Search

PURPOSE/AIM OF THE STUDY Pericytes (PCs), located abluminal of endothelial cells on capillaries, are essential for vascular development and stability. They display a heterogeneous morphology depending on organ localization, differentiation state and function. Consequently, PCs show a diverse gene expression profile, impeding the usage of a unique PC marker and therefore the distinct identification of PCs. Inducible reporter mouse models represent an important tool for investigating the fate of PCs under physiological and pathophysiological conditions. PC-specific expression efficiency of the fluorescence reporter tdTomato following tamoxifen induction was analyzed and compared in two inducible Cre recombinase-expressing mouse models under control of the NG2 and PDGFRb promotor. MATERIAL AND METHODS The NG2-CreERTM-tdTomato and the PDGFRb-P2A-CreERT2-tdTomato mice were treated with tamoxifen at three defining time points of retinal vascular development: postnatal days (P)5, P10/11/12 and P48/49/50/51. TdTomato reporter induction efficiency was determined by analyzing retinal whole mounts utilizing confocal microscopy, using the antibodies Anti-neural/glial antigen 2 (PCs), Anti-Collagen IV (basement membrane) and Anti-Glutamine Synthetase (Muller glial cells). RESULTS Tamoxifen induction at the three different time points resulted in PC-specific expression of tdTomato in both reporter models. In the NG2-CreERTM-tdTomato mouse, the induction efficiency ranged from 21.9 to 35.5%. In the PDGFRb-P2A-CreERT2-tdTomato mouse an induction efficiency between 78.9 and 94.1% was achieved. TdTomato expression in the retina was restricted to PCs and vascular smooth muscle cells in the NG2-CreERTM-tdTomato mouse, however, in the PDGFRb-P2A-CreERT2-tdTomato mouse, tdTomato was also expressed in Muller glial cells. CONCLUSION Both reporter mouse models represent promising tools for fate mapping studies of PCs. While the NG2-CreERTM-tdTomato mouse reveals very specific labeling of PCs in the retina, its induction efficiency is lower compared to the PDGFRb-P2A-CreERT2-tdTomato mouse. Although the latter revealed a high percentage of tdTomato-positive PCs in the retina, additional labeling of Muller cells potentially hampers analysis of reporter-positive PCs.

Introduction. Inflammatory bowel disease (IBD), which includes Crohn’s disease (CD) and ulcerative colitis (UC), is a multifactorial intestinal disorder but its precise etiology remains elusive. As the cells of the intestinal mucosa have high energy demands, mitochondria may play a role in IBD pathogenesis. The present study is aimed at evaluating the expression levels of mitochondrial oxidative phosphorylation (OXPHOS) complexes in IBD. Material and Methods. 286 intestinal biopsy samples from the terminal ileum, ascending colon, and rectum from 124 probands (34 CD, 33 UC, and 57 controls) were stained immunohistochemically for all five OXPHOS complexes and the voltage-dependent anion-selective channel 1 protein (VDAC1 or porin). Expression levels were compared in multivariate models including disease stage (CD and UC compared to controls) and age (pediatric/adult). Results. Analysis of the terminal ileum of CD patients revealed a significant reduction of complex II compared to controls, and a trend to lower levels was evident for VDAC1 and the other OXPHOS complexes except complex III. A similar pattern was found in the rectum of UC patients: VDAC1, complex I, complex II, and complex IV were all significantly reduced, and complex III and V showed a trend to lower levels. Reductions were more prominent in older patients compared to pediatric patients and more marked in UC than CD. Conclusion. A reduced mitochondrial mass is present in UC and CD compared to controls. This is potentially a result of alterations of mitochondrial biogenesis or mitophagy. Reductions were more pronounced in older patients compared to pediatric patients, and more prominent in UC than CD. Complex I and II are more severely compromised than the other OXPHOS complexes. This has potential therapeutic implications, since treatments boosting biogenesis or influencing mitophagy could be beneficial for IBD treatment. Additionally, substances specifically stimulating complex I activity should be tested in IBD treatment.

Psoriasis is an inflammatory skin disease characterized by increased neo-vascularization, keratinocyte hyperproliferation, a pro-inflammatory cytokine milieu and immune cell infiltration. Diacerein is an anti-inflammatory drug, modulating immune cell functions, including expression and production of cytokines, in different inflammatory conditions. Therefore, we hypothesized that topical diacerein has beneficial effects on the course of psoriasis. The current study aimed to evaluate the effect of topical diacerein on imiquimod (IMQ)-induced psoriasis in C57BL/6 mice. Topical diacerein was observed to be safe without any adverse side effects in healthy or psoriatic animals. Our results demonstrated that diacerein significantly alleviated the psoriasiform-like skin inflammation over a 7-day period. Furthermore, diacerein significantly diminished the psoriasis-associated splenomegaly, indicating a systemic effect of the drug. Remarkably, we observed significantly reduced infiltration of CD11c+ dendritic cells (DCs) into the skin and spleen of psoriatic mice with diacerein treatment. As CD11c+ DCs play a pivotal role in psoriasis pathology, we consider diacerein to be a promising novel therapeutic candidate for psoriasis.

Pericytes (PCs), located abluminal of endothelial cells on capillaries, are essential for vascular development and stability. They display a heterogeneous morphology depending on organ localization, differentiation state, and function. Consequently, PCs show a diverse gene expression profile, impeding the usage of a unique PC marker and therefore the distinct identification of PCs. Inducible reporter mouse models represent an important tool for investigating the fate of PCs under physiological and pathophysiological conditions. PC-specific expression efficiency of the fluorescence reporter tdTomato following tamoxifen induction was analyzed and compared in two inducible Cre recombinase-expressing mouse models under control of the NG2 and PDGFRb promotor.The NG2-CreER™-tdTomato and the PDGFRb-P2A-CreERTamoxifen induction at the three different time points resulted in PC-specific expression of tdTomato in both reporter models. In the NG2-CreER™-tdTomato mouse, the induction efficiency ranged from 21.9 to 35.5%. In the PDGFRb-P2A-CreERBoth reporter mouse models represent promising tools for fate-mapping studies of PCs. While the NG2-CreER™-tdTomato mouse reveals very specific labeling of PCs in the retina, its induction efficiency is lower compared to the PDGFRb-P2A-CreER

The regulatory (neuro)peptide galanin is widely distributed in the central and peripheral nervous systems, where it mediates its effects via three G protein-coupled receptors (GAL1-3R). Galanin has a vast diversity of biological functions, including modulation of feeding behavior. However, the clinical application of natural galanin is not practicable due to its rapid in vivo breakdown by peptidases and lack of receptor subtype specificity. Much effort has been put into the development of receptor-selective agonists and antagonists, and while receptor selectivity has been attained to some degree, most ligands show overlapping affinity. Therefore, we aimed to develop a novel ligand with specificity to a single galanin receptor subtype and increased stability. To achieve this, a lanthionine amino acid was enzymatically introduced into a galanin-related peptide. The residue’s subsequent cyclization created a conformational constraint which increased the peptide’s receptor specificity and proteolytic resistance. Further exchange of certain other amino acids resulted in a novel methyllanthionine-stabilized galanin receptor agonist, a G1pE-T3N-S6A-G12A-methyllanthionine[13–16]-galanin-(1–17) variant, termed M89b. M89b has exclusive specificity for GAL2R and a prolonged half-life in serum. Intranasal application of M89b to unfasted rats significantly reduced acute 24 h food intake inducing a drop in body weight. Combined administration of M89b and M871, a selective GAL2R antagonist, abolished the anorexigenic effect of M89b, indicating that the effect of M89b on food intake is indeed mediated by GAL2R. This is the first demonstration of in vivo activity of an intranasally administered lanthipeptide. Consequently, M89b is a promising candidate for clinical application as a galanin-related peptide-based therapeutic.

Current research into neuropeptides is bringing to light many remarkable functions of these endocrine/neurocrine/paracrine factors, such as their roles in modulating immune responses. Galanin is a neuropeptide expressed in both neural and non‐neural tissues and exerts its effects through three G protein–coupled receptors, GAL1,2,3‐R. It has been demonstrated that galanin has modulatory effects on immune cells, including neutrophils and natural killer cells. Because monocytes express GAL2‐R, and therefore are expected to be a target of galanin, we analyzed the effect of galanin on the expression of cytokines and chemokines by monocytes. Galanin increased the expression of IL‐1β up to 1.5‐fold, TNF‐α, IL‐10, IL‐18, and CCL3 up to twofold, and CXCL8 up to fourfold in nonactivated monocytes, but had no major effect on activated monocytes. A cross‐correlation analysis of cytokine expression profiles, irrespective of the activation status of the monocytes, revealed that galanin changed the cross‐correlation of the expression of certain cytokines. Galanin abolished several significant correlations in IFN‐γ–stimulated monocytes. For example, treatment with 10 nM galanin changed the Spearman's rank coefficient of IL‐18 and CXCL8 from 0.622 (P ≤ 0.01) to 0.126. These results further emphasize the importance of neuroregulatory peptides, such as galanin and their therapeutic potential to treat inflammatory diseases., Neuropeptides have various functions, including modulating immune responses. Because monocytes express one of the receptors for the neuropeptide galanin, the authors analyzed the effect of galanin on the expression of cytokines and chemokines by monocytes. Their data indicate that galanin modulates the immune responses of monocytes by regulating cytokine expression networks and that it exerts different functions in naive and inflammation‐induced cells.

Galanin is a neuropeptide which mediates its effects via three G-protein coupled receptors (GAL1-3 ). Administration of a GAL3 antagonist reduces alcohol self-administration in animal models while allelic variation in the GAL3 gene has been associated with an increased risk of alcohol use disorders in diverse human populations. Based on the association of GAL3 with alcoholism, we sought to characterize drug-seeking behavior in GAL3 -deficient mice for the first time. In the two-bottle free choice paradigm, GAL3 -KO mice consistently showed a significantly increased preference for ethanol over water when compared to wildtype littermates. Furthermore, male GAL3 -KO mice displayed significantly increased responding for ethanol under operant conditions. These differences in alcohol seeking behavior in GAL3 -KO mice did not result from altered ethanol metabolism. In contrast to ethanol, GAL3 -KO mice exhibited similar preference for saccharin and sucrose over water, and a similar preference for a high fat diet over a low fat diet as wildtype littermates. No differences in cognitive and locomotor behaviors were observed in GAL3 -KO mice to account for increased alcohol seeking behavior. Overall, these findings suggest genetic ablation of GAL3 in mice increases alcohol consumption.

Glaucoma is the second leading cause of blindness worldwide. This multifactorial, neurodegenerative group of diseases is characterized by the progressive loss of retinal ganglion cells (RGCs) and their axons, leading to irreversible visual impairment and blindness. There is a huge unmet and urging need for the development of new and translatable strategies and treatment options to prevent this progressive loss of RGC. Accumulating evidence points towards a critical role of neuroinflammation, in particular microglial cells, in the pathogenesis of glaucoma. Leukotrienes are mediators of neuroinflammation and are involved in many neurodegenerative diseases. Therefore, we tested the leukotriene receptors CysLT1R/GPR17-selective antagonist Montelukast (MTK) for its efficacy to modulate the reactive state of microglia in order to ameliorate RGCs loss in experimental glaucoma. Ocular hypertension (OHT) was induced unilaterally by injection of 8 μm magnetic microbead (MB) into the anterior chamber of female Brown Norway rats. The contralateral, untreated eye served as control. Successful induction of OHT was verified by daily IOP measurement using a TonoLab rebound tonometer. Simultaneously to OHT induction, one group received daily MTK treatment and the control group vehicle solution by oral gavage. Animals were sacrificed 13-15 days after MB injection. Retina and optic nerves (ON) of OHT and contralateral eyes were analyzed by immunohistochemistry with specific markers for RGCs (Brn3a), microglial cells/macrophages (Iba1 and CD68), and cysteinyl leukotriene pathway receptors (CysLT1R and GPR17). Protein labeling was documented by confocal microscopy and analyzed with ImageJ plugins. Further, mRNA expression of genes of the inflammatory and leukotriene pathway was analyzed in retinal tissue. MTK treatment resulted in a short-term IOP reduction at day 2, which dissipated by day 5 of OHT induction in MTK treated animals. Furthermore, MTK treatment resulted in a decreased activation of Iba1+ microglial cells in the retina and ON, and in a significantly increased RGC survival in OHT eyes. Within the retina, GPR17 and CysLT1R expression was demonstrated in single RCGs and in microglial cells respectively. Further, increased mRNA expression of pro-inflammatory genes was detected in OHT induced retinas. In the ON, OHT induction increased the number of GPR17+ cells, showing a significant reduction following MTK treatment. This study shows for the first time a significantly increased RGC survival in an acute OHT model following treatment with the leukotriene receptor antagonist MTK. These results strongly suggest a neuroprotective effect of MTK and a potential new therapeutic strategy for glaucoma treatment.

The neuropeptide galanin is distributed in the central and peripheral nervous systems and in non-neuronal peripheral organs, including the skin. Galanin acts via three G protein-coupled receptors which, except galanin receptor 1, are expressed in various skin structures. The galanin system has been associated with inflammatory processes of the skin and of several other organs. Psoriasis is an inflammatory skin disease with increased neovascularization, keratinocyte hyperproliferation, a proinflammatory cytokine milieu, and immune cell infiltration. In this study, we showed that galanin receptor 3 is present in endothelial cells in human and murine dermal vessels and is co-expressed with nestin in neo-vessels of psoriatic patients. Moreover, in a murine psoriasis model, we showed that C57/BL6 mice lacking galanin receptor 3 display a milder course of psoriasis upon imiquimod treatment, leading to decreased disease severity, delayed neo-vascularization, reduced infiltration of neutrophils, and significantly lower levels of proinflammatory cytokines compared with wild-type mice. In contrast, galanin receptor 2-knockout animals did not differ significantly from wild type mice at both the macroscopic and molecular levels in their inflammatory response to imiquimod treatment. Our data indicate that galanin receptor 3, but not galanin receptor 2, plays an important role in psoriasis-like skin inflammation.

Association Between Non-Alcoholic Fatty Liver Disease (NAFLD) and Iron Metabolism in Obese Children and Adolescents : Results of the Beta-JUDO Study

BACKGROUND: Pharmacological treatment options for adolescents with obesity are very limited. Glucagon-like-peptide-1 (GLP-1) receptor agonist could be a treatment option for adolescent obesity. OBJECTIVE: To investigate the effect of exenatide extended release on body mass index (BMI)-SDS as primary outcome, and glucose metabolism, cardiometabolic risk factors, liver steatosis, and other BMI metrics as secondary outcomes, and its safety and tolerability in adolescents with obesity. METHODS: Six-month, randomized, double-blinded, parallel, placebo-controlled clinical trial in patients (n = 44, 10-18 years, females n = 22) with BMI-SDS > 2.0 or age-adapted-BMI > 30 kg/m2 according to WHO were included. Patients received lifestyle intervention and were randomized to exenatide extended release 2 mg (n = 22) or placebo (n = 22) subcutaneous injections given once weekly. Oral glucose tolerance tests (OGTT) were conducted at the beginning and end of the intervention. RESULTS: Exenatide reduced (P De två första författarna delar förstaförfattarskapet

Inhibition of intestinal glucose resorption can serve as an effective strategy for the prevention of an increase in blood glucose levels. We have recently shown that various extracts prepared from guava (Psidium guajava) inhibit sodium-dependent glucose cotransporter 1 (SGLT1)- and glucose transporter 2 (GLUT2)-mediated glucose transport in vitro (Caco-2 cells) and in vivo (C57BL/6N mice). However, the efficacy in humans remains to be confirmed. For this purpose, we conducted a parallelized, randomized clinical study with young healthy adults. Thirty-one volunteers performed an oral glucose tolerance test (OGTT) in which the control group received a glucose solution and the intervention group received a glucose solution containing a guava fruit extract prepared by supercritical CO2 extraction. The exact same extract was used for our previous in vitro and in vivo experiments. Blood samples were collected prior to and up to two hours after glucose consumption to quantitate blood glucose and insulin levels. Our results show that, in comparison to the control group, consumption of guava fruit extract resulted in a significantly reduced increase in postprandial glucose response over the basal fasting plasma glucose levels after 30 min (&Delta, control 2.60 &plusmn, 1.09 mmol/L versus &Delta, intervention 1.96 &plusmn, 0.96 mmol/L, p = 0.039) and 90 min (&Delta, control 0.44 &plusmn, 0.74 mmol/L versus &Delta, intervention &minus, 0.18 &plusmn, 0.88 mmol/L, p = 0.023). In addition, we observed a slightly reduced, but non-significant insulin secretion (&Delta, control 353.82 &plusmn, 183.31 pmol/L versus &Delta, intervention 288.43 &plusmn, 126.19 pmol/L, p = 0.302). Interestingly, storage time and repeated freeze-thawing operations appeared to negatively influence the efficacy of the applied extract. Several analytical methods (HPLC-MS, GC-MS, and NMR) were applied to identify putative bioactive compounds in the CO2 extract used. We could assign several substances at relevant concentrations including kojic acid (0.33 mg/mL) and 5-hydroxymethylfurfural (2.76 mg/mL). Taken together, this clinical trial and previous in vitro and in vivo experiments confirm the efficacy of our guava fruit extract in inhibiting intestinal glucose resorption, possibly in combination with reduced insulin secretion. Based on these findings, the development of food supplements or functional foods containing this extract appears promising for patients with diabetes and for the prevention of insulin resistance. Trial registration: 415-E/2319/15-2018 (Ethics Commissions of Salzburg).

The regulatory peptide galanin is broadly distributed in the central- and peripheral nervous systems as well as in non-neuronal tissues, where it exerts its diverse physiological functions via three G-protein-coupled receptors (GAL1-3-R). Regulatory peptides are important mediators of the cross-communication between the nervous- and immune systems and have emerged as a focus of new therapeutics for a variety of inflammatory diseases. Studies on inflammatory animal models and immune cells revealed both pro- and anti-inflammatory functions of galanin. Here, we probed specific immune-related functions of the galanin system and found galanin and GAL1-R and GAL2-R mRNA to be expressed in a range of human immune cells. In particular, macrophages displayed differentiation- and polarization-dependent expression of galanin and its receptors. Exposure to exogenous galanin affected the cytokine/chemokine expression profile of macrophages differently, depending on their differentiation and polarization, and mainly modulated the expression of chemokines (CCL2, CCL3, CCL5 and CXCL8) and anti-inflammatory cytokines (TGF-β, IL-10 and IL-1Ra), especially in type-1 macrophages. Cytokine/chemokine expression levels in interferon-gamma- and lipopolysaccharide-polarized macrophages were upregulated whereas in unpolarized macrophages they were downregulated upon galanin treatment for 20 hours. This study illuminates the regulation of important cytokines/chemokines in macrophages by galanin, depending on specific cell activation.

Neurogenic inflammation mediated by peptidergic sensory nerves has a crucial impact on the pathogenesis of various joint diseases. Galanin is a regulatory sensory neuropeptide, which has been shown to attenuate neurogenic inflammation, modulate neutrophil activation, and be involved in the development of adjuvant arthritis, but our current understanding about its targets and physiological importance is incomplete. Among the receptors of galanin (GAL1–3), GAL3 has been found to be the most abundantly expressed in the vasculature and on the surface of some immune cells. However, since there are minimal in vivo data on the role of GAL3 in joint diseases, we analyzed its involvement in different inflammatory mechanisms of the K/BxN serum transfer-model of autoimmune arthritis employing GAL3 gene-deficient mice. After arthritis induction, GAL3 knockouts demonstrated increased clinical disease severity and earlier hindlimb edema than wild types. Vascular hyperpermeability determined by in vivo fluorescence imaging was also elevated compared to the wild-type controls. However, neutrophil accumulation detected by in vivo luminescence imaging or arthritic mechanical hyperalgesia was not altered by the lack of the GAL3 receptor. Our findings suggest that GAL3 has anti-inflammatory properties in joints by inhibiting vascular hyperpermeability and consequent edema formation.

Antibodies are an integral biomedical tool, not only for research but also as therapeutic agents. However, progress can only be made with sensitive and specific antibodies. The regulatory (neuro)peptide galanin and its three endogenous receptors (GAL1-3-R) are widely distributed in the central and peripheral nervous systems, and in peripheral non-neuronal tissues. The galanin system has multiple biological functions, including feeding behavior, pain processing, nerve regeneration and inflammation, to name only a few. Galanin could serve as biomarker in these processes, and therefore its receptors are potential drug targets for various diseases. For that reason, it is of paramount interest to precisely measure galanin peptide levels in tissues and to determine the cellular and subcellular localization of galanin receptors. A plethora of antibodies and antibody-based tools, including radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA) kits, are commercially available to detect galanin and its receptors. However, many of them lack rigorous validation which casts doubt on their specificity. A goal of the present study was to raise awareness of the importance of validation of antibodies and antibody-based tools, with a specific focus on the galanin system. To that end, we tested and report here about commercially available antibodies against galanin and galanin receptors that appear specific to us. Furthermore, we investigated the validity of commercially available galanin ELISA kits. As the tested ELISAs failed to meet the validation requirements, we developed and validated a specific sandwich ELISA which can be used to detect full-length galanin in human plasma.

Galanin is a neuropeptide which mediates its effects via three G-protein coupled receptors (GAL

In the background of obesity, among other factors, regulatory alterations in energy balance affecting peptide systems may also be assumed. Regulation of energy balance does not only involve maintenance of body weight but also that of metabolic rate and core temperature. The contribution of alarin, a new member of the potentially orexigenic galanin peptide family, to the regulation of energy metabolism has been recently suggested. Our aim was to analyze the thermoregulatory effects of alarin in rats. Adult male Wistar rats received full-length alarin (alarin 1-25), its truncated form (alarin 6-25Cys) or scrambled alarin in various doses intracerebroventricularly at different ambient temperatures. Oxygen consumption, heat loss (assessed by tail skin temperature) and core temperature of rats were recorded in an indirect calorimeter system. Upon alarin injection at 25 °C, an increase in oxygen consumption and continuous tail skin vasoconstriction induced a slow rise in core temperature that reached 0.5 °C by 120 and 1.0 °C by 180 min. At cooler or slightly warmer temperatures similar responses were seen. Neither the truncated nor the scrambled alarin elicited any significant thermoregulatory response, however, the truncated form antagonized the hyperthermic actions of the full-length peptide. Alarin appears to elicit a slow hypermetabolic, hyperthermic response in rats. Such a thermoregulatory response would characterize a catabolic (anorexic and hypermetabolic) mediator. Further investigations are needed to clarify the complex role of alarin in energy homeostasis.

The neuropeptide galanin (GAL) is widely distributed in the central and peripheral nervous systems. It is a modulator of various physiological and pathological processes, and it mediates its effects via three G protein-coupled receptors (GAL1-3 receptors). A role for GAL as a modulator of mood and anxiety was suggested, because GAL and its receptors are highly expressed in limbic brain structures of rodents. In recent years, numerous studies of animal models have suggested an involvement of GAL and GAL1 and GAL2 receptors in anxiety- and depression-related behavior. However, to date, there is sparse literature implicating GAL3 receptors in behavioral functions. Therefore, we studied the behavior of GAL3 receptor-deficient (GAL3-KO) mice to elucidate whether GAL3 receptors are involved in mediating behavior-associated actions of GAL. The GAL3-KO mouse line exhibited normal breeding and physical development. In addition to behavioral tests, phenotypic characterization included analysis of hematology, amino acid profiles, metabolism, and sudomotor function. In contrast to WT littermates, male GAL3-KO mice exhibited an anxiety-like phenotype in the elevated plus maze, open field, and light/dark box tests, and they were less socially affiliated than WT animals to a stranger mouse in a social interaction test. In conclusion, our data suggest involvement of GAL3 receptors in GAL-mediated effects on mood, anxiety, and behavior, making it a possible target for alternative treatment strategies for mood disorders.

Hypothalamic neuropeptides influence the main components of energy balance: metabolic rate, food intake, body weight as well as body temperature, by exerting either an overall anabolic or catabolic effect. The contribution of alarin, the most recently discovered member of the galanin peptide family to the regulation of energy metabolism has been suggested. Our aim was to analyze the complex thermoregulatory and food intake-related effects of alarin in rats. Adult male Wistar rats received different doses of alarin (0.3; 1; 3 and 15μg corresponding approximately to 0.1, 0.33, 1, and 5 nmol, respectively) intracerebroventricularly. Regarding thermoregulatory analysis, oxygen consumption (indicating metabolic rate), core temperature and heat loss (assessed by tail skin temperature) were recorded in an Oxymax indirect calorimeter system complemented with thermocouples and Benchtop thermometer. In order to investigate potential prostaglandin-mediated mechanisms of the hyperthermic effect of alarin, effects of intraperitoneally applied non-selective (indomethacin, 2mg/kg) or selective cyclooxygenase inhibitor (COX-2 inhibitor meloxicam, 1; 2mg/kg) were tested. Effects of alarin on daytime and nighttime spontaneous food intake, as well as, 24-h fasting-induced re-feeding were recorded in an automated FeedScale system. Alarin increased oxygen consumption with simultaneous suppression of heat loss leading to a slow coordinated rise in core temperature. Both applied COX-inhibitors suppressed this action. Alarin failed to induce daytime food intake, but suppressed spontaneous nighttime and also fasting-induced re-feeding food intake. Alarin appears to elicit a slow anorexigenic and prostaglandin-mediated, fever-like hyperthermic response in rats. Such a combination would characterize a catabolic mediator. The potential involvement of alarin in sickness behavior may be assumed.

Allergic contact dermatitis (ACD) is an inflammatory skin disease induced by allergen exposure and characterized by erythema, oedema and immune cell infiltration. The sensory peptide galanin (GAL) and its three receptors (GAL1-3 ) are involved in regulating inflammation. As GAL and its receptors are expressed in human and murine skin and GAL expression is increased in oxazolone-induced contact allergy, it could play a role in dermatitis. As GAL reduces neurogenic plasma extravasation in the mouse skin via GAL3 activation, the role of GAL3 in the oxazolone-induced dermatitis model was explored. Following topical challenge with oxazolone, GAL3 gene-deficient mice showed a trend towards reduced ear thickness. Plasma extravasation and neutrophil infiltration increased considerably upon oxazolone challenge in both GAL3 knockout animals and wild-type controls without any observable effect of the gene deletion. We conclude that a lack of GAL3 does not influence oxazolone-induced ACD.

Neurogenic inflammatory components mediated by peptidergic sensory nerves have a crucial impact on the symptoms of inflammatory diseases. Galanin is a regulatory sensory neuropeptide, which was shown to attenuate neurogenic inflammation, but our current understanding about its endogenous targets, and physiologic importance is incomplete. Among the endogenous receptors of galanin (GAL1, GAL2, GAL3) we found GAL3 to be the most abundantly expressed on the vasculature and GAL2 on different types of immune cells including polymorphonuclear neutrophils and natural killer cells. Therefore, we evaluated if galanin exerts direct or indirect effects on these immune cells. Our data revealed that galanin can be regarded as an immunomodulatory peptide as it can sensitize polymorphonuclear neutrophils and natural killer cells towards proinflammatory cytokines. Since there are only scarce in vivo data concerning the role of GAL3 in inflammatory disease conditions, we analysed its involvement in the K/BxN serum transfer model of autoimmune arthritis and the oxazolone-model of allergic contact dermatitis, employing GAL3 gene-deficient mice. After arthritis induction, GAL3-knockout mice demonstrated increased clinical disease severity and earlier hindlimb edema than wildtype mice. Vascular hyperpermeability was also elevated compared to wildytpes, but neutrophil myeloperoxidase activity and arthritic hyperalgesia were not significantly different. In contrast, disease severity, vascular, and immune components were not affected in allergic contact dermatitis in GAL3 knockouts in comparison with wildtypes. Our findings suggest GAL3 activation as a substantial anti-inflammatory pathway in neutrophil-dominated autoimmune arthritis, modulating the early neurogenic vascular hyperpermeability and consequent edema formation. However, the involvement of GAL3 activation in the T-cell dependent allergic contact dermatitis remains unsupported.

Introduction One of the causes of preeclampsia (PE) is due to an imbalance of inflammatory and anti-inflammatory proteins, as well as hyperactivity of the sympathetic nervous system. Previous studies have already revealed the involvement of the neuropeptides galanin (GAL) and oxytocin (OXT) in inflammatory processes and their modulatory role with respect to the autonomous system 1, 2. Despite those findings, evidence for a relation of these hormones in the pathogenesis of PE is missing so far. Objectives Aim of the study is to characterise the above mentioned peptide systems regarding their expression patterns in the placenta, umbilical cord and plasma of PE patients and healthy controls. Patients and methods Placental (maternal and fetal side) and umbilical cord tissues have been collected during caesarean section from control and PE patients. Furthermore blood samples have been taken at the day of the surgery and four days postpartum. Biopsies have been analysed for the expression of GAL, GAL1,3- and OXT-receptors (GAL1, GAL3, OXTR). Additionally, differentiated analysis of maternal and fetal tissue was performed to reveal potential different expression patterns. Results Preliminary results of the IHC revealed a clear expression of GAL3 in trophoblasts of PE placentas, whereas controls showed an increase expression of GAL3 on endothelial cells. Moreover an increased expression of GAL1 and GAL on endothelial cells has been detected in controls compared to PE. Different patterns were also visible regarding the OXTR with higher expression levels in controls compared to PE. Furthermore, PE placentas showed local expression differences (fetal > maternal side), which was not apparent in controls. Conclusion The different expression patterns of the GAL-system and the OXTR still have to be verified by Western Blot analyses, qPCR and IHC. However, their preliminary detection in placental tissue could be a first hint regarding the involvement of those systems in the inflammatory processes of PE.

BackgroundEpidemiological case-control studies have revealed associations between mitochondrial haplogroups and the onset and/or progression of various multifactorial diseases. For instance, mitochondrial haplogroup T was previously shown to be associated with vascular diseases, including coronary artery disease and diabetic retinopathy. In contrast, haplogroup H, the most frequent haplogroup in Europe, is often found to be more prevalent in healthy control subjects than in patient study groups. However, justifications for the assumption that haplogroups are functionally distinct are rare. Therefore, we attempted to compare differences in mitochondrial function between haplogroup H and T cybrids.Methodology/principal findingsMitochondrial haplogroup H and T cybrids were generated by fusion of HEK293 cells devoid of mitochondrial DNA with isolated thrombocytes of individuals with the respective haplogroups. These cybrid cells were analyzed for oxidative phosphorylation (OXPHOS) enzyme activities, mitochondrial DNA (mtDNA) copy number, growth rate and susceptibility to reactive oxygen species (ROS). We observed that haplogroup T cybrids have higher survival rate when challenged with hydrogen peroxide, indicating a higher capability to cope with oxidative stress.Conclusions/significanceThe results of this study show that functional differences exist between HEK293 cybrid cells which differ in mitochondrial genomic background.

BackgroundOnset and development of the multifactorial disease age-related macular degeneration (AMD) are highly interrelated with mitochondrial functions such as energy production and free radical turnover. Mitochondrial dysfunction and overproduction of reactive oxygen species may contribute to destruction of the retinal pigment epithelium, retinal atrophy and choroidal neovascularization, leading to AMD. Consequently, polymorphisms of the mitochondrial genome (mtDNA) are postulated to be susceptibility factors for this disease. Previous studies from Australia and the United States detected associations of mitochondrial haplogroups with AMD. The aim of the present study was to test these associations in Middle European Caucasians.Methodology/principal findingsMitochondrial haplogroups (combinations of mtDNA polymorphisms) and mitochondrial CR polymorphisms were analyzed in 200 patients with wet AMD (choroidal neovascularization, CNV), in 66 patients with dry AMD, and in 385 controls from Austria by means of multiplex primer extension analysis and sequencing, respectively. In patients with CNV, haplogroup H was found to be significantly less frequent compared to controls, and haplogroup J showed a trend toward a higher frequency compared to controls. Five CR polymorphisms were found to differ significantly in the two study populations compared to controls, and all, except one (T152C), are linked to those haplogroups.Conclusions/significanceIt can be concluded that haplogroup J is a risk factor for AMD, whereas haplogroup H seems to be protective for AMD.