Hypothalamic UDP Increases in Obesity and Promotes Feeding via P2Y6-Dependent Activation of AgRP Neurons.

Activation of orexigenic AgRP-expressing neurons in the arcuate nucleus of the hypothalamus potently promotes feeding, thus defining new regulators of AgRP neuron activity could uncover potential novel targets for obesity treatment. Here, we demonstrate that AgRP neurons express the purinergic receptor 6 (P2Y6), which is activated by uridine-diphosphate (UDP). In vivo, UDP induces ERK phosphorylation and cFos expression in AgRP neurons and promotes action potential firing of these neurons in brain slice recordings. Consequently, central application of UDP promotes feeding, and this response is abrogated upon pharmacologic or genetic inhibition of P2Y6 as well as upon pharmacogenetic inhibition of AgRP neuron activity. In obese animals, hypothalamic UDP content is elevated as a consequence of increased circulating uridine concentrations. Collectively, these experiments reveal a potential regulatory pathway in obesity, where peripheral uridine increases hypothalamic UDP concentrations, which in turn can promote feeding via PY6-dependent activation of AgRP neurons.

Deep immune profiling of chronic rhinosinusitis in allergic and non-allergic cohorts using mass cytometry.

Chronic rhinosinusitis (CRS) is a persistent nasal and paranasal sinus mucosa inflammation comprising two phenotypes, namely CRS with nasal polyps (CRSwNP) and without (CRSsNP). CRSwNP can be associated with asthma and hypersensitivity to non-steroidal anti-inflammatory drug (NSAID) in a syndrome known as NSAID-exacerbated respiratory disease (N-ERD). Furthermore, CRS frequently intertwines with respiratory allergies. This study investigated levels of 33 different nasal and serum cytokines and phenotypic characteristics of peripheral blood mononuclear cells (PBMCs) within cohorts of CRS patients (n = 24), additionally examining the influence of comorbid respiratory allergies by mass cytometry. N-ERD patients showed heightened type 2 nasal cytokine levels. Mass cytometry revealed increased activated naive B cell levels in CRSwNP and N-ERD, while resting naive B cells were higher in CRSsNP. Th2a cell levels were significantly elevated in allergic subjects, but not in CRS groups. In conclusion, there are distinct immunological features in PBMCs of CRS phenotypes and allergy.

Pharmacokinetics of isavuconazole at different target sites in healthy volunteers after single and multiple intravenous infusions.

BACKGROUND: Invasive aspergillosis is a severe fungal infection that affects multiple organ systems including the CNS and the lungs. Isavuconazole, a novel triazole antifungal agent, has demonstrated promising activity against Aspergillus spp. However, data on the penetration of isavuconazole into the CNS and ELF and intracellular accumulation remain limited. MATERIALS AND METHODS: We conducted a prospective single-centre pharmacokinetic (PK) study in 12 healthy volunteers. Subjects received seven doses of 200 mg isavuconazole to achieve an assumed steady-state. After the first and final infusion, plasma sampling was conducted over 8 and 12 h, respectively. All subjects underwent one lumbar puncture and bronchoalveolar lavage, at either 2, 6 or 12 h post-infusion of the final dose. PBMCs were collected in six subjects from blood to determine intracellular isavuconazole concentrations at 6, 8 or 12 h. The AUC/MIC was calculated for an MIC value of 1 mg/L, which marks the EUCAST susceptibility breakpoint for Aspergillus fumigatus and Aspergillus flavus. RESULTS: C max and AUC0-24h of isavuconazole in plasma under assumed steady-state conditions were 6.57 ±â€Š1.68 mg/L (mean ±â€ŠSD) and 106 ±â€Š32.1 h·mg/L, respectively. The average concentrations measured in CSF, ELF and in PBMCs were 0.07 ±â€Š0.03, 0.94 ±â€Š0.46 and 27.1 ±â€Š17.8 mg/L, respectively. The AUC/MIC in plasma, CSF, ELF and in PBMCs under steady-state conditions were 106 ±â€Š32.1, 1.68 ±â€Š0.72, 22.6 ±â€Š11.0 and 650 ±â€Š426 mg·h/L, respectively. CONCLUSION: Isavuconazole demonstrated moderate penetration into ELF, low penetrability into CSF and high accumulation in PBMCs. Current dosing regimens resulted in sufficient plasma exposure in all subjects to treat isolates with MICs ≤ 1 mg/L.

Prevention of M2 polarization and temporal limitation of differentiation in monocytes by extracellular ATP.

BACKGROUND: Elevated levels of extracellular adenosine triphosphate (ATP) modulate immunologic pathways and are considered to be a danger signal in inflammation, lung fibrosis and cancer. Macrophages can be classified into two main types: M1 macrophages are classically activated, pro-inflammatory macrophages, whereas M2 macrophages are alternatively activated, pro-fibrotic macrophages. In this study, we examined the effect of ATP on differentiation of native human monocytes into these macrophage subtypes. We characterized M1 and M2 like macrophages by their release of Interleukin-1beta (IL-1ß) and Chemokine (C-C motif) ligand 18 (CCL18), respectively. RESULTS: Monocytes were stimulated with ATP or the P2X7 receptor agonist Benzoylbenzoyl-ATP (Bz-ATP), and the production of various cytokines was analyzed, with a particular focus on CCL18 and IL-1ß, along with the expression of different purinergic receptors. Over a 72 h period of cell culture, monocytes spontaneously differentiated to M2 like macrophages, as indicated by an increased release of CCL18. Immediate stimulation of monocytes with ATP resulted in a dose-dependent reduction in CCL18 release, but had no effect on the concentration of IL-1ß. In contrast, delayed stimulation with ATP had no effect on either CCL18 or IL-1ß release. Similar results were observed in a model of inflammation using lipopolysaccharide-stimulated human monocytes. Stimulation with the P2X7 receptor agonist Bz-ATP mimicked the effect of ATP on M2-macrophage differentiation, indicating that P2X7 is involved in ATP-induced inhibition of CCL18 release. Indeed, P2X7 was downregulated during spontaneous M2 differentiation, which may partially explain the ineffectiveness of late ATP stimulation of monocytes. However, pre-incubation of monocytes with PPADS, Suramin (unselective P2X- and P2Y-receptor blockers) and KN62 (P2X7-antagonist) failed to reverse the reduction of CCL18 by ATP. CONCLUSIONS: ATP prevents spontaneous differentiation of monocytes into M2-like macrophages in a dose- and time-dependent manner. These effects were not mediated by P2X and P2Y receptors.

Lack of adipocyte purinergic P2Y6 receptor greatly improves whole body glucose homeostasis.

Uridine diphosphate (UDP)-activated purinergic receptor P2Y6 (P2Y6R) plays a crucial role in controlling energy balance through central mechanisms. However, P2Y6R's roles in peripheral tissues regulating energy and glucose homeostasis remain unexplored. Here, we report the surprising finding that adipocyte-specific deletion of P2Y6R protects mice from diet-induced obesity, improving glucose tolerance and insulin sensitivity with reduced systemic inflammation. These changes were associated with reduced JNK signaling and enhanced expression and activity of PPARα affecting downstream PGC1α levels leading to beiging of white fat. In contrast, P2Y6R deletion in skeletal muscle reduced glucose uptake, resulting in impaired glucose homeostasis. Interestingly, whole body P2Y6R knockout mice showed metabolic improvements similar to those observed with mice lacking P2Y6R only in adipocytes. Our findings provide compelling evidence that P2Y6R antagonists may prove useful for the treatment of obesity and type 2 diabetes.

[Diagnosis and treatment of asthma: a guideline for respiratory specialists 2023 - published by the German Respiratory Society (DGP) e. V.] / S2k-Leitlinie zur fachärztlichen Diagnostik und Therapie von Asthma 2023.

The management of asthma has fundamentally changed during the past decades. The present guideline for the diagnosis and treatment of asthma was developed for respiratory specialists who need detailed and evidence-based information on the new diagnostic and therapeutic options in asthma. The guideline shows the new role of biomarkers, especially blood eosinophils and fractional exhaled NO (FeNO), in diagnostic algorithms of asthma. Of note, this guideline is the first worldwide to announce symptom prevention and asthma remission as the ultimate goals of asthma treatment, which can be achieved by using individually tailored, disease-modifying anti-asthmatic drugs such as inhaled steroids, allergen immunotherapy or biologics. In addition, the central role of the treatment of comorbidities is emphasized. Finally, the document addresses several challenges in asthma management, including asthma treatment during pregnancy, treatment of severe asthma or the diagnosis and treatment of work-related asthma.

European Respiratory Society Guidelines for the Diagnosis of Asthma in Adults.

Although asthma is very common affecting 5-10% of the population, the diagnosis of asthma in adults remains a challenge in the real world that results in both over- and under-diagnosis. A task force (TF) was set up by the European Respiratory Society to systematically review the literature on the diagnostic accuracy of tests used to diagnose asthma in adult patients and provide recommendation for clinical practice.The TF defined eight PICO (Population, Index, Comparator, and Outcome) questions that were assessed using the GRADE (Grading of Recommendations, Assessment, Development and Evaluation) approach, The TF utilised the outcomes to develop an evidenced-based diagnostic algorithm, with recommendations for a pragmatic guideline for everyday practice that was directed by real-life patient experiences.The TF support the initial use of spirometry followed, and if airway obstruction is present, by bronchodilator reversibility testing. If initial spirometry fails to show obstruction, further tests should be performed in the following order: FeNO, PEF variability or in secondary care, bronchial challenge. We present the thresholds for each test that are compatible with a diagnosis of asthma in the presence of current symptoms.The TF reinforce the priority to undertake spirometry and recognise the value of measuring blood eosinophils and serum IgE to phenotype the patient. Measuring gas trapping by body plethysmography in patients with preserved FEV1/FVC ratio deserves further attention. The TF draw attention on the difficulty of making a correct diagnosis in patients already receiving inhaled corticosteroids, the comorbidities that may obscure the diagnosis, the importance of phenotyping, and the necessity to consider the patient experience in the diagnostic process.

P2Y12-dependent activation of hematopoietic stem and progenitor cells promotes emergency hematopoiesis after myocardial infarction.

Emergency hematopoiesis is the driving force of the inflammatory response to myocardial infarction (MI). Increased proliferation of hematopoietic stem and progenitor cells (LSK) after MI enhances cell production in the bone marrow (BM) and replenishes leukocyte supply for local cell recruitment to the infarct. Decoding the regulation of the inflammatory cascade after MI may provide new avenues to improve post-MI remodeling. In this study, we describe the influence of adenosine diphosphate (ADP)-dependent P2Y12-mediated signaling on emergency hematopoiesis and cardiac remodeling after MI. Permanent coronary ligation was performed to induce MI in a murine model. BM activation, inflammatory cell composition and cardiac function were assessed using global and platelet-specific gene knockout and pharmacological inhibition models for P2Y12. Complementary in vitro studies allowed for investigation of ADP-dependent effects on LSK cells. We found that ADP acts as a danger signal for the hematopoietic BM and fosters emergency hematopoiesis by promoting Akt phosphorylation and cell cycle progression. We were able to detect P2Y12 in LSK, implicating a direct effect of ADP on LSK via P2Y12 signaling. P2Y12 knockout and P2Y12 inhibitor treatment with prasugrel reduced emergency hematopoiesis and the excessive inflammatory response to MI, translating to lower numbers of downstream progeny and inflammatory cells in the blood and infarct. Ultimately, P2Y12 inhibition preserved cardiac function and reduced chronic adverse cardiac remodeling after MI. P2Y12-dependent signaling is involved in emergency hematopoiesis after MI and fuels post-ischemic inflammation, proposing a novel, non-canonical value for P2Y12 antagonists beyond inhibition of platelet-mediated atherothrombosis.

Blocking P2X purinoceptor 4 signalling alleviates cigarette smoke induced pulmonary inflammation.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is associated with elevated ATP levels in the extracellular space. Once released, ATP serves as danger signal modulating immune responses by activating purinergic receptors. Accordingly, purinergic signalling has been implicated in respiratory inflammation associated with cigarette smoke exposure. However, the role of P2X4-signalling has not been fully elucidated yet. METHODS: Here, we analysed the P2X4 mRNA expression in COPD patients as well as cigarette smoke-exposed mice. Furthermore, P2X4-signalling was blocked by either using a specific antagonist or genetic depletion of P2rx4 in mice applied to an acute and prolonged model of cigarette smoke exposure. Finally, we inhibited P2X4-signalling in macrophages derived from THP-1 before stimulation with cigarette smoke extract. RESULTS: COPD patients exhibited an increased P2X4 mRNA expression in cells isolated from the bronchoalveolar lavage fluid and peripheral mononuclear cells. Similarly, P2rx4 expression was elevated in lung tissue of mice exposed to cigarette smoke. Blocking P2X4-signalling in mice alleviated cigarette smoke induced airway inflammation as well as lung parenchyma destruction. Additionally, human macrophages derived from THP-1 cells released reduced concentrations of proinflammatory cytokines in response to cigarette smoke extract stimulation when P2X4 was inhibited. CONCLUSION: Taken together, we provide evidence that P2X4-signalling promotes innate immunity in the immunopathologic responses induced by cigarette smoke exposure.

Comparison of heart rates at fixed percentages and the ventilatory thresholds in patients with interstitial lung disease.

Heart rate (HR) responses to maximal exercise are commonly used for the prescription of training intensities in pulmonary rehabilitation. Those intensities are usually based on fixed percentages of peak HR (HRpeak), heart rate reserve (HRR), or peak work load (Wpeak), and rarely on HRs at the individual ventilatory thresholds (VT1 and VT2) derived from cardiopulmonary exercise testing (CPET). For patients suffering from interstitial lung disease (ILD), data on cardiorespiratory responses to CPET are scarce. Thus, the aim of this study was to record cardiorespiratory responses to CPET and to compare fixed HR percentages with HRs at VT1 and VT2 in ILD patients. A total of 120 subjects, 80 ILD patients and 40 healthy controls, underwent a symptom-limited CPET. From the ILD patient, 32 suffered from idiopathic pulmonary fibrosis (IPF), 37 from connective tissue disease (CTD), and 11 from sarcoidosis. HRs at fixed percentages, that is, at 70%HRpeak, at 70%Wpeak, and at 60%HRR were significantly lower in the ILD patients compared with the control group (p-values: 0.001, 0.044, and 0.011). Large percentages of HR values at 70%Wpeak and 60%HRR ranged between the HRs at VT1 and VT2 in ILD subgroups and controls as well. HRs at 70%HRpeak were lower than HRs at VT1 in 66% of the IPF patients, 54% of the CTD patients, and 55% of patients with sarcoidosis compared with 18% in the control group. Our findings demonstrate a considerable scattering of fixed HR percentages compared with HRs at the individual VTs derived from CPET in ILD patients. These findings may provide valuable information for the prescription of exercise intensity in pulmonary rehabilitation of ILD patients.

Comparison of pulmonary function test, diffusion capacity, blood gas analysis and CT scan in patients with and without persistent respiratory symptoms following COVID-19.

BACKGROUND: Long-lasting symptoms following SARS-CoV2-infection have been described in several studies. However, there is only limited knowledge about the ongoing pathophysiology and the association with pathological findings in medical examinations. METHODS: In this post hoc analysis of a prospective trial, 135 patients following COVID-19 were enrolled and grouped with respect to the presence or absence of respiratory ongoing symptoms following COVID-19. Pulmonary function test (PFT), diffusion capacity measurement (TLCO SB and TLCO/VA), blood gas analysis (BGA), laboratory tests and high-resolution computed tomography (HRCT) of patients with persistent respiratory symptoms were compared to those of asymptomatic patients. RESULTS: In this analysis, 71% (96/135) of all patients (mean age 49 years; range 20-91 years) reported long-lasting symptoms after a median (IQR) of 85 days (60-116) following COVID-19 whereby 57.8% (78/135) complained about persistent pulmonary symptoms. Pathological findings in blood test, PFT, TLCO, BGA and/or HRCT were found in 71.8% and 64.1% of patients with and without long-lasting respiratory symptoms respectively. Patients with persistent respiratory symptoms were significantly younger and presented a significant lower FVC (%), TLC (L), and TLCO SB compared to asymptomatic patients (p < 0.05). The multiple logistic regression results in a significant effect of age (p = 0.004) and TLCO SB (p = 0.042). CONCLUSION: Following COVID-19, a large proportion of patients experience ongoing symptoms, whereby the respiratory symptoms are the predominant complaints. Compared to asymptomatic patients, patients with ongoing symptoms were younger and presented a significant lower FVC, TLC and TLCO SB. The multiple logistic regression demonstrated only a significant association between the TLCO SB as the only PFT parameter and the perceived symptoms.

Toward personalization of asthma treatment according to trigger factors.

Asthma is a severe and chronic disabling disease affecting more than 300 million people worldwide. Although in the past few drugs for the treatment of asthma were available, new treatment options are currently emerging, which appear to be highly effective in certain subgroups of patients. Accordingly, there is a need for biomarkers that allow selection of patients for refined and personalized treatment strategies. Recently, serological chip tests based on microarrayed allergen molecules and peptides derived from the most common rhinovirus strains have been developed, which may discriminate 2 of the most common forms of asthma, that is, allergen- and virus-triggered asthma. In this perspective, we argue that classification of patients with asthma according to these common trigger factors may open new possibilities for personalized management of asthma.

Creatine supplementation impairs airway inflammation in an experimental model of asthma involving P2 × 7 receptor.

Creatine (Cr) is a substrate for adenosine triphosphate synthesis, and it is the most used dietary supplement among professional and recreative athletes and sportsmen. Creatine supplementation may increase allergic airway response, but the cellular and molecular mechanisms are unknown. We used murine model of OVA-induced chronic asthma and showed that Cr supplementation increased total proteins, ATP level, lymphocytes, macrophages, and IL-5 levels in BALF, as well as IL-5 in the supernatant of re-stimulated mediastinal lymph nodes. IL-5 and IL-13 expression by epithelial cells and by peribronchial leukocytes were increased by Cr. Cr augmented the expression of P2 × 7 receptor by peribronchial leukocytes and by epithelial cells, and increased the accumulation of eosinophils in peribronchial space and of collagen fibers in airway wall. In human cells, while Cr induced a release of ATP, IL-6, and IL-8 from BEAS-2B cells, whole blood cells, such as eosinophils, and CD4+ T cells, P2 × 7 receptor inhibitor (A740003) reduced such effects, as denoted by reduced levels of ATP, IL-6, and IL-8. Therefore, Cr supplementation worsened asthma pathology due to activation of airway epithelial cells and peribronchial leukocytes, involving purinergic signaling.

Effectiveness of mepolizumab therapy in patients with severe eosinophilic asthma: Austrian real-life data.

BACKGROUND: Mepolizumab was effective in several randomized clinical trials (RCTs) in patients with severe eosinophilic asthma, but evidence for symptom control in a real-world population is scarce. OBJECTIVE: To assess asthma symptom control, lung function, use of oral corticosteroids, and biomarkers after mepolizumab initiation in real-world clinical practice. METHODS: Thirty-five adult patients with severe eosinophilic asthma and inadequate asthma symptom control, including former smokers and patients with cardiac disease, were enrolled in a prospective single-arm real-world study. Asthma control tests (ACT), exacerbations, spirometry (pre-bronchodilator forced expiratory volume in 1 s [FEV1]), and oral corticosteroid doses were documented. Further assessments included peripheral blood eosinophil counts and adverse events. RESULTS: After mepolizumab initiation asthma symptom control was significantly improved with the median ACT score of 12.5 at baseline (interquartile range [IQR ]10.5-19.5) rising to 19 (15-22.5) after 4 weeks. The improvement was maintained throughout the observation period of 20 weeks. Likewise, exacerbations were reduced. After 8 weeks of mepolizumab daily OCS doses were reduced from 6.25 mg daily (0-20) at baseline to 2.5 mg daily (0-11.9) at week 8 (P < 0.001). FEV1 remained generally unchanged during the course of the study. Eosinophil counts rapidly declined and remained at a low level during the observation period. No new safety signals were observed in this study. CONCLUSION: Mepolizumab improved asthma symptom control and had a steroid-sparing effect. Efficacy in this real-world study was comparable to RCTs, despite a history of smoking and comorbidities in many of the patients included.

Nucleotide signalling during inflammation.

Inflammatory conditions are associated with the extracellular release of nucleotides, particularly ATP. In the extracellular compartment, ATP predominantly functions as a signalling molecule through the activation of purinergic P2 receptors. Metabotropic P2Y receptors are G-protein-coupled, whereas ionotropic P2X receptors are ATP-gated ion channels. Here we discuss how signalling events through P2 receptors alter the outcomes of inflammatory or infectious diseases. Recent studies implicate a role for P2X/P2Y signalling in mounting appropriate inflammatory responses critical for host defence against invading pathogens or tumours. Conversely, P2X/P2Y signalling can promote chronic inflammation during ischaemia and reperfusion injury, inflammatory bowel disease or acute and chronic diseases of the lungs. Although nucleotide signalling has been used clinically in patients before, research indicates an expanding field of opportunities for specifically targeting individual P2 receptors for the treatment of inflammatory or infectious diseases.

Purinergic Signaling During Immune Cell Trafficking.

Migration and positioning of immune cells is fundamental for their differentiation and recruitment at sites of infection. Besides the fundamental role played by chemokines and their receptors, recent studies demonstrate that a complex network of purinergic signaling events plays a key role in these trafficking events. This process includes the release of nucleotides (such as ATP and ADP) and subsequent autocrine and paracrine signaling events through nucleotide receptors. At the same time, surface-expressed ectoapyrases and nucleotidases convert extracellular nucleotides to adenosine, and adenosine signaling events play additional functional roles in leucocyte trafficking. In this review we revisit classical paradigms of inflammatory cell trafficking in the context of recent studies implicating purinergic signaling events in this process.

P2X7 Deficiency Blocks Lesional Inflammasome Activity and Ameliorates Atherosclerosis in Mice.

BACKGROUND: Extracellular adenosine triphosphate (ATP) binds as a danger signal to purinergic receptor P2X7 and promotes inflammasome assembly and interleukin-1ß expression. We hypothesized a functional role of the signal axis ATP-P2X7 in inflammasome activation and the chronic inflammation driving atherosclerosis. METHODS: P2X7-competent and P2X7-deficient macrophages were isolated and stimulated with lipopolysaccharide, ATP, or both. To assess whether P2X7 may have a role in atherosclerosis, P2X7 expression was analyzed in aortic arches from low density lipoprotein receptor-/- mice consuming a high-cholesterol or chow diet. P2X7+/+ and P2X7-/- low density lipoprotein receptor-/- mice were fed a high-cholesterol diet to investigate the functional role of P2X7 knockout in atherosclerosis. Human plaques were derived from carotid endarterectomy and stained against P2X7. RESULTS: Lipopolysaccharide or ATP stimulation alone did not activate caspase 1 in isolated macrophages. However, priming with lipopolysaccharide, followed by stimulation with ATP, led to an activation of caspase 1 and interleukin-1ß in P2X7-competent macrophages. In contrast, P2X7-deficient macrophages showed no activation of caspase 1 after sequential stimulation while still expressing a basal amount of interleukin-1ß. P2X7 receptor was higher expressed in murine atherosclerotic lesions, particularly by lesional macrophages. After 16 weeks of a high-cholesterol diet, P2X7-deficient mice showed smaller atherosclerotic lesions than P2X7-competent mice (0.162 cm2±0.023 [n=9], P2X7-/- low density lipoprotein receptor-/- : 0.084 cm2±0.01 [n=11], P=0.004) with a reduced amount of lesional macrophages. In accord with our in vitro findings, lesional caspase 1 activity was abolished in P2X7-/- mice. In addition, intravital microscopy revealed reduced leukocyte rolling and adhesion in P2X7-deficient mice. Last, we observe increased P2X7 expression in human atherosclerotic lesions, suggesting that our findings in mice are relevant for human disease. CONCLUSIONS: P2X7 deficiency resolved plaque inflammation by inhibition of lesional inflammasome activation and reduced experimental atherosclerosis. Therefore, P2X7 represents an interesting potential new target to combat atherosclerosis.

Purinergic receptor Y2 (P2Y2)- dependent VCAM-1 expression promotes immune cell infiltration in metabolic syndrome.

Sterile inflammation of visceral fat, provoked by dying adipocytes, links the metabolic syndrome to cardiovascular disease. Danger-associated molecular patterns, such as adenosine triphosphate (ATP), are released by activated or dying cells and orchestrate leukocyte infiltration and inflammation via the purinergic receptor P2Y2. The gene expression of ATP receptor P2Y2 did not change in several tissues in the course of obesity, but was increased within epididymal fat. Adipose tissue from P2Y 2-/- mice consuming high-fat diet (HFD) contained less crown-like structures with a reduced frequency of adipose tissue macrophages (ATMs). This was likely due to decreased leukocyte migration because of missing VCAM-1 exposition on P2Y2 deficient hypertrophic adipose tissue endothelial cells. Accordingly, P2Y 2-/- mice showed blunted traits of the metabolic syndrome: they gained less weight compared to P2Y 2+/+ controls, while intake of food and movement behaviour remained unchanged. Liver and adipose tissue were smaller in P2Y 2-/- animals. Insulin tolerance testing (ITT) performed in obese P2Y 2-/- mice revealed a better insulin sensitivity as well as lower plasma C-peptide and cholesterol levels. We demonstrate that interfering with somatic P2Y2 signalling prevents excessive immune cell deposition in diet-induced obesity (DIO), both attenuating adipose tissue inflammation and ameliorating the metabolic phenotype. Thus, blocking the P2Y2 cascade may be a promising strategy to limit metabolic disease and its sequelae.

C1P Attenuates Lipopolysaccharide-Induced Acute Lung Injury by Preventing NF-κB Activation in Neutrophils.

Recently, ceramide-1-phosphate (C1P) has been shown to modulate acute inflammatory events. Acute lung injury (Arnalich et al. 2000. Infect. Immun. 68: 1942-1945) is characterized by rapid alveolar injury, lung inflammation, induced cytokine production, neutrophil accumulation, and vascular leakage leading to lung edema. The aim of this study was to investigate the role of C1P during LPS-induced acute lung injury in mice. To evaluate the effect of C1P, we used a prophylactic and therapeutic LPS-induced ALI model in C57BL/6 male mice. Our studies revealed that intrapulmonary application of C1P before (prophylactic) or 24 h after (therapeutic) LPS instillation decreased neutrophil trafficking to the lung, proinflammatory cytokine levels in bronchoalveolar lavage, and alveolar capillary leakage. Mechanistically, C1P inhibited the LPS-triggered NF-κB levels in lung tissue in vivo. In addition, ex vivo experiments revealed that C1P also attenuates LPS-induced NF-κB phosphorylation and IL-8 production in human neutrophils. These results indicate C1P playing a role in dampening LPS-induced acute lung inflammation and suggest that C1P could be a valuable candidate for treatment of ALI.