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This study aimed to determine whether a trunk orthosis with joints providing resistive force (TORF) modifies sagittal malalignment during level walking in patients with lumbar spinal stenosis (LSS). Fifteen patients, 6 months after undergoing surgery for LSS, performed level walking at a self-selected speed while wearing a TORF. Dynamic sagittal alignment, including sagittal vertical axis, lumbar lordosis, and pelvic tilt, and spatiotemporal data as well as lower limb kinematic and kinetic data were recorded using a three-dimensional motion analysis system and six force plates. Statistical analysis was performed to compare these data with and without the TORF, respectively. Compared to the condition without the TORF, the use of the TORF significantly decreased positive sagittal vertical axis (p < 0.05) and increased the lumbar lordosis and pelvic tilt (p < 0.05). Peak hip flexion angle and extension moment during loading response (LR) significantly increased (p < 0.05), and peak hip extension angle and flexion moment during PS statistically decreased (p < 0.05). There was no difference in spatiotemporal data between the two conditions. Our findings suggest that TORF may modify the dynamic sagittal global alignment and lower limb kinematic and kinetics in postoperative LSS patients during level walking. [ABSTRACT FROM AUTHOR]

Background and Aim: This study aimed to compare the efficacy and safety of endoscopic ultrasound-guided gallbladder drainage and percutaneous transhepatic gallbladder drainage as a bridge to surgery in patients with acute cholecystitis unfit for urgent cholecystectomy. Methods: This retrospective study included 46 patients who underwent cholecystectomy following endoscopic ultrasound-guided gallbladder drainage (EUS-GBD) or percutaneous transhepatic gallbladder drainage (PTGBD) for acute cholecystitis in NTT Tokyo Medical Center. We surveyed 35 patients as the EUS-GBD group and 11 patients as the PTGBD group, and compared the rate of technical success of the cholecystectomy and periprocedural adverse events. A 7-F, 10-cm double pigtail plastic stent was used for ultrasound-guided gallbladder drainage. Results: The rate of technical success of cholecystectomy was 100% in both groups. Regarding postsurgical adverse events, no significant difference was noted between the two groups (EUS-GBD group, 11.4%, vs. PTGBD group, 9.0%; p = 0.472). Conclusions: EUS-GBD as a BTS seems to be an alternative for patients with AC because it can ensure lower adverse events. On the other hand, there are two major limitations in this study––the sample size is small and there is a risk of selection bias. [ABSTRACT FROM AUTHOR]

We aimed to investigate the risk factors of spontaneous osseous fusion (SOF) of the atlantoaxial joint after closed reduction under general anesthesia followed by halo fixation (remodeling therapy) for chronic atlantoaxial rotatory fixation, and to elucidate the recovery mechanism of the rotatory range of motion (ROM) after halo removal. Twelve patients who underwent remodeling therapy were retrospectively reviewed. Five patients with SOF were categorized as the fusion group and seven patients without SOF as the non-fusion group. Three dimensional CT was used to detect direct osseous contact (DOC) of facet joints before and during halo fixation, while dynamic CT at neutral and maximally rotated head positions was performed to measure rotatory ROM after halo removal. The duration from onset to initial visit was significantly longer (3.2 vs. 5.7 months, p = 0.04), incidence of DOC during halo fixation was higher (0/7 [0%] vs. 4/5 [80%], p = 0.004), and segmental rotatory ROM of Occiput/C1 (Oc/C1) at final follow-up was larger (9.8 vs. 20.1 degrees, p = 0.003) in the fusion group. Long duration from the onset to the initial visit might induce irreversible damage to the articular surface of the affected facet, which was confirmed as DOC during halo fixation and resulted in SOF. Long duration from the onset to the initial visit and DOC during halo fixation could be used to suggest the risk for SOF. Nonetheless, rotatory ROM of Oc/C1 increased to compensate for SOF. [ABSTRACT FROM AUTHOR]

In patients with dropped head syndrome (DHS), cervical malalignment is one of the risk factors for impaired horizontal gaze and restrictions to ambulation. The characteristics of gait in patients with DHS have not been clarified biomechanically from the viewpoint of dynamic alignment and lower limb kinematics. This study aimed to clarify kinematic and kinetic differences during level walking in patients with DHS compared to the healthy elderly. Twelve patients with DHS and healthy elderly individuals performed level walking at a self-selected speed. Spatiotemporal, kinematic, and kinetic data were recorded using a three-dimensional motion analysis system. Statistical analysis was performed to compare these data between the two groups, respectively. Compared with the healthy elderly, stride length and peak hip-joint extension angle in patients with DHS were significantly shorter and smaller. The thorax was also significantly tilted backwards. Peak ankle-joint plantar-flexion moment was significantly smaller despite larger dorsiflexion angle compared with the healthy elderly. The walking of DHS patients demonstrated kinematic and kinetic characteristics of the lower limb joints and alignment of the thorax and pelvis corresponding to their short stride and walking speed. [ABSTRACT FROM AUTHOR]

Dropped head syndrome (DHS) exhibits cervical deformity due to weakness of the cervical extensor group, and sarcopenia is characterized by progressive and systemic reduction in skeletal muscle mass. These clinical finding are associated with reduced activity of daily living, reduced quality of life, and increased risk of mortality. We collected and reviewed prospective registry data for 16 patients with idiopathic DHS continuously collected without dropping out and 32 healthy individuals who matched their gender and age. The prevalence of sarcopenia and body composition data were compared. There were no differences in the prevalence of sarcopenia, appendicular muscle mass, and leg muscle mass between DHS patients and the healthy elderly. Trunk muscle mass in DHS patients was significantly lower than that in healthy individuals. A significant correlation was found between appendicular muscle mass and trunk muscle mass in healthy subjects but not in DHS patients. Sarcopenia was not associated with the onset of idiopathic DHS. The prevalence of sarcopenia was not high in patients with idiopathic DHS due to the preservation of their appendicular skeletal muscle mass. Patients with DHS were characterized by a significant loss of trunk muscle mass that may be related to the disease but not aging. [ABSTRACT FROM AUTHOR]

The call for second generation malaria vaccines needs not only the identification of novel candidate antigens or adjuvants but also a better understanding of immune responses and the underlying protective processes. Plasmodium parasites have evolved a range of strategies to manipulate the host immune system to guarantee survival and establish parasitism. These immune evasion strategies hamper efforts to develop effective malaria vaccines. In the case of a malaria vaccine targeting the N-terminal domain of P. falciparum serine repeat antigen 5 (SE36), now in clinical trials, we observed reduced responsiveness (lowered immunogenicity) which may be attributed to immune tolerance/immune suppression. Here, immunogenicity data and insights into the immune responses to SE36 antigen from epidemiological studies and clinical trials are summarized. Documenting these observations is important to help identify gaps for SE36 continued development and engender hope that highly effective blood-stage/multi-stage vaccines can be achieved. [Display omitted] • Clinical trials show that SE36 antigen is a promising blood-stage vaccine candidate. • Repeated malaria infection lowers the humoral response in some SE36 vaccinees. • Concurrent parasitemia during vaccination resulted to reduced immunogenicity. • SE36 binds to host vitronectin akin to molecular camouflage. • Immune tolerance may explain the limited observed polymorphism of SE36. [ABSTRACT FROM AUTHOR]

To prevent infections associated with medical implants, various antimicrobial silver-coated implant materials have been developed. However, these materials do not always provide consistent antibacterial effects in vivo despite having dramatic antibacterial effects in vitro, probably because the antibacterial effects involve silver-ion-mediated reactive oxygen species generation. Additionally, the silver application process often requires extremely high temperatures, which damage non-metal implant materials. We recently developed a bacteria-resistant coating consisting of hydroxyapatite film on which ionic silver is immobilized via inositol hexaphosphate chelation, using a series of immersion and drying steps performed at low heat. Here we applied this coating to a polymer, polyetheretherketone (PEEK), and analyzed the properties and antibacterial activity of the coated polymer in vitro and in vivo. The ionic silver coating demonstrated significant bactericidal activity and prevented bacterial biofilm formation in vitro. Bio-imaging of a soft tissue infection mouse model in which a silver-coated PEEK plate was implanted revealed a dramatic absence of bacterial signals 10 days after inoculation. These animals also showed a strong reduction in histological features of infection, compared to the control animals. This innovative coating can be applied to complex structures for clinical use, and could prevent infections associated with a variety of plastic implants. [ABSTRACT FROM AUTHOR]

Study Design: Retrospective multicenter study.Objective: We aimed to assess the outcomes following posterior cervical decompression for cervical spondylotic myelopathy (CSM) in a large sample of patients older than 80 years.Summary Of Background Data: Although age is a predictor of surgical outcomes for CSM, most previous studies have only investigated the effect of age for patients aged 65 to 75 years, and surgical outcomes remain unclear for older patients.Methods: Patients older than 50 years who underwent posterior cervical decompression for CSM were enrolled from 17 institutions. The patients were grouped into a young-old group (<80 yrs old) and an old-old group (≥80 yrs). Clinical information, comorbidities, perioperative complications, Japanese Orthopaedic Association (JOA) scores, Japanese Orthopaedic Association Cervical Myelopathy Evaluation Questionnaire (JOACMEQ), visual analog scale (VAS) scores, and radiographic parameters were compared between the two groups. All patients were followed for at least 1 year after surgery.Results: The young-old and old-old groups included 491 and 77 patients, respectively. The JOA score and quality of life measured by JOACMEQ improved significantly after surgery in both groups. Compared with the young-old group, the old-old group had lower preoperative JOA scores (9.6 vs. 11.0), lower final JOA scores (12.6 vs. 14.0), and lower recovery rates (42% vs. 50%). However, there were no significant differences in perioperative complications or in the VAS scores at the final assessments. The old-old group had a higher mean preoperative sagittal Cobb angle at C2-C5, though this was not statistically significant postoperatively.Conclusion: Posterior decompression surgery is safe and beneficial for patients older than 80 years with CSM, despite having a more limited neurological recovery compared with younger patients.Level Of Evidence: 3. [ABSTRACT FROM AUTHOR]

We devised a simple and rapid technique for establishing axenic cultures of centric diatoms by sieving and washing resting stage cells. This technique is an effective means for establishing the axenic cultures of species that have endogenous resting spores. The primary advantage of this method is in the isolation of cells when the target species is absent from the water column. [ABSTRACT FROM AUTHOR]

Pull-out force and insertion torque have not been generally used as intraoperative measures for the evaluation of pedicle screw stability because of their invasiveness. On the other hand, resonance frequency analysis is a non-invasive and repeatable technique that has been clinically used in dentistry to evaluate implant stability e.g. by the Osstell apparatus. In this study, the characteristics of the implant stability quotient (ISQ) value obtained by the Osstell apparatus in the field of spinal surgery were investigated. Biomechanical test materials simulating human bone were used to provide a comparative platform for evaluating each fixation strength measure, including pull-out force, insertion torque, and the ISQ value. To perform pull-out force measurement and to repeat pedicle screw insertion and removal, loosening was artificially created, and its effect was investigated. The grade of loosening was quantified on a micro-CT image after pedicle screw removal. In the comparison of the 3 fixation strength measures, the correlations of the ISQ value with the pull-out force (R2 = 0.339 p <0.0001) and the insertion torque (R2 = 0.337 p <0.0001) were lower than the correlation between pull-out force and insertion torque (R2 = 0.918 p <0.0001). On a micro-CT study, the material volume of the internal threads disappeared after destruction of its integrity due to repeated pedicle screw insertion and removal. Material integrity destruction of the internal threads decreased only the pull-out force and the insertion torque, but it did not affect the ISQ value. The ISQ value only decreased when the material volume of the internal threads disappeared, probably because the ISQ value reflects the resistance against a force in the perpendicular direction of the screw, unlike the conventional measures of fixation strength, such as pull-out force and insertion torque, which reflect axial load. [ABSTRACT FROM AUTHOR]

Purpose: Hinge-like hyper-mobility is occasionally observed at the atlanto-occipital (O-C1) joint. However, it has not been clear if this kind of hinge-like hyper-mobility at the O-C1 joint should be regarded as "pathologic", or referred to as "instability". To solve this issue, we aimed to establish a reliable radiographic assessment method for this specific type of O-C1 instability and figure out the "standard value" for the range of motion (ROM) of the O-C1 joint.Methods: To figure out the standard range of the O-C1 angle, we acquired magnetic resonance imaging (MRI) sagittal views of the cervical spine for 157 healthy volunteers [average: 37.4 year-old (yo)] without spine diseases, at neutral, maximum flexion and maximum extension positions.Results: The average value (AVE) for ROM of O-C1 angle was 9.91°. The standard value for ROM of O-C1 angle was calculated as 0°-21°. There was no statistically significant gender difference. We also found that the older population (≧ 40 yo) significantly had a larger ROM of O-C1 angle (AVE: 11.72°) compared to the younger population (< 40 yo) (AVE: 8.99°).Conclusions: We consider that hinge-like instability at O-C1 joint, which cannot be assessed by measuring Powers ratio, can be assessed by measuring the range of O-C1 angles using dynamic-MRI. Evaluation of O-C1 instability is important especially when we perform surgical treatment for diseases with upper cervical instability (such as retro-odontoid pseudotumor). We consider that the current study provides important information in such a case. [ABSTRACT FROM AUTHOR]

Background Pemphigus foliaceus (PF) is an autoimmune blistering disease caused by autoantibodies (Abs) against desmoglein 1 (Dsg1). PF sera contain polyclonal Abs which are heterogeneous mixture of both pathogenic and non-pathogenic Abs, as shown by isolation of monoclonal Abs (mAbs). Objective To investigate how pathogenic and non-pathogenic anti-Dsg1 Abs contribute to blister formation in PF. Methods Using organ-cultured human skin, we compared the effect of a single pathogenic anti-Dsg1 IgG mAb, a single non-pathogenic anti-Dsg1 IgG mAb, and their mixture on blister formation as analyzed by histology, subcellular localization of IgG deposits and desmosomal proteins by confocal microscopy, and desmosomal structure by electron microscopy. In addition, we measured keratinocyte adhesion by an in vitro dissociation assay. Results 24 h after injection, a single pathogenic anti-Dsg1 IgG caused a subcorneal blister with IgG and Dsg1 localized linearly on the cell surface of keratinocytes. A single non-pathogenic anti-Dsg1 IgG bound linearly on the keratinocytes but did not induce blisters. A pathogenic and a non-pathogenic IgG mAb injected together caused an aberrant granular pattern of IgG and Dsg1 in the lower epidermis with blister formation in the superficial epidermis. Electron microscopy demonstrated that the mixture of mAbs shortened desmosomal lengths more than a single mAb in the basal and spinous layers. Furthermore, although Dsg1 clustering required both cross-linking of Dsg1 molecules by the non-pathogenic IgG plus a pathogenic antibody, the latter could be in the form of a monovalent single chain variable fragment, suggesting that loss of trans-interaction of Dsg1 is required for clustering. Finally, a p38MAPK inhibitor blocked Dsg1 clustering. When pathogenic strength was measured by the dissociation assay, a mixture of pathogenic and non-pathogenic IgG mAbs disrupted keratinocyte adhesion more than a single pathogenic mAb. This pathogenic effect was only partially suppressed by the p38MAPK inhibitor. Conclusion These findings indicate that a polyclonal mixture of anti-Dsg1 IgG antibodies enhances pathogenic activity for blister formation associated with p38MAPK-dependent Dsg1 clustering and that not only pathogenic antibodies but also non-pathogenic antibodies coordinately contribute to blister formation in PF. [ABSTRACT FROM AUTHOR]

Musculoskeletal infections, including surgical-site and implant-associated infections, often cause progressive inflammation and destroy areas of the soft tissue. Treating infections, especially those caused by multi-antibiotic resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA) remains a challenge. Although there are a few animal models that enable the quantitative evaluation of infection in soft tissues, these models are not always reproducible or sustainable. Here, we successfully established a real-time, in vivo, quantitative mouse model of soft-tissue infection in the superficial gluteus muscle (SGM) using bioluminescence imaging. A bioluminescent strain of MRSA was inoculated into the SGM of BALB/c adult male mice, followed by sequential measurement of bacterial photon intensity and serological and histological analyses of the mice. The mean photon intensity in the mice peaked immediately after inoculation and remained stable until day 28. The serum levels of interleukin-6, interleukin-1 and C-reactive protein at 12 hours after inoculation were significantly higher than those prior to inoculation, and the C-reactive protein remained significantly elevated until day 21. Histological analyses showed marked neutrophil infiltration and abscesses containing necrotic and fibrous tissues in the SGM. With this SGM mouse model, we successfully visualized and quantified stable bacterial growth over an extended period of time with bioluminescence imaging, which allowed us to monitor the process of infection without euthanizing the experimental animals. This model is applicable to in vivo evaluations of the long-term efficacy of novel antibiotics or antibacterial implants. [ABSTRACT FROM AUTHOR]

Although minimally invasive transforaminal lumbar interbody fusion (MIS-TLIF) has widely been developed in patients with lumbar diseases, surgeons risk exposure to fluoroscopic radiation. However, to date, there is no studies quantifying the effective dose during MIS-TLIF procedure, and the radiation dose distribution is still unclear. In this study, the surgeons' radiation doses at 5 places on the bodies were measured and the effective doses were assessed during 31 consecutive 1- to 3-level MIS-TLIF surgeries. The operating surgeon, assisting surgeon, and radiological technologist wore thermoluminescent dosimeter on the unshielded thyroid, chest, genitals, right middle finger, and on the chest beneath a lead apron. The doses at the lens and the effective doses were also calculated. Mean fluoroscopy times were 38.7, 53.1, and 58.5 seconds for 1, 2, or 3 fusion levels, respectively. The operating surgeon's mean exposures at the lens, thyroid, chest, genitals, finger, and the chest beneath the shield, respectively, were 0.07, 0.07, 0.09, 0.14, 0.32, and 0.05 mSv in 1-level MIS-TLIF; 0.07, 0.08, 0.09, 0.18, 0.34, and 0.05 mSv in 2-level; 0.08, 0.09, 0.14, 0.15, 0.36, and 0.06 mSv in 3-level; and 0.07, 0.08, 0.10, 0.15, 0.33, and 0.05 mSv in all cases. Mean dose at the operating surgeon's right finger was significantly higher than other measurements parts (P<0.001). The operating surgeon's effective doses (0.06, 0.06, and 0.07 mSv for 1, 2, and 3 fusion levels) were low, and didn't differ significantly from those of the assisting surgeon or radiological technologist. Revision MIS-TLIF was not associated with higher surgeons' radiation doses compared to primary MIS-TLIF. There were significantly higher surgeons' radiation doses in over-weight than in normal-weight patients. The surgeons' radiation exposure during MIS-TLIF was within the safe level by the International Commission on Radiological Protection's guidelines. The accumulated radiation exposure, especially to surgeon's hands, should be carefully monitored. [ABSTRACT FROM AUTHOR]

Information on diatom resting stages is fundamentally important to understanding the population dynamics of diatoms including bloom formation. The distribution of viable diatom resting stage cells in bottom sediments of the eastern Bering Sea in July 2009 was investigated by the most probable number (MPN) method. The abundances of diatom resting stage cells ranged from 1.7×103 to 1.2×106 MPNcellscm−3 wet sediment, comparable to those in shallow eutrophic areas where diatom blooms frequently occur. Common species during the spring phytoplankton bloom in the eastern Bering Sea were also dominant in sediments as resting stage cells. It should be noted that relatively high numbers of ice algae species, especially ribbon-shaped chain forming pennate diatoms, were found in the sediments. The life cycle strategy using resting stage cells allows planktonic and ice algal species to survive unfavorable environmental conditions such as the dark winter season, and potentially contribute to form blooms of several types (subsurface of ice, ice edge, plankton) through vertical mixing. [ABSTRACT FROM AUTHOR]

Abstract: The effectiveness of influenza vaccines is still controversial, and the role of adjuvants in such vaccines is briefly reviewed in this paper. Inactivated whole virus vaccines may include components that function as adjuvants, meaning that additive adjuvants are often not required. MF59 and AS03 showed higher adjuvanticity than aluminum salts in several clinical studies. Recent research has suggested that immune cell recruitment is the main mechanism underlying adjuvant actions in general, and that aluminum salts induce this recruitment via inflammation at the injected site. The aspect of how oil-based adjuvants, such as MF59 and AS03, recruit immune cells remains to be clarified. [Copyright &y& Elsevier]

The demand is currently high for new vaccination strategies, particularly to help combat problematic intracellular pathogens, such as HIV and malarial parasites. In the past decade, the identification of host receptors that recognize pathogen-derived nucleic acids has revealed an essential role for nucleic acid sensing in the triggering of immunity to intracellular pathogens. This Review first addresses our current understanding of the nucleic acid-sensing immune machinery. We then explain how the study of nucleic acid-sensing mechanisms not only has revealed their central role in driving the responses mediated by many current vaccines, but is also revealing how they could be harnessed for the design of new vaccines. [ABSTRACT FROM AUTHOR]

Abstract: To evaluate the responses to the spring phytoplankton bloom, short-term changes in population structure and vertical distribution of mesopelagic copepods (Gaetanus simplex, Gaidius variabilis, Pleuromamma scutullata, Paraeuchaeta elongata, P. birostrata, Heterorhabdus tanneri and Heterostylites major) were studied in the Oyashio region. Samples were collected with a 60μm mesh VMPS from 9 strata between 0 and 1000m both day and night on five occasions during March–April 2007. All the species except Heterorhabdidae species performed reproduction during the spring phytoplankton bloom, while no recruitment to copepodid stages was detected because the newly born individuals were eggs or nauplii. The shallower-living species, G. simplex, P. scutullata and P. elongata had nocturnal ascent diel vertical migration (DVM). While suspension feeding copepods cease DVM after 11 April (P. scutullata) or 23 April (G. simplex), carnivorous P. elongata continued DVM over the study period. Since the gut contents of G. simplex showed a nocturnal increment even in the period of no DVM (23 and 29 April), they might be feeding at depth without DVM. Thus, the cessation of DVM in mesopelagic suspension feeding copepods would be induced by the increase of sinking particles (e.g. food for suspension feeders) during the spring phytoplankton bloom. [Copyright &y& Elsevier]

Paraneoplastic pemphigus (PNP) is an autoimmune blistering disease associated with lymphoproliferative neoplasms and characterized by antibodies against plakins and desmoglein 3 (Dsg3). Anti-Dsg3 antibodies have a primary role in blister formation in PNP. In this study, we used phage display to clone monoclonal anti-Dsg3 antibodies from a PNP patient to further characterize their pathogenicity. We isolated 20 unique Dsg3-reactive mAbs, which we classified into four groups according to the heavy-chain complementarity-determining region 3 (CDR3) region. Genetic analyses demonstrated that three antibody groups used the VH1-46 gene (18 clones) and one group used the VH1-02 gene (2 clones). The results of an in vitro keratinocyte dissociation assay and a human skin organ culture injection assay showed that three antibodies displayed pathogenic activity in blister formation with different potencies. Epitope mapping using domain-swapped Dsg3/Dsg2 showed that these pathogenic mAbs bound Ca2+-dependent conformational epitopes in the middle portion of the extracellular region of Dsg3 (EC2 and EC3 domains), in contrast to most previously characterized pathogenic pemphigus vulgaris antibodies, which bound to the EC1 domain of Dsg3. These mAbs reflect the unique polyclonal nature of anti-Dsg3 antibodies in PNP and represent an important tool for detailing the pathophysiological mechanisms of blister formation in PNP. [ABSTRACT FROM AUTHOR]

I shii K.-I., I wataki M., M atsuoka K. and I mai I. 2011. Proposal of identification criteria for resting spores of species (Bacillariophyceae) from a temperate coastal sea. 50: 351--362. DOI: 10.2216/10-36.1 is a large, ecologically important genus of marine planktonic diatoms. Many species of form resting spores to survive environmental stress. Species identification based on morphological characteristics of resting spores is difficult. In the present study, we proposed a flow diagram for the identification of species based on morphology. Resting spores of 18 species were identified in water and sediment samples from southern Japan and the East China Sea based on morphological characteristics of resting spores and/or the vegetative cells containing them. A single ring of puncta was found to be the most important morphological feature of resting spores for species identification, by which the positions of primary and secondary valves were correctly determined. Other morphological characteristics such as features of the primary valve face, margins of the primary and secondary valve mantles, and secondary valve face are also useful for species identification. A combination of these morphological characteristics was used to construct a flow diagram for identification of the 18 species found in our samples with light microscopy. This flow diagram is most applicable to Japanese waters, but the approach should be refined and improved to understand the biology and ecology of species in any marine ecosystem. [ABSTRACT FROM AUTHOR]

Abstract: Background: Epidermolysis bullosa acquisita (EBA) is an acquired autoimmune mechanobullous disease. EBA patients possess autoantibodies against type VII collagen which is composed of a collagenous domain flanked by non-collagenous NC1 and NC2 domains. It was reported that major epitopes reside within the NC1 domain and minor epitopes reside within NC2 domain. Objective: The aim of this study is to develop a sensitive and specific ELISA to facilitate the diagnosis of EBA. Methods: We developed ELISAs using recombinant NC1 domain produced by mammalian expression system and recombinant NC2 domain produced by mammalian or bacterial expression system to characterize autoantibodies in EBA. Next, we developed an ELISA using a combination of the NC1 (mammalian expression) and NC2 (bacterial expression). We tested the ELISAs with 49 EBA sera, 55 normal control sera, 20 pemphigus vulgaris and 20 bullous pemphigoid sera. Results: When we evaluated the 49 EBA sera using the NC1 and NC2 ELISAs, 38 (77.5%) reacted with NC1 domain only, 7 sera (14.2%) reacted with both NC1 and NC2 domains, and one serum (2%) reacted with NC2 domain only. Therefore, to increase the sensitivity of the assay, we developed an ELISA coated with a mixture of recombinant NC1 and NC2 domains, resulting in 93.8% sensitivity and 98.1% specificity. By analyzing the time course of two EBA patients, ELISA scores fluctuated in parallel with their disease activity. Conclusion: We conclude that the NC1+NC2 ELISA can be a practical assay for the diagnosis and follow up of the antibody titers of EBA patients. [Copyright &y& Elsevier]

Multipotent somatic stem cells have been identified in various adult tissues. However, the stem/progenitor cells of the peripheral nerves have been isolated only from fetal tissues. Here, we isolated Schwann-cell precursors/immature Schwann cells from the injured peripheral nerves of adult mice using a floating culture technique that we call "Schwann-spheres." The Schwann-spheres were derived from de-differentiated mature Schwann cells harvested 24 hours to 6 weeks after peripheral nerve injury. They had extensive self-renewal and differentiation capabilities. They strongly expressed the immature- Schwann-cell marker p75, and differentiated only into the Schwann-cell lineage. The spheres showed enhanced myelin formation and neurite growth compared to mature Schwann cells in vitro. Mature Schwann cells have been considered a promising candidate for cell-transplantation therapies to repair the damaged nervous system, whereas these "Schwannspheres" would provide a more potential autologous cell source for such transplantation. [ABSTRACT FROM AUTHOR]

Summary: Particulates such as silica crystal (silica) and aluminum salts (alum) activate the inflammasome and induce the secretion of proinflammatory cytokines in macrophages. These particulates also induce the production of immunoglobulin E via a T helper 2 (Th2) cell-associated mechanism. However, the mechanism involved in the induction of type 2 immunity has not been elucidated. Here, we showed that silica and alum induced lipopolysaccharide-primed macrophages to produce the lipid mediator prostaglandin E2 (PGE2) and interleukin-1β (IL-1β). Macrophages deficient in the inflammasome components caspase 1, NALP3, and ASC revealed that PGE2 production was independent of the NALP3 inflammasome. PGE2 expression was markedly reduced in PGE synthase-deficient (Ptges −/−) macrophages, and Ptges −/−mice displayed reduced antigen-specific serum IgE concentrations after immunization with alum or silica. Our results indicate that silica and alum regulate the production of PGE2 and that the induction of PGE2 by particulates controls the immune response in vivo. [Copyright &y& Elsevier]

The article focuses on a study which investigated the risk factors for developing subaxial subluxations (SAS) following C1-C2 arthrodesis for atlantoaxial (AA) subluxation in patients with rheumatoid arthritis (RA). The study included 58 patients with RA who underwent C1-C2 arthrodesis. The patients preoperative and sequential postoperative radiographs were evaluated. Study authors concluded that excessive correction of AA angle can cause loss of cervical lordosis.

Autoantibodies in pemphigus foliaceus (PF) and vulgaris (PV) bind to desmoglein (Dsg) 1 and 3, respectively, and cause loss of keratinocyte adhesion. To characterize the pathogenicity and genetics of such antibodies we have used phage display to isolate monoclonal antibodies (mAbs) from patients. PCR is used to clone the heavy and light chain variable region of the peripheral B cells into a vector that creates a phage particle with the antibody expressed on its surface and the cDNA encoding that antibody inside. The library of phage produced from a PF or PV patient are then panned on a plate containing Dsg1 or Dsg3 to isolate clones. The cDNA of each clone is sequenced to characterize the genetics of the expressed mAb. The mAb from each unique clone is tested for pathogenicity either by injecting into normal human skin organ culture or into neonatal mice. Pathogenic antibodies cause typical pemphigus blisters. In both PV and PF patients the heavy chain (VH) genes used for Dsg-binding antibodies are severely restricted. PV and PF patients have both pathogenic and non-pathogenic mAbs. The immunochemical characteristics of the antibodies (including pathogenicity) sort with the VH, not the VL, gene. These monoclonal pathogenic antibodies can be used to screen peptide libraries to find short peptides that block antibody binding. In summary, the antibody response is restricted and, therefore, it may be feasible to target the specific pathogenic antibodies for therapy. [ABSTRACT FROM AUTHOR]

During microbial infection and tissue injury, nucleic acids and their metabolites, such as nucleotides, nucleosides and uric acids, can be released from dying host cells and may modify immune responses. These nucleic acids and/or their metabolites are in fact recognized by specific host receptors, such as Toll-like receptors (TLRs), RIG-like receptors (RLRs) and NOD-like receptors (NLRs), purinergic receptors such as P2X and P2Y receptors, and adenosine receptors such as A2A receptors. The resultant responses may vary depending on the balance between immune responses to and metabolism of nucleic acids, thereby contributing not only to the host defense, but also to the homeostatic clearance of host dying cells, or even to deleterious autoimmune diseases. [Copyright &y& Elsevier]

Abstract: In viral infections the host innate immune system is meant to act as a first line defense to prevent viral invasion or replication before more specific protection by the adaptive immune system is generated. In the innate immune response, pattern recognition receptors (PRRs) are engaged to detect specific viral components such as viral RNA or DNA or viral intermediate products and to induce type I interferons (IFNs) and other pro-inflammatory cytokines in the infected cells and other immune cells. Recently these innate immune receptors and their unique downstream pathways have been identified. Here, we summarize their roles in the innate immune response to virus infection, discrimination between self and viral nucleic acids and inhibition by virulent factors and provide some recent advances in the coordination between innate and adaptive immune activation. [Copyright &y& Elsevier]

Mammalian innate immunity possesses a distinct system to recognize aberrant DNA inside the cell. One class of DNA sensors is the Toll-like receptor 9, which is expressed in the specialized immune cells, binds to single-stranded DNA in the endosome to transmit cellular signaling through myeloid differentiation primary response protein 88 (MyD88). Another class of DNA sensors exists in the cytoplasm of most type of cells in the tissue, detecting double-stranded DNA to signal through TANK-binding kinase-1 (TBK1)-mediated type-I interferon production and apoptosis-associated speck-like protein containing a CARD (ASC)-mediated IL-1β secretion. Since DNA sensors have potential to recognize aberrant DNA of both self and nonself origin, their physiological roles in microbial infection, tissue damage, autoimmune diseases, and DNA-based therapeutic applications are being intensively investigated. [Copyright &y& Elsevier]

Pemphigus foliaceus (PF) is a blistering disease caused by autoantibodies to desmoglein 1 (Dsg1) that cause loss of epidermal cell adhesion. To better understand PF pathophysiology, we used phage display to isolate anti-Dsg1 mAbs as single-chain variable fragments (scFvs) from a PF patient. Initial panning of the library isolated only non-pathogenic scFvs. We then used these scFvs to block non-pathogenic epitopes and were able to isolate two unique scFvs, each of which caused typical PF blisters in mice or human epidermis models, showing that a single mAb can disrupt Dsg1 function to cause disease. Both pathogenic scFvs bound conformational epitopes in the N terminus of Dsg1. Other PF sera showed a major antibody response against the same or nearby epitopes defined by these pathogenic scFvs. Finally, we showed restriction of the heavy-chain gene usage of all anti-Dsg1 clones to only five genes, which determined their immunological properties despite promiscuous light-chain gene usage. These mAbs will be useful for studying Dsg1 function and mechanisms of blister formation in PF and for developing targeted therapies and tools to monitor disease activity.Journal of Investigative Dermatology (2008) 128, 939–948; doi:10.1038/sj.jid.5701132; published online 15 November 2007 [ABSTRACT FROM AUTHOR]

Extracellular ATP, an autocrine or paracrine intercellular transmitter, is known to induce apoptosis in macrophages. However, the precise signaling mechanisms of ATP-induced apoptosis remain to be elucidated. Here we showed that activation of p38 mitogen-activated protein kinase (MAPK) plays a critical role in ATP-induced apoptosis. p38 activation and apoptosis in macrophages were induced by ATP. ATP-induced apo- ptosis was mediated in part by production of reactive oxygen species (ROS) derived from NOX2/gp91phox, a component of the NADPH oxidase complex expressed in macrophages and neutrophils. Furthermore, ATP-induced ROS generation, p38 activation, and apoptosis were almost completely inhibited by selective P2X7 receptor antagonists. We also found that ATP-induced apoptosis were diminished in ASK1-deficient macrophages accompanied by the lack of p38 activation. These results demonstrate that ROS-mediated activation of the ASK1- p38 MAPK pathway downstream of P2X7 receptor is required for ATP-induced apoptosis in macrophages. [ABSTRACT FROM AUTHOR]

Successful vaccines contain not only protective antigen(s) but also an adjuvant component that triggers innate immune activation and is necessary for their optimal immunogenicity. In the case of DNA vaccines, this consists of plasmid DNA; however, the adjuvant element(s) as well as its intra- and inter-cellular innate immune signalling pathway(s) leading to the encoded antigen-specific T- and B-cell responses remain unclear. Here we demonstrate in vivo that TANK-binding kinase 1 (TBK1), a non-canonical IκB kinase, mediates the adjuvant effect of DNA vaccines and is essential for its immunogenicity in mice. Plasmid-DNA-activated, TBK1-dependent signalling and the resultant type-I interferon receptor-mediated signalling was required for induction of antigen-specific B and T cells, which occurred even in the absence of innate immune signalling through a well known CpG DNA sensor—Toll-like receptor 9 (TLR9) or Z-DNA binding protein 1 (ZBP1, also known as DAI, which was recently reported as a potential B-form DNA sensor). Moreover, bone-marrow-transfer experiments revealed that TBK1-mediated signalling in haematopoietic cells was critical for the induction of antigen-specific B and CD4+ T cells, whereas in non-haematopoietic cells TBK1 was required for CD8+ T-cell induction. These data suggest that TBK1 is a key signalling molecule for DNA-vaccine-induced immunogenicity, by differentially controlling DNA-activated innate immune signalling through haematopoietic and non-haematopoietic cells. [ABSTRACT FROM AUTHOR]

It has long been known that malaria infection causes host immune modulation by various mechanisms. However, the role of Toll-like receptors (TLRs) in mediating innate immune responses to parasite-derived components during the blood stages of malaria has only recently been described. TLRs might have an important role in pathogenesis during malaria infection, as supported by genetic analyses in mice and humans. Moreover, recent findings revealed that sporozoites can partially differentiate in lymph nodes and that liver stages induce the formation of previously unknown parasite-filled vesicles (merosomes) that could function as immune escape machinery. Elucidation of the mechanisms by which the host innate immune system responds to, and/or is manipulated by, Plasmodium infection will hopefully lead to discoveries of potential targets that will ultimately prevent and/or intervene in malaria infection. [Copyright &y& Elsevier]

Cyclic AMP regulates multiple neuronal functions, including neurite outgrowth and axonal regeneration. GPR3, GPR6, and GPR12 make up a family of constitutively active G protein-coupled receptors (GPCRs) that share greater than 50% identity and 65% similarity at the amino acid level. They are highly expressed in the central nervous system, and their expression in various cell lines results in constitutive stimulation of cAMP production. When the constitutively active GPCRs were overexpressed in rat cerebellar granule neurons in culture, the transfected neurons exhibited significantly enhanced neurite outgrowth and overcame growth inhibition caused by myelin-associated glycoprotein. GPR12-mediated neurite outgrowth was the most prominent and was shown to depend on Gs and cAMP-dependent protein kinase, Moreover, the GPR12-mediated rescue from myelin-associated glycoprotein inhibition was attributable to cAMP-dependent protein kinase-mediated inhibition of the small GTPase, RhoA. Among the three receptors, GPR3 was revealed to be enriched in the developing rat cerebellar granule neurons. When the endogenous GPR3 was knocked down, significant reduction of neurite growth was observed, which was reversed by expression of either GPR3 or GPR12. Taken together, our results indicate that expression of the constitutively active GPCRs up-regulates cAMP production in neurons, stimulates neurite outgrowth, and counteracts myelin inhibition. Further characterization of the GPCRs in developing and injured mammalian neurons should provide insights into how basal cAMP levels are regulated in neurons and could establish a firm scientific foundation for applying receptor biology to treatment of various neurological disorders. [ABSTRACT FROM AUTHOR]

DNA in microbes or host cells is normally sequestered from the immune system, and therefore inert, but becomes an active immunostimulatory molecule during infection or tissue damage. Recent evidence suggests that Toll-like receptor (TLR)9, currently the only known immune sensor for DNA, recognizes more diverse elements in its ligand than initially thought, and must cooperate with additional host factors to provoke an optimal innate immune response in the physiological environment. Moreover, the innate immune system possesses a TLR9-independent, as-yet-undefined intracellular recognition machinery of double-stranded DNA that induces type I interferons through distinct signaling pathways. TLR9-dependent and TLR9-independent immune recognition of DNA might play crucial roles in DNA-associated protective immunity and in pathological autoimmunity. [Copyright &y& Elsevier]

Introducing double-stranded DNA (dsDNA) into the cytoplasm of macrophages and dendritic cells triggers the activation of these professional antigen-presenting cells (APCs). This process is characterized by the up-regulation of costimulatory molecules and the production of various cytokines, chemokines, and antibacterial/viral factors. Current findings indicate that interferon-β (IFN-β) plays a key role in the stimulatory cascade triggered by dsDNA. Both immune and non-immune cells respond to intracytoplasmic dsDNA by up-regulating IFN-β) expression, a process that reduces host susceptibility to infection. The immune activation induced by dsDNA is independent of MyD88, TRIF and DNA-PKcs, indicating that a Toll-like receptor-independent mechanism underlies the cellular activation mediated by intracytoplasmic dsDNA. [ABSTRACT FROM AUTHOR]

The innate immune system recognizes nucleic acids during infection or tissue damage; however, the mechanisms of intracellular recognition of DNA have not been fully elucidated. Here we show that intracellular administration of double-stranded B-form DNA (B-DNA) triggered antiviral responses including production of type I interferons and chemokines independently of Toll-like receptors or the helicase RIG-I. B-DNA activated transcription factor IRF3 and the promoter of the gene encoding interferon-β through a signaling pathway that required the kinases TBK1 and IKKi, whereas there was substantial activation of transcription factor NF-κB independent of both TBK and IKKi. IPS-1, an adaptor molecule linking RIG-I and TBK1, was involved in B-DNA-induced activation of interferon-β and NF-κB. B-DNA signaling by this pathway conferred resistance to viral infection in a way dependent on both TBK1 and IKKi. These results suggest that both TBK1 and IKKi are required for innate immune activation by B-DNA, which might be important in antiviral innate immunity and other DNA-associated immune disorders. [ABSTRACT FROM AUTHOR]

Patients with pemphigus vulgaris (PV) have circulating anti-desmoglein (Dsg) 3 immunoglobulin G (IgG) autoantibodies that induce blister formation. We developed anin vitroquantitative assay to evaluate the pathogenic strength of anti-Dsg3 IgG autoantibodies in blister formation. To obtain intercellular adhesion mediated dominantly by Dsg3, we used primary cultured normal human keratinocytes expressing low level of Dsg2 in the presence of exfoliative toxin A that specifically digests Dsg1. After incubation with various antibodies, monolayers released by dispase were subjected to mechanical stress by pipetting, and the number of cell fragments were counted. When anti-Dsg3 monoclonal antibodies (mAb) obtained from pemphigus model mice were tested, pathogenic AK23 mAb yielded significantly higher number of cell fragments than AK7 or AK20 non-pathogenic mAb. Dissociation scores, defined with AK23 mAb as the positive control, were significantly higher with active stage PV sera (n=10, 77.4±21.4) than controls (n=11, 16.0±9.6; p=0.003). When pair sera obtained from 6 PV patients in active stage and in remission were compared, the dissociation scores reflected well the disease activity as those in active stage were four to 17 times higher than those in remission. When sera from different patients showing similar ELISA scores but different clinical severity were tested (n=6), the dissociation scores with sera from severe disease activity were significantly higher than those with sera in remission. These findings indicate that this dissociation assay will provide a simple and objective biological method to measure the pathogenic strength of pemphigus autoantibodies. [ABSTRACT FROM AUTHOR]

Pemphigus is a life-threatening blistering disorder of the skin and mucous membranes caused by pathogenic autoantibodies to desmosomal adhesion proteins desmoglein 3 (Dsg3) and Dsg1. Mechanisms of antibody pathogenicity are difficult to characterize using polyclonal patient sera. Using antibody phage display, we have isolated repertoires of human anti-Dsg mAbs as single-chain variable-region fragments (scFvs) from a patient with active mucocutaneous pemphigus vulgaris. ScFv mAbs demonstrated binding to Dsg3 or Dsg1 alone, or both Dsg3 and Dsg1. Inhibition ELISA showed that the epitopes defined by these scFvs are blocked by autoantibodies from multiple pemphigus patients. Injection of scFvs into neonatal mice identified 2 pathogenic scFvs that caused blisters histologically similar to those observed in pemphigus patients. Similarly, these 2 scFvs, but not others, induced cell sheet dissociation of cultured human keratinocytes, indicating that both pathogenic and nonpathogenic antibodies were isolated. Genetic analysis of these mAbs showed restricted patterns of heavy and light chain gene usage, which were distinct for scFvs with different desmoglein-binding specificities. Detailed characterization of these pemphigus mAbs should lead to a better understanding of the immunopathogenesis of disease and to more specifically targeted therapeutic approaches. [ABSTRACT FROM AUTHOR]

Objective: To examine the effect of suppressive oligodeoxynucleotides (ODNs) on the pulmonary inflammation induced by immunostimulatory CpG DNA.Design: Prospective, randomized, controlled study.Setting: Research laboratories.Subjects: RAW 264.7 murine macrophage-like cell line and BALB/c mice.Interventions: RAW 264.7 cells were incubated with bacterial DNA or CpG ODN, alone or combined with suppressive ODN. The in vivo effect of suppressive ODN was determined using an acute lung injury model. CpG ODN alone or combined with suppressive ODN was instilled into the mouse lung.Measurements and Main Results: Production of tumor necrosis factor (TNF)-alpha and macrophage inflammatory protein (MIP)-2 by RAW 264.7 cells were measured by enzyme-linked immunosorbent assay (ELISA), whereas their messenger RNA levels were determined by reverse transcriptase-polymerase chain reaction. Synthetic ODN containing CpG motifs (CpG ODN) mimicked the ability of bacterial DNA to stimulate the production of TNF-alpha and MIP-2. Suppressive ODN significantly inhibited the activation of RAW 264.7 cells by both bacterial DNA and CpG ODN. In the lung injury model, production of proinflammatory cytokines (TNF-alpha and IL-6) and chemokines (MIP-2 and KC) in bronchoalveolar lavage (BAL) fluids was measured by ELISA. Neutrophil accumulation in the alveolar spaces was also evaluated. Instillation of CpG ODN into the lungs of normal mice triggered the synthesis of TNF-alpha, IL-6, MIP-2, and KC. Suppressive ODN significantly blocked the production of these proinflammatory cytokines and chemokines and also reduced neutrophil mobilization into the alveolar spaces by CpG DNA.Conclusions: Proinflammatory cytokines and chemokines are up-regulated by CpG motifs in bacterial DNA. Suppressive ODN significantly inhibits the inflammatory response induced by CpG DNA in murine macrophages and the lung. This study supports the use of suppressive ODN to reduce the deleterious inflammatory responses induced by bacterial DNA. [ABSTRACT FROM AUTHOR]

Desmosomes are intercellular adhesive junctions that exhibit cell- and differentiation-specific differences in their molecular composition. In complex epithelia, desmosomes contain multiple representatives of the desmosomal cadherin family, which includes three desmogleins and three desmocollins. Rules governing the assembly of desmosomal cadherin isoforms into desmosomes of different cell types are unknown. Here we compared the assembly properties of desmoglein 2 (Dsg2) and desmocollin 2 (Dsc2), which are widely expressed, with Dsg1 and Dsc1, which are expressed in the differentiated layers of complex epithelia, by introducing myc-tagged forms into simple and squamous epithelial cells that do not express Dsg1 or Dsc1. Dsc2.myc and Dsg2.myc assembled efficiently into desmosomes in every cell type in spite of significant shifts in the stoichiometric relationship between desmogleins and desmocollins. In contrast, Dsc1a.myc, Dsc1b.myc, and Dsg1.myc did not stably incorporate into desmosomes in any line. Coexpression of Dsc1a.myc or Dsc1b.myc and Dsg1.myc did not lead to their colocalization and failed to enhance incorporation of either cadherin into desmosomes. Dsg1.myc, but not Dsc1a, Dsc1b, disrupted desmosome assembly in a cell-type-specific manner, and disruption correlated with the recruitment of Dsg1.myc, but not Dsc1a or Dsc1b, into a Triton-insoluble pool. The plakoglobin:E-cadherin ratio decreased in Dsg1-expressing cells with disrupted desmosomes, but a decrease was also observed in a Dsc1a line. Thus, a modest reduction of plakoglobin associated with E-cadherin is apparently not sufficient to disrupt desmosome assembly. Our results demonstrate that desmosome assembly tolerates large shifts in cadherin stoichiometry, but is sensitive to isoform-specific differences exhibited by desmogleins and desmocollins. [ABSTRACT FROM AUTHOR]

Chronic bone loss is an under-recognized complication of malaria, the underlying mechanism of which remains incompletely understood. We have previously shown that persistent accumulation of Plasmodium products in the bone marrow leads to chronic inflammation in osteoblast (OB) and osteoclast (OC) precursors causing bone loss through MyD88, an adaptor molecule for diverse inflammatory signals. However, the specific contribution of MyD88 signaling in OB or OC precursors in malaria-induced bone loss remains elusive. To assess the direct cell-intrinsic role of MyD88 signaling in adult bone metabolism under physiological and infection conditions, we used the Lox-Cre system to specifically deplete MyD88 in the OB or OC lineages. Mice lacking MyD88 primarily in the maturing OBs showed a comparable decrease in trabecular bone density by microcomputed tomography to that of controls after Plasmodium yoelii non-lethal infection. In contrast, mice lacking MyD88 in OC precursors showed significantly less trabecular bone loss than controls, suggesting that malaria-mediated inflammatory mediators are primarily controlled by MyD88 in the OC lineage. Surprisingly, however, depletion of MyD88 in OB, but not in OC, precursors resulted in reduced bone mass with decreased bone formation rates in the trabecular areas of femurs under physiological conditions. Notably, insulin-like growth factor-1, a key molecule for OB differentiation, was significantly lower locally and systemically when MyD88 was depleted in OBs. Thus, our data demonstrate an indispensable intrinsic role for MyD88 signaling in OB differentiation and bone formation, while MyD88 signaling in OC lineages plays a partial role in controlling malaria-induced inflammatory mediators and following bone pathology. These findings may lead to the identification of novel targets for specific intervention of bone pathologies, particularly in malaria-endemic regions. [ABSTRACT FROM AUTHOR]

Desmoglein 3 is the autoimmune target of pemphigus vulgaris. Most, if not all, pathogenic autoantibodies are raised against conformational epitopes on desmoglein 3. In this study, we examined whether posttranslational modification in endoplasmic reticulum is required for the proper three-dimensional structure formation of recombinant pemphigus vulgaris antigen. Previously, we have produced by baculovirus expression a secreted form of desmoglein 3, PVIg, which represents equivalent conformational epitopes of the native antigen and showed that PVIg is able to immunoadsorb heterogeneous autoantibodies form pemphigus vulgaris patients sera. To elucidate the role of proteolytic processing, we constructed a mutant PVig, molecule, PVIg moecule, PVIg-fXa, whose putative endoproteolytic cleavage site was replaced by the recogntion sequence of serum coagulation factor Xa. PVIg-fXa was produced without proteolytic processing; however, exogenous treatment of PVIg-fXa with factor Xa resulted in cleavage of the prosequence. Interestingly, not only the processd PVIg-fXa,but also the unprocessed form, showed the immunoadsorptive activity. Furthermore, to elucidate the role of endoplasmic reticulum signal peptide at the amino terminus, we constructed another mutant, PVIg-Δsig, which lacks the signal peptide and prosequence. PVIg-Δsig was not secreted and accumulated in the cytosol. PVIg-Δsig failed to show the immunoadsorption. Together with or previous finding on the role of glycosylation, these observation indicate that the conformational epitopes of the recombinant pemphigus antigen are not affected either by glycosylation or proteolytic processing, although they need to be formed in the endoplasmic reticulum, and emphasize the importance of conformation of the antigen in pathogenic autoantibody binding. [ABSTRACT FROM AUTHOR]

The target molecule of pemphigus autoantibodies is a transmembrane desmosomal component, desmoglein 3 (Dsg3) in pemphigus vulgaris (PV) and Dsg1 in pemphigus foliaceus (PF). In this study, we examined the effects of calcium and glycosylation on the antigenicity of the pemphigus antigens and on the generation of conformational epitopes. We used recombinant baculovirus proteins, PVIg and PFIg, which are considered to reflect accurately the native conformation of the extracellular domain of their respective proteins Dsg3 and Dsg1. These baculoproteins could immunoadsorb heterogeneous autoantibodies from the corresponding sera of PV and PF patients, completely blocking indirect immunofluorescence staining of normal human skin. Chelating calcium from the solution containing the baculoproteins using ethylenediaminetetraacetic acid (EDTA) or ethyleneglycol-bis(β-aminoethyl ether)-N,N,N' ,N'-tetraacetic acid (EGTA) abolished immunoadsorption by both PVIg and PFIg; however, immunoadsorption by the baculoproteins was restored after dialysis against 1 mM calcium. Nonglycosylated forms of both baculoproteins produced in the presence of tunicamycin retained their immunoadsorptive ability. Furthermore, immunoadsorption by the baculoproteins was prevented irreversibly by treatment with low pH, high pH, and boiling, but not with the non-ionic detergent Nonidet P-40. These findings indicate that formation of the conformational epitopes on the pemphigus antigens is dependent on calcium but independent of glycosylation, and provide direct evidence that calcium plays an important role in determining the antigenic properties of the pemphigus antigens. [ABSTRACT FROM AUTHOR]

The discovery and optimization of a novel series of G9a/GLP (EHMT2/1) inhibitors are described. Starting from known G9a/GLP inhibitor 5 , efforts to explore the structure-activity relationship and optimize drug properties led to a novel compound 13 , the side chain of which was converted to tetrahydroazepine. Compound 13 showed increased G9a/GLP inhibitory activity compared with compound 5. In addition, compound 13 exhibited improved human ether-a-go-go related gene (hERG) inhibitory activity over compound 5 and also improved pharmacokinetic profile in mice (oral bioavailability: 17 to 40%). Finally, the co-crystal structure of G9a in complex with compound 13 provides the basis for the further development of tetrahydroazepine-based G9a/GLP inhibitors. [ABSTRACT FROM AUTHOR]

Assesses primary neural system/progenitor cells (NSPC) that were transplanted into injured spinal cord. Bioluminescence in the lentivirally transduced NSPC; Viability studies in spinal cord injury; Importance of the timing of NSPC transplantation to the function of integrated cells for the injured spinal cord.

CpG methylation of DNA silences TLR9-mediated innate immune recognition. In this issue of Immunity, suggest that the innate immune recognition of RNA by TLR3, TLR7, or TLR8 is in fact controlled by modification of nucleotides, including methylation. [Copyright &y& Elsevier]

Bone marrow is a dynamic organ composed of stem cells that constantly receive signals from stromal cells and other hematopoietic cells in the niches of the bone marrow to maintain hematopoiesis and generate immune cells. Perturbation of the bone marrow microenvironment by infection and inflammation affects hematopoiesis and may affect immune cell development. Little is known about the effect of malaria on the bone marrow stromal cells that govern the hematopoietic stem cell (HSC) niche. In this study, we demonstrate that the mesenchymal stromal CXCL12-abundant reticular (CAR) cell population is reduced during acute malaria infection. The reduction of CXCL12 and interleukin-7 signals in the bone marrow impairs the lymphopoietic niche, leading to the depletion of common lymphoid progenitors, B cell progenitors, and mature B cells, including plasma cells in the bone marrow. We found that interferon-γ (IFNγ) is responsible for the upregulation of Sca1 on CAR cells, yet the decline in CAR cell and B cell populations in the bone marrow is IFNγ-independent. In contrast to the decline in B cell populations, HSCs and multipotent progenitors increased with the expansion of myelopoiesis and erythropoiesis, indicating a bias in the differentiation of multipotent progenitors during malaria infection. These findings suggest that malaria may affect host immunity by modulating the bone marrow niche. [ABSTRACT FROM AUTHOR]

Cerebral malaria (CM) is a life-threatening complication of the malaria disease caused by Plasmodium falciparum infection and is responsible for the death of half a million people annually. The molecular pathogenesis underlying CM in humans is not completely understood, although sequestration of infected erythrocytes in cerebral microvessels is thought to play a major role. In contrast, experimental cerebral malaria (ECM) models in mice have been thought to be distinct from human CM, and are mainly caused by inflammatory mediators. Here, to understand the spatial distribution and the potential sequestration of parasites in the whole-brain microvessels during a mouse model of ECM, we utilized the new tissue-clearing method CUBIC (Clear, Unobstructed, Brain/Body Imaging Cocktails and Computational analysis) with light-sheet fluorescent microscopy (LSFM), and reconstructed images in three dimensions (3D). We demonstrated significantly greater accumulation of Plasmodium berghei ANKA (Pb ANKA) parasites in the olfactory bulb (OB) of mice, compared with the other parts of the brain, including the cerebral cortex, cerebellum and brainstem. Furthermore, we show that Pb ANKA parasites preferentially accumulate in the brainstem when the OB is surgically removed. This study therefore not only highlights a successful application of CUBIC tissue-clearing technology to visualize the whole brain and its microvessels during ECM, but it also shows CUBIC's future potential for visualizing pathological events in the whole ECM brain at the cellular level, an achievement that would greatly advance our understanding of human cerebral malaria. [ABSTRACT FROM AUTHOR]

Langerhans cells (LCs) are mainly present in the epidermis and mucosa, and have important roles during skin infection. Migration of LCs to lymph nodes is essential for antigen presentation. However, due to the difficulties in isolating and culturing human LCs, it is not fully understood how LCs move and interact with the extracellular matrix (ECM) through their adhesion molecules such as integrin, during the immune responses. In this study, we aimed to investigate LC motility, cell shape and the role of integrin under inflammatory conditions using monocyte‐derived Langerhans cells (moLCs) as a model. As a result, lipopolysaccharide (LPS) stimulation increased adhesion on fibronectin coated substrate and integrin α5 expression in moLCs. Time‐lapse imaging of moLCs revealed that stimulation with LPS elongated cell shape, whilst decreasing their motility. Additionally, this decrease in motility was not observed when pre‐treated with a neutralising antibody targeting integrin α5. Together, our data suggested that activation of LCs decreases their motility by promoting integrin α5 expression to enhance their affinity to the fibronectin, which may contribute to their migration during inflammation. [ABSTRACT FROM AUTHOR]

This article discusses a rare case of co-occurrence of bullous pemphigoid (BP) and mucosal-dominant pemphigus vulgaris (MDPV) in a Japanese man. The patient presented with tense blisters and erosions on the skin and mucosal lesions on the lips, tongue, and buccal mucosa. Initially, the mucosal lesions were attributed to BP, but further analysis revealed that they were caused by pathogenic anti-desmoglein 3 (Dsg3) antibodies. The article emphasizes the importance of confirming the pathogenicity of anti-Dsg antibodies and highlights the usefulness of a bead aggregation assay in monitoring disease activity and selecting appropriate treatments. [Extracted from the article]