NTRK expression is common in xanthogranuloma and is associated with the solitary variant.

BACKGROUND: Previously identified mutually-exclusive driver genes in juvenile xanthogranuloma (JXG) and adult xanthogranuloma (AXG) include mutations in MAP kinase pathway genes such as MAP2K1, BRAF, ARAF, KRAS, NRAS, PIK3CD as well as fusions in BRAF and ALK, with a subset of cases with no identified driver yet. NTRK fusion has been identified in rare cases. METHODS: We identified two consecutive index cases of localized JXG or AXG with NTRK1 fusion by next-generation sequencing (NGS) and confirmed by pan-NTRK immunostain. We expanded the study to a total of 50 cases of JXG and AXG using screening by pan-NTRK immunostain. We confirmed the specificity of our approach with negative results in 5 cases of histiocytic neoplasia lacking an NTRK fusion by NGS and 14 cases of non-neoplastic histiocytic disease. RESULTS: We found 23 cases of JXG or AXG with overexpression of NTRK by immunostain, and these cases were restricted to localized disease (23 of 43 cases, 53.5%) rather than disseminated disease (zero of seven cases). CONCLUSIONS: NTRK expression is common in JXG or AXG and associated with localized rather than disseminated disease. We speculate that the potential importance of this in JXG and AXG has not been previously appreciated due to the tendency to focus sequencing studies on disseminated disease. We confirm the presence of an NTRK1 fusion in two positive cases by NGS, however, additional genetic studies are necessary to further explore this.

Intra-accumbal administration of shRNAs against CART peptides cause increases in body weight and cocaine-induced locomotor activity in rats.

In order to examine the effect of cocaine and amphetamine regulated transcript (CART) peptide depletion in adult rats, CART shRNAs or scrambled control shRNAs were administered bilaterally into the nucleus accumbens (NAc). There was an increase in body weight of the shRNA injected rats compared with the rats injected with the scrambled RNA. This is compatible with the data showing a role for the peptide in body weight and food intake. Also at this time, there was about a two-and-a-half fold increase in cocaine-mediated locomotion in the shRNA injected rats compared to the control rats. This finding is critical support for the hypothesis that endogenous CART peptides in the NAc inhibit the actions of cocaine and other psychostimulants. In immunohistochemical experiments on these same animals, there was a decrease in the staining density of CART peptide in the NAc of the shRNA injected rats. These data show that shRNA can reduce CART peptides in the NAc and that endogenous CART peptides influence body weight and cocaine-induced locomotor activity (LMA).

The cocaine- and amphetamine-regulated transcript mediates ligand-independent activation of ERα, and is an independent prognostic factor in node-negative breast cancer.

Personalized medicine requires the identification of unambiguous prognostic and predictive biomarkers to inform therapeutic decisions. Within this context, the management of lymph node-negative breast cancer is the subject of much debate with particular emphasis on the requirement for adjuvant chemotherapy. The identification of prognostic and predictive biomarkers in this group of patients is crucial. Here, we demonstrate by tissue microarray and automated image analysis that the cocaine- and amphetamine-regulated transcript (CART) is expressed in primary and metastatic breast cancer and is an independent poor prognostic factor in estrogen receptor (ER)-positive, lymph node-negative tumors in two separate breast cancer cohorts (n=690; P=0.002, 0.013). We also show that CART increases the transcriptional activity of ERα in a ligand-independent manner via the mitogen-activated protein kinase pathway and that CART stimulates an autocrine/paracrine loop within tumor cells to amplify the CART signal. Additionally, we demonstrate that CART expression in ER-positive breast cancer cell lines protects against tamoxifen-mediated cell death and that high CART expression predicts disease outcome in tamoxifen-treated patients in vivo in three independent breast cancer cohorts. We believe that CART profiling will help facilitate stratification of lymph node-negative breast cancer patients into high- and low-risk categories and allow for the personalization of therapy.

Understanding the Global Problem of Drug Addiction is a Challenge for IDARS Scientists.

IDARS is an acronym for the International Drug Abuse Research Society. Apart from our scientific and educational purposes, we communicate information to the general and scientific community about substance abuse and addiction science and treatment potential. Members of IDARS are research scientists and clinicians from around the world, with scheduled meetings across the globe. IDARS is developing a vibrant and exciting international mechanism not only for scientific interactions in the domain of addiction between countries but also ultimately as a resource for informing public policy across nations. Nonetheless, a lot more research needs to be done to better understand the neurobiological basis of drug addiction - A challenge for IDARS scientists.

CART Peptides Regulate Psychostimulants and May be Endogenous Antidepressants.

CART peptides are endogenous neurotransmitters that are involved in a variety of physiologic functions. Injection of CART 55-102 into the nucleus accumbens produces no effect, but when co-administered with cocaine, it reduces the locomotor and rewarding properties of cocaine. In a human study, subjects carrying a missense mutation of the CART gene exhibited increased anxiety and depression. Also, several animal studies support the idea that CART is involved in anxiety and depression, and they also suggest several possible mechanisms by which this may occur. Thus, there is interesting evidence that CART peptides play a role in anxiety and depression, and that CART peptides may be endogenous antidepressants.

Cocaine and amphetamine-regulated transcript-containing neurons in the nucleus accumbens project to the ventral pallidum in the rat and may inhibit cocaine-induced locomotion.

We have previously demonstrated that cocaine- and amphetamine-regulated transcript (CART) peptide colocalizes with GABA, dynorphin, D1 receptors, and substance P in some neurons in the nucleus accumbens (NAcc). One of the main nuclei that receive accumbal efferents is the ventral pallidum (VP), and both dynorphin and substance P have been shown to be present in the cell bodies and terminals of this projection. Thus, we investigated whether CART peptide is also present in the VP in terminals that originate in the accumbens. The anterograde tracer Phaseolus vulgaris leukoagglutinin (PHA-L) colocalized with CART in neuronal processes in the VP when injected into the NAcc. Also, CART colocalized with the retrograde tracer r-BDA in accumbens cell bodies after the tracer was injected into the VP. Using electron microscopic immunocytochemistry, we examined CART terminals in the VP and found that CART-immunoreactive terminals formed symmetric synapses consistent with inhibitory GABAergic synapses. These synapses closely resemble GABAergic synapses in the substantia nigra pars reticulata (SNr), another nucleus that receives some CART-containing accumbal efferents. Lastly, we found that intra-pallidal injection of CART 55-102 inhibited cocaine-induced locomotion, indicating that CART peptide in the VP can have functional effects.

Regulation of cocaine- and amphetamine-regulated transcript mRNA expression by calcium-mediated signaling in GH3 cells.

Cocaine- and amphetamine-regulated-transcript (CART) peptides are associated with multiple physiological processes, including, feeding, body weight, and the response to drugs of abuse. CART mRNA and peptide levels and the expression of the CART gene appears to be under the control of a number of extra- and intra-cellular factors including the transcription factor, cAMP response element binding protein (CREB). Similar to the effects of CART, Ca(2+) signaling leads to the phosphorylation of CREB and has been associated with both feeding and the actions of psychostimulants; therefore, we hypothesized that Ca(2+) may play a role in CART gene regulation. We used real-time PCR (rtPCR) and GH3 cells to examine the effect of ionomycin, which increases intracellular Ca(2+), on CART mRNA levels. Ionomycin increased CART mRNA in a dose- and time-dependent manner. The effect of ionomycin appeared transient as CART mRNA had returned to control levels 3 h following treatment. Calmidazolium and KN93, inhibitors of calmodulin and Ca(2+)-modulated protein (CaM) kinases respectively, attenuated the effect of ionomycin (10 microM) on CART mRNA levels suggesting a calmodulin-dependent mechanism. Western immunoblotting indicated that ionomycin increased phosphorylated cAMP response element binding protein (pCREB) levels and electrophoretic mobility shift assay/supershift assay using antibodies against pCREB demonstrated increased levels of a CART oligo/pCREB protein complex. Finally, we showed that injection of ionomycin into the rat nucleus accumbens increases CART mRNA levels. To our knowledge, this is the first study providing evidence that the CART gene is, in part, regulated by Ca(2+)/CaM/CREB-dependent cell signaling.

CART peptides: regulators of body weight, reward and other functions.

Over the past decade or so, CART (cocaine- and amphetamine-regulated transcript) peptides have emerged as major neurotransmitters and hormones. CART peptides are widely distributed in the CNS and are involved in regulating many processes, including food intake and the maintenance of body weight, reward and endocrine functions. Recent studies have produced a wealth of information about the location, regulation, processing and functions of CART peptides, but additional studies aimed at elucidating the physiological effects of the peptides and at characterizing the CART receptor(s) are needed to take advantage of possible therapeutic applications.

Unique responses of midbrain CART neurons in macaques to ovarian steroids.

CART (cocaine and amphetamine regulated transcript) is a neuropeptide involved in the control of several physiological processes, such as response to psychostimulants, food intake, depressive diseases and neuroprotection. It is robustly expressed in the brain, mainly in regions that control emotional and stress responses and it is regulated by estrogen in the hypothalamus. There is a distinct population of CART neurons located in the vicinity of the Edinger-Westphal nucleus of the midbrain that also colocalize urocortin-1. The aims of this study were 1) to determine the distribution of CART immunoreactive neurons in the monkey midbrain, 2) to examine the effects of estrogen (E) and progesterone (P) on midbrain CART mRNA and peptide expression and 3) to determine whether midbrain CART neurons contain steroid receptors. Adult female rhesus monkeys (Macaca mulatta) were spayed and either treated with placebo (OVX), estrogen alone (E), progesterone alone (P) or E+P. Animals were prepared (a) for RNA extraction followed by microarray analysis and quantitative (q) RT-PCR (n=3/group); (b) for immunohistochemical analysis of CART and CART+tryptophan hydroxylase (TPH), CART+estrogen receptors (ER) or CART+progesterone receptors (n=5/group) and (c) for Western blots (n=3/group). Both E- and E+P-administration decreased CART gene expression on the microarray and with qRT-PCR. Stereological analysis of CART immunostaining at five levels of the Edinger-Westphal nucleus indicated little effect of E or E+P administration on the area of CART immunostaining. However, P administration increased CART-immunopositive area in comparison to the OVX control group with Student's t-test, but not with ANOVA. CART 55-102 detection on Western blot was unchanged by hormone administration. ERbeta and PR were detected in CART neurons and CART fibers appeared to innervate TPH-positive serotonin neurons in the dorsal raphe. In summary, E decreased CART mRNA, but this effect did not translate to the protein level. Moreover, P administration alone had a variable effect on CART mRNA, but it caused an increase in CART immunostaining. Together, the data suggest that CART neurons in the midbrain have a unique steroid response, which may be mediated by nuclear receptors, neuroactive steroids or interneurons.

CART deficiency increases body weight but does not alter bone strength.

The regulation of bone metabolism mediated by leptin is a complex process that is not clearly understood. Recent studies suggest that CART (cocaine-amphetamine related transcript) is a significant neuronal co-factor when combined with leptin. CART deficiency is thought to result in low trabecular bone mass, but since leptin exerts contrasting effects on trabecular and cortical bone it is possible that cortical bone may not respond to the absence of CART signaling in the same manner as trabecular bone. We tested the hypothesis that CART deficiency decreases cortical bone mass, density, and strength by examining femora of adult wild-type mice (CART(+/+)) and CART-deficient mice (CART(-/-)). DEXA densitometry (PIXImus system) was used to measure whole-bone mineral content (BMC) and mineral density (BMD) from right femora, and pQCT used to calculate densitometric and geometric parameters from the femur midshaft. Femora were also tested in three-point bending, and sections of the tibia analyzed histologically to determine bone marrow adipocyte density (N.At./M.Ar) and endocortical osteoclast number (N.Oc/B.Pm). The control mice weighed less than the mice lacking CART (P<0.001), but mechanical testing data showed no differences (p>0.05) in ultimate force, energy to fracture, stiffness, or intrinsic properties such as ultimate stress, ultimate strain, or modulus. CART-deficient mice did not differ from normal controls in whole-femur BMC (p=0.09), BMD (p=0.19), midshaft cortical bone thickness (p=0.67), midshaft cortical bone area (p=0.59) or N.Oc/B.Pm (p=0.94), although CART deficiency was associated with a three-fold increase in bone marrow adipocyte density (p<0.001). Our data suggest that while the central, neuroendocrine regulation of bone mass via CART signaling may have effects on trabecular mass, absence of CART expression does not significantly alter cortical bone geometry, density, or strength.

Frequency of maternal licking and grooming correlates negatively with vulnerability to cocaine and alcohol use in rats.

Because licking and grooming behavior of dams with pups can influence some behaviors of pups when they are adults, we tested if licking and grooming scores in a maternal separation protocol correlated with cocaine or ethanol self-administration in the pups as adults. The protocol produced litters that were separated from dams for 0 (MS0), 15 (MS15) or 180 (MS180) min, and a nonhandled (NH) group as well. Self-administration of both drugs as shown in earlier studies was lowest in the MS15 group, highest in the NH group and intermediate in the other groups. Licking and grooming scores correlated negatively with drug intake and suggests that maternal care of pups can influence drug use in pups when they are adults.

Cocaine administration increases the fraction of CART cells in the rat nucleus accumbens that co-immunostain for c-Fos.

In order to further test whether or not psychostimulant drugs activate CART peptide-containing cells in the nucleus accumbens, we examined the fraction of CART positive cells that co-immunostained for c-Fos after administration of saline or cocaine (10 and 25 mg/kg i.p.). There was about a 45% increase in the fraction of cells that stained for both CART and c-Fos after administration of cocaine, but there was no change in the fraction after administration of saline. Moreover, the increase was not found 24h after injection and is therefore reversible. These results support the notion that psychostimulant drugs activate CART cells in the nucleus accumbens, even under conditions where it is difficult to show a change in CART levels.

High-energy-resolution inelastic electron and proton scattering and the multiphonon nature of mixed-symmetry 2+ states in 94Mo.

High-energy-resolution inelastic electron scattering (at the S-DALINAC) and proton scattering (at iThemba LABS) experiments permit a thorough test of the nature of proposed one- and two-phonon symmetric and mixed-symmetric 2+ states of the nucleus 94Mo. The combined analysis reveals the one-phonon content of the mixed-symmetry state and its isovector character suggested by microscopic nuclear model calculations. The purity of two-phonon 2+ states is extracted.

CART-peptide immunoreactivity in enteric nerves in patients with Hirschsprung's disease.

AIMS: Cocaine- and amphetamine-regulated transcript (CART)-peptide is found in the brain and participates in the control of feeding behavior. It is also expressed in the peripheral nervous system and is suggested to have neuromodulatory and/or neurotrophic effects in rat intestine. The aims of this study were to investigate the presence of CART-peptide in the normal ganglionic as well as aganglionic intestine from patients with Hirschsprung's disease and the peptide's possible coexistence with other neurotransmitters. METHODS: Intestinal specimens from nine patients with Hirschsprung's disease were examined using immunohistochemistry. A double immunostaining technique was used in order to elucidate the presence of CART-peptide in NOS and VIP-containing enteric neurons. RESULTS: In ganglionic intestine, CART-peptide was found in numerous nerve fibers, predominantly within the smooth muscle layers and in myenteric nerve cell bodies. A high degree of co-localization of CART with NOS and VIP was seen. Only very few CART immunoreactive nerve fibers and no nerve cell bodies were found in the aganglionic intestine. CONCLUSIONS: This is the first report on the presence of CART-peptide in the human intestine. In the ganglionic intestine CART was detected mainly in myenteric neurons, while only very few CART-IR nerve fibers were found in the aganglionic intestine. This, together with the coexistence of CART with NOS and VIP, indicates an intrinsic origin of the CART-containing neurons and suggests that CART may influence NO and VIP-induced effects.

Histogenesis of sarcomatoid urothelial carcinoma of the urinary bladder: evidence for a common clonal origin with divergent differentiation.

The histogenesis of sarcomatoid urothelial carcinoma, a rare neoplasm with bidirectional epithelial and mesenchymal differentiation, has been a matter of controversy. To clarify its origin, we analysed the status of X-chromosome inactivation in sarcomatoid urothelial carcinomas from 10 female patients and examined losses of heterozygosity (LOH) in these specimens and in additional 20 tumours from male patients. Six polymorphic microsatellite markers where genetic alterations occur frequently in early or advanced stages of urothelial carcinomas, including D3S3050, D8S261, IFNA, D9S177, D11S569 and TP53, were investigated in the current study. The identical pattern of non-random X-chromosome inactivation in both carcinomatous and sarcomatous components was identified in five of eight informative female patients, and the remaining three informative cases showed a random, but concordant, pattern of X-chromosome inactivation. The concordant X-chromosome inactivation results in all eight informative cases support the concept of a monoclonal origin of both components of this biphasic neoplasm. Among the tumours demonstrating loss of heterozygosity, high incidences of an identical pattern of allelic loss between carcinomatous and sarcomatous components were identified in genetic alterations associated with early carcinogenesis: 86% at D8S261, 78% at D11S569, 75% at D9S177 and 57% at IFNA. In contrast, concordant LOH patterns were less frequently observed for microsatellites related to advanced carcinogenesis: only 40% at D3S3050 and 40% at TP53. The significant overlap of loss of heterozygosity supports a monoclonal cell origin and suggests that clonal divergence may occur during tumour progression and differentiation. Divergent patterns of discordant allelic loss of microsatellite markers imply that heterogeneous pathogenetic pathways may exist in the evolution of this enigmatic neoplasm.

Maternal separation alters drug intake patterns in adulthood in rats.

Maternal separation/handling (MS/H) is an animal model of early life stress that causes profound neurochemical and behavioral alterations in pups that persist into adulthood. Many recent studies have used the MS/H model to study changes in drug effects in adulthood that are linked to behavioral treatments and stressors in the perinatal period. The drug effects focused on in this review are the reinforcing properties of the abused drugs, cocaine and alcohol. A striking finding is that variations in maternal separation and handling cause changes in ethanol and cocaine self-administration. Further, these changes indicate that various manipulations in the perinatal period can have long lasting effects of interest to biochemical pharmacologists. This article will review recent studies on ethanol and cocaine self-administration using the MS/H model and the neurochemical alterations that may play a role in the effects of MS/H on ethanol and cocaine self-administration. Studying the MS/H model can provide important clues into the vulnerability to drug abuse and perhaps identify a crucial window of opportunity for therapeutic intervention.

Maternal separation alters serotonergic transporter densities and serotonergic 1A receptors in rat brain.

RATIONALE: The basic mechanisms underlying the association between early life maternal separation and adulthood psychiatric disorders are largely unknown. One possible candidate is the central serotonergic system, which is also abnormal in psychiatric illnesses. Neuroadaptational changes in serotonergic transporter and serotonergic 1A receptors may underlie links between early life stress and adulthood psychiatric disorders. OBJECTIVE: The aim of this study was to investigate the consequences of a rat model of maternal separation on serotonergic transporter and serotonergic 1A receptor densities and function in adult rat forebrain. METHODS: Rat pups were separated from dams from postnatal day 2 to postnatal day 14, each day, for zero time, 15 min and 180 min to determine the time-course of effects. A non-handled group was added to control for the effects of handling by an experimenter compared with the animal facility-reared group. Quantitative [(125)I]3beta-(4-iodophenyl)tropan-2beta-carboxylic acid methyl ester and [(125)I]-mPPI autoradiography was used to determine serotonergic transporter and serotonergic 1A densities, respectively. Adult rats were challenged with saline or serotonergic 1A agonist (+) 8-hydroxy-2-(di-n-propylamino)tetralin, 0.4 mg/kg, s.c.) and plasma adrenocorticotropic hormone and corticosterone were determined. RESULTS: serotonergic transporter and serotonergic 1A densities were significantly lower in the non-handled group in the paraventricular, arcuate, dorsomedial and ventromedial nuclei of the hypothalamus. The non-handled group also displayed lower serotonergic transporter and serotonergic 1A densities in the basolateral anterior, basolateral ventral and basomedial amygdaloid nuclei. Serotonergic transporter densities were also decreased in the CA3 area of the hippocampus in the non-handled group. In contrast, the maternal separation 15 min group displayed the highest serotonergic transporter and serotonergic 1A densities in the basomedial nucleus of amygdala, basolateral anterior nucleus of amygdala, basolateral ventral nucleus of amygdala and basomedial nucleus of amygdala amygdaloid nuclei. CONCLUSIONS: Early life maternal separation and the extent of handling can alter adult brain serotonergic transporter and serotonergic 1A levels and function in the forebrain. Alterations in these serotonergic systems by early rearing conditions might increase vulnerability for behavioral disorders in adulthood.

Adolescents carrying a missense mutation in the CART gene exhibit increased anxiety and depression.

A Leu34Phe mutation in the cocaine- and amphetamine-regulated transcript (CART) gene has been associated with severe early-onset obesity in an affected family. It has been shown that, in a cell culture system, the mutation resulted in altered CART peptide levels, and animal studies suggest that CART is involved in anxiety as well. The availability of the affected family allowed for testing of anxiety and other traits in humans carrying a mutation in CART gene. This study shows that a small group of adolescents with the mutation exhibit higher anxiety and depression scores than control subjects.

Anti-phospholipid antibodies following vaccination with recombinant hepatitis B vaccine.

This study was undertaken to evaluate the possible role of hepatitis B recombinant vaccine inducing the synthesis of IgG and IgM anti-cardiolipin antibodies (aCL), antibodies against beta(2)GPI (anti-beta(2)GPI), lupus anti-coagulant (LA), anti-nuclear antibodies and antibodies against extractable nuclear antigens (anti-ENA). The study population consisted of 85 healthy students (63 female, 22 male; mean age 20.8 years), vaccinated with three doses of recombinant DNA hepatitis B vaccine. One month after vaccination with the first dose of hepatitis B vaccine a minority of vaccinated individuals showed changes in IgG or IgM aCL or anti-beta(2)GPI or LA activity (P < 0.001). Among subjects in whom changes of IgG anti-beta(2)GPI were observed, a significantly higher number of increased (8/85) than decreased (2/85) values were found (P < 0.01). Analyses of paired data showed that differences in aCL or anti-beta(2)GPI levels before vaccination or 1 month later did not reach statistical significance. In two people aCL transitorily reached medium positivity after the first dose of hepatitis B vaccine with a drop 5 months later. Similar evident anti-beta(2)GPI fluctuation was also observed in one person. Another participant was initially low positive for IgG anti-beta2GPI and the levels were increasing after vaccination. Two participants became positive for anti-nuclear antibodies during 6 months' follow-up. There were no sex-dependent differences in tested antibodies observed and no associations between levels of aPL and levels of anti-HBV antibodies. We conclude that HBV can induce aPL, although rarely. In genetically susceptible individuals or together with some other triggers such combination might confer the risk of developing a continuous autoimmune response in an individual.

A cocaine-and-amphetamine-regulated-transcript peptide projection from the lateral hypothalamus to the ventral tegmental area.

Cocaine-and-amphetamine-regulated-transcript peptides play a role in the modulation of feeding and psychomotor stimulant-like behaviors. The ventral tegmental area and the lateral hypothalamus are likely structures where cocaine-and-amphetamine-regulated-transcript peptides mediate both of these functions. Although lateral hypothalamus inputs to the ventral tegmental area have long been known, the chemical nature of this pathway remains poorly understood. To address this issue, we tested the possibility that cocaine-and-amphetamine-regulated-transcript peptide-containing neurons in the lateral hypothalamus project to the ventral tegmental area using the retrograde transport of cholera toxin subunit B combined with cocaine-and-amphetamine-regulated-transcript peptide immunostaining. The largest density of retrogradely-labeled neurons in the hypothalamus after cholera toxin subunit B injection in the ventral tegmental area was found, ipsi- and contralaterally, in the lateral hypothalamus/perifornical area, although substantial numbers of retrogradely-labeled cells were also found in the medial preoptic area, lateral preoptic area, paraventricular nucleus, dorsomedial hypothalamus and ventromedial hypothalamus. More than 80% of the retrogradely-labeled cocaine-and-amphetamine-regulated-transcript peptide-immunoreactive neurons in the hypothalamus were found in the lateral hypothalamus/perifornical area both ipsilateral and contralateral to the injection sites. Although retrogradely-labeled neurons were seen in the amygdala, locus coeruleus, and raphe nucleus, none of them displayed cocaine-and-amphetamine-regulated-transcript peptide immunoreactivity. Therefore, the hypothalamic projection to the ventral tegmental area provides a substrate whereby cocaine-and-amphetamine-regulated-transcript peptides could mediate the rewarding aspects of feeding and psychomotor stimulant-like behaviors. These findings, combined with the fact that the lateral hypothalamus receives strong inputs from the shell of the nucleus accumbens and ventral pallidum, suggest that these structures are part of integrative functional loops that control reward and appetitive behaviors.