CNS-infiltrating CD4+ T lymphocytes contribute to murine spinal nerve transection-induced neuropathic pain

We previously reported leukocytic infiltration into the lumbar spinal cord in a rodent spinal nerve L5 transection (L5Tx) neuropathic pain model. Here, we further investigated the role of infiltrating T lymphocytes in the etiology of persistent pain following L5Tx. T lymphocyte-deficient nude mice showed no evident mechanical hypersensitivity after day 3 of L5Tx compared to wild-type BALB/c mice. Through FACS analysis, we determined that significant leukocytic infiltration (CD45(hi)) into the lumbar spinal cord peaked at day 7 post L5Tx. These infiltrating leukocytes contained predominantly CD4(+) but not CD8(+) T lymphocytes. B lymphocytes, natural killer cells and macrophages were not detected at day 7 post L5Tx. No differences in the activation of peripheral CD4(+) T lymphocytes were detected in either the spleen or lumbar lymph nodes between L5Tx and sham surgery groups. Further, CD4 KO mice displayed significantly decreased mechanical hypersensitivity after day 7 of L5Tx, and adoptive transfer of CD4(+) leukocytes reversed this effect. Decreased immunoreactivity of glial fibrillary acidic protein observed in CD4 KO mice post L5Tx indicated possible T lymphocyte-glial interactions. These results strongly support a contributing role of spinal cord-infiltrating CD4(+) T lymphocytes versus peripheral CD4(+) T lymphocytes in the maintenance of nerve injury-induced neuropathic pain.     

The contributing role of CD14 in toll-like receptor 4 dependent neuropathic pain

We have previously demonstrated that CNS toll-like receptor 4 (TLR4) plays a key role in the development of behavioral hypersensitivity in a rodent model of neuropathic pain, spinal nerve L5 transection (L5Tx). TLR4 is a well-known receptor for lipopolysaccharide (LPS) in innate immune responses. In the current study, we further investigated the role of CD14, an accessory molecule in the LPS-TLR4 signaling pathway, in the development of L5Tx-induced neuropathic pain. CD14 knockout (KO) mice displayed significantly decreased behavioral sensitivity (mechanical allodynia and thermal hyperalgesia) as early as day 1 post-L5Tx, indicating a nociceptive role of CD14. By flow cytometric analyses, we observed significantly elevated microglial surface CD14 expression in the ipsilateral lumbar spinal cord 3 days post-L5Tx, as well as remarkable increases in microglial size (via forward scatter (FSC)) and granularity (via side scatter (SSC)). Further, intrathecal injection of soluble CD14 induced significantly greater mechanical hypersensitivity in wild type (C3H/HeN) mice compared with TLR4-deficient (C3H/HeJ) mice. Together, these data demonstrate that CD14 plays a contributing role in TLR4-dependent nerve injury-induced neuropathic pain.     

Absence of Notch1 in murine myeloid cells attenuates the development of experimental autoimmune encephalomyelitis by affecting Th1 and Th17 priming

Inhibition of Notch signalling in T cells attenuates the development of experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis. Growing evidence indicates that myeloid cells are also key players in autoimmune processes. Thus, the present study evaluates the role of the Notch1 receptor in myeloid cells on the progression of myelin oligodendrocyte glycoprotein (MOG)_35‐55‐induced EAE, using mice with a myeloid‐specific deletion of the Notch1 gene (MyeNotch1KO). We found that EAE progression was less severe in the absence of Notch1 in myeloid cells. Thus, histopathological analysis revealed reduced pathology in the spinal cord of MyeNotch1KO mice, with decreased microglia/astrocyte activation, demyelination and infiltration of CD4⁺ T cells. Moreover, these mice showed lower Th1 and Th17 cell infiltration and expression of IFN‐γ and IL‐17 mRNA in the spinal cord. Accordingly, splenocytes from MyeNotch1KO mice reactivated in vitro presented reduced Th1 and Th17 activation, and lower expression of IL‐12, IL‐23, TNF‐α, IL‐6, and CD86. Moreover, reactivated wild‐type splenocytes showed increased Notch1 expression, arguing for a specific involvement of this receptor in autoimmune T cell activation in secondary lymphoid tissues. In summary, our results reveal a key role of the Notch1 receptor in myeloid cells for the initiation and progression of EAE.     

Mouse strains that lack spinal dynorphin upregulation after peripheral nerve injury do not develop neuropathic pain

Several experimental models of peripheral neuropathy show that a significant upregulation of spinal dynorphin A and its precursor peptide, prodynorphin, is a common consequence of nerve injury. A genetically modified mouse strain lacking prodynorphin does not exhibit sustained neuropathic pain after nerve injury, supporting a pronociceptive role of elevated levels of spinal dynorphin. A null mutation of the gamma isoform of protein kinase C (PKCgamma KO [knockout]), as well as an inbred mouse strain, 129S6, also does not manifest behavioral signs of neuropathic pain following peripheral nerve injury. The objective of this study was to extend our observations to these genetic models to test the hypothesis that elevated levels of spinal dynorphin are essential for the maintenance of abnormal pain. In PKCgamma wild-type mice and the outbred mouse strain ICR, ligation of the L5 and L6 spinal nerves (SNL) elicited both tactile hypersensitivity and thermal hyperalgesia. Both strains showed a significant elevation in dynorphin in the lumbar spinal dorsal horn following SNL. Spinal administration of an anti-dynorphin A antiserum blocked the thermal and tactile hypersensitivity in both strains of mice. However, the PKCgamma KO mice and the 129S6 mice (which express PKCgamma) did not show abnormal pain after SNL; neither strain showed elevated levels of spinal dynorphin. The multiple phenotypic deficits in PKCgamma KO mice confound the interpretation of the proposed role of PKCgamma-expressing spinal neurons in neuropathic pain states. Additionally, the data show that the regulation of spinal dynorphin expression is a common critical feature of expression of neuropathic pain.     

Activation of microglia and p38 mitogen-activated protein kinase in the dorsal column nucleus contributes to tactile allodynia following peripheral nerve injury

The activation of glial cells in the CNS has been suggested to be involved in abnormal pain sensation after peripheral nerve injury. Previous studies demonstrated phosphorylation of p38 mitogen-activated protein kinase (MAPK) in spinal cord glial cells after peripheral nerve injury, and such phosphorylation has been suggested to be involved in the development of neuropathic pain. The aim of this study was to examine the dorsal column nuclei for phosphorylation of p38 MAPK following peripheral nerve injury and to explore a possibility of its contribution to neuropathic pain. Immunohistochemical labeling for phosphorylated p38 (p-p38) MAPK was performed in histological sections of the rat spinal cord and medulla oblongata after the fifth lumbar (L5) spinal nerve ligation (SNL). The number of p-p38 MAPK-immunoreactive (IR) cells was significantly increased in the L5 dorsal horn and the gracile nucleus ipsilateral to the injury at days 3-21 after SNL. Double immunofluorescence labeling with cell-specific markers revealed that p-p38 MAPK-IR cells co-expressed OX-42, suggesting their microglial identity. Increased immunofluorescence labeling for OX-42 indicated that microglial cells were activated by SNL in the L5 dorsal horn and the gracile nucleus ipsilateral to the injury. Continuous infusion of a p38 MAPK inhibitor into the cisterna magna for 14 days beginning on the day of SNL suppressed the development of tactile allodynia, but not thermal hyperalgesia induced by nerve injury. These results demonstrate that SNL activates p38 MAPK pathway in microglia in the gracile nucleus as well as in the spinal cord dorsal horn. Activation of p38 MAPK in medullary microglia may contribute to the pathogenesis of neuropathic pain.     

A transcranial magnetic stimulation study of corticospinal excitability during the observation of meaningless, goal-directed, and social behaviour

Peripheral nerve injury causes a progressive series of morphological changes in spinal microglia, and extracellular ATP stimulates proliferation of microglia and may be involved in neuropathic pain. We defined the precise expression of P2X7 in the spinal cord following peripheral nerve injury. We found that both P2X7 mRNA and protein increased in the spinal cord, with a peak at 7 d after injury. Double labeling studies revealed that cells expressing increased P2X7 mRNA and protein after nerve injury were predominantly microglia in dorsal horn. Pharmacological blockades by intrathecal administration of a P2X7 antagonist (A 438079 hydrochloride) suppressed the development of mechanical hypersensitivity. We present distinct evidence that increases in the number of P2X7 receptors in spinal microglia may play an important role in neuropathic pain.     

Role of astrocytic S100beta in behavioral hypersensitivity in rodent models of neuropathic pain

S100beta is a calcium-binding peptide produced mainly by astrocytes that exerts paracrine and autocrine effects on neurons and glia. We have previously shown that S100beta is markedly elevated at the mRNA level in the spinal cord following peripheral inflammation, intraplantar administration of complete Freund's adjuvant in the rat. The purpose of the present study was to further investigate the role of astrocytic S100beta in mediating behavioral hypersensitivity in rodent models of persistent pain. First, we assessed the lumbar spinal cord expression of S100beta at the mRNA and protein level using real-time RT-PCR, Western blot and immunohistochemistry analysis following L5 spinal nerve transection in rats, a rodent model of neuropathic pain. Second, we assessed behavioral hypersensitivity (mechanical allodynia) in wild type and genetically modified mice lacking or overexpressing S100beta following L5 spinal nerve transection. Third, we assessed the expression level of S100beta protein in the CD1 wild type mice after nerve injury. We report that lumbar spinal S100beta mRNA steadily increased from days 4-28 after nerve injury. S100beta protein in the lumbar spinal cord was significantly increased in both rats and mice at day 14 following nerve injury as compared with sham control groups. S100beta genetically deficient mice displayed significantly increased tactile thresholds (reduced response to non-noxious stimuli) after nerve injury as compared with the wild type group. S100beta overexpressing mice displayed significantly decreased tactile threshold responses (enhanced response to non-noxious stimuli). Together, these results from both series of experiments using a peripheral nerve injury model in two different species implicate the involvement of glial-derived S100beta in the pathophysiology of neuropathic pain.     

Tolerogenic dendritic cells produced by lentiviral‐mediated CD40‐ and interleukin‐23p19‐specific shRNA can ameliorate experimental autoimmune encephalomyelitis by suppressing T helper type 17 cells

Down‐regulation of soluble or membrane‐bound co‐stimulatory molecules by RNAi in dendritic cells can prevent the activation of immune responses. Therefore, this study was designed to evaluate the therapeutic efficacy of bone marrow‐derived DCs (BMDCs) transduced with lentiviral vectors to permanently expressed shRNA specific for CD40 (CD40LV‐DCs) and/or p19 subunit of interleukin (IL)‐23 (p19LV‐DCs) mRNAs in experimental autoimmune encephalomyelitis (EAE). In‐vitro studies showed that double‐transduced BMDCs (CD40⁺p19LV‐DCs) resemble tolerogenic DCs due to profound down‐regulation of CD40, lower expression of proinflammatory cytokines (IL‐6 and IL‐12), increased IL‐10 production and stronger stimulation of myelin oligodendrocyte glycoprotein (MOG)_35–55‐specific T cells for production of IL‐10 compared with CD40LV‐DCs, p19LV‐DCs and BMDCs transduced with control lentiviral vector (CoLV‐DCs). Moreover, injection of transduced CD40⁺p19LV‐ BMDCs in EAE mice resulted in more reduction in clinical score, significant reduction in IL‐17 or increased production of IL‐10 by mononuclear cells derived from the lymph nodes or spinal cord compared with CoLV‐DCs‐treated EAE mice. In conclusion, simultaneous knock‐down of CD40 and IL‐23 production by BMDCs may represent a promising therapeutic tool for the treatment of IL‐17‐dependent autoimmune diseases, including multiple sclerosis.     

Cytotoxic CD8+ T cell ablation enhances the capacity of regulatory T cells to delay viral elimination in Theiler's murine encephalomyelitis

Theiler's murine encephalomyelitis (TME) of susceptible mouse strains is a commonly used infectious animal model for multiple sclerosis. The study aim was to test the hypothesis whether cytotoxic T cell responses account for the limited impact of regulatory T cells on antiviral immunity in TME virus‐induced demyelinating disease (TMEV‐IDD) resistant C57BL/6 mice. TME virus‐infected C57BL/6 mice were treated with (i) interleukin‐2/‐anti‐interleukin‐2‐antibody‐complexes to expand regulatory T cells (“Treg‐expansion”), (ii) anti‐CD8‐antibodies to deplete cytotoxic T cells (“CD8‐depletion”) or (iii) with a combination of Treg‐expansion and CD8‐depletion (“combined treatment”) prior to infection. Results showed that “combined treatment”, but neither sole “Treg‐expansion” nor “CD8‐depletion,” leads to sustained hippocampal infection and virus spread to the spinal cord in C57BL/6 mice. Prolonged infection reduces myelin basic protein expression in the spinal cord together with increased accumulation of β‐amyloid precursor protein in axons, characteristic of myelin loss and axonal damage, respectively. Chronic spinal cord infection upon “combined treatment” was also associated with increased T and B cell recruitment, accumulation of CD107b⁺ microglia/macrophages and enhanced mRNA expression of interleukin (IL)‐1α, IL‐10 and tumor necrosis factor α. In conclusion, data revealed that the suppressive capacity of Treg on viral elimination is efficiently boosted by CD8‐depletion, which renders C57BL/6 mice susceptible to develop chronic neuroinfection and TMEV‐IDD.     

Dynamic Changes of Microglia/Macrophage M1 and M2 Polarization in Theiler's Murine Encephalomyelitis

Microglia and macrophages play a central role for demyelination in Theiler's murine encephalomyelitis (TME) virus infection, a commonly used infectious model for chronic‐progressive multiple sclerosis. In order to determine the dynamic changes of microglia/macrophage polarization in TME, the spinal cord of Swiss Jim Lambert (SJL) mice was investigated by gene expression profiling and immunofluorescence. Virus persistence and demyelinating leukomyelitis were confirmed by immunohistochemistry and histology. Electron microscopy revealed continuous myelin loss together with abortive myelin repair during the late chronic infection phase indicative of incomplete remyelination. A total of 59 genes out of 151 M1‐ and M2‐related genes were differentially expressed in TME virus‐infected mice over the study period. The onset of virus‐induced demyelination was associated with a dominating M1 polarization, while mounting M2 polarization of macrophages/microglia together with sustained prominent M1‐related gene expression was present during the chronic‐progressive phase. Molecular results were confirmed by immunofluorescence, showing an increased spinal cord accumulation of CD16/32⁺ M1‐, arginase‐1⁺ M2‐ and Ym1⁺ M2‐type cells associated with progressive demyelination. The present study provides a comprehensive database of M1‐/M2‐related gene expression involved in the initiation and progression of demyelination supporting the hypothesis that perpetuating interaction between virus and macrophages/microglia induces a vicious circle with persistent inflammation and impaired myelin repair in TME.     

MicroRNA‐155 Deficiency Suppresses Th17 Cell Differentiation and Improves Locomotor Recovery after Spinal Cord Injury

Spinal cord injury (SCI) is considered to be primarily associated with loss of motor function and leads to activate diverse cellular mechanisms in the central nervous system to attempt to repair the damaged spinal cord tissue. Mir‐155 has been reported to be involved in both innate and adaptive immune responses. But the role of Mir‐155 in spinal cord injury is still unknown. In our current study, Mir‐155 deficiency displays increased myelin sparring and enhanced SC repair process. The number of T cells, B cells and neutrophils are all significantly lower in Mir‐155^?/? group than that in WT group after SCI. IL‐17A‐producing cells and the expression of IL‐17A are markedly lower in Mir‐155^?/? mice than that in WT mice. We also found higher production of IL‐17 by WT CD4⁺ T cells than Mir‐155^?/? CD4⁺ T cells in vitro. In our further DC‐T cell coculture system, Mir‐155 deficiency in DCs results in significantly less IL‐17 production from T cells. Furthermore, the inhibited Th17 differentiation induced by Mir‐155 deficiency is partly dependent on increased expression of SOCS1. In conclusion, our present work provides evidence to support the concept that Mir‐155 deficiency suppresses Th17 cell differentiation and improves locomotor recovery after SCI.     

IL‐12‐mediated STAT4 signaling and TCR signal strength cooperate in the induction of CD40L in human and mouse CD8+ T cells

CD40L is one of the key molecules bridging the activation of specific T cells and the maturation of professional and nonprofessional antigen‐presenting cells including B cells. CD4⁺ T cells have been regarded as the major T‐cell subset that expresses CD40L upon cognate activation; however, we demonstrate here that a putative CD8⁺ helper T‐cell subset expressing CD40L is induced in human and murine CD8⁺ T cells in vitro and in mice immunized with antigen‐pulsed dendritic cells. IL‐12 and STAT4‐mediated signaling was the major instructive cytokine signal boosting the ability of CD8⁺ T cells to express CD40L both in vitro and in vivo. Additionally, TCR signaling strength modulated CD40L expression in CD8⁺ T cells after primary differentiation in vitro as well as in vivo. The induction of CD40L in CD8⁺ T cells regulated by IL‐12 and TCR signaling may enable CD8⁺ T cells to respond autonomously of CD4⁺ T cells. Thus, we propose that under proinflammatory conditions, a self‐sustaining positive feedback loop could facilitate the efficient priming of T cells stimulated by high affinity peptide displaying APCs.     

1,25‐dihydroxyvitamin D3‐induced dendritic cells suppress experimental autoimmune encephalomyelitis by increasing proportions of the regulatory lymphocytes and reducing T helper type 1 and type 17 cells

Dendritic cells (DCs), a bridge for innate and adaptive immune responses, play a key role in the development of multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE), an animal model for MS. Administration of tolerogenic DCs has been used as an immunotherapy in autoimmune diseases. Deficiency of vitamin D is an environmental risk factor of MS. In this study, we induced tolerogenic DCs by 1,25‐dihydroxyvitamin D₃ and transferred the tolerogenic DCs (VD₃‐DCs) into EAE mice by adoptive transfer. We found that VD₃‐DCs inhibited the infiltrations of T helper type 1 (Th1) and Th17 cells into spinal cord and increased the proportions of regulatory T cells (CD4⁺ CD25⁺ Foxp3⁺), CD4⁺ IL‐10⁺ T cells and regulatory B cells (CD19⁺ CD5⁺ CD1d⁺) in peripheral immune organs, which resulted in attenuated EAE. However, the proportions of T helper type 1 (Th1) and Th17 cells in spleen and lymph nodes and the levels of pro‐inflammatory cytokines and IgG in serum also increased after transfer of VD₃‐DCs. We conclude that transfer of VD₃‐DCs suppressed EAE by increasing proportions of regulatory T cells, CD4⁺ IL‐10⁺ T cells and regulatory B cells in spleen and reducing infiltration of Th1 and Th17 cells into spinal cord, which suggests a possible immunotherapy method using VD₃‐DCs in MS.     

Remote astrocytic and microglial activation modulates neuronal hyperexcitability and below-level neuropathic pain after spinal injury in rat

In this study, we evaluated whether astrocytic and microglial activation mediates below-level neuropathic pain following spinal cord injury. Male Sprague–Dawley (225–250 g) rats were given low thoracic (T13) spinal transverse hemisection and behavioral, electrophysiological and immunohistochemical methods were used to examine the development and maintenance of below-level neuropathic pain. On postoperation day 28, both hind limbs showed significantly decreased paw withdrawal thresholds and thermal latencies as well as hyperexcitability of lumbar (L4-5) spinal wide dynamic range (WDR) neurons on both sides of spinal dorsal horn compared to sham controls (* P<0.05). Intrathecal treatment with propentofylline (PPF, 10 mM) for 7 consecutive days immediately after spinal injury attenuated the development of mechanical allodynia and thermal hyperalgesia in both hind limbs in a dose-related reduction compared to vehicle treatments (* P<0.05). Intrathecal treatment with single injections of PPF at 28 days after spinal injury, attenuated the existing mechanical allodynia and thermal hyperalgesia in both hind limbs in a dose related reduction (* P<0.05). In electrophysiological studies, topical treatment of 10 mM PPF onto the spinal surface attenuated the neuronal hyperexcitability in response to mechanical stimuli. In immunohistochemical studies, astrocytes and microglia in rats with spinal hemisection showed significantly increased GFAP and OX-42 expression in both superficial and deep dorsal horns in the lumbar spinal dorsal horn compared to sham controls (* P<0.05) that was prevented in a dose-related manner by PPF. In conclusion, our present data support astrocytic and microglial activation that contributes to below-level central neuropathic pain following spinal cord injury.     

Suppressive and pro‐inflammatory roles for IL‐4 in the pathogenesis of experimental drug‐induced liver injury

The pathogenesis of immune‐mediated drug‐induced liver injury (DILI) following halogenated anesthetics, carbamazepine or alcohol has not been fully elucidated. Detecting cytochrome P450 2E1 (CYP2E1) IgG4 auto‐antibodies in anesthetic DILI patients suggests a role for IL‐4 in this hapten‐mediated process. We investigated IL‐4‐mediated mechanisms using our model of experimental DILI induced by immunizing BALB/c (WT) and IL‐4^?/? (KO) mice with S100 liver proteins covalently modified by a trifluoroacetyl chloride (TFA) hapten formed following halogenated anesthetic metabolism by CYP2E1. WT mice developed more hepatitis, TFA and S100 antibodies (p<0.01), as well as T‐cell proliferation to CYP2E1 and TFA (p<0.01) than KO mice. Additionally, WT CD4⁺ T cells adoptively transferred hepatitis to naïve Rag^?/? mice (p<0.01). Pro‐inflammatory cytokines were expectedly decreased in TFA hapten‐stimulated KO splenocyte supernatants (p<0.001); however, IL‐2 and IFN‐γ (p<0.05), as well as IL‐6 and IL‐10 (p<0.001) levels were elevated in CYP2E1‐stimulated KO splenocyte supernatants, suggesting dual IL‐4‐mediated pro‐inflammatory and regulatory responses. Anti‐IL‐10 administered to KO mice increased hepatitis, TFA and CYP2E1 antibodies in KO mice confirming a critical role for IL‐4. This is the first demonstration of dual roles for IL‐4 in the pathogenesis of immune‐mediated DILI by suppressing auto‐antigen‐induced regulatory responses while promoting hapten‐induced pro‐inflammatory responses.     

The development of atopic dermatitis is independent of Immunoglobulin E up-regulation in the K14-IL-4 SKH1 transgenic mouse model

Background We have successfully generated an IgE‐associated (extrinsic/allergic) mouse model of atopic dermatitis in K14‐IL‐4‐Tg/CByB6 mice. The newly described subset of non‐IgE‐associated (intrinsic/non‐allergic) atopic dermatitis in human patients raises the question on the role of IgE in the pathogenesis. Objective The aim of this study was to develop a non‐IgE‐associated atopic dermatitis model in K14‐IL‐4‐Tg/SKH1 mice. Methods K14‐IL‐4‐Tg/CByB6 mice were crossed with SKH1 mice to produce K14‐IL‐4‐Tg/SKH1 mice. Phenotypes of clinical and histological, cytokine expression in the skin lesions, and total serum IgE in K14‐IL‐4‐Tg/CByB6 and K14‐IL‐4‐Tg/SKH1 mice were compared. The CD40 and CD40L on T and B cells were also studied to differentiate their roles in IgE production. Results K14‐IL‐4‐Tg/SKH1mice had a normal total serum IgE level and manifested a chronic inflammatory skin phenotype identical to that of K14‐IL‐4‐Tg/CByB6 IgE‐mediated mice in clinical morphology, histology, infiltration of mononuclear cells/eosinophils/mast cells, mast cell degranulation, and up‐regulation of chronic lesional cytokine mRNA expression of IL‐1β, IL‐3, IL‐4, IL‐6, IL‐10, IL‐12, IL‐13, IFN‐γ, TNF‐α, and TNF‐β. We also found that the inability of CD4⁺ T cells of the K14‐IL‐4‐Tg/SKH1mice to up‐regulate CD40L expression upon stimulation might account for their inability to up‐regulate the IgE level. B cell abnormality was ruled out as CD19⁺ B cells of K14‐IL‐4‐Tg/SKH1 mice synthesized the same amount of IgE in vitro compared with K14‐IL‐4‐Tg/CByB6 mice in the presence of IL‐4 and soluble CD40L. Our studies further suggested that the defect of early growth response‐1 in T cells might be responsible for the impaired CD40L up‐regulation in K14‐IL‐4‐Tg/SKH1 mice. Conclusion K14‐IL‐4‐Tg/SKH1 mice developed skin inflammation that resembled human intrinsic atopic dermatitis. Therefore, this model may be suitable to study the pathogenesis of intrinsic atopic dermatitis.     

Haptoglobin deficiency facilitates the development of autoimmune inflammation

Haptoglobin (HP) is an acute phase protein synthesized by liver cells in response to IL‐6. HP has been demonstrated to modulate the immune response and to have anti‐inflammatory activities. To analyze HP's effect on autoimmune inflammation, we here studied the course of EAE induced by immunization of Hp knockout (Hp^?/?) and syngeneic WT mice with myelin oligodendrocyte glycoprotein peptide (MOG_35–55). Hp^?/?mice suffered from a more severe disease that was associated with increased expression of IL‐17A, IL‐6, and IFN‐γ mRNA in the CNS and with a denser cellular infiltrate in the spinal cord. During the recovery phase, a significantly higher number of myeloid DC, CD8⁺ cells, IL‐17⁺ CD4⁺ and IFN‐γ⁺ CD4⁺ cells persisted in the CNS of Hp^?/? mice. Absence of HP affected the priming and differentiation of T cells after MOG_35–55 immunization, as levels of Th2 cytokines produced in response to MOG stimulation by Hp^?/? T cells were reduced. These results suggest that HP plays a modulatory and protective role on autoimmune inflammation of the CNS.     

Purinoceptors in microglia and neuropathic pain

Emerging evidence indicates that microglia play a critical role in the pathogenesis of neuropathic pain, a debilitating chronic pain condition that can occur after peripheral nerve damage caused by disease, infection, or physical injury. Microglia are immunocompetent cells of the central nervous system and express various ionotropic P2X and metabotropic P2Y purinoceptors. After injury to a peripheral nerve, microglia in the spinal cord become activated and upregulate expression of the P2X4 receptor. Recent findings suggest that activation of P2X4 receptors evokes release of brain-derived neurotrophic factor from microglia and that this mediates microglia–neuron signaling leading to pain hypersensitivity. Thus, P2X4 receptors and the intracellular signaling mediators in microglia are promising therapeutic targets for the development of novel pharmacological agents in the management of neuropathic pain.