Evaluation of airway involvement and treatment in patients with relapsing polychondritis.

Airway involvement in relapsing polychondritis (RP) can be debilitating and life threatening, often requiring interventional procedures. If standard therapies including systemic corticosteroid and immunosuppressive agents are ineffective, airway stenting is often required. Recently, biologics have been reported to be effective for RP, and the early administration of biologics may avoid airway stenting. To evaluate survival rates and treatment approaches, medical records of RP patients with airway involvement were reviewed. These cases were divided into the following groups: with and without malacia, stenting and non-stenting, and with and without biologics. Kaplan-Meier was used to calculate survival rates and log rank tests were used to analyze biologics groups. A total of 77 patients were enrolled. Airway stenting was performed in 13 patients, all of which developed airway malacia. The stenting group had significantly lower survival rates than the non-stenting group (p < 0.001). Stent-related complications were granulation tissue (85%) and mucostasis (69%). In the non-stenting group, a lower mortality rate was observed. A significantly higher survival rate was seen in patients administered biologics than without (p = 0.014). The early administration of biologics shows promise in preventing severe airway disorders that require airway stenting.

Establishment of a novel experimental system using single cell-derived pleomorphic rhabdomyosarcoma cell lines expressing K-RasG12V and deficient in p53.

Pleomorphic rhabdomyosarcoma (PRMS) predominantly arises in adult skeletal musculature and is usually associated with poor prognosis. Thus, effective treatments must be developed. PRMS is a rare tumor; therefore, it is critical to develop an experimental system to understand the cellular and molecular mechanisms of PRMS. We previously demonstrated that PRMS develops after p53 gene deletion and oncogenic K-Ras expression in the skeletal muscle tissue. In that study, oncogenic K-Ras-expressing cells were diverse and the period until disease onset was difficult to control. In this study, we developed an experimental system to address this problem. Single cell-derived murine cell lines, designated as RMS310 and RMSg2, were established by limiting the dilution of cells from a lung metastatic tumor colony that were positive for various cancer stem cells and activated skeletal muscle-resident stem/progenitor cell marker genes by RT-PCR. All cell lines stably recapitulated the histological characteristics of human PRMS as bizarre giant cells, desmin-positive cells, and lung metastases in C57BL/6 mice. All subclones of the RMSg2 cells by the limiting dilution in vitro could seed PRMS subcutaneously, and as few as 500 RMSg2 cells were sufficient to form tumors. These results suggest that the RMSg2 cells are multipotent cancer cells that partially combine the properties of skeletal muscle-resident stem/progenitor cells and high tumorigenicity. Thus, our model system's capacity to regenerate tumor tissue in vivo and maintain stable cells in vitro makes it useful for developing therapeutics to treat PRMS.

Serum-Induced Expression of Brain Natriuretic Peptide Contributes to Its Increase in Patients with HFpEF.

Brain natriuretic peptide (BNP) levels are increased in both patients with heart failure with preserved (HFpEF) and reduced ejection fraction (HFrEF), but the reasons for this remain unclear. Our purpose was to examine whether serum-induced BNP (iBNP) expression partly contributes to increased BNP in patients with HFpEF. BNP reporter cardiomyocytes from pBNP-luc-KI mice were stimulated with serum from patients with HFpEF or HFrEF (n = 114 and n = 82, respectively). Luciferase activity was examined as iBNP and the iBNP-to-BNP ratio was evaluated. Patient characteristics and clinical parameters were compared, and multivariate regression analysis was performed to determine independent predictors of the iBNP-to-BNP ratio. Female sex and frequencies of atrial fibrillation, hypertension and the use of a calcium channel blocker (CCB) were higher in HFpEF. The iBNP-to-BNP ratio was significantly higher in HFpEF (26.9) than in HFrEF (16.1, p < 0.001). Multivariate regression analysis identified the existence of HFpEF as an independent predictor of the iBNP-to-BNP ratio after adjusting for all other measurements (ß = 0.154, p = 0.032). Age, hemoglobin, CCB usage and deceleration time were also independent predictors (ß = 0.167, p = 0.025; ß = 0.203, p = 0.006; ß = 0.138, p = 0.049; and ß = 0.143, p = 0.049, respectively). These results indicate that the elevated BNP in patients with HFpEF is partly due to iBNP from the heart.

Mechanical model of steady-state and inflammatory conditions in patients with relapsing polychondritis: A review.

ABSTRACT: Relapsing polychondritis (RP) is a multisystem inflammatory disorder, considered to associate with immune aberration.Increased T helper type-1 cell-related cytokines were reported in RP patients. mRNA expressions of a regulatory T cell cytokine interleukin (IL)-10 increased, whereas pro-inflammatory cytokines IL1ß and IL6 mRNA expressions decreased in freshly isolated peripheral blood mononuclear cells of RP patients compared with those in healthy individuals. Upon in vitro stimulation with mitogen, IL10 mRNA expressions decreased, and IL1ß and IL6 mRNA expressions increased in RP patients.This short-time dynamic change of gene expressions from anti-inflammatory to pro-inflammatory features of immune cells may be associated with the "relapsing" disease course of patients with RP. IL1ß mRNA expressions of peripheral blood mononuclear cells exhibited positive correlations with serum matrix metalloproteinase (MMP)-3 concentrations in patients with respiratory involvement. Such positive correlation was not found in those without respiratory involvement.In a metagenomic analysis, an altered composition of gut microbes was found, suggesting that microbe metabolites such as short-chain fatty acids may affect T cell responses of the patients.In this review, the relationships among RP-related inflammatory molecules were summarized. The data support a hypothesis that the immune conditions are different between steady-state and inflammation in RP patients.

Nationwide cross-sectional survey of patients with relapsing polychondritis in 2019 demonstrates reduction of airway involvement compared with that in 2009.

We conducted retrospective cohort studies of patients with relapsing polychondritis (RP) twice in 2009 and 2019, using a physician questionnaire. We compared the patients' clinical statuses between the years. Age and gender were comparable between the two surveys. Mean disease duration was longer in 2019 survey (8.3 years) than that in 2009 survey (4.8 years, P < 0.001). The mortality rate declined in 2019 survey compared with those in 2009 survey (from 9.2 to 1.6%, P < 0.001). Incidence of airway involvement decreased in 2019 survey compared with that in 2009 survey (from 49 to 37%, P = 0.012). In 2019 survey, we found more frequent use of biological agents and immunosuppressants in patients with airway involvement. When we focused on RP patients with airway involvement, physicians in 2019 chose methotrexate and calcineurin inhibitors preferentially, compared with azathioprine and cyclophosphamide. Of note is that increased use of infliximab was observed in RP patients with airway involvement, but not in those without. Reduction of airway involvement and mortality in patients with RP was observed in 2019 survey. The reduction may associate with the frequent use of biologics including infliximab in RP patients with airway involvement.

Nicotine treatment regulates PD-L1 and PD-L2 expression via inhibition of Akt pathway in HER2-type breast cancer cells.

The immune checkpoint molecules such as PD-L1 and PD-L2 have a substantial contribution to cancer immunotherapy including breast cancer. Microarray expression profiling identified several molecular subtypes, namely luminal-type (with a good-prognosis), HER2-type (with an intermediate-prognosis), and triple-negative breast cancer (TNBC)-type (with a poor-prognosis). We found that PD-L1 and PD-L2 mRNA expressions were highly expressed in TNBC-type cell lines (HCC1937, MDA-MB-231), moderately expressed in HER2-type cell line (SK-BR-3), and poorly expressed in luminal-type cell lines (MDA-MB-361, MCF7). The PD-L1 and PD-L2 expression in SK-BR-3 cells, but not those in HCC1937 and MDA-MB-231 cells, decreased by nicotine stimulation in a dose-dependent manner. In addition, nicotine treatment decreased the phosphorylation of Akt in SK-BR-3 cells, but not in other cell lines. These results show that nicotine regulates the expression of immune checkpoint molecules, PD-L1 and PD-L2, via inhibition of Akt phosphorylation. This findings may provide the new therapeutic strategies for the treatment of breast cancer.

Serum MMP3 Correlated With IL1ß Messenger RNA Expressions of Peripheral Blood Mononuclear Cells in Patients With Relapsing Polychondritis With Respiratory Involvement.

OBJECTIVE: Respiratory involvement was intimately associated with poorer prognosis in patients with relapsing polychondritis (RP). We previously reported that high serum matrix metalloproteinase-3 (MMP3) was frequently observed in patients with RP with respiratory involvement. Elevated MMP3 secreted through local inflammation may be associated with the development of airway lesions. METHODS: We collected peripheral blood mononuclear cells (PBMCs) and sera from 30 patients with RP and 14 healthy individuals. Interleukin (IL) 1ß, IL6, and tumor necrosis factor (TNF) α messenger RNA (mRNA) expressions were analyzed in freshly isolated and cultured PBMCs with phytohemagglutinin and phorbol myristate acetate stimulation by real-time reverse transcription polymerase chain reaction and serum MMP3 by enzyme-linked immunosorbent assay (ELISA). RESULTS: We confirmed our previous finding that patients with respiratory involvements showed higher serum MMP3 compared with patients lacking respiratory involvement. IL1ß mRNA expression was significantly higher in patients with RP than in healthy individuals after mitogenic stimulation. TNFα mRNA expression after stimulation was significantly lower in patients with RP compared with in healthy individuals. We performed correlation analyses between MMP3 and cytokine mRNA expressions in patients with RP. In patients with respiratory involvement, MMP3 correlated with IL1ß and IL6 after stimulation. In patients without respiratory involvement, no positive correlations between MMP3 and cytokine mRNA expressions were observed regardless of culture condition. We did not find any positive correlations between MMP3 and TNFα mRNA expression in patients with RP. CONCLUSION: It is possible that IL1ß mRNA expression associates by some means with airway inflammation via the secretion of MMP3 in patients with RP. Involvement of proinflammatory cytokines, including IL1ß, was suggested for the pathophysiology of airway lesions in patients with RP.

Dementia model mice exhibited improvements of neuropsychiatric symptoms as well as cognitive dysfunction with neural cell transplantation.

Elderly patients with dementia suffer from cognitive dysfunctions and neuropsychiatric symptoms (NPS) such as anxiety and depression. Alzheimer's disease (AD) is a form of age-related dementia, and loss of cholinergic neurons is intimately associated with development of AD symptoms. We and others have reported that neural cell transplantation ameliorated cognitive dysfunction in AD model mice. It remains largely unclear whether neural cell transplantation ameliorates the NPS of AD. It would be interesting to determine whether NPS correlates with cognitive dysfunctions before and after neural cell transplantation in AD model mice. Based on the revalidation of our previous data from a Morris water maze test, we found that neural cell transplantation improved anxiety and depression significantly and marginally affected locomotion activity in AD mice. A correlation analysis revealed that the spatial learning function of AD mice was correlated with their NPS scores both before and after cell transplantation in a similar manner. In contrast, in the mice subjected to cell transplantation, spatial reference memory function was not correlated with NPS scores. These results suggested the neural cell transplantation in the AD model mice significantly improved NPS to the same degree as cognitive dysfunctions, possibly via distinct mechanisms, such as the cholinergic and GABAergic systems.

Defective Reelin/Dab1 signaling pathways associated with disturbed hippocampus development of homozygous yotari mice.

Homozygous Dab1 yotari mutant mice, Dab1yot (yot/yot) mice, have an autosomal recessive mutation of Dab1 and show reeler-like phenotype including histological abnormality of the cerebellum, hippocampus, and cerebral cortex. We here show abnormal hippocampal development of yot/yot mice where granule cells and pyramidal cells fail to form orderly rows but are dispersed diffusely in vague multiplicative layers. Possibly due to the positioning failure of granule cells and pyramidal cells and insufficient synaptogenesis, axons of the granule cells did not extend purposefully to connect with neighboring regions in yot/yot mice. We found that both hippocampal granule cells and pyramidal cells of yot/yot mice expressed proteins reactive with the anti-Dab1 antibody. We found that Y198- phosphorylated Dab1 of yot/yot mice was greatly decreased. Accordingly the downstream molecule, Akt was hardly phosphorylated. Especially, synapse formation was defective and the distribution of neurons was scattered in hippocampus of yot/yot mice. Some of neural cell adhesion molecules and hippocampus associated transcription factors of the neurons were expressed aberrantly, suggesting that the Reelin-Dab1 signaling pathway seemed to be importantly involved in not only neural migration as having been shown previously but also neural maturation and/or synaptogenesis of the mice. It is interesting to clarify whether the defective neural maturation is a direct consequence of the dysfunctional Dab1, or alternatively secondarily due to the Reelin-Dab1 intracellular signaling pathways.

Female dominance of both spatial cognitive dysfunction and neuropsychiatric symptoms in a mouse model of Alzheimer's disease.

Alzheimer's disease (AD) is a prevalent neurological disorder affecting memory function in elderly persons. Indeed, AD exhibits abnormality in cognitive behaviors and higher susceptibility to neuropsychiatric symptoms (NPS). Various factors including aging, sex difference and NPS severity, are implicated during in development of AD. In this study, we evaluated behavioral abnormalities of AD model, PDAPP transgenic mice at young age using the Morris Water Maze test, which was established to assess hippocampal-dependent learning and memory. We found that female AD model mice exhibited spatial learning dysfunction and highly susceptible to NPS such as anxiety and depression, whereas spatial reference memory function was comparable in female PDAPP Tg mice to female wild type (WT) mice. Spatial learning function was comparable in male AD model mice to male WT mice. Multiple regression analysis showed that spatial learning dysfunction was associated with NPS severity such as anxiety and depression. Furthermore, the analysis showed that spatial reference memory function was associated with status of depression, but not anxiety. Thus, these results suggest female dominance of spatial learning dysfunction in the AD model mice accompanying increased NPS severity. The understandings of AD model may be useful for the development of therapeutic agents and methods in human AD.

Generation of Transgenic Mice that Conditionally Overexpress Tenascin-C.

Tenascin-C (TNC) is an extracellular matrix glycoprotein that is expressed during embryogenesis. It is not expressed in normal adults, but is up-regulated under pathological conditions. Although TNC knockout mice do not show a distinct phenotype, analyses of disease models using TNC knockout mice combined with in vitro experiments revealed the diverse functions of TNC. Since high TNC levels often predict a poor prognosis in various clinical settings, we developed a transgenic mouse that overexpresses TNC through Cre recombinase-mediated activation. Genomic walking showed that the transgene was integrated into and truncated the Atp8a2 gene. While homozygous transgenic mice showed a severe neurological phenotype, heterozygous mice were viable, fertile, and did not exhibit any distinct abnormalities. Breeding hemizygous mice with Nkx2.5 promoter-Cre or α-myosin heavy chain promoter Cre mice induced the heart-specific overexpression of TNC in embryos and adults. TNC-overexpressing mouse hearts did not have distinct histological or functional abnormalities. However, the expression of proinflammatory cytokines/chemokines was significantly up-regulated and mortality rates during the acute stage after myocardial infarction were significantly higher than those of the controls. Our novel transgenic mouse may be applied to investigations on the role of TNC overexpression in vivo in various tissue/organ pathologies using different Cre donors.

Precise Regulation of the Basal PKCγ Activity by DGKγ Is Crucial for Motor Coordination.

Diacylglycerol kinase γ (DGKγ) is a lipid kinase to convert diacylglycerol (DG) to phosphatidic acid (PA) and indirectly regulates protein kinase C γ (PKCγ) activity. We previously reported that the basal PKCγ upregulation impairs cerebellar long-term depression (LTD) in the conventional DGKγ knockout (KO) mice. However, the precise mechanism in impaired cerebellar LTD by upregulated PKCγ has not been clearly understood. Therefore, we first produced Purkinje cell-specific DGKγ KO (tm1d) mice to investigate the specific function of DGKγ in Purkinje cells and confirmed that tm1d mice showed cerebellar motor dysfunction in the rotarod and beam tests, and the basal PKCγ upregulation but not PKCα in the cerebellum of tm1d mice. Then, the LTD-induced chemical stimulation, K-glu (50 mM KCl + 100 µM, did not induce phosphorylation of PKCα and dissociation of GluR2 and glutamate receptor interacting protein (GRIP) in the acute cerebellar slices of tm1d mice. Furthermore, treatment with the PKCγ inhibitor, scutellarin, rescued cerebellar LTD, with the phosphorylation of PKCα and the dissociation of GluR2 and GRIP. In addition, nonselective transient receptor potential cation channel type 3 (TRPC3) was negatively regulated by upregulated PKCγ. These results demonstrated that DGKγ contributes to cerebellar LTD by regulation of the basal PKCγ activity.

Elucidation of predictors of disease progression in patients with relapsing polychondritis at the onset: potential impact on patient monitoring.

BACKGROUND: In patients with relapsing polychondritis (RP), organ involvement developed in those with progressive and/or long disease courses. For their management, elucidation of a subgroup suggesting disease progression is awaited. METHODS: We previously conducted a physician's questionnaire-based retrospective study to elucidate major clinical features of Japanese patients with RP. We here evaluated organ involvement at disease onset and at the last follow-up. We then counted cumulative numbers of involved organs at the last follow-up in 229 RP patients and compared them with involved organ numbers at disease onset, as possible indicators of disease progression. We assigned their prognosis at the last follow-up into "patient prognostic stages" from no medication (stage 1) to death (stage 5). We utilized nonparametric tests for group comparisons. RESULTS: Involved organ numbers per-patient were 1.13 ± 0.03 at disease onset and 3.25 ± 0.10 at the last follow-up (disease duration was 4.69 ± 0.33 years), and increased along with the patient prognostic stages.At disease onset, 135 and 48 patients had auricular involvement (59% of 229 patients, defined as auricular-onset subgroup; AO) and respiratory involvement (21%, respiratory-onset subgroup; RO), respectively. 46 patients presented with other conditions (20%, miscellaneous-onset subgroup; MO) including CNS, ocular, and inner ear involvement, among others.RO patients showed worse (poorer) prognostic stages than AO patients. MO patients developed respiratory and/or auricular involvement thereafter and then showed significantly higher mortality rate (15%; 7/46) than AO patients (5.9%; 8/135).In RP patients who did not develop respiratory involvement until the last follow-up (throughout the disease course; 117 patients), mortality rate was 19% in 26 MO patients and 3.3% in 91 AO patients. Accordingly, RO patients and MO patients associated with relatively poor prognosis compared with AO patients. CONCLUSIONS: Allocation of patients to RO and MO subgroups was suggested to associate with poorer prognosis of RP than AO subgroups, especially AO subgroups without respiratory involvement throughout. All RP patients deserve careful monitoring but special attention should be paid to MO patients because of their diverse and accelerated disease progression.

Interaction between SDF1 and CXCR4 Promotes Photoreceptor Differentiation via Upregulation of NFκB Pathway Signaling Activity in Pax6 Gene-Transfected Photoreceptor Precursors.

BACKGROUND: CCL2 (also known as monocyte chemoattractant protein 1) and CX3CR1 (also known as Fractalkine receptor)-deficient mice have damaged photoreceptors. OBJECTIVES: We examined the interaction of SDF1 and CXCR4 on the differentiation of retinal progenitors into rhodopsin-positive photoreceptors. METHODS: Cloned retinal progenitors were obtained by Pax6 gene transfection of mouse iPS cells followed by serial dilution. Clones were selected by expression of nestin, Musashi1, Six3, and Chx10 mRNA. Cell surface protein expression was analyzed by flow cytometry. The levels of mRNA and intracellular protein were examined by real-time PCR and immunochemistry, respectively. Transient transfection experiments of retinal progenitors were conducted using a human rhodopsin promoter luciferase plasmid. RESULTS: We selected 10 clones that expressed Six3, Chx10, Crx, Rx1, Nrl, CD73, and rhodopsin mRNA, which, except for rhodopsin, are photoreceptor precursor markers. Clones expressed both CD73 and CXCR4 on the cell surface and differentiated into rhodopsin-positive photoreceptors, which was reinforced by the addition of exogenous SDF1. A CXCR4 inhibitor AMD3100 blocked SDF1-mediated differentiation of progenitors into photoreceptors. SDF1 enhanced human rhodopsin promoter transcription activity, possibly via the NFκB pathway. Addition of SDF1 to the cell culture induced nuclear translocation of NFκB on retinal progenitor cell clones. Neonatal and newborn mouse retinas expressed SDF1 and CXCR4. Cells in the outer nuclear layer where photoreceptors are located expressed CXCR4 at P14 and P56. Cells in the inner nuclear layer expressed SDF1. CONCLUSIONS: These findings suggest that retinal progenitor cell differentiation was at least partly regulated by SDF1 and CXCR4 via upregulation of NFκB activity.

Facile preparation of aminosilane-alginate hybrid beads for enzyme immobilization: Kinetics and equilibrium studies.

A simple, facile and potential platform for enzyme immobilization using alginate-based beads has been demonstrated by simultaneous gelation and modification of alginate using calcium chloride (CaCl2) and 3-aminopropyltriethoxysilane (APTES). In this method, sodium alginate solution containing enzyme was simply dripped into a crosslinker solution containing CaCl2 and APTES, leading to the formation of APTES-alginate hybrid beads (AP-beads). The optical observation, FT-IR analysis and amino group measurements provided evidence that APTES was successfully adsorbed to the alginate chain via electrostatic interaction. On the assumption that the binding of Ca2+ ion to polymannuronate residues of alginate via bidentate bridging coordination is competitive with APTES, the equilibrium isotherm and kinetics for the adsorption of APTES to AP-beads was found to follow extended Langmuir isotherm in binary system. Formate dehydrogenase (FDH) as a model enzyme was successfully immobilized in AP-beads and the immobilization yield of ca. 100% could be achieved under optimal conditions of CaCl2 and APTES concentrations in crosslinker solution. Furthermore, the AP-beads were reused efficiently for 9 cycles without loss of FDH activity. The above results demonstrated that AP-beads were effective support for enzyme immobilization.

Hyper-Formation of GABA and Glycine Co-Releasing Terminals in the Mouse Cerebellar Nuclei after Deprivation of GABAergic Inputs from Purkinje Cells.

GABA and glycine are inhibitory neurotransmitters. However, the mechanisms underlying the formation of GABAergic and glycinergic synapses remain unclear. The influence of GABAergic input deprivation on inhibitory terminal formation was investigated using Purkinje cell (PC)-specific vesicular GABA transporter (VGAT) knockout (L7-VGAT) mice, in which GABA release from PCs diminishes in an age-dependent manner. We compared the late development of GABAergic and glycinergic terminals in the cerebellar nucleus (CN) between control and L7-VGAT mice. In the control CN, the density of glutamate decarboxylase (GAD)-positive dots remained unchanged between postnatal 2 months (P2M) and 13 months (P13M), whereas glycine transporter 2 (GlyT2)-positive dots increased in density during this time frame. No difference in the density of GlyT2-positive dots was observed between control and L7-VGAT mice at P2M, but the density was significantly higher in the L7-VGAT fastigial nuclei (FN) than the control FN at P13M. When VGAT was absent from PC terminals, GlyT2-positive dots included GAD and VGAT and formed synapses. These results indicated that GABAergic terminals were formed by P2M, glycinergic terminals were actively formed after P2M, and more glycinergic terminals were formed in the L7-VGAT FN than in the control FN, suggesting that the increased glycinergic terminals may derive from interneurons within the FN and may also release GABA. These results suggest that the deprivation of GABAergic inputs from PCs may accelerate the formation of co-releasing terminals derived from interneurons and that the inhibitory terminal numbers and types may be regulated by the quantity of functional GABAergic inputs.

BNP as a Major Player in the Heart-Kidney Connection.

Brain natriuretic peptide (BNP) is an important biomarker for patients with heart failure, hypertension and cardiac hypertrophy. Although it is known that BNP levels are relatively higher in patients with chronic kidney disease and no heart disease, the mechanism remains unknown. Here, we review the functions and the roles of BNP in the heart-kidney interaction. In addition, we discuss the relevant molecular mechanisms that suggest BNP is protective against chronic kidney diseases and heart failure, especially in terms of the counterparts of the renin-angiotensin-aldosterone system (RAAS). The renal medulla has been reported to express depressor substances. The extract of the papillary tips from kidneys may induce the expression and secretion of BNP from cardiomyocytes. A better understanding of these processes will help accelerate pharmacological treatments for heart-kidney disease.

Roles of Reelin/Disabled1 pathway on functional recovery of hemiplegic mice after neural cell transplantation; Reelin promotes migration toward motor cortex and maturation to motoneurons of neural grafts.

Reelin is a large glycoprotein which regulates central nervous system (CNS) development. Dysfunctions of Reelin were reported on certain neuropsychiatric diseases. We examined involvement of Reelin pathway in functional recovery of hemiplegic mice after neural transplantation. Reelin was expressed 1 day after cryogenic injury of right motor cortex. We transplanted neural stem/progenitor cells (NSPCs) from wild-type mice into ipsilateral striatum of hemiplegic mice. The grafts migrated from the striatum and reached the injured cortex 14 days after transplantation. The transplantation significantly improved their motor functions (P < .05). The NSPCs migrating toward the cortex expressed Reelin receptors, Apoer and Vldlr, and phosphorylated Disabled1 (Dab1), a downstream signaling molecule of Reelin. The grafts expressed Ncadherin and active form of Integrin ß1, both of which were known to become active with Reelin stimulation. At day 28, the grafts expressed Ctip2, Crim1, Foxp2, and Fezf2, all of which were forebrain motoneuron associated markers, and Nfm and Synapsin1 on the damaged cortex. We then transplanted NSPCs of yotari mice (yot/yot genotype) having nonfunctional Dab1 by a mutation of its gene. Majority of the grafts from yotari mice (>80%) did not migrate and thus remained at the striatum. The grafts did not express the forebrain motoneuron associated markers nor the cell adhesion molecules including Ncadherin and active Integrin ß1. Reelin pathway was involved in graft migration by regulating certain adhesion molecules and in their differentiation to functional motoneurons accompanying synapse formation. We suggested involvement of Reelin pathway for neural regeneration and functional recovery of hemiplegic mice in adulthood after neural transplantation.

Cerebellar Neurodegeneration and Neuronal Circuit Remodeling in Golgi pH Regulator-Deficient Mice.

The Golgi apparatus plays an indispensable role in posttranslational modification and transport of proteins to their target destinations. Although it is well established that the Golgi apparatus requires an acidic luminal pH for optimal activity, morphological and functional abnormalities at the neuronal circuit level because of perturbations in Golgi pH are not fully understood. In addition, morphological alteration of the Golgi apparatus is associated with several neurodegenerative diseases, including Parkinson's disease, Alzheimer's disease, and amyotrophic lateral sclerosis. Here, we used anatomical and electrophysiological approaches to characterize morphological and functional abnormalities of neuronal circuits in Golgi pH regulator (GPHR) conditional knock-out mice. Purkinje cells (PCs) from the mutant mice exhibited vesiculation and fragmentation of the Golgi apparatus, followed by axonal degeneration and progressive cell loss. Morphological analysis provided evidence for the disruption of basket cell (BC) terminals around PC soma, and electrophysiological recordings showed selective loss of large amplitude responses, suggesting BC terminal disassembly. In addition, the innervation of mutant PCs was altered such that climbing fiber (CF) terminals abnormally synapsed on the somatic spines of mutant PCs in the mature cerebellum. The combined results describe an essential role for luminal acidification of the Golgi apparatus in maintaining proper neuronal morphology and neuronal circuitry.

Relative abundance of Megamonas hypermegale and Butyrivibrio species decreased in the intestine and its possible association with the T cell aberration by metabolite alteration in patients with Behcet's disease (210 characters).

OBJECTIVES: We have previously demonstrated that the phylum Actinobacteria, the family Lactobacillaceae, and the genus Bifidobacterium increased in relative abundance of gut microbiota in patients with Behcet's disease (BD). The phylum Firmicutes and the class Clostridia were predominant in the feces of normal individuals. The class Clostridia includes short-chain fatty acid-producing bacteria, important for the balance between regulatory T cells and helper T type 17 (Th17) cells. It is possible that the bacterial compositional alteration causes low intestinal short-chain fatty acid concentrations, leading to skewed immune functions in patients with BD. METHODS: To test the hypothesis, we examined species composition and gene functions from the 16S rRNA data by utilizing PICRUSt software. RESULTS: We have shown that relative abundance of Eggerthella lenta, Acidaminococcus species, Lactobacillus mucosae, Bifidobacterium bifidum, Lactobacillus iners, Streptococcus species, and Lactobacillus salivarius increased significantly in patients with BD. Relative abundance of Megamonas hypermegale, Butyrivibrio species, Streptococcus infantis, and Filifactor species increased significantly in normal individuals compared with BD patients. In the functional annotation analysis by PICRUSt, we found prevalent gene functions of the pentose phosphate pathway and the inosine monophosphate biosynthesis in patients with BD. The data suggested that BD gut microbes altered nucleic acid and fatty acid synthesis. CONCLUSIONS: These compositional and functional alterations of gut microbes may accompany unfavorable molecular exchanges between intestinal immunocompetent cells and gut microbes, and these interactions may have an association with the immune aberration in patients with BD.