Oligodendroglial connexin 47 regulates neuroinflammation upon autoimmune demyelination in a novel mouse model of multiple sclerosis.

In multiple sclerosis plaques, oligodendroglial connexin (Cx) 47 constituting main gap junction channels with astroglial Cx43 is persistently lost. As mice with Cx47 single knockout exhibit no demyelination, the roles of Cx47 remain undefined. We aimed to clarify the effects of oligodendroglia-specific Cx47 inducible conditional knockout (icKO) on experimental autoimmune encephalomyelitis (EAE) induced by myelin oligodendrocyte glycoprotein peptide (MOG35-55) in PLP/CreERT;Cx47fl/fl mice at 14 d after tamoxifen injection. Cx47 icKO mice demonstrated exacerbation of acute and chronic relapsing EAE with more pronounced demyelination than Cx47 flox (fl)/fl littermates. CD3+ T cells more abundantly infiltrated the spinal cord in Cx47 icKO than in Cx47 fl/fl mice throughout the acute to chronic phases. CXCR3-CCR6+CD4+ and IL17+IFNγ-CD4+ helper T (Th) 17 cells isolated from spinal cord and brain tissues were significantly increased in Cx47 icKO mice compared with Cx47 fl/fl mice, while MOG35-55-specific proliferation and proinflammatory cytokine production of splenocytes were unaltered. Microarray analysis of isolated microglia revealed stronger microglial activation toward proinflammatory and injury-response phenotypes with increased expressions of chemokines that can attract Th17 cells, including Ccl2, Ccl3, Ccl4, Ccl7, and Ccl8, in Cx47 icKO mice compared with Cx47 fl/fl mice. In Cx47 icKO mice, NOS2+ and MHC class II+ microglia were more enriched immunohistochemically, and A1-specific astroglial gene expressions and astroglia immunostained for C3, a representative A1 astrocyte marker, were significantly increased at the acute phase, compared with Cx47 fl/fl mice. These findings suggest that oligodendroglia-specific Cx47 ablation induces severe inflammation upon autoimmune demyelination, underscoring a critical role for Cx47 in regulating neuroinflammation.

Brain gray matter astroglia-specific connexin 43 ablation attenuates spinal cord inflammatory demyelination.

BACKGROUND: Brain astroglia are activated preceding the onset of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). We characterized the effects of brain astroglia on spinal cord inflammation, focusing on astroglial connexin (Cx)43, because we recently reported that Cx43 has a critical role in regulating neuroinflammation. METHODS: Because glutamate aspartate transporter (GLAST)+ astroglia are enriched in the brain gray matter, we generated Cx43fl/fl;GLAST-CreERT2/+ mice that were brain gray matter astroglia-specific Cx43 conditional knockouts (Cx43 icKO). EAE was induced by immunization with myelin oligodendroglia glycoprotein (MOG) 35-55 peptide 10 days after tamoxifen injection. Cx43fl/fl mice were used as controls. RESULTS: Acute and chronic EAE signs were significantly milder in Cx43 icKO mice than in controls whereas splenocyte MOG-specific responses were unaltered. Histologically, Cx43 icKO mice showed significantly less demyelination and fewer CD45+ infiltrating immunocytes, including F4/80+ macrophages, and Iba1+ microglia in the spinal cord than controls. Microarray analysis of the whole cerebellum revealed marked upregulation of anti-inflammatory A2-specific astroglia gene sets in the pre-immunized phase and decreased proinflammatory A1-specific and pan-reactive astroglial gene expression in the onset phase in Cx43 icKO mice compared with controls. Astroglia expressing C3, a representative A1 marker, were significantly decreased in the cerebrum, cerebellum, and spinal cord of Cx43 icKO mice compared with controls in the peak phase. Isolated Cx43 icKO spinal microglia showed more anti-inflammatory and less proinflammatory gene expression than control microglia in the pre-immunized phase. In particular, microglial expression of Ccl2, Ccl5, Ccl7, and Ccl8 in the pre-immunized phase and of Cxcl9 at the peak phase was lower in Cx43 icKO than in controls. Spinal microglia circularity was significantly lower in Cx43 icKO than in controls in the peak phase. Significantly lower interleukin (IL)-6, interferon-γ, and IL-10 levels were present in cerebrospinal fluid from Cx43 icKO mice in the onset phase compared with controls. CONCLUSIONS: The ablation of Cx43 in brain gray matter astroglia attenuates EAE by promoting astroglia toward an anti-inflammatory phenotype and suppressing proinflammatory activation of spinal microglia partly through depressed cerebrospinal fluid proinflammatory cytokine/chemokine levels. Brain astroglial Cx43 might be a novel therapeutic target for MS.

Early postnatal allergic airway inflammation induces dystrophic microglia leading to excitatory postsynaptic surplus and autism-like behavior.

Microglia play key roles in synaptic pruning, which primarily occurs from the postnatal period to adolescence. Synaptic pruning is essential for normal brain development and its impairment is implicated in neuropsychiatric developmental diseases such as autism spectrum disorders (ASD). Recent epidemiological surveys reported a strong link between ASD and atopic/allergic diseases. However, few studies have experimentally investigated the relationship between allergy and ASD-like manifestations, particularly in the early postnatal period, when allergic disorders occur frequently. Therefore, we aimed to characterize how allergic inflammation in the early postnatal period influences microglia and behavior using mouse models of short- and long-term airway allergy. Male mice were immunized by an intraperitoneal injection of aluminum hydroxide and ovalbumin (OVA) or phosphate-buffered saline (control) on postnatal days (P) 3, 7, and 11, followed by intranasal challenge with OVA or phosphate-buffered saline solution twice a week until P30 or P70. In the hippocampus, Iba-1-positive areas, the size of Iba-1-positive microglial cell bodies, and the ramification index of microglia by Sholl analysis were significantly smaller in the OVA group than in the control group on P30 and P70, although Iba-1-positive microglia numbers did not differ significantly between the two groups. In Iba-1-positive cells, postsynaptic density protein 95 (PSD95)-occupied areas and CD68-occupied areas were significantly decreased on P30 and P70, respectively, in the OVA group compared with the control group. Immunoblotting using hippocampal tissues demonstrated that amounts of PSD95, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor 2, and N-methyl-D-aspartate (NMDA) receptor 2B were significantly increased in the OVA group compared with the control group on P70, and a similar increasing trend for PSD95 was observed on P30. Neurogenesis was not significantly different between the two groups on P30 or P70 by doublecortin immunohistochemistry. The social preference index was significantly lower in the three chamber test and the number of buried marbles was significantly higher in the OVA group than in the control group on P70 but not on P30, whereas locomotion and anxiety were not different between the two groups. Compared with the control group, serum basal corticosterone levels were significantly elevated and hippocampal glucocorticoid receptor (GR) amounts and nuclear GR translocation in microglia, but not in neurons or astrocytes, were significantly decreased in the OVA group on P70 but not on P30. Gene set enrichment analysis of isolated microglia revealed that genes related to immune responses including Toll-like receptor signaling and chemokine signaling pathways, senescence, and glucocorticoid signaling were significantly upregulated in the OVA group compared with the control group on P30 and P70. These findings suggest that early postnatal allergic airway inflammation induces dystrophic microglia that exhibit defective synaptic pruning upon short- and long-term allergen exposure. Furthermore, long-term allergen exposure induced excitatory postsynaptic surplus and ASD-like behavior. Hypothalamo-pituitary-adrenal axis activation and the compensatory downregulation of microglial GR during long-term allergic airway inflammation may also facilitate these changes.

Comparison of Maximum Mouth Opening Following Mandibular Bilateral Sagittal Splitting Ramus osteotomies in Class III Females Using Two Different Osteosynthesis Methods.

BACKGROUND: Limitation of mouth opening is a common complaint following orthognathic surgery. OBJECTIVES: This investigation reports on the progress of maximum incisal opening following orthognathic surgery and compares the impact of two different osteosynthesis methods on mouth opening in female patients with Class III dentofacial deformities. METHODS: Forty cases of skeletal class III malocclusion were divided into twenty cases treated using the conventional single miniplate osteosynthesis method (P1) and twenty cases using the additional L-shaped miniplate osteosynthesis method (P2). No significant differences in pre-operative clinical status were detected between the P1 and P2 group, and all patients were managed with elastics in the post-operative period. Independent mouth opening exercises were initiated seven days after surgery, and inter-incisal distance was measured as maximum mouth opening (MMO) at 1 week, 2 weeks, and 1, 2, 3, 6 months. Statistical analysis was performed in order to analyse differences in MMO between the P1 and P2 groups (Prism 7 GraphPad software, San Diego, CA). Values of P < .05 were considered to be significant. RESULTS: MMO significantly increased from 2 weeks after surgery in both groups. The MMO of P2 was significantly larger than that of P1 in all experimental periods after surgery. MMO was statistically improved in P2 at 2 months after surgery, while MMO in P1 was significantly smaller than the pre-operative MMO, even at 6 months post-operative. A minimum MMO of 40 mm was achieved by all patients. CONCLUSION: L-shaped miniplate osteosynthesis was more useful for early limitation of mandibular opening improvement than conventional single miniplate osteosynthesis.

Connexin 30 deficiency attenuates A2 astrocyte responses and induces severe neurodegeneration in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride Parkinson's disease animal model.

BACKGROUND: The first pathology observed in Parkinson's disease (PD) is 'dying back' of striatal dopaminergic (DA) terminals. Connexin (Cx)30, an astrocytic gap junction protein, is upregulated in the striatum in PD, but its roles in neurodegeneration remain elusive. We investigated Cx30 function in an acute PD model by administering 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to wild-type (WT) and Cx30 knockout (KO) mice. METHODS: On days 1 and 7 after MPTP administration, we evaluated changes in astrocytic Cx30, Cx43, glial fibrillary acidic protein, and ionised calcium-binding adapter molecule 1 expression by immunostaining and biochemical analysis. Loss of DA neurons was evaluated by tyrosine hydroxylase immunostaining. Gene expression was analysed using A1, A2, pan-reactive astrocyte microarray gene sets, and M1, M2, and M1/M2 mixed microglial microarray gene sets. Real-time PCR and in situ hybridisation were performed to evaluate glial cell-derived neurotrophic factor (Gdnf) and S100a10 expression. Striatal GDNF protein levels were determined by enzyme-linked immunosorbent assay. RESULTS: MPTP treatment induced upregulation of Cx30 and Cx43 levels in the striatum of WT and KO mice. DA neuron loss was accelerated in Cx30 KO compared with WT mice after MPTP administration, despite no change in the striatal concentration of methyl-4-phenylpyridinium+. Astrogliosis in the striatum of Cx30 KO mice was attenuated by MPTP, whereas microglial activation was unaffected. Microarrays of the striatum showed reduced expression of pan-reactive and A2 astrocyte genes after MPTP treatment in Cx30 KO compared with WT mice, while M1, M2, and M1/M2 mixed microglial gene expression did not change. MPTP reduced the number of striatal astrocytes co-expressing Gdnf mRNA and S100ß protein or S100a10 mRNA and S100ß protein and also reduced the level of GDNF in the striatum of Cx30 KO compared with WT mice. CONCLUSIONS: These findings indicate that Cx30 plays critical roles in astrocyte neuroprotection in an MPTP PD model.

HLA-DRB1*04:05 allele is associated with intracortical lesions on three-dimensional double inversion recovery images in Japanese patients with multiple sclerosis.

BACKGROUND: Cortical lesions (CLs) frequently observed in Caucasian patients with multiple sclerosis (MS) contribute to disability. However, it remains unclear whether CLs are associated with clinical features and genetic risk factors, such as HLA-DRB1*15:01 and -DRB1*04:05 in Asian MS patients. OBJECTIVE: To elucidate the frequency of CLs and their association with HLA-DRB1 and DPB1 alleles in Japanese MS patients. METHODS: Three-dimensional double inversion recovery imaging and clinical information were retrospectively obtained from 92 Japanese MS patients. RESULTS: CLs of any type, intracortical lesions (ICLs), and leukocortical lesions (LCLs) were detected in 39.1%, 26.1%, and 28.3% of patients, respectively. MS patients with ICLs had a significantly higher frequency of secondary progression and greater Expanded Disability Status Scale (EDSS) scores than those without ICLs. Similar trends were observed with CLs and LCLs. The number of all three lesion types positively correlated with EDSS scores. The frequency and number of ICLs were significantly higher in HLA-DRB1*15:01 carriers than in HLA-DRB1*15:01 non-carriers, but significantly lower in HLA-DRB1*04:05 carriers than in HLA-DRB1*04:05 non-carriers. Multivariate logistic regression analysis revealed a negative association of HLA-DRB1*04:05 with ICLs. CONCLUSION: ICLs are associated with greater disease severity in Japanese MS patients and are partly suppressed by the HLA-DRB1*04:05 allele.

Early strong intrathecal inflammation in cerebellar type multiple system atrophy by cerebrospinal fluid cytokine/chemokine profiles: a case control study.

BACKGROUND: The pathology of multiple system atrophy cerebellar-type (MSA-C) includes glial inflammation; however, cerebrospinal fluid (CSF) inflammatory cytokine profiles have not been investigated. In this study, we determined CSF cytokine/chemokine/growth factor profiles in MSA-C and compared them with those in hereditary spinocerebellar ataxia (SCA). METHODS: We collected clinical data and CSF from 20 MSA-C patients, 12 hereditary SCA patients, and 15 patients with other non-inflammatory neurological diseases (OND), and measured 27 cytokines/chemokines/growth factors using a multiplexed fluorescent bead-based immunoassay. The size of each part of the hindbrain and hot cross bun sign (HCBS) in the pons was studied by magnetic resonance imaging. RESULTS: Granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin (IL)-6, IL-7, IL-12, and IL-13 levels were significantly higher in MSA-C and SCA compared with OND. In MSA-C, IL-5, IL-6, IL-9, IL-12, IL-13, platelet-derived growth factor-bb, macrophage inflammatory protein (MIP)-1α, and GM-CSF levels positively correlated with anteroposterior diameters of the pontine base, vermis, or medulla oblongata. By contrast, in SCA patients, IL-12 and MIP-1α showed significant negative correlations with anteroposterior diameters of the pontine base, and unlike MSA-C, there was no cytokine with a positive correlation in SCA. IL-6 was significantly higher in MSA-C patients with the lowest grade of HCBS compared with those with the highest grade. Macrophage chemoattractant protein-1 (MCP-1) had a significant negative correlation with disease duration only in MSA-C patients. Tumor necrosis factor-alpha, IL-2, IL-15, IL-4, IL-5, IL-10, and IL-8 were all significantly lower in MSA-C and SCA compared with OND, while IL-1ra, an anti-inflammatory cytokine, was elevated only in MSA-C. IL-1ß and IL-8 had positive correlations with Unified Multiple System Atrophy Rating Scale part 1 and 2, respectively, in MSA-C. CONCLUSIONS: Although CSF cytokine/chemokine/growth factor profiles were similar between MSA-C and SCA, pro-inflammatory cytokines, such as IL-6, GM-CSF, and MCP-1, correlated with the disease stage in a way higher at the beginning only in MSA-C, reflecting early stronger intrathecal inflammation.

Cerebral blood flow laterality derived from arterial spin labeling as a biomarker for assessing the disease severity of parkinson's disease.

PURPOSE: To evaluate cerebral blood flow (CBF) laterality derived from arterial spin labeling (ASL) in early-stage Parkinson's disease (PD) patients compared with those with advanced stages. MATERIALS AND METHODS: Thirty-eight patients with PD (21 patients in early stages, 17 patients in advanced stages) were retrospectively studied. The CBF maps derived from 3T ASL data were co-registered to the corresponding 3DT1WI using SPM 12 software. Caudate nucleus (CN), putamen (PT), globus pallidus (GP), and thalamus (TH) were manually traced on the representative axial slices of 3DT1WI. CBF of the CN, PT, GP, and TH was measured using corresponding pixels on the co-registered CBF maps. A laterality index (LI) was calculated as the ratio of the contralateral CBF to primary affected side CBF. Each LI was compared between early and advanced stages of PD using the Mann-Whitney U-test. The LIs were also compared between each stage of PD. RESULTS: In the CN, the LIs were significantly higher in early stages (mean LI ± SD, 95% confidence interval = 1.06 ± 0.14, 1.00-1.13) than in advanced stages (0.94 ± 0.14, 0.87-1.01; P < 0.05). We also observed a tendency toward decreased LIs with disease severity (1.10 ± 0.14, 0.99-1.21 for Hoehn and Yahr stage I; 1.04 ± 0.14, 0.92-1.12 for stage II; 0.96 ± 0.11, 0.89-1.10 for stage III; 0.93 ± 0.17, 0.81-1.05 for stage IV). CONCLUSION: The evaluation of CBF laterality pattern in the CN using ASL may be useful for assessing the disease severity of PD patients. LEVEL OF EVIDENCE: 3 J. MAGN. RESON. IMAGING 2017;45:1821-1826.

Structural changes in Parkinson's disease: voxel-based morphometry and diffusion tensor imaging analyses based on 123I-MIBG uptake.

OBJECTIVE: Patients with Parkinson's disease (PD) may exhibit symptoms of sympathetic dysfunction that can be measured using 123I-metaiodobenzylguanidine (MIBG) myocardial scintigraphy. We investigated the relationship between microstructural brain changes and 123I-MIBG uptake in patients with PD using voxel-based morphometry (VBM) and diffusion tensor imaging (DTI) analyses. METHODS: This retrospective study included 24 patients with PD who underwent 3 T magnetic resonance imaging and 123I-MIBG scintigraphy. They were divided into two groups: 12 MIBG-positive and 12 MIBG-negative cases (10 men and 14 women; age range: 60-81 years, corrected for gender and age). The heart/mediastinum count (H/M) ratio was calculated on anterior planar 123I-MIBG images obtained 4 h post-injection. VBM and DTI were performed to detect structural differences between these two groups. RESULTS: Patients with low H/M ratio had significantly reduced brain volume at the right inferior frontal gyrus (uncorrected p < 0.0001, K > 90). Patients with low H/M ratios also exhibited significantly lower fractional anisotropy than those with high H/M ratios (p < 0.05) at the left anterior thalamic radiation, the left inferior fronto-occipital fasciculus, the left superior longitudinal fasciculus, and the left uncinate fasciculus. CONCLUSIONS: VBM and DTI may reveal microstructural changes related to the degree of 123I-MIBG uptake in patients with PD. KEY POINTS: • Advanced MRI methods may detect brain damage more precisely. • Voxel-based morphometry can detect grey matter changes in Parkinson's disease. • Diffusion tensor imaging can detect white matter changes in Parkinson's disease.

Presenilins regulate calcium homeostasis and presynaptic function via ryanodine receptors in hippocampal neurons.

Presenilin (PS) plays a central role in the pathogenesis of Alzheimer's disease, and loss of PS causes progressive memory impairment and age-related neurodegeneration in the mouse cerebral cortex. In hippocampal neurons, PS is essential for neurotransmitter release, NMDA receptor-mediated responses, and long-term potentiation. PS is also involved in the regulation of calcium homeostasis, although the precise site of its action is less clear. Here we investigate the mechanism by which PS regulates synaptic function and calcium homeostasis using acute hippocampal slices from PS conditional knockout mice and primary cultured postnatal hippocampal neurons, in which PS is inducibly inactivated. Using two different calcium probes, Fura-2 and Mag-Fura-2, we found that inactivation of PS in primary hippocampal neurons does not affect calcium concentration in the endoplasmic reticulum. Rather, in the absence of PS, levels of ryanodine receptor (RyR) are reduced in the hippocampus, measured by Western analysis and radioligand binding assay, although the mRNA expression is unaffected. RyR-mediated function is also impaired, as indicated by reduced RyR agonist-induced calcium release from the ER and RyR-mediated synaptic responses in the absence of PS. Furthermore, knockdown of RyR expression in wild-type hippocampal neurons by two independent shRNAs to levels comparable with the RyR protein reduction in PS-deficient hippocampal neurons mimics the defects exhibited in calcium homeostasis and presynaptic function. Collectively, our findings show that PS regulates calcium homeostasis and synaptic function via RyR and suggest that disruption of intracellular calcium homeostasis may be an early pathogenic event leading to presynaptic dysfunction in Alzheimer's disease.

Astroglial connexin 43 is a novel therapeutic target for chronic multiple sclerosis model.

In chronic stages of multiple sclerosis (MS) and its animal model, experimental autoimmune encephalitis (EAE), connexin (Cx)43 gap junction channel proteins are overexpressed because of astrogliosis. To elucidate the role of increased Cx43, the central nervous system (CNS)-permeable Cx blocker INI-0602 was therapeutically administered. C57BL6 mice with chronic EAE initiated by MOG35-55 received INI-0602 (40 mg/kg) or saline intraperitoneally every other day from days post-immunization (dpi) 17-50. Primary astroglia were employed to observe calcein efflux responses. In INI-0602-treated mice, EAE clinical signs improved significantly in the chronic phase, with reduced demyelination and decreased CD3+ T cells, Iba-1+ and F4/80+ microglia/macrophages, and C3+GFAP+ reactive astroglia infiltration in spinal cord lesions. Flow cytometry analysis of CD4+ T cells from CNS tissues revealed significantly reduced Th17 and Th17/Th1 cells (dpi 24) and Th1 cells (dpi 50). Multiplex array of cerebrospinal fluid showed significantly suppressed IL-6 and significantly increased IL-10 on dpi 24 in INI-0602-treated mice, and significantly suppressed IFN-γ and MCP-1 on dpi 50 in the same group. In vitro INI-0602 treatment inhibited ATP-induced calcium propagations of Cx43+/+ astroglial cells to similar levels of those of Cx43-/- cells. Astroglial Cx43 hemichannels represent a novel therapeutic target for chronic EAE and MS.

Loss of leucine-rich repeat kinase 2 causes impairment of protein degradation pathways, accumulation of alpha-synuclein, and apoptotic cell death in aged mice.

Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common genetic cause of Parkinson's disease. LRRK2 is a large protein containing a small GTPase domain and a kinase domain, but its physiological role is unknown. To identify the normal function of LRRK2 in vivo, we generated two independent lines of germ-line deletion mice. The dopaminergic system of LRRK2(-/-) mice appears normal, and numbers of dopaminergic neurons and levels of striatal dopamine are unchanged. However, LRRK2(-/-) kidneys, which suffer the greatest loss of LRRK compared with other organs, develop striking accumulation and aggregation of alpha-synuclein and ubiquitinated proteins at 20 months of age. The autophagy-lysosomal pathway is also impaired in the absence of LRRK2, as indicated by accumulation of lipofuscin granules as well as altered levels of LC3-II and p62. Furthermore, loss of LRRK2 dramatically increases apoptotic cell death, inflammatory responses, and oxidative damage. Collectively, our findings show that LRRK2 plays an essential and unexpected role in the regulation of protein homeostasis during aging, and suggest that LRRK2 mutations may cause Parkinson's disease and cell death via impairment of protein degradation pathways, leading to alpha-synuclein accumulation and aggregation over time.

Iguratimod Ameliorates the Severity of Secondary Progressive Multiple Sclerosis in Model Mice by Directly Inhibiting IL-6 Production and Th17 Cell Migration via Mitigation of Glial Inflammation.

We previously reported a novel secondary progressive multiple sclerosis (SPMS) model, progressive experimental autoimmune encephalomyelitis (pEAE), in oligodendroglia-specific Cx47-inducible conditional knockout (Cx47 icKO) mice. Based on our prior study showing the efficacy of iguratimod (IGU), an antirheumatic drug, for acute EAE treatment, we aimed to elucidate the effect of IGU on the SPMS animal model. We induced pEAE by immunizing Cx47 icKO mice with myelin oligodendrocyte glycoprotein peptide 35-55. IGU was orally administered from 17 to 50 days post-immunization. We also prepared a primary mixed glial cell culture and measured cytokine levels in the culture supernatant after stimulation with designated cytokines (IL-1α, C1q, TNF-α) and lipopolysaccharide. A migration assay was performed to evaluate the effect of IGU on the migration ability of T cells toward mixed glial cell cultures. IGU treatment ameliorated the clinical signs of pEAE, decreased the demyelinated area, and attenuated glial inflammation on immunohistochemical analysis. Additionally, IGU decreased the intrathecal IL-6 level and infiltrating Th17 cells. The migration assay revealed reduced Th17 cell migration and IL-6 levels in the culture supernatant after IGU treatment. Collectively, IGU successfully mitigated the clinical signs of pEAE by suppressing Th17 migration through inhibition of IL-6 production by proinflammatory-activated glial cells.

Early and extensive alterations of glial connexins, distal oligodendrogliopathy type demyelination, and nodal/paranodal pathology are characteristic of multiple system atrophy.

The pathological hallmark of multiple system atrophy (MSA) is aberrant accumulation of phosphorylated α-synuclein in oligodendrocytes, forming glial cytoplasmic inclusions (GCIs). Extensive demyelination occurs particularly in the olivopontocerebellar and striatonigral pathways, but its precise mechanism remains elusive. Glial connexins (Cxs), which form gap junction channels between astrocytes and oligodendrocytes, play critical roles in myelin maintenance, and have not been studied in MSA. Therefore, we immunohistochemically investigated glial Cx changes in the cerebellar afferent fibers in 15 autopsied patients with MSA. We classified demyelinating lesions into three stages based on Klüver-Barrera staining: early (Stage I), intermediate (Stage II), and late (Stage III) stages showing subtle, moderate, and severe myelin reduction, respectively. Myelin-associated glycoprotein, but not myelin oligodendrocyte glycoprotein, was preferentially decreased in Stage I, suggesting distal oligodendrogliopathy type demyelination. Accumulation of phosphorylated α-synuclein in oligodendrocytes was frequently seen in Stage I but less frequently observed in Stages II and III. Tubulin polymerization-promoting protein (TPPP/p25α)-positive oligodendrocytes were preserved in Stage I but successively decreased in Stages II and III. Even at Stage I, Cx32 was nearly absent from myelin, despite the relative preservation of other nodal proteins, such as neurofascin, claudin-11/oligodendrocyte-specific protein, and contactin-associated protein 1, which successively decreased in the later stages. Cx32 was re-distributed in the oligodendrocyte cytoplasm and co-localized with GCIs. Cx47 gradually decreased at the oligodendrocyte surface in a stage-dependent manner but was not co-localized with GCIs. Astrocytic Cx43 was down-regulated in Stage I but up-regulated in Stages II and III, reflecting astrogliosis. Cx43/Cx47 gap junctions significantly decreased from Stage I to III. Activated microglia/macrophages and T cells infiltrated in Stage I rather than Stages II and III. Therefore, early and extensive alterations of glial Cxs, particularly Cx32 loss, occur in MSA and may accelerate distal oligodendrogliopathy type demyelination and nodal/paranodal dysfunction through disruption of inter-glial communication.

R1441C mutation in LRRK2 impairs dopaminergic neurotransmission in mice.

Dominantly inherited mutations in leucine-rich repeat kinase 2 (LRRK2) are a common genetic cause of Parkinson's disease (PD). The importance of the R1441 residue in the pathogenesis is highlighted by the identification of three distinct missense mutations. To investigate the pathogenic mechanism underlying LRRK2 dysfunction, we generated a knockin (KI) mouse in which the R1441C mutation is expressed under the control of the endogenous regulatory elements. Homozygous R1441C KI mice appear grossly normal and exhibit no dopaminergic (DA) neurodegeneration or alterations in steady-state levels of striatal dopamine up to 2 years of age. However, these KI mice show reductions in amphetamine (AMPH)-induced locomotor activity and stimulated catecholamine release in cultured chromaffin cells. The introduction of the R1441C mutation also impairs dopamine D2 receptor function, as suggested by decreased responses of KI mice in locomotor activity to the inhibitory effect of a D2 receptor agonist, quinpirole. Furthermore, the firing of nigral neurons in R1441C KI mice show reduced sensitivity to suppression induced by quinpirole, dopamine, or AMPH. Together, our data suggest that the R1441C mutation in LRRK2 impairs stimulated dopamine neurotransmission and D2 receptor function, which may represent pathogenic precursors preceding dopaminergic degeneration in PD brains.

Early decrease in intermediate monocytes in peripheral blood is characteristic of multiple system atrophy-cerebellar type.

To identify biomarkers for multiple system atrophy-cerebellar type (MSA-C), we used flow cytometry to measure surface marker expression of peripheral blood monocytes from patients with MSA-C or hereditary spinocerebellar degeneration (hSCD) and from healthy controls (HCs). The percentage of intermediate monocytes was significantly lower in MSA-C patients than in hSCD patients and HCs and showed significant positive correlations with disease duration and unified MSA rating scale scores. The percentage of CD62L+ intermediate monocytes was significantly lower in MSA-C patients than in hSCD patients and HCs. Early decrease of peripheral blood intermediate monocytes is characteristic of MSA-C and is a biomarker.

Comparison of Postoperative Stability Between BSSRO and Le Fort 1 Osteotomy with BSSRO in Skeletal Class III Malocclusion with Severe Open Bite.

INTRODUCTION: We aimed to investigate postoperative stability after orthognathic surgery in patients with skeletal class III malocclusion with severe open bite by comparison between bilateral sagittal splitting osteotomy (BSSRO) and BSSRO with Le Fort 1 osteotomy. MATERIALS AND METHODS: Seventeen patients with skeletal class III malocclusion with severe open bite who were needed more than 6 degree counterclockwise rotation of distal segment by only BSSRO in preoperative cephalometric prediction. The subjects were divided into group A, where 9 patients were treated by BSSRO, and group B, where 8 patients were treated by BSSRO with Le Fort 1 osteotomy. Patient's characteristics of age, gender, preoperative over jet (OJ) and over bite (OB) were not found to be significantly different between the two groups. Counterclockwise rotation of distal segment in preoperative cephalometric prediction by only BSSRO was not found to be significantly different between group A of 7.6 (6-10.6) degree and group B of 9 (6-13) degree. The amount of rotation was reduced to 5.4 (3-10) degree by bimaxillary surgery using BSSRO and Le Fort 1 osteotomy in group B. OJ and OB were measured as occlusal stability factor. Distance between ANS-to-PNS plane and the edge of upper incisor (NF-U1Ed), and distance between Menton and edge of lower incisor (Me-L1Ed) were measured as skeletal stability factor using cephalometric analysis. These lengths were measured at pre-surgery (T0), 2 weeks after surgery (T1) and 1 year after surgery (T2), and these differences between the two groups were statistically analyzed. RESULTS: OJ and OB kept a good relation at any experimental periods. The change of Me-L1Ed was significantly larger in group A (1.21 mm at T0-T1, 1.02 mm at T0-T2) than in group B (0.14 mm at T0-T1, 0.16 mm at T0-T2). The change of NF-U1Ed was not significantly different between group A (1.07 mm at T0-T1, 0.57 mm at T0-T2) and group B (0.51 mm at T0-T1, - 0.05 mm at T0-T2). CONCLUSION: In case with more than 6 degree counterclockwise rotation of distal segment, skeletal stability was better after bimaxillary surgery than only BSSRO; however, OJ and OB kept a good relation.

Temporal expression profiling of DAMPs-related genes revealed the biphasic post-ischemic inflammation in the experimental stroke model.

The neuroinflammation in the ischemic brain could occur as sterile inflammation in response to damage-associated molecular patterns (DAMPs). However, its long-term dynamic transcriptional changes remain poorly understood. It is also unknown whether this neuroinflammation contributes to the recovery or just deteriorates the outcome. The purpose of this study is to characterize the temporal transcriptional changes in the post-stroke brain focusing on DAMPs-related genes by RNA-sequencing during the period of 28 days. We conducted the RNA-sequencing on day 1, 3, 7, 14, 28 post-stroke in the mouse photothrombosis model. The gross morphological observation showed the ischemic lesion on the ipsilateral cortex turned into a scar with the clearance of cellular debris by day 28. The transcriptome analyses indicated that post-stroke period of 28 days was classified into four categories (I Baseline, II Acute, III Sub-acute-#1, IV Sub-acute-#2 phase). During this period, the well-known genes for DAMPs, receptors, downstream cascades, pro-inflammatory cytokines, and phagocytosis were transcriptionally increased. The gene ontology (GO) analysis of biological process indicated that differentially expressed genes (DEGs) are genetically programmed to achieve immune and inflammatory pathways. Interestingly, we found the biphasic induction of various genes, including DAMPs and pro-inflammatory factors, peaking at acute and sub-acute phases. At the sub-acute phase, we also observed the induction of genes for phagocytosis as well as regulatory and growth factors. Further, we found the activation of CREB (cAMP-response element binding protein), one of the key players for neuronal plasticity, in peri-ischemic neurons by immunohistochemistry at this phase. Taken together, these findings raise the possibility the recurrent inflammation occurs at the sub-acute phase in the post-stroke brain, which could be involved in the debris clearance as well as neural reorganization.

Factors related to patients' nutritional state after orthognathic surgery.

PURPOSE: The purpose of this study was to evaluate patients' nutritional state after orthognathic surgery. METHODS: The subjects were 40 female patients with dentofacial deformity aged 17-33 years who were undergoing bilateral sagittal splitting ramus osteotomy. Twenty patients were treated with intermaxillary fixation, and 20 patients were treated without intermaxillary fixation. Age and body mass index (kg/m2) were assessed as physical factors, operation time, blood loss, and amount of mandibular movement with or without intermaxillary fixation were assessed as operation stress factors, and the following laboratory data, total protein, serum albumin, total cholesterol, total lymphocytes, and cholinesterase were assessed as nutritional state factors at 1 and 2 weeks after surgery. Statistical analysis was performed for body weight loss and relationship between body weight loss and examination factors. RESULTS: Body weight significantly decreased 2.3% at 1 week and 3.9% at 2 weeks after surgery rather than preoperation. All laboratory data except total lymphocyte were decreased at 1 week after surgery and still remained significantly decreased at 2 weeks after surgery. There was a statistically significant relationship between body weight loss at 1 week after surgery and operation time. CONCLUSIONS: These results indicate that long operation time caused body weight loss in orthognathic surgery.

A novel model for treatment of hypertrophic pachymeningitis.

Objective: Immunoglobulin (Ig)G4-related disease is a major cause of hypertrophic pachymeningitis (HP), presenting as a progressive thickening of the dura mater. HP lacks an animal model to determine its underlying mechanisms. We developed a suitable animal model for the treatment of HP. Methods: We longitudinally evaluated dura in mice with a mutation (Y136F) in the linker for activation of T cells (LAT), which induced type 2 T helper (Th2) cell proliferation and IgG1 (IgG4 human equivalent) overexpression. Mice were therapeutically administered daily oral irbesartan from 3 to 6 weeks of age. Human IgG4-related, anti-neutrophil cytoplasmic antibody-related, and idiopathic HP dura were also immunohistochemically examined. Results: LATY136F mice showing dural gadolinium enhancement on magnetic resonance imaging had massive infiltration of B220+ B cells, IgG1+ cells, CD138+ plasma cells, CD3+ T cells, F4/80+ macrophages, and polymorphonuclear leukocytes in the dura at 3 weeks of age, followed by marked fibrotic thickening. In dural lesions, transforming growth factor (TGF)-ß1 was produced preferentially in B cells and macrophages while TGF-ß receptor I (TGF-ß RI) was markedly upregulated on fibroblasts. Quantitative western blotting revealed significant upregulation of TGF-ß1, TGF-ß RI, and phosphorylated SMAD2/SMAD3 in dura of LATY136F mice aged 13 weeks. A similar upregulation of TGF-ß RI, SMAD2/SMAD3, and phosphorylated SMAD2/SMAD3 was present in autopsied dura of all three types of human HP. Irbesartan abolished dural inflammatory cell infiltration and fibrotic thickening in all treated LATY136F mice with reduced TGF-ß1 and nonphosphorylated and phosphorylated SMAD2/SMAD3. Interpretation: TGF-ß1/SMAD2/SMAD3 pathway is critical in HP and is a potential novel therapeutic target.