Platelet-activating Factor Receptor Research Articles

Characterization of platelet-activating factor-induced cutaneous edema and erythema in dogs.

OBJECTIVE To characterize platelet-activating factor (PAF)-induced edema and erythema in the skin of dogs and compare those reactions with histamine-induced cutaneous reactions. ANIMALS 6 healthy Beagles. PROCEDURES Experiments were performed at ≥ 2-week intervals. Each dog received ID injections (5 μg/site) of PAF C16, PAF C18, lyso-PAF, and histamine. Edema (mean diameter) and erythema scores (none, mild, moderate, or severe) were assessed 30 minutes after the injections. Dogs received ID injections of PAF and histamine each with various concentrations of WEB 2086 (PAF receptor antagonist) or underwent ID testing with PAF and histamine before and 3 hours after oral administration of cetirizine hydrochloride or prednisolone (at 2 doses each). RESULTS ID injections of PAF C16 and PAF C18, but not lyso-PAF, induced comparable levels of edema and erythema. The PAF-induced edema and erythema peaked at 30 minutes and lasted for 6 hours after the injection; histamine-induced edema and erythema peaked at 30 minutes and lasted for 3 hours after the injection. Edema sizes and erythema scores were significantly smaller and lower, respectively, for PAF than for histamine. The WEB 2086 inhibited PAF-induced but not histamine-induced edema and erythema. Cetirizine slightly, but significantly, repressed PAF-induced edema and erythema as well as histamine-induced cutaneous reactions. Prednisolone suppressed both PAF-induced and histamine-induced edema and erythema. CONCLUSIONS AND CLINICAL RELEVANCE In canine skin, the duration of PAF-induced inflammation was longer than that of histamine-induced inflammation. The PAF- and histamine-induced cutaneous reactions were effectively suppressed by oral administration of prednisolone. The importance of PAF in dogs with anaphylaxis and allergic disorders warrants further investigation.

Mechanisms of anaphylaxis in human low-affinity IgG receptor locus knock-in mice

Anaphylaxis can proceed through distinct IgE- or IgG-dependent pathways, which have been investigated in various mouse models. We developed a novel mouse strain in which the human low-affinity IgG receptor locus, comprising both activating (hFcγRIIA, hFcγRIIIA, and hFcγRIIIB) and inhibitory (hFcγRIIB) hFcγR genes, has been inserted into the equivalent murine locus, corresponding to a locus swap. We sought to determine the capabilities of hFcγRs to induce systemic anaphylaxis and identify the cell types and mediators involved. hFcγR expression on mouse and human cells was compared to validate the model. Passive systemic anaphylaxis was induced by injection of heat-aggregated human intravenous immunoglobulin and active systemic anaphylaxis after immunization and challenge. Anaphylaxis severity was evaluated based on hypothermia and mortality. The contribution of receptors, mediators, or cell types was assessed based on receptor blockade or depletion. The human-to-mouse low-affinity FcγR locus swap engendered hFcγRIIA/IIB/IIIA/IIIB expression in mice comparable with that seen in human subjects. Knock-in mice were susceptible to passive and active anaphylaxis, accompanied by downregulation of both activating and inhibitory hFcγR expression on specific myeloid cells. The contribution of hFcγRIIA was predominant. Depletion of neutrophils protected against hypothermia and mortality. Basophils contributed to a lesser extent. Anaphylaxis was inhibited by platelet-activating factor receptor or histamine receptor 1 blockade. Low-affinity FcγR locus-switched mice represent an unprecedented model of cognate hFcγR expression. Importantly, IgG-related anaphylaxis proceeds within a native context of activating and inhibitory hFcγRs, indicating that, despite robust hFcγRIIB expression, activating signals can dominate to initiate a severe anaphylactic reaction.

Rupatadine: global safety evaluation in allergic rhinitis and urticaria

ABSTRACTIntroduction: Rupatadine is a second-generation H1-antihistamine with dual affinity for histamine H1 and PAF receptors. Rupatadine is indicated for the treatment of allergic rhinitis and urticaria.Areas covered: A Medline search was conducted to identify preclinical and clinical studies of rupatadine. This was supplemented with additional articles obtained from online sources. The focus of this review is on the safety profile of rupatadine.Expert opinion: The review of these data indicates that rupatadine is highly selective for histamine H1-receptors, exhibits additional PAF antagonism in in vitro and in vivo studies, does not cross the blood-brain barrier, and has similar adverse events comparable with other second-generation antihistamines. Rupatadine is a safe and well tolerated drug in patients over 2 years old, with no central nervous system or cardiovascular effects and it can be taken with or without foods.

PAFR activation of NF-κB p65 or p105 precursor dictates pro- and anti-inflammatory responses during TLR activation in murine macrophages.

Platelet-activating factor receptor (PAFR) is a G protein-coupled receptor (GPCR) implicated in many diseases. Toll-like receptors (TLRs) play a critical role in shaping innate and adaptive immune responses. In this study, we investigated whether PAFR signaling changes the macrophages responsiveness to agonists of TLR2 (Pam3Cys), TLR4 (LPS), and TLR3 agonist Poly(I:C). Exogenous PAF inhibited the production of pro-inflammatory cytokines (IL-12p40, IL-6, and TNF-α) and increased anti-inflammatory IL-10 in macrophages challenged with Pam3Cys and LPS, but not with Poly (I:C). PAF did not affect mRNA expression of MyD88, suggesting that PAF acts downstream the adaptor. PAF inhibited LPS-induced phosphorylation of NF-κB p65 and increased NF-κB p105 phosphorylation, which is processed in the proteasome to generate p50 subunit. The PAF potentiation of IL-10 production was dependent on proteasome processing but independent of NF-κB transactivation domain. Inhibition of p50 abolished the PAF-induced IL-10 production. These findings indicate that the impaired transcriptional activity of the p65 subunit and the enhanced p105 phosphorylation induced by PAF are responsible for down regulation of pro-inflammatory cytokines and up regulation of IL-10, respectively, in LPS-challenged macrophages. Together, our data unveil a heretofore unrecognized role for PAFR in modulating activation of NF-κB in macrophages.

Dysfunctional epileptic neuronal circuits and dysmorphic dendritic spines are mitigated by platelet-activating factor receptor antagonism.

Temporal lobe epilepsy or limbic epilepsy lacks effective therapies due to a void in understanding the cellular and molecular mechanisms that set in motion aberrant neuronal network formations during the course of limbic epileptogenesis (LE). Here we show in in vivo rodent models of LE that the phospholipid mediator platelet-activating factor (PAF) increases in LE and that PAF receptor (PAF-r) ablation mitigates its progression. Synthetic PAF-r antagonists, when administered intraperitoneally in LE, re-establish hippocampal dendritic spine density and prevent formation of dysmorphic dendritic spines. Concomitantly, hippocampal interictal spikes, aberrant oscillations, and neuronal hyper-excitability, evaluated 15–16 weeks after LE using multi-array silicon probe electrodes implanted in the dorsal hippocampus, are reduced in PAF-r antagonist-treated mice. We suggest that over-activation of PAF-r signaling induces aberrant neuronal plasticity in LE and leads to chronic dysfunctional neuronal circuitry that mediates epilepsy.

Nuclear localization of platelet-activating factor receptor controls retinal neovascularization.

Platelet-activating factor (PAF) is a pleiotropic phospholipid with proinflammatory, procoagulant and angiogenic actions on the vasculature. We and others have reported the presence of PAF receptor (Ptafr) at intracellular sites such as the nucleus. However, mechanisms of localization and physiologic functions of intracellular Ptafr remain poorly understood. We hereby identify the importance of C-terminal motif of the receptor and uncover novel roles of Rab11a GTPase and importin-5 in nuclear translocation of Ptafr in primary human retinal microvascular endothelial cells. Nuclear localization of Ptafr is independent of exogenous PAF stimulation as well as intracellular PAF biosynthesis. Moreover, nuclear Ptafr is responsible for the upregulation of unique set of growth factors, including vascular endothelial growth factor, in vitro and ex vivo. We further corroborate the intracrine PAF signaling, resulting in angiogenesis in vivo, using Ptafr antagonists with distinct plasma membrane permeability. Collectively, our findings show that nuclear Ptafr translocates in an agonist-independent manner, and distinctive functions of Ptafr based on its cellular localization point to another dimension needed for pharmacologic selectivity of drugs.

An antagonist of the platelet-activating factor receptor inhibits adherence of both nontypeable Haemophilus influenzae and Streptococcus pneumoniae to cultured human bronchial epithelial cells exposed to cigarette smoke.

BackgroundCOPD is emerging as the third largest cause of human mortality worldwide after heart disease and stroke. Tobacco smoking, the primary risk factor for the development of COPD, induces increased expression of platelet-activating factor receptor (PAFr) in the lung epithelium. Nontypeable Haemophilus influenzae (NTHi) and Streptococcus pneumoniae adhere to PAFr on the luminal surface of human respiratory tract epithelial cells.ObjectiveTo investigate PAFr as a potential drug target for the prevention of infections caused by the main bacterial drivers of acute exacerbations in COPD patients, NTHi and S. pneumoniae.MethodsHuman bronchial epithelial BEAS-2B cells were exposed to cigarette smoke extract (CSE). PAFr expression levels were determined using immunocytochemistry and quantitative polymerase chain reaction. The epithelial cells were challenged with either NTHi or S. pneumoniae labeled with fluorescein isothiocyanate, and bacterial adhesion was measured using immunofluorescence. The effect of a well-evaluated antagonist of PAFr, WEB-2086, on binding of the bacterial pathogens to BEAS-2B cells was then assessed. In silico studies of the tertiary structure of PAFr and the binding pocket for PAF and its antagonist WEB-2086 were undertaken.ResultsPAFr expression by bronchial epithelial cells was upregulated by CSE, and significantly associated with increased bacterial adhesion. WEB-2086 reduced the epithelial adhesion by both NTHi and S. pneumoniae to levels observed for non-CSE-exposed cells. Furthermore, it was nontoxic toward the bronchial epithelial cells. In silico analyses identified a binding pocket for PAF/WEB-2086 in the predicted PAFr structure.ConclusionWEB-2086 represents an innovative class of candidate drugs for inhibiting PAFr-dependent lung infections caused by the main bacterial drivers of smoking-related COPD.

SIRT1 prevents pulmonary thrombus formation induced by arachidonic acid via downregulation of PAF receptor expression in platelets

SIRT1, a class III histone deacetylase, is critically involved in cellular response to stress and modulates cardiovascular risk factors. However, its role in thrombus formation is largely unknown. Thus, this study investigated the effect of SIRT1 on pulmonary thrombus formation, and then identified its role in the modulation of platelet aggregation. In isolated human platelets, cell aggregation was increased by various platelet activators, such as platelet activating factor (PAF), arachidonic acid (AA), ADP, and thrombin. AA- and PAF-mediated platelet aggregations were suppressed by WEB2086, a PAF receptor (PAFR) antagonist. Pulmonary thrombus formation induced by PAF or AA was also attenuated by WEB2086, suggesting that PAFR plays a key role in AA-induced platelet aggregation. In platelets isolated from SIRT1-TG mice as well as in platelets treated with resveratrol or reSIRT1, PAFR expression was decreased, whereas this expressional downregulation by SIRT1 activators was inhibited in platelets treated with MG132 (a proteasome inhibitor) or NH4Cl (a lysosome inhibitor). Furthermore, platelet aggregation induced by AA was markedly attenuated by resveratrol and reSIRT1. Likewise, the increased pulmonary thrombus formation in mice treated with AA was also attenuated by SIRT1 activators. In line with these results, pulmonary thrombus formation was markedly attenuated in SIRT1-TG mice. Taken together, this study showed that SIRT1 downregulates PAFR expression on platelets via proteasomal and lysosomal pathways, and that this downregulation inhibits platelet aggregation in vitro and pulmonary thrombus formation in vivo.

Platelet Activating Factor (PAF) Receptor Deletion or Antagonism Attenuates Severe HSV-1 Meningoencephalitis.

Herpes simplex virus type 1 (HSV-1) is a human pathogen that may cause severe encephalitis. The exacerbated immune response against the virus contributes to the disease severity and death. Platelet activating factor (PAF) is a mediator capable of inducing increase in vascular permeability, production of cytokines on endothelial cells and leukocytes. We aimed to investigate the activation of PAF receptor (PAFR) and its contribution to the severity of the inflammatory response in the brain following HSV-1 infection. C57BL/6 wild-type (WT) and PAFR deficient (PAFR-/-) mice were inoculated intracranially with 104 plaque-forming units (PFU) of HSV-1. Visualization of leukocyte recruitment was performed using intravital microscopy. Cells infiltration in the brain tissue were analyzed by flow cytometry. Brain was removed for chemokine assessment by ELISA and for histopathological analysis. The pharmacological inhibition by the PAFR antagonist UK-74,505 was also analyzed. In PAFR-/- mice, there was delayed lethality but no difference in viral load. Histopathological analysis of infected PAFR-/- mice showed that brain lesions were less severe when compared to their WT counterparts. Moreover, PAFR-/- mice showed less TCD4+, TCD8+ and macrophages in brain tissue. This reduction of the presence of leukocytes in parenchyma may be mechanistically explained by a decrease in leukocytes rolling and adhesion. PAFR-/- mice also presented a reduction of the chemokine CXCL9 in the brain. In addition, by antagonizing PAFR, survival of C57BL/6 infected mice increased. Altogether, our data suggest that PAFR plays a role in the pathogenesis of experimental HSV-1 meningoencephalitis, and its blockade prevents severe disease manifestation.

Mediators of vascular leak in dengue infections

Although plasma leakage is the hallmark of severe dengue infections the factors that cause increased vascular permeability have not been identified. As platelet activating factor (PAF), lipid mediators and cytokines are associated with increase in vascular permeability, we set out to investigate the role of these mediators in acute dengue infection. Levels of inflammatory lipid mediators and cytokines were initially evaluated in 25 patients with confirmed acute dengue infection and 12 healthy individuals. After determining which inflammatory mediators were associated with severe dengue, serial assays of the above lipid mediators, and cytokines were done in an additional 36 patients in serum samples collected 12 hours apart throughout the course of their hospital stay. We found that PAF levels were significantly higher in patients (p=0.001) when compared to healthy individuals and that PAF levels rose just before the onset of the critical phase. PAF values of >100ng/ml were associated with reduced expression of gap junction proteins leading to increased vascular permeability. A similar effect was seen with serum from patients with dengue infection which was reversed by a PAF receptor antagonist. S1P levels which are associated with increasing the integrity of the endothelial barrier were significantly reduced in patients with DHF. Serum IL-10, TNFα and IL-1β levels were elevated in patients with DHF and followed the same patterns as PAF. In conclusion, our results show that PAF is likely to be a potent mediator of vascular leak in dengue and use of PAF blockers could be therapeutically useful.

Platelet-activating factor receptor activation promotes prostate cancer cell growth, invasion and metastasis via ERK1/2 pathway.

Platelet-activating factor (PAF) and its receptor (PAFR), have been reported to participate in many cellular processes of cancer. However, little is known about their function in prostate cancer. In the present study, we found that PAFR was overexpressed in prostate cancer cells. PAF stimulation dose-dependently promoted the invasion, migration and growth of prostate cancer cells invitro, while knockdown of PAFR inhibited the effect of PAF on prostate cancer cells. We further found that PAFR promoted prostate cancer cell growth and metastasis invivo. Moreover, we found that PAFR activation increased MMP-3 expression and decreased E-cadherin expression of prostate cancer cells invitro and invivo. Finally, we found that PAFR time-dependently induced activation of ERK1/2, and ERK1/2 pathway contributed to PAFR-mediated prostate cancer cell invasion, migration and growth. Together, our findings demonstrate that PAFR can activate ERK1/2 pathway, and subsequently increase MMP-3 expression and decrease E-cadherin expression, which finally promote prostate cancer cell growth, invasion and metastasis. Thus, PAFR may act as a potential target for therapeutic use of prostate cancer.

Influence of platelet-activating factor receptor (PAFR) on Brucella abortus infection: implications for manipulating the phagocytic strategy of B. abortus.

BackgroundBrucella abortus is an intracellular pathogen which can infect and persist in host cells through multiple interactions. Above all, its interaction to host cell receptor is important to understand the pathogenic mechanisms of B. abortus. Accordingly, we demonstrated that platelet-activating factor receptor (PAFR) affects host cell response against B. abortus infection.ResultsFirst of all, B. abortus infection to macrophage induces secretion of platelet-activating factor (PAF), which is a PAFR agonist. The stimulation of PAFR by PAF remarkably increases B. abortus uptake into macrophages. It induces Janus kinase 2 (JAK2) and p38α phosphorylation, indicating that PAFR-mediated activation of JAK2 signaling leads to enhanced uptake of B. abortus. Moreover, the dynamics of F-actin polymerization revealed that PAFR-mediated B. abortus uptake is related with the reorganization of F-actin and JAK2. Upon B. abortus phagocytosis, reduced PAFR in the membrane and subsequently increased levels of PAFR colocalization with endosomes were observed which indicate that B. abortus uptake into macrophages allowed PAFR trafficking to endosomes.ConclusionsThis study demonstrated that PAFR has a compelling involvement in B. abortus uptake as a promoter of phagocytosis, which is associated with JAK2 activation. Thus, our findings establish a novel insight into a receptor-related phagocytic mechanism of B. abortus.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-016-0685-8) contains supplementary material, which is available to authorized users.

590 UVB generates microvesicle particles via Platelet-activating Factor-receptor signaling: a novel pathway by which a skin-specific stimulus exerts systemic effects

590 UVB generates microvesicle particles via Platelet-activating Factor-receptor signaling: a novel pathway by which a skin-specific stimulus exerts systemic effects

Lipopolysaccharide Cross-Tolerance Delays Platelet-Activating Factor-Induced Sudden Death in Swiss Albino Mice: Involvement of Cyclooxygenase in Cross-Tolerance.

Lipopolysaccharide (LPS) signaling through Toll-like receptor-4 (TLR-4) has been implicated in the pathogenesis of many infectious diseases. Some believe that TLR-mediated pathogenicity is due, in part, to the lipid pro-inflammatory mediator platelet-activating factor (PAF), but this has been questioned. To test the direct contribution of PAF in endotoxemia in murine models, we injected PAF intraperitoneally into Swiss albino mice in the presence and absence of LPS. PAF alone (5 μg/mouse) caused death within 15–20 min, but this could be prevented by pretreating mice with PAF-receptor (PAF-R) antagonists or PAF-acetylhydrolase (PAF-AH). A low dose of LPS (5 mg/kg body wt) did not impair PAF-induced death, whereas higher doses (10 or 20 mg/kg body wt) delayed death, probably via LPS cross-tolerance. Cross-tolerance occurred only when PAF was injected simultaneously with LPS or within 30 min of LPS injection. Tolerance does not appear to be due to an abundant soluble mediator. Histologic examination of lungs and liver and measurement of circulating TNF-α and IL-10 levels suggested that the inflammatory response is not diminished during cross-tolerance. Interestingly, aspirin, a non-specific cyclooxygenase (COX) inhibitor, partially blocked PAF-induced sudden death, whereas NS-398, a specific COX-2 inhibitor, completely protected mice from the lethal effects of PAF. Both COX inhibitors (at 20 mg/kg body wt) independently amplified the cross-tolerance exerted by higher dose of LPS, suggesting that COX-derived eicosanoids may be involved in these events. Thus, PAF does not seem to have a protective role in endotoxemia, but its effects are delayed by LPS in a COX-sensitive way. These findings are likely to shed light on basic aspects of the endotoxin cross-tolerance occurring in many disease conditions and may offer new opportunities for clinical intervention.

AChE Inhibition-based Multi-target-directed Ligands, a Novel Pharmacological Approach for the Symptomatic and Disease-modifying Therapy of Alzheimer's Disease.

Alzheimer's disease (AD) is the most common form of dementia in elder people, characterised by a progressive decline in memory as a result of an impairment of cholinergic neurotransmission. To date acetylcholinesterase inhibitors (AChEIs) have become the most prescribed drugs for the symptomatic treatment of mild to moderate AD. However, the traditional “one molecule-one target” paradigm is not sufficient and appropriate to yield the desired therapeutic efficacy since multiple factors, such as amyloid-β (Aβ) deposits, neuroinflammation, oxidative stress, and decreased levels of acetylcholine (ACh) have been thought to play significant roles in the AD pathogenesis. New generation of multi-target drugs is earnestly demanded not only for ameliorating symptoms but also for modifying the disease. Herein, we delineated the catalytic and non-catalytic functions of AChE, and summarized the works of our group and others in research and development of novel AChEI-based multi-target-directed ligands (MTDLs), such as dual binding site AChEIs and multi-target AChEIs inhibiting Aβ aggregation, regulating Aβ procession, antagonizing platelet-activating factor (PAF) receptor, scavenging oxygen radical, chelating metal ions, inhibiting monoamine oxidase B (MAO-B), blocking N-methyl-D-aspartic acid (NMDA) receptor and others.

Implications For Platelet Activating Factor Receptor Antagonism For Managing Bladder Cancer Progression

Cigarette smoking is the number one risk factor for bladder cancer development and epidemiological data suggests that nearly half of all bladder cancer patients have a history of smoking. In addition to stimulating growth of a primary tumor, it has been shown that there is a correlation between smoking and tumor metastasis. Platelet activating factor (PAF) is an inflammatory cell recruiter that is expressed on the endothelium during times of inflammation and through binding with the PAF‐receptor (PAFR) facilitates transendothelial migration of circulating inflammatory cells. PAF has been implicated in primary tumor growth and interactions with the PAFR have been shown to stimulate cellular growth. PAF is generated through hydrolysis of membrane phospholipids by the activity of calcium‐independent phospholipase A2 (iPLA2) which removes the sn‐‐2 fatty acid yielding a free fatty acid, arachidonic acid, and a lysophospholipid. Acetylation of the newly formed lysophospholipid results in PAF production. Under normal circumstances, PAF is maintained at a low concentration by PAF‐acetylhydrolase (PAF‐AH) resulting in the inactive lyso‐PAF. In the present study, we show that cigarette smoke extract (CSE) exposure of bladder cancer cells inhibits PAF‐AH activity and increases iPLA‐2 activity resulting in increased PAF accumulation. In addition to PAF accumulation, CSE increases PAFR expression. Furthermore, by immunohistochemical analysis of tumor biopsy samples from bladder cancer patients we detected increased PAF, iPLA2 and the PAFR in tumor regions when compared to surrounding tissue. This data highlights a pathway that is upregulated in tumor areas of bladder cancer patients and that is influenced by CSE which could facilitate primary tumor growth and increase metastatic potential. Targeting of the PAF‐PAFR interaction could serve as a beneficial therapeutic target for managing further growth of a developing tumor.

Cigarette Smoke‐Induced Bladder Cancer Initiation

Tobacco smoking is a major risk factor associated with the development and metastasis of bladder cancer. Muscle‐invasive bladder cancer with a high risk of metastasis is the most common cause of mortality amongst bladder cancer patients, with a 5‐year survival rate below 10%. Cigarette smoking and increased metastasis correlate, but the underlying mechanism is unknown. Platelet activating factor (PAF) is a potent inflammatory mediator that we have shown to be involved in attachment of tumor cells to the endothelium lining the vasculature following cigarette smoke extract exposure. Attachment to endothelial cells occurs through the interaction between the tumor cell PAF‐receptor (PAFR) and PAF on the endothelial cell surface. The PAF/PAFR interaction facilitates transendothelial cell migration of tumor cells into the circulation to establish secondary tumor sites. PAF has also been shown to promote growth of the primary tumor via angiogenesis, allowing for increased nutrient supply to the developing tumor. Pigment epithelium‐derived factor (PEDF) is an anti‐angiogenic protein that has been shown to be down regulated in higher grade tumors. Preliminary experiments in which we exposed mice to cigarette smoke for 4 weeks indicated signs of urothelial cell hyperplasia, increased basal PAF expression, reduced PEDF expression and nuclear localization of β‐catenin. These observed changes indicate that PAF accumulation and PEDF down regulation could increase growth of primary bladder tumors as well as increase the potential for metastasis. This work highlights two potentially exciting therapeutic targets for managing bladder cancer: targeting the PAF‐PAFR interaction or increasing expression of PEDF.

Platelet-activating factor receptor (PAFR) plays a crucial role in experimental global cerebral ischemia and reperfusion.

Stroke is one of the most frequent causes of death and disability worldwide leading to a significant clinical and socioeconomic burden. Although different mechanisms are involved in the pathogenesis of stroke, inflammatory response occurs after ischemia and contributes to the expansion of brain injury. Platelet-activating factor receptor (PAF) plays crucial roles in both physiological and pathological conditions in the brain. PAF receptor (PAFR) may be expressed on cellular and nuclear membranes of various cell types, especially leukocytes, platelets, endothelial cells, neuronal cells and microglia. Herein, using mice lacking the PAFR receptor (PAFR(-/-)), we investigate a potential role for this receptor during experimental transient global cerebral ischemia and reperfusion (BCCAo). In PAFR deficiency, we observed a significant improvement in the neurological deficits, which were associated with a reduction of brain infarcted area as evaluated by triphenyltetrazolium chloride (TTC). Moreover, a decrease in the percentage of necrotic cavities areas and in the frequency of ischemic neurons was also found by employing histometric analysis. In addition, in PAFR(-/-) mice there was prevention of caspase-3 activation and decreased vascular permeability and brain edema. Decreased brain levels of the cytokines tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta) and the chemokine (C-X-C motif) ligand 1 (CXCL1) by ELISA were also detected in PAFR(-/-) BCCAo animals. Taken together, our results suggest that PAFR activation might be crucial for the global brain ischemia and reperfusion injury.

UVB Generates Microvesicle Particle Release in Part Due to Platelet-activating Factor Signaling.

The lipid mediator platelet-activating factor (PAF) and oxidized glycerophosphocholine PAF agonists produced by ultraviolet B (UVB) have been demonstrated to play a pivotal role in UVB-mediated processes, from acute inflammation to delayed systemic immunosuppression. Recent studies have provided evidence that microvesicle particles (MVPs) are released from cells in response to various signals including stressors. Importantly, these small membrane fragments can interact with various cell types by delivering bioactive molecules. The present studies were designed to test if UVB radiation can generate MVP release from epithelial cells, and the potential role of PAF receptor (PAF-R) signaling in this process. We demonstrate that UVB irradiation of the human keratinocyte-derived cell line HaCaT resulted in the release of MVPs. Similarly, treatment of HaCaT cells with the PAF-R agonist carbamoyl PAF also generated equivalent amounts of MVP release. Of note, pretreatment of HaCaT cells with antioxidants blocked MVP release from UVB but not PAF-R agonist N-methyl carbamyl PAF (CPAF). Importantly, UVB irradiation of the PAF-R-negative human epithelial cell line KB and KB transduced with functional PAF-Rs resulted in MVP release only in PAF-R-positive cells. These studies demonstrate that UVB can generate MVPs in vitro and that PAF-R signaling appears important in this process.

PAFR in adipose tissue macrophages is associated with anti-inflammatory phenotype and metabolic homoeostasis.

Metabolic dysfunction is associated with adipose tissue inflammation and macrophage infiltration. PAFR (platelet-activating factor receptor) is expressed in several cell typesand binds to PAF (platelet-activating factor) and oxidized phospholipids. Engagement of PAFR in macrophages drives them towards the anti-inflammatory phenotype. In the present study, we investigated whether genetic deficiency of PAFR affects the phenotype of ATMs (adipose tissue macrophages) and its effect on glucose and insulin metabolism. PARFKO (PAFR-knockout) and WT (wild-type) mice were fed on an SD (standard diet) or an HFD (high-fat diet). Glucose and insulin tolerance tests were performed by blood monitoring. ATMs were evaluated by FACS for phenotypic markers. Gene and protein expression was investigated by real-time reverse transcription-quantitative PCR and Western blotting respectively. Results showed that the epididymal adipose tissue of PAFRKO mice had increased gene expression of Ccr7, Nos2, Il6 and Il12, associated with pro-inflammatory mediators, and reduced expression of the anti-inflammatory Il10. Moreover, the adipose tissue of PAFRKO mice presented more pro-inflammatory macrophages, characterized by an increased frequency of F4/80(+)CD11c(+) cells. Blood monocytes of PAFRKO mice also exhibited a pro-inflammatory phenotype (increased frequency of Ly6C(+) cells) and PAFR ligands were detected in the serum of both PAFRKO and WT mice. Regarding metabolic parameters, compared with WT, PAFRKO mice had: (i) higher weight gain and serum glucose concentration levels; (ii) decreased insulin-stimulated glucose disappearance; (iii) insulin resistance in the liver; (iv) increased expression of Ldlr in the liver. In mice fed on an HFD, some of these changes were potentiated, particularly in the liver. Thus it seems that endogenous ligands of PAFR are responsible for maintaining the anti-inflammatory profile of blood monocytes and ATMs under physiological conditions. In the absence of PAFR signalling, monocytes and macrophages acquire a pro-inflammatory phenotype, resulting in adipose tissue inflammation and metabolic dysfunction.