Background and Aims: Cardiovascular disorders may be closely related to abnormal bone metabolism. The progression of atherosclerosis and vascular calcification has an inverse relation with a decrease in bone mineral density, and this is known as so-called “bone-vascular relationship”. This cross-sectional study is performed to elucidate the relationship between bone metabolic markers and atherosclerosis.

Aim: Trimethylamine-N-oxide (TMAO) recently attracted much attention as it was shown to promote atherosclerosis. TMAO is produced from carnitine in meet through the metabolism by gut microbiota and the liver. On the other hand, TMAO level is known to be very high in fish meet. We therefore examined the effect of TMAO on atherosclerosis in Japanese, who daily consume fish.

Aim: Reactive hyperemia index (RHI) assessed by reactive hyperemia-peripheral arterial tonometry (RH-PAT), is associated with flow mediated dilatation (FMD). Recent studies suggested that the RHI may be predictable for mortality in subjects at high risks for cardiovascular events. On the other hand, carotid intima-media thickness (IMT), pulse wave velocity (PWV) and/or Cardio ankle vascular index (CAVI) are known to be used for screening vascular disorders. Since obesity and metabolic syndrome are at risks of cardiovascular events, we aimed to examine the association of the RHI, CAVI, and plaque score (PS) in carotid arteries with body mass index (BMI).

Background: In addition to clinical risk factors, asymptomatic ischemic changes and microbleeds in MRI are known as strong risk factors for stroke event. But it is unclear how accurately it predicts future stroke event when these factors are taken into account all together.

Background: Previously, we demonstrated that the transplantation of human mesenchymal stem cells (B10) decreased the lesion size, neurological deficits, and expression of proinflammatory factors in a rat model of transient middle cerebral artery occlusion (MCAO). Given the multipotency of B10, other mechanisms of B10 transplantation-induced neurological improvements could also exist.

Background: CystatinC (CysC) is an endogenous cysteine protease inhibitor, and expressed in all nucleated cells. This protein is mainly localized in lysosome, but is also secreted in body fluids. L68Q mutation of CysC increases its amyloid aggregation property, leading to Hereditary CysC Cerebral Amyloid Angiopathy. Moreover, CysC is known as a major component of Bunina bodies, an ALS-specific neuronal inclusion body, suggesting its relationship with neurodegeneration.

Background: Ataxia and Male Sterility (AMS) mouse is a cerebellar Purkinje cell (PC) degeneration model, arose spontaneously from MRL/lpr mouse. Genetic studies showed a point mutation in the coding region of Nna1 gene, and homozygous animals show the disease phenotype. The degeneration starts from 3 weeks, and almost all PC are lost within 4 weeks of age along with appearance of ataxia.

Aims: Recent studies suggested that subclasses of high-density lipoprotein (HDL) may be a better biomarker to predict the risk of atherosclerotic disorders. We aimed to examine the association of HDL2- and HDL3-cholesterol (HDL2-C and HDL3-C) with carotid intima-media thickness (IMT) using a new method to quantify the HDL-C subclasses. Methods: Participants were 657 Japanese subjects (434 women) who received a health examination (mean age: 73 years). Serum samples were analyzed by the homogenous assay for HDL-C and HDL3-C. HDL2-C was calculated indirectly by subtracting HDL3-C from HDL-C. HDL3-C measured by this assay was well correlated with that measured by ultracentrifugation (r=0.898, p < 0.001). The maximum IMT (max-IMT) and plaque score (PS) were evaluated by ultrasonography following the standard protocol. Results: HDL3-C was associated with age both in men (r = -0.322, p < 0.0001) and women (r = -0.315, p < 0.0001). In a simple regression analysis, max-IMT showed an inverse association with HDL3-C, whereas no significant association was observed with HDL2-C. A multiple linear regression analysis indicated, however, that the association between HDL3-C and max-IMT was not significant in both aged and younger populations when age was included in the analysis. Further, not only HDL2-C but also HDL3-C was not a significant predictor of 'atherosclerotic arteries' defined as the max-IMT >= 1.5 mm. Similar results were observed in the analysis on PS. Conclusions: Neither HDL3-C nor HDL2-C was significantly associated with carotid atherosclerosis in the Japanese population in this study.

Oligomer formation is considered as a critical process for the neurotoxic effects of Alzheimer's amyloid beta (A beta) peptide. Previously we have demonstrated that lysophosphatidylcholine (LPC) increases the oligomer formation of A beta(1-42), the major A beta peptide found Alzheimer's disease (AD) lesions. In this study, we have investigated whether LPC affects the neurotoxic effects of Ab1-42 in a neuronal cell line (A1) culture. Dimethyl thiazolyl diphenyl tetrazolium (MTT) assay revealed that up to 10 mu M concentration, LPC did not affect A1 cell viability. A beta(1-42) decreased the cell viability, and such effect was dose dependently enhanced by LPC. However, neither LPC nor A beta(1-42), alone or in combination increased lactate dehydrogenase (LDH) release from A1 cells after 24-h treatment. Terminal deoxynucleotidyl transferase dUTP-biotin nick-end-labeling (TUNEL) assay showed that LPC increased A beta(1-42)-induced apoptotic cell number. To determine the underlying mechanisms, the proteins implicated in apoptosis pathways including Bcl-2-and caspase-family were analyzed by Western blotting. The results demonstrated that A beta(1-42) decreased Bcl-2 in A1 cells at 24 h, whereas LPC had no effect at any time point. Both LPC and A beta(1-42) increased Bax level at 24 h, and their combined stimulation showed a synergistic effect. Similar synergistic effect of LPC and A beta(1-42) on caspase9 activation was observed. Dot blot immunoassay and Western blotting showed that LPC augmented A beta(1-42) oligomer formation in cell culture medium. Removing LPC-induced early-formed A beta(1-42) oligomer from the culture medium by immunoprecipitation decreased active caspase9 level and neurotoxicity, as revealed by Western blotting and MTT assay. Furthermore, dihydroethidium (DHE) assay showed that A beta(1-42) increased reactive oxygen species level in A1 cells, such effect was further enhanced by LPC. Thus, our results demonstrated that LPC increased the oligomer formation process of A beta(1-42) peptide in culture condition, and consequently increased apoptotic neuronal death. Such process might be important for the pathogenesis of AD, and inhibition of LPC generation could be a therapeutic target for the disease. (C) 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

Objective - Inflammation has received increasing attention as a cause of stroke. Although several lines of evidence suggest that inflammatory processes have a role in arteriosclerotic vascular events, their involvement remains to be determined. The purpose of this study was to examine the associations between serum high-sensitive C-reactive protein (hs-CRP) levels and cerebral small vessel (CSV)-related lesions as a manifestation of arteriosclerosis. Materials and Methods - Neurologically normal subjects without any history of neurologic or psychiatric diseases were enrolled (n = 519). All the participants underwent magnetic resonance imaging (MRI), and their CSV-related lesions (i.e., lacunar infarcts, cerebral microbleeds, deep white matter hyperintensity, and periventricular hyperintensity) were evaluated. The serum levels of hs-CRP were evaluated as common inflammatory markers. Results - Subjects with higher C-reactive protein (CRP) levels had more lacunar infarcts (P = 0.02). After adjusting for the traditional cardiovascular risk factors, higher hsCRP levels were still associated with the presence of lacunar infarcts [ odds ratio for the highest vs the lowest tertile of hs-CRP, 3.57 (95% confidence interval: 1.30-9.80)]. These associations did not change when the logarithmically transformed values for hs-CRP were included. Furthermore, subjects with higher CRP levels had more cerebral microbleeds (P = 0.03), more severe deep white matter hyperintensity (P = 0.04), and periventricular hyperintensity (P = 0.04); however, these associations were not observed after adjusting for the cardiovascular risk factors. Conclusions - Higher levels of hs-CRP were associated with lacunar infarcts. Thus, inflammatory processes may be involved in the pathogenesis of small-vessel disease.

Neuro-Behçet's disease (NBD) is a serious complication of Behçet's disease. Generally, NBD patients with a chronic course are refractory to immunosuppressive treatment, resulting in the deterioration of personality. In this study, levels of B cell-activating factor belonging to the TNF family (BAFF) were measured in the cerebrospinal fluid (CSF) from 18 patients with NBD, 27 patients with epidemic aseptic meningitis (AM), 24 patients with multiple sclerosis (MS) and 34 healthy controls. BAFF levels in patients with NBD were significantly elevated compared with healthy controls, but showed no statistically significant elevation compared with either of the disease controls. In contrast, CSF IL-6 levels were slightly elevated in patients with NBD and significantly elevated in patients with AM and MS compared with healthy controls. Patients with NBD were subdivided into two groups according to their clinical course (eight patients with a slowly progressive course presenting with psychosis and dementia and 10 patients with an acute course including aseptic meningitis, brainstem involvement and myelopathy). BAFF levels were significantly increased in those with a slowly progressive course compared with those with an acute course. CSF BAFF levels did not correlate with serum BAFF levels, CSF cell counts or CSF IL-6 levels in patients with NBD. These data suggested that BAFF was produced within the central nervous system and may be associated with the development of NBD, particularly with a progressive course.

OBJECTIVES:Dopamine neurotransmission is a critical factor for executive function, which is controlled by the prefrontal cortex in humans. Although the contribution of genetic factors to the regulation of brain dopaminergic activity is widely acknowledged, identification of a genotype-phenotype association has not yet been clearly established. In this study, we therefore evaluated the effects of five functional single-nucleotide polymorphisms (SNPs) in specific genes related to dopamine neurotransmission on executive function in a general population.MATERIALS AND METHODS:Participants of the health examination at the Shimane Institute of Health Science were recruited for this study (n = 964). To evaluate executive function, the Frontal Assessment Battery (FAB) was administered. SNPs were genotyped using the TaqMan method.RESULTS:A significant association was found between an SNP in the catechol-O-methyltransferase (COMT) gene (rs4680) encoding the low-activity Met allele and FAB score (P = 0.003). Of note, the flexibility subset of the FAB was associated with the SNP in COMT (P = 0.003) after adjustment for confounding factors. The generalized multifactor dimensionality reduction method identified that the combination of two SNPs in the COMT gene (rs4680) and the dopamine D4 receptor gene (rs1800955) had a significant effect on FAB score.CONCLUSIONS:Our study indicates a contribution of rs4680 in the COMT gene to the variability in executive function, as assessed by the FAB. In addition, we have indicated that a complex gene-gene interaction between SNPs in the genes related to dopamine neurotransmission may influence executive function in a general population.

OBJECTIVES:To study the involvement of cystatin C in the progression of ischemic white matter lesions (WMLs).MATERIALS AND METHODS:Cystatin C levels in the cerebrospinal fluid (CSF) of patients with cerebrovascular disease, and also in primary and established human neural cell cultures were investigated. For pathologic analysis, cystatin C immunoreactivity was investigated in the white matter of patients with severe WMLs, mild WMLs or controls.RESULTS:Cystatin C levels in the CSF of patients with Fazekas WML grade 3 [14 with hypertension; W/HT(+) and nine without hypertension; W/HT(-)] were lower than those in 38 patients with grade 0-1 (P = 0.0022 and P < 0.0001 respectively). Immunohistochemical study showed that the cystatin C immunoreactivity was found in astrocytes, and the number of astrocytes in the white matter in the severe WML group was decreased when compared with that in controls (P = 0.0027) and in the mild WML group (P = 0.0024). In human neural cell cultures, treatments with thrombin, matrix metalloproteinases and interleukin 1 beta increased the expression of cystatin C mRNA in human astrocytes and hybrid neurons, but an enzyme-linked immunosorbent assay revealed that only thrombin significantly increased the production and secretion of cystatin C in astrocytes.CONCLUSIONS:These results suggest that low levels of CSF cystatin C in ischemic WMLs might be due to the decreased number of astrocytes that secrete cystatin C in response to the stimuli of proteases and inflammatory cytokines.

OBJECTIVES:We evaluated the expression of chemokine-like factor (CKLF) in biopsied muscle fibers in inflammatory myopathies, non-inflammatory myopathies and neurologically diseased controls.MATERIALS AND METHODS:We studied the expression of CKLF in 15 polymyositis (PM), five dermatomyositis (DM), 15 non-inflammatory myopathies and nine neurologically diseased patients by immunohistochemistry.RESULTS:Chemokine-like factor was mostly expressed in small diameter muscle fibers surrounded by infiltrated lymphocytes of inflammatory myopathies patients. Parts of them were also positive for the staining of the developmental form of myosin heavy chain, a maker of regenerating muscle fibers. Thrombin immunoreactivity was observed in endomysium in PM and perimysium in DM. In vitro differentiation study showed a constitutive expression of CKLF in myoblasts that was abolished in myotubes during differentiation process and was induced again by thrombin. Thrombin regulates CKLF expression through protease-activated receptor-1 in myotubes. Treatment of a protein kinase C inhibitor partially blocked CKLF expression in myoblasts, while it remarkably inhibited that in myotubes.CONCLUSION:Chemokine-like factor expression is differentially regulated in myoblasts and myotubes. Thrombin could be a strong regulator for its expression. As CKLF is immunohistochemically positive in regenerating muscle fibers, we postulate here that CKLF is a useful marker for regenerating muscle fibers in inflammatory myopathies.

In the present study, we investigated the expression of protease-activated receptors (PARs), receptors for thrombin, in substantia nigra pars compacta (SNpc) of Parkinson disease (PD) brains and cultures of human neurons, astrocytes, oligodendrocytes, and microglia as determined by immunocytochemistry and reverse transcriptase-polymerase chain reaction (RT-PCR). Expression of PAR-I was demonstrated only in glial fibrillary acidic protein-positive astrocytes in SNpc, and the number of astrocytes expressing PAR-1 increased in SNpc of PD as compared with nonneurologic control brain. Immunoreactivity for thrombin and prothrombin was stronger in astrocytes and the vessel walls in SNpc of PD brains. PAR-1 was expressed in human astrocytes and neurons, but not in oligodendrocytes or microglia as determined by RT-PCR. We investigated thrombin-mediated activation of human astrocytes. Thrombin treatment activates human astrocytes and induces morphologic change and a marked increase in proliferation of astrocytes. Increased expression of glial cell line-derived growth factor and glutathione peroxidase (GPx) but no change in the expression of nerve growth factor and inflammatory cytokines/chemokine (IL-1 beta, IL-6, IL-8, MCPA) was found in thrombin/PAR-activated astrocytes. Next, we studied the neuroprotective effect exerted by thrombin-activated astrocytes in human cerebral neuron X human neuroblastoma hybrid neurons. Although thrombin showed neurotoxicity against human hybrid neurons in a dose-dependent manner, the conditioned media derived from thrombin-pretreated astrocyte cultures promoted the survival of human hybrid neurons. The protective effect was completely inhibited with a GPx inhibitor, mercaptosuccinic acid, indicating that GPx released from thrombin/PAR-activated astrocytes is responsible for neuroprotection of hybrid neurons against thrombin cytotoxicity. The present study suggests that the increased expression of PAR-I in astrocytes in SNpc of PD brain is the restorative move taken by the brain to provide neuroprotection against neuronal degeneration and cell death of dopaminergic neurons caused by noxious insults during the progression of PD pathology.

We report augmentation of somatosensory evoked potentials (SEP) in a patient with frontal alien hand signs after left medial frontal lobe damage. The SEP components occurring later than 30 msec post stimuli were enhanced over the parietal and frontal scalp sites of the lesioned hemisphere. This finding suggests that deficits in inhibitory control of somatosensory processing in parietal and frontal lobes contributes in some way to frontal alien hand signs.

Cystatin C, a cysteine protease inhibitor, is implicated in pathogenesis of late-onset Alzheimer's disease and other neurological disorders. Our recent study showed that cystatin C injection into rat hippocampus induced neuronal cell death in granule cell layer of dentate gyrus in vivo. We further confirmed that cystatin C neurotoxicity was inhibited by simultaneous coapplication of cathepsin B, a cysteine protease. In vitro cytotoxicity was also studied in cultures of human CNS neurons, mixed cultures with astrocytes and A1 human hybrid neurons. Cystatin C induced neuronal cell death in a dose-dependent manner, which accompanied increased number of TUNEL (+) cells, up-regulation of active caspase-3 and DNA ladder. The results of the present study indicate that cystatin C participates in the process of apoptotic neuronal cell death in experimental conditions by means of inhibitory activity of cysteine proteases, and that cystatin C might be involved in the pathogenesis in human neurological disorders including Alzheimer's disease.