A prospective pilot trial of the health data monitoring system without educational content for patients with hypertension.

In Japan, a hypertension treatment aid application (CureApp Co., Ltd.) with educational content on hypertension was the first in the world to show significant blood pressure (BP) reduction in hypertensive patients in 2021. Omron Healthcare Corporation and I developed the Health data Monitoring System (HMS) without educational content, which allows patients to check their home BP values periodically and allows physicians to check their home BP data before the hospital visit. As a pilot study, nineteen patients with hypertension used HMS for six months. The percentage of patients achieving their antihypertensive goal increased from 16% to 37%. Mean home systolic BP decreased from 138.1 ± 11.8 mmHg to 130.2 ± 7.8 mmHg. The increase in number of home BP measurements was significantly associated with the amount of homed systolic BP reduction in men. In conclusion, HMS without educational content may be an adjunct to hypertension treatment.

Rufy3 is an adapter protein for small GTPases that activates a Rac guanine nucleotide exchange factor to control neuronal polarity.

RUN and FYVE domain-containing 3 (Rufy3) is an adapter protein for small GTPase proteins and is bound to activated Rap2, a Ras family protein in the developing neuron. Previously, we reported the presence of a rapid cell polarity determination mechanism involving Rufy3, which is likely required for in vivo neuronal development. However, the molecular details of this mechanism are unclear. To this end, here we produced Rufy3 knock-out (Rufy3-KO) mice to study the role of Rufy3 in more detail. Examining Rufy3-KO neurons, we found that Rufy3 is recruited via glycoprotein M6A to detergent-resistant membrane domains, which are biochemically similar to lipid rafts. We also clarified that Rufy3, as a component of a ternary complex, induces the assembly of Rap2 in the axonal growth cone, whereas in the absence of Rufy3, the accumulation of a Rac guanine nucleotide exchange factor, T-cell lymphoma invasion and metastasis 2 (Tiam2/STEF), is inhibited downstream of Rap2. We also found that Rufy3 regulates the cellular localization of Rap2 and Tiam2/STEF. Taken together, we conclude that Rufy3 is a physiological adapter for Rap2 and activates Tiam2/STEF in glycoprotein M6A-regulated neuronal polarity and axon growth.

Semaphorin3A-induced axonal transport mediated through phosphorylation of Axin-1 by GSK3ß.

The establishment of neuronal polarity is necessary for proper neuronal wiring. Semaphorin3A (Sema3A), originally identified as a repulsive axon guidance molecule, exerts a wide variety of biological functions through signaling pathways including sequential phosphorylation of collapsin response mediator protein by cyclin-dependent kinase-5 (Cdk5) and glycogen synthase kinase-3ß (GSK3ß). Sema3A acts on its receptor neuropilin-1 to regulate axonal transport. To delineate mechanism by which Sema3A induces axonal transport, we investigate whether GSK3ß is involved in mediating Sema3A-induced axonal transport. 4-Benzyl-2-methyl-1,2,4-thiadiazolidine-3,5-dione, an inhibitor of GSK3ß, suppressed Sema3A-induced antero- and retrograde axonal transport. Introduction of either GSK3ß mutants, GSK3ß-L128A or K85M, suppressed Sema3A-induced axonal transport. On the other hand, introduction of GSK3ß-R96A did not affect the Sema3A effect, suggesting that unprimed substrates are primarily involved in Sema3A-induced axonal transport. Overexpression of a partial fragment of frequently rearranged in advanced T-cell lymphomas 1 (FRATtide), which interferes the interaction between GSK3ß and Axis inhibitor-1 (Axin-1), also suppressed Sema3A-induced transport. siRNA knockdown of Axin-1, an unprimed substrate of GSK3ß, suppressed Sema3A-induced antero- and retrograde axonal transport. These results indicate that GSK3ß and Axin-1 are involved in Sema3A-induced bidirectional axonal transport. This finding should provide a clue for understanding of mechanisms of a wide variety of biological activities of Sema3A.

Plexin-A4-dependent retrograde semaphorin 3A signalling regulates the dendritic localization of GluA2-containing AMPA receptors.

The dendritic targeting of neurotransmitter receptors is vital for dendritic development and function. However, how such localization is established remains unclear. Here we show that semaphorin 3A (Sema3A) signalling at the axonal growth cone is propagated towards the cell body by retrograde axonal transport and drives AMPA receptor GluA2 to the distal dendrites, which regulates dendritic development. Sema3A enhances glutamate receptor interacting protein 1-dependent localization of GluA2 in dendrites, which is blocked by knockdown of cytoplasmic dynein heavy chain. PlexinA (PlexA), a receptor component for Sema3A, interacts with GluA2 at the immunoglobulin-like Plexin-transcription-factor domain (PlexA-IPT) in somatodendritic regions. Overexpression of PlexA-IPT suppresses dendritic localization of GluA2 and induces aproximal bifurcation phenotype in the apical dendrites of CA1 hippocampal neurons. Thus, we propose a control mechanism by which retrograde Sema3A signalling regulates the glutamate receptor localization through trafficking of cis-interacting PlexA with GluA2 along dendrites.

GSK3ß/axin-1/ß-catenin complex is involved in semaphorin3A signaling.

Semaphorin3A (Sema3A) exerts a wide variety of biological functions by regulating reorganization of actin and tubulin cytoskeletal proteins through signaling pathways including sequential phosphorylation of collapsin response mediator protein 1 (CRMP1) and CRMP2 by cyclin-dependent kinase-5 and glycogen synthase kinase-3ß (GSK3ß). To delineate how GSK3ß mediates Sema3A signaling, we here determined the substrates of GSK3ß involved. Introduction of either GSK3ß mutants, GSK3ß-R96A, L128A, or K85M into chick dorsal root ganglion (DRG) neurons suppressed Sema3A-induced growth cone collapse, thereby suggesting that unprimed as well as primed substrates are involved in Sema3A signaling. Axin-1, a key player in Wnt signaling, is an unprimed substrate of GSK3ß. The phosphorylation of Axin-1 by GSK3ß accelerates the association of Axin-1 with ß-catenin. Immunocytochemical studies revealed that Sema3A induced an increase in the intensity levels of ß-catenin in the DRG growth cones. Axin-1 siRNA knockdown suppressed Sema3A-induced growth cone collapse. The reintroduction of RNAi-resistant Axin-1 (rAxin-1)-wt rescued the responsiveness to Sema3A, while that of nonphosphorylated mutants, rAxin S322A/S326A/S330A and T485A/S490A/S497A, did not. Sema3A also enhanced the colocalization of GSK3ß, Axin-1, and ß-catenin in the growth cones. The increase of ß-catenin in the growth cones was suppressed by the siRNA knockdown of Axin-1. Furthermore, either Axin-1 or ß-catenin RNAi knockdown suppressed the internalization of Sema3A. These results suggest that Sema3A induces the formation of GSK3ß/Axin-1/ß-catenin complex, which regulates signaling cascade of Sema3A via an endocytotic mechanism. This finding should provide clue for understanding of mechanisms of a wide variety of biological functions of Sema3A.

Semaphorin3A facilitates axonal transport through a local calcium signaling and tetrodotoxin-sensitive voltage-gated sodium channels.

Semaphorin3A (Sema3A), a secreted factor that navigates axons and dendrites of developing neurons, facilitates axonal transport. However, little is known about the mechanism underlying Sema3A-induced facilitation and its functional implications. Here we show that Sema3A induces facilitation of axonal transport via local calcium signaling in growth cone. The facilitation of axonal transport was blocked by inhibitors of voltage-gated sodium channels (tetrodotoxin, TTX), L-type voltage-gated calcium channel, and ryanodine receptor (RyR). Sema3A evoked intracellular Ca(2+) elevation in growth cone by local application of Sema3A to growth cone. Sema3A also activated RyR in growth cone as well as cell body. Notably, TTX suppressed Sema3A-induced RyR activation in cell body but not in growth cone. Our results identify a novel mechanism of Sema3A-induced axonal transport, and further suggest that Sema3A-induced local calcium signaling in growth cone is propagated to cell body in a TTX-sensitive manner.

Cav2.1 in cerebellar Purkinje cells regulates competitive excitatory synaptic wiring, cell survival, and cerebellar biochemical compartmentalization.

In the adult cerebellum, each Purkinje cell (PC) is innervated by a single climbing fiber (CF) in proximal dendrites and 10(5)-10(6) parallel fibers (PFs) in distal dendrites. This organized wiring is established postnatally through heterosynaptic competition between PFs and CFs and homosynaptic competition among multiple CFs. Using PC-specific Cav2.1 knock-out mice (PC-Cav2.1 KO mice), we have demonstrated recently that postsynaptic Cav2.1 plays a key role in the homosynaptic competition by promoting functional strengthening and dendritic translocation of single "winner" CFs. Here, we report that Cav2.1 in PCs, but not in granule cells, is also essential for the heterosynaptic competition. In PC-Cav2.1 KO mice, the extent of CF territory was limited to the soma and basal dendrites, whereas PF territory was expanded reciprocally. Consequently, the proximal somatodendritic domain of PCs displayed hyperspiny transformation and fell into chaotic innervation by multiple CFs and numerous PFs. PC-Cav2.1 KO mice also displayed patterned degeneration of PCs, which occurred preferentially in aldolase C/zebrin II-negative cerebellar compartments. Furthermore, the mutually complementary expression of phospholipase Cß3 (PLCß3) and PLCß4 was altered such that their normally sharp boundary was blurred in the PCs of PC-Cav2.1 KO mice. This blurring was caused by an impaired posttranscriptional downregulation of PLCß3 in PLCß4-dominant PCs during the early postnatal period. A similar alteration was noted in the banded expression of the glutamate transporter EAAT4 in PC-Cav2.1 KO mice. Therefore, Cav2.1 in PCs is essential for competitive synaptic wiring, cell survival, and the establishment of precise boundaries and reciprocity of biochemical compartments in PCs.

Distinct modes of AMPA receptor suppression at developing synapses by GluN2A and GluN2B: single-cell NMDA receptor subunit deletion in vivo.

During development there is an activity-dependent switch in synaptic N-Methyl-D-aspartate (NMDA) receptor subunit composition from predominantly GluN2B to GluN2A, though the precise role of this switch remains unknown. By deleting GluN2 subunits in single neurons during synaptogenesis, we find that both GluN2B and GluN2A suppress AMPA receptor expression, albeit by distinct means. Similar to GluN1, GluN2B deletion increases the number of functional synapses, while GluN2A deletion increases the strength of unitary connections without affecting the number of functional synapses. We propose a model of excitatory synapse maturation in which baseline activation of GluN2B-containing receptors prevents premature synapse maturation until correlated activity allows induction of functional synapses. This activity also triggers the switch to GluN2A, which dampens further potentiation. Furthermore, we analyze the subunit composition of synaptic NMDA receptors in CA1 pyramidal cells, provide electrophysiological evidence for a large population of synaptic triheteromeric receptors, and estimate the subunit-dependent open probability.

Computational analysis of the effects of antineoplastic agents on axonal transport.

Axonal transport plays a crucial role in neuronal morphogenesis, survival, and function. Despite its importance, however, the molecular mechanisms of axonal transport remain mostly unknown because a simple and quantitative assay system for axonal transport has been lacking. In order to better characterize the molecular mechanisms involved in axonal transport, we here developed a computer-assisted monitoring system. Using lipophilic fluorochrome chloromethylbenzamido dialkylcarbocyanine (CM-DiI) as a labeling dye, we have successfully labeled membranous organelles in cultured chick dorsal root ganglia neurons. We confirmed that sodium azide, an ATPase inhibitor, and nocodazole, a microtubule-destabilizing agent, markedly suppressed anterograde and retrograde axonal transport of CM-DiI-labeled particles. We further tested the effects of several anti-neoplastic drugs on axonal transport. Paclitaxel, vincristine, cisplatin, and oxaliplatin, all of which are known to be neurotoxic and to cause neurological symptoms, suppressed anterograde and retrograde axonal transport. Another series of anti-neoplastic drugs, including methotrexate and 5-fluorouracil, did not affect the axonal transport. This is the first report of an automated monitoring system for axonal transport. This system will be useful for toxicity assays, characterizing axonal transport, or screening drugs that may modify neuronal functions.

Mucosal lesions may be a minor complication of SAPHO syndrome: a study of 11 Japanese patients with SAPHO syndrome.

Since the term synovitis-acne-pustulosis-hyperostosis-osteitis (SAPHO) syndrome was proposed by Chamot et al. (Rev Rhum Mal Osteoartic 54:187-196, 1987), clinical reviews concerning this syndrome have been mainly reported from Europe. We carried out a retrospective analysis of 11 Japanese patients with SAPHO syndrome, and reviewed the clinical features of our series in comparison with those in a European large case study. In this study the major features of SAPHO syndrome were chronic osteitis of the anterior chest wall and pustulotic arthro-ostitis with middle age onset, and mucosal lesions seemed to be a minor complication of SAPHO syndrome. The non-erosive peripheral large joints arthritis and the particular HLA types (HLA-B51, B52, or A26), which had been reported to be increased in Behcet's disease, were frequently seen in SAPHO syndrome with mucosal lesions. This study also suggests that SAPHO syndrome with mucosal lesions may be part of a broader disease spectrum including Behcet's disease.

Semaphorin3A alleviates skin lesions and scratching behavior in NC/Nga mice, an atopic dermatitis model.

Topical steroids and antihistamines are commonly used for the treatment of atopic dermatitis (AD). However, in a substantial number of patients with AD, these treatments are not sufficiently effective. In AD patients, C-fibers in the epidermis increase and sprout, inducing hypersensitivity, which is considered to aggravate the disease. Semaphorin3A (Sema3A), an axon guidance molecule, is a potent inhibitor of neurite outgrowth of sensory neurons. To investigate the effect of Sema3A on AD, we administered recombinant Sema3A intracutaneously into the skin lesions of NC/Nga mice, an animal model of AD. Sema3A dose-dependently improved skin lesions and attenuated the scratching behavior in NC/Nga mice. Histological examinations revealed a decrease in: (a) epidermal thickness; (b) the density of invasive nerve fibers in the epidermis; (c) inflammatory infiltrates, including mast cells and CD4+ T cells; and (d) the production of IL-4 in the Sema3A-treated lesions. Because the interruption of the itch-scratch cycle likely contributes to the improvement of the AD-like skin lesions, Sema3A is promising in the treatment of patients with refractory AD, as well as overall itching dermatosis.

Frequent and variable abnormalities in p14 tumor suppressor gene in glioma cell lines.

Ten glioma cell lines were examined for abnormalities of exon 1beta of the p14 gene and then for abnormalities of the entire p14 gene with the use of previous findings of other exons. Abnormalities of exon 1beta and the entire p14 gene were detected in eight of ten cases: homozygous deletion of the entire gene in six cases, hemizygous deletion of exon 1beta with homozygous deletion of downstream exons in one case, and hemizygous deletion of the entire coding region with a missense mutation (A97V) at the C-terminal nucleolar localization domain in one case. The remaining two cases revealed no such abnormalities. p14 gene expression was observed in the latter two cases and one case with A97V mutation in the hemizygously deleted coding region, but not in the others, including one case with only exon 1beta. In the three cases with p14 gene expression, immunocytochemistry revealed p14 nucleolar staining, suggesting the retention of the functional activity of p14 protein and, in the case with the A97V mutation, an insufficient mutational effect as well. The present findings of the frequent and variable p14 gene abnormalities, including rare-type ones with or without sufficient mutational effect in glioma cell lines, might be of value for better understanding of the p14 gene and its related pathways in glioma carcinogenesis.

Gene-environment association of an ITGB2 sequence variant with obesity in ethnic Japanese.

Mice lacking the integrin alphaMbeta2 (Mac-1, CD11b/CD18) develop an obese phenotype on western diet rich in fat. However, no association has been found between variations in the human genes encoding the integrin alphaMbeta2 and obesity. This study was aimed to investigate the association between a single-nucleotide polymorphism (SNP) (rs235326) in the gene encoding human integrin beta2 subunit (ITGB2) with obesity. Our subject cohort comprised 651 people of Japanese ethnicity, of which 274 were Japanese Americans living in Hawaii, and the remaining 377 were native Japanese, two populations in the same genetic background with or without westernized life style. We genotyped the rs235326 polymorphism using a TaqMan assay. In the Japanese-American population, the risk of obesity was found to be 3.29-fold higher (a 95% confidence interval of 1.25-8.67, P = 0.02) in TT homozygotes than in C carriers, using a recessive model and logistic regression analysis that had been adjusted for age. This association was not found in native Japanese individuals. These results indicate that the rs235326 polymorphism in the ITGB2 gene is associated with obesity in Japanese living in the United States whose diet has become "westernized."

Regulation of spine development by semaphorin3A through cyclin-dependent kinase 5 phosphorylation of collapsin response mediator protein 1.

Collapsin response mediator protein 1 (CRMP1) is one of the CRMP family members that mediates signal transduction of axonal guidance and neuronal migration. We show here evidence that CRMP1 is involved in semaphorin3A (Sema3A)-induced spine development in the cerebral cortex. In the cultured cortical neurons from crmp1+/- mice, Sema3A increased the density of clusters of synapsin I and postsynaptic density-95, but this increase was markedly attenuated in crmp1-/- mice. This attenuation was also seen in cyclin-dependent kinase 5 (cdk5)-/- neurons. Furthermore, the introduction of wild-type CRMP1 but not CRMP1-T509A/S522A, (Thr 509 and Ser 522 were replaced by Ala), a mutant that cannot be phosphorylated by Cdk5, into crmp1-/- neurons rescued the defect in Sema3A responsiveness. The Golgi-impregnation method showed that the crmp1-/- layer V cortical neurons showed a lower density of synaptic bouton-like structures and that this phenotype had genetic interaction with sema3A. These findings suggest that Sema3A-induced spine development is regulated by phosphorylation of CRMP1 by Cdk5.

Good syndrome with thymic adenosquamous carcinoma--report of a case.

A 68-year-old man with recurrent bilateral severe pneumonia and invasive thymic carcinoma was admitted to our hospital. An extended thymo-thymectomy with lymph nodes dissection was performed for an irregular shaped anterior mediastinum mass. The tumor was mainly composed of type C, adenosquamous carcinoma, and found to have a small area of types B2 and B3 thymoma. History and laboratory findings were compatible with the diagnosis of Good syndrome. Although there are some reports of thymic carcinoma arising from thymoma, this is the first report of co-existence of adenosquamous carcinomas and thymoma with Good syndrome as far as reviewed articles. Thymic carcinoma with severe infection should be examined carefully for co-existence of thymoma, and co-existence of thymoma and thymic carcinoma suggests a close histogenetic relationship between the 2 tumors.

Regulation of dendritic branching and spine maturation by semaphorin3A-Fyn signaling.

A member of semaphorin family, semaphorin3A (Sema3A), acts as a chemorepellent or chemoattractant on a wide variety of axons and dendrites in the development of the nervous systems. We here show that Sema3A induces clustering of both postsynaptic density-95 (PSD-95) and presynaptic synapsin I in cultured cortical neurons without changing the density of spines or filopodia. Neuropilin-1 (NRP-1), a receptor for Sema3A, is present on both axons and dendrites. When the cultured neurons are exposed to Sema3A, the cluster size of PSD-95 is markedly enhanced, and an extensive colocalization of PSD-95 and NRP-1 or actin-rich protrusion is seen. The effects of Sema3A on spine morphology are blocked by PP2, an Src type tyrosine kinase inhibitor, but not by the PP3, the inactive-related compound. In the cultured cortical neurons from fyn(-/-) mice, dendrites bear few spines, and Sema3A does not induce PSD-95 cluster formation on the dendrites. Sema3A and its receptor genes are highly expressed during the synaptogenic period of postnatal days 10 and 15. The cortical neurons in layer V, but not layer III, show a lowered density of synaptic bouton-like structure on dendrites in sema3A- and fyn-deficient mice. The neurons of the double-heterozygous mice show the lowered spine density, whereas those of single heterozygous mice show similar levels of the spine density as the wild type. These findings suggest that the Sema3A signaling pathway plays an important role in the regulation of dendritic spine maturation in the cerebral cortex neurons.

Intracellular activation of trypsinogen in rat pancreatic acini after supramaximal secretagogue stimulation: cysteine protease and serine protease activity.

OBJECTIVE: The aim of the study was to explore the mechanism by which trypsinogen becomes activated during acute pancreatitis. METHODS: Given the ability of cholecystokinin (CCK) to induce pancreatitis in vivo, the effects of high-dose CCK on preparations of isolated pancreatic acini were examined using immunofluorescence techniques for the detection of trypsinogen activation. Acini were pretreated with weak base or serine or cysteine protease inhibitors before CCK hyperstimulation. RESULTS: CCK was found to stimulate the generation of trypsinogen activation peptide (TAP), a marker for trypsinogen processing. The generation of TAP was inhibited by pretreatment with a weak base, chloroquine (40 micromol/L). TAP generation was also inhibited by pretreatment with serine protease inhibitor FUT-175 (1 micromol/L) but not cysteine protease inhibitor E64 (0.1 mmol/L) or E64-d (0.1 mmol/L). Although treatment with a high dose of E64-d (1 mmol/L) reduced activation, it also caused cell injury. CONCLUSIONS: High-dose CCK stimulated the intracellular activation of trypsinogen within isolated pancreatic acini. This event appears to require an acidic subcellular compartment and serine protease activity. The role for thiol proteases in this model remains unclear.

Effects of pancreatic duct ligation on pancreatic response to bombesin.

To examine mechanisms that might be related to biliary pancreatitis, we examined the effects of pancreatic duct ligation (PDL) with pancreatic stimulation in vivo. PDL alone caused no increase in pancreatic levels of trypsinogen activation peptide (TAP), trypsin, or chymotrypsin and did not initiate pancreatitis. Although bombesin caused zymogen activation within the pancreas, the increases were slight and it did not cause pancreatitis. However, the combination of PDL with bombesin resulted in prominent increases in pancreatic TAP, trypsin, chymotrypsin, and the appearance of TAP in acinar cells and caused pancreatitis. Disruption of the apical actin network in the acinar cell was observed when PDL was combined with bombesin but not with PDL or bombesin alone. These studies suggest that when PDL is combined with pancreatic acinar cell stimulation, it can promote zymogen activation, the retention of active enzymes in acinar cells, and the development of acute pancreatitis.