High-grade gliomas, particularly glioblastoma, are covered by authors led by Suzy Baker and Paul Mischel. The review by Rankin, Zhu and Baker focuses on mouse models of high-grade gliomas, describing their generation and diversity and how they can be used for assessing: (i) the incremental integration of specific cell pathway abnormalities in different cell types at different times during central nervous system development;

(ii) the acquisition of further genomic abnormalities during disease HCS assay progression; (iii) glioma biology with respect to interactions between tumour and stroma; and (iv) preclinical testing. Masui, Cloughesy and Mischel describe how, for adult glioblastoma, established genetic abnormalities have already been translated into assays with diagnostic or therapeutic utility and how emerging biological concepts about the molecular subclassification of the disease might yield more. There is an emphasis on the complexities of the glioma cell’s molecular circuitry and how targeted therapies need to take account of this if they are to be effective. Neuropathologists should take a leading role in the evolution of brain tumour diagnostics, assuming responsibility for the presentation of histopathological and molecular data, alongside clinical recommendations,

in a comprehensive diagnostic report. To do this, they will require knowledge of how advances in our selleck screening library understanding of brain tumour biology can be translated into robust and pertinent assays. These reviews provide some of that knowledge, and I hope that you enjoy reading them as much as I

have. “
“This chapter contains sections titled: How to Use this Atlas Specimen Aspartate Derivation and Preparation Recommended Reading Internet Sources “
“Diagnostic Pathology – Neuropathology’ is one of 23 titles in the Amirsys Publishing ‘Diagnostic Pathology’ series. Titles range from ‘Normal Histology’ through the various organ systems, to specialized titles such as ‘Transplant Pathology’ and ‘Familial Cancer Syndromes’. The volume dedicated to neuropathology includes various well-known international names in the field of neuropathology, including the late Bernd Scheithauer, to whose memory the book is dedicated. The book is divided into three separate parts covering 139 specific diagnoses. Part 1 covers neoplastic lesions and is subdivided into five sections: brain and spinal cord, sellar region, meninges, cranial, spinal and peripheral nerves, and familial tumour syndromes. Part 2 covers non-neoplastic pathology and is subdivided into four sections: benign cysts, infections, inflammatory and reactive lesions, vascular diseases and cortical dysplasia. Part 3 is a short reference section containing an antibody index and molecular factors index. All of the chapters have a similar layout.

However, minor, albeit significant, changes were observed in the percentage of pre-marginal zone, marginal zone, T2 and B1 B cells. Although the

meaning of this observation is presently unclear, this finding suggests that Treg cells may also contribute to maintaining overall homeostasis of splenic B-cell populations. In addition to disrupting Treg-cell activity SAHA HDAC ic50 with administration of anti-GITR mAb, a large number of studies have examined the role of Treg cells in immune responses using a depleting anti-CD25 mAb.51–55 High-dose anti-CD25 treatment deletes most but not all Treg cells, because a minority of Foxp3+ T cells in secondary lymphoid tissues are CD25.1–47,52 BALB/c mice were injected with 250 μg of either anti-CD25 mAb (PC61) or control rIgG on days −2, +1, +5 with injections continued twice weekly until the mice were killed. Mice were immunized with SRBC on day 0 and splenic GCs were examined on days 8–24. As opposed to continuous anti-GITR mAb treatment, extended anti-CD25 mAb treatment did not lead to mortality, probably because of the protective activity of residual CD25− Treg cells. Similar to mice treated with anti-GITR mAb, however, injection of anti-CD25 mAb resulted in a larger total GC response and a progressive imbalance

of switched to IgM+ GC B cells (see Supplementary material, Fig. S2). Regardless of the means by which Treg-cell activity was inactivated, therefore, GC responses were markedly dysregulated. Although both anti-GITR mAb and anti-CD25 mAb treatments are well BTK inhibitor accepted methods for inactivating Treg cells in vivo, it is possible that the mAbs may have direct effects on GC B cells. To rule out this possibility, GC B cells were tested at days 8, 12 and 18 post-immunization for expression of GITR and CD25. As shown in Supplementary material, Fig. S3, GC B cells were negative for these molecules at all time-points tested.

To ensure that Treg-cell control of GC responses was strain independent, C57BL/6 mice were similarly challenged with SRBC and treated with either anti-GITR mAb or control Branched chain aminotransferase rIgG (Fig. 2). Even though control-treated C57BL/6 mice generated a smaller splenic GC reaction after SRBC immunization compared with BALB/c mice (Fig. 2a,b), the response was again characterized by a steady ratio of IgM+ to switched B cells at all time-points (Fig. 2c). Importantly, anti-GITR mAb administration resulted in a larger proportion and total number of GC B cells (Fig. 2b), especially at the early time-points, and a disproportionate percentage and number of switched GC B cells throughout the response (Fig. 2c). Similar to findings in BALB/c mice, there was also a significant increase in the percentage of IgG1+ GC B cells at day 8 in anti-GITR mAb compared with rIgG-treated mice (data not shown).

The prevalence of CVID increases with age [5]. It can also be difficult to distinguish developing CVID from delayed maturation of the immune system in so-called transient hypogammaglobulinaemia, which is relatively common especially in younger children [6]. The majority of CVID patients present PD-0332991 cell line with recurrent bacterial infections

of the respiratory tract. In some patients with CVID, ultimately T-lymphocyte function deteriorates as well [7]. Gastrointestinal disease, lymphoproliferative disorders, autoimmune phenomena, and granulomatous inflammation are seen in subgroups of patients; in some patients these precede the recurrent infections [8]. Up to 73% of CVID patients develop chronic structural pulmonary complications. Although the incidence is lower, these pulmonary abnormalities are already

present in children with CVID [9, 10]. Patients are treated with life-long replacement of immunoglobulins, but even with adequate immunoglobulin substitution chronic lung disease will develop in the majority of patients [11]. The exact aetiology of CVID is unknown, but causative gene mutations have been reported in a few families, including CD19 [12], CD20, B cell activating factor receptor (BAFF-R), the inducible costimulator (ICOS), and CD80 genes [13] and around 10% of CVID mTOR inhibitor patients show disease-modifying heterozygous amino acid substitutions in the transmembrane and calcium-modulating cyclophilin ligand (CAML) interactor (TACI) [13, 14]. Immunophenotyping of lymphocyte subpopulations is an important tool in the diagnosis GBA3 of immunological and haematological diseases. When absolute numbers of lymphocyte subpopulations

fall outside predetermined reference ranges, this indicates possible disease. Lymphocyte subpopulations are also increasingly used to classify patients with CVID into subgroups with different clinical prognosis according to the composition of their B-lymphocyte compartment [15–17]. These classifications were mainly developed with data obtained in adults, however. Because of their maturing immune system, these classifications may not be equally applicable in children: age-matched reference values that have been determined for B-lymphocyte subpopulations in children show great changes in the composition of the B-lymphocyte compartment during development [18–26]. Not only do the absolute number of CD19+ B-lymphocytes show a massive expansion shortly after birth, the relative distribution between naive (CD19+CD27-IgD+), natural effector (CD19+CD27+IgD+), switched memory (CD19+CD27+IgD-) [18, 20, 23, 24, 26], and CD21low (CD19+CD21lowCD38low) B-lymphocytes [24], as well as class-switched plasmablasts (CD19+CD38+++IgM-) and transitional B cells (CD19+CD38++IgM++) [18] also change significantly with increasing age. The most important shifts in B-lymphocyte subpopulations take place in the first weeks to months after birth, but development continues until adulthood.

31–33 Interestingly, ICCs in the lamina propria respond to ATP but not to muscarinic agonist carbachol, Alectinib manufacturer while ICCs in the detrusor respond to carbachol via M3 receptor, indicating a parasympathetic control of ICCs in the detrusor.34 This implies the two types of ICCs have different functional

roles in the bladder physiology. Spontaneous electrical activity and Ca2+-transients in the ICCs and close structural connections with nerves and SMCs26,35 have suggested that the ICCs may be pacemaking cells of SCs in the bladder. Indeed, the c-Kit tyrosine kinase inhibitor imatinib mesylate inhibited SCs.36,37 However, the frequency of spontaneous Ca2+-transients differed between the ICCs and neighboring SMCs.38 This evidence contradicts the notion that ICCs in the bladder function as pacemakers. ICCs in the detrusor are positive for cyclooxygenase Y-27632 research buy related to prostaglandins synthesis,39,40 and a recent study showed that ICCs in the detrusor have numerous vesicles, indicating a secretory function.41 Therefore, ICCs

may control the SMC activity by releasing a modulator. This is an attractive hypothesis. There are at least three factors that may contribute to changes in the SCs due to SCI or BOO: myogenic alterations, local mediators in the detrusor and urotheliogenic modulation (Table 1). SMCs have spontaneous electrical and contractile activity. However, electrical coupling is normally limited to some neighboring cells, and action potentials may spread through gap junctional intercellular communication.42 In BOO, cell-to-cell communication between primary cultured SMCs of bladders stained with a fluorescent dye was enhanced in SMCs from rats with BOO compared with those from control rats.43 This enhancement was inhibited by a gap-junction inhibitor. Enhanced cell-to-cell communication may, therefore, contribute to the enhanced SCs associated with BOO. The expression of

connexin 43 gap junctions between SMCs is increased in the human bladder with DO mainly due to SCI.44 This implies that intercellular communication between SMCs is enhanced and may result in enhanced SCs in SCI. Alterations in ion channel activity may be involved in the generation of enhanced SCs Montelukast Sodium in BOO. Downregulation of large and small conductance calcium-activated potassium channels and the TREK-1 potassium channel, and upregulation of calcium-activated chloride channels may cause enhanced SCs.45–47 However, there have been contradictory findings, namely, upregulated expression of potassium channels has also been identified in bladders with BOO.22 The reason for this contradiction is unknown. Further studies of alterations to potassium channels are required. There is an intracellular signal transduction mechanism that can increase the contractile ability of SMCs, that is, calcium sensitization that involves the rhoA/rho-kinase pathway.48 The expression of rhoA and rho-kinase was upregulated in obstructed rat bladders.

In addition, systemic cytokine/chemokine responses can be identified in patients with periodontitis [3–5]. Interleukin (IL)-1β, tumour necrosis factor (TNF)-α and IL-6 are principal pro-inflammatory cytokines with pleotropic biological activities on immune and non-immune cells, as well as in osteogenic pathways). IL-8 (CXCL8) is the major neutrophil chemokine, while macrophage chemotactic protein (MCP)-1 (CCL2), a major chemoattractant and maturation signal for macrophages, and regulated upon activation, normal T cell expressed and secreted (RANTES; CCL5) is a member of the IL-8 superfamily of cytokines.

It is a selective attractant for memory T lymphocytes and monocytes. These chemokines have all been detected in the serum of patients with microbial infections [6–10], including periodontitis [11–14]. Selleck Target Selective Inhibitor Library However, chronic stimulation of these biomolecules generally represents dysregulated responses, and is associated frequently with systemic disease sequelae [15–21]. In some cases, particularly with polymicrobial infections at mucosal surfaces, innate immune mechanisms may function exceptionally well to manage surface colonization by commensal opportunistic pathogens and maintain homeostasis [22–25]. Nevertheless, with respect to a number of chronic inflammatory diseases, the interaction between Trichostatin A the challenge (e.g. bacteria) and

the inflammatory and innate immune response can result in collateral damage of the local tissues. Adverse pregnancy outcomes provide a potential example of these ramifications of a dysregulated

host response. Ascending vaginal infections trigger the local production of various inflammatory mediators and matrix metalloproteinases (MMP), resulting in amnionitis that impact placental functions negatively and lead potentially to fetal infection [26–32]. Reports described relationships between the presence of inflammatory mediators in amniotic fluid and uterine contractions and/or birth in humans and non-human primates. Proinflammatory cytokines/chemokines, immunomodulatory and immunosuppressive 4��8C cytokines and prostanoids [e.g. prostaglandin E2 (PGE2)] are produced by the amniotic and decidual membranes and can be found in fetal circulation and amniotic fluid, often associated with premature delivery. Expanding literature supports that the levels of many of these cytokines/chemokines in serum are also reflective of, and potentially contribute to, the risk for premature rupture of membranes (PROM) with preterm labour and delivery [26,32–35]. Consequently, relationships between serum and local cytokine levels and their association with adverse pregnancy outcomes are possible. Periodontitis is a chronic oral infection with polymicrobial biofilms triggering a localized immunoinflammatory lesion.

There was no significant difference in the post-surgical seizure outcome between patients with Palmini type I and type

II cortical dysplasia in the UCLA cohort[70] and in other epilepsy centers.[71] However, some studies reported less favorable outcomes in patients with Palmini type I cortical dysplasia,[72, 73] and other studies reported opposite results,[74] although a significant proportion of these patients also had HS. Such inconsistent results among various studies also appear to be a major problem in elucidating the clinicopathological correlation of cortical dysplasia as being discussed in HS, and may be due, at least in part, to the difference in inclusion and exclusion criteria. Recently a Smoothened Agonist purchase consensus histological classification scheme of FCD was proposed at the initiative BGB324 price of the Task Force on FCD in the ILAE Diagnostic Methods Commission.[56] The major changes from Palmini’s classification to the ILAE classification included separation of “isolated” FCD type I from those associated with other epileptogenic

principal lesions; that is, HS, tumors, vascular malformations, and any other lesion acquired during early life, such as trauma, ischemic injury and encephalitis, and classifying these “associated” counterparts as FCD type III, forming a three-tiered classification system (Table 6). Histological definition PI-1840 of FCD type I was reorganized in the ILAE classification. Another change was also made in the terminology; the term “giant neurons” in Palmini’s classification

is now designated as “hypertrophic neurons” in the ILAE classification, which is defined as large pyramidal neurons resembling those of neocortical layer 5 abnormally located in layers 1, 2, 3 or 4. Hypertrophic neurons can be observed in all types of FCD. Of note, the term “giant cells” refers to large gemistocytic astrocyte-like cells observed in TSC-tubers, which are morphologically identical to BCs observed in FCD type IIb. Although the etiology and pathogenesis of each FCD type are yet to be elucidated, this new classification seems applicable in terms of good interobserver and intraobserver agreement[75] to the future clinicopathological correlation study for evaluating post-surgical seizure outcomes in patients with “isolated” FCD types I and II without any other epileptogenic lesions. One study using ILAE classification demonstrated poorer post-surgical outcomes in patients with FCD type III than in patients with isolated FCD (FCD types I and II).

These results suggest that endogenous

mCRAMP regulates antigen-specific IgG1 production in vivo by suppressing CD4+ T-cell IL-4 expression, although whether this is a direct effect or indirect through another cell type is yet to be determined. mCRAMP is an AMP that is beginning to be appreciated as a potent and important immunomodulatory molecule. PI3K activity While our data begin to elucidate the role of mCRAMP in the adaptive immune response, more information is needed to fully understand its role in the different microenvironments within the host. It is clear that the cell type producing and/or responding to mCRAMP will partially determine the effect observed. Additional studies are needed to fully understand the role of mCRAMP and other AMPs in the adaptive immune

response. C57BL/6 mice were purchased from the Jackson Laboratory. Decitabine purchase Camp-deficient 129/SVJ mice (Camp−/−, KO) were backcrossed to B6 mice for ten generations and identified by PCR analysis as described previously 8. All mice were maintained under pathogen-free conditions and under approved animal protocols from the Institutional Animal Care and Use Committee at the University of Alabama at Birmingham. The 38 amino acid mCRAMP peptide (ISRLAGLLRKGGEKIGEKLKKIGQKIKNFFQKLVPQPE) was synthesized by Alpha Diagnostic Int. (San Antonio, TX, USA) and the lyophilized peptides were resuspended in 0.01% acetic acid to generate 100 μM working stocks, which were stored at −80°C until time of use. B-cell purification and activation was performed as described previously 40. Purified splenic B cells were obtained using a CD43 magnetic P-type ATPase bead depletion strategy (Miltenyi Biotec). B cells (5×104) were cultured in 96-well flat-bottom plates in 200 μL of complete medium (cRPMI). B

cells were stimulated with 20 μg/mL LPS (Sigma-Aldrich), 1 ng/mL recombinant mouse IL-4 (eBioscience), 10 ng/mL recombinant mouse IFN-γ (eBioscience), and/or CD40L-expressing Sf9 cells (a gift from Dr. Virginia Sanders, The Ohio State University) at a B cell-to-Sf9 ratio of 10:1. Culture supernatants were collected and stored at −80°C until further analysis. Flow cytometry and cell sorting was performed as described previously 41. Intracellular staining was performed using the Cytofix/Cytoperm kit (BD Biosciences). FITC-labeled anti-γ1, anti-CD23, anti-Mac-1; PE-labeled anti-CD5, anti-Mac 1, anti-IL-4; APC-labeled anti-B220, PE-Cy7-anti-CD4, PB-anti-B220, PE-anti-IL-4, and PE-rIgG1 isotype antibodies were purchased from BD Pharmingen. Anti-CD21 (clone 7G6) antibody was purified and labeled with PE in our laboratory. Cy5-labeled goat anti-mouse IgM antibody was purchased from Jackson ImmunoResearch. FcR blocker Ab93 was generated in our laboratory 42. Experiments were performed on a FACSCalibur (BD Biosciences), cell sorting using a FACSAria (BD Biosciences), and analysis using FlowJo software (Tree Star). Seven- to nine-wk-old female mice were immunized i.v. with 1×108 heat-killed Streptococcus pneumonia (R36A) or i.p.

We retrospectively reviewed medical records of 637 Korean patients

with onychomycosis between December 2000 and December 2006. We examined six clinical factors to evaluate the effects on the CR, DC and RR: age, sex, clinical type, treatment pattern, presence of diabetes mellitus (DM) and the extent of nail involvement. On the view of the clinical nail appearance and potassium hydroxide (KOH) preparation, we designated the CR, DC and RR. In addition, SB525334 we examined the differences in the CR, DC and RR in terms of the above-mentioned clinical factors. A total of 207 eligible patients were finally analysed. The CR as a whole was 78.3%, the DC was 31.7 ± 18.4 weeks and the RR was 36.0%. There were significant differences in the CR, DC and RR according to the extent of nail involvement. Age

affects the CR and DC, and DM also affects the DC and RR. We found that the extent of nail involvement, age and DM affect the CR, DC and RR of onychomycosis. “
“The following case report describes a patient with acute liver failure who presented in multiple organ failure and required emergency liver Vemurafenib manufacturer transplantation. A complicated postoperative course lead to sepsis which did not respond to conventional anti bacterial therapy. Despite antifungal prophylaxis with an azole invasive candidiasis was diagnosed and the patient was successfully treated with anidulafungin. The difficulties in diagnosis and treatment of invasive fungal infections in this population are highlighted. “
“The production of Secretory Aspartyl Proteases (Sap) is an important virulence factor of Candida albicans. Many studies have shown that a challenge with sub-inhibitory concentrations of antifungals lead species of Candida to the secretion of higher concentrations of Sap. Nevertheless, published studies only reported the secretion of such enzymes by cells growing in planktonic phase, with few mention of biofilms. The present study

evaluated the alterations in the secretion of Sap by C. albicans MRIP grown in biofilms and exposed to sub-inhibitory concentrations of fluconazole. The MICs for fluconazole of seven clinical strains were determined for planktonic cells. Biofilm and planktonic cells were grown in the presence of ½ MIC, ¼ MIC, and no medication (control). The relative metabolic activity, indirectly related to cell loads, were estimated by the absorbance of reduced XTT and the Sap activity was evaluated by bovine albumin test. It was observed that 72 h-old biofilms under the influence of ½ MIC had fewer cells than ¼ MIC and control. The production of Sap was inversely proportional to the cell content, with higher secretion in ½ MIC, followed by ¼ MIC and control. Biofilms of C. albicans challenged by sub-MICs of fluconazole tend to secrete higher quantities of Sap.

, 2005; Liu et al., 2007; Agashe et al., 2009) and multiplex PCR (amplification of two or more gene targets simultaneously; Okazaki et al., 2005; Colmenero et al., 2010; Sharma et al., 2011a, b) have been exploited for EPTB diagnosis. The DNA-PCR is unable to differentiate viable and nonviable organisms, while bacterial MK 2206 mRNA with a mean half-life of 3–5 min is more prone to destruction than the genomic DNA; thus, a positive mRNA signal would indicate the presence of viable organisms (Rana et al., 2011). The mRNA-based reverse transcriptase-PCR (RT-PCR) is a rapid method to differentiate viable and nonviable M. tuberculosis

and has also been used for the diagnosis of EPTB as well as to monitor drug resistance (Eltringham et al., 1999; Rana et al., 2011). Real-time PCR is a novel and robust assay primarily used to quantify the nucleic acid molecules in EPTB specimens (Baba et al., 2008; Rosso et al., 2011). The main advantages of real-time PCR are shortened turnaround time, quantification of bacterial load and automation of the procedure that reduces hands-on time and decreased risk of cross-contamination (Kalantri et al.,

2011; Rosso et al., 2011). During PCR amplification, several inhibitors such as host proteins, blood and even eukaryotic DNA in extrapulmonary specimens are known to interfere Selleckchem AZD6738 with the sensitivity of PCR and give false-negative results (Gan et al., 2002; Haldar et al., 2011; Sun et al., 2011). A multi-step process is often required to eliminate PCR inhibitors and to obtain highly purified DNA. To achieve this, numerous techniques for DNA sample preparation have been recommended such as freeze-boiling, chelex/proteinase K treatment and sequence capture method (Honore-Bouakline et al., 2003). Chakravorty & Tyagi (2005) introduced a novel multi-purpose universal sample processing (USP) technique

using chaotropic property of guanidinium hydrochloride as a principle Liothyronine Sodium component and that can be used for inhibitor-free PCR for both PTB and EPTB specimens. The addition of cetyltrimethylammonium bromide or silica membranes in the DNA purification has also been shown to effectively remove the PCR inhibitors and, hence could improve the PCR sensitivity in EPTB specimens (Böddinghaus et al., 2001; Honore-Bouakline et al., 2003; Rafi et al., 2007). However, the additional purification steps could lead to substantial loss of mycobacterial DNA, and to circumvent this problem, a short-culture augmentation step for 2–3 days has been proposed before performing PCR test (Cheng et al., 2005), which could enhance the mycobacterial load, while concomitantly diluting PCR inhibitors. Recently, Santos et al. (2009) compared nine different DNA extraction systems (seven manual and two automatic) in an experimental model of pleural TB for analysis with real-time PCR.

For intracellular staining click here of GM-CSF, isolated leukocytes were incubated with 50 ng/mL PMA, 500 ng/mL ionomycin, Golgi-Plug (1 μL/mL) containing brefeldin A in RPMI-1640 at 37°C for 4 h. Thereafter,

cells were stained with rat antimouse CD4-FITC, rat antimouse CD45-V450, fixed and permeabilized with Cytofix/Cytoperm (BD), and stained with rat antimouse GM-CSF-PE (BD). Apoptotic and dead CD4+ T cells were detected by staining with 7-AAD and CD4-allophycocyanin. Fas expression on CD4+ T cells was analyzed by staining with hamster antimouse Fas-PE and CD4-FITC. Controls were stained with isotype-matched control antibodies. All antibodies and reagents were obtained from BD Biosciences (Heidelberg, Germany) unless otherwise mentioned. Flow cytometry was performed on a FACScan (BD Biosciences), and the data were analyzed with WinMDI or Cell Quest software. Primary astrocytes this website were isolated from 1- to 2-day-old newborn mice and cultured as published before [43]. To obtain pure astrocytes, cells were harvested from astrocyte cultures and stained with rat antimouse CD11b-PE. Pure astrocytes (CD11b−) were then separated from CD11b+ microglia with a FACSVantage cell sorter (BD). Neuronal cultures were obtained according to Lenz et al. [44]

with slight modifications. Briefly, pregnant female mice were sacrificed by cervical dislocation at gestational day 18.5, and dissociated cells of each embryonic brain were cultivated in flasks coated with poly-L-lysine in Neurobasal medium supplemented with B27 (Invitrogen) and 500 μM L-glutamine (Gibco). The purity of cultures for neurons was ≥98%, as determined by immunofluorescence

staining for ZD1839 mouse neuron-specific class III β-tubulin. DNA was isolated from sorted astrocytes and microglia, respectively, as well as from cultured neurons using a DNA isolation kit (Qiagen, Germany). For the detection of FasL expressed on the surface of astrocytes, mixed astrocyte/microglia cultures were stained with mouse antimouse FasL-PE and CD11b-FITC. Controls were stained with isotype-matched control antibodies. For histology on paraffin sections, mice anesthetized with methoxyflurane were perfused with 0.1 M PBS followed by 4% paraformaldehyde in PBS, spinal cords were processed and stained with hemalum and eosin, cresyl violet, and luxol fast blue. In addition, paraffin sections were used for immunohistochemical demonstration of GFAP, neurofilament, Mac3, and CD3 (Serotec, Düsseldorf, Germany) by an ABC protocol as described [45]. Total mRNA was isolated from the spinal cords of nonimmunized and MOG35–55- immunized mice (RNeasy kit, Qiagen, Germany) at day 15 and day 22 p.i., respectively. SuperScript reverse transcriptase kit with oligo (dT) primers (Invitrogen, Germany) was used to generate cDNA from total mRNA.