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N-acyl-phosphatidylethanolamine phospholipase D (NAPE–PLD) is considered to be the principal enzyme that produces N-acylethanolamines (NAEs), a family of signaling lipids. NAEs are involved in numerous physiological processes such as appetite, satiety, pain, inflammation, fertility, stress, and anxiety. Furthermore, aberrant NAE levels are associated with metabolic syndrome and non-alcoholic steatohepatitis (NASH). Several inhibitors for NAPE–PLD have been reported. But most of the inhibitors showed poor to moderate potency for NAPE–PLD in vitro. Recently, Mario van der Stelt et al describe the SAR of NAPE–PLD inhibitors that afforded LEI–401 in vitro. However, no attempt was instigated to produce a consensus pharmacophore model of LEI–401 as inhibitors of NAPE–PLD. Pharmacophore modeling is an efficient and useful approach to identify important patterns in a series of molecules for optimizations. The consensus pharmacophore model revealed the importance of structural features and their correlation with the biological activity. &nbsp

Objective To determine if models of human 'receptive' and 'non-receptive endometrium' differ in their responses to nitric oxide (NO) supplementation by measuring the levels of the enzymes of the endocannabinoid system (ECS) (fatty acid amide hydrolase (FAAH) and N-acylphosphatidylethanolamine-specific phospholipase D (NAPE-PLD)), which control the 'anandamide tone' essential for successful pregnancy. Design A study of FAAH and NAPE-PLD expression (using human endometrium) through the menstrual cycle and an in vitro using a model of 'receptive' (Ishikawa) and 'non-receptive' (HEC-1A) human endometrial cell lines treated with the NO-donating compound S-nitroso-N-acetylpenicillamine (SNAP). Results Immunoreactivity measured by optimised H-score for both FAAH and NAPE-PLD was reduced in secretory (receptive) endometrium compared to proliferative (non-receptive) endometrium (P = 0.0009 and FAAH and NAPE transcript levels were significantly higher in untreated Ishikawa cells than in HEC-1A cells (P = 0.0228 and 0.0001, respectively). Treatment of cultures with SNAP resulted in an increase in the amount of FAAH mRNA produced by Ishikawa cells and a decrease in NAPE-PLD mRNA. No effect of SNAP was observed in HEC-1A cells. Similarly, FAAH protein was significantly decreased in endometria representative of the receptive endometrium. Conclusion These data suggest that NO most likely affects the expression of ECS enzymes in the implantation site of a receptive endometrium; a phenomenon not seen in a non-receptive endometrium. These effects are most marked with FAAH expression, suggesting that FAAH may play the more critical role in ensuring the correct 'anandamide tone' for successful embryo implantation than NAPE-PLD. Lay summary Embryo implantation into the wall of the uterus is only successful when the inner wall of the uterus (the endometrium) is ‘receptive’, because if it is ‘non-receptive’, implantation will fail. Previous work showed that enzymes of the 'endocannabinoid system' are critical for implantation by maintaining the correct level of a fat called anandamide. This is by balancing its synthesis (by N-acylphosphatidylethanolamine specific phospholipase D, NAPE-PLD) and degradation (by fatty acid amide hydrolase, FAAH). Using immortalised cell lines as models of ‘receptive’ and ‘non-receptive’ human endometrium, we demonstrate a key stimulator of implantation, nitric oxide, has a positive effect on implantation by both increasing the mRNA levels of the degrading enzyme (FAAH) and decreasing the expression of the synthesising enzyme (NAPE-PLD). These effects are most marked with the degrading enzyme, suggesting that FAAH plays a more critical role than NAPE-PLD in ensuring the correct 'anandamide tone' for successful embryo implantation.

Lichen simplex chronicus (LSC) of the vulva is an acquired, persistent, severely itchy dermatosis usually located in easily reachable areas of the body like the extremities, nape of the neck, and genitalia. It also has a strong association with atopy and is said to be initiated and perpetuated by the itch-scratch-itch cycle. Two cases of rugose labia as the sole presentation of LSC of vulva are being presented.

Background: Histone deacetylase (HDAC) inhibition has been found to be effective in the treatment of inflammatory bowel disease. Previous studies have reported that Cinnamyl sulfonamide hydroxamate derivatives possess non-selective HDAC inhibition. Objective: The present study was designed to screen three selected Cinnamyl sulfonamide hydroxamate derivatives, NMJ-1, NMJ-2, and NMJ3, for in vitro anti-inflammatory response by assessing the expression of pNF-κB in lipopolysaccharide (LPS)-induced inflammatory changes on RAW 264.7 cells, and in vivo anti-inflammatory response in acetic acid (AA) and 2.4-dinitrochlorobenzene (DNCB)-induced colitis models in Wistar rats. Method: AA-induced colitis was produced in Wistar rats by intra-colonic administration of 1 ml AA. DNCBinduced colitis was produced by spraying 250 μL DNCB in acetone (20g/L) on the nape of the rats for 14 days, followed by the intracolonic administration on day 15. Drugs were administered for three days after the induction of colitis. Results: In vitro anti-inflammatory effect was observed by NMJ1 and NMJ2 through a significant decrease in pNF-κB overexpression-induced by LPS. Similar effect was observed in anti-colitis response by NMJ2 in both models by reversing the colitis-induced changes in length, weight, anti-oxidant profile and histopathology of the colon. Conclusion: NMJ2 was found to be most effective among the tested compounds as an anti-inflammatory agent in both in vitro and in vivo inflammatory studies.

1,2-diacyl-sn-glycero-3-phospho-N-acyl-ethanolamines (NAPE) are low abundance phospholipids but important constituents of intracellular membranes of plant tissues, responsible to generate bioactive...

BACKGROUND Receptor interacting protein kinase 1 (RIPK1)-mediated cell death, including apoptosis and necroptosis, belongs to programmed cell death. It has been reported that RIPK1-mediated necroptosis exists in lesions of cerebral hemorrhage (CH). Electroacupuncture, a treatment derived from traditional Chinese medicine, could improve neurological impairment in patients with brain injury. AIM To investigate the protective role of cross electro-nape acupuncture (CENA) in CH, and clarify the potential mechanism. METHODS CH rat models were established, and CENA was applied to the experimental rats. Neurological functions and encephaledema were then measured. Necrotic cells in the brain of rats with CH were evaluated by propidium iodide staining. Necroptosis was assessed by immunofluorescence. Activation of the necroptosis-related pathway was detected by western blot. Extraction of brain tissue, cerebrospinal fluid and serum samples was conducted to measure the expression and secretion of inflammatory cytokines by quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay. RESULTS The necroptotic marker p-MLKL was detectable in the brains of rats with CH. Next, we found that CENA could ameliorate neurological functions in rat models of CH. Moreover, the upregulation of RIPK1-mediated necroptosis-related molecules in the brains of rats with CH were inhibited by CENA. Further investigation revealed that CENA partially blocked the interaction between RIPK1 and RIPK3. Finally, in vivo assays showed that CENA decreased the expression of the inflammatory cytokines tumor necrosis factor-α, interleukin-6 and interleukin-8 in CH rat models. CONCLUSION These findings revealed that CENA exerts a protective role in CH models by inhibiting RIPK1-mediated necroptosis.

Acne keloidalis nuchae (AKN) is a chronic inflammatory condition that almost exclusively affects the occipital and nape areas. Although not completely understood, its etiopathogenesis seems to be multifactorial, including association with metabolic syndrome (MetS). Despite being commonly seen in patients with MetS, obesity per se as a possible related factor for AKN has yet to be studied. The aim of this study was to evaluate the relationship between obesity and AKN in a series of patients. Eight male patients, with a median age of 38 years (range 15–48), were included. Overweight/obesity was found in 8/8 (100%), with median BMI of 32.2 (range 27.7–43.85 kg/m2), including 2 patients with class 3 obesity. We hypothesize adipose tissue accumulation in the nape, the only moving part of the scalp, leads to redundant skin folds, more friction, and inflammation, triggering AKN. This paper highlights the possible relationship between AKN and overweight/obesity, hypothesizing a mechanism for their contribution to the etiopathogenesis of this scalp disorder. To the best of our knowledge, this is the first study focused specifically in this association. While general physicians should pay attention to the appearing of AKN in overweight/obese patients, dermatologists must be aware that AKN patients should be assessed beyond the skin.

Arachidonylethanolamide (anandamide) acts as an endogenous ligand of cannabinoid receptors, while other N-acylethanolamines (NAEs), such as palmitylethanolamide and oleylethanolamide, show analgesic, anti-inflammatory, and appetite-suppressing effects through other receptors. In mammalian tissues, NAEs, including anandamide, are produced from glycerophospholipid via N-acyl-phosphatidylethanolamine (NAPE). The ɛ isoform of cytosolic phospholipase A2 (cPLA2) functions as an N-acyltransferase to form NAPE. Since the cPLA2 family consists of six isoforms (α, β, γ, δ, ɛ, and ζ), the present study investigated a possible involvement of isoforms other than ɛ in the NAE biosynthesis. Firstly, when the cells overexpressing one of the cPLA2 isoforms were labeled with [14C]ethanolamine, the increase in the production of [14C]NAPE was observed only with the ɛ-expressing cells. Secondly, when the cells co-expressing ɛ and one of the other isoforms were analyzed, the increase in [14C]N-acyl-lysophosphatidylethanolamine (lysoNAPE) and [14C]NAE was seen with the combination of ɛ and γ isoforms. Furthermore, the purified cPLA2γ hydrolyzed not only NAPE to lysoNAPE, but also lysoNAPE to glycerophospho-N-acylethanolamine (GP-NAE). Thus, the produced GP-NAE was further hydrolyzed to NAE by glycerophosphodiesterase 1. These results suggested that cPLA2γ is involved in the biosynthesis of NAE by its phospholipase A1/A2 and lysophospholipase activities.

Este trabalho tem como objetivo principal aprimorar uma ferramenta computacional de predição de cobertura radioelétrica para simular casos práticos de uma campanha de medições em campo, com o intuito de comparar as previsões com os valores reais, avaliando dessa forma o desempenho do algoritmo que utiliza o método DMFT-SSPE. Dentre as melhorias efetuadas, é possível destacar dois eixos estruturantes. O primeiro sobre o desenvolvimento de metodologias simples e eficientes para extrair parâmetros de cenários realísticos, como diagrama de radiação de antenas comerciais e perfis topográficos de qualquer localidade. O segundo sobre a implementação de uma técnica que adiciona efeitos de retroespalhamento do campo eletromagnético, que é fundamental quando se analisa a propagação em terrenos irregulares, tornando assim os resultados computacionais mais precisos. A campanha de medições integra um estudo de engenharia realizado ao longo de uma via férrea de metrô, em ambiente urbano, que mensurou a cobertura de rádio na faixa de frequência de VHF. Nela foram utilizados equipamentos profissionais que coletaram a base de dados necessária para o desenvolvimento deste trabalho. Através de uma análise estatística dos níveis de potência recebida e da rugosidade dos enlaces foi possível tanto avaliar a acurácia das soluções NAPE e WAPE quanto direcionar esforços para pesquisas futuras. Por fim, mas não menos importante, a criação de uma interface gráfica do usuário, para apresentação dos parâmetros de entrada e dos resultados de saída da simulação, finaliza as contribuições deste estudo. Palavras-chave: DMFT-SSPE, retroespalhamento, rugosidade, NAPE, WAPE, VHF, casos práticos, predição de cobertura. The main objective of this work is to improve a computational tool of radio coverage prediction to simulate practical cases of a measurement campaign, in order to compare the predictions with real values, evaluating the performance of the algorithm that uses the DMFT-SSPE method. Among the improvements made, it is possible to highlight two structuring topics. The first is the development of simple and efficient methodologies to extract parameters from realistic scenarios, such as radiation pattern of commercial antennas and topographic profiles of any location. The second is the implementation of a technique that considers backscattering effects of the electromagnetic field, which is very important when analyzing the propagation in irregular terrain, making the computational results more accurate. The measurement campaign is part of an engineering study done along a subway track in an urban environment, which measured the radio coverage in the VHF frequency range. Professional equipment was used to collect the database needed for the development of this work. Through a statistical analysis of received power levels and terrain roughness it was possible to evaluate the accuracy of the NAPE and WAPE solutions and direct future research efforts. Last but not least, the creation of a graphical user interface, for displaying the input parameters and output results, concludes the contributions of this study. Keywords: DMFT-SSPE, backscattering, roughness, NAPE, WAPE, VHF, practical cases, coverage prediction.

Dermatomyofibroma is a rare cutaneous mesenchymal tumor of benign fibroblastic and myofibroblastic derivations. It predominantly affects young women, and it usually presents as a reddish-brown plaque or nodule, which is commonly located over the upper trunk. We report the case of a 41-year-old female patient who presented with progressive linear dermatomyofibroma over the nape of her neck. This case report expands the knowledge about the clinical and histopathological features of this rare, benign and cutaneous tumor.

Endocannabinoids and endocannabinoid-related compounds (ERCs) are involved in many physiological processes. They are released on demand from phosphoinositide and N-acylphosphatidyl ethanolamine (NAPE) precursors and comprise 2-monoacylglycerols (2-MGs) and FA ethanolamides (FEAs). Despite the abundance of advanced quantitative methods, however, their determined concentrations in blood plasma are inconsistent because 2-MGs and FEAs undergo artifactual de novo formation, chemical isomerization, and degradation during sample collection and storage. For a comprehensive survey of these compounds in blood and plasma, we have developed and validated an ultra-HPLC-MS/MS method to quantify 24 endocannabinoids, ERCs, and their phospholipid precursors. Immediate acidification of EDTA-blood to pH 5.8 blocked artifactual FEA formation for at least 4 h on ice. The 2-MGs were stabilized after plasma harvest with 0.5 M potassium thiocyanate at pH 4.7. FEA and MG plasma concentrations in six healthy volunteers ranged between 0.04–3.48 and 0.63–6.18 ng/ml, respectively. Interestingly, only 1–5% of circulating FEAs were present in their free form, while the majority was bound to NAPEs. Similarly, 97% of 2-arachidonoylglycerol (2-AG) was bound to a potential phosphoinositide pool. The herein-described stabilization and extraction methods may now be used to reliably and comprehensively quantify endocannabinoids, ERCs, and their phospholipid precursors in clinical studies.

Adverse alterations in the composition of the gut microbiota have been implicated in the development of obesity and a variety of chronic diseases. Re-engineering the gut microbiota to produce beneficial metabolites is a potential strategy for treating these chronic diseases. N-acyl-phosphatidylethanolamines (NAPEs) are a family of bioactive lipids with known anti-obesity properties, Previous studies showed that administration of Escherichia coli Nissle 1917 (EcN) engineered with Arabidopsis thaliana NAPE synthase to produce NAPEs imparted resistance to obesity induced by a high fat diet that persisted after ending their administration. In prior studies, mice were pre-treated with ampicillin prior to administering engineered EcN for 8 weeks in drinking water. If use of antibiotics and long-term administration are required for beneficial effects, implementation of this strategy in humans might be problematic. Studies were therefore undertaken to determine if less onerous protocols could still impart persistent resistance and sustained NAPE biosynthesis. Administration of engineered EcN for only two weeks without pre-treatment with antibiotics sufficed to establish persistent resistance. Sustained NAPE biosynthesis by EcN was required as antibiotic treatment after administration of the engineered EcN markedly attenuated its effects. Finally, heterologous expression of human Phospholipase A/acyl-transferase-2 (PLAAT2) in EcN provided similar resistance to obesity as heterologous expression of A. thaliana NAPE synthase, confirming that NAPEs are the bioactive mediator of this resistance.

To observe the clinical efficacy of Gao’s nape acupuncture plus swallowing training in treating pharyngeal deglutition disorder after stroke. One hundred patients with post-stroke pharyngeal deglutition disorder were randomized into a treatment group and a control group, with 50 cases in each group. The two groups both received routine neurological intervention. In addition, the treatment group was given Gao’s nape acupuncture plus swallowing training, while the control group was intervened by swallowing training alone. After eight-week treatment, the two groups were observed in terms of the changes in repetitive saliva swallowing test (RSST), modified water swallowing test (MWST), standardized swallowing assessment (SSA) and swallowing-related quality of life (SWAL-QOL). The clinical efficacies of the two groups were also compared. After treatment, the RSST grading, and scores of MWST, SSA and SWAL-QOL changed significantly in both groups (P

Some patients with hidradenitis suppurativa (HS) develop severe inflammatory lesions of the nape. Through a single-center, cross-sectional study with a total of 377 patients, we sought to compare patients with and without nape involvement, to determine whether disease severity is greater in these patients and to describe their clinical characteristics. Thirty patients (90% male) were identified as having nape involvement. Patients with versus without nape involvement had greater disease severity, earlier disease onset, a family history of HS, lower body mass, and higher Dermatological Life Quality Index and pain scores. Amongst them, involvement of the trunk and gluteal regions predominated. A significant univariate relationship existed between patients with nape involvement and male sex, early disease onset, family history, scalp and gluteal involvement, Canoui-Poitrine phenotypes II and III, and Dowling-Degos disease. A significant multivariate relationship existed with early disease onset, Dowling-Degos disease, and phenotypes II and III.

N-acylethanolamines (NAEs) are lipid signaling mediators, which can be synthesized from dietary fatty acids via n-acylphosphatidylethanolamine-phospholipase D (NAPE-PLD) and in turn influence physiological outcomes; however, the roles of NAPE-PLD upon dietary fatty acid modulation are not fully understood. Presently, we examine if NAPE-PLD is necessary to increase NAEs in response to dietary fatty acid manipulation. Post-weaning male wild-type (C57Bl/6), NAPE-PLD (-/+) and NAPE-PLD (-/-) mice received isocaloric fat diets containing either beef tallow, corn oil, canola oil or fish oil (10% wt/wt from fat) for 9 weeks. Brain docosahexaenoic acid (DHA) levels were higher (P.01) in NAPE-PLD (-/+) (10.01±0.31 μmol/g) and NAPE-PLD (-/-) (10.89±0.61 μmol/g) than wild-type (7.72±0.61 μmol/g) consuming fish oil. In NAPE-PLD (-/-) mice, brain docosahexaenoylethanolamide (DHEA) levels were higher (P.01) after fish oil feeding suggesting that NAPE-PLD was not necessary for DHEA synthesis. Liver and jejunum arachidonoylethanolamide, 1,2-arachidonoylglycerol and DHEA levels reflected their corresponding fatty acid precursors suggesting that alternate pathways are involved in NAE synthesis. NAPE-PLD (-/-) mice had lower oleoylethanolamide levels in the jejunum and a leaner phenotype compared to wild-type mice. Overall, these results demonstrate that dietary fatty acid can augment tissue NAEs in the absence of NAPE-PLD.

Oleoylethanolamide (OEA), a fatty acid ethanolamide (FAE), is a lipid mediator that controls food intake and lipid metabolism. Accumulating data imply the importance of intestinal OEA in controlling satiety in addition to gastrointestinal peptide hormones. Although the biochemical pathway of FAE production has been illustrated, the enzymes responsible for the cleavage of OEA from its precursor N-acyl-phosphatidylethanolamine (NAPE) must be identified among reported candidates in the gut. In this study, we assessed the involvement of NAPE-specific phospholipase D (NAPE-PLD), which can directly release FAEs from NAPE, in intestinal OEA synthesis and lipid metabolism. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPER-associated protein 9 (Cas9)-mediated deletion of the NAPE-PLD gene in intestinal epithelial-like Caco-2 cells reduced OEA levels, regardless of their differentiation states. Transcriptome analysis revealed that deletion of NAPE-PLD activates a transcriptional program for nutrient transportation, including lipids and lipoproteins, and inactivates cell-cycle or mitosis-related genes in Caco-2 cells. In addition, the basolateral secretion of lipoproteins was increased in NAPE-PLD-deleted cells although lipoprotein size was not affected. By contrast, cellular lipid levels were reduced in NAPE-PLD-deleted cells. Overall, these results indicate that NAPE-PLD plays important roles in OEA synthesis and fat absorption by regulating lipoprotein production in the intestinal epithelial cells.-Igarashi, M., Watanabe, K., Tsuduki, T., Kimura, I., Kubota, N. NAPE-PLD controls OEA synthesis and fat absorption by regulating lipoprotein synthesis in an in vitro model of intestinal epithelial cells.

BACKGROUND: Hidradenitis suppurativa (HS) is a chronic inflammatory disorder affecting the hair follicle which typically affects the axilla, inguinal, sub-mammary and perianal areas. Atypical HS, such as facial HS, dissecting cellulitis of the scalp (DCS), nape HS and HS lymphedema has also been described. There is scarce evidence about the treatment of atypical HS. AIMS: To describe the efficacy of biologic drugs and adjuvant therapeutic interventions used in patients with atypical HS who had an inadequate response to systemic antibiotics. METHODS: Prospective case series study of patients with atypical HS (facial HS, DCS, nape HS and HS lymphedema) treated with biologic drugs in a HS Clinic setting. Disease activity indexes, socio-demographic, clinical and safety variables were collected. RESULTS: Two patients met criteria for Facial HS, three met criteria for DCS / nape HS and three patients met criteria for HS lymphedema. Patients with facial HS achieved rapid improvement without requiring other therapies. Patients with DCS, nape HS and HS lymphedema showed variable response, with decrease of activity indexes, and requiring adjuvant treatments. CONCLUSIONS: Biologic drugs are useful in the management of special HS locations. In the case of facial HS, biologic therapy seems to be quite effective as monotherapy. In the cases of DCS, HS lymphedema and nape HS, treatment combination or intensification may be needed. This article is protected by copyright. All rights reserved.

Title: Impact of nape massage on postural stability Objectives: The aim of this work is to find out if sport nape massage has an effect on postural stability. Methods: The research was realized in form of quaziexperiment which was attended by 5 probands. Result of the measurement is consist of comparing the difference of postural stability before and after the nape massage. The results were obtained by Footscan pressure platform in Laboratory of sports motor skills at UK FTVS. At first we measured proband's weight and height, after that we measured postural stability. Then we performed nape massage on proband. And after the massage we measured again the postural stability. Results: In the overall summary of the results of all measured stands, there was an improvement after massage in the parameter TTW (total travelled way) by 6,12 %. This value indicates a slight improvement, but statistically nothing significant happened. The results of lower limbs load ratio were also positive, when after applying nape massage there was improvement by 35,5 %. This means that in our research with five probands, nape massage had a positive effect on level of static postural stability. Keywords: balance, footscan, sport massage, nape area

N-acylphosphatidylethanolamines (NAPEs) are glycerophospholipid precursors for bioactive lipid amides and potential regulators of membrane function. They are hydrolyzed by NAPE-specific phospholipase D (NAPE-PLD) and have been implicated in neurodegenerative disorders such as Parkinson's disease. Here, we used siRNA-mediated silencing of NAPE-PLD in human SH-SY5Y cells and NAPE-PLD-/- mice to determine whether NAPEs influence the membrane association of LRRK2, a multifunctional protein kinase that is frequently mutated in persons with sporadic Parkinson's disease. NAPE-PLD deletion caused a significant accumulation of non-metabolized NAPEs, which was accompanied by a shift of LRRK2 from membrane to cytosol and a reduction in total LRRK2 content. Conversely, exposure of intact SH-SY5Y cells to bacterial PLD lowered NAPE levels and enhanced LRRK2 association with membranes. The results suggest that NAPE-PLD activity may contribute to the control of LRRK2 localization by regulating membrane NAPE levels.

International audience; Parrots are highly social birds that are recognized for their primate-like cognitive abilities but their way to express emotions remain overlooked. Herein we explored potential facial indicators of emotions in cockatoos. We predicted that facial feather ruffling is an indicator of a cockatoo's emotional state and hypothesized that specific facial feather positions would be present more during positive valence and low arousal situations. We observed feather position on the crest, cheek and nape during the daily routine of a group of five captive, non-breeding, Sulphur-crested cockatoos. The data show that cheek and nape feather ruffling occurred significantly more during activities associated with low arousal levels and positive valence such as maintenance behaviours, positive and quiet social contact and resting. Our data suggest that ruffling feathers over the bill (i.e. cheek feather ruffling) and nape ruffling may provide visual indicators of calm/relaxed states in cockatoos. Subtle movement of facial feathers may be an effective close-ranged visual signal to communicate birds' affective states or their intention to engage in specific activities. This work provides a novel approach to assessing the positive welfare of captive cockatoos and to understanding emotional communication in non-mammalian species.

Houhai acupoint (HA) is a site for acupuncture stimulation, located in the fossa between the anus and tail base in animals. To evaluate HA as a potential immunization site, the immune responses were compared when HA and the conventional site nape were vaccinated in rats. The results showed that injection of a porcine epidemic diarrhea virus (PEDV) vaccine in HA induced significantly higher IgG, IgG1, IgG2, splenocyte proliferation and mRNA expression of IL-2, IL-4 and IFN-γ than in the nape. To search for the underlying mechanisms, the draining lymph nodes for HA and the nape were investigated. When rats were injected in HA with Indian ink, 11 lymph nodes including caudal mesenteric lymph node and bilateral gluteal lymph nodes, posterior inguinal lymph nodes, lumbar lymph nodes, internal iliac lymph nodes and popliteal lymph nodes were visibly stained with the ink and injection of a model antigen ovalbumin (OVA) in HA resulted in detection of OVA by western blotting while in the same lymph nodes only a pair of lymph nodes (central brachial lymph nodes) were observed when Indian ink or OVA was injected in the nape. IL-2 mRNA expression was detected in all the lymph nodes when PEDV vaccine was injected. Therefore, the enhanced immune response elicited by vaccination in HA may be attributed to more lymphocytes activated.

SummaryMechanisms of pruritus are implicated in the dysregulation of the metabolites in the spinal cord. We investigated pruritus behavioral testing in three groups of young adult male C57Bl/6 mice, including one group treated with normal saline, while the other groups intradermally injected with α-Me-5-HT (histamine-independent pruritogen), compound 48/80 (histamine-dependent pruritogen) at the nape skin of the neck, respectively. Proton nuclear magnetic resonance spectroscopy (MRS) was used to compare spinal metabolites from the vertebral cervical among three groups, and to study the association of spinal metabolite ratio and pruritus intensity. The MRS-measured N-acetylaspartate-to-myoinositol ratio (NAA/Ins) was significantly correlated with the number of scratches between normal saline group and 48/80 group or α-Me-5-HT group (both PP=0.017), indicating that Glu/Ins may be a surrogate marker of histamine-dependent pruritogen, while GABA/Ins was highly significantly different between normal saline group and α-Me-5-HT group (P=0.008), suggesting that GABA/Ins may be a surrogate marker of histamine-independent pruritogen. MRS may reflect the extent of pruritus intensity elicited by α-Me-5-HT and compound 48/80 with sensitivity similar to the number of scratches, and above potential markers need to be further validated in pre-clinical and clinical treatment trials.

N-acylethanolamines (NAEs), which include the endocannabinoid anandamide, represent an important family of signaling lipids in the brain. The lack of chemical probes that modulate NAE biosynthesis in living systems hamper the understanding of the biological role of these lipids. Using a high-throughput screen, chemical proteomics and targeted lipidomics, we report here the discovery and characterization of LEI-401 as a CNS-active N-acylphosphatidylethanolamine phospholipase D (NAPE-PLD) inhibitor. LEI-401 reduced NAE levels in neuroblastoma cells and in the brain of freely moving mice, but not in NAPE-PLD KO cells and mice, respectively. LEI-401 activated the hypothalamus–pituitary–adrenal axis and impaired fear extinction, thereby emulating the effect of a cannabinoid CB1 receptor antagonist, which could be reversed by a fatty acid amide hydrolase inhibitor. Our findings highlight the distinctive role of NAPE-PLD in NAE biosynthesis in the brain and suggest the presence of an endogenous NAE tone controlling emotional behavior.

Mesenchymal stem cells (MSCs), such as adipose-derived stem cells (ADSCs) and skeletal muscle-derived stem cells (MDSCs), are potential sources for cell-based therapeutic strategies. However, there is little knowledge about the lipid composition of these stem cells and the mechanisms of their differentiation. Lipids have important biological and physiological functions that are critical for understanding the regulation and control of stem cell fate. This study sought to analyze the lipidome of rabbit ADSCs and MDSCs and their adipogenic and osteogenic differentiation. The MSCs were isolated and were characterized by flow cytometry. Lipids were extracted from both MSCs and differentiated cells, and the lipids were subsequently analyzed with a hybrid triple quadrupole time-of-flight mass spectrometer. The results showed a total of 1687 lipid species. MSCs exhibited different lipid profiles as well as changes in lipid composition after differentiation. Furthermore, the expression levels of N-acyl-phosphatidylethanolamine (NAPE) 54:7+NH4 (-FA 17:0(NH4)) and phosphatidylcholine (PC) 42:6+Na were higher in the adipogenic lineages in of both MSC types, and NAPE 58:2+NH4 (-FA 17:0 (NH4)) and NAPE 56:2+NH4 (-FA 17:0 (NH4)) had higher levels in the osteogenic lineages, suggesting lipid similarities in cells differentiated from different stem cell sources.

To evaluate the therapeutic effects of nape acupuncture combined with rehabilitative swallowing training for dysphagia caused by pseudobulbar palsy, and to compare it with rehabilitative swallowing training alone, and to observe the improvement in quality of life after the therapy.One hundred patients were randomly divided into two groups: the rehabilitative swallowing training group (control group, n=50) and the nape acupuncture combined with rehabilitative swallowing training group (experimental group, n=50). Each group had 8 weeks' therapy, 5 times a week. Patients in the control group received rehabilitative swallowing training, while those in the experimental group received nape acupuncture therapy based on swallowing rehabilitation. The outcomes were assessed by the repetitive saliva-swallowing test (RSST), water swallow test (WST), standardized swallowing assessment (SSA), and a swallow quality-of-life questionnaire (SWAL-QOL). Correlations of onset age, onset frequency and lesion location with the efficacy of the acupuncture treatment were also observed.The scores for RSST, WST, and SSA in both groups were lower than before the therapy (P0.001), although the changes were more marked in the experimental group than in the control group (RSST and WST, P0.005; SSA, P0.001). Both groups recorded changes in SWAL-QOL index after the therapy (P0.001); and the experimental group scored higher than the control group (P0.001). The efficacy of acupuncture was not correlated with location (P0.05), but was related to onset age (P0.05) and onset frequency (P0.01).Nape acupuncture combined with rehabilitative swallowing training has an effect on dysphagia caused by pseudobulbar palsy and improves quality of life.

N-acyl-phosphatidylethanolamine phospholipase D (NAPE-PLD) (EC 3.1.4.4) catalyzes the final step in the biosynthesis of N-acyl-ethanolamides (NAEs). Reduced NAPE-PLD expression and activity may contribute to obesity and inflammation, but a major obstacle to elucidating the role of NAPE-PLD and NAE biosynthesis in various physiological processes has been the lack of effective NAPE-PLD inhibitors. The endogenous bile acid lithocholic acid (LCA) inhibits NAPE-PLD activity (IC50 68 μM) but LCA is also a highly potent ligand for TGR5 (EC50 0.52 μM). Recently, the first selective small molecule inhibitor of NAPE-PLD, ARN19874, was reported (IC50 34 μM). To identify more potent inhibitors of NAPE-PLD, we screened compounds using a quenched fluorescent NAPE analog, PED-A1, as a substrate for recombinant mouse NAPE-PLD. Screened compounds included a panel of bile acids as well as a library of experimental compounds (the Spectrum Collection). Muricholic acids and several other bile acids inhibited NAPE-PLD with potency similar to LCA. Fourteen potent NAPE-PLD inhibitors were identified in the Spectrum Collection, with the two most potent (IC50 ~2 μM) being symmetrically substituted dichlorophenes: hexachlorophene and bithionol. Structure activity relationship assays using additional substituted dichlorophenes identified key moieties needed for NAPE-PLD inhibition. Both hexachlorophene and bithionol showed significant selectivity for NAPE-PLD compared to non-target lipase activities such as S. chromofuscus PLD activity or serum lipase activity. Both also effectively inhibited NAPE-PLD activity in cultured HEK293 cells.

This study aimed to evaluate the influence of the individual nutritional status on the salivary concentration of N-acylethanolamines (NAEs), including linoleoylethanolamide (LEA), oleoylethanolamide (OEA), and palmitoylethanolamide (PEA), and their precursors N-acylphosphatidylethanolamines (NAPEs) upon mastication of biscuits containing different fats. Three types of biscuits were formulated with 10% extra-virgin olive oil (EVOB), 10% palm oil (PALMB) or 10% paraffin oil (0% lipids, CONB). Twenty-five healthy subjects, 12 normal weight (NW, 9 F, 30.4 ± 8.7 years) and 13 obese (OB, 8 F, 35.5 ± 10.7 years) participated in a randomized crossover study. Fasting subjects collected unstimulated saliva (US) and stimulated saliva by masticating a parafilm piece (PP), and CONB, EVOB and PALMB. NAPEs, LEA, OEA and PEA were quantified in saliva samples by liquid chromatography-high-resolution mass spectrometry. The results showed that salivary NAPE and NAE concentrations in OB were higher than in NW in both US (NAPEs: 280.0 ± 45.4 ng mL-1vs. 121.8 ± 24.4 ng mL-1, p = 0.015; NAEs: 10.8 ± 1.4 ng mL-1vs. 4.8 ± 0.8 ng mL-1, p = 0.002, respectively) and PP (NAPEs: 259.8 ± 47.1 ng mL-1vs. 121.7 ± 16.9 ng mL-1, p = 0.049; NAEs: 6.1 ± 0.8 ng mL-1vs. 3.8 ± 0.4 ng mL-1, p = 0.03, respectively). NAPE and LEA levels were similar in US and PP, while the levels of OEA and PEA were lower in PP vs. US. Compared to PP, biscuit mastication increased the salivary NAPEs, LEA, OEA and overall NAEs in NW and OB. NAPEs increased in the order of EVOB = CONB > PALMB in NW and EVOB > CONB = PALMB in OB. LEA, OEA and overall NAEs increased similarly with all the biscuits in NW and in the order of EVOB > PALMB > CONB in OB. In contrast, the PEA concentration did not vary in saliva upon biscuit mastication in NW and neither with EVOB in OB, while it lowered with CONB and PALMB in OB. In conclusion, OB showed higher salivary levels of NAPEs and NAEs than NW. Mastication itself did not vary salivary NAPEs and LEA but reduced OEA, PEA and overall NAEs. Biscuit mastication increased salivary NAPEs and all NAEs, but PEA. Altogether, the data suggested that NAPEs and NAEs were released in saliva from biscuits at levels influenced by the individual nutritional status and biscuit type. These findings may have implications in molecular mechanisms underpinning gustatory processes in humans.

Craniofacial hyperhidrosis occurs in the forehead, temples, upper lips, nose, nape, and scalp. It is less common in these areas than in the armpits, palmar region, or plantar region. It is possible to notice isolated sweating, or it can be associated with other areas.

Diverse metabolic disorders have been associated with an alteration of N-acylethanolamine (NAE) levels. These bioactive lipids are synthesized mainly by N-acylphosphatidylethanolamine-selective phospholipase D (NAPE-PLD) and influence host metabolism. We have previously discovered that NAPE-PLD in the intestine and adipose tissue is connected to the pathophysiology of obesity. However, the physiological function of NAPE-PLD in the liver remains to be deciphered. To study the role of liver NAPE-PLD on metabolism, we generated a new mouse model of inducible Napepld hepatocyte-specific deletion (Napepld∆Hep mice). In this study, we report that Napepld∆Hep mice develop a high-fat diet-like phenotype, characterized by an increased fat mass gain, hepatic steatosis and we show that Napepld∆Hep mice are more sensitive to liver inflammation. We also demonstrate that the role of liver NAPE-PLD goes beyond the mere synthesis of NAEs, since the deletion of NAPE-PLD is associated with a marked modification of various bioactive lipids involved in host homeostasis such as oxysterols and bile acids. Collectively these data suggest that NAPE-PLD in hepatocytes is a key regulator of liver bioactive lipid synthesis and a dysregulation of this enzyme leads to metabolic complications. Therefore, deepening our understanding of the regulation of NAPE-PLD could be crucial to tackle obesity and related comorbidities.

Our objective was to explore the physiological role of the intestinal endocannabinoids in the regulation of appetite upon short-term exposure to high-fat-diet (HFD) and understand the mechanisms responsible for aberrant gut-brain signaling leading to hyperphagia in mice lacking Napepld in the intestinal epithelial cells (IECs). We generated a murine model harboring an inducible NAPE-PLD deletion in IECs ( NapepldΔIEC). After an overnight fast, we exposed wild-type (WT) and NapepldΔIEC mice to different forms of lipid challenge (HFD or gavage), and we compared the modification occurring in the hypothalamus, in the vagus nerve, and at endocrine level 30 and 60 min after the stimulation. NapepldΔIEC mice displayed lower hypothalamic levels of N-oleoylethanolamine (OEA) in response to HFD. Lower mRNA expression of anorexigenic Pomc occurred in the hypothalamus of NapepldΔIEC mice after lipid challenge. This early hypothalamic alteration was not the consequence of impaired vagal signaling in NapepldΔIEC mice. Following lipid administration, WT and NapepldΔIEC mice had similar portal levels of glucagon-like peptide-1 (GLP-1) and similar rates of GLP-1 inactivation. Administration of exendin-4, a full agonist of GLP-1 receptor (GLP-1R), prevented the hyperphagia of NapepldΔIEC mice upon HFD. We conclude that in response to lipid, NapepldΔIEC mice displayed reduced OEA in brain and intestine, suggesting an impairment of the gut-brain axis in this model. We speculated that decreased levels of OEA likely contributes to reduce GLP-1R activation, explaining the observed hyperphagia in this model. Altogether, we elucidated novel physiological mechanisms regarding the gut-brain axis by which intestinal NAPE-PLD regulates appetite rapidly after lipid exposure.

A 58-year-old Caucasian male presents to the physician due to generalized fatigue, arthralgias of the metacarpophalangeal joints, and abdominal discomfort for 2 months. He has hypertension, hyperlipidemia, and type 2 diabetes mellitus. His grandfather died of liver cancer at the age of 62 years. He is 5 ft 7 in. tall and weighs 205 lb. BMI is 32.11 kg/m2. His temperature is 98 °F, pulse is 106/min, and blood pressure is 130/80 mmHg. Physical examination reveals bronze hyperpigmented skin over the nape of the neck and extremities (Figs. 3.1 and 3.2). The liver is palpated 5 cm below the right costal margin. Wood’s light reveal lime green fluorescence of the patient’s urine.

In mammals, the endocannabinoid anandamide is biosynthesized from N-arachidonoyl-phosphatidylethanolamine by the enzyme N-acyl-phosphatidylethanolamine-hydrolyzing phospholipase D (NAPE-PLD). Importantly, NAPE-PLD also produces other bioactive N-acylethanolamines, including anti-inflammatory palmitoylethanolamide and appetite-suppressive oleoylethanolamide, from their corresponding N-acyl-phosphatidylethanolamines (NAPEs). The analyses of NAPE-PLD-deficient mice revealed remarkable accumulation of NAPEs in brain and several peripheral tissues, suggesting the central role in NAPE degradation. On the other hand, the endogenous levels of N-acylethanolamines, including anandamide, were significantly decreased or almost unchanged, probably depending on co-existing NAPE-PLD-independent alternative pathways. The phenotypes of NAPE-PLD-deficient mice suggested the physiological and pathophysiological roles of NAPE-PLD in appetite, energy metabolism, and inflammation. So far, the reported inhibitors are not highly specific for NAPE-PLD, and potent specific inhibitors are under development.

N-acyl-phosphatidylethanolamine (NAPE)-hydrolyzing phospholipase D (NAPE-PLD) is a zinc metallohydrolase enzyme that converts NAPEs to bioactive N-acyl-ethanolamides (NAEs). Altered NAPE-PLD activity may contribute to pathogenesis of obesity, diabetes, atherosclerosis, and neurological diseases. Selective measurement of NAPE-PLD activity is challenging, however, due to alternative phospholipase pathways for NAPE hydrolysis. Previous methods to measure NAPE-PLD activity involved addition of exogenous NAPE followed by thin-layer chromatography (TLC) or liquid chromatography-tandem mass spectrometry (LC/MS/MS), which are time- and resource-intensive. Recently, NAPE-PLD activity in cells has been assayed using the fluorogenic NAPE analogs PED-A1 and PED6, but these substrates also detect the activity of serine hydrolase-type lipases PLA1 and PLA2. To create a fluorescence assay that selectively measured cellular NAPE-PLD activity, we synthesized an analog of PED-A1 (flame-NAPE) where the sn-1 ester bond was replaced with a N-methyl amide to create resistance to PLA1 hydrolysis. Recombinant NAPE-PLD produced fluorescence when incubated with either PED-A1 or flame-NAPE, while PLA1 only produced fluorescence when incubated with PED-A1. Furthermore, fluorescence in HepG2 cells using PED-A1 could be partially blocked by either biothionol (a selective NAPE-PLD inhibitor) or tetrahydrolipstatin (an inhibitor of a broad spectrum of serine hydrolase-type lipases). In contrast, fluorescence assayed in HepG2 cells using flame-NAPE could only be blocked by biothionol. In multiple cell types, the phospholipase activity detected using flame-NAPE was significantly more sensitive to biothionol inhibition than that detected using PED-A1. Thus, using flame-NAPE to measure phospholipase activity provides a rapid and selective method to measure NAPE-PLD activity in cells and tissues.

N-acylphosphatidylethanolamine phospholipase D (NAPE-PLD) catalyzes the cleavage of membrane NAPEs into bioactive fatty-acid ethanolamides (FAEs). Along with this precursor role, NAPEs might also serve autonomous signaling functions. Here, we report that injections of 6-hydroxydopamine (6-OHDA) into the mouse striatum cause a local increase in NAPE and FAE levels, which precedes neuronal cell death. NAPE, but not FAE, accumulation is enhanced in mice lacking NAPE-PLD, which display a substantial reduction in 6-OHDA-induced neurotoxicity, as shown by increased survival of substantia nigra dopamine neurons, integrity of striatal dopaminergic fibers, and striatal dopamine metabolite content. Reduced damage is accompanied by attenuation of the motor response evoked by apomorphine. Furthermore, NAPE-PLD silencing protects cathecolamine-producing SH-SY5Y cells from 6-OHDA-induced reactive oxygen species formation, caspase-3 activation and death. Mechanistic studies in mice suggest the existence of multiple molecular contributors to the neuroprotective effects of NAPE-PLD deletion, including suppression of Rac1 activity and attenuated transcription of several genes (Cadps, Casp9, Egln1, Kcnj6, Spen, and Uchl1) implicated in dopamine neuron survival and/or Parkinson’s disease. The findings point to a previously unrecognized role for NAPE-PLD in the regulation of dopamine neuron function, which may be linked to the control of NAPE homeostasis in membranes.

N-acyl-phosphatidylethanolamine (NAPE) is known to be a precursor for various bioactive N-acylethanolamines including the endocannabinoid anandamide. NAPE is produced in mammals through the transfer of an acyl chain from certain glycerophospholipids to phosphatidylethanolamine (PE) by Ca2+-dependent or -independent N-acyltransferases. The e isoform of mouse cytosolic phospholipase A2 (cPLA2e) was recently identified as a Ca2+-dependent N-acyltransferase (Ca-NAT). In the present study, we first showed that two isoforms of human cPLA2e function as Ca-NAT. We next purified both mouse recombinant cPLA2e and its two human orthologues to examine their catalytic properties. The enzyme absolutely required Ca2+ for its activity and the activity was enhanced by phosphatidylserine (PS). PS enhanced the activity 25-fold in the presence of 1 mM CaCl2 and lowered the EC50 value of Ca2+ >8-fold. Using a PS probe, we showed that cPLA2e largely co-localizes with PS in plasma membrane and organelles involved in the endocytic pathway, further supporting the interaction of cPLA2e with PS in living cells. Finally, we found that the Ca2+-ionophore ionomycin increased [14C]NAPE levels >10-fold in [14C]ethanolamine-labeled cPLA2e-expressing cells while phospholipase A/acyltransferase-1, acting as a Ca2+-independent N-acyltransferase, was insensitive to ionomycin for full activity. In conclusion, PS potently stimulated the Ca2+-dependent activity and human cPLA2e isoforms also functioned as Ca-NAT.

Bacteria inhabit a vast range of biological niches and have evolved diverse mechanisms to cope with environmental stressors. The genus Acinetobacter comprises a complex group of Gram-negative bacteria. Some of these bacteria such as A. baumannii are nosocomial pathogens. They are often resistant to multiple antibiotics and are associated with epidemic outbreaks. A. radioresistens is generally considered to be a commensal bacterium on human skin or an opportunistic pathogen. Interestingly, this species has exceptional resistance to a range of environmental challenges which contributes to its persistence in clinical environment and on human skin. We studied changes in its lipid composition induced by the onset of stationary phase. This strain produced triglycerides (TG) as well as four common phospholipids: phosphatidylethanolamine (PE), phosphatidylglycerol (PG), cardiolipin (CL) and lysocardiolipin (LCL). It also produced small amounts of acyl-phosphatidylglycerol (APG). As the bacterial growth entered the stationary phase, the lipidome switched from one dominated by PE and PG to another dominated by CL and LCL. Surprisingly, bacteria in the stationary phase produced N-acyl-phosphatidylethanolamine (NAPE) and another rare lipid we tentatively name as 1-phosphatidyl-2-acyl-glycero-3-phosphoethanolamine (PAGPE) based on tandem mass spectrometry. It is possible these tri-acylated lipids play an important role in coping with nutrient depletion.