Mouse Myelin Basic Protein Research Articles

Ginsenoside Rg1 protects the blood-brain barrier and myelin sheath to prevent postoperative cognitive dysfunction in aged mice.

In this study, the postoperative cognitive dysfunction (POCD) mouse model was established to observe the changes in inflammation, blood-brain barrier permeability, and myelin sheath, and we explore the effect of ginsenoside Rg1 pretreatment on improving POCD syndrome. The POCD model of 15- to 18-month-old mice was carried out with internal fixation of tibial fractures under isoflurane anesthesia. Pretreatment was performed by continuous intraperitoneal injection of ginsenoside Rg1(40 mg/kg/day) for 14 days before surgery. The cognitive function was detected by the Morris water maze. The contents of interleukin-1β and tumor necrosis factor-α in the hippocampus, cortex, and serum were detected by ELISA. The permeability of blood-brain barrier was observed by Evans blue. The mRNA levels and protein expression levels of 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase), myelin basic protein (MBP), beta-catenin, and cyclin D1 in the hippocampus were analyzed by quantitative PCR and western blotting. The protein expression levels of ZO-1 and Wnt1 in the hippocampus were analyzed by western blotting. Finally, the localizations of CNPase and MBP in the hippocampus were detected by immunofluorescence. Ginsenoside Rg1 can prevent POCD, peripheral and central inflammation, and blood-brain barrier leakage, and reverse the downregulation of ZO-1, CNPase, MBP, and Wnt pathway-related molecules in aged mice. Preclinical studies suggest that ginsenoside Rg1 improves postoperative cognitive function in aged mice by protecting the blood-brain barrier and myelin sheath, and its specific mechanism may be related to the Wnt/β-catenin pathway.

C-Abl-induced Olig2 phosphorylation regulates the proliferation of oligodendrocyte precursor cells.

Oligodendrocytes (OLs), the myelinating cells in the central nervous system (CNS), are differentiated from OL progenitor cells (OPCs). The proliferation of existing OPCs is indispensable for myelination during CNS development and remyelination in response to demyelination stimulation. The transcription factor Olig2 is required for the specification of OLs and is expressed in the OL lineage. However, the post-translational modification of Olig2 in the proliferation of OPCs is poorly understood. Herein, we identified that c-Abl directly phosphorylates Olig2 mainly at the Tyr137 site, and that Olig2 phosphorylation is essential for OPC proliferation. The expression levels of c-Abl gradually decreased with brain development; moreover, c-Abl was highly expressed in OPCs. OL-specific c-Abl knockout at the developmental stage led to an insufficient proliferation of OPCs, a decreased expression of myelin-related genes, and myelination retardation. Accordingly, a c-Abl-specific kinase inhibitor suppressed OPC proliferation in vitro. Furthermore, we observed that OL-specific c-Abl knockout reduced OPC proliferation and remyelination in a cuprizone model of demyelination. In addition, we found that nilotinib, a clinically used c-Abl inhibitor, decreased the expression of myelin basic protein (Mbp) and motor coordination in mice, indicating a neurological side effect of a long-term administration of the c-Abl inhibitor. Thus, we identified the important role of c-Abl in OLs during developmental myelination and remyelination in a disease model.

Abstract P772: Complement C3a Receptor Deletion is Neuroprotective in a Murine Model of Vascular Cognitive Impairment

Background: Chronic cerebral hypoperfusion (CCH) leads to vascular contributions to cognitive impairment and dementia (VCID). We and others have reported that either the genetic deficiency of complement C3a receptor (C3aR) or its pharmacological inhibition in rodents protect against cerebral ischemia. Hypothesis: CCH augments VCID via overactivation of C3aR in brain. Methods: Male C3aR knockout (C3aR -/- ) and wild-type (C3aR +/+ ) mice (Age:12 weeks; N=8-10/gp) were subjected to either VCID with bilateral common carotid artery stenosis (BCAS) or sham surgery. At 4-mo post-BCAS, changes in cerebral blood flow (CBF), hippocampal atrophy (HA), white matter degeneration (WMD) and ventricular size were determined with laser speckle contrast analysis and magnetic resonance imaging. Behavioral outcomes were evaluated using Morris water maze (MWM) and novel object recognition (NOR). Histopathology with Luxol-Fast Blue (LFB) staining, and protein biochemistry with Western blotting were also performed. Results: BCAS resulted in reduced CBF, inducing HA, WMD and ventricle enlargement in both groups compared to their sham controls. These deleterious findings were attenuated in C3aR -/- compared to C3aR +/+ mice. Moreover, the C3aR -/- -BCAS group performed significantly better than C3aR +/+ -BCAS group in MWM and NOR tests; however, mice subjected to BCAS never outperformed their sham-groups. WMD demonstrated by LFB-staining and reduced expression of myelin basic protein and tight junction proteins (ZO-1 and Occludin) in mice subjected to BCAS was mitigated in C3aR -/- as compared to C3aR +/+ mice. C3aR +/+ -BCAS also showed significantly higher expression of C3a in plasma and C3aR in brain lysates compared to their sham-controls. Conclusion: CCH augments C3aR activation leading to poor outcomes in VCID. Therefore, future studies in cell-specific mutant mice are warranted to understand the pathological role of C3aR in VCID.

Bu-Shen-Yi-Sui Capsule, an Herbal Medicine Formula, Promotes Remyelination by Modulating the Molecular Signals via Exosomes in Mice with Experimental Autoimmune Encephalomyelitis.

Multiple sclerosis (MS) is a common inflammatory demyelinating disorder of the central nervous system. Bu-shen-yi-sui capsule (BSYSC) could significantly reduce the relapse rate, prevent the progression of MS, and enhance remyelination following neurological injury in experimental autoimmune encephalomyelitis (EAE), an established model of MS; however, the mechanism underlying the effect of BSYSC on remyelination has not been well elucidated. This study showed that exosomes carrying biological information are involved in the pathological process of MS and that modified exosomes can promote remyelination by modulating related proteins and microRNAs (miRs). Here, the mechanism by which BSYSC promoted remyelination via exosome-mediated molecular signals was investigated in EAE mice and oligodendrocyte progenitor cells (OPCs) in vitro. The results showed that BSYSC treatment significantly improved the body weight and clinical scores of EAE mice, alleviated inflammatory infiltration and nerve fiber injury, protected the ultrastructural integrity of the myelin sheath, and significantly increased the expression of myelin basic protein (MBP) in EAE mice. In an in vitro OPC study, BSYSC-containing serum, especially 20% BSYSC, promoted the proliferation and migration of OPCs and induced OPCs to differentiate into mature oligodendrocytes that expressed MBP. Furthermore, BSYSC treatment regulated the expression of neuropilin- (NRP-) 1 and GTX, downregulated the expression of miR-16, let-7, miR-15, miR-98, miR-486, and miR-182, and upregulated the level of miR-146 in serum exosomes of EAE mice. In conclusion, these results suggested that BSYSC has a neuroprotective effect and facilitates remyelination and that the mechanism underlying the effect of BSYSC on remyelination probably involves regulation of the NRP-1 and GTX proteins and miRs in serum exosomes, which drive promyelination.

Identification of a novel variant of Golli myelin basic protein BG21 in the uniquely neuroprotective white-footed mouse

The myelin basic protein (MBP) gene is a complex gene which codes for several distinct forms of MBP. The various forms of MBP are functionally involved in the development of the nervous system, T-cell regulation, and myelination. Several neurological disorders have been linked to MBP abnormality, further demonstrating its functional significance in the nervous system. The white-footed mouse (Peromyscus leucopus) exhibits profound neuroprotective characteristics, is asymptomatic to various disease-states, and has a lifespan twice that of the house mouse (Mus musculus). We used M. musculus mouse MBP as a reference to explore MBP in P. leucopus mice. Through genetic and downstream proteomic data analysis, we identified a novel variant of the BG21 isoform of MBP in P. leucopus mice. Variation in this isoform is present at the genetic level between the two species of mice. Our results show differences within the open reading frame of the transcripts accompanied by corresponding differences in protein structure prediction. These data introduce the potential of MBP variation as one of many causal variables contributing to the unique presentation of enhanced neuroprotection and longevity in P. leucopus mice.

Myotube-derived factor promotes oligodendrocyte precursor cell proliferation

Muscle cells secrete numerous molecules that function as endocrine hormones and regulate the functions of distant organs. Myelination in the central nervous system (CNS) is regulated by peripheral hormones. However, the effects of muscle-derived molecules on myelination have not been sufficiently analyzed. In this study, we show that muscle-releasing factors promote proliferation of oligodendrocyte precursor cells (OPCs), which is an element of myelination process. Supernatants of mouse myotube cultures stimulated bromodeoxyuridine (BrdU) incorporation into mouse OPCs. Mouse myotube supernatants did not enhance mouse OPC transmigration and myelin basic protein (MBP) expression. RNA sequencing identified candidate genes with hormonal functions that were expressed in mouse myotubes. These data support the possibility that hormonal molecules secreted by myotubes contribute to OPC proliferation and myelination.

ATX-MS-1467 Induces Long-Term Tolerance to Myelin Basic Protein in (DR2 × Ob1)F1 Mice by Induction of IL-10-Secreting iTregs.

IntroductionAntigen-specific immunotherapy could provide a targeted approach for the treatment of multiple sclerosis that removes the need for broad-acting immunomodulatory drugs. ATX-MS-1467 is a mixture of four peptides identified as the main immune-dominant disease-associated T-cell epitopes in myelin basic protein (MBP), an autoimmune target for activated autoreactive T cells in multiple sclerosis. Previous animal studies have shown that ATX-MS-1467 treatment prevented the worsening of signs of disease in experimental autoimmune encephalitis (EAE) in the humanized (DR2 × Ob1)F1 mouse in a dose-dependent fashion.Methods and ResultsOur study extends these observations to show that subcutaneous treatment with 100 µg of ATX-MS-1467 after induction of EAE in the same mouse model reversed established clinical disability (p < 0.0001) and histological markers of inflammation and demyelination (p < 0.001) compared with vehicle-treated animals; furthermore, in longitudinal magnetic resonance imaging analyses, disruption of blood–brain barrier integrity was reversed, compared with vehicle-treated animals (p < 0.05). Chronic treatment with ATX-MS-1467 was associated with an enduring shift from a pro-inflammatory to a tolerogenic state in the periphery, as shown by an increase in interleukin 10 secretion, relative to interleukin 2, interleukin 17 and interferon γ, a decrease in splenocyte proliferation and an increase in interleukin 10+ Foxp3− T cells in the spleen.ConclusionOur results suggest that ATX-MS-1467 can induce splenic iTregs and long-term tolerance to MBP with the potential to partially reverse the pathology of multiple sclerosis, particularly during the early stages of the disease.FundingEMD Serono, Inc., a business of Merck KGaA.

Viral-mediated oligodendroglial alpha-synuclein expression models multiple system atrophy.

MSA is a fatal neurodegenerative disorder characterized by a combination of autonomic dysfunction, cerebellar ataxia, and l-dopa unresponsive parkinsonism. The hallmark of MSA is the accumulation of α-synuclein, forming cytoplasmic inclusions in oligodendrocytes. Adeno-associated viruses allow efficient targeting of disease-associated genes in selected cellular ensembles and have proven efficient for the neuronal overexpression of α-synuclein in the substantia nigra in the context of PD. We aimed to develop viral-based models of MSA. Chimeric viral vectors expressing either human wild-type α-synuclein or green fluorescent protein under the control of mouse myelin basic protein were injected in the striatum of rats and monkeys. Rats underwent a longitudinal motor assessment before histopathological analysis at 3 and 6 months. Injection of vectors expressing α-synuclein in the striatum resulted in >80% oligodendroglial selectivity in rats and >60% in monkeys. Rats developed progressive motor deficits that were l-dopa unresponsive when assessed at 6 months. Significant loss of dopaminergic neurons occurred at 3 months, further progressing at 6 months, together with a loss of striatal neurons. Prominent α-synuclein accumulation, including phosphorylated and proteinase-K-resistant α-synuclein, was detected in the striatum and substantia nigra. Viral-mediated oligodendroglial expression of α-synuclein allows replicating some of the key features of MSA. This flexible strategy can be used to investigate, in several species, how α-synuclein accumulation in selected oligodendroglial populations contributes to the pathophysiology of MSA and offers a new framework for preclinical validation of therapeutic strategies. © 2017 International Parkinson and Movement Disorder Society.

Myelin Basic Protein Citrullination, a Hallmark of Central Nervous System Demyelination, Assessed by Novel Monoclonal Antibodies in Prion Diseases.

Myelin basic protein (MBP) citrullination by peptidylarginine deiminase (PAD) enzymes leads to incomplete protein-lipid bilayer interactions and vulnerability to proteolytic enzymes, resulting in disorganization of the myelin sheath in the central nervous system. Therefore, citrullinated MBP (citMBP) has been suggested as a hallmark of demyelination, but how citMBP is implicated in prion diseases remains unknown. For the first time, we developed mouse monoclonal anti-citMBP IgG1 (clones 1B8, 1H1, and 3C6) and IgM (clone 3G5) antibodies that recognize human citMBP at its R25, R122, and R130 residues and at its C-terminal region (or the corresponding sites in mouse MBP), respectively. Using a biochemical, immunohistochemical, and immunogold-silver staining for electron microscopy techniques, we found that MBP residue R23 (corresponding to human R25) was specifically citrullinated, was stained as intense punctae in the corpus callosum, the striatum, and the cerebellar white matter, and was predominantly localized in disorganized myelin in the brains of scrapie-infected mice. In the brains of Creutzfeldt-Jakob disease (CJD) patients, MBP residues R25, R122, and R130 were markedly citrullinated and were stained as fibrils and punctae. In particular, white matter regions, such as the midbrain and the medulla, exhibited high levels of citMBP compared to other regions. However, the high levels of citMBP were not correlated with PAD2 expression. The clone 3G5 recognized significantly increased expression of the 18.5kDa and/or 21.5kDa variants of MBP in prion disease. Our findings suggest that significantly increased levels of citMBP may reflect demyelinating neuropathology, and that these newly developed antibodies may be useful for identifying demyelination.

Phosphorylated TandeMBP: A unique protein substrate for protein phosphatase assay

To analyze a variety of protein phosphatases, we developed phosphorylated TandeMBP (P-TandeMBP), in which two different mouse myelin basic protein isoforms were fused in tandem, as a protein phosphatase substrate. P-TandeMBP was prepared efficiently in four steps: (1) phosphorylation of TandeMBP by a protein kinase mixture (Ca2+/calmodulin-dependent protein kinase Iδ, casein kinase 1δ, and extracellular signal-regulated kinase 2); (2) precipitation of both P-TandeMBP and protein kinases to remove ATP, Pi, and ADP; (3) acid extraction of P-TandeMBP with HCl to remove protein kinases; and (4) neutralization of the solution that contains P-TandeMBP with Tris. In combination with the malachite green assay, P-TandeMBP can be used to detect protein phosphatase activity without using radioactive materials. Moreover, P-TandeMBP served as an efficient substrate for PPM family phosphatases (PPM1A, PPM1B, PPM1D, PPM1F, PPM1G, PPM1H, PPM1K, and PPM1M) and PPP family phosphatase PP5. Various phosphatase activities were also detected with high sensitivity in gel filtration fractions from mouse brain using P-TandeMBP. These results indicate that P-TandeMBP might be a powerful tool for the detection of protein phosphatase activities.

Could Intrathymic Injection of Myelin Basic Protein Suppress Inflammatory Response After Co-culture of T Lymphocytes and BV-2 Microglia Cells?

Background:The interaction between activated microglia and T lymphocytes can yield abundant pro-inflammatory cytokines. Our previous study proved that thymus immune tolerance could alleviate the inflammatory response. This study aimed to investigate whether intrathymic injection of myelin basic protein (MBP) in mice could suppress the inflammatory response after co-culture of T lymphocytes and BV-2 microglia cells.Methods:Totally, 72 male C57BL/6 mice were randomly assigned to three groups (n = 24 in each): Group A: intrathymic injection of 100 μl MBP (1 mg/ml); Group B: intrathymic injection of 100 μl phosphate-buffered saline (PBS); and Group C: sham operation group. Every eight mice in each group were sacrificed to obtain the spleen at postoperative days 3, 7, and 14, respectively. T lymphocytes those were extracted and purified from the spleens were then co-cultured with activated BV-2 microglia cells at a proportion of 1:2 in the medium containing MBP for 3 days. After identified the T lymphocytes by CD3, surface antigens of T lymphocytes (CD4, CD8, CD152, and CD154) and BV-2 microglia cells (CD45 and CD54) were detected by flow cytometry. The expressions of pro-inflammatory factors of BV-2 microglia cells (interleukin [IL]-1β, tumor necrosis factor-α [TNF-α], and inducible nitric oxide synthase [iNOS]) were detected by quantitative real-time polymerase chain reaction (PCR). One-way analysis of variance (ANOVA) and the least significant difference test were used for data analysis.Results:The levels of CD152 in Group A showed an upward trend from the 3rd to 7th day, with a downward trend from the 7th to 14th day (20.12 ± 0.71%, 30.71 ± 1.14%, 13.50 ± 0.71% at postoperative days 3, 7, and 14, respectively, P < 0.05). The levels of CD154 in Group A showed a downward trend from the 3rd to 7th day, with an upward trend from the 7th to 14th day (10.00 ± 0.23%, 5.28 ± 0.69%, 14.67 ± 2.71% at postoperative days 3, 7, and 14, respectively, P < 0.05). The ratio of CD4+/CD8 + T in Group A showed a downward trend from the 3rd to 7th day, with the minimum at postoperative day 7, then an upward trend from the 7th to 14th day (P < 0.05). Meanwhile, the levels of CD45 and CD54 in Group A were found as the same trend as the ratio of CD4+/CD8 + T (CD45: 83.39 ± 2.56%, 82.74 ± 2.09%, 87.56 ± 2.11%; CD54: 3.80 ± 0.24%, 0.94 ± 0.40%, 3.41 ± 0.33% at postoperative days 3, 7, and 14, respectively, P < 0.05). The expressions of TNF-α, IL-1β, and iNOS in Group A were significantly lower than those in Groups B and C, and the values at postoperative day 7 were the lowest compared with those at postoperative days 3 and 14 (P < 0.05). No significant difference was found between Groups B and C.Conclusions:Intrathymic injection of MBP could suppress the immune reaction that might reduce the secondary immune injury of brain tissue induced by an inflammatory response.

Hes3 expression in the adult mouse brain is regulated during demyelination and remyelination

Hes3 is a component of the STAT3-Ser/Hes3 Signaling Axis controlling the growth and survival of neural stem cells and other plastic cells. Pharmacological activation of this pathway promotes neuronal rescue and behavioral recovery in models of ischemic stroke and Parkinson's disease. Here we provide initial observations implicating Hes3 in the cuprizone model of demyelination and remyelination. We focus on the subpial motor cortex of mice because we detected high Hes3 expression. This area is of interest as it is impacted both in human demyelinating diseases and in the cuprizone model. We report that Hes3 expression is reduced at peak demyelination and is partially restored within 1 week after cuprizone withdrawal. This raises the possibility of Hes3 involvement in demyelination/remyelination that may warrant additional research. Supporting a possible role of Hes3 in the maintenance of oligodendrocyte markers, a Hes3 null mouse strain shows lower levels of myelin basic protein in undamaged adult mice, compared to wild-type controls. We also present a novel method for culturing the established oligodendrocyte progenitor cell line oli-neu in a manner that maintains Hes3 expression as well as its self-renewal and differentiation potential, offering an experimental tool to study Hes3. Based upon this approach, we identify a Janus kinase inhibitor and dbcAMP as powerful inducers of Hes3 gene expression. We provide a new biomarker and cell culture method that may be of interest in demyelination/remyelination research.

Different isoforms of deiminated myelin basic protein have different adsorption capacities to the myelin lipids

Myelin basic protein (MBP) is one of the most abundant proteins of the mammalian myelin sheath and plays a structural role in maintaining the stability of myelin. This lipid-rich membrane is essential for proper nerve conduction and the maintenance of neuronal health. MBP is localized on the cytoplasmic side of the membrane. There are 4 isoforms of MBP in the myelin sheath [1]. The mouse MBP transcription unit has been shown to contain 11 exons [2]. The gene contains three different transcription start sites that regulate production of the different MBP-related products [3]. The most downstream transcription start site 3 controls the classic MBP isoforms, with molecular sizes ranging from 14 to 21 kDa. In addition, each protein isoform can exist in the form of several different charge isomers. Alteration of MBP cationicity may represent a regulatory mechanism for normal myelin assembly or a degradative mechanism in demyelinating disorders [4]. MBP shows extensive posttranslational modifications, including deimination of arginine residues. The fractions isolated by cation exchange chromatography on a CM52 column [5] have been named C1, C2, C3, ... , C8. The first big peak to elute is the least cationic isomer, i.e., MBP C8, which is further divided in two subfractions C8a and C8b [6]. It has been shown that fraction C8b contains only MBP, but the main component of fraction C8a has been identified as stathmin [6], which is a microtubule-destabilizing protein; it prevents healing in the demyelinating brain [7]. Deiminated MBP is structurally less ordered and more susceptible to proteolytic attack than the native form. The reduction in cationicity of deiminated MBP impedes membrane assembly and exposes an immunodominant epitope in the membrane-bound protein to proteases [5]. This exposure may then cause the deiminated, highly immunogenic epitope to be released, priming the innate cells of the CNS [4]. In a previous study, we have found that citrullination (i.e., deimination of arginine residues) of myelin basic protein leads to abnormal formation of the myelin membrane [8]. The citrullinated peptide 45-89 released from modified deiminated MBP activates microglial cells and subsequent generation of nitric oxide induces damage of oligodendrocytes, which may be a first step in the initiation of demyelinating disorders [9].

Age-dependent redistribution and hypersialylation of the central myelin paranodal loop membrane protein Opalin in the mouse brain

Opalin/Tmem10 is a myelin-associated sialylglycoprotein that is specific to only the mammalian central nervous system. However, little is known about the properties or function of this protein. Here, we analyzed the expression and glycosylation patterns of Opalin in the postnatal mouse brain. Immunolocalization patterns of Opalin were similar to those of myelin basic protein in juvenile and adolescent mice. On the other hand, in the adult mouse brain, decreasing immunoreactivity for Opalin was observed in the hindbrain region especially in the cerebellar white matter compared with the corpus callosum in the forebrain. In addition, Opalin showed increasing molecular size with mouse aging. This age-dependent increase in Opalin molecular weight was mainly owing to hypersialylation of O-glycans. These results indicate that the regional redistribution and the degree of sialylation of Opalin protein are age-dependently regulated in mouse brains.

Myelin Basic Protein Cleaves Cell Adhesion Molecule L1 and Promotes Neuritogenesis and Cell Survival

The cell adhesion molecule L1 is a Lewis(x)-carrying glycoprotein that plays important roles in the developing and adult nervous system. Here we show that myelin basic protein (MBP) binds to L1 in a Lewis(x)-dependent manner. Furthermore, we demonstrate that MBP is released by murine cerebellar neurons as a sumoylated dynamin-containing protein upon L1 stimulation and that this MBP cleaves L1 as a serine protease in the L1 extracellular domain at Arg(687) yielding a transmembrane fragment that promotes neurite outgrowth and neuronal survival in cell culture. L1-induced neurite outgrowth and neuronal survival are reduced in MBP-deficient cerebellar neurons and in wild-type cerebellar neurons in the presence of an MBP antibody or L1 peptide containing the MBP cleavage site. Genetic ablation of MBP in shiverer mice and mutagenesis of the proteolytically active site in MBP or of the MBP cleavage site within L1 as well as serine protease inhibitors and an L1 peptide containing the MBP cleavage site abolish generation of the L1 fragment. Our findings provide evidence for novel functions of MBP in the nervous system.

TandeMBP: generation of a unique protein substrate for protein kinase assays

Myelin basic protein (MBP) is one of the major components of central nervous system myelin and has multiple sites for protein phosphorylation. Therefore, it has been widely used as a substrate for in vitro assays of various protein kinases. In this study, to obtain more efficient substrates for protein kinase assays than commercially available MBP from bovine brain, we produced various recombinant MBPs using Escherichia coli expression systems. Three splice isoforms of mouse MBP were expressed in E. coli and successfully purified using a new protocol consisting of HCl extraction, urea treatment and affinity purification with HiTrap Chelating HP column. The recombinant MBP isoforms thus obtained served as more efficient substrates for protein kinases than MBP isolated from bovine brain. To generate an even better substrate for protein kinase assays, we produced a hybrid protein composed of two different MBP isoforms connected in tandem, designated TandeMBP. TandeMBP was readily expressed in E. coli and could be purified by the newly developed simple procedure. TandeMBP was phosphorylated by various Ser/Thr protein kinases more efficiently than the other MBP isoforms. Taken together, TandeMBP will become a powerful tool for in vitro assays to analyse various protein kinase activities.

The role of staphylococcal enterotoxin A in achieving oral tolerance to myelin basic protein in adult mice

Background: Oral tolerance is the biological process explaining the non-responsiveness of gut lymphoid tissue to intestinal content. Our study tested a new approach for the enhancement of oral tolerance to a multiple sclerosis-triggering auto-antigen–myelin basic protein, by its oral administration with the Staphylococcal enterotoxin A.Methods: Immune tolerance thus stimulated was assessed in adult BALB/c mice, by measuring different cytokines from the supernatant of mesenteric lymph nodes cells (IFN-γ, IL-4, IL-10, IL-17, and TGF-β), and in a SJL/E mouse model of experimental autoimmune encephalomyelitis, by evaluating the development of regulatory T cells in mesenteric lymph nodes and the clinical outcome of the intervention.Results: We obtained a significant rise in the levels of IL-10 and TGF-β compared with control and a significant decrease of IFN-γ, IL-4 (p < 0.05). Regulatory T cells were increased compared with control (p < 0.05). These results were attributable both to myelin basic protein and to Staphylococcal enterotoxin A. The clinical outcome of experimental autoimmune encephalomyelitis was influenced only by the administration of myelin basic protein.Conclusion: In our experiment, Staphylococcal enterotoxin A enhanced the immune tolerance to myelin basic protein in the gut mucosa, but had no impact on the clinical evolution of experimental autoimmune encephalomyelitis.

N-terminal Domain of Myelin Basic Protein Inhibits Amyloid β-Protein Fibril Assembly

Accumulation of amyloid β-protein (Aβ) into brain parenchymal plaques and the cerebral vasculature is a pathological feature of Alzheimer disease and related disorders. Aβ peptides readily form β-sheet-containing oligomers and fibrils. Previously, we reported a strong interaction between myelin basic protein (MBP) and Aβ peptides that resulted in potent inhibition of fibril assembly (Hoos, M. D., Ahmed, M., Smith, S. O., and Van Nostrand, W. E. (2007) J. Biol. Chem. 282, 9952-9961; Hoos, M. D., Ahmed, M., Smith, S. O., and Van Nostrand, W. E. (2009) Biochemistry 48, 4720-4727). MBP is recognized as a highly post-translationally modified protein. In the present study, we demonstrate that human MBP purified from either brain or a bacterial recombinant expression system comparably bound to Aβ and inhibited Aβ fibril assembly indicating that post-translational modifications are not required for this activity. We also show that purified mouse brain MBP and recombinantly expressed mouse MBP similarly inhibited Aβ fibril formation. Through a combination of biochemical and ultrastructural techniques, we demonstrate that the binding site for Aβ is located in the N-terminal 64 amino acids of MBP and that a stable peptide (MBP1) comprising these residues was sufficient to inhibit Aβ fibrillogenesis. Under conditions comparable with those used for Aβ, the fibrillar assembly of amylin, another amyloidogenic peptide, was not inhibited by MBP1, although MBP1 still bound to it. This observation suggests that the potent inhibitory effect of MBP on fibril formation is not general to amyloidogenic peptides. Finally, MBP1 could prevent the cytotoxic effects of Aβ in primary cortical neurons. Our findings suggest that inhibition of Aβ fibril assembly by MBP, mediated through its N-terminal domain, could play a role in influencing amyloid formation in Alzheimer disease brain and corresponding mouse models.

Haloperidol activates quiescent oligodendroglia precursor cells in the adult mouse brain

Recent human studies suggest that abnormal development of oligodendrocytes (OLs) is an important component in the pathophysiology of schizophrenia. However, less information is available regarding effects of antipsychotics on OLs' development. In the present study, young adult C57BL/6 mice were given haloperidol (HAL; 2 mg/kg/day) in their drinking water for three or six weeks. At the conclusion of the drug treatment, mice were sacrificed and the numbers of NG2- and Olig2-expressing cells in the brain regions of the corpus callosum, hippocampus and cerebral cortex were quantified. NG2 is a specific marker for oligodendroglia precursor cells (OPCs); Olig2 marks glial progenitors. HAL treatment for three weeks increased the number of NG2-expressing cells in the corpus callosum; HAL treatment for three and six weeks increased the numbers of Olig2-expressing cells in all three brain regions and increased the levels of Olig2 expression in the same brain regions. These results suggest that HAL treatment activates adult OPCs, which divide infrequently under normal conditions but respond to a variety of insulting factors by proliferation and differentiation. However, our further observations showed no changes in the number of mature OLs and the amount of myelin basic protein in HAL-treated mice, suggesting the drug treatment has no effect on the maturation of OLs. In addition, HAL treatment did not increase the numbers of GFAP- and CD68-expressing cells, suggesting that no gliosis and inflammatory responses occurred while the drug activated the quiescent OPCs in adult brain. These results suggest that HAL treatment may target the development of OLs.

Influence of high level of antibodies to myelin basic protein in female mice on the postnatal development and behavioral reactions of the progeny

Physical development, behavioral reactions, and training capacity were studied in the progeny of female BALB/c mice with high levels of antibodies to myelin basic protein. The proposed protocol of immunization ensures high levels of antibodies to myelin basic protein in this mouse strain. High level of antibodies to myelin basic protein in pregnant females causes an increase in the blood level of these antibodies in the progeny. Inhibitory effect of antibodies to myelin basic protein on physical development, training process, and memory in mouse pups was detected.