Neuropeptide Y and leptin receptor expression in the hypothalamus of rats with chronic immobilization stress

In this study, Sprague-Dawley rats were immobilized to a frame for 3 hours a day for 21 days to establish a model of chronic immobilization stress. The body weight and food intake of rats subjected to chronic immobilization stress were significantly decreased compared with the control group. Dual-labeling immunofluorescence revealed that the expression of leptin receptor and the co-localization coeffient in these leptic receptor neurons in the arcuate nucleus of the hypothalamus were both upregulated, while the number of neuropeptide Y neurons was decreased. Chronic immobilization stress induced high expression of leptin receptor in the arcuate nucleus and suppressed the synthesis and secretion of neuropeptide Y, thereby disrupting the pathways in the arcuate nucleus that regulate feeding behavior, resulting in diminished food intake and reduced body weight.

Effects of acupuncture therapy on plasma neuropeptide Y levels and resuscitation in patients with very early stage acute cerebral infarction A randomized controlled study

BACKGROUND： It is known that acupuncture therapy can decrease plasma neuropeptide Y （NPY） levels in patients with cerebral infarction, but different types of acupuncture therapy used in various stages of cerebral infarction have not been evaluated. OBJECTIVE： To explore the effect of acupuncture therapy on resuscitation （Xingnao Kaiqiao） and plasma NPY levels in patients with very early stage acute cerebral infarction. DESIGN, TIME AND SETTING： This case-controlled study was performed at the Affiliated Hospital of the Medical College of the Chinese People＇s Armed Police Force between September 2004 and October 2005. PARTICIPANTS： Sixty patients with acute cerebral infarction of ≤ 6 hours were used in this study. Patients were randomly divided into an acupuncture therapy group （n = 30） and a routine treatment group （n = 30）. Another 30 healthy subjects were used as the control group. METHODS： The acupuncture therapy of Xingnao Kaiqiao used in the acupuncture therapy group was based on routine western medical treatment and was performed at bilateral Neiguan （PCG） using the twirling, reinforcing-reducing method, Renzhong （DU26） using heavy bird-pecking needling, Sanyinjiao （SPG） using reinforcing and reducing by lifting and thrusting the needle, Jiquan （HT1）, Weizhong （BL40） and Chize （LU5） using reinforcing and reducing by lifting and thrusting the needle. The acupuncture lasted for 14 days. Patients in the routine treatment group underwent routine medical treatment and no intervention was given to subjects in the control group. MAIN OUTCOME MEASURES： A 4 mL venous blood sample was obtained at different time points, i.e., immediately after hospitalization, the next morning, 7 and 14 days after treatment, to measure plasma NPY levels pre- and post-treatment using the radio-immunity method. RESULTS： The plasma NPY levels were significantly higher in both the routine treatment group and the acupuncture therapy group than in the control group pre- and post-treatment （P 〈 0.01）. In particular, the plasma NPY levels in both the acupuncture therapy group and the routine treatment group were increased 7 days post-treatment but decreased from 7-14 days post-treatment. In addition, the plasma NPY levels were significantly lower in the acupuncture therapy group than in the routine treatment group on day 7 and 14 post-treatment （P 〈 0.01）. CONCLUSION： Acupuncture therapy of Xingnao Kaiqiao can decrease plasma NPY levels in patients with very early stage acute cerebral infarction. In addition, the therapeutic effect of acupuncture with a prolonged therapy time is superior to routine treatment.

Influence of Ganoderma lucidum spores on the levels of neuropeptide Y and somatostatin in brains of seizure rats

BACKGROUND： Previous research has revealed that somatostatin can induce epilepsy, and that the levels of neuropeptide Y may increase and become more active in brain areas with epileptic seizures. OBJECTIVE： To observe the effect of Ganoderma luciclum spore powder on the neuropeptide Y and somatostatin content in the cerebral cortex and hippocampal regions of seizure rats induced by pentylenetetrazol （PTZ）. Furthermore, to verify any effect of Ganoderma lucidum spore powder on inhibition of epileptic seizures. DESIGN, TIME AND SETTING： A randomized group animal study was performed in August 2007 in the School of Basic Medical Sciences, Jiamusi University （Jiamusi, Heilongjiang, China）. MATERIALS： Thirty healthy, male, Wistar rats, aged 12 weeks and weighing 180-220 g, were taken as the experimental animals. PTZ （Sigma Company, United States） was used to induce epilepsy. Ganoderma lucidum spores （Leyss, ex Fr variety） were purchased from Jiamusi City Wild Growing Case of the Ganoderma Lucidum （China）. Rabbit anti-somatostatin antibodies and secondary antibodies were purchased from Wuhan Boster Company （China）. Neuropeptide Y radioimmunoassay kit was purchased from Beijing Furui Biotechnology Company （China）. METHODS： Thirty rats were randomly divided into three groups： a control group, an epilepsy model group and a Ganoderma lucidum spore-treated group. Each group contained 10 animals. Rats in the epilepsy model group were treated with intraperitoneal injections of PTZ and gastric perfusion of physiologic saline. In the Ganoderma lucidum spore-treated group, intraperitoneal injection of PTZ and gastric perfusion of Ganoderma lucidum spore powder were administered. The blank control group was only administered with the physiological saline by intraperitoneal injection and gastric perfusion. MAIN OUTCOME MEASURES： Immunohistochemical staining and radioimmunoassay methods were used to observe the changes of somatostatin and neuropeptide Y content in brain tissue of epileptic rats, as well as the morphology of neurons. RESULTS： All 30 rats were involved in the result analysis, without any loss. The number of somatostatin immunoreacted positive cells in the cerebral cortex and hippocampus was significantly increased in the epilepsy model group compared to the blank control group （P 〈 0.01）. The number of somatostatin immunoreacted positive cells in cerebral cortex and hippocampus was significantly decreased in the Ganoderma lucidum spore-treated group compared to the epilepsy model group （P 〈 0.01）. The contents of neuropeptide Y in cerebral cortex and hippocampus were significantly increased in the epilepsy model group compared to the blank control group （P 〈 0.01）. The contents of neuropeptide Y in the cerebral cortex and hippocampus were significantly decreased in the Ganoderma lucidum spore-treated group compared to the epilepsy model group （P 〈 0.05）. The epilepsy seizures in the Ganoderma lucidum spore-treated group were obviously reduced compared to the epilepsy model group. CONCLUSION： Ganoderma lucidum spore powder was able to reduce the somatostatin and neuropeptide Y content in the cerebral cortex and hippocampus effectively, so as to achieve an anti-epileptic function and protect neurons from being damaged.

Anti-epileptic effects of neuropeptide Y gene transfection into the rat brain

Neuropeptide Y gene transfection into normal rat brain tissue can provide gene overexpression, which can attenuate the severity of kainic acid-induced seizures. In this study, a recombinant adeno-associated virus carrying the neuropeptide Y gene was transfected into brain tissue of rats with kainic acid-induced epilepsy through stereotactic methods. Following these transfections, we verified overexpression of the neuropeptide Y gene in the epileptic brain. Electroencephalograms showed that seizure severity was significantly inhibited and seizure latency was significantly prolonged up to 4 weeks after gene transfection. Moreover, quantitative fluorescent PCR and western blot assays revealed that the mRNA and protein expression of the N-methyI-D-aspartate receptor subunits NR1, NR2A, and NR2B was inhibited in the hippocampus of epileptic rats. These findings indicate that neuropeptide Y may inhibit seizures via down-regulation of the functional expression of N-methyI-D-aspartate receptors.

Neuropeptide Y gene transfection inhibits post-epileptic hippocampal synaptic reconstruction

Exogenous neuropeptide Y has antiepileptic effects; however, the underlying mechanism and optimal administration method for neuropeptide Y are still unresolved. Previous studies have used intracerebroventricular injection of neuropeptide Y into animal models of epilepsy. In this study, a recombinant adeno-associated virus expression vector carrying the neuropeptide Y gene was injected into the lateral ventricle of rats, while the ipsilateral hippocampus was injected with kainic acid to establish the epileptic model. After transfection of neuropeptide Y gene, mossy fiber sprouting in the hippocampal CA3 region of epileptic rats was significantly suppressed, hippocampal synaptophysin （p38） mRNA and protein expression were inhibited, and epileptic seizures were reduced. These experimental findings indicate that a recombinant adeno-associated virus expression vector carrying the neuropeptide Y gene reduces mossy fiber sprouting and inhibits abnormal synaptophysin expression, thereby suppressing post-epileptic synaptic reconstruction.

Metformin inhibits food intake and neuropeptide Y gene expression in the hypothalamus

Metformin may reduce food intake and body weight, but the anorexigenic effects of metformin are still poorly understood. In this study, Sprague-Dawley rats were administered a single intracere- broventricular dose of metformin and compound C, in a broader attempt to investigate the regula- tory effects of metformin on food intake and to explore the possible mechanism. Results showed that central administration of metformin significantly reduced food intake and body weight gain, par- ticularly after 4 hours. A reduction of neuropeptide Y expression and induction of AMP-activated protein kinase phosphorylation in the hypothalamus were also observed 4 hours after metformin administration, which could be reversed by compound C, a commonly-used antagonist of AMP-activated protein kinase. Furthermore, metformin also improved lipid metabolism by reducing plasma low-density lipoprotein. Our findings suggest that under normal physiological conditions, central regulation of appetite by metformin is related to a decrease in neuropeptide Y gene expres- sion, and that the activation of AMP-activated protein kinase may simply be a response to the anorexigenic effect of metformin.

Neuropeptide Y protects cerebral cortical neurons by regulating microglial immune function

Neuropeptide Y has been shown to inhibit the immunological activity of reactive microglia in the rat cerebral cortex, to reduce N-methyl-D-aspartate current（INMDA） in cortical neurons, and protect neurons. In this study, after primary cultured microglia from the cerebral cortex of rats were treated with lipopolysaccharide, interleukin-1β and tumor necrosis factor-α levels in the cell culture medium increased, and mRNA expression of these cytokines also increased. After primary cultured cortical neurons were incubated with the lipopolysaccharide-treated microglial conditioned medium, peak INMDA in neurons increased. These effects of lipopolysaccharide were suppressed by neuropeptide Y. After addition of the neuropeptide Y Y1 receptor antagonist BIBP3226, the effects of neuropeptide Y completely disappeared. These results suggest that neuropeptide Y prevents excessive production of interleukin-1β and tumor necrosis factor-α by inhibiting microglial reactivity. This reduces INMDA in rat cortical neurons, preventing excitotoxicity, thereby protecting neurons.

Neuropeptide Y and nestin expression in the hippocampal CA3 region following restrained and inverted stress in rats

Our preliminary study demonstrated that neuropeptide Y （NPY）/nestin-positive cells exhibit a consistent spatial distribution in the hippocampus of normal adult rats. However, following severe acute and chronic stress-induced impaired learning and memory, synchronous decreased expression of nestin and NPY takes place in the hippocampus, and the underlying mechanisms remain unclear. In the present study, acute and chronic stress rat models were established using combined restrained and inverted stress. Results showed that learning and memory significantly decreased in acute and chronic stress rats. In addition, hippocampal cells were damaged, in particular in the acute stress rats, and nestin and NPY expression, as well as the number of NPY/nestin-positive cells in the CA3 region, significantly decreased. Furthermore, mature neurofilament 200-positive neurons were absent in the chronic stress rats. The NPY and cytoskeletal protein system equally contributed to stress-induced early learning and memory deficits as well as sustained cerebral iniurv in the adult hippocampus.

Mechanisms of action of neuropeptide Y on stem cells and its potential applications in orthopaedic disorders

Musculoskeletal disorders are the leading causes of disability and result in reduced quality of life.The neuro-osteogenic network is one of the most promising fields in orthopaedic research.Neuropeptide Y(NPY)system has been reported to be involved in the regulations of bone metabolism and homeostasis,which also provide feedback to the central NPY system via NPY receptors.Currently,potential roles of peripheral NPY in bone metabolism remain unclear.Growing evidence suggests that NPY can regulate biological actions of bone marrow mesenchymal stem cells,hematopoietic stem cells,endothelial cells,and chondrocytes via a local autocrine or paracrine manner by different NPY receptors.The regulative activities of NPY may be achieved through the plasticity of NPY receptors,and interactions among the targeted cells as well.In general,NPY can influence proliferation,apoptosis,differentiation,migration,mobilization,and cytokine secretion of different types of cells,and play crucial roles in the development of bone delayed/non-union,osteoporosis,and osteoarthritis.Further basic research should clarify detailed mechanisms of action of NPY on stem cells,and clinical investigations are also necessary to comprehensively evaluate potential applications of NPY and its receptor-targeted drugs in management of musculoskeletal disorders.

Effect of neuropeptide Y on white matter demyelination and serum interleukin-4 and gamma-interferon levels in the guinea pig with experimental allergic encephalomyelitis

BACKGROUND： Neuropeptide Y （NPY） may influence differentiation of Th cells immunological pathology of experimental allergic encephalomyelitis （EAE） is differentiation of Th cells It is assumed that the related to abnormal OBJECTIVE： To investigate the effect of NPY on white matter demyelination, the serum levels interleukin-4 （IL-4） and gamma-interferon （IFN-γ ）, as well as EAE pathogenesis in an EAE guinea pig model following NPY injection into the lateral cerebral ventricle. DESIGN, TIME AND SETTING： A randomized controlled animal study, which was performed in the Infection Immunity Animal Laboratory, Affiliated Hospital of Luzhou Medical College, China, from October 2005 to April 2006. MATERIALS： Thirty healthy female guinea pigs of 8-12 weeks of age, and 10 healthy female rats of three months of age were used. NPY was provided by Sigma Company, USA. NPY kit was provided by Beijing Huaying Biotechnology Institute, China. METHODS： Thirty guinea pigs were randomly divided into three groups： normal control group, EAE model group, and NPY intervention group （n =10 per group）. Normal control group and EAE model group： Saline （10μ L, once） was injected into the lateral cerebral ventricle. After one week, the same volume of Freund＇s adjuvant complete was either injected subcutaneously into two post-palms or EAE was modeled. NPY intervention group： EAE was modeled after one week and NPY was injected （10 μ L of 6 nmol NPY, once） into the lateral cerebral ventricle. Myelin basic protein （MBP） antigen made from rat spinal cord homogenate and Freund＇s adjuvant complete were injected subcutaneously into both post-palms （0.2 mL per palm） to establish the EAE model. MAIN OUTCOME MEASURES： White matter demyelination of the cerebrum, cerebellum, brain stem, and spinal cord were observed by light microscopy after HE staining. Levels of serum IFN-γ and IL-4 were detected by the double antibody sandwich ABC-ELISA technique. NPY content was detected by radioimmunoassay. RESULTS： Pathological alterations in the NPY intervention groups were reduced compared to those in the EAE model group, suggesting a reduction and remission of white matter demyelination with NPY treatment. When compared to the model group, the serum IL-4 level was increased in the NPY intervention group during the high-frequent EAE stage （P 〈 0.01）, but the serum IFN-γ level was decreased （P 〈 0.01）. Furthermore, the EAE latency was prolonged （P 〈 0.01）, the neurological scores were decreased in the high-frequent EAE stage （P 〈 0.01）, and the death rate was decreased （P 〈 0.05）. NPY content and the serum IL-4 level at the peak stage were positively correlated with those in the latent phase （r =0.863-0.900, P 〈 0.01）, but negatively correlated with neurological scores at the peak stage （r=- -0.068 to -0.863, P 〈 0.05-0.01）. The IFN-γ level at the peak stage was negatively correlated to that in the latent phase （r = -0.683-0.650, P 〈 0.05）, but positively correlated to neurological scores at the peak stage （r =0.975, 0.845, P 〈 0.05）. CONCLUSION： NPY injection into the lateral cerebral ventricle can promote the secretion of IL-4, inhibit the production of IFN-γ, relieve white matter demyelination, and inhibit EAE attack in an experimental model of EAE.

Role of Neuropeptide Y and Peroxisome Proliferator-activated Receptor γ Coactivator-1α in Stress Cardiomyopathy

Death following situations of intense emotional stress has been linked to the cardiac pathology described as stress cardiomyopathy, whose pathomechanism is still not clear. In this study, we sought to determine, via an animal model, whether the transcriptional coactivator peroxisome proliferator-activated receptor γ coactivator-1alpha (PGC-1α) and the amino peptide neuropeptide Y (NPY) play a role in the pathogenesis of this cardiac entity. Male Sprague-Dawley rats in the experimental group were subjected to immobilization in a plexy glass box for 1 h, which was followed by low voltage elec-tric foot shock for about 1h at 10s intervals in a cage fitted with metallic rods. After 25 days the rats were sacrificed and sections of their hearts were processed. Hematoxylin-eosin staining of cardiac tissues revealed the characteristic cardiac lesions of stress cardiomyopathy such as contraction band necrosis, inflammatory cell infiltration and fibrosis. The semi-quantitative RT-PCR analysis for PGC-1α mRNA expression showed significant overexpression of PGC1-α in the stress-subjected rats (P<0.05). Fluorescence immunohistochemistry revealed a higher production of NPY in the stress-subjected rats as compared to the control rats (P=0.0027). Thus, we are led to conclude that following periods of intense stress, an increased expression of PGC1-α in the heart and an overflow of NPY may lead to stress car-diomyopathy and even death in susceptible victims. Moreover, these markers can be used to identify stress cardiomyopathy as the cause of sudden death in specific cases.

Associations of Gonadotropin-Releasing Hormone Receptor (GnRHR) and Neuropeptide Y(NPY) Genes’Polymorphisms with Egg-Laying Traits in Wenchang Chicken

Single nucleotide polymorphisms （SNP） of chicken gonadotropin-releasing hormone receptor （GnRHR） and neuropeptide Y （NPY） were selected to identify the genotypes of Wenchang （Chinese indigenous breed） chicken with restricton fragment length polymorphisms. The associations of the SNPs with the total egg production （NE）, average days of continual laying （ADCL）, and number of double-yolked eggs （DYE） traits were analyzed. The frequency of restriction enzyme A/a alleles in the population was for GnRHR 0.69 （Bpu1102 Ⅰ A） and 0.31 （Bpu1102 Ⅰ a） and for NPY 0.46 （Dra Ⅰ B） and 0.54 （Dra Ⅰ b）. Trait data from a total of 120 hens, which were purebred introduced from Hainan Province, China from one generation were recorded. Two significant effects of genes＇ marker were found： for GnRHR and number of eggs （dominant; t= 2.67, df= 116） and NPY and number of eggs （additive; t= 1.97, df= 116）. The current research supports the effects of GnRHR and NPY genes on egg-laying traits of chickens.

Identification and characterization of a novel neuropeptide(neuropeptide Y-HS) from leech salivary gland of Haemadipsa sylvestris

The present study was designed to identify immunomodulatory components from the leech salivary gland of Haemadipsa sylvestris. The Sephadex G-50, ResourceTM S column chromatography and reverse-phase high performance liquid chromatography(RP-HPLC) were used to isolate and purify the salivary gland extracts(SGE). Structural analysis of isolated compounds was based on Edman degradation and matrix assisted laser desorption ionization time-of-flight mass spectrometer(MALDI-TOF-MS). The c DNA encoding the precursor of the compound was cloned from the c DNA library of the salivary gland of H. sylvestris. The levels of inflammatory mediators, including tumor necrosis factor-α(TNF-α), interferon ?(IFN-?), interleukin-6(IL-6), and monocyte chemotactic protein-1(MCP-1) were assayed using an enzyme-linked immunosorbent assay(ELISA). The effects on cell proliferation and cell viability were observed using MTT assay. A novel neuropeptide Y(Neuropeptide Y-HS) from the leech salivary gland of H. sylvestris was purified and characterized. It was composed of 36 amino acid residues and the amino acid sequence was determined to be FLEPPERPAVFTSVEQMKSYIKALNDYYLLLGRPRF-NH2, containing an amidated C-terminus. It showed significant inhibitory effects on the production of inflammatory cytokines including TNF-?, IFN-?, IL-6, and MCP-1. Neuropeptide Y was identified from leeches for the first time. The presence of neuropeptide Y-HS in leech salivary gland may help get blood meal from hosts and inhibit inflammation.

Plasma Neuropeptide Y Levels in Vasovagal Syncope in Children

Background： Vasovagal syncope （VVS） is the most common cause of syncope in children. Neuropeptide Y （NPY） plays an important role in the regulation of blood pressure （BP）, as well as myocardial contractility. This study aimed to explore the role of plasma NPY in VVS in children. Methods： Fifty-six children who were diagnosed with VVS （VVS group） using head-up tilt test （HUT） and 31 healthy children who were selected as controls （control group） were enrolled. Plasma NPY concentrations were detected. The independent t-test was used to compare the data of the VVS group with those of the control group. The changes in plasma NPY levels in the VVS group during the HUT, as well as hemodynamic parameters, such as heart rate （HR）, BP, total peripheral vascular resistance （TPVR）, and cardiac output （CO）, were evaluated using the paired t-test. Furthermore, the correlations between plasma NPY levels and hemodynamic parameters were analyzed using bivariate correlation analysis. Results： The BP, HR, and plasma NPY （0.34 ± 0.12 pg/ml vs. 0.46 ± 0.13 pg/ml） levels in the supine position were statistically low in the VVS group compared to levels in the control group （all P 〈 0.05）. Plasma NPY levels were positively correlated with the HR （Pearson, R = 0.395, P 〈 0.001） and diastolic BP （Pearson, R = 0.311, P = 0.003） when patients were in the supine position. When patients in the VVS group were in the supine position, elevated TPVR （4.6 ± 3.7 mmHg·min-1·L-1 vs. 2.5 ± 1.0 mmHg·min-1·L-1, respectively, P 〈 0.001;1 mmHg = 0.133 kPa） and reduced CO （1.0 ± 0.7 L/min vs. 2.4 ± 1.3 L/min, respectively, P 〈 0.001） were observed in the positive-response period compared with baseline values. The plasma NPY levels were positively correlated with TPVR （Spearman, R = 0.294, P = 0.028） but negatively correlated with CO in the positive-response period during HUT （Spearman, R = -0.318, P = 0.017）. Conclusions： Plasma NPY may contribute to the pathogenesis of VVS by increasing the TPVR and decreasing the CO during orthostatic regulation.

Neuropeptide Y receptors:a promising target for cancer imaging and therapy 5th China-Europe Symposium on Biomaterials in Regenerative Medicine(CESB 2015)Hangzhou,China April 7–10,2015

Neuropeptide Y(NPY)was first identified from porcine brain in 1982,and plays its biological functions in humans through NPY receptors(Y1,Y2,Y4 and Y5).NPY receptors are known to mediate various physiological functions and involve in a majority of human diseases,such as obesity,hypertension,epilepsy and metabolic disorders.Recently,NPY receptors have been found to be overexpressed in many cancers,so they emerged as promising target in cancer diagnosis and therapy.This review focuses on the latest research about NPY and NPY receptors,and summarizes the current knowledge on NPY receptors expression in cancers,selective ligands for NPY receptors and their application in cancer imaging and therapy.

Neuropeptide Y and melanocortin receptors in fish:regulators of energy homeostasis

Energy homeostasis,which refers to the physiological processes that the energy intake is exquisitely coordinated with energy expenditure,is critical for survival.Therefore,multiple and complex mechanisms have been involved in the regulation of energy homeostasis.The central melanocortin system plays an important role in modulating energy homeostasis.This system includes the orexigenic neurons,expressing neuropeptide Y/Agouti-related protein(NPY/AgRP),and the anorexigenic neurons expressing proopiomelanocortin(POMC).The downstream receptors of NPY,AgRP and post-translational products of POMC are G protein-coupled receptors(GPCRs).This review summarizes the compelling evidence demonstrating that NPY and melanocortin receptors are involved in energy homeostasis.Subsequently,the comparative studies on physiology and pharmacology of NPY and melanocortin receptors in humans,rodents and teleosts are summarized.Also,we provide a strategy demonstrating the potential application of the new ligands and/or specific variants of melanocortin system in aquaculture.

Electroacupuncture Alleviates Anxiety-like Behavior in Pain Aversion Rats by Attenuating the Expression of Neuropeptide Y in Anterior Cingulate Cortex

Background:Pain is considered as a multidimensional conscious experience that includes a sensory component and a negative affective-motivational component.Electroacupuncture(EA)is widely used to treat pain and paininduced negative emotions,however,little is known about the mechanisms underlying the effect of EA.Objective:This study investigated the effect of EA on alleviating the anxiety-like behaviors in pain aversion rats and its anterior cingulate cortex(ACC)regulation mechanism.Methods:After a Freund’s complete adjuvant(CFA)-conditioned place aversion(C-CPA)model was established in rats,EA treatment(2/100 Hz,30 min,once/day,4 days totally)was applied at bilateral Zusanli(ST36)and Kunlun(BL60)acupoints.Von Frey filaments were used to measure changes of pain withdrawal threshold(PWT)at indicated time points.Elevated zero maze(EZM)was used to investigate the changes of pain-related anxiety and CPA was used to investigate the changes of pain aversion.The protein expression levels of GAD67,PV,and NPY in ACC were detected by Western blotting.Results:Compared with the control group,the staying time in the"CFA-paired compartment"was significantly reduced,and the PWT was decreased in model group.In the EZM assessment,the distance and the time in open arm,as well as the number of open arm entries of model group were significantly lower than those in the control group.In the CPA assessment,the time spent in the"CFA-paired compartment"was significantly decreased in model group compared with control group,and EA reversed the changes in pain sensation and in pain-related emotions.Western blotting showed that the NPY level,but not the levels of GAD67 and PV,was significantly increased in the ACC of the model group compared to that of the control group.The increased expression of NPY in the ACC was significantly downregulated by EA,while sham EA produced no such effect.Conclusion:EA can effectively relieve the pain and pain-related emotions,and its mechanism may be achieved by down-regulating the expression of NPY in the ACC.

The effect of traditional Chinese medicine──Tian Gui Recipeon obesity and anovulation in androgen-sterilized rats

Objective: A traditional Chinese medicine Tian Gui Recipe (TGR) has been used to effectively treat clomiphene resistant anovulatory disease and obesity, especially in polycystic ovary syndrome (PCOS) cases with hyperinsulinemia. The effect of TGR on obesity and anovulation was investigated in androgen-sterilized rats (ASR). Methods: Female SD rats at the age of 9 days were divided into 3 groups. Group ASR (n=15): anovulation was demonstrated at the age of 70 days by vaginal smear in rats while 1. 25 mg of testosterone propionate was injected subcutaneously on its 9th neonatal day. Group A+H (n = 25): TGR were administered to ASR at the age of 80 days for 3 weeks. Rats with regular estrous cycle and ovulation after herbal treatment were included in this group. Group C (n = 15): normal ovulated rats were recruited. Around the age of 112 days or on proestrous day, all rats were sacrificed under anesthesia. Serum leptin, testosterone (T), estrogen (E2), follicular stimulating hormone (FSH), luteinizing hormone (LH) levels were measured with radioimmunoassay (RIA). Double immunofluorescent staining combined with confocal laser scanning microscope and dual in situ hybridization were carried out on sections through areas of arcuate nucleus (ARC) to determine whether estrogen receptor (ER) immunoreactivity (IR) or long form of leptin receptor (OB-Rb) mRNA were expressed in neuropeptide Y (NPY) immuno-and mRNA-containing neurons, and its relationship to proopiomelanocortin (POMC) mRNA-containing neurons. Immunohistochemistry and in situ hybridization were used to observe the levels of ER, NPY, gonadotrophin releasing hormone (GnRH) and gene expression levels on NPY, OB-Rb, POMC. Meanwhile, the criteria of energy state, including daily food intake, retroperitoneal fat depot pad and body weight, were measured and evaluated. Values of immunohistochemistry and in situ hybridization were expressed in mean optic density (MOD). Results: Seventeen out of the 25 rats (68%) in Group A+H were ovulated after herbal treatment. ASR characterized with significantly high metabolic rate, energy imbalance and obesity (P<0. 01, P<0. 01, P<0. 01). The mean serum leptin, T, E2 levels of group ASR were significantly higher than those of Group C and Group A+H (P< 0. 01,P<0. 01 ), while FSH, LH levels were lower (P<0. 01, P<0. 01 ). ERs were proved to be distributing in NPY-containing neurons, OB-Rb coexpressed in NPY-syn- thesizing neurons, and both NPY and POMC mRNA expressing neurons overlapped in ARC. Compared to Group C, levels of NPY mRNA and NPY, POMC mRNA expressions were increased (P<0. 01, P<0. 01, P<0. 01 ), OB-Rb mRNA, ER and GnRH immunoreactive expression were decreased significantly in the hypothalamus of ASR (P<0. 01, P<0. 01, P<0. 01 ). These dramatic neuroendocrine changes became normal in ovulated ASR after the herbal treatment (all P>0. 05). Conclusion: The elevated peripheral E2 caused by high T and leptin levels in ASR may down-regulate their corresponding receptors oriented on hypothalamic NPY- containing neurons respectively, therefore challenge the NPY mRNA and NPY, POMC mRNA overexpressions. The overexpression, of NPY, POMC gene transcription and translation stimulate food intake, promote the deposit of adipocyte tissue, and inhibit GnRH expression and GnRH/FSH, LH release, which contribute to the occurrence of obesity and anovulation in ASR. TGR may reverse these abnormal neuroendocrine cascades by lowering the levels of T, E2 and leptin.

Identification of differentially expressed genes in ulcerative colitis and verification in a colitis mouse model by bioinformatics analyses

BACKGROUND Ulcerative colitis(UC)is an inflammatory bowel disease that is difficult to diagnose and treat.To date,the degree of inflammation in patients with UC has mainly been determined by measuring the levels of nonspecific indicators,such as C-reactive protein and the erythrocyte sedimentation rate,but these indicators have an unsatisfactory specificity.In this study,we performed bioinformatics analysis using data from the National Center for Biotechnology Information-Gene Expression Omnibus(NCBI-GEO)databases and verified the selected core genes in a mouse model of dextran sulfate sodium(DSS)-induced colitis.AIM To identify UC-related differentially expressed genes(DEGs)using a bioinformatics analysis and verify them in vivo and to identify novel biomarkers and the underlying mechanisms of UC.METHODS Two microarray datasets from the NCBI-GEO database were used,and DEGs between patients with UC and healthy controls were analyzed using GEO2R and Venn diagrams.We annotated these genes based on their functions and signaling pathways,and then protein-protein interactions(PPIs)were identified using the Search Tool for the Retrieval of Interacting Genes.The data were further analyzed with Cytoscape software and the Molecular Complex Detection(MCODE)app.The core genes were selected and a Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis was performed.Finally,colitis model mice were established by administering DSS,and the top three core genes were verified in colitis mice using real-time polymerase chain reaction(PCR).RESULTS One hundred and seventy-seven DEGs,118 upregulated and 59 downregulated,were initially identified from the GEO2R analysis and predominantly participated in inflammation-related pathways.Seven clusters with close interactions in UC formed:Seventeen core genes were upregulated[C-X-C motif chemokine ligand 13(CXCL13),C-X-C motif chemokine receptor 2(CXCR2),CXCL9,CXCL5,C-C motif chemokine ligand 18,interleukin 1 beta,matrix metallopeptidase 9,CXCL3,formyl peptide receptor 1,complement component 3,CXCL8,CXCL1,CXCL10,CXCL2,CXCL6,CXCL11 and hydroxycarboxylic acid receptor 3]and one was downregulated[neuropeptide Y receptor Y1(NYP1R)]in the top cluster according to the PPI and MCODE analyses.These genes were substantially enriched in the cytokinecytokine receptor interaction and chemokine signaling pathways.The top three core genes(CXCL13,NYP1R,and CXCR2)were selected and verified in a mouse model of colitis using real-time PCR Increased expression was observed compared with the control mice,but only CXCR2 expression was significantly different.CONCLUSION Core DEGs identified in UC are related to inflammation and immunity inflammation,indicating that these reactions are core features of the pathogenesis of UC.CXCR2 may reflect the degree of inflammation in patients with UC.

Effect of Amlodipine on the Growth of Vascular SmoothMuscle Cells of Spontaneously Hypertensive Rats

The effect of anti-hypertensive drug amlodipine on regression of cardiovascular hypertrophy due to hypertension was studied by using cultured smooth muscle cells derived from arteries of spontaneously hypertensive rats (SHR) and measuring [3H]-TdR and [3H]-Leucine binding. 48 h after adding amlodipine.[3H] -TdR binding in arterial smooth muscle cells from SHR in vitro was reduced by 50.5% and [3H]-Leucine binding was reduced by 56. 2% as compared with neuropeptide Y (NPY)-treated group. However, there was no significant change in cell number. The results showed that amlodipine could effectively inhibit increase of DNA and protein synthesis of vascular smooth muscle cell (VSMC) due to NPY. It indicates that amlodipine is of great significance on regression of genesis and development of cardiovascular hypertrophy due to hypertension.