Genetic deciphering of the antagonistic activities of the melanin-concentrating hormone and melanocortin pathways in skin pigmentation.

The genetic origin of human skin pigmentation remains an open question in biology. Several skin disorders and diseases originate from mutations in conserved pigmentation genes, including albinism, vitiligo, and melanoma. Teleosts possess the capacity to modify their pigmentation to adapt to their environmental background to avoid predators. This background adaptation occurs through melanosome aggregation (white background) or dispersion (black background) in melanocytes. These mechanisms are largely regulated by melanin-concentrating hormone (MCH) and α-melanocyte-stimulating hormone (α-MSH), two hypothalamic neuropeptides also involved in mammalian skin pigmentation. Despite evidence that the exogenous application of MCH peptides induces melanosome aggregation, it is not known if the MCH system is physiologically responsible for background adaptation. In zebrafish, we identify that MCH neurons target the pituitary gland-blood vessel portal and that endogenous MCH peptide expression regulates melanin concentration for background adaptation. We demonstrate that this effect is mediated by MCH receptor 2 (Mchr2) but not Mchr1a/b. mchr2 knock-out fish cannot adapt to a white background, providing the first genetic demonstration that MCH signaling is physiologically required to control skin pigmentation. mchr2 phenotype can be rescued in adult fish by knocking-out pomc, the gene coding for the precursor of α-MSH, demonstrating the relevance of the antagonistic activity between MCH and α-MSH in the control of melanosome organization. Interestingly, MCH receptor is also expressed in human melanocytes, thus a similar antagonistic activity regulating skin pigmentation may be conserved during evolution, and the dysregulation of these pathways is significant to our understanding of human skin disorders and cancers.

Transcriptomic analysis links diverse hypothalamic cell types to fibroblast growth factor 1-induced sustained diabetes remission.

In rodent models of type 2 diabetes (T2D), sustained remission of hyperglycemia can be induced by a single intracerebroventricular (icv) injection of fibroblast growth factor 1 (FGF1), and the mediobasal hypothalamus (MBH) was recently implicated as the brain area responsible for this effect. To better understand the cellular response to FGF1 in the MBH, we sequenced >79,000 single-cell transcriptomes from the hypothalamus of diabetic Lepob/ob mice obtained on Days 1 and 5 after icv injection of either FGF1 or vehicle. A wide range of transcriptional responses to FGF1 was observed across diverse hypothalamic cell types, with glial cell types responding much more robustly than neurons at both time points. Tanycytes and ependymal cells were the most FGF1-responsive cell type at Day 1, but astrocytes and oligodendrocyte lineage cells subsequently became more responsive. Based on histochemical and ultrastructural evidence of enhanced cell-cell interactions between astrocytes and Agrp neurons (key components of the melanocortin system), we performed a series of studies showing that intact melanocortin signaling is required for the sustained antidiabetic action of FGF1. These data collectively suggest that hypothalamic glial cells are leading targets for the effects of FGF1 and that sustained diabetes remission is dependent on intact melanocortin signaling.

FR - Nevus melanocíticos eruptivos asociados a medicamentos / Drug-Induced Eruptive Melanocytic Nevi

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The thermogenic effect of nesfatin-1 requires recruitment of the melanocortin system.

Nesfatin-1 is a bioactive polypeptide expressed both in the brain and peripheral tissues and involved in the control of energy balance by reducing food intake. Central administration of nesfatin-1 significantly increases energy expenditure, as demonstrated by a higher dry heat loss; yet, the mechanisms underlying the thermogenic effect of central nesfatin-1 remain unknown. Therefore, in this study, we sought to investigate whether the increase in energy expenditure induced by nesfatin-1 is mediated by the central melanocortin pathway, which was previously reported to mediate central nesfatin-1´s effects on feeding and numerous other physiological functions. With the application of direct calorimetry, we found that intracerebroventricular nesfatin-1 (25 pmol) treatment increased dry heat loss and that this effect was fully blocked by simultaneous administration of an equimolar dose of the melanocortin 3/4 receptor antagonist, SHU9119. Interestingly, the nesfatin-1-induced increase in dry heat loss was positively correlated with body weight loss. In addition, as assessed with thermal imaging, intracerebroventricular nesfatin-1 (100 pmol) increased interscapular brown adipose tissue (iBAT) as well as tail temperature, suggesting increased heat production in the iBAT and heat dissipation over the tail surface. Finally, nesfatin-1 upregulated pro-opiomelanocortin and melanocortin 3 receptor mRNA expression in the hypothalamus, accompanied by a significant increase in iodothyronine deiodinase 2 and by a nonsignificant increase in uncoupling protein 1 and peroxisome proliferator-activated receptor gamma coactivator-1 alpha mRNA in the iBAT. Overall, we clearly demonstrate that nesfatin-1 requires the activation of the central melanocortin system to increase iBAT thermogenesis and, in turn, overall energy expenditure.

Effects of melanocortin-4 receptor agonists and antagonists on expression of genes related to reproduction in spotted scat, Scatophagus argus.

Melanocortin-4 receptor (Mc4r) function related to reproduction in fish has not been extensively investigated. Here, we report on gene expression changes by real-time PCR following treatment with Mc4r agonists and antagonists in the spotted scat (Scatophagus argus). Using in vitro incubated hypothalamus, the Mc4r nonselective agonist NDP-MSH ([Nle4, D-Phe7]-α-melanocyte stimulating hormone; 10-6 M) and selective agonist THIQ (N-[(3R)-1, 2, 3, 4-Tetrahydroisoquinolinium-3-ylcarbonyl]- (1R)-1-(4-chlorobenzyl)-2-[4-cyclohexyl-4-(1H-1,2,4-triazol-1-ylmethyl) piperidin-1-yl]-2-oxoethylamine; 10-7 M) significantly increased the expression of gnrh (Gonadotropin releasing hormone), while the Mc4r nonselective antagonist SHU9119 (Ac-Nle-[Asp-His-DPhe/DNal(2')-Arg-Trp-Lys]-NH2; 10-6 M) and selective antagonist Ipsen 5i (compound 5i synthesized in Ipsen Research Laboratories; 10-6 M) significantly inhibited gnrh expression after 3 h of incubation. In incubated pituitary tissue, NDP-MSH and THIQ significantly increased the expression of fshb (Follicle-stimulating hormone beta subunit) and lhb (Luteinizing hormone beta subunit), while SHU9119 and Ipsen 5i significantly decreased fshb and lhb expression after 3 h of incubation. During the in vivo experiment, THIQ (1 mg/kg bw) significantly increased gnrh expression in hypothalamic tissue, as well as the fshb and lhb expression in pituitary tissue 12 h after abdominal injection. Furthermore, Ipsen 5i (1 mg/kg bw) significantly inhibited gnrh expression in hypothalamic tissue, as well as fshb and lhb gene expression in pituitary tissue 12 h after abdominal injection. In summary, Mc4r singling appears to stimulate gnrh expression in the hypothalamus, thereby modulating the synthesis of Fsh and Lh in the pituitary. In addition, Mc4r also appears to directly regulate fshb and lhb levels in the pituitary in spotted scat. Our study suggests that Mc4r, through the hypothalamus and pituitary, participates in reproductive regulation in fish.

D-TRP(8)-γMSH Prevents the Effects of Endotoxin in Rat Skeletal Muscle Cells through TNFα/NF-KB Signalling Pathway.

Sepsis induces anorexia and muscle wasting secondary to an increase in muscle proteolysis. Melanocyte stimulating hormones (MSH) is a family of peptides that have potent anti-inflammatory effects. Melanocortin receptor-3 (MC3-R) has been reported as the predominant anti-inflammatory receptor for melanocortins. The aim of this work was to analyse whether activation of MC3-R, by administration of its agonist D-Trp(8)-γMSH, is able to modify the response of skeletal muscle to inflammation induced by lipopolysaccharide endotoxin (LPS) or TNFα. Adult male rats were injected with 250 µg/kg LPS and/or 500 µg/kg D-Trp(8)-γMSH 17:00 h and at 8:00 h the following day, and euthanized 4 hours afterwards. D-Trp(8)-γMSH decreased LPS-induced anorexia and prevented the stimulatory effect of LPS on hypothalamic IL-1ß, COX-2 and CRH as well as on serum ACTH and corticosterone. Serum IGF-I and its expression in liver and gastrocnemius were decreased in rats injected with LPS, but not in those that also received D-Trp(8)-γMSH. However, D-Trp(8)-γMSH was unable to modify the effect of LPS on IGFBP-3. In the gastrocnemius D-Trp(8)-γMSH blocked LPS-induced decrease in pAkt, pmTOR, MHC I and MCH II, as well as the increase in pNF-κB(p65), FoxO1, FoxO3, LC3b, Bnip-3, Gabarap1, atrogin-1, MuRF1 and in LC3a/b lipidation. In L6 myotube cultures, D-Trp(8)-γMSH was able to prevent TNFα-induced increase of NF-κB(p65) phosphorylation and decrease of Akt phosphorylation as well as of IGF-I and MHC I expression. These data suggest that MC3-R activation prevents the effect of endotoxin on skeletal wasting by modifying inflammation, corticosterone and IGF-I responses and also by directly acting on muscle cells through the TNFα/NF-κB(p65) pathway.

Glucose Enhances Basal or Melanocortin-Induced cAMP-Response Element Activity in Hypothalamic Cells.

Melanocyte-stimulating hormone (MSH)-induced activation of the cAMP-response element (CRE) via the CRE-binding protein in hypothalamic cells promotes expression of TRH and thereby restricts food intake and increases energy expenditure. Glucose also induces central anorexigenic effects by acting on hypothalamic neurons, but the underlying mechanisms are not completely understood. It has been proposed that glucose activates the CRE-binding protein-regulated transcriptional coactivator 2 (CRTC-2) in hypothalamic neurons by inhibition of AMP-activated protein kinases (AMPKs), but whether glucose directly affects hypothalamic CRE activity has not yet been shown. Hence, we dissected effects of glucose on basal and MSH-induced CRE activation in terms of kinetics, affinity, and desensitization in murine, hypothalamic mHypoA-2/10-CRE cells that stably express a CRE-dependent reporter gene construct. Physiologically relevant increases in extracellular glucose enhanced basal or MSH-induced CRE-dependent gene transcription, whereas prolonged elevated glucose concentrations reduced the sensitivity of mHypoA-2/10-CRE cells towards glucose. Glucose also induced CRCT-2 translocation into the nucleus and the AMPK activator metformin decreased basal and glucose-induced CRE activity, suggesting a role for AMPK/CRTC-2 in glucose-induced CRE activation. Accordingly, small interfering RNA-induced down-regulation of CRTC-2 expression decreased glucose-induced CRE-dependent reporter activation. Of note, glucose also induced expression of TRH, suggesting that glucose might affect the hypothalamic-pituitary-thyroid axis via the regulation of hypothalamic CRE activity. These findings significantly advance our knowledge about the impact of glucose on hypothalamic signaling and suggest that TRH release might account for the central anorexigenic effects of glucose and could represent a new molecular link between hyperglycaemia and thyroid dysfunction.

Hypothalamic leptin action is mediated by histone deacetylase 5.

Hypothalamic leptin signalling has a key role in food intake and energy-balance control and is often impaired in obese individuals. Here we identify histone deacetylase 5 (HDAC5) as a regulator of leptin signalling and organismal energy balance. Global HDAC5 KO mice have increased food intake and greater diet-induced obesity when fed high-fat diet. Pharmacological and genetic inhibition of HDAC5 activity in the mediobasal hypothalamus increases food intake and modulates pathways implicated in leptin signalling. We show HDAC5 directly regulates STAT3 localization and transcriptional activity via reciprocal STAT3 deacetylation at Lys685 and phosphorylation at Tyr705. In vivo, leptin sensitivity is substantially impaired in HDAC5 loss-of-function mice. Hypothalamic HDAC5 overexpression improves leptin action and partially protects against HFD-induced leptin resistance and obesity. Overall, our data suggest that hypothalamic HDAC5 activity is a regulator of leptin signalling that adapts food intake and body weight to our dietary environment.

Central proopiomelanocortin but not neuropeptide Y mediates sympathoexcitation and hypertension in fat fed conscious rabbits.

OBJECTIVE: High-fat diet (HFD)-induced hypertension in rabbits is neurogenic because of the central sympathoexcitatory actions of leptin. Hypothalamic melanocortin and neuropeptide Y (NPY) neurons are recognized as the major signalling pathways through which leptin exerts its central effects. In this study, we assessed the effects of specific antagonists and agonists to melanocortin and NPY receptors on HFD-induced sympathoexcitation and hypertension. METHODS: Rabbits were instrumented with intracerebroventricular cannula, renal sympathetic nerve activity (RSNA) electrode, and blood pressure telemetry transmitter. RESULTS: After 3 weeks HFD (13.5% fat, n = 12) conscious rabbits had higher RSNA (+3.8  nu, P = 0.02), blood pressure (+8.6  mmHg, P < 0.001) and heart rate (+15  b/min, P = 0.01), and brain-derived neurotrophic factor levels in the hypothalamus compared with rabbits fed a control diet (4.2% fat, n = 11). Intracerebroventricular administration of the melanocortin receptor antagonist SHU9119 reduced RSNA (-2.7  nu) and blood pressure (-8.5  mmHg) in HFD but not control rabbits, thus reversing 100% of the hypertension and 70% of the sympathoexcitation induced by a HFD. By contrast, blocking central NPY Y1 receptors with BVD10 increased RSNA only in HFD rabbits. Intracerebroventricular α-melanocortin stimulating hormone increased RSNA and heart rate (P < 0.001) in HFD rabbits but had no effect in control rabbits. CONCLUSION: These findings suggest that obesity-induced hypertension and increased RSNA are dependent on the balance between greater activation of melanocortin signalling through melanocortin receptors and lesser activation of NPY sympathoinhibitory signalling. The amplification of the sympathoexcitatory effects of α-melanocortin stimulating hormone also indicates that the underlying mechanism is related to facilitation of leptin-melanocortin signalling, possibly involving chronic activation of brain-derived neurotrophic factor.

Hypothalamic Paraventricular and Arcuate Nuclei Contribute to Elevated Sympathetic Nerve Activity in Pregnant Rats: Roles of Neuropeptide Y and α-Melanocyte-Stimulating Hormone.

Pregnancy increases sympathetic nerve activity (SNA), but the mechanisms are unknown. Here, we investigated the contributions of the hypothalamic paraventricular and arcuate nuclei in α-chloralose-anesthetized pregnant and nonpregnant rats. Baseline arterial pressure (AP) was lower, and heart rate (HR), lumbar sympathetic activity, and splanchnic SNA were higher in pregnant rats compared with nonpregnant rats. Inhibition of the paraventricular nucleus via bilateral muscimol nanoinjections decreased AP and HR more in pregnant rats than in nonpregnant rats and decreased lumbar SNA only in pregnant rats. Similarly, after arcuate muscimol nanoninjections, the decreases in AP, HR, and lumbar, renal, and splanchnic sympathetic nerve activities were greater in pregnant rats than in nonpregnant rats. Major arcuate neuronal groups that project to the paraventricular nucleus express inhibitory neuropeptide Y (NPY) and excitatory α-melanocyte-stimulating hormone. Inhibition of paraventricular melanocortin 3/4 receptors with SHU9119 also decreased AP, HR, and lumbar SNA in pregnant rats but not in nonpregnant rats. Conversely, paraventricular nucleus NPY expression was reduced in pregnant animals, and although blockade of paraventricular NPY Y1 receptors increased AP, HR, and lumbar sympathetic activity in nonpregnant rats, it had no effects in pregnant rats. Yet, the sympathoinhibitory, depressor, and bradycardic effects of paraventricular NPY nanoinjections were similar between groups. In conclusion, the paraventricular and arcuate nuclei contribute to increased basal SNA during pregnancy, likely due in part to decreased tonic NPY inhibition and increased tonic α-melanocyte-stimulating hormone excitation of presympathetic neurons in the paraventricular nucleus.

Serine-727 phosphorylation activates hypothalamic STAT-3 independently from tyrosine-705 phosphorylation.

Transcriptional activity of signal transducer and activator of transcription-3 (STAT-3) is a key element in the central regulation of appetite and energy homeostasis. Activation of hypothalamic STAT-3 has been attributed to cytokine-promoted phosphorylation at tyrosine-705 (Tyr-705). In nonhypothalamic cells, STAT-3 is also phosphorylated at serine-727 (Ser-727), but the functional significance of Ser-727 in the regulation of hypothalamic STAT-3 is not known. We used 2 hypothalamic cell lines and analyzed the effects of various hormones on STAT-3-dependent reporter gene activity and observed that IFN-γ, epidermal growth factor (EGF), and bradykinin (BK) induce similar STAT-3 reporter activation. EGF and BK solely increased Ser-727 and IFN-γ increased Tyr-705 phosphorylation of STAT-3. Specific inhibition of ERK-1/2 activity blocked EGF- and BK-induced STAT-3 activation and Ser-727 phosphorylation. BK-induced ERK-1/2 activation occurred via EGF receptor transactivation. Consequently, the BK-mediated effects on STAT-3 were blocked by a specific EGF receptor antagonist. Next, we analyzed the effects of IFN-γ and EGF on the expression of the STAT-3-dependent genes thyroliberin-releasing hormone and suppressors of cytokine signaling-3. EGF but not IFN-γ enhanced thyroliberin-releasing hormone expression via STAT-3. With regard to suppressors of cytokine signaling-3, we observed prolonged expression induced by IFN-γ and a transient effect of EGF that required coactivation of the activator protein-1. Thus, EGF-promoted Ser-727 phosphorylation by ERK-1/2 is not only sufficient to fully activate hypothalamic STAT-3, but, in terms of targeted genes and required cofactors, entails distinct modes of STAT-3 actions compared with IFN-γ-induced Tyr-705 phosphorylation.

Cinnamomum cassia essential oil inhibits α-MSH-induced melanin production and oxidative stress in murine B16 melanoma cells.

Essential oils extracted from aromatic plants exhibit important biological activities and have become increasingly important for the development of aromatherapy for complementary and alternative medicine. The essential oil extracted from Cinnamomum cassia Presl (CC-EO) has various functional properties; however, little information is available regarding its anti-tyrosinase and anti-melanogenic activities. In this study, 16 compounds in the CC-EO have been identified; the major components of this oil are cis-2-methoxycinnamic acid (43.06%) and cinnamaldehyde (42.37%). CC-EO and cinnamaldehyde exhibited anti-tyrosinase activities; however, cis-2-methoxycinnamic acid did not demonstrate tyrosinase inhibitory activity. In murine B16 melanoma cells stimulated with α-melanocyte-stimulating hormone (α-MSH), CC-EO and cinnamaldehyde not only reduced the melanin content and tyrosinase activity of the cells but also down-regulated tyrosinase expression without exhibiting cytotoxicity. Moreover, CC-EO and cinnamaldehyde decreased thiobarbituric acid-reactive substance (TBARS) levels and restored glutathione (GSH) and catalase activity in the α-MSH-stimulated B16 cells. These results demonstrate that CC-EO and its major component, cinnamaldehyde, possess potent anti-tyrosinase and anti-melanogenic activities that are coupled with antioxidant properties. Therefore, CC-EO may be a good source of skin-whitening agents and may have potential as an antioxidant in the future development of complementary and alternative medicine-based aromatherapy.

Conditional viral tracing reveals that steroidogenic factor 1-positive neurons of the dorsomedial subdivision of the ventromedial hypothalamus project to autonomic centers of the hypothalamus and hindbrain.

Excitation of neurons in the ventromedial hypothalamus (VMH), especially those residing in the dorsomedial part of the nucleus (VMHdm), evokes sympathetic nervous system (SNS) outflow, modulating a number of physiological functions including feeding and blood glucose homeostasis. However, the anatomical basis of VMH-mediated SNS activation has thus far proved elusive. To understand how VMH neurons exercise output functions and describe an anatomical link between these neurons and the SNS, we identified downstream neural targets of the VMHdm by injecting an adenoviral vector encoding Cre recombinase (Cre)-regulated farnesylated green fluorescent protein (GFPf ) into the VMHdm of mice that express Cre in neurons expressing the VMH-specific transcription factor steroidogenic factor 1 (SF1). We confirm previously described projection patterns of the VMHdm and report the existence of a formerly unidentified projection pathway to a number of autonomic centers in the brainstem. These VMH efferents travel caudally through the periaqueductal gray (PAG) and then ventrally through the lateral lemniscus to the ventral surface of the brain, where they eventually reach caudal autonomic centers including the C1 catecholamine cell group of the rostral ventrolateral medulla (RVLM) and the nucleus of the solitary tract (NTS), where VMH efferents make close contacts with catecholaminergic neurons. We also found that VMHdm fibers reach a number of brainstem areas, including the retrotrapezoid nucleus (RTN), which are important in regulating respiration. Thus, the present study indicates that the VMH may modulate sympathetic and autonomic activity via synaptic contacts in the RTN, NTS, and RVLM and provides significant anatomical evidence to support a role of the VMH in respiratory regulation.

Antioxidative and melanogenesis-inhibitory activities of caffeoylquinic acids and other compounds from moxa.

The MeOH extract of moxa, the processed leaves of Artemisia princeps PAMP. (Asteraceae), exhibited potent 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activity and melanogenesis-inhibitory activity in α-melanocyte-stimulating hormone (α-MSH)-stimulated B16 melanoma cells. Eight caffeoylquinic acids, 1 and 6-12, five flavonoids, 13-17, two benzoic acid derivatives, 18 and 19, three coumarin derivatives, 20-22, four steroids, 23-26, and six triterpenoids, 27-32, were isolated from the MeOH extract. Upon evaluation of compounds 1, 6-23, and four semisynthetic caffeoylquinic acid esters, 2-5, for their DPPH radical-scavenging activity, 15 compounds, 1-13, 17, and 19, showed potent activities (IC(50) 3.1-16.8 µM). The 15 compounds exhibited, moreover, potent inhibitory activities (51.1-92.5% inhibition) against peroxidation of linoleic acid emulsion at 10 µg/ml concentration. In addition, when 27 compounds, 1-8, 10, 12, 13, 15-18, 20-25, and 27-32, were evaluated for their inhibitory activity against melanogenesis in α-MSH-stimulated B16 melanoma cells, five caffeoylquinic acids, i.e., chlorogenic acid (1), ethyl chlorogenate (3), propyl chlorogenate (4), isopropyl chlorogenate (5), and butyl chlorogenate (6), along with homoorientin (17) and vanillic acid (18), exhibited inhibitory activities with 33-62% reduction of melanin content at 100 µM concentration with no or almost no toxicity to the cells (89-114% of cell viability at 100 µM). Western blot analysis showed that compound 6 reduced the protein levels of microphtalmia-associated transcription factor (MITF), tyrosinase, tyrosine-related protein 1 (TRP-1), and TRP-2 mostly in a concentration-dependent manner, suggesting that this compound inhibits melanogenesis on α-MSH-stimulated B16 melanoma cells by, at least in part, inhibiting the expression of MITF, followed by decreasing the expression of tyrosinase, TRP-1, and TRP-2. Furthermore, four compounds, 13, 15, 16, and 30, exhibited cytotoxicities against HL60 human leukemia cell line (IC(50) 7.0-11.1 µM), and nine compounds, 14-16, 23, 26-28, 31, and 32, showed inhibitory effects (IC(50) 272-382 mol ratio/32 pmol 12-O-tetradecanoylphohrbol-13-acetate (TPA)) against Epstein-Barr virus early antigen (EBV-EA) activation induced by TPA in Raji cells.

Intracerebroventricular administration of C-type natriuretic peptide suppresses food intake via activation of the melanocortin system in mice.

C-type natriuretic peptide (CNP) and its receptor are abundantly distributed in the brain, especially in the arcuate nucleus (ARC) of the hypothalamus associated with regulating energy homeostasis. To elucidate the possible involvement of CNP in energy regulation, we examined the effects of intracerebroventricular administration of CNP on food intake in mice. The intracerebroventricular administration of CNP-22 and CNP-53 significantly suppressed food intake on 4-h refeeding after 48-h fasting. Next, intracerebroventricular administration of CNP-22 and CNP-53 significantly decreased nocturnal food intake. The increment of food intake induced by neuropeptide Y and ghrelin was markedly suppressed by intracerebroventricular administration of CNP-22 and CNP-53. When SHU9119, an antagonist for melanocortin-3 and melanocortin-4 receptors, was coadministered with CNP-53, the suppressive effect of CNP-53 on refeeding after 48-h fasting was significantly attenuated by SHU9119. Immunohistochemical analysis revealed that intracerebroventricular administration of CNP-53 markedly increased the number of c-Fos-positive cells in the ARC, paraventricular nucleus, dorsomedial hypothalamus, ventromedial hypothalamic nucleus, and lateral hypothalamus. In particular, c-Fos-positive cells in the ARC after intracerebroventricular administration of CNP-53 were coexpressed with α-melanocyte-stimulating hormone immunoreactivity. These results indicated that intracerebroventricular administration of CNP induces an anorexigenic action, in part, via activation of the melanocortin system.

ß-Cryptoxanthin suppresses UVB-induced melanogenesis in mouse: involvement of the inhibition of prostaglandin E2 and melanocyte-stimulating hormone pathways.

OBJECTIVE: ß-cryptoxanthin (ß-CPX) is a carotenoid that is widely contained in the fruits of citrus plants. We evaluated the effect of ß-CPX on UVB-induced pigmentation and mRNA expression related to melanogenesis in mouse skin. In addition, changes in melanogenic molecules were evaluated in cultured melanocytes stimulated with prostaglandin (PG) E(2), melanocyte-stimulating hormone (MSH) and endothelin (ET)-1. METHODS: Mice were irradiated with UVB and were given ß-CPX (0.1, 1 and 10 mg/kg) orally for 14 days. Pigmentation was evaluated by skin colour change and microscopic observation. Total RNA was obtained from the skin and the expression of melanogenic mRNA was evaluated by RT-PCR. In cell culture studies, human melanocytes were cultured with ß-CPX and melanogenic stimulants (PGE(2), MSH and ET-1) for 6-10 days. Melanin contents, dendricity, melanogenic mRNA and phosphorylation of cyclic AMP response element-binding protein (CREB) were evaluated. KEY FINDINGS: ß-CPX (10 mg/kg) significantly suppressed skin pigmentation and mRNA expression of cyclooxygenase-2, ET-1 receptors, low-affinity neurotrophin receptor, PGE(2) receptor (EP1), melanocortin 1 receptor (MC1R), tyrosinase (Tyr), tyrosinase-related protein (Tyrp) 1 and microphthalmia transcription factor. ß-CPX (10 µg/ml) suppressed melanogenesis induced by PGE(2), MSH and ET-1. In the PGE(2)-stimulated melanocytes, mRNA expressions of EP-1, Tyr and Tyrp1 and phosphorylation of CREB protein were suppressed. In the ET-1-stimulated cells, only expression of CREB protein was suppressed. In the MSH-induced cells, mRNA expression of MC1R and Tyrp1 and protein expression of CREB were suppressed. CONCLUSION: Oral administration of ß-CPX was found to suppress UVB-induced melanogenesis. Suppression of melanogenic enzymes, receptors of melanogenic stimulators, expression and phosphorylation of CREB are thought to be involved in the mechanism.

The neuroendocrine circuitry controlled by POMC, MSH, and AGRP.

Obesity is one of the most challenging health problems worldwide. Over the past few decades, our knowledge concerning mechanisms of weight regulation has increased tremendously leading to the identification of the leptin-melanocortin pathway. The filling level of energy stores is signaled to the brain, and the information is integrated by hypothalamic nuclei, resulting in a well-orchestrated response to food intake and energy expenditure to ensure constant body weight. One of the key players in this system is proopiomelanocortin (POMC), a precursor of a variety of neuropeptides. POMC-derived alpha- and beta-MSH play an important role in energy homeostasis by activating melanocortin receptors expressed in the arcuate nucleus (MC3R) and in the nucleus paraventricularis (MC4R). Activation of these two G protein-coupled receptors is antagonized by agouti-related peptide (AgRP). Naturally occurring mutations in this system were identified in patients suffering from common obesity as well as in patients demonstrating a phenotype of severe early-onset obesity, adrenal insufficiency, red hair, and pale skin. Detailed understanding of the complex system of POMC-AgRP-MC3R-MC4R and their interaction with other hypothalamic as well as peripheral signals is a prerequisite to combat the obesity epidemic.

Cannabinoid CB1 receptor restrains accentuated activity of hypothalamic corticotropin-releasing factor and brainstem tyrosine hydroxylase neurons in endotoxemia-induced hypophagia in rats.

It is well known that endocannabinoids play an important role in the regulation of food intake and body weight. Endocannabinoids and cannabinoid receptors are found in the hypothalamus and brainstem, which are central areas involved in the control of food intake and energy expenditure. Activation of these areas is related to hypophagia observed during inflammatory stimulus. This study investigated the effects of cannabinoid (CB1) receptor blockade on lipopolysaccharide (LPS)-induced hypophagia. Male Wistar rats were pretreated with rimonabant (10 mg/kg, by gavage) or vehicle; 30 min later they received an injection of either LPS (100 µg/kg, intraperitoneal) or saline. Food intake, body weight, corticosterone response, CRF and CART mRNA expression, Fos-CRF and Fos-α-MSH immunoreactivity in the hypothalamus and Fos-tyrosine hydroxylase (TH) immunoreactivity in the brainstem were evaluated. LPS administration decreased food intake and body weight gain and increased plasma corticosterone levels and CRF mRNA expression in the PVN. We also observed an increase in Fos-CRF and Fos-TH double-labeled neurons after LPS injection in vehicle-pretreated rats, with no changes in CART mRNA or Fos-α-MSH immunoreactive neurons in the ARC. In saline-treated animals, rimonabant pretreatment decreased food intake and body weight gain but did not modify hormone response or Fos expression in the hypothalamus and brainstem compared with vehicle-pretreated rats. Rimonabant pretreatment potentiated LPS-induced hypophagia, body weight loss and Fos-CRF and Fos-TH expressing neurons. Rimonabant did not modify corticosterone, CRF mRNA or Fos-α-MSH responses in rats treated with LPS. These data suggest that the endocannabinoid system, mediated by CB1 receptors, modulates hypothalamic and brainstem circuitry underlying the hypophagic effect during endotoxemia to prevent an exaggerated food intake decrease. This article is part of a Special Issue entitled 'Central Control of Food Intake'.

Molecular engineering of short half-life small peptides (VIP, αMSH and γ3MSH) fused to latency-associated peptide results in improved anti-inflammatory therapeutics.

OBJECTIVE: To facilitate the targeting to inflammation sites of small anti-inflammatory peptides, with short half-lives, by fusion with the latency-associated peptide (LAP) of transforming growth factor ß1 through a cleavable matrix metalloproteinase (MMP) linker. This design improves efficacy, overcoming the limitations to their clinical use. METHODS: We generated latent forms of vasoactive intestinal peptide (VIP), α-melanocyte-stimulating hormone (MSH) and γ(3)MSH by fusion to LAP through an MMP cleavage site using recombinant DNA technology. The biological activities of these latent therapeutics were studied in vivo using monosodium urate (MSU)-induced peritonitis and collagen-induced arthritis (CIA) models. We assessed gene therapy and purified protein therapy. RESULTS: The recruitment of the polymorphonuclear cells induced by MSU injection into mouse peritoneal cavity was reduced by 35% with γ(3)MSH (1 nmol), whereas administration of a much lower dose of purified latent LAP-MMP-γ(3)MSH (0.03 nmol) attenuated leucocyte influx by 50%. Intramuscular gene delivery of plasmids coding LAP-MMP-VIP and LAP-MMP-αMSH at disease onset reduced the development of CIA compared with LAP-MMP, which does not contain any therapeutic moiety. Histological analysis confirmed a significantly lower degree of inflammation, bone and cartilage erosion in groups treated with LAP-MMP-VIP or LAP-MMP-αMSH. Antibody titres to collagen type II and inflammatory cytokine production were also reduced in these two groups. CONCLUSION: Incorporation of small anti-inflammatory peptides within the LAP shell and delivered as recombinant protein or through gene therapy can control inflammatory and arthritic disease. This platform delivery can be developed to control human arthritides and other autoimmune diseases.

The HPA axis modulates the CNS melanocortin control of liver triacylglyceride metabolism.

The central melanocortin system regulates lipid metabolism in peripheral tissues such as white adipose tissue. Alterations in the activity of sympathetic nerves connecting hypothalamic cells expressing melanocortin 3/4 receptors (MC3/4R) with white adipocytes have been shown to partly mediate these effects. Interestingly, hypothalamic neurons producing corticotropin-releasing hormone and thyrotropin-releasing hormone co-express MC4R. Therefore we hypothesized that regulation of hypothalamo-pituitary adrenal (HPA) and hypothalamo-pituitary thyroid (HPT) axes activity by the central melanocortin system could contribute to its control of peripheral lipid metabolism. To test this hypothesis, we chronically infused rats intracerebroventricularly (i.c.v.) either with an MC3/4R antagonist (SHU9119), an MC3/4R agonist (MTII) or saline. Rats had been previously adrenalectomized (ADX) and supplemented daily with 1mg/kg corticosterone (s.c.), thyroidectomized (TDX) and supplemented daily with 10 µg/kgL-thyroxin (s.c.), or sham operated (SO). Blockade of MC3/4R signaling with SHU9119 increased food intake and body mass, irrespective of gland surgery. The increase in body mass was accompanied by higher epididymal white adipose tissue (eWAT) weight and higher mRNA content of lipogenic enzymes in eWAT. SHU9119 infusion increased triglyceride content in the liver of SO and TDX rats, but not in those of ADX rats. Concomitantly, mRNA expression of lipogenic enzymes in liver was increased in SO and TDX, but not in ADX rats. We conclude that the HPA and HPT axes do not play an essential role in mediating central melanocortinergic effects on white adipose tissue and liver lipid metabolism. However, while basal hepatic lipid metabolism does not depend on a functional HPA axis, the induction of hepatic lipogenesis due to central melanocortin system blockade does require a functional HPA axis.