Inactivation of the mouse melanocortin-3 receptor results in increased fat mass and reduced lean body mass.

Genetic and pharmacological studies have defined a role for the melanocortin-4 receptor (Mc4r) in the regulation of energy homeostasis. The physiological function of Mc3r, a melanocortin receptor expressed at high levels in the hypothalamus, has remained unknown. We evaluated the potential role of Mc3r in energy homeostasis by studying Mc3r-deficient (Mc3r(-/-)) mice and compared the functions of Mc3r and Mc4r in mice deficient for both genes. The 4-6-month Mc3r-/- mice have increased fat mass, reduced lean mass and higher feed efficiency than wild-type littermates, despite being hypophagic and maintaining normal metabolic rates. (Feed efficiency is the ratio of weight gain to food intake.) Consistent with increased fat mass, Mc3r(-/-) mice are hyperleptinaemic and male Mc3r(-/-) mice develop mild hyperinsulinaemia. Mc3r(-/-) mice did not have significantly altered corticosterone or total thyroxine (T4) levels. Mice lacking both Mc3r and Mc4r become significantly heavier than Mc4r(-/-) mice. We conclude that Mc3r and Mc4r serve non-redundant roles in the regulation of energy homeostasis.

Mutant human presenilin 1 protects presenilin 1 null mouse against embryonic lethality and elevates Abeta1-42/43 expression.

Mutations in presenilin 1 (PS1) are linked to early onset of familial Alzheimer's disease (FAD) and are shown to foster production of Abeta1-42/43 in FAD patients and transgenic mice. PS1 null mice are embryonic lethal and exhibit axial skeleton malformation and CNS defects. We show that transgenic mouse lines expressing either the wild-type human PS1 protein or human PS1 with the A246E FAD mutation can rescue the PS1 knockout mouse from embryonic lethality to similar degrees, indicating that the mutation does not lead to loss of PS1 function during development. Furthermore, a 50% reduction of PS1 activity in PS1(+/-) mice does not lead to Abeta1-42/43 increase, whereas expression of human mutant PS1 on murine PS1 null background is sufficient to elevate Abeta1-42/43, supporting a gain-of-function activity as the result of the PS1 mutation.

The mechanisms of Ang II-induced hypertensive vascular remodeling under suppression of CD68 in macrophages.

OBJECTIVE: High blood pressure (hypertension) is one of the most common cardiovascular diseases. In recent years, there were more and more studies on the function of inflammation in hypertension. CD68 mainly mediates the activation of cytokine interleukin-17 (IL-17) signaling pathway and participates in inflammatory responses. It has been studied the function of CD68 and IL-17 in hypertension, but it has not been reported whether it affected hypertension and vascular remodeling when macrophage CD68 expression inhibited. In this study, antisense-CD68 mice were used to study the effect and mechanism of angiotensin II-induced hypertensive vascular remodeling under specific suppression of macrophage CD68. MATERIALS AND METHODS: Fifty 8-week-old male antisense-CD681 and C57 mice were divided into control and experimental group (angiotensin II group, 1000 ng•kg-1•min-1). After infusion of angiotensin II for 28 days, hematoxylin-eosin (HE) staining and immunohistochemical staining were used to observe the remodel of vascular. The changes of aortic inflammatory factors were detected by Real-time PCR (RT-PCR) and Western blotting. RESULTS: By specifically inhibiting the expression of macrophage CD68, macrophage infiltration was mitigated in Ang II-induced hypertensive vascular remodeling model mouse, which also down-regulated the expression of vascular tissue inflammatory factor and activation of vascular smooth muscle cell p65. CONCLUSIONS: CD68 regulates the Ang II-induced hypertensive vascular remodeling through mediating macrophage inflammatory factor release.

A selective human beta3 adrenergic receptor agonist increases metabolic rate in rhesus monkeys.

Activation of beta3 adrenergic receptors on the surface of adipocytes leads to increases in intracellular cAMP and stimulation of lipolysis. In brown adipose tissue, this serves to up-regulate and activate the mitochondrial uncoupling protein 1, which mediates a proton conductance pathway that uncouples oxidative phosphorylation, leading to a net increase in energy expenditure. While chronic treatment with beta3 agonists in nonprimate species leads to uncoupling protein 1 up-regulation and weight loss, the relevance of this mechanism to energy metabolism in primates, which have much lower levels of brown adipose tissue, has been questioned. With the discovery of L-755,507, a potent and selective partial agonist for both human and rhesus beta3 receptors, we now demonstrate that acute exposure of rhesus monkeys to a beta3 agonist elicits lipolysis and metabolic rate elevation, and that chronic exposure increases uncoupling protein 1 expression in rhesus brown adipose tissue. These data suggest a role for beta3 agonists in the treatment of human obesity.

[Prognosis of the central nervous system involvement in patients with hemophagocytic lymphohistiocytosis].

Objective: To investigate the characteristics and prognostic factor of central nervous system (CNS) involvement in patients with hemophagocytic lymphohistiocytosis (HLH) . Methods: From January 2006 to October 2015, 152 patients with HLH, 88 patients had CNS involvement, their clinical data were collected, and survival was analyzed using the Kaplan-Meier life table method, univariate and multivariate Cox regression model analyses were applied to identify the risk factors of prognosis. Results: ①57.9% patients complicated with neurological symptoms, cerebrospinal fluid abnormalities were observed in 37.0% patients, 57.5% patients had abnormal neuroradiology. ②36 patients survived well, 3 patients lost to follow-up, 49 dead, 1 survival patient had epilepsy. ③The 3-year overall survival rate of 88 patients was 44%. ④abnormal CSF and unreceived IT bore a significant, independent adverse prognostic value (P<0.05) . Conclusion: CNS involvement in HLH has a high frequency and poor prognosis, few patients remained neurologic sequelae; abnormal CSF related to poor prognosis, positive intrathecal injections could improve the prognosis.

Effects of alpha-melanocyte-stimulating hormone on magnocellular oxytocin neurones and their activation at intromission in male rats.

The peptides alpha-melanocyte-stimulating hormone (alpha-MSH) and oxytocin have very similar effects on several behaviours, including male sexual behaviour. Both induce penile erection and enhance copulatory behaviour when given centrally, suggesting that their central actions are not independent. Here, we used intromission as a physiological stimulus to investigate whether some central effects of alpha-MSH during male sexual behaviour are mediated by oxytocin neurones. We used the expression of the immediate-early gene product Fos to investigate oxytocin neurone activation at intromission and after intracerebroventricular (i.c.v.) administration of alpha-MSH (1 microg/5 microl) and studied the effects of i.c.v. administration of a MC4 receptor antagonist on Fos expression and on the latency of male rats to exhibit sexual behaviour in the presence of a receptive female. In rats that showed intromission, Fos was expressed in magnocellular oxytocin neurones in both the paraventricular nucleus (PVN) and the supraoptic nucleus (SON), but there was no significant activation of parvocellular oxytocin neurones of the PVN. Similarly, alpha-MSH increased Fos expression in magnocellular oxytocin neurones but had little or no effect in parvocellular oxytocin neurones. In male rats that achieved intromission, central injection of a MC4 receptor antagonist significantly attenuated the increase in Fos expression in magnocellular oxytocin neurones in both the PVN and the SON and increased mount and intromission latencies compared to vehicle-injected controls. Together, the results indicate that magnocellular oxytocin neurones are involved in the central regulation of male sexual behaviour, and that some of the central effects of alpha-MSH are likely to be mediated by magnocellular oxytocin neurones.

[X-linked lymphoproliferative syndrome type 1 complicated with secondary hemophagocytic lymphohistiocytosis and ileal perforation: case report and literature review].

OBJECTIVE: To analyze and summarize the clinical characteristics, laboratory tests and treatment of X-linked lymphoproliferative syndrome type 1 (XLP-1). METHOD: A retrospective study was done in 2012 on an XLP-1 patient to collect the data on clinical manifestation, laboratory examination, gene and protein expression, complications and prognosis. Literatures were reviewed in Pubmed with the key word"X-linked lymphoproliferative syndrome". RESULT: The patient with persistent high fever, jaundice, abdominal distension, hepatosplenomegaly and lymphadenectasis, rash and suspicious positive family history; the patient eventually died of hemophagocytic lymphohistiocytosis (HLH), with intestinal perforation, intestinal infection and bleeding after being infected with EB virus. This patient with SH2D1A gene exon 1 large fragment of the coding region of the nucleotide deletion and insertion mutations causing missense mutations (p.Leu25Lys) and nonsense mutations (stop codon TAG was inserted after missense mutation so that the protein encoded by the early termination of the 25 amino acids), which led to SAP protein missing. The expression of SAP in his mother was also partly missing. Retrieval of reports on XLP-1 was conducted through literature search (included totally 157 cases) at home and abroad, positive family history accounted for 60.6%(40/66); lymphoma incidence accounted for 49.7%(72/145); low gamma globulin occurred in 24.8%(39/157) of cases; secondary HLH ratio accounted for 43.3%(68/157); XLP-1 in patients with hemorrhagic enteritis and gastritis was low, accounted for only 2.6%(3/116). CONCLUSION: XLP-1 patients occasionally develop necrotic enteritis complicated with ileal perforation.XLP-1 with large fragment deletion of SH2D1A gene might be associated with serious gastrointestinal manifestations.

Orexigenic action of peripheral ghrelin is mediated by neuropeptide Y and agouti-related protein.

Ghrelin, a stomach-derived orexigenic hormone, has stimulated great interest as a potential target for obesity control. Pharmacological evidence indicates that ghrelin's effects on food intake are mediated by neuropeptide Y (NPY) and agouti-related protein (AgRP) in the central nervous system. These include intracerebroventricular application of antibodies to neutralize NPY and AgRP, and the application of an NPY Y1 receptor antagonist, which blocks some of the orexigenic effects of ghrelin. Here we describe treatment of Agrp(-/-);Npy(-/-) and Mc3r(-/-);Mc4r(-/-) double knockout mice as well as Npy(-/-) and Agrp(-/-) single knockout mice with either ghrelin or an orally active nonpeptide ghrelin agonist. The data demonstrate that NPY and AgRP are required for the orexigenic effects of ghrelin, as well as the involvement of the melanocortin pathway in ghrelin signaling. Our results outline a functional interaction between the NPY and AgRP pathways. Although deletion of either NPY or AgRP caused only a modest or nondetectable effect, ablation of both ligands completely abolished the orexigenic action of ghrelin. Our results establish an in vivo orexigenic function for NPY and AgRP, mediating the effect of ghrelin.

Distribution of orexin receptor mRNA in the rat brain.

The expression pattern of mRNA encoding two orexin receptors (OX1R and OX2R) in the rat brain was examined. OX1R and OX2R exhibited marked differential distribution. Within the hypothalamus, OX1R mRNA is most abundant in the ventromedial hypothalamic nucleus whereas OX2R is predominantly expressed in the paraventricular nucleus. High levels of OX1R mRNA were also detected in tenia tecta, the hippocampal formation, dorsal raphe, and locus coeruleus. OX2R mRNA is mainly expressed in cerebral cortex, nucleus accumbens, subthalamic and paraventricular thalamic nuclei, anterior pretectal nucleus. The presence of orexin receptor mRNA in the hypothalamus is in support of its proposed role in feeding regulation. Broad central distribution of orexin receptors may indicate additional functions for orexins.

Differential expression of mRNA for leptin receptor isoforms in the rat brain.

Leptin plays an important role in the control of food intake and energy metabolism by interacting with its receptor (OB-R) in the brain. Several alternatively spliced isoforms of OB-R have been identified. To study the expression patterns and the potential biological function of these OB-Rs in the brain, the distribution of mRNA encoding OB-R isoforms was examined by in situ hybridization. In agreement with previous studies, strong signals for OB-R mRNA were detected in the hypothalamus, thalamus and choroid plexus. In addition, intense signals were observed in several other brain areas including piriform cortex, granule cell layer of the cerebellum and substantia nigra. With isoform-specific probes, a differential expression pattern of OB-Rs was revealed: OB-Ra and OB-Rb, but not OB-Rc and OB-Rf, are abundantly expressed in the hypothalamus, whereas OB-Ra, OB-Rc and OB-Rf, but not OB-Rb, are significantly expressed in the choroid plexus. The preferential expression of OB-Rb in the hypothalamus is in support of its role in mediating the satiety effect of leptin. The co-expression of OB-Ra with OB-Rb in the hypothalamus may suggest a possible interaction between the two isoforms. Finally, the detection of OB-R mRNA in a number of other brain regions may indicate the involvement of leptin in additional as yet undefined physiological functions.

Evidence of altered hypothalamic pro-opiomelanocortin/ neuropeptide Y mRNA expression in tubby mice.

The tubby mouse is characterized by an autosomal recessive mutation which results in the development of maturity-onset obesity and sensorineural hearing loss and retinal degeneration. Although the tubby mutation which leads to a splicing defect of the tub gene has been identified recently, the mechanism by which it causes the obesity syndrome has not been established. In this study, the potential dysfunction of several hypothalamic neuroendocrine pathways involved in the central regulation of energy metabolism was investigated in tubby mice. In comparison with the wild-type controls, a significant reduction (20%) of pro-opiomelanocortin (POMC) mRNA expression was observed in the arcuate nucleus (ARC) of the mature, obese but not in the juvenile, non-obese tubby mice. Similarly, an age and body mass-dependent induction (about 30-fold) of neuropeptide Y (NPY) mRNA was observed in the dorsomedial (DMH) and ventromedial (VMH) hypothalamic nuclei of the tubby mice. However, NPY mRNA in the ARC was decreased by approximately 30 to 40% in both juvenile and mature tubby mice. The hypothalamic expression patterns of corticotropin releasing hormone (CRH) and the long form leptin receptor (OB-Rb) were not significantly altered in the mutant mice. These results suggest that the altered hypothalamic POMC and/or NPY functions may be important contributing factors for the development of obesity in this animal model.

Distribution of mRNA encoding the growth hormone secretagogue receptor in brain and peripheral tissues.

Growth hormone release is under tight control by two hypothalamic hormones: growth hormone-releasing hormone and somatostatin. In addition, synthetic growth hormone secretagogues have also been shown to regulate growth hormone release through the growth hormone secretagogue receptor (GHS-R), suggesting the existence of an additional physiological regulator for growth hormone release. To understand the physiological role of the GHS-R in more detail, we mapped the expression of mRNA for the receptor by in situ hybridization and RNase protection assays using rat and human tissues. In the rat brain, the major signals were detected in multiple hypothalamic nuclei as well as in the pituitary gland. Intense signals were also observed in the dentate gyrus of the hippocampal formation. Other brain areas that displayed localized and discrete signals for the receptor include the CA2 and CA3 regions of the hippocampus, the substantia nigra, ventral tegmental area, and dorsal and median raphe nuclei. In resemblance to the results from rat brain, RNase protection assays using human tissues revealed specific signals in pituitary, hypothalamus and hippocampus. Moreover, a weak signal was noted in the pancreas. The demonstration of hypothalamic and pituitary localization of the GHS-R is consistent with its role in regulating growth hormone release. The expression of the receptor in other central and peripheral regions may implicate its involvement in additional as yet undefined physiological functions.

Cloning, sequencing, and expression of the rhesus monkey beta 2 adrenergic receptor.

The beta 2 adrenergic receptor (beta 2AR) plays a key role in the signal transduction mechanism for epinephrine and norepinephrine. The gene for beta 2 adrenergic receptors has been cloned for several species, but has remained undetermined for rhesus monkey. In this study, we report the isolation of the gene encoding the rhesus beta 2AR from both cDNA and genomic DNA sources. Sequence analysis of the gene reveals an intronless open reading frame that encodes a 415-amino-acid protein. The rhesus receptor is highly homologous to that from other species, especially to the human receptor (97% sequence identity). Functional characterization by ligand binding and agonist-mediated cAMP accumulation indicates that the rhesus beta 2 receptor shares a very similar pharmacological profile with the human beta 2 receptor. Therefore, the rhesus monkey represents a valid animal model for developing therapeutic agents targeted at the corresponding human beta 2 receptor.

Induction of neuropeptide Y expression in dorsomedial hypothalamus of diet-induced obese mice.

Alterations of hypothalamic neuropeptide Y(NPY) and melanocortinergic functions in diet-induced obese (DIO) C57BL/6J mice were investigated by in situ hybridization. Compared with controls, the DIO mice displayed a profound induction (approximately 40-fold) of NPY expression in the dorsomedial (DMH) and ventromedial (VMH) hypothalamic nuclei, whereas the level of NPY mRNA in the arcuate nucleus (ARC) was reduced by 44%. The expression of pro-opiomelanocortin (POMC) and agouti-related protein was not significantly altered in the ARC of obese mice. Both excess body weight gain and altered hypothalamic NPY expression were reversible. We propose that the highly induced NPY expression in DMH and/or VMH may be a contributing etiological factor for the development of obesity and leptin resistance in the DIO mice.

Alterations of immunoreactive substance P and enkephalins in rat spinal cord after electroacupuncture.

A form of electrically-induced analgesia known as electroacupuncture was administered to rats bilaterally at the point "Huan-tiao." Compared with untreated rats, treated rats showed altered pain thresholds characterized as low, intermediate, and high. From immunocytochemical studies, the spinal cords taken from the treated rats exhibited differences in immunoreactivity for substance P (SP), methionine- and leucine-enkephalins (ME and LE respectively). By densitometry, the altered levels of immunoreactive (IR) peptides correlated with the pain thresholds in specific ways. That is, high pain threshold correlated with the visualization of increased IR-SP adn IR-LE within neuronal processes throughout the dorsal horn substantia gelatinosa. In the same specimens, decreased IR-ME could be seen. In contrast, low pain threshold correlated with decreased IR-SP and IR-ME. IR-LE showed a concomitant decrease in the medial substantia gelatinosa region, and slight, insignificant changes laterally. The data suggest that different degrees of analgesia induced by electroacupuncture result from the variable release of SP, ME, and LE in spinal regions associated with nociception. In terms of current models of pain processing, the data do not entirely support an axo-axonic interaction between enkephalin interneurons and SP terminals. Some modifications and an alternative model are considered.

Fluoxetine increases the extracellular levels of serotonin in the nucleus accumbens.

The effects of an IP injection of the monoamine uptake inhibitor fluoxetine on the extracellular concentration of serotonin (5-HT), dopamine (DA), 5-hydroxyindoleacetic acid (5-HIAA), 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in the nucleus accumbens of awake and freely moving rats were examined using a push-pull perfusion technique. Baseline values of 5-HT, 5-HIAA, DA, DOPAC and HVA in the perfusates were approximately 0.07, 13, 0.8, 49 and 12 pmol/hr, respectively. The IP administration of 5 and 10 mg/kg fluoxetine dose-dependently elevated the amounts of 5-HT 3- and 13-fold, respectively, in the push-pull perfusate, with the maximum reached within one hour after drug administration. Moreover, 10 mg/kg fluoxetine also significantly decreased the levels of 5-HIAA in the perfusate as much as 50% within 2-3 hours. On the other hand, no significant effect of 5 or 10 mg/kg fluoxetine was observed on the contents of DA, DOPAC and HVA in the push-pull perfusates. The data indicate that fluoxetine, in accord with its role as a 5-HT uptake inhibitor, increases the physiologically active pool of 5-HT in the nucleus accumbens under in vivo conditions.

Serotonin microinfusion into the ventral tegmental area increases accumbens dopamine release.

The effects of microinfusion of serotonin (5-HT) agents as well as glutamate and muscimol into the ventral tegmental area (VTA) on dopamine (DA) release in the ipsilateral nucleus accumbens (ACC) were investigated in freely moving rats, using a push-pull perfusion procedure. The baseline values for DA, 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 5-hydroxyindoleacetic acid (5-HIAA) were approximately 0.24, 8.4, 2.1 and 2.7 pmol/15 min, respectively, in the push-pull perfusate of the ACC. When microinfused into the VTA, glutamate (0.74 microgram) significantly (p less than 0.05) increased the contents of DOPAC (110%) and HVA (90%) over baseline levels in the perfusate. On the other hand, 0.5 microgram muscimol (a gamma-amino-n-butyric acid, GABA, agonist) significantly, (p less than 0.05) decreased both DA (40%) and DOPAC (20%) levels relative to baseline values. Administration of 2 micrograms 5-HT into the VTA caused a significant (p less than 0.05) elevation in the perfusate levels of DOPAC (80%) and HVA (70%) over baseline values. A similar effect was obtained with a nonselective 5-HT1 agonist but not with a selective 5-HT1A agonist. The results suggest that 5-HT innervations in the VTA may have an excitatory action possibly via 5-HT1B rather than 5-HT1A receptors on the mesolimbic DA system projecting to the ACC and that this DA system may also be regulated by glutamatergic and GABAergic (via GABAA receptors) inputs.

Amphetamine-induced in vivo release of dopamine in the striatum is influenced by local pH.

The effect of local pH on the in vivo efflux of endogenous dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) following administration of d-amphetamine (AMPH) was examined in the striatum of the anesthetized rat using two bilaterally placed push-pull cannulae. At both pH 7.3 and 6.4, the baseline efflux values for DA and DOPAC were approximately 0.2 and 25 pmoles/15 min, respectively. Subcutaneous injection of 2 mg/kg AMPH induced a 3-fold increase of DA release at pH 7.3 and a 21-fold increase of DA release at pH 6.4. In both cases, the maximum was reached at about 30 min after the drug administration. Following the administration of AMPH, the efflux of DOPAC was reduced to the same degree (20% of control values) under both pH conditions. In vitro data showed that the lower pH did not alter the recovery of DA or DOPAC. In addition, release of DA produced by local perfusion with 5 uM AMPH was also greater at the lower pH (50-fold increase over baseline) than at the physiological pH (10-fold increase over baseline). The stimulated DA release produced by local perfusion with 35 mM K+, however, was the same at both pH values. Preliminary experiments also indicated that there was a pH effect for AMPH-induced serotonin (5-HT) release but that the difference in the amount of 5-HT in the two media was not nearly as large as that obtained for DA. The markedly elevated level of extracellular DA at the lower pH might be due to a higher affinity of the DA uptake system for AMPH, thereby producing greater inhibition of DA uptake as well as enhanced DA release. The data also suggest an enhanced affinity of AMPH for 5-HT uptake sites at the lower pH.

Effects of K+-stimulation and precursor loading on the in vivo release of dopamine, serotonin and their metabolites in the nucleus accumbens of the rat.

The in vivo efflux of endogenous 3,4-dihydroxyphenylethylamine (DA 5-hydroxytryptamine (5-HT), 3,4-dihydroxyphenyl-acetic acid (DOPAC), homovanillic acid (HVA) and 5-hydroxy-indoleacetic acid (5-HIAA) in the nucleus accumbens of the anesthetized rat was studied using a push-pull cannula. Local perfusion for 10 minutes with 35 mM K+ significantly (P less than 0.01) increased the release of DA and 5-HT, but not their metabolites, from their respective control levels of 0.95 and 0.04 pmol/15 min to 2.5 and 0.23 pmol/15 min. Exposure to 35 mM K+ a second and third time resulted in a decrement in the amount of stimulated release for both DA and 5-HT. This decrease was prevented by local perfusion for 10 minutes with 50 uM L-tyrosine and -tryptophan starting 30 minutes before each episode of depolarization. The baseline amounts of DOPAC, HVA and 5-HIAA observed in the perfusates were several fold higher than the basal levels found for 5-HT and DA. In the absence of precursors, the efflux of DOPAC, HVA and 5-HIAA decreased approximately 60, 40 and 25%, respectively, from the first to the last baseline fraction collected. Addition of precursors prevented the decrease for DOPAC and 5-HIAA but not for HVA. The data indicated that (a) the in vivo release of DA and 5-HT, along with their metabolites, could be simultaneously measured with the present procedure, and (b) when using the push-pull cannula, local perfusion with precursors may be necessary following periods of sustained and/or repeated stimulation in order to replenish the monoamine transmitter pools.