The emerging literature implicates a role for glia/cytokines in persistent pain. However, the mechanisms by which these non-neural elements contribute to CNS activity-dependent plasticity and pain are unclear. Using a trigeminal model of inflammatory hyperalgesia, here we provide evidence that demonstrates a mechanism by which glia interact with neurons, leading to activity-dependent plasticity and hyperalgesia. In response to masseter inflammation, there was an upregulation of glial fibrillary acidic proteins (GFAPs), a marker of astroglia, and interleukin-1␤ (IL-1␤), a prototype proinflammatory cytokine, in the region of the trigeminal nucleus specifically related to the processing of deep orofacial input. The activated astroglia exhibited hypertrophy and an increased level of connexin 43, an astroglial gap junction protein. The upregulated IL-1␤ was selectively localized to astrocytes but not to microglia and neurons. Local anesthesia of the masseter nerve prevented the increase in GFAP and IL-1␤ after inflammation, and substance P, a prototype neurotransmitter of primary afferents, induced similar increases in GFAP and IL-1␤, which was blocked by a nitric oxide synthase inhibitor

Chronic pain conditions are difficult to treat and are major health problems. Bone marrow stromal cells (BMSCs) have generated considerable interest as a candidate for cell-based therapy. BMSCs are readily accessible and are easy to isolate and expand ex vivo. Clinical studies show that direct injection of BMSCs does not produce unwanted side effects and is well tolerated and safe. Here, we show that a single systemic (intravenous) or local injection (into the lesion site) of rat primary BMSCs reversed pain hypersensitivity in rats after injury and that the effect lasted until the conclusion of the study at 22 weeks. The pain hypersensitivity was rekindled by naloxone hydrochloride, an opioid receptor antagonist that acts peripherally and centrally, when tested at 1-5 weeks after BMSC infusion. In contrast, naloxone methiodide, a peripherally acting opioid receptor antagonist, only rekindled hyperalgesia in the first 3 weeks of BMSC treatment. Focal downregulation of brainstem mu opioid receptors by RNA interference (RNAi) reversed the effect of BMSCs, when RNAi was introduced at 5-but not 1-week after BMSC transplantation. Thus, BMSCs produced longterm relief of pain and this effect involved activation of peripheral and central opioid receptors in distinct time domains. The findings prompt studies to elucidate the cellular mechanisms of the BMSC-induced pain relieving effect and translate these observations into clinical settings. STEM

SummaryThe cytotoxicity of Bordetella bronchiseptica to infected cells is known to be dependent on a B. bronchiseptica type III secretion system. Although the precise mechanism of the type III secretion system is unknown, BopN, BopD and Bsp22 have been identified as type III secreted proteins. In order to identify other proteins secreted via the type III secretion machinery in Bordetella , a type III mutant was generated, and its secretion profile was compared with that of the wild-type strain. The results showed that the wild-type strain, but not the type III mutant, secreted a 40-kDa protein into the culture supernatant. This protein was identified as BopB by the analysis of its N-terminal amino acid sequence. Severe cytotoxicity such as necrosis was induced in L2 cells by infection with the wild-type B. bronchiseptica . In contrast, this effect was not observed by the BopB mutant infection. The haemolytic activity of the BopB mutant was greatly impaired compared with that of the wild-type strain. The results of a digitonin assay strongly suggested that BopB was translocated into HeLa cells infected with the wild-type strain. Taken together, our results demonstrate that Bordetella secretes BopB via a type III secretion system during infection. BopB may play a role in the formation of pores in the host plasma membrane which serve as a conduit for the translocation of effector proteins into host cells.

The inflammatory response is one of several host alert mechanisms that recruit neutrophils from the circulation to the area of infection. We demonstrate that Bordetella, a bacterial pathogen, exploits an antiinflammatory cytokine, interleukin-10 (IL-10), to evade the host immune system. We identified a Bordetella effector, BopN, that is translocated into the host cell via the type III secretion system, where it induces enhanced production of IL-10. Interestingly, the BopN effector translocates itself into the nucleus and is involved in the down-regulation of mitogen-activated protein kinases. Using pharmacological blockade, we demonstrated that BopN-induced IL-10 production is mediated, at least in part, by its ability to block the extracellular signal-regulated kinase pathway. We also showed that BopN blocks nuclear translocation of nuclear factor κB p65 (NF-κBp65) but, in contrast, promotes nuclear translocation of NF-κBp50. A BopN-deficient strain was unable to induce IL-10 production in mice, resulting in the elimination of bacteria via neutrophil infiltration into the pulmonary alveoli. Furthermore, IL-10–deficient mice effectively eliminated wild-type as well as BopN mutant bacteria. Thus, Bordetella exploits BopN as a stealth strategy to shut off the host inflammatory reaction. These results explain the ability of Bordetella species to avoid induction of the inflammatory response.

Bordetella pertussis and B. parapertussis are the etiological agents of pertussis, yet the former has a higher incidence and is the cause of a more severe disease, in part due to pertussis toxin. To identify other factors contributing to the different pathogenicity of the two species, we analyzed the capacity of structurally different lipooligosaccharide (LOS) from B. pertussis and LPS from B. parapertussis to influence immune functions regulated by dendritic cells. Either B. pertussis LOS and B. parapertussis LPS triggered TLR4 signaling and induced phenotypic maturation and IL-10, IL-12p40, IL-23, IL-6, and IL-1β production in human monocyte-derived dendritic cells (MDDC). B. parapertussis LPS was a stronger inducer of all these activities as compared with B. pertussis LOS, with the notable exception of IL-1β, which was equally produced. Only B. parapertussis LPS was able to induce IL-27 expression. In addition, although MDDC activation induced by B. parapertussis LPS was greatly dependent on soluble CD14, B. pertussis LOS activity was CD14-independent. The analysis of the intracellular pathways showed that B. parapertussis LPS and B. pertussis LOS equally induced IκBα and p38 MAPK phosphorylation, but B. pertussis LOS triggered ERK1/2 phosphorylation more rapidly and at higher levels than B. parapertussis LPS. Furthermore, B. pertussis LOS was unable to induce MyD88-independent gene induction, which was instead activated by B. parapertussis LPS, witnessed by STAT1 phosphorylation and induction of the IFN-dependent genes, IFN regulatory factor-1 and IFN-inducible protein-10. These differences resulted in a divergent regulation of Th cell responses, B. pertussis LOS MDDC driving a predominant Th17 polarization. Overall, the data observed reflect the different structure of the two LPS and the higher Th17 response induced by B. pertussis LOS may contribute to the severity of pertussis in humans.

The mandibular condylar cartilage plays an important role as a stress absorber during function. However, relatively little information is available on its dynamic properties under compression. We hypothesized that these properties are region-specific and depend on loading frequency. To characterize the viscoelastic properties of the condylar cartilage, we performed dynamic indentation tests over a wide range of loading frequencies. Ten porcine mandibular condyles were used; the articular surface was divided into 4 regions, anteromedial, anterolateral, posteromedial, and posterolateral. The dynamic complex, storage, and loss moduli increased with frequency, and these values were the highest in the anteromedial region. Loss tangent decreased with frequency from 0.68 to 0.17, but a regional difference was not found. The present results suggest that the dynamic compressive modulus is region-specific and is dependent on the loading frequency, which might have important implications for the transmission of load in the temporomandibular joint.

Bordetella parapertussis is one of the bacteria that causes whooping cough. However, little attention has been paid to this bacterium because it causes a milder illness than Bordetella pertussis and the rate of detection is low, even though research suggests that pertussis vaccines have limited efficacy against B. parapertussis infection. However, recent studies have revealed high rates of detection in patients with whooping cough in some field studies. In this review, the relevant studies of B. parapertussis are summarized and it is demonstrated that it is now necessary to pay greater attention to infections by this bacterium.

The protective immunity induced by infection withWhooping cough caused by Bordetella pertussis is a serious disease in children. Commercial pertussis vaccines, which consist of killed B. pertussis cells or derived antigens, are very effective and have reduced the incidence of whooping cough very considerably. However, in addition to B. pertussis, Bordetella parapertussis also causes symptoms typical of whooping cough (22). The illness caused by B. parapertussis is sometimes as severe as that caused by B. pertussis (10). Outbreaks of infection by B. parapertussis have been reported in several countries (8,11,18). B. parapertussis is closely related to B. pertussis in terms of virulence and attachment factors, such as filamentous hemagglutinin (FHA), adenylate cyclase toxin, heat-labile toxin, and pertactin (PRN) (29). However, several reports suggest that pertussis vaccine has no or limited ability to protect against B. parapertussis (9, 13, 15, 27, 32). Stehr et al. reported that the efficacy of the acellular pertussis component diphtheria-tetanus-pertussis (DTP) vaccine and the whole-cell pertussis component DTP vaccine in children was 31% and Ϫ6%, respectively (27). Khelef et al. suggested that immunization with antigens derived from B. pertussis induce no protection against B. parapertussis in mice (13). These reports suggested that reciprocal protective immunity between the two species might not be induced. However, in these studies, subcutaneous or peritoneal injections were commonly used as methods of immunization. Mills et al. suggested that there might be a difference, in terms of the profiles of the protective immune response against B. pertussis, between the response after immunization by injection with vaccines and the response during convalescence after infection with B. pertussis (20). We postulated that immunization by natural infection of the two species might clarify the relationship between protection against B. pertussis and protection against B. parapertussis. To test our hypothesis, we infected mice by exposing them to an aerosol of B. pertussis or B. parapertussis. After mice had recovered, convalescent mice were investigated for protective responses against the two species of Bordetella, for levels of antigen-specific antibodies, and for splenocyte proliferation and cytokine secretion responses in vitro after stimulation by antigens. MATERIALS AND METHODSMice. Specific-pathogen-free female dd-Y mice were obtained from Japan SLC (Hamamatsu, Japan). All mice were 3.5 weeks old at the start of experiments.Bacterial strains and culture conditions. The phase I strain of B. pertussis strain 18-323 and B. parapertussis strain 23054 were used in this study. Cells were grown on Bordet-Gengou (BG) agar supplemented with 20% (vol/vol) defibrinated horse blood at 37°C.Bacterial antigens. Killed whole-cell B. pertussis or B. parapertussis antigens were prepared as described below. B. pertussis or B. parapertussis was cultured on BG plates for 30 h at 37°C. Cells were harvested in phosphate-buffered sali...