Effects of bovine serum albumin (BSA) on the excited-state properties of meso-tetrakis(sulfonatophenyl) porphyrin (TPPS4).

To infer changes in the photophysical properties of porphyrins due to complexation with albumin, a combination of Z-scan and conventional spectroscopic techniques was employed. We measured the characteristics of excited states of meso-tetrakis(sulfonatophenyl) porphyrin bound to bovine serum albumin and observed that the binding reduces the intersystem crossing quantum yield and increases the internal conversion one. A reverse saturable absorption process was observed in the nanosecond timescale. These results are important for prediction of the efficiency of this complex in medical and optical applications, because associating porphyrins to proteins enables better accumulation in tumors and improves its stability in optical devices, but at the same time, decreases its triplet quantum yield.

Neuroimmune-Glia Interactions in the Sensory Ganglia Account for the Development of Acute Herpetic Neuralgia.

Herpetic neuralgia is the most important symptom of herpes zoster disease, which is caused by Varicella zoster Nevertheless, the pathophysiological mechanisms involved in herpetic neuralgia are not totally elucidated. Here, we examined the neuroimmune interactions at the sensory ganglia that account for the genesis of herpetic neuralgia using a murine model of Herpes Simplex Virus Type-1 (HSV-1) infection. The cutaneous HSV-1 infection of mice results in the development of a zosteriform-like skin lesion followed by a time-dependent increase in pain-like responses (mechanical allodynia). Leukocytes composed mainly of macrophages and neutrophils infiltrate infected DRGs and account for the development of herpetic neuralgia. Infiltrating leukocytes are responsible for driving the production of TNF, which in turn mediates the development of herpetic neuralgia through downregulation of the inwardly rectifying K+ channel Kir4.1 in satellite glial cells. These results revealed that neuroimmune-glia interactions at the sensory ganglia play a critical role in the genesis of herpetic neuralgia. In conclusion, the present study elucidates novel mechanisms involved in the genesis of acute herpetic pain and open new avenues for its control.SIGNIFICANCE STATEMENT Acute herpetic neuralgia is the most important symptom of herpes zoster disease and it is very difficult to treat. Using a model of peripheral infection of mice with HSV-1, we have characterized for the first time the neuroimmune-glia interactions in the sensory ganglia that account for the development of acute herpetic neuralgia. Among these mechanisms, leukocytes composed mainly of macrophages and neutrophils infiltrate infected sensory ganglia and are responsible for driving the production of TNF. TNF, via TNFR1, mediates herpetic neuralgia development through downregulation of the inwardly rectifying K+ channel Kir4.1 in satellite glial cells. This study elucidates novel mechanisms involved in the genesis of acute herpetic neuralgia and open new avenues for its control.

Photodynamic inactivation of Bovine herpesvirus type 1 (BoHV-1) by porphyrins.

In this work, the photodynamic efficiency of anionic meso-tetrakis sulfonophenyl (TPPS4), cationic meso-tetrakis methylpyridiniumyl (TMPyP) and their zinc complexes (ZnTPPS4 and ZnTMPyP) in the inactivation of Bovine herpesvirus type 1 (BoHV-1) was evaluated. At a non-cytotoxic concentration, all porphyrins showed significant antiviral activity after irradiation using a halogen lamp. The efficiency of the cationic porphyrins was higher than that of the anionic ones. Porphyrin complexation with zinc increases its lipophilicity and the number of absorbed photons, dramatically reducing the time for complete virus inactivation. The high superposition of the compound optical absorption and light source emission spectra played a key role in the virus inactivation efficiency. The results demonstrated the high effectivity of the photodynamic inactivation of BoHV-1. This method can be used as an auxiliary in the treatment of disorders attributed to BoHV-1 infection, and the porphyrins are promising photosensitizers for this application.

Spinal cord oligodendrocyte-derived alarmin IL-33 mediates neuropathic pain.

Neuropathic pain from injury to the peripheral and CNS represents a major health care issue. We have investigated the role of IL-33/IL-33 receptor (ST2) signaling in experimental models of neuropathic pain in mice. Chronic constriction injury (CCI) of the sciatic nerve induced IL-33 production in the spinal cord. IL-33/citrine reporter mice revealed that oligodendrocytes are the main cells expressing IL-33 within the spinal cord together with a minor expression by neurons, microglia. and astrocytes. CCI-induced mechanical hyperalgesia was reduced in IL-33R (ST2)(-/ -) mice compared with wild-type (WT) mice. Intrathecal treatment of WT mice with soluble IL-33 receptor (IL-33 decoy receptor) markedly reduced CCI-induced hyperalgesia. Consistent with these observations, intrathecal injection of IL-33 enhanced CCI hyperalgesia and induced hyperalgesia in naive mice. IL-33-mediated hyperalgesia during CCI was dependent on a reciprocal relationship with TNF-α and IL-1ß. IL-33-induced hyperalgesia was markedly attenuated by inhibitors of PI3K, mammalian target of rapamycin, MAPKs (p38, ERK, and JNK), NF-κB, and also by the inhibitors of glial cells (microglia and astrocytes). Furthermore, targeting these signaling pathways and cells inhibited IL-33-induced TNF-α and IL-1ß production in the spinal cord. Our study, therefore, reveals an important role of oligodendrocyte-derived IL-33 in neuropathic pain.

Targeting the minor pocket of C5aR for the rational design of an oral allosteric inhibitor for inflammatory and neuropathic pain relief.

Chronic pain resulting from inflammatory and neuropathic disorders causes considerable economic and social burden. Pharmacological therapies currently available for certain types of pain are only partially effective and may cause severe adverse side effects. The C5a anaphylatoxin acting on its cognate G protein-coupled receptor (GPCR), C5aR, is a potent pronociceptive mediator in several models of inflammatory and neuropathic pain. Although there has long been interest in the identification of C5aR inhibitors, their development has been complicated, as for many peptidomimetic drugs, mostly by poor drug-like properties. Herein, we report the de novo design of a potent and selective C5aR noncompetitive allosteric inhibitor, DF2593A, guided by the hypothesis that an allosteric site, the "minor pocket," previously characterized in CXC chemokine receptors-1 and -2, is functionally conserved in the GPCR class. In vitro, DF2593A potently inhibited C5a-induced migration of human and rodent neutrophils. In vivo, oral administration of DF2593A effectively reduced mechanical hyperalgesia in several models of acute and chronic inflammatory and neuropathic pain, without any apparent side effects. Mechanical hyperalgesia after spared nerve injury was also reduced in C5aR(-/-) mice compared with WT mice. Furthermore, treatment of C5aR(-/-) mice with DF2593A did not produce any further antinociceptive effect compared with C5aR(-/-) mice treated with vehicle. The successful medicinal chemistry strategy confirms that a conserved minor pocket is amenable for the rational design of selective inhibitors and the pharmacological results support that the allosteric blockade of the C5aR represents a highly promising therapeutic approach to control chronic inflammatory and neuropathic pain.

Activation of the STING adaptor attenuates experimental autoimmune encephalitis.

Cytosolic DNA sensing activates the stimulator of IFN genes (STING) adaptor to induce IFN type I (IFN-αß) production. Constitutive DNA sensing to induce sustained STING activation incites tolerance breakdown, leading to autoimmunity. In this study, we show that systemic treatments with DNA nanoparticles (DNPs) induced potent immune regulatory responses via STING signaling that suppressed experimental autoimmune encephalitis (EAE) when administered to mice after immunization with myelin oligodendrocyte glycoprotein (MOG), at EAE onset, or at peak disease severity. DNP treatments attenuated infiltration of effector T cells into the CNS and suppressed innate and adaptive immune responses to myelin oligodendrocyte glycoprotein immunization in spleen. Therapeutic responses were not observed in mice treated with cargo DNA or cationic polymers alone, indicating that DNP uptake and cargo DNA sensing by cells with regulatory functions was essential for therapeutic responses to manifest. Intact STING and IFN-αß receptor genes, but not IFN-γ receptor genes, were essential for therapeutic responses to DNPs to manifest. Treatments with cyclic diguanylate monophosphate to activate STING also delayed EAE onset and reduced disease severity. Therapeutic responses to DNPs were critically dependent on IDO enzyme activity in hematopoietic cells. Thus, DNPs and cyclic diguanylate monophosphate attenuate EAE by inducing dominant T cell regulatory responses via the STING/IFN-αß/IDO pathway that suppress CNS-specific autoimmunity. These findings reveal dichotomous roles for the STING/IFN-αß pathway in either stimulating or suppressing autoimmunity and identify STING-activating reagents as a novel class of immune modulatory drugs.

Characterization of a Cryptic and Intriguing Low Molecular Weight Plasmid.

The complete nucleotide sequence of cryptic plasmid pVCM04 isolated from Salmonella enterica serovar Enteritidis was determined and analyzed. pVCM04 contains 3853 bp with 53.6 % GC content and has twelve ORFs with more than 50 amino acids. Five of these sequences showed homology with replication and mobilization proteins. ORF1 and ORF2 showed homology with replication proteins, while ORFs 3-5 showed homology with mobilization proteins. The pVCM04 possesses a region associated with the theta-type replication mechanism. BLASTn search analysis revealed unexpectedly no similarity with sequences deposited in GenBank. The nucleotide sequence of pVCM04 can be divided into two arms: the region between nucleotides 552-1774 (encoding RepA and RepB) and the region between nucleotides 1775-3853 (encoding MobA, MobB and MobC). Codon bias pattern is distinct between mobA and repA, so the program Modeltest was used to select the best evolutionary model to study these genes. The result of ModelTest (model GTR+G for mobA and model HKY+G for repA) suggests that these genes would be subject to different selective pressures. Considering the differences in the codon usage, the selection of two different evolutionary models, and the absence of plasmids with homology to pVCM04 in GenBank, we believe that pVCM04 is a chimeric molecule and represents a new plasmid lineage.

Fractalkine mediates inflammatory pain through activation of satellite glial cells.

The activation of the satellite glial cells (SGCs) surrounding the dorsal root ganglion (DRG) neurons appears to play a role in pathological pain. We tested the hypothesis that fractalkine, which is constitutively expressed by primary nociceptive neurons, is the link between peripheral inflammation and the activation of SGCs and is thus responsible for the genesis of the inflammatory pain. The injection of carrageenin into the rat hind paw induced a decrease in the mechanical nociceptive threshold (hypernociception), which was associated with an increase in mRNA and GFAP protein expression in the DRG. Both events were inhibited by anti-fractalkine antibody administered directly into the DRG (L5) [intraganglionar (i.gl.)]. The administration of fractalkine into the DRG (L5) produced mechanical hypernociception in a dose-, time-, and CX3C receptor-1 (CX3CR1)-dependent manner. Fractalkine's hypernociceptive effect appears to be indirect, as it was reduced by local treatment with anti-TNF-α antibody, IL-1-receptor antagonist, or indomethacin. Accordingly, the in vitro incubation of isolated and cultured SGC with fractalkine induced the production/release of TNF-α, IL-1ß, and prostaglandin E2. Finally, treatment with i.gl. fluorocitrate blocked fractalkine (i.gl.)- and carrageenin (paw)-induced hypernociception. Overall, these results suggest that, during peripheral inflammation, fractalkine is released in the DRG and contributes to the genesis of inflammatory hypernociception. Fractalkine's effect appears to be dependent on the activation of the SGCs, leading to the production of TNFα, IL-1ß, and prostanoids, which are likely responsible for the maintenance of inflammatory pain. Thus, these results indicate that the inhibition of fractalkine/CX3CR1 signaling in SGCs may serve as a target to control inflammatory pain.

Morphine peripheral analgesia depends on activation of the PI3Kgamma/AKT/nNOS/NO/KATP signaling pathway.

Morphine is one of the most prescribed and effective drugs used for the treatment of acute and chronic pain conditions. In addition to its central effects, morphine can also produce peripheral analgesia. However, the mechanisms underlying this peripheral action of morphine have not yet been fully elucidated. Here, we show that the peripheral antinociceptive effect of morphine is lost in neuronal nitric-oxide synthase null mice and that morphine induces the production of nitric oxide in primary nociceptive neurons. The activation of the nitric-oxide pathway by morphine was dependent on an initial stimulation of PI3Kgamma/AKT protein kinase B (AKT) and culminated in increased activation of K(ATP) channels. In the latter, this intracellular signaling pathway might cause a hyperpolarization of nociceptive neurons, and it is fundamental for the direct blockade of inflammatory pain by morphine. This understanding offers new targets for analgesic drug development.

Synergic vascular photodynamic activity by methylene blue-curcumin supramolecular assembly.

A supramolecular assembly was obtained by combining methylene blue (MB) with a natural plant extract, curcumin (Curc), in a stoichiometric ratio of 1:4 in aqueous solution (90% PBS + 10% ethanol) at room temperature. The MB-Curc supramolecular assembly was evidenced by absorption and fluorescence spectroscopies, and the stoichiometry and bonding constant were obtained using Cielens model. Its stability and photostability were evaluated by chromatographic analysis and UV-Vis absorption. The MB-Curc avoids the aggregation of both isolated compounds and efficiently produces singlet oxygen (ΦΔ= 0.52 ± 0.03). Its potential for photodynamic antiangiogenic treatments was evaluated through the vascular effect observed in chicken chorioallantoic membrane (CAM) assay. The results showed intense damage in CAM vascular network by MB-Curc after irradiation, which is higher than the effect of isolated compounds, indicating a synergistic vascular effect. This combination can be essential to prevent cancer revascularization after photodynamic application and improve the efficacy of this approach. The characteristics exhibited by MB-Curc make it a potential candidate for use in cancer treatments through photodynamic antiangiogenic therapy.

Stimulation of peripheral kappa opioid receptors inhibits inflammatory hyperalgesia via activation of the PI3Kγ/AKT/nNOS/NO signaling pathway.

BACKGROUND: In addition to their central effects, opioids cause peripheral analgesia. There is evidence showing that peripheral activation of kappa opioid receptors (KORs) inhibits inflammatory pain. Moreover, peripheral µ-opioid receptor (MOR) activation are able to direct block PGE(2)-induced ongoing hyperalgesia However, this effect was not tested for KOR selective activation. In the present study, the effect of the peripheral activation of KORs on PGE(2)-induced ongoing hyperalgesia was investigated. The mechanisms involved were also evaluated. RESULTS: Local (paw) administration of U50488 (a selective KOR agonist) directly blocked, PGE(2)-induced mechanical hyperalgesia in both rats and mice. This effect was reversed by treating animals with L-NMMA or N-propyl-L-arginine (a selective inhibitor of neuronal nitric oxide synthase, nNOS), suggesting involvement of the nNOS/NO pathway. U50488 peripheral effect was also dependent on stimulation of PI3Kγ/AKT because inhibitors of these kinases also reduced peripheral antinociception induced by U50488. Furthermore, U50488 lost its peripheral analgesic effect in PI3Kγ null mice. Observations made in vivo were confirmed after incubation of dorsal root ganglion cultured neurons with U50488 produced an increase in the activation of AKT as evaluated by western blot analyses of its phosphorylated form. Finally, immunofluorescence of DRG neurons revealed that KOR-expressing neurons also express PI3Kγ (≅ 43%). CONCLUSIONS: The present study indicates that activation of peripheral KORs directly blocks inflammatory hyperalgesia through stimulation of the nNOS/NO signaling pathway which is probably stimulated by PI3Kγ/AKT signaling. This study extends a previously study of our group suggesting that PI3Kγ/AKT/nNOS/NO is an important analgesic pathway in primary nociceptive neurons.

Endothelin-1 induces neutrophil recruitment in adaptive inflammation via TNFα and CXCL1/CXCR2 in mice.

Endothelin mediates neutrophil recruitment during innate inflammation. Herein we address whether endothelin-1 (ET-1) is involved in neutrophil recruitment in adaptive inflammation in mice, and its mechanisms. Pharmacological treatments were used to determine the role of endothelin in neutrophil recruitment to the peritoneal cavity of mice challenged with antigen (ovalbumin) or ET-1. Levels of ET-1, tumour necrosis factor α (TNFα), and CXC chemokine ligand 1 (CXCL1) were determined by enzyme-linked immunosorbent assay. Neutrophil migration and flow cytometry analyses were performed 4 h after the intraperitoneal stimulus. ET-1 induced dose-dependent neutrophil recruitment to the peritoneal cavity. Treatment with the non-selective ET(A)/ET(B) receptor antagonist bosentan, and selective ET(A) or ET(B) receptor antagonists BQ-123 or BQ-788, respectively, inhibited ET-1- and ovalbumin-induced neutrophil migration to the peritoneal cavity. In agreement with the above, the antigen challenge significantly increased levels of ET-1 in peritoneal exudates. The ET-1- and ovalbumin-induced neutrophil recruitment were reduced in TNFR1 deficient mice, and by treatments targeting CXCL1 or CXC chemokine receptor 2 (CXCR2); further, treatment with bosentan, BQ-123, or BQ-788 inhibited ET-1- and antigen-induced production of TNFα and CXCL1. Furthermore, ET-1 and ovalbumin challenge induced an increase in the number of cells expressing the Gr1(+) markers in the granulocyte gate, CD11c(+) markers in the monocyte gate, and CD4(+) and CD45(+) (B220) markers in the lymphocyte gate in an ET(A)- and ET(B)-dependent manner, as determined by flow cytometry analysis, suggesting that ET-1 might be involved in the recruitment of neutrophils and other cells in adaptive inflammation. Therefore, the present study demonstrates that ET-1 is an important mediator for neutrophil recruitment in adaptive inflammation via TNFα and CXCL1/CXCR2-dependent mechanism.

Meningeal dendritic cells drive neuropathic pain through elevation of the kynurenine metabolic pathway in mice.

Neuropathic pain is one of the most important clinical consequences of injury to the somatosensory system. Nevertheless, the critical pathophysiological mechanisms involved in neuropathic pain development are poorly understood. In this study, we found that neuropathic pain is abrogated when the kynurenine metabolic pathway (KYNPATH) initiated by the enzyme indoleamine 2,3-dioxygenase 1 (IDO1) is ablated pharmacologically or genetically. Mechanistically, it was found that IDO1-expressing dendritic cells (DCs) accumulated in the dorsal root leptomeninges and led to an increase in kynurenine levels in the spinal cord. In the spinal cord, kynurenine was metabolized by kynurenine-3-monooxygenase-expressing astrocytes into the pronociceptive metabolite 3-hydroxykynurenine. Ultimately, 3-hydroxyanthranilate 3,4-dioxygenase-derived quinolinic acid formed in the final step of the canonical KYNPATH was also involved in neuropathic pain development through the activation of the glutamatergic N-methyl-D-aspartate receptor. In conclusion, these data revealed a role for DCs driving neuropathic pain development through elevation of the KYNPATH. This paradigm offers potential new targets for drug development against this type of chronic pain.

Post-traumatic stress disorder increases pain sensitivity by reducing descending noradrenergic and serotoninergic modulation.

Exposure to stress might influence pain sensitivity; however, little is known about whether post-traumatic stress disorder (PTSD)-like symptoms alter pain sensitivity and how it can happen. Male rats were exposed to the inescapable footshock paired with either social isolation or a control condition (not exposed to footshock but subjected to social isolation). After 7, 14, or 21 days, memory retention was evaluated. In the following three days, animals underwent the following tests: open-field, social interaction and formalin tests. Another group of animals were subjected to the object recognition test and to von Frey filaments. In other cohorts of animals, saline, fluoxetine, or desipramine were injected intrathecally and immunohistochemistry was performed to investigate whether PTSD-like symptoms alter the expression of c-Fos in serotonergic and noradrenergic neurons. Inescapable footshock induced the development of PTSD-like symptoms. Animals with PTSD-like symptoms showed an increase in the number of flinches in the formalin test and a reduction in mechanical threshold in the von Frey test at both retention intervals. The social interaction was negatively correlated with the nociceptive response in the formalin test. Fluoxetine or desipramine prevented the nociceptive response to chemical stimulus in the formalin test. In addition, in animals with PTSD-like symptoms, there was a reduction in c-Fos expression in serotonergic and noradrenergic neurons. Our results are important for the association of increased sensitivity to pain as one of the clinical manifestations that are present in the development of PTSD, and a possible treatment for increased pain sensitivity in male individuals with PTSD.

The PI3Kγ/AKT signaling pathway mediates peripheral antinociceptive action of dipyrone.

Dipyrone (DIP), also known as metamizole, is an over-the-counter analgesic used in Europe and Latin America. Evidence suggesting that inflammatory pain attenuation by DIP is associated with a direct impact on peripheral primary nociceptive neurons through the stimulation of nitric oxide signaling pathway. However, the molecular mechanism by which DIP activates this pathway remains unknown. The PI3Kγ/AKT signaling cascade activation is one of the well-known molecular mechanisms that promote nitric oxide production in sensory neurons. Herein, we investigated the role of the PI3Kγ/AKT signaling cascade in the context of peripheral analgesic effect of DIP. DIP was administered into PGE2 pre-sensitized paws of rats and mechanical hyperalgesia was determined using electronic von Frey test after 1 h. Nonselective or selective pharmacological inhibitors of PI3Kγ and AKT were also administered in DIP-treated rats under paws sensitized with PGE2. Intraplantar injection of DIP attenuated PGE2-induced hyperalgesia in a dose-dependent manner. Treatment with nonselective (wortmannin or LY294002) or selective (AS605240) pharmacological inhibitors of PI3Kγ reduced the peripheral antihypernociceptive effect of DIP. Consistently, AKT selective inhibitor also reversed analgesic DIP effects. Corroborating these data, we found that DIP induced AKT phosphorylation in cultured dorsal root ganglion neurons, which was prevented in the presence of PI3Kγ selective inhibitor. Taken together, these findings provide evidence that peripheral analgesic effect of DIP is dependent on the activation of PI3Kγ/AKT signaling pathway.

Caspase-1 is involved in the genesis of inflammatory hypernociception by contributing to peripheral IL-1ß maturation.

BACKGROUND: Caspase-1 is a cysteine protease responsible for the processing and secretion of IL-1ß and IL-18, which are closely related to the induction of inflammation. However, limited evidence addresses the participation of caspase-1 in inflammatory pain. Here, we investigated the role of caspase-1 in inflammatory hypernociception (a decrease in the nociceptive threshold) using caspase-1 deficient mice (casp1-/-). RESULTS: Mechanical inflammatory hypernociception was evaluated using an electronic version of the von Frey test. The production of cytokines, PGE2 and neutrophil migration were evaluated by ELISA, radioimmunoassay and myeloperoxidase activity, respectively. The interleukin (IL)-1ß and cyclooxygenase (COX)-2 protein expression were evaluated by western blotting. The mechanical hypernociception induced by intraplantar injection of carrageenin, tumour necrosis factor (TNF)α and CXCL1/KC was reduced in casp1-/- mice compared with WT mice. However, the hypernociception induced by IL-1ß and PGE2 did not differ in WT and casp1-/- mice. Carrageenin-induced TNF-α and CXCL1/KC production and neutrophil recruitment in the paws of WT mice were not different from casp1-/- mice, while the maturation of IL-1ß was reduced in casp1-/- mice. Furthermore, carrageenin induced an increase in the expression of COX-2 and PGE2 production in the paw of WT mice, but was reduced in casp1-/- mice. CONCLUSION: These results suggest that caspase-1 plays a critical role in the cascade of events involved in the genesis of inflammatory hypernociception by promoting IL-1ß maturation. Because caspase-1 is involved in the induction of COX-2 expression and PGE2 production, our data support the assertion that caspase-1 is a key target to control inflammatory pain.

Non-Peptidergic Nociceptive Neurons Are Essential for Mechanical Inflammatory Hypersensitivity in Mice.

Small nerve fibers that bind the isolectin B4 (IB4+ C-fibers) are a subpopulation of primary afferent neurons that are involved in nociceptive sensory transduction and do not express the neuropeptides substance P and calcitonin-gene related peptide (CGRP). Several studies have attempted to elucidate the functional role of IB4+-nociceptors in different models of pain. However, a functional characterization of the non-peptidergic nociceptors in mediating mechanical inflammatory hypersensitivity in mice is still lacking. To this end, in the present study, the neurotoxin IB4-Saporin (IB4-Sap) was employed to ablate non-peptidergic C-fibers. Firstly, we showed that intrathecal (i.t.) administration of IB4-Sap in mice depleted non-peptidergic C-fibers, since it decreased the expression of purinoceptor 3 (P2X3) and transient receptor potential cation channel subfamily V member 1 (TRPV1) in the dorsal root ganglia (DRGs) as well as IB4 labelling in the spinal cord. Non-peptidergic C-fibers depletion did not alter the mechanical nociceptive threshold, but it inhibited the mechanical inflammatory hypersensitivity induced by glial cell-derived neurotrophic factor (GDNF), but not nerve growth factor (NGF). Depletion of non-peptidergic C-fibers abrogated mechanical inflammatory hypersensitivity induced by carrageenan. Finally, it was found that the inflammatory mediators PGE2 and epinephrine produced a mechanical inflammatory hypersensitivity that was also blocked by depletion of non-peptidergic C-fibers. These data suggest that IB4-positive nociceptive nerve fibers are not involved in normal mechanical nociception but are sensitised by inflammatory stimuli and play a crucial role in mediating mechanical inflammatory hypersensitivity.

The NOD2 signaling in peripheral macrophages contributes to neuropathic pain development.

Neuropathic pain is one of the most important types of chronic pain. It is caused by neuronal damage. Clinical and experimental studies suggest a critical role for neuroimmune interactions in the development of neuropathic pain. In this article, we have shown that the cytoplasmic receptor Nod-like receptor-2, NOD2, and its adaptor-signaling molecule RIPK2 participate in the development of neuropathic pain after peripheral nerve injury (spared nerve injury model). The activation of NOD2 signaling in peripheral macrophage mediates the development of neuropathic pain through the production of pronociceptive cytokines (tumor necrosis factor and IL-1ß). This study found that peripheral nerve injury promoted a systemic increase in the NOD2 ligand. These results highlight a previously undetermined role for NOD2 signaling in the development of neuropathic pain, suggesting a new potential target for preventing neuropathic pain.

15d-prostaglandin J2 inhibits inflammatory hypernociception: involvement of peripheral opioid receptor.

The 15-deoxy-Delta(12,14)-prostaglandin J(2) (15d-PGJ(2)) is an endogenous ligand of peroxisome proliferator-activated receptors gamma (PPAR-gamma) and is now recognized as a potent anti-inflammatory mediator. However, information regarding the influence of 15d-PGJ(2) on inflammatory pain is still unknown. In this study, we evaluated the effect of 15d-PGJ(2) upon inflammatory hypernociception and the mechanisms involved in this effect. We observed that intraplantar administration of 15d-PGJ(2) (30-300 ng/paw) inhibits the mechanical hypernociception induced by both carrageenan (100 mug/paw) and the directly acting hypernociceptive mediator, prostaglandin E(2) (PGE(2)). Moreover, 15d-PGJ(2) [100 ng/temporomandibular joint (TMJ)] inhibits formalin-induced TMJ hypernociception. On the other hand, the direct administration of 15d-PGJ(2) into the dorsal root ganglion was ineffective in blocking PGE(2)-induced hypernociception. In addition, the 15d-PGJ(2) antinociceptive effect was enhanced by the increase of macrophage population in paw tissue due to local injection of thioglycollate, suggesting the involvement of these cells on the 15d-PGJ(2)-antinociceptive effect. Moreover, the antinociceptive effect of 15d-PGJ(2) was also blocked by naloxone and by the PPAR-gamma antagonist 2-chloro-5-nitro-N-phenylbenzamide (GW9662), suggesting the involvement of peripheral opioids and PPAR-gamma receptor in the process. Similar to opioids, the 15d-PGJ(2) antinociceptive action depends on the nitric oxide/cGMP/protein kinase G (PKG)/K(ATP)(+) channel pathway because it was prevented by the pretreatment with the inhibitors of nitric-oxide synthase (N(G)-monomethyl-l-arginine acetate), guanylate cyclase]1H-(1,2,4)-oxadiazolo(4,2-alpha)quinoxalin-1-one[, PKG [indolo[2,3-a]pyrrolo[3,4-c]carbazole aglycone (KT5823)], or with the ATP-sensitive potassium channel blocker glibenclamide. Taken together, these results demonstrate for the first time that 15d-PGJ(2) inhibits inflammatory hypernociception via PPAR-gamma activation. This effect seems to be dependent on endogenous opioids and local macrophages.

Role of IL-18 in overt pain-like behaviour in mice.

There are evidences that targeting IL-18 might be beneficial to inhibit inflammatory symptoms, including hypernociception (decrease in nociceptive threshold). The mechanism of IL-18 mechanical hypernociception depends on endothelin in rats and mice. However, the role of IL-18 in overt pain-like behaviour remains undetermined. Therefore, we addressed the role of IL-18 in writhing response induced by intraperitoneal (i.p.) injection of phenyl-p-benzoquinone (PBQ) and acetic acid in mice. Firstly, it was detected that PBQ and acetic acid i.p. injection induced a dose-dependent number of writhes in Balb/c mice. Subsequently, it was observed that the PBQ - but not the acetic acid-induced writhes were diminished in IL-18 deficient ((-/-)) mice. Therefore, considering that IFN-gamma, endothelin and prostanoids mediate IL-18-induced mechanical hypernociception, we also investigated the role of these mediators in the same model of writhing response in which IL-18 participates. It was noticed that PBQ-induced writhes were diminished in IFN-gamma(-/-) mice and by the treatment with bosentan (mixed endothelin ETA/ETB receptor antagonist), BQ 123 (cyclo[DTrp-DAsp-Pro-DVal-Leu], selective endothelin ETA receptor antagonist), BQ 788 (N-cys-2,6 dimethylpiperidinocarbonyl-l-methylleucyl-d-1-methoxycarboyl-d-norleucine, selective endothelin ETB receptor antagonist) or indomethacin (cycloxigenase inhibitor). Thus, IL-18, IFN-gamma, endothelin acting on endothelin ETA and ETB receptors, and prostanoids mediate PBQ-induced writhing response in mice. To conclude, these results further advance the understanding of the physiopathology of overt pain-like behaviour, and suggest for the first time a role for IL-18 in writhing response in mice.