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Whole genome analysis has identified rare copy number variations (CNV) that are strongly involved in the pathogenesis of psychiatric disorders, and 3q29 deletion has been found to have the largest effect size. The 3q29 deletion mice model (3q29-del mice) has been established as a good pathological model for schizophrenia based on phenotypic analysis; however, circadian rhythm and sleep, which are also closely related to neuropsychiatric disorders, have not been investigated. In this study, our aims were to reevaluate the pathogenesis of 3q29-del by recreating model mice and analyzing their behavior and to identify novel new insights into the temporal activity and temperature fluctuations of the mouse model using a recently developed small implantable accelerometer chip, Nano-tag. We generated 3q29-del mice using genome editing technology and reevaluated common behavioral phenotypes. We next implanted Nano-tag in the abdominal cavity of mice for continuous measurements of long-time activity and body temperature. Our model mice exhibited weight loss similar to that of other mice reported previously. A general behavioral battery test in the model mice revealed phenotypes similar to those observed in mouse models of schizophrenia, including increased rearing frequency. Intraperitoneal implantation of Nano-tag, a miniature acceleration sensor, resulted in hypersensitive and rapid increases in the activity and body temperature of 3q29-del mice upon switching to lights-off condition. Similar to the 3q29-del mice reported previously, these mice are a promising model animal for schizophrenia. Successive quantitative analysis may provide results that could help in treating sleep disorders closely associated with neuropsychiatric disorders.

Aim A Japanese individual with schizophrenia harboring a novel exonic deletion in RELN was recently identified by genome-wide copy-number variation analysis. Thus, the present study aimed to generate and analyze a model mouse to clarify whether Reln deficiency is associated with the pathogenesis of schizophrenia. Methods A mouse line with a novel RELN exonic deletion (Reln-del) was established using the CRISPR/Cas9 method to elucidate the underlying molecular mechanism. Subsequently, general behavioral tests and histopathological examinations of the model mice were conducted and phenotypic analysis of the cerebellar granule cell migration was performed. Results The phenotype of homozygous Reln-del mice was similar to that of reeler mice with cerebellar atrophy, dysplasia of the cerebral layers, and abrogated protein levels of cerebral reelin. The expression of reelin in heterozygous Reln-del mice was approximately half of that in wild-type mice. Conversely, behavioral analyses in heterozygous Reln-del mice without cerebellar atrophy or dysplasia showed abnormal social novelty in the three-chamber social interaction test. In vitro reaggregation formation and neuronal migration were severely altered in the cerebellar cultures of homozygous Reln-del mice. Conclusion The present results in novel Reln-del mice modeled after our patient with a novel exonic deletion in RELN are expected to contribute to the development of reelin-based therapies for schizophrenia.

Schizophrenia (SCZ) is known to be a heritable disorder; however, its multifactorial nature has significantly hampered attempts to establish its pathogenesis. Therefore, in this study, we performed genome-wide copy-number variation (CNV) analysis of 2940 patients with SCZ and 2402 control subjects and identified a statistically significant association between SCZ and exonic CNVs in the ARHGAP10 gene. ARHGAP10 encodes a member of the RhoGAP superfamily of proteins that is involved in small GTPase signaling. This signaling pathway is one of the SCZ-associated pathways and may contribute to neural development and function. However, the ARHGAP10 gene is often confused with ARHGAP21, thus, the significance of ARHGAP10 in the molecular pathology of SCZ, including the expression profile of the ARHGAP10 protein, remains poorly understood. To address this issue, we focused on one patient identified to have both an exonic deletion and a missense variant (p.S490P) in ARHGAP10. The missense variant was found to be located in the RhoGAP domain and was determined to be relevant to the association between ARHGAP10 and the active form of RhoA. We evaluated ARHGAP10 protein expression in the brains of reporter mice and generated a mouse model to mimic the patient case. The model exhibited abnormal emotional behaviors, along with reduced spine density in the medial prefrontal cortex (mPFC). In addition, primary cultured neurons prepared from the mouse model brain exhibited immature neurites in vitro. Furthermore, we established induced pluripotent stem cells (iPSCs) from this patient, and differentiated them into tyrosine hydroxylase (TH)-positive neurons in order to analyze their morphological phenotypes. TH-positive neurons differentiated from the patient-derived iPSCs exhibited severe defects in both neurite length and branch number; these defects were restored by the addition of the Rho-kinase inhibitor, Y-27632. Collectively, our findings suggest that rare ARHGAP10 variants may be genetically and biologically associated with SCZ and indicate that Rho signaling represents a promising drug discovery target for SCZ treatment.

Paranodal axo-glial junctions (PNJs) play an essential role in the organization and maintenance of molecular domains in myelinated axons. To understand the importance of PNJs better, we investigated cerebroside sulfotransferase (CST; a sulfatide synthetic enzyme)-deficient mice, which partially lack PNJs in both the central nervous system (CNS) and the peripheral nervous system (PNS). Previously, we reported that axonal mitochondria at the nodes of Ranvier in the PNS were large and swollen in CST-deficient mice. Although we did not observed significant defects in the nodal regions in several areas of the CNS, myelinated internodal regions showed many focal swellings in Purkinje cell axons in the cerebellum, and the number and the size of swellings increased with age. In the present analysis of various stages of the swellings in 4-12-week-old mutant mice, calbindin-positive axoplasm swellings started to appear at an early stage. After that, accumulation of neurofilament and mitochondria gradually increased, whereas deposition of amyloid precursor protein became prominent later. Ultrastructural analysis showed accumulations of tubular structures closely resembling smooth endoplasmic reticulum (ER). Staining of cerebellar sections of the mutant mice for type I inositol 1,4,5-trisphosphate receptor (IP3 R1) revealed high immunoreactivity within the swellings. This IP3 R1 deposition was the initial change and was not observed in development prior to the onset of myelination. This suggests that local calcium regulation through ER was involved in these axonal swellings. Therefore, in addition to the biochemical composition of the internodal myelin sheath, PNJs might also affect maintenance of axonal homeostasis in Purkinje cells.

The autophagy pathway has recently been implicated in several neurodegenerative diseases. Recently, it was reported that chorein-depleted cells showed accumulation of autophagic markers and impaired autophagic flux. Here, we demonstrate that chorein overexpression preserves cell viability from starvation-induced cell death in human embryonic kidney 293 (HEK293) cells. Subsequent coimmunoprecipitation and reverse coimmunoprecipitation assays using extracts from chorein that stably overexpressed HEK293 cells revealed that chorein interacts with α-tubulin and histone deacetylase 6, a known α-tubulin deacetylater and central component of basal autophagy. Indeed, acetylated α-tubulin immunoreactivity was significantly decreased in chorein that stably overexpressed HEK293 cells. These results suggest that chorein/histone deacetylase 6/α-tubulin interactions may play an important role in starvation-induced cell stress, and their disruption may be one of the molecular pathogenic mechanisms of chorea-acanthocytosis.-Sasaki, N., Nakamura, M., Kodama, A., Urata, Y., Shiokawa, N., Hayashi, T., Sano, A. Chorein interacts with α-tubulin and histone deacetylase 6, and overexpression preserves cell viability during nutrient deprivation in human embryonic kidney 293 cells.

Paranodal axo-glial junctions (PNJs) play an essential role in the organization and maintenance of molecular domains in myelinated axons. To understand the importance of PNJs better, we investigated cerebroside sulfotransferase (CST; a sulfatide synthetic enzyme)-deficient mice, which partially lack PNJs in both the central nervous system (CNS) and the peripheral nervous system (PNS). Previously, we reported that axonal mitochondria at the nodes of Ranvier in the PNS were large and swollen in CST-deficient mice. Although we did not observed significant defects in the nodal regions in several areas of the CNS, myelinated internodal regions showed many focal swellings in Purkinje cell axons in the cerebellum, and the number and the size of swellings increased with age. In the present analysis of various stages of the swellings in 4-12-week-old mutant mice, calbindin-positive axoplasm swellings started to appear at an early stage. After that, accumulation of neurofilament and mitochondria gradually increased, whereas deposition of amyloid precursor protein became prominent later. Ultrastructural analysis showed accumulations of tubular structures closely resembling smooth endoplasmic reticulum (ER). Staining of cerebellar sections of the mutant mice for type I inositol 1,4,5-trisphosphate receptor (IP3 R1) revealed high immunoreactivity within the swellings. This IP3 R1 deposition was the initial change and was not observed in development prior to the onset of myelination. This suggests that local calcium regulation through ER was involved in these axonal swellings. Therefore, in addition to the biochemical composition of the internodal myelin sheath, PNJs might also affect maintenance of axonal homeostasis in Purkinje cells.

子宮頸管ポリープに上皮内癌および微小浸潤扁平上皮癌を認めた各症例を経験したので報告する. 症例1は61歳. 人間ドックでの子宮頸部スメア異常を主訴として, 当科を受診. 外来受診時のスメアはclass IVであり, 径5mmの頸管ポリープを切除したところ, 組織学的に微小浸潤扁平上皮癌と診断された. 症例2は37歳. 集団検診にて子宮頸部スメア異常を指摘され当科を紹介初診となった. 初診時のスメアはclass III aであった. 径6mmの頸管ポリープ表面に白色上皮を認め, ポリープ切除を施行したところ, 組織学的に上皮内癌と診断された. 子宮頸管ポリープでは悪性の頻度は小さいが, ときに癌を合併することがあり, 頸管ポリープを切除した場合には組織学的診断を必ず行うことが必要であるといえる.

単純子宮全摘出術の4年後に腟断端に再発した子宮頸部悪性腺腫 (adenoma malignum) を報告する. 症例は41歳, 1回経妊1回経産婦で, 腟断端腫瘤の生検標本に腺癌が認められたため, 当科へ紹介され受診した. 初診時, 腟断端左側に数個の小豆大腫瘤を認め転移性腺癌を疑い, CTによる全身検索を施行したが, 原発巣と考えられる腫瘍は他に認められなかった. 4年前の摘出子宮標本を再鏡検したところ, 一部の頸管腺は既存の頸管腺領域を越えて小嚢胞を形成しており, その組織像は腟断端腫瘤の生検組織像ときわめて類似するものであった. 免疫組織化学的検索では, 腫瘍細胞の腺腔側の細胞膜上にCEA陽性所見が認められ, 4年前の子宮頸部標本をadenoma malignumと診断した.

The property of intensive 45Ca2+ uptake by A-431 human epidermoidal carcinoma cells was indicated to be an influx, not binding to the cell surface, since the two apparent dissociation constants (Kd) between 45Ca2+ and cells were almost the same when measured in either the presence or absence of 1 mM [ethylenebis (oxyethylenenitrilo)]tetraacetic acid (EGTA); these constants were approximately 5-10 x 10(-6) and 1 x 10(-4) M, respectively, which are much higher than the chelating constant of EGTA for Ca2+ (approximately 10(-11) M). Furthermore, addition of A23187, a calcium ionophore, rapidly released the 45Ca2+ incorporated into cells at both 37 degrees C and 0 degrees C. The 45Ca2+ associated with the cells was slowly released or exchanged when cells were incubated in medium depleted of Ca2+, or in that containing 1 mM non-radioactive Ca2+. The ability of A-431 cells to respond to extracellular ATP by elevating their level of intracellular calcium ions, as well as by producing inositol trisphosphate (InsP3), was suppressed in cells depleted of cellular calcium. These data suggest that calcium ions are extensively incorporated or exchanged with those outside the cells, maintained as stored calcium, and involved in production of InsP3, when A-431 cells are stimulated by ATP to trigger the signal transduction system.

The alpha catalytic subunits of Na+/K(+)-ATPase were isolated from the kidney and brain of rats (alpha 1 and alpha 2, respectively). The antisera raised against these subunits were used as probes to analyze the isoform of catalytic subunits of Na+/K(+)-ATPase in various tissues of rats. Of 27 rat tissues examined, most had a catalytic subunit identical to alpha 1 but some, such as the nervous and muscle tissues, had both alpha 1 and alpha 2 isoforms as judged by their reactivities to antisera and their electrophoretic mobility. We found that the submandibular gland contained a new electrophoretic variant of immunoreactive alpha subunit (designated alpha(S) in this report) in addition to alpha 1 identical to those found in kidney and brain. The new variant, alpha(S), strongly cross-reacted with anti-alpha 1 antiserum, but to a lesser extent with anti-alpha 2 antiserum. The alpha(S) had a molecular mass which was found to be slightly less (approx. 90 kDa) than brain and kidney alpha 1. We examined whether or not the alpha(S) is formed by proteolytic cleavage of alpha subunits during preparation and concluded that this is not the case. The alpha(S) reacted with [gamma-32P]ATP, resulting in the formation of radioactive alpha subunit which was stabilized by 2 mM ouabain but which was labile in the presence of 70 mM potassium chloride. Since N-terminal amino acid sequence of alpha(S) protein [G()DKY()PAAVS] corresponds exactly and uniquely with the sequence of the alpha 1 chain between residues 1 and 11, it is very probable that alpha(S) protein originated from alpha 1 protein following the post-translational processing.

A short step synthesis of cyclopentatrienylidene retinals [(9)-(11)] is reported and the formation of the novel rhodopsin analogue (12) from the isomer (9) is described.

The structure of a new chromophore in II has been confirmed by the synthesis of the tetraene III containing the tetrahydroindene skeleton.

The simultaneous determination of vitamin D2 and its isomers by high-speed liquid chromatography (HSLC) is described. Preferred operating conditions for the resolution and quantitation of vitamin D2, 5, 6-traps-vitamin D2, ergosterol, lumisterol2, isovitamin D2, isotachysterol2, tachysterol2, and previtamin D2 are as follows: column, “Zorbax” SIL (25cm×2.1mm i.d.); pressure, 100-120kg/cm2; temperature, ambient; detector, UV 254nm; mobile phase, 0.15% methanol+2% ether in pentane (for the simultaneous determination), 10% ether in hexane (for quantitation of the practical sets of isomers), or 55% CHCl3 (distilled) in pentane (for separation of isovitamin D2 and 5, 6-traps-vitamin D2); sample size, 1 μl (>1-5 ng); internal standard, p-cresol or α-naphthol. High-speed, good resolution, precise and accurate trace analysis, greater analysis flexibility, no necessity for preparing derivatives, and gentle operating conditions are main potential advantages of our method which has proved to be very efficient and surpasses all analytical procedures hitherto proposed.

The title compound (3) which involves a new conjugated system, has been synthesised in a geometrically pure form the indanone (4).

The photochemical behaviour of 9-cis-retro-γ-retinal (II) was investigated by high-performance liquid chromatography. A new isomer, 9, 13-di-cis-retro-γ-retinal (III) was isolated from the irradiation products of II.

During the studies on a photoreaction of retinal, involving various kinds of (Z)-(E) isomerization, a heretofore unknown photo product of retinal was isolated in a pure state and was characterized unambiguously. Thus, direct irradiation of all-(E)-retinal (I) and of all-(E)-β-ionylidenecrotonaldehyde (II) in acetonitrile solution gave the corresponding 6e-electrocyclized photoproducts, (III) and (IV), both via the possible 7-(Z)-isomer intermediates of the parent conjugated poly enals. Unlike the lower members in the retinal series, it was also con firmed that sigmatropic rearrangement or photo-Diels-Alder reaction hardly proceeds in these higher members of the series mentioned above.

The photochemical reaction of cyclopentatrienylidene 11-cis-locked-rhodopsin derived from cyclopentatrienylidene 11-cis-locked-retinal and cattle opsin was spectrophotometrically studied. The difference absorption spectrum between the cyclopentatrienylidene 11-cis-locked-rhodopsin and its retinal oxime had its maximum at 495 nm (P-495). Irradiation of P-495 at -196 degrees C with either blue light or orange light caused no spectral change, supporting the cis-trans isomerization hypothesis for formation of bathorhodopsin. Upon irradiation of P-495 at 0 degree C with orange light, however, its absorption spectrum shifted to a shorter wavelength owing to formation of a hypsochromic product. The difference absorption spectrum between this product (P-466) and its retinal oxime showed its maximum at 466 nm. Analysis of retinal isomers by high-performance liquid chromatography showed that this spectral shift was not accompanied by photoisomerization of the chromophore. P-466 could almost completely be photoconverted to the original pigment (P-495) by irradiation at 0 degree C with blue light with little formation of the other isomeric form of its chromophore. The alpha-band of the circular dichroism spectrum of P-495 was very small in comparison with that of rhodopsin, while that of P-466 was comparable to it. These facts suggest that P-495 has a planar conformation in the side chain of the chromophore and that P-466 has a twisted one, probably at the C8-C9 single bond. Cyclic-GMP phosphodiesterase in frog rod outer segment was activated by neither P-495 nor P-466. This result suggests that the isomerization of the retinylidene chromophore of rhodopsin is indispensable in the phototransduction process.

9-cis-Retro-gamma-rhodopsin (lambda max = 420 nm) was prepared from 9-cis-retro-gamma-retinal and cattle opsin. After cooling to liquid nitrogen temperature (77 K), the pigment was irradiated with light at 380 nm. The spectrum shifted to the longer wavelengths, owing to formation of a batho product. This fact indicates that the conjugated double bond system from C-5 to C-8 of the chromophoric retinal in rhodopsin was not necessary for formation of bathorhodopsin. Reirradiation of the batho product with light at wavelengths longer than 520 nm yielded a mixture composed of presumably 9- or 11-cis forms of retro-gamma-rhodopsin. These three isomers are interconvertible by light at liquid nitrogen temperature. Thus the retro-gamma-rhodopsin system is similar in photochemical reaction at 77 K to cattle rhodopsin system. Each system has its own batho product. Based on these results, it was infered that the formation of batho-rhodopsin is due to photoisomerization of the chromophoric retinal of rhodopsin and is not due to translocation of a proton on the ring or on the side chain from C-6 to C-8 of the chromophoric retinal to the Schiff-base nitrogen.

Spectral characterization of 5,6-epoxyretinal isomers (II) prepared by a regioselective epoxidation of the parent retinal isomers have been described. High-performance liquid chromatographic separation of epoxyretinals and a photoisomerization behavior of all-trans-5,6-epoxyretinals (IIa) have been investigated.

A study of nonbacterial thrombotic endocarditis has been carried out in a series of 3,404 autopsies performed upon atomic bomb survivors in Hiroshima in the period 1953-1970. The prevalence of the lesion was 2.4 per cent, with a greater frequency among the elderly and among females, and with a significant relationship to malignant neoplasms. In contrast to other reported series, there was a greater prevalence among cancers of the colon, rectum, and female genitourinary tract. No relationship was noted between the presence of NBTE and exposure to ionizing radiation. Histologic findings in the heart-valve leaflets in close proximity to the verrucae, like experimental studies reported by others, suggest that in association with severe systemic disease there appears a process consisting of degenerative changes in valve collagen and ground substance, with subsequent denudation of endothelium, localized almost entirely to the apposing leaflet surfaces of the left-heart valves. The verrucae of nonbacterial thrombotic endocarditis are then formed upon this abnormal leaflet surface. While the relationship between systemic disease and the pathologic changes observed in cardiac valve tissue is unclear, and although it is not known whether a "hypercoagulable state" may accentuate the tendency for thrombi to form upon these abnormal valves, there is no doubt that this lesion represents a clinically important complication of severe systemic disease. It also seems likely that in some cases NBTE may complicate an illness which may otherwise be curable. Increasing awareness of this pathologic entity among clinicians, coupled with appropriate laboratory techniques, most likely echocardiography, will permit more frequent diagnosis in living patients.

A high-performance liquid chromatographic method for the simultaneous determination of geometric isomers of retinal in a mixture has been developed. The method can be applied to problems such as determination of optimal conditions for the preparation of a particular isomer, or the investigation of novel photoproducts from retinal. Aprotic solvent effects on cis-trans isomerization are discussed.

The alpha and alpha(+) isoforms of Na+,K(+)-ATPase were isolated from the kidney and brain of rats and purified. Their antisera were raised to analyze the alpha isoforms in rat tissues. We found that the submandibular gland (SMG) contains a new immunoreactive alpha subunit isoform, designated alpha(S) in this report, in addition to alpha identical with those found in the kidney or brain. The new alpha(S) strongly reacted with anti-alpha-antiserum but to a much lesser extent with anti-alpha(+)-antiserum. The alpha(S) had a slightly lower molecular weight (approximately 90,000) than the brain and kidney alpha isoforms. Various fractions of SMG tissues were added to the SMG microsomes and incubated in order to test whether or not the alpha(S) is formed artificially; no increase of alpha(S) was observed by these treatments, suggesting that the alpha(S) was not the product formed from alpha during the preparation of microsome sample, but was rather a protein originally present in the SMG. The alpha(S) protein was not detected in the SMG of 2- or 5-week-old rats, but it gradually increased in rats older than 8 weeks, reaching the maximum in 30-week-old animals. The Na+,K(+)-ATPase activity in the SMG increased concomitantly with the increase of alpha(S), indicating that Na+,K(+)-ATPase comprising alpha(S) also shows enzyme activity; it is speculated that alpha(S) may have some unique and unknown function(s) in older rats.

Homologues of retinoic acid are converted by acid-catalysed cyclization into hydrindene compounds bearing a non-conjugated (αβ-saturated) enoic acid (or lactone) side-chain. Explanations are given for the colour reaction including structural elucidation of the quenched products and their mechanism of formation.

The 11-cis-locked-cyclopentatrienylideneretinals (7a), (7b), (8a), (8b), and (9) have been obtained in a short-path synthesis from the β-ionyl sulphone (3). Their characteristic spectral properties (u.v. and 1H n.m.r.) are discussed.