative stress (OS) promotes endothelial dysfunction (ED) which is a critical early step toward diabetic macrovascular complications. Nuclear factor erythroid 2-related factor 2 (NRF2) is a master regulator of cellular antioxidant defense system and combats diabetes-induced OS. Previously, we found that impaired NRF2 antioxidant signaling contributed to diabetes-induced endothelial OS and dysfunction in mice. The present study has investigated the effect of microRNA-200a (miR-200a) on NRF2 signaling and diabetic ED. In aortic endothelial cells (ECs) isolated from C57BL/6 wild-type (WT) mice, high glucose (HG) reduced miR-200a levels and increased the expression of kelch-like ECH-associated protein 1 (Keap1) - a target of miR-200a and a negative regulator of NRF2. This led to the inactivation of NRF2 signaling and exacerbation of OS and inflammation. miR-200a mimic (miR-200a-M) or inhibitor modulated KEAP1/NRF2 antioxidant signaling and manipulated OS and inflammation under HG conditions. These effects were completely abolished by knockdown of Keap1, indicating that Keap1 mRNA is a major target of miR-200a. Moreover, the protective effect of miR-200a-M was completely abrogated in aortic ECs isolated from C57BL/6 Nrf2 knockout (KO) mice, demonstrating that NRF2 is required for miR-200a's actions. In vivo, miR-200a-M inhibited aortic Keap1 expression, activated NRF2 signaling, and attenuated hyperglycemia-induced OS, inflammation and ED in the WT, but not Nrf2 KO, mice. Therefore, the present study has uncovered miR-200a/KEAP1/NRF2 signaling that controls aortic endothelial antioxidant capacity, which protects against diabetic ED.

Combination gut hormones: prospects and questions for the future of obesity and diabetes therapy

Abstract: Obesity represents an important public health challenge for the twenty-first century: globalised, highly prevalent and increasingly common with time, this condition is likely to reverse some of the hard-won gains in mortality accomplished in previous centuries. In the search for safe and effective therapies for obesity and its companion, type 2 diabetes mellitus (T2D), the gut hormone glucagon-like peptide-1 (GLP-1) has emerged as a forerunner and analogues thereof are now widely used in treatment of obesity and T2D, bringing proven benefits in improving glycaemia and weight loss and, notably, cardiovascular outcomes. However, GLP-1 alone is subject to limitations in terms of efficacy, and as a result, investigators are evaluating other gut hormones such as glucose-dependent insulinotropic peptide (GIP), glucagon and peptide YY (PYY) as possible partner hormones that may complement and enhance GLP-1's therapeutic effects. Such combination gut hormone therapies are in pharmaceutical development at present and are likely to make it to market within the next few years. This review examines the physiological basis for combination gut hormone therapy and presents the latest clinical results that underpin the excitement around these treatments. We also pose, however, some hard questions for the field which need to be answered before the full benefit of such treatments can be realised.

miRNA-mRNA profile and regulatory network in stearic acid-treated β-cell dysfunction.

Abstract: Chronic exposure of pancreatic β-cells to saturated fatty acid (palmitic or stearic acid) is a leading cause of impaired insulin secretion. However, the molecular mechanisms underlying stearic-acid-induced β-cell dysfunction remain poorly understood. Emerging evidence indicates that miRNAs are involved in various biological functions. The aim of this study was to explore the differential expression of miRNAs and mRNAs, specifically in stearic-acid-treated- relative to palmitic-acid-treated β-cells, and to establish their co-expression networks. β-TC-6 cells were treated with stearic acid, palmitic acid or normal medium for 24 h. Differentially expressed miRNAs and mRNAs were identified by high-throughput sequencing and bioinformatic analysis. Co-expression network, gene ontology (GO) and pathway analyses were then conducted. Changes in the expression of selected miRNAs and mRNAs were verified in β-TC-6 cells and mouse islets. Sequencing analysis detected 656 known and 1729 novel miRNAs. miRNA-mRNA network and Venn-diagram analysis yielded two differentially expressed miRNAs and 63 mRNAs exclusively in the stearic-acid group. miR-374c-5p was up-regulated by a 1.801 log2(fold-change) and miR-297b-5p was down-regulated by a -4.669 log2(fold-change). We found that miR-297b-5p and miR-374c-5p were involved in stearic-acid-induced lipotoxicity to β-TC-6 cells. Moreover, the effects of miR-297b-5p and miR-374c-5p on the alterations of candidate mRNAs expressions were verified. This study indicates that expression changes of specific miRNAs and mRNAs may contribute to stearic-acid-induced β-cell dysfunction, which provides a preliminary basis for further functional and molecular mechanism studies of stearic-acid-induced β-cell dysfunction in the development of type 2 diabetes.