u-0126 and Body-Weight

Fat mass and obesity-associated (FTO) gene is strongly associated with obesity which brings a major health threat. Altered expression of its encoded protein FTO in the hypothalamus has been identified to contribute to central control of appetite and body weight. However, its molecular mechanisms remain elusive.. Mouse hypothalamic POMC cell line N43/5 was treated with FTO inhibitor rhein, FTO shRNA, or extracellular signal-regulated kinase 1/2 (ERK1/2) inhibitor U0126 to inhibit FTO or ERK1/2. Rhein and U0126 were injected into lateral ventricle of the mice by intracerebroventricular cannulation. Western blotting and immunofluorescent assays were performed to monitor protein level.. This study identified that inhibition of FTO in N43/5 cells led to phosphorylation of signal transducer and activator of transcription 3 (STAT3) at S727 site and induced p-STAT3-S727 nuclear translocation. We further showed that FTO inhibition promoted phosphorylation of ERK1/2; specific inhibition of ERK1/2 signaling by U0126 could abolish the effect of FTO inhibition on STAT3-S727 phosphorylation and nuclear translocation. Furthermore, we found that inhibition of hypothalamic FTO promoted STAT3-S727 phosphorylation in the hypothalamic arcuate nucleus, and the mice showed reductions in food intake and body weight. In addition, inhibition of hypothalamic ERK1/2 could abolish the effects of FTO inhibition on STAT3-S727 phosphorylation, reductions of food intake and body weight.. Our in vitro and in vivo data suggest that the inhibition of hypothalamic FTO could activate STAT3 through ERK1/2, which is potentially associated with reductions in food intake and body weight.

Topics: Alpha-Ketoglutarate-Dependent Dioxygenase FTO; Animals; Body Weight; Eating; Hypothalamus; Leptin; MAP Kinase Signaling System; Mice; Mitogen-Activated Protein Kinase 3; Obesity; Phosphorylation; STAT3 Transcription Factor

Myricetin has been reported to promote osteogenic differentiation and inhibit osteoclastogenesis. The aim of this study was to investigate the potential effects of myricetin on glucocorticoid-induced osteoporosis (GIOP) both in vivo and in vitro. Male Sprague-Dawley (SD) rats were given dexamethasone (DEX, 0.1 mg/kg, s.c.) once daily and myricetin (1 mg/kg or 2.5 mg/kg, i.p.) once every other day for a total of five weeks. Body weight was recorded once a week. Bone mineral density (BMD), the activities or levels of bone turnover markers, and histological changes were assessed. MC3T3-E1 cells were incubated with DEX (1 μM) and myricetin (20 μM). Osteoblast proliferation, differentiation and mineralization were evaluated. U0126 was added to evaluate the involvement of the ERK signaling pathway. The results showed that myricetin increased body weight gain and inhibited DEX-induced reduction in BMD, enhanced alkaline phosphatase (ALP) activity, and upregulated osteocalcin (OCN), bone morphogenetic protein 2 (BMP2) and runt-related transcription factor 2 (Runx2) levels, whereas reduced tartrate-resistant acid phosphatase (TRAP) activity and C-terminal telopeptide of type I collagen (CTx) level. In addition, myricetin ameliorated histological changes in the femurs. In our in vitro studies, myricetin promoted osteoblast differentiation and mineralization in DEX-treated MC3T3-E1 cells, accompanied by increases in BMP2, Runx2, ALP, OCN, collagen type I alpha 1 (COL1A1) and osteopontin (OPN) levels. The promotion effects of myricetin on osteogenic differentiation and matrix mineralization were reversed by U0126. These results suggest that myricetin may alleviate DEX-induced osteoporosis by promoting osteogenic differentiation and matrix mineralization via the ERK signaling pathway.

Topics: 3T3 Cells; Alkaline Phosphatase; Animals; Body Weight; Bone Morphogenetic Protein 2; Butadienes; Cell Differentiation; Cell Proliferation; Core Binding Factor Alpha 1 Subunit; Extracellular Signal-Regulated MAP Kinases; Flavonoids; Glucocorticoids; Male; MAP Kinase Signaling System; Mice; Nitriles; Osteoblasts; Osteocalcin; Osteogenesis; Osteoporosis; Rats; Rats, Sprague-Dawley; Signal Transduction

To explore the mechanisms underlying endothelin-1 (ET-1) elevations induced by excessive fluoride exposure.. We measured serum and bone fluoride ion content and plasma ET-1 levels and compared these parameters among different groups in an animal model. We also observed morphological changes in the aorta and endothelium of rabbits. In cell experiments, human umbilical vein endothelial cells (HUVECs) were treated with varying concentrations of NaF for 24h, with or without 10 µM U0126 pretreatment for 1 h. ET-1 levels in culture fluid and intracellular reactive oxygen species (ROS) levels, as well as ET1 gene, endothelin-converting enzyme-1 (ECE-1), extracellular signal-regulating kinase 1/2 (ERK1/2), pERK1/2 expression levels and RAS activation were measured and compared among the groups.. Plasma ET-1 levels of rabbits increased significantly in fluorinated groups compared with those in the control group. The rabbit thoracic aortas became slightly hardened in fluorinated groups compared with those in the control group, and some vacuoles were present in the endothelial cell cytoplasm of the rabbits in fluorinated groups. In our cell experiments, ET1 gene and ECE-1 expression levels in HUVECs and ET-1 expression levels in the cell culture supernatants increased significantly in some experimental groups compared with those in the control group. These trends paralleled the changes in intracellular ROS levels, RAS activation, and the pERK1/2-to-ERK1/2 ratio. After U0126 was added, ECE-1 expression and ET-1 levels decreased significantly.. Excessive fluoride exposure leads to characteristic endothelial damage (vacuoles), thoracic aorta hardening, and plasma ET-1 level elevations in rabbits. In addition, the ROS-RAS-MEK1/2-pERK1/2/ERK1/2 pathway plays a crucial-and at least partial-role in ET-1 over-expression, which is promoted by excessive fluoride exposure.

Topics: Animals; Aorta; Body Weight; Butadienes; Cell Proliferation; Diet; Drinking Water; Endothelin-1; Endothelin-Converting Enzymes; Endothelium, Vascular; Extracellular Signal-Regulated MAP Kinases; Fluorides; Gene Expression Regulation; Human Umbilical Vein Endothelial Cells; Intracellular Space; Ions; Male; Nitriles; Phosphorylation; Rabbits; ras Proteins; Reactive Oxygen Species; Vacuoles

Leptin is a key regulator of glucose metabolism in mammals, but the mechanisms of its action have remained elusive. We now show that signaling by extracellular signal-regulated kinase (ERK) and its upstream kinase MEK in the ventromedial hypothalamus (VMH) mediates the leptin-induced increase in glucose utilization as well as its insulin sensitivity in the whole body and in red-type skeletal muscle of mice through activation of the melanocortin receptor (MCR) in the VMH. In contrast, activation of signal transducer and activator of transcription 3 (STAT3), but not the MEK-ERK pathway, in the VMH by leptin enhances the insulin-induced suppression of endogenous glucose production in an MCR-independent manner, with this effect of leptin occurring only in the presence of an increased plasma concentration of insulin. Given that leptin requires 6 h to increase muscle glucose uptake, the transient activation of the MEK-ERK pathway in the VMH by leptin may play a role in the induction of synaptic plasticity in the VMH, resulting in the enhancement of MCR signaling in the nucleus and leading to an increase in insulin sensitivity in red-type muscle.

Topics: Animals; Body Weight; Butadienes; Chromones; Enzyme Inhibitors; Extracellular Signal-Regulated MAP Kinases; Glucose; Insulin; Leptin; Male; Melanocyte-Stimulating Hormones; Mice; Morpholines; Muscle, Skeletal; Nitriles; Phosphorylation; Receptors, Corticotropin; Signal Transduction; Ventromedial Hypothalamic Nucleus

Glucagon-like peptide-1 receptor (GLP-1R) activation within the nucleus tractus solitarius (NTS) suppresses food intake and body weight (BW), but the intracellular signals mediating these effects are unknown. Here, hindbrain (fourth i.c.v.) GLP-1R activation by Exendin-4 (Ex-4) increased PKA and MAPK activity and decreased phosphorylation of AMPK in NTS. PKA and MAPK signaling contribute to food intake and BW suppression by Ex-4, as inhibitors RpcAMP and U0126 (fourth i.c.v.), respectively, attenuated Ex-4's effects. Hindbrain GLP-1R activation inhibited feeding by reducing meal number, not meal size. This effect was attenuated with stimulation of AMPK activity by AICAR (fourth i.c.v.). The PKA, MAPK, and AMPK signaling responses by Ex-4 were present in immortalized GLP-1R-expressing neurons (GT1-7). In conclusion, hindbrain GLP-1R activation suppresses food intake and BW through coordinated PKA-mediated suppression of AMPK and activation of MAPK. Pharmacotherapies targeting these signaling pathways, which mediate intake-suppressive effects of CNS GLP-1R activation, may prove efficacious in treating obesity.

Topics: AMP-Activated Protein Kinase Kinases; Animals; Body Weight; Butadienes; Cyclic AMP-Dependent Protein Kinases; Eating; Exenatide; Glucagon-Like Peptide-1 Receptor; Male; Mitogen-Activated Protein Kinase Kinases; Nitriles; Peptides; Phosphorylation; Protein Kinases; Rats; Rats, Sprague-Dawley; Receptors, Glucagon; Signal Transduction; Solitary Nucleus; Venoms

The extracellular signal-regulated kinase (ERK) cascade has long been known to be central to the activation of cellular processes such as proliferation, differentiation, and oncogenic transformation. The mitogen-activated protein (MAP) serine/threonine family of protein kinases, of which ERK is a member, is activated by a mechanism that includes protein kinase cascades. Mitogen-activated protein kinases (MAPKs) are well-conserved enzymes connecting cell surface receptors to intracellular regulatory targets; they are activated in response to a wide variety of stimuli. The aim of this study was to investigate the effects of PD98059, a highly selective inhibitor of MAP/ERK kinase1 (MEK1) activation, on the development of lung inflammation and fibrosis. Lung injury was induced by intratracheal instillation of bleomycin (1 mg/kg), and PD98059 (10 mg/kg, 10% dimethyl sulfoxide, i.p.) was administrated 1 h after bleomycin instillation and daily for 7 days. PD98059 treatment shows therapeutic effects on pulmonary damage, decreasing many inflammatory and apoptotic parameters, such as (1) cytokine production; (2) IkBα degradation and NF-kB nuclear translocation; (3) iNOS expression; (4) nitrotyrosine and PAR localization; and (5) the degree of apoptosis, as evaluated by Bax and Bcl-2 balance, FAS ligand expression, and terminal deoxynucleotidyl transferase dUTP nick-end labeling staining. In particular, to assess whether PD98059 treatment influences MAPKs pathway, we have also investigated the expression of activated ERK and JNK after bleomycin-induced pulmonary fibrosis, showing that the inhibition of the cascade reduces the inflammatory processes that lead to the appearance of the fibrosis. Taken together, all our results clearly show that PD98059 reduces the lung injury and inflammation due to the intratracheal bleomycin administration in mice.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Bleomycin; Body Weight; Butadienes; Fas Ligand Protein; Flavonoids; I-kappa B Proteins; Instillation, Drug; Interleukin-1beta; JNK Mitogen-Activated Protein Kinases; Lung; Male; MAP Kinase Kinase 1; Mice; Mice, Inbred Strains; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinase Kinases; Neutrophils; NF-KappaB Inhibitor alpha; Nitric Oxide Synthase Type II; Nitriles; Phosphorylation; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerases; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-bcl-2; Pulmonary Edema; Pulmonary Fibrosis; Transcription Factor RelA; Tumor Necrosis Factor-alpha; Tyrosine

In response to nutrient stimuli, the mediobasal hypothalamus (MBH) drives multiple neuroendocrine and behavioral mechanisms to regulate energy balance. While central leucine reduces food intake and body weight, the specific neuroanatomical sites of leucine sensing, downstream neural substrates, and neurochemical effectors involved in this regulation remain largely unknown. Here we demonstrate that MBH leucine engages a neural energy regulatory circuit by stimulating POMC (proopiomelanocortin) neurons of the MBH, oxytocin neurons of the paraventricular hypothalamus, and neurons within the brainstem nucleus of the solitary tract to acutely suppress food intake by reducing meal size. We identify central p70 S6 kinase and Erk1/2 pathways as intracellular effectors required for this response. Activation of endogenous leucine intracellular metabolism produced longer-term reductions in meal number. Our data identify a novel, specific hypothalamus-brainstem circuit that links amino acid availability and nutrient sensing to the control of food intake.

Topics: Animals; Anorexia; Body Weight; Bone Morphogenetic Protein Receptors, Type I; Brain Stem; Butadienes; Dose-Response Relationship, Drug; Drug Administration Schedule; Eating; Enzyme Inhibitors; Feeding Behavior; Green Fluorescent Proteins; Hypothalamus; In Vitro Techniques; Injections, Intraventricular; Keto Acids; Leucine; Male; Melanocortins; Melanocyte-Stimulating Hormones; Mice; Mice, Inbred C57BL; Mice, Transgenic; Neural Pathways; Neurons; Nitriles; Oxytocin; Pro-Opiomelanocortin; Proto-Oncogene Proteins c-fos; Rats; Rats, Sprague-Dawley; Signal Transduction; Threonine; Time Factors; Tyrosine; Vasotocin

The extracellular signal-regulated kinase (ERK) pathway, which modulates the activity of many transcriptional factors leading to the proliferation of various cells, is activated in lesions in regions of selective vulnerability after traumatic brain injury (TBI). In the present study, using the ERK inhibitor U0126, we investigated the role of the ERK pathway in histopathological and behavioral outcomes after TBI. Adult male Sprague-Dawley rats, weighing 300-400 g were subjected to lateral fluid percussion brain injury. The ERK inhibitor U0126 was injected intravenously before injury at 100, 200 and 400 microg/kg. The severity of CA3 neuronal damage was evaluated by the number of surviving CA3 neurons 7 days after injury. The contusional lesion volume 72 h after injury was analysed using a computer-assisted analysis system. Three different motor skill tasks were measured on days 1-5, 7, 14 and 21 after injury. Pretreatment with U0126 significantly reduced both CA3 neuronal damage and contusional lesion volume after injury. In addition, administration of U0126 ameliorated motor function recovery on days 3, 4 and 5 after injury. Therefore, inhibition of ERK phosphorylation could be a potentially effective therapeutic target after TBI.

Topics: Animals; Behavior, Animal; Blotting, Western; Body Weight; Brain; Brain Injuries; Butadienes; Cell Count; Cell Survival; Cerebral Cortex; Enzyme Inhibitors; Extracellular Signal-Regulated MAP Kinases; Hippocampus; Immunohistochemistry; Male; Neurons; Nitriles; Postural Balance; Rats; Rats, Sprague-Dawley