Raspberry ketone promotes FNDC5 protein expression via HO-1 upregulation in 3T3-L1 adipocytes.

Obesity is a global health problem and a risk factor for cardiovascular diseases and cancers. Exercise is an effective intervention to combat obesity. Fibronectin type III domain containing protein 5 (FNDC5)/irisin, a myokine, can stimulate the browning of white adipose tissue by increasing uncoupling protein 1 (UCP1) expression, and therefore may represent a link between the beneficial effects of exercise and improvement in metabolic diseases. Thus, upregulating the endogenous expression of FNDC5/irisin by administering medication would be a good approach for treating obesity. Herein, we evaluated the efficacy of raspberry ketone (RK) in inducing FNDC5/irisin expression and the underlying mechanisms. The expression of brown fat-specific proteins (PR domain containing 16 (PRDM16), CD137, and UCP1), heme oxygenase-1 (HO-1), FNDC5, and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) in differentiated 3T3-L1 adipocyte was analyzed by western blotting or immunofluorescence. The level of irisin in the culture medium was also assayed using an enzyme-linked immunosorbent assay kit. Results showed that RK (50 µM) significantly induced the upregulation of FNDC5 protein in differentiated 3T3-L1 adipocytes; however, the irisin level in the culture media was unaffected. Moreover, RK significantly increased the levels of PGC1α, brown adipocyte markers (PRDM16, CD137, and UCP1), and HO-1. Furthermore, the upregulation of PGC1α and FNDC5 and the browning effect induced by RK were significantly reduced by SnPP or FNDC5 siRNA, respectively. In conclusion, RK can induce FNDC5 protein expression via the HO-1 signaling pathway, and this study provides new evidence for the potential use of RK in the treatment of obesity.

Effects of RU486 in Treatment of Traumatic Stress-Induced Glucocorticoid Dysregulation and Fear-Related Abnormalities: Early versus Late Intervention.

Central glucocorticoid receptor (GR) activity is enhanced following traumatic events, playing a key role in the stress-related cognitive abnormalities of posttraumatic stress disorder (PTSD). GR antagonists are expected to have potential as pharmacological agents to treat PTSD-related symptoms such as anxiety and fear memory disruption. However, an incubation period is usually required and stress-induced abnormalities do not develop immediately following the trauma; thus, the optimal intervention timing should be considered. Single prolonged stress (SPS) was employed as a rodent PTSD model to examine the effects of early or late (1-7 versus 8-14 days after the SPS) sub-chronic RU486 (a GR antagonist) administration. Behaviorally, fear conditioning and anxiety behavior were assessed using the fear-conditioning test and elevated T-maze (ETM), respectively. Neurochemically, the expressions of GR, FK506-binding proteins 4 and 5 (FKBP4 and FKBP5), and early growth response-1 (Egr-1) were assessed in the hippocampus, medial prefrontal cortex (mPFC), amygdala, and hypothalamus, together with the level of plasma corticosterone. Early RU486 administration could inhibit SPS-induced behavioral abnormalities and glucocorticoid system dysregulation by reversing the SPS-induced fear extinction deficit, and preventing SPS-reduced plasma corticosterone levels and SPS-induced Egr-1 overexpression in the hippocampus. Early RU486 administration following SPS also increased the FKBP5 level in the hippocampus and hypothalamus. Finally, both early and late RU486 administration inhibited the elevated hippocampal FKBP4 level and hypothalamus GR level in the SPS rats. Early intervention with a GR antagonist aids in the correction of traumatic stress-induced fear and anxiety dysregulation.

Ethyl pyruvate ameliorates heat stroke-induced multiple organ dysfunction and inflammatory responses by induction of stress proteins and activation of autophagy in rats.

Objective: Heat stroke (HS) elicits the systemic inflammatory responses that result in multiple organ dysfunction (MOD). Heat shock response and autophagy are activated during heat stress for removal of damaged organelles and proteins, emerging as a major regulator of cellular homeostasis. Ethyl pyruvate (EP) is a derivative of pyruvic acid and possesses antioxidant and anti-inflammatory effects. This study aims to investigate the effects of EP on MOD in HS rats and explore the possible mechanisms.Method: Anesthetized rats were placed in a heating chamber (42 °C) to elevate the core body temperature attaining to 42.9 °C. Rats were then moved to room temperature and monitored for 6 h. EP (60 mg/kg, i.v.) was administered 30 min prior to heat exposure.Results: Results showed that EP significantly reduced HS-induced increases in plasma levels of LDH, CPK, GPT and CK-MB, reversed the decrease of platelet counts, and alleviated intestinal mucosal and pulmonary damage. Moreover, EP reduced pro-inflammatory protein, including TNF-α, IL-6, IL-1ß, HMGB1 and iNOS, and induced stress proteins, heme oxygenase-1 (HO-1), heat shock protein (HSP) 70 and HSP90 in the liver of HS rats. The levels of HS-activated autophagy-regulatory proteins were affected by EP, in which the phosphorylated mTOR and AKT were reduced, and the phosphorylated AMPK increased, accompanied with upregulation in ULK1, Atg7, Atg12 and LC3II, and downregulation of p62.Conclusion: In conclusion, EP ameliorated HS-induced inflammatory responses and MOD, and the underlying mechanism is associated with the induction of the stress proteins HO-1 and HSP70 as well as restorage of autophagy.

Raloxifene inhibits adipose tissue inflammation and adipogenesis through Wnt regulation in ovariectomized rats and 3 T3-L1 cells.

BACKGROUND: Loss of ovarian function, as in menopause or after ovariectomy (OVX), is closely associated with obesity and white adipose tissue (WAT) inflammation. Estrogen replacement protects against postmenopausal obesity but increases the risks of carcinogenesis. In the present study, we investigated the effects of long-term treatment of raloxifene (RAL), a selective estrogen receptor modulator, on the features of estrogen deficiency-induced obesity and explored the involvement of canonical and non-canonical Wnt regulation in vivo and in vitro. METHODS: Adult female rats received bilateral OVX and divided into 5 groups: (1) Sham, (2) OVX, (3) OVX + E2: OVX rats were administered with E2 (50 µg/kg, s.c., 3 times/week), (4) OVX + RAL: OVX rats were treated with RAL (gavage, 1 mg/kg/day) suspended in 0.8% carboxymethylcellulose (CMC), (5) OVX + CMC: 0.8% CMC as vehicle control. All treatments were given for 8 weeks beginning at 1 week after OVX. In 3 T3-L1 cells, the effects of RAL on adipogenesis and lipopolysaccharide (LPS)-induced inflammation were evaluated. RESULTS: Treatment with RAL significantly decreased body weight, visceral fat pad mass, adipocyte size and plasma levels of glucose but increased plasma adiponectin. RAL reduced the elevation of HIF-1α, VEGF-A and proinflammatory cytokines (MCP-1 and TNF-α) expression by inhibition of NF-κB p65 and JNK cascades in retroperitoneal WAT. This anti-inflammatory capacity of RAL may result from upregulation of secreted frizzle-related protein 5 (SFRP5), an adipokine that repressed Wnt5a signaling. Furthermore, RAL inhibited adipogenic factors such as PPAR-γ, C/EBP-α, and FABP4, and preserved canonical Wnt10b/ß-catenin protein expression. In 3 T3-L1 adipocytes, RAL (20 µM) diminished lipid accumulation and inhibited adipogenic factors accompanied with the induction of ß-catenin, which were effectively reversed by the ß-catenin inhibitor IWR-1-endo. In addition, RAL reduced LPS-induced NF-κB p65 and p-IκB expression as well as TNF-α secretion. Suppression of SFRP5 by small interfering RNA significantly abrogated the anti-inflammatory effects of RAL. CONCLUSIONS: Distinct activation of canonical ß-catenin on inhibition of adipogenesis and non-canonical SFRP5 on suppression of WAT inflammation may contribute to the beneficial effects of RAL. Therefore, this study provides a rationale for the therapeutic potential of RAL for postmenopausal obesity.

Lemnalol Modulates the Electrophysiological Characteristics and Calcium Homeostasis of Atrial Myocytes.

Sepsis, an inflammatory response to infection provoked by lipopolysaccharide (LPS), is associated with high mortality, as well as ischemic stroke and new-onset atrial arrhythmia. Severe bacterial infections causing sepsis always result in profound physiological changes, including fever, hypotension, arrhythmia, necrosis of tissue, systemic multi-organ dysfunction and finally death. LPS challenge-induced inflammatory responses during sepsis may increase the likelihood of the arrhythmogenesis. Lemnalol is known to possess potent anti-inflammatory effects. This study examined whether Lemnalol (0.1 µM) could modulate the electrophysiological characteristics and calcium homeostasis of atrial myocytes under the influence of LPS (1µg/mL). Under challenge with LPS, Lemnalol-treated LA myocytes, had a longer AP duration at 20%, 50% and 90% repolarization of the amplitude, compared to the LPS-treated cells. LPS-challenged LA myocytes showed increased late sodium current, Na+-Ca2+ exchanger current, transient outward current, rapid component of delayed rectifier potassium current, tumor necrosis factor-α, NF-κB and increased phosphorylation of ryanodine receptor (RyR), but a lower L-type Ca2+ current than the control LA myocytes. Exposure to Lemnalol reversed the LPS-induced effects. The LPS-treated and control groups of LA myocytes, with or without the existence of Lemnalol. showed no apparent alterations in the sodium current amplitude or Cav1.2 expression. The expression of sarcoendoplasmic reticulum calcium transport ATPase (SERCA2) was reduced by LPS treatment, while Lemnalol ameliorated the LPS-induced alterations. The phosphorylation of RyR was enhanced by LPS treatment, while Lemnalol attenuated the LPS-induced alterations. In conclusion, Lemnalol modulates LPS-induced alterations of LA calcium homeostasis and blocks the NF-κB pathways, which may contribute to the attenuation of LPS-induced arrhythmogenesis.

Genistein suppresses leptin-induced proliferation and migration of vascular smooth muscle cells and neointima formation.

Obesity is a strong risk factor for the development of cardiovascular diseases and is associated with a marked increase in circulating leptin concentration. Leptin is a peptide hormone mainly produced by adipose tissue and is regulated by energy level, hormones and various inflammatory mediators. Genistein is an isoflavone that exhibits diverse health-promoting effects. Here, we investigated whether genistein suppressed the atherogenic effect induced by leptin. The A10 cells were treated with leptin and/or genistein, and then the cell proliferation and migration were analysed. The reactive oxygen species (ROS) and proteins levels were also measured, such as p44/42MAPK, cell cycle-related protein (cyclin D1 and p21) and matrix metalloproteinase-2 (MMP-2). Immunohistochemistry and morphometric analysis were used for the neointima formation in a rat carotid artery injury model. Genistein (5 µM) significantly inhibited both the proliferation and migration of leptin (10 ng/ml)-stimulated A10 cells. In accordance with these finding, genistein decreased the leptin-stimulated ROS production and phosphorylation of the p44/42MAPK signal transduction pathway. Meanwhile, genistein reversed the leptin-induced expression of cyclin D1, and cyclin-dependent kinase inhibitor, p21. Genistein attenuated leptin-induced A10 cell migration by inhibiting MMP-2 activity. Furthermore, the leptin (0.25 mg/kg)-augmented neointima formation in a rat carotid artery injury model was attenuated in the genistein (5 mg/kg body weight)-treated group when compared with the balloon injury plus leptin group. Genistein was capable of suppressing the atherogenic effects of leptin in vitro and in vivo, and may be a promising candidate drug in the clinical setting.

Activation of autophagy is involved in the protective effect of 17ß-oestradiol on endotoxaemia-induced multiple organ dysfunction in ovariectomized rats.

Oestrogens have been reported to attenuate acute inflammation in sepsis. In this study, the effects of long-term oestrogen replacement with 17ß-oestradiol (E2 ) on endotoxaemia-induced circulatory dysfunction and multiple organ dysfunction syndrome were evaluated in ovariectomized (Ovx) rats. E2 (50 µg/kg, s.c., 3 times/week) was administered for 8 weeks, followed by the induction of endotoxaemia by intravenous infusion of lipopolysaccharides (LPS; 30 mg/kg/4 hrs). Oestrogen deficiency induced by ovariectomy for 9 weeks augmented the LPS-induced damage, including endotoxic shock, myocardial contractile dysfunction, renal dysfunction and rhabdomyolysis. Cardiac levels of NF-κB p65, iNOS and oxidized glutathione, free radical production in skeletal muscles, myoglobin deposition in renal tubules, and plasma levels of plasminogen activator inhibitor-1, TNF-α, and IL-6 were more pronounced in the Ovx + LPS group than in the Sham + LPS group. Long-term treatment of E2 prevented this amplified damage in Ovx rats. Six hours after LPS initiation, activation of the autophagic process, demonstrated by increases in Atg12 and LC3B-II/LC3B-I ratios, and induction of haem oxygenase (HO)-1 and heat-shock protein (HSP) 70 protein expression in myocardium were increased significantly in the Ovx + E2  + LPS group. These results suggest that activation of autophagy and induction of HO-1 and HSP70 contribute to the protective effect of long-term E2 replacement on multiple organ dysfunction syndrome in endotoxaemia.

Excavatolide B Modulates the Electrophysiological Characteristics and Calcium Homeostasis of Atrial Myocytes.

Severe bacterial infections caused by sepsis always result in profound physiological changes, including fever, hypotension, arrhythmia, necrosis of tissue, systemic multi-organ dysfunction, and finally death. The lipopolysaccharide (LPS) provokes an inflammatory response under sepsis, which may increase propensity to arrhythmogenesis. Excavatolide B (EXCB) possesses potent anti-inflammatory effects. However, it is not clear whether EXCB could modulate the electrophysiological characteristics and calcium homeostasis of atrial myocytes. This study investigated the effects of EXCB on the atrial myocytes exposed to lipopolysaccharide. A whole-cell patch clamp and indo-1 fluorimetric ratio technique was employed to record the action potential (AP), ionic currents, and intracellular calcium ([Ca2+]i) in single, isolated rabbit left atrial (LA) cardiomyocytes, with and without LPS (1 µg/mL) and LPS + EXCB administration (10 µM) for 6 ± 1 h, in order to investigate the role of EXCB on atrial electrophysiology. In the presence of LPS, EXCB-treated LA myocytes (n = 13) had a longer AP duration at 20% (29 ± 2 vs. 20 ± 2 ms, p < 0.05), 50% (52 ± 4 vs. 40 ± 3 ms, p < 0.05), and 90% (85 ± 5 vs. 68 ± 3 ms, p < 0.05), compared to the LPS-treated cells (n = 12). LPS-treated LA myocytes showed a higher late sodium current, Na⁺/Ca2+ exchanger current, transient outward current, and delayed rectifier potassium current, but a lower l-type Ca2+ current, than the control LA myocytes. Treatment with EXCB reversed the LPS-induced alterations of the ionic currents. LPS-treated, EXCB-treated, and control LA myocytes exhibited similar Na⁺ currents. In addition, the LPS-treated LA myocytes exhibited a lower [Ca2+]i content and higher sarcoplasmic reticulum calcium content, than the controls. EXCB reversed the LPS-induced calcium alterations. In conclusion, EXCB modulates LPS-induced LA electrophysiological characteristics and calcium homeostasis, which may contribute to attenuating LPS-induced arrhythmogenesis.

Heat shock protein 70 and AMP-activated protein kinase contribute to 17-DMAG-dependent protection against heat stroke.

Heat shock protein 70 (Hsp70) preconditioning induces thermotolerance, and adenosine monophosphate (AMP)-activated protein kinase (AMPK) plays a role in the process of autophagy. Here, we investigated whether 17-dimethylaminoethylamino-17-demethoxy-geldanamycin (17-DMAG) protected against heat stroke (HS) in rats by up-regulation of Hsp70 and phosphorylated AMPK (pAMPK). To produce HS, male Sprague-Dawley rats were placed in a chamber with an ambient temperature of 42°C. Physiological function (mean arterial pressure, heart rate and core temperature), hepatic and intestinal injury, inflammatory mediators and levels of Hsp70, pAMPK and light chain 3 (LC3B) in hepatic tissue were measured in HS rats or/and rats pre-treated with 17-DMAG. 17-DMAG pre-treatment significantly attenuated hypotension and organ dysfunction induced by HS in rats. The survival time during HS was also prolonged by 17-DMAG treatment. Hsp70 expression was increased, whereas pAMPK levels in the liver were significantly decreased in HS rats. Following pre-treatment with 17-DMAG, Hsp70 protein levels increased further, and pAMPK levels were enhanced. Treatment with an AMPK activator significantly increased the LC3BII/LC3BI ratio as a marker of autophagy in HS rats. Treatment with quercetin significantly suppressed Hsp70 and pAMPK levels and reduced the protective effects of 17-DMAG in HS rats. Both of Hsp70 and AMPK are involved in the 17-DMAG-mediated protection against HS. 17-DMAG may be a promising candidate drug in the clinical setting.

Effects of Acetaminophen on Left Atrial Contractility.

BACKGROUND: It has been observed that acetaminophen shows cardioprotective efficacy in mammals. In this study, we investigated the electromechanical effects of acetaminophen on the left atrium (LA). METHODS: Conventional microelectrodes were used to record the action potentials (AP) in rabbit LA preparations. The action potential duration (APD) at repolarization levels of 90%, 50% and 20% of the AP amplitude (APD90, APD50, and APD20, respectively), resting membrane potential, and contractile force were measured during 2 Hz electrical stimulation before and after sequential acetaminophen administration to the LA. RESULTS: Acetaminophen (0.1, 0.3, 1, and 3 mM) reduced APD20 from 9.4 ± 1.2 to 8.0 ± 1.1 (p < 0.05), 7.1 ± 0.8 (p < 0.05), 7.8 ± 1.1, and 6.8 ± 1.2 ms (p < 0.05), respectively, and APD50 from 20.2 ± 1.9 to 17.4 ± 2.0, 15.6 ± 1.8 (p < 0.05), 15.8 ± 2.2 (p < 0.05), and 14.1 ± 2.4 ms (p < 0.05), respectively, in a concentration-dependent manner. APD90 was reduced from 72.0 ± 3.6 to 64.7 ± 4.2, 61.9 ± 4.3, 60.5 ± 3.7, and 53.4 ± 4.4 ms (p < 0.05), respectively. Acetaminophen increased LA contractility from 45 ± 9 to 52 ± 10 (p < 0.05), 55 ± 9 (p < 0.01), 58 ± 9 (p < 0.01), and 60 ± 9 mg (p < 0.01), respectively, in a concentration-dependent manner. In the presence of the NOS inhibitor L-NAME or PKG-I inhibitor DT-2, additional acetaminophen treatment did not significantly increase LA contractility. CONCLUSIONS: Acetaminophen modulated the electromechanical characteristics of LA by inhibiting the NOS and PKG I pathway, and then contributed to the positive inotropic effect.

Effects of Subchronic Buspirone Treatment on Depressive Profile in Socially Isolated Rats: Implication of Early Life Experience on 5-HT1A Receptor-Related Depression.

The heterogeneity of etiology may serve as a crucial factor in the challenges of treatment, including the low response rate and the delay in establishing therapeutic effect. In the present study, we examined whether social experience since early life is one of the etiologies, with the involvement of the 5-HT1A receptors, and explored the potentially therapeutic action of the subchronic administration of buspirone, a partial 5-HT1A agonist. Rats were isolation reared (IR) since their weaning, and the depressive profile indexed by the forced-swim test (FST) was examined in adulthood. Nonspecific locomotor activity was used for the IR validation. Buspirone administration (1 mg/kg/day) was introduced for 14 days (week 9-11). The immobility score of the FST was examined before and after the buspirone administration. Tissue levels of serotonin (5-HT) and its metabolite 5-HIAA were measured in the hippocampus, the amygdala, and the prefrontal cortex. Efflux levels of 5-HT, dopamine (DA), and norepinephrine (NE) were detected in the hippocampus by brain dialysis. Finally, the full 5-HT1A agonist 8-OH-DPAT (0.5 mg/kg) was acutely administered in both behavioral testing and the dialysis experiment. Our results showed (i) increased immobility time in the FST for the IR rats as compared to the social controls, which could not be reversed by the buspirone administration; (ii) IR-induced FST immobility in rats receiving buspirone was corrected by the 8-OH-DPAT; and (iii) IR-induced reduction in hippocampal 5-HT levels can be reversed by the buspirone administration. Our data indicated the 5-HT1A receptor-linked early life social experience as one of the mechanisms of later life depressive mood.

Alpha-Lipoic Acid Induces Adipose Tissue Browning through AMP-Activated Protein Kinase Signaling in Vivo and in Vitro.

Background: AMP-activated protein kinase (AMPK) is a key enzyme for cellular energy homeostasis and improves metabolic disorders. Brown and beige adipose tissues exert thermogenesis capacities to dissipate energy in the form of heat. Here, we investigated the beneficial effects of the antioxidant alpha-lipoic acid (ALA) in menopausal obesity and the underlying mechanisms. Methods: Female Wistar rats (8 weeks old) were subjected to bilateral ovariectomy (Ovx) and divided into four groups: Sham (n=8), Ovx (n=11), Ovx+ALA2 (n=10), and Ovx+ALA3 (n=6) (ALA 200 and 300 mg/kg/day, respectively; gavage) for 8 weeks. 3T3-L1 cells were used for in vitro study. Results: Rats receiving ALA2 and ALA3 treatment showed significantly lower levels of body weight and white adipose tissue (WAT) mass than those of the Ovx group. ALA improved plasma lipid profiles including triglycerides, total cholesterol, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol. Hematoxylin & eosin staining of inguinal WAT showed that ALA treatment reduced Ovx-induced adipocyte size and enhanced uncoupling protein 1 (UCP1) expression. Moreover, plasma levels of irisin were markedly increased in ALA-treated Ovx rats. Protein expression of brown fat-specific markers including UCP1, PRDM16, and CIDEA was downregulated by Ovx but markedly increased by ALA. Phosphorylation of AMPK, its downstream acetyl-CoA carboxylase, and its upstream LKB1 were all significantly increased by ALA treatment. In 3T3-L1 cells, administration of ALA (100 and 250 µM) reduced lipid accumulation and enhanced oxygen consumption and UCP1 protein expression, while inhibition of AMPK by dorsomorphin (5 µM) significantly reversed these effects. Conclusion: ALA improves estrogen deficiency-induced obesity via browning of WAT through AMPK signaling.

Escitalopram reversibility of the impacts following chronic stress on central 5-HT profiles - Implications to depression and anxiety.

Stress is considered a crucial determinant influencing health capacity in modern society. Long-term stress makes individuals more susceptible to mental dysfunctions, among which depression and anxiety are two major mental disorders. The success of using selective serotonin reuptake inhibitors (SSRIs) to treat these two disorders highlights the involvement of the central serotonergic (5-HT) system. Later studies suggest both presynaptic and postsynaptic 5-HT profiles should be considered for the effects of SSRIs, making it difficult to interpret the etiological and therapeutic mechanisms underlying depression and anxiety. The present study aims to examine whether the intervention of escitalopram (Es, 5 mg/kg daily for 14 days) can reverse the behavioral phenotypes of both depression-like [by sucrose preference test (SPT) and forced swim test (FST)] and anxiety-like [by avoidance latency and escape latency in elevated-T maze (ETM)] behaviors, and the brain area-dependent neurochemical changes of 5-HT profiles of the terminal regions regarding both synaptic efflux and tissue levels in rats of chronic mild stress (CMS). Our results showed that: (i) Even mild stresses when presented in an unpredictable and long-term manner, can induce both depression-like and anxiety-like behaviors. (ii) Depressive profile indexed by SPT was more sensitive to reflect the Es effect than that of FST. (iii) Es did not significantly affect the CMS-induced anxiety-like symptoms indexed by ETM. (iv) Changes in the protein expression of 5-HT1A receptors in the prefrontal cortex and hippocampus were compatible with the treatment outcome. Our results contributed to the understanding of stress-induced mood dysfunction and the involvement of central 5-HT.

Ethyl pyruvate reduces acute lung injury via regulation of iNOS and HO-1 expression in endotoxemic rats.

BACKGROUND: Ethyl pyruvate (EP) has been shown to attenuate lipopolysaccharide (LPS)-induced acute lung injury (ALI). Induction of heme oxygenase-1 (HO-1) and suppression of inducible nitric oxide synthase (iNOS) expression provide cytoprotection in lung and vascular injury. The aim of this study is to evaluate whether the beneficial effect of EP on lung inflammation is related to HO-1 induction in a rat model of LPS-induced ALI. MATERIALS AND METHODS: Rats were administered LPS (30 mg/kg) by intravenous infusion for 4 h to induce ALI. EP (20, 40, and 60 mg/kg/4 h i.v. infusion) or vehicle was given 1 h after LPS initiation. RESULTS: EP 40 and 60 mg/kg attenuated plasma levels of TNF-α and IL-6 caused by LPS, and further increased IL-10 levels compared with the LPS group. At 6 h after LPS initiation, iNOS protein expression in lungs and plasma NO metabolite levels were markedly increased, which were reduced by EP 60 mg/kg. LPS caused a significant HO-1 induction, whereas administration of EP 60 mg/kg significantly induced higher HO-1 expression compared with the LPS group. The beneficial effects of EP on cytokines and iNOS expression were reversed by HO-1 inhibitor SnPP. EP significantly suppressed phosphorylated p38 MAPK and increased phosphorylated ERK1/2 protein levels in the lung tissue. The edema and infiltration of neutrophils into lungs was reduced by EP. CONCLUSION: EP reduced LPS-induced ALI, which may be mediated by induction of HO-1. The underlying mechanisms are associated with suppression of p38 MAPK and increase of ERK1/2 signaling pathway activation.

Wogonin suppresses arrhythmias, inflammatory responses, and apoptosis induced by myocardial ischemia/reperfusion in rats.

Wogonin is a flavonoid isolated from Scutellaria baicalensis Georgi, a traditional Chinese medicine, and it possesses antioxidant and anti-inflammatory effects. The aim of this study is to investigate the in vivo effect of wogonin on myocardial ischemia/reperfusion injury in an open-chest anesthetized rat model, which was induced by 45-minute left coronary artery occlusion and 2-hour reperfusion. Rats were treated with wogonin (5, 10, and 20 mg/kg, intraperitoneal) 40 minutes before ischemia or treatment with 10 mg/kg of wogonin 15 minutes after occlusion. Pretreatment with 10 mg/kg of wogonin significantly delayed the occurrence of ventricular premature contractions and tachycardia, and it suppressed the incidence of ventricular tachycardia and ventricular fibrillation, and mortality elicited by ischemia when compared with that in the control group, accompanied by reducing the arrhythmia scores. After 2-hour reperfusion, pretreatment and posttreatment with wogonin significantly reduced the infarct size and plasma levels of creatine kinase muscle-brain fraction and lactate dehydrogenase. Wogonin also significantly reduced the elevation of plasma tissue necrosis factor-α and superoxide anion production in the myocardium with ischemia/reperfusion. The expression of monocyte chemoattractant protein-1, phosphorylated p38 mitogen-activated protein kinase, p65 and IκBα, and active caspase-3 in ischemic myocardium pronouncedly increased in the control group; these were significantly attenuated by treatment with wogonin. In conclusion, wogonin demonstrated in vivo cardioprotective effects by the attenuation of the severity of ischemia-induced arrhythmias and irreversible ischemia/reperfusion injury, which is associated with its antioxidant capacity and anti-inflammatory effects. The suppression of nuclear factor-κB and p38 mitogen-activated protein kinase activation and the inhibition of monocyte chemoattractant protein-1 expression contribute to the beneficial effects of wogonin.

Purinergic P2X receptor regulates N-methyl-D-aspartate receptor expression and synaptic excitatory amino acid concentration in morphine-tolerant rats.

BACKGROUND: The present study examined the effect of P2X receptor antagonist 2',3'-O-(2,4,6-trinitrophenyl) adenosine 5'-triphosphate (TNP-ATP) on morphine tolerance in rats. METHODS: Male Wistar rats were implanted with two intrathecal catheters with or without a microdialysis probe, then received a continuous intrathecal infusion of saline (control) or morphine (tolerance induction) for 5 days. RESULTS: Long-term morphine infusion induced antinociceptive tolerance and up-regulated N-methyl-d-aspartate receptor subunits NR1 and NR2B expression in both total lysate and synaptosome fraction of the spinal cord dorsal horn. TNP-ATP (50 µg) treatment potentiated the antinociceptive effect of morphine, with a 5.5-fold leftward shift of the morphine dose-response curve in morphine-tolerant rats, and this was associated with reversal of the up-regulated NR1 and NR2B subunits in the synaptosome fraction. NR1/NR2B-specific antagonist ifenprodil treatment produced a similar effect as TNP-ATP; it also potentiated the antinociceptive effect of morphine. On day 5, morphine challenge resulted in a significant increase in aspartate and glutamate concentration in the cerebrospinal fluid dialysates of morphine-tolerant rats, and this effect was reversed by TNP-ATP treatment. Moreover, the amount of immunoprecipitated postsynaptic density-95/NR1/NR2B complex was increased in morphine-tolerant rats, and this was prevented by the TNP-ATP treatment. CONCLUSIONS: The findings suggest that attenuation of morphine tolerance by TNP-ATP is attributed to down-regulation of N-methyl-d-aspartate receptor subunits NR1 and NR2B expression in the synaptosomal membrane and inhibition of excitatory amino acids release in morphine-tolerant rats. The TNP-ATP regulation on the N-methyl-d-aspartate receptor expression may be involved in a loss of scaffolding proteins postsynaptic density-95.

Effects of early life social experience on fear extinction and related glucocorticoid profiles - behavioral and neurochemical approaches in a rat model of PTSD.

People may agonize over an intrusive fear-inducing memory even when the traumatic event has passed, which is the principle manifestation of posttraumatic stress disorder (PTSD). However, many traumatized people do not present symptoms of PTSD, implying that certain hidden factors help those individuals to cope with the traumatic stress. Increasing evidence suggests that early life experience may serve as a predisposing factor in the development of PTSD. For example, early life social deprivation disrupts the glucocorticoid system, one of the biological abnormalities of PTSD. By employing isolation rearing (IR) with a subsequent single prolonged stress (SPS) paradigm, we examined the hypothesis that early-life social experience may change the outcome of traumatic stress in both behavioral and neurochemical profiles. Behaviorally, the performance of rats on a Pavlovian fear conditioning test was measured to evaluate their retrieval ability of fear memory extinction. Neurochemically, plasma corticosterone levels and glucocorticoid receptor (GR), FK506-binding proteins 4 and 5 (FKBP4 and FKBP5) and early growth response-1 (Egr-1) expression were measured in GR-abundant brain areas, including the hypothalamus, medial prefrontal cortex, and hippocampus. Our results demonstrated an area-dependent IR effect on the SPS outcomes. IR prevented the SPS-impaired fear extinction retrieval ability and averted the SPS-elevated expression of GR, FKBP4, and Egr-1 in the hippocampus, whereas it did not change the SPS-reduced plasma corticosterone levels and SPS-enhanced GR activity in the mPFC and hypothalamus. The present study provides some new insights to support the hypothesis that early-life experience may play a role in the occurrence of PTSD.

Early life social experience affects adulthood fear extinction deficit and associated dopamine profile abnormalities in a rat model of PTSD.

Individuals may develop fear extinction deficits after life-threatening traumatic events; such deficits indicate posttraumatic stress disorder (PTSD). Because the occurrence of this disorder differs among people who have experienced trauma, hidden underlying factors should be determined. Increasing evidence suggests the involvement of neuronal dysregulation of information processes or cognitive function during development. This neuronal dysregulation is caused by disturbances in dopamine (DA) transmission within the fear circuit, which comprises the medial prefrontal cortex (mPFC), amygdala, and hippocampus. Single prolonged stress (SPS) combined with an isolation rearing (IR) paradigm was used to randomly assign rats to four groups [social rearing-no SPS (SR-NS), SR-SPS, IR-NS, and IR-SPS], and their performance in prepulse inhibition (PPI) and on Pavlovian fear conditioning tests was assessed. Tissue DA levels and the expression of DA receptors (D1R and D2R) in the fear circuit were measured at the end of the experiment. Our results indicated that PPI deficits and fear extinction problems were specific to rats subjected to IR and SPS, respectively. Furthermore, IR-induced PPI deficits were not influenced by SPS, but SPS-induced fear extinction retrieval impairment could be adjusted according to previous IR experiences. Neurochemically, tissue DA levels and D1R expression in the mPFC and amygdala were nonspecifically reduced by IR and SPS, whereas D2R expression in the mPFC and amygdala was higher in IR-SPS than in SR-SPS rats. These findings suggest that early life experiences may influence fear responses in adulthood through a change in DA profiles within the fear circuit.

Involvement of the p62/Nrf2/HO-1 pathway in the browning effect of irisin in 3T3-L1 adipocytes.

Irisin has gained attention because of its potential applications in the treatment of metabolic diseases. Accumulating evidence indicates that irisin attenuates obesity via the browning of white adipose tissue; however, the underlying mechanisms are unclear. Here, we evaluated the effects of irisin on adipocyte browning and the underlying mechanisms. The western blotting and immunofluorescence analyses demonstrated that irisin significantly induced the up-regulation of brown fat-specific proteins (PGC1α, PRDM16, and UCP-1) and HO-1 in 3T3-L1 adipocytes. Moreover, irisin significantly increased the levels of cytosolic p62 and nuclear Nrf2. These effects of irisin in the adipocytes were attenuated by treatment with SnPP or p62 siRNA. In addition, the browning effect of irisin was observed in BAT-WT-1 cells. These findings suggest that irisin induced browning effect via the p62/Nrf2/HO-1 signalling pathway and that it may be a potential candidate for preventing or treating obesity.

Genistein ameliorated obesity accompanied with adipose tissue browning and attenuation of hepatic lipogenesis in ovariectomized rats with high-fat diet.

Estrogen deficiency in postmenopausal women is linked to the higher prevalence of obesity, type 2 diabetes and metabolic syndromes. Development of beige adipocytes (browning of WAT) increases energy expenditure and could be a promising strategy for obesity management. This study aimed to investigate the effects of phytoestrogen genistein (GEN) on white adipose tissue (WAT) inflammation, browning and hepatic lipogenesis in ovariectomized rats with high-fat diet (HFD) and further explore the underlying mechanism. Female Wistar rats received ovariectomy (Ovx) and HFD (45% fat) and then were administered with 17ß-estradiol (E2, 3 times/week, subcutaneously) or GEN (15 mg/kg or 30 mg/kg, gavage, once daily) for 4 weeks. Administration of GEN decreased Ovx-induced body weight gain and adiposity and improved insulin sensitivity as well as increased insulin signaling p-IRS1 and p-AKT in retroperitoneal WAT. Adipocyte hypertrophy and production of proinflammatory cytokines MCP-1, TNF-α and IL-6 were reduced by GEN. It also suppressed the activation of NF-κB pathway evidenced by attenuation of p65 and phospho-IκB levels. Additionally, GEN elevated myokine irisin and promoted WAT browning by increasing UCP-1, PRDM-16, PGC-1α and CIDEA proteins and Ppargc1a, Ucp-1 and Tbx-1 mRNA in inguinal WAT which is associated with up-regulation of nuclear estrogen receptor-α. Plasma levels of triglyceride and cholesterol were reduced by GEN treatment accompanied with inhibition of lipogenic proteins (p-ACC, SREBP-1, FAS and CD36) in the liver. Long-term treatment with GEN attenuated estrogen-deficiency-induced obesity, WAT inflammation and hepatic lipogenesis and promoted the induction of WAT browning. It may provide a promising approach to prevent obesity during menopause.