Search

The tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has emerged as a promising anticancer agent with high tumor-selective cytotoxicity. The congenital and acquired resistance of some cancer types including malignant melanoma and osteosarcoma impede the current TRAIL therapy of these cancers. Since fine tuning of the intracellular Ca2+ level is essential for cell function and survival, Ca2+ dynamics could be a promising target for cancer treatment. Recently, we demonstrated that mitochondrial Ca2+ removal increased TRAIL efficacy toward malignant melanoma and osteosarcoma cells. Here we report that mitochondrial Ca2+ overload leads to tumor-selective sensitization to TRAIL cytotoxicity. Treatment with the mitochondrial Na+/Ca2+ exchanger inhibitor CGP-37157 and oxidative phosphorylation inhibitor antimycin A and FCCP resulted in a rapid and persistent mitochondrial Ca2+ rise. These agents also increased TRAIL sensitivity in a tumor-selective manner with a switching from apoptosis to a nonapoptotic cell death. Moreover, we found that mitochondrial Ca2+ overload led to increased mitochondrial fragmentation, while mitochondrial Ca2+ removal resulted in mitochondrial hyperfusion. Regardless of their reciprocal actions on the mitochondrial dynamics, both interventions commonly exacerbated TRAIL-induced mitochondrial network abnormalities. These results expand our previous study and suggest that an appropriate level of mitochondrial Ca2+ is essential for maintaining the mitochondrial dynamics and the survival of these cells. Thus, disturbing mitochondrial Ca2+ homeostasis may serve as a promising approach to overcome the TRAIL resistance of these cancers with minimally compromising the tumor-selectivity.

Ca2+ has emerged as a new target for cancer treatment since tumor-specific traits in Ca2+ dynamics contributes to tumorigenesis, malignant phenotypes, drug resistance, and survival in different tumor types. However, Ca2+ has a dual (pro-death and pro-survival) function in tumor cells depending on the experimental conditions. Therefore, it is necessary to minimize the onset of the pro-survival Ca2+ signals caused by the therapy. For this purpose, a better understanding of pro-survival Ca2+ pathways in cancer cells is critical. Here we report that Ca2+ protects malignant melanoma (MM) and osteosarcoma (OS) cells from tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) cytotoxicity. Simultaneous measurements using the site-specific Ca2+ probes showed that acute TRAIL treatment rapidly and dose-dependently increased the cytosolic Ca2+ concentration ([Ca2+]cyt) and mitochondrial Ca2+ concentration ([Ca2+]mit) Pharmacological analyses revealed that the [Ca2+]mit remodeling was under control of mitochondrial Ca2+ uniporter (MCU), mitochondrial permeability transition pore (MPTP), and a Ca2+ transport pathway sensitive to capsazepine and AMG9810. Ca2+ chelators and the MCU inhibitor ruthenium 360, an MPTP opener atractyloside, capsazepine, and AMG9810 all decreased [Ca2+]mit and sensitized these tumor cells to TRAIL cytotoxicity. The Ca2+ modulation enhanced both apoptotic and non-apoptotic cell death. Although the [Ca2+]mit reduction potentiated TRAIL-induced caspase-3/7 activation and cell membrane damage within 24 h, this potentiation of cell death became pronounced at 72 h, and not blocked by caspase inhibition. Our findings suggest that in MM and OS cells mitochondrial Ca2+ removal can promote apoptosis and non-apoptotic cell death induction by TRAIL. Therefore, mitochondrial Ca2+ removal can be exploited to overcome the resistance of these cancers to TRAIL.

The ventral striatum is involved in motivated behavior. Akin to the dorsal striatum, the ventral striatum contains two parallel pathways: the striatomesencephalic pathway consisting of dopamine receptor Type 1-expressing medium spiny neurons (D1-MSNs) and the striatopallidal pathway consisting of D2-MSNs. These two genetically identified pathways are thought to encode opposing functions in motivated behavior. It has also been reported that D1/D2 genetic selectivity is not attributed to the anatomical discrimination of two pathways. We wanted to determine whether D1- and D2-MSNs in the ventral striatum functioned in an opposing manner as previous observations claimed, and whether D1/D2 selectivity corresponded to a functional segregation in motivated behavior of mice. To address this question, we focused on the lateral portion of ventral striatum as a region implicated in food-incentive, goal-directed behavior, and recorded D1 or D2-MSN activity by using a gene-encoded ratiometric Ca 2+ indicator and by constructing a fiberphotometry system, and manipulated their activities via optogenetic inhibition during ongoing behaviors. We observed concurrent event-related compound Ca 2+ elevations in ventrolateral D1- and D2-MSNs, especially at trial start cue-related and first lever press-related times. D1 or D2 selective optogenetic inhibition just after the trial start cue resulted in a reduction of goal-directed behavior, indicating a shared coding of motivated behavior by both populations at this time. Only D1-selective inhibition just after the first lever press resulted in the reduction of behavior, indicating D1-MSN-specific coding at that specific time. Our data did not support opposing encoding by both populations in food-incentive, goal-directed behavior. SIGNIFICANCE STATEMENT An opposing role of dopamine receptor Type 1 or Type 2-expressing medium spiny neurons (D1-MSNs or D2-MSNs) on striatum-mediated behaviors has been widely accepted. However, this idea has been questioned by recent reports. In the present study, we measured concurrent Ca 2+ activity patterns of D1- and D2-MSNs in the ventrolateral striatum during food-incentive, goal-directed behavior in mice. According to Ca 2+ activity patterns, we conducted timing-specific optogenetic inhibition of each type of MSN. We demonstrated that both D1- and D2-MSNs in the ventrolateral striatum commonly and positively encoded action initiation, whereas only D1-MSNs positively encoded sustained motivated behavior. These findings led us to reconsider the prevailing notion of a functional segregation of MSN activity in the ventral striatum., (Copyright © 2017 the authors 0270-6474/17/372724-11$15.00/0.)

Impaired motivation is present in a variety of neurological disorders, suggesting that decreased motivation is caused by broad dysfunction of the nervous system across a variety of circuits. Based on evidence that impaired motivation is a major symptom in the early stages of Huntington's disease, when dopamine receptor type 2-expressing striatal medium spiny neurons (D2-MSNs) are particularly affected, we hypothesize that degeneration of these neurons would be a key node regulating motivational status. Using a progressive, time-controllable, diphtheria toxin-mediated cell ablation/dysfunction technique, we find that loss-of-function of D2-MSNs within ventrolateral striatum (VLS) is sufficient to reduce goal-directed behaviours without impairing reward preference or spontaneous behaviour. Moreover, optogenetic inhibition and ablation of VLS D2-MSNs causes, respectively, transient and chronic reductions of goal-directed behaviours. Our data demonstrate that the circuitry containing VLS D2-MSNs control motivated behaviours and that VLS D2-MSN loss-of-function is a possible cause of motivation deficits in neurodegenerative diseases., Competing Interests: The authors declare no competing financial interests.

Approaches to investigate gene functions in experimental biology are becoming more diverse and reliable. Furthermore, several kinds of tissues and organs that possess their original identities can be generated in petri dishes from stem cells including embryonic, adult and induced pluripotent stem cells. Researchers now have several choices of experimental methods and their combinations to analyze gene functions in various biological systems. Here, as an example we describe one of the better protocols, which combines three-dimensional embryonic stem cell culture with small regulatory RNA-mediated technologies, clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9), and inducible RNA interference (RNAi). This protocol allows investigation of genes of interest to better understand gene functions in target tissues (or organs) during in vitro development.

Background: Functional magnetic resonance imaging (fMRI) in mice is typically performed under anesthesia due to difficulties in holding the head of awake mice stably with a conventional three-point fixation method that uses a tooth-bar and earplugs. Although some studies have succeeded in fMRI in awake mice by attaching a head-post on the skull, this cannot be applied to fMRI using a high signal-to-noise ratio (SNR) cryogenic MRI-detector, CryoProbe, because it covers the head of a mouse closely., New Method: We developed head-fixation implements for awake mice that are applicable to fMRI using CryoProbe., Results: A head-bar was surgically attached to the skull of a mouse that was then habituated to a mock fMRI-environment, two hours/day for eight days with physiological examinations of body-weight, fecal weight, electromyogram (EMG), and electrocardiogram. EMG power decreased with just one day of habituation, whereas heart rate decreased after at least seven days of habituation. Estimated head motions of awake mice during fMRI were significantly smaller than a voxel size. Unexpectedly, temporal SNR of fMRI signals for awake mice was higher than that for anesthetized mice held by a conventional method. Functional connectivity in the brain of both anesthetized and awake mice showed bilateral and unilateral networks. COMPARISON WITH EXISTING METHOD(S): fMRI using CryoProbe had been performed on anesthetized mice previously. Our method does not use anesthetics during habituation or fMRI., Conclusion: Our method would be beneficial for translational research using fMRI in mice and humans because human fMRI is typically performed without anesthetics., (Copyright © 2016 Elsevier B.V. All rights reserved.)

We previously demonstrated that mouse embryonic stem cell (mESC)-derived retinal epithelium self-forms an optic cup-like structure. In the developing retina, the dorsal and ventral sides differ in terms of local gene expression and morphological features. This aspect has not yet been shown in vitro Here, we demonstrate that mESC-derived retinal tissue spontaneously acquires polarity reminiscent of the dorsal-ventral (D-V) patterning of the embryonic retina. Tbx5 and Vax2 were expressed in a mutually exclusive manner, as seen in vivo Three-dimensional morphometric analysis showed that the in vitro-formed optic cup often contains cleft structures resembling the embryonic optic fissure. To elucidate the mechanisms underlying the spontaneous D-V polarization of mESC-derived retina, we examined the effects of patterning factors, and found that endogenous BMP signaling plays a predominant role in the dorsal specification. Further analysis revealed that canonical Wnt signaling, which was spontaneously activated at the proximal region, acts upstream of BMP signaling for dorsal specification. These observations suggest that D-V polarity could be established within the self-formed retinal neuroepithelium by intrinsic mechanisms involving the spatiotemporal regulation of canonical Wnt and BMP signals., (© 2016. Published by The Company of Biologists Ltd.)

Recent active research and new regulatory guidance on pharmaceutical cocrystals have increased the rate of their development as promising approaches to improve handling, storage stability, and bioavailability of poorly soluble active pharmaceutical ingredients (APIs). However, their complex structure and the limited amount of available information related to their performance may require development strategies that differ from those of single-component crystals to ensure their clinical safety and efficacy. This article highlights current methods of characterizing pharmaceutical cocrystals and approaches to controlling their quality. Different cocrystal regulatory approaches between regions are also discussed. The physical characterization of cocrystals should include elucidating the structure of their objective crystal form as well as their possible variations (e.g., polymorphs, hydrates). Some solids may also contain crystals of individual components. Multiple processes to prepare pharmaceutical cocrystals (e.g., crystallization from solutions, grinding) vary in their applicable ingredients, scalability, and characteristics of resulting solids. The choice of the manufacturing method affects the quality control of particular cocrystals and their formulations. In vitro evaluation of the properties that govern clinical performance is attracting increasing attention in the development of pharmaceutical cocrystals. Understanding and mitigating possible factors perturbing the dissolution and/or dissolved states, including solution-mediated phase transformation (SMPT) and precipitation from supersaturated solutions, are important to ensure the bioavailability of orally administrated lower-solubility APIs. The effect of polymer excipients on the performance of APIs emphasizes the relevance of formulation design for appropriate use.

Dieting often leads to body weight cycling involving repeated weight loss and regain. However, little information is available regarding rapid-response serum markers of overnutrition that predict body weight alterations during weight cycling. Here, we report the rapid response of serum leukocyte cell-derived chemotaxin 2 (LECT2), a hepatokine that induces insulin resistance in skeletal muscle, during diet-induced weight cycling in mice. A switch from a high-fat diet (HFD) to a regular diet (RD) in obese mice gradually decreased body weight but rapidly decreased serum LECT2 levels within 10 days. In contrast, a switch from a RD to a HFD rapidly elevated serum LECT2 levels. Serum LECT2 levels showed a positive correlation with liver triglyceride contents but not with adipose tissue weight. This study demonstrates the rapid response of LECT2 preceding body weight alterations during weight cycling in mice and suggests that measurement of serum LECT2 may be clinically useful in the management of obesity., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

The eyes are subdivided from the rostral diencephalon in early development. How the neuroectoderm regulates this subdivision, however, is largely unknown. Taking advantage of embryonic stem cell (ESC) culture using a Rax reporter line to monitor rostral diencephalon formation, we found that ESC-derived tissues at day 7 grown in Glasgow Minimum Expression Media (GMEM) containing knockout serum replacement (KSR) exhibited higher levels of expression of axin2, a Wnt target gene, than those grown in chemically defined medium (CDM). Surprisingly, Wnt agonist facilitated eye field-like tissue specification in CDM. In contrast, the addition of Wnt antagonist diminished eye field tissue formation in GMEM+KSR. Furthermore, the morphological formation of the eye tissue anlage, including the optic vesicle, was accompanied by Wnt signaling activation. Additionally, using CDM culture, we developed an efficient method for generating Rax+/Chx10+ retinal progenitors, which could become fully stratified retina. Here we provide a new avenue for exploring the mechanisms of eye field specification in vitro., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

This article contains data related to the research article entitled "Specification of embryonic stem cell-derived tissues into eye fields by Wnt signaling using rostral diencephalic tissue-inducing culture" Sakakura (2016) [1]. Mouse embryonic stem cells (ESC) were used for the generation of optic vesicle-like tissues in vitro. In this article we described data in which a Rax::GFP knock-in ESC line was used to monitor the formation of optic tissues. In addition, we also described the data of regional marker expression of Rax, Sox2 and Pax6 in vivo around the forebrain and the eye tissues for comparative purposes. These data can be valuable to researchers interested in investigating forebrain and eye tissue development.

Aim: Although sorafenib is a standard drug for advanced hepatocellular carcinoma (HCC), little is known about a patient's clinical course after treatment. We investigated the effect of post-progression survival (PPS) and progression-free survival (PFS) on overall survival (OS) in patients whose advanced HCC was treated by sorafenib., Methods: We searched in the PubMed database for reports with survival data of patients with HCC treated with sorafenib monotherapy, and selected reports with 20 or more patients each that provided data for both OS and PFS or time to progression (TTP). Median PPS (mPPS) was defined as the period obtained by subtracting median PFS or TTP (mPFS/TTP) from median OS (mOS). We identified 56 reports with 5803 patients. We investigated the correlation of mOS and either mPPS or mPFS/TTP using weighted linear regression., Results: Median PPS correlated with mOS (r = 0.834) very strongly, whereas mPFS/TTP did not correlate with mOS as highly as PPS did (r = 0.546). When we stratified survival data by Child-Pugh classification, a significantly greater average percentage of mPPS to mOS was seen in Child-Pugh class A (54.4 ± 17.6%) than in Child-Pugh class B (32.0 ± 11.6%) (P = 0.015)., Conclusion: PPS highly correlated with OS, and its importance should be more emphasized for advanced HCC patients treated after sorafenib therapy, whereas we need to take more care in interpreting the results of PFS to evaluate treatment efficacy in clinical trials of advanced HCC., (© 2015 The Japan Society of Hepatology.)

The purposes of this study were to elucidate the type-specific characteristics of salt, cocrystal, and solvate formulations upon dissolution and precipitation, and to clarify their effect on enhancing oral absorption. Several types of solid formulations (dantrolene sodium salt [DAN-NA], pioglitazone hydrochloride salt [PIO-HCL], megestrol acetate saccharin cocrystal [MEG-SA], and an in-house compound ZR ethanolate [ZR-ETH]) that induce supersaturation of BCS class II drugs were compared to their crystalline free forms. An in vitro miniscale dissolution test in biorelevant media was used to characterize their dissolution profiles and residue forms. Both salts (DAN-NA and PIO-HCL) rapidly reached the maximum concentration within 5min, whereas the cocrystal (MEG-SA) did so slowly. After the maximum concentration had been reached, the dissolved concentrations of DAN-NA, PIO-HCL, and MEG-SA decreased, but that of ZR-ETH did not. Time-dependent XRPD analysis revealed that the initial solid state of each salt dissolved within 5min, whereas the cocrystal remained for more than 10min, and the solvate remained for 4h. It also revealed that PIO-HCL and MEG-SA precipitated to the stable free form, while DAN-NA precipitated to the metastable form, which maintains a higher concentration than the stable free form continuously. In vivo absorption in beagle dogs was also examined. The plasma AUC of DAN-NA, MEG-SA, and ZR-ETH was respectively 1.5-, 2.1-, and 11-fold more than each free form. On the other hand, the absorption of PIO-HCL was not enhanced compared with its free form. The results in the present study clarified that not only the precipitation rate and the form of precipitation but also the retention of the initial solid state in the absorption process contribute to enhancing the in vivo absorption of Class II drugs from solid formulations such as salts, solvates, and cocrystals., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The epiblast (foremost embryonic ectoderm) generates all three germ layers and therefore has crucial roles in the formation of all mammalian body cells. However, regulation of epiblast gene expression is poorly understood because of the difficulty of manipulating epiblast tissues in vivo. In the present study, using the self-organizing properties of mouse embryonic stem cell (ESC), we generated and characterized epiblast-like tissue in three-dimensional culture. We identified significant genome-wide gene expression changes in this epiblast-like tissue by transcriptomic analysis. In addition, we identified the particular significance of the Erk/Mapk and integrin-linked kinase pathways, and genes related to ectoderm/epithelial formation, using the bioinformatics resources IPA and DAVID. Here, we focused on Fgf5, which ranked in the top 10 among the discovered genes. To develop a functional analysis of Fgf5, we created an efficient method combining CRISPR/Cas9-mediated genome engineering and RNA interference (RNAi). Notably, we show one-step generation of various Fgf5 reporter lines including heterozygous and homozygous knockins (the GET method). For time- and dose-dependent depletion of fgf5 over the course of development, we generated an ESC line harboring Tol2 transposon-mediated integration of an inducible short hairpin RNA interference system (pdiRNAi). Our findings raised the possibility that Fgf/Erk signaling and apicobasal epithelial integrity are important factors in epiblast development. In addition, our methods provide a framework for a broad array of applications in the areas of mammalian genetics and molecular biology to understand development and to improve future therapeutics.

Insulin-like growth factors (IGFs) are involved in growth and tissue development, including diseases such as type-2 diabetes and cancers. However, their roles in lineage specification, especially in early mammalian neural development, are poorly understood. Here, we analyzed the protein expression of IGF-2 in early mouse embryo, and it was preferentially detected in anterior mesodermal tissue, adjacent to the neural plate. We utilized a self-organizing neural tissue culture system and analyzed the direct effect of IGF-2 on the general neural marker Sox1. Interestingly, using recombinant IGF-2 and a chemical inhibitor of its receptor (IGF-1R), we found that the IGF-2/IGF-1R pathway positively regulated Sox1 expression in embryonic stem (ES) cell-derived neural tissue. Furthermore, to visualize the expression patterns of other neural markers, we used reporter ES cell lines and we found that the IGF-2/IGF-1R signaling upregulated the expression of the posterior neural marker Irx3. In contrast, the anterior neural marker Six3 was downregulated by IGF-2/IGF-1R signaling. Together, our results demonstrate that IGF-2/IGF-1R signaling has different effects on neural marker expression, which may influence the early regional identity of ES cell-derived neural tissues.

Wnt/ß-catenin signaling is crucial for maintenance of pluripotent state of embryonic stem cell (ESC). However, it is unclear how Wnt/ß-catenin signaling affects the differentiation ability of ESC, especially with regard to rostral forebrain cells. Here, using Rax, rostral forebrain marker, and Wnt/ß-catenin reporter lines, we report ratio of Rax(+) and Wnt responding tissue (Wnt(+)) patterns, which were affected by seeding number of ESC in three-dimensional culture system. Surprisingly, we found ß-catenin level and localization are heterogeneous in ESC colony by immunostaining and time-laps imaging of ß-catenin-mEGFP signals. Moreover, activation of Wnt signaling in ESC promoted expression level and nuclear localization of ß-catenin, and mRNA levels of Wnt antagonists, axin2 and dkk1, leading to upregulating Wnt/ß-catenin reporter in ESC state and Rax expression at differentiation culture day 7. Together, our results suggest that activation of Wnt signaling in ESC promotes the differentiation efficacy of rostral forebrain cells. Wnt-priming culture method may provide a useful tool for applications in the areas of basic science and molecular therapeutics for regenerative medicine.

Lignocellulose, composed of cellulose, hemicellulose, and lignin, in the secondary cell wall constitutes wood and is the most abundant form of biomass on Earth. Enhancement of wood accumulation may be an effective strategy to increase biomass as well as wood strength, but currently only limited research has been undertaken. Here, we demonstrated that OsSWN1, the orthologue of the rice NAC Secondary-wall Thickening factor (NST) transcription factor, effectively enhanced secondary cell wall formation in the Arabidopsis inflorescence stem and poplar (Populus tremula×Populus tremuloides) stem when expressed by the Arabidopsis NST3 promoter. Interestingly, in transgenic Arabidopsis and poplar, ectopic secondary cell wall deposition in the pith area was observed in addition to densification of the secondary cell wall in fiber cells. The cell wall content or density of the stem increased on average by up to 38% and 39% in Arabidopsis and poplar, respectively, without causing growth inhibition. As a result, physical strength of the stem increased by up to 57% in poplar. Collectively, these data suggest that the reinforcement of wood by NST3pro:OsSWN1 is a promising strategy to enhance wood-biomass production in dicotyledonous plant species.

Anterior pituitary is critical for endocrine systems. Its hormonal responses to positive and negative regulators are indispensable for homeostasis. For this reason, generating human anterior pituitary tissue that retains regulatory hormonal control in vitro is an important step for the development of cell transplantation therapy for pituitary diseases. Here we achieve this by recapitulating mouse pituitary development using human embryonic stem cells. We find that anterior pituitary self-forms in vitro following the co-induction of hypothalamic and oral ectoderm. The juxtaposition of these tissues facilitated the formation of pituitary placode, which subsequently differentiated into pituitary hormone-producing cells. They responded normally to both releasing and feedback signals. In addition, after transplantation into hypopituitary mice, the in vitro-generated corticotrophs rescued physical activity levels and survival of the hosts. Thus, we report a useful methodology for the production of regulator-responsive human pituitary tissue that may benefit future studies in regenerative medicine.

Acute gastric volvulus is a torsion of the stomach by more than 180° and a life-threatening condition. We present a 50-year-old male patient with acute abdominal pain who has Down syndrome/trisomy 21. Computed tomography showed a significant distended stomach with features of a severe gastric volvulus. Emergency operation in form of reduction and gastropexy was performed. We are not aware of any similar cases published in the English literature, where as gastric volvulus occurred in a patient with Down syndrome.

Background: Many methods to prevent postoperative shivering (POS) has been reported. However, there are few reports demonstrating the effect of flurbiprofen on POS which affects the set point in the thermocenter of the hypothalamus., Methods: One hundred and forty six patients undergoing lung lobectomy or segmentectomy under video-assisted thoracic surgery were divided into a flurbiprofen-treated group (Group F) and a non-treated group (Group N). We retrospectively investigated the incidence of POS associated with total intravenous anesthesia with epidural anesthesia compared with or without flurbiprofen. We weighed the incidence of POS against age, body mass index, the effective site concentration of fentanyl on extubation, the mean dose of remifentanil, the minimum rectal temperature, the surgical duration and total hemorrhage volume based on the anesthetic chart Chi-square and Student t-test were used for statistical analysis., Results: Although the surgical duration in Group F was shorter than that in Group N (223±83 vs. 165±80 (min), P<0.01), the incidence of POS in Group F was higher than that in Group N (1/32 vs. 28/114, P<0.01). There were no significant differences in another items between the two groups., Conclusions: The results of the study indicates that flurbiprofen has a possible beneficial effect in preventing POS.

Neuronal heterotopia refers to brain malformations resulting from deficits of neuronal migration. Individuals with heterotopias show a high incidence of neurological deficits, such as epilepsy. More recently, it has come to be recognized that focal heterotopias may also show a range of psychiatric problems, including cognitive and behavioral impairments. However, because focal heterotopias are not always located in the brain areas responsible for the symptoms, the causal relationship between the symptoms and heterotopias remains elusive. In this study, we showed that mice with focal heterotopias in the somatosensory cortex generated by in utero electroporation exhibited spatial working memory deficit and low competitive dominance behavior, which have been shown to be closely associated with the activity of the medial prefrontal cortex (mPFC) in rodents. Analysis of the mPFC activity revealed that the immediate-early gene expression was decreased and the local field potentials of the mPFC were altered in the mice with heterotopias compared with the control mice. Moreover, activation of these ectopic and overlying sister neurons using the DREADD (designer receptor exclusively activated by designer drug) system improved the working memory deficits. These findings suggest that cortical regions containing focal heterotopias can affect distant brain regions and give rise to behavioral abnormalities. Significance statement: Recent studies reported that patients with heterotopias have a variety of clinical symptoms, such as cognitive disturbance, psychiatric symptoms, and autistic behavior. However, the causal relationship between the symptoms and heterotopias remains elusive. Here we showed that mice with focal heterotopias in the somatosensory cortex generated by in utero electroporation exhibited behavioral deficits that have been shown to be associated with the mPFC activity in rodents. The existence of heterotopias indeed altered the neural activities of the mPFC, and direct manipulation of the neural activity of the ectopic neurons and their sister neurons in the overlying cortex improved the behavioral deficit. Thus, our results indicate that focal heterotopias could affect the activities of distant brain areas and cause behavioral abnormalities., (Copyright © 2015 the authors 0270-6474/15/3512433-14$15.00/0.)

Cortical spreading depression (CSD) is a phenomenon associated with a propagating large shift in direct current (DC) potential followed by suppression of electrophysiological activity. For temporal analysis of CSD propagation, electrophysiological recording is the most reliable tool. However, it is difficult to completely identify the spatial area of the brain influenced by CSD, because recording sites are technically limited. Histological post hoc identification of activated neurons by labeling the induction of an immediate early gene (IEG) could determine areas of CSD propagation. We found that cortical application of potassium chloride induced expression of Npas4 IEG mRNA in the ipsilateral dorsal cortex. Interestingly, induction of Npas4 was never observed in the ipsilateral hippocampus and there was a clear boundary to the area of Npas4 expression. To determine whether the boundary of the area of Npas4 mRNA expression was the limit of CSD propagation, we recorded local field potentials from multiple sites that crossed the boundary of Npas4 expression. We found that the area of Npas4 mRNA expression coincided with the area of DC-potential shift propagation. We propose that induction of Npas4 identifies the area influenced by CSD propagation., (Copyright © 2015 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved.)

Embryonic stem (ES) cells have a remarkable capacity to self-organize complex, multi-layered optic cups in vitro via a culture technique called SFEBq. During both SFEBq and in vivo optic cup development, Rax (Rx) expressing neural retina epithelial (NRE) tissues utilize Fgf and Wnt/β-catenin signalling pathways to differentiate into neural retina (NR) and retinal-pigmented epithelial (RPE) tissues, respectively. How these signaling pathways affect gene expression during optic tissue formation has remained largely unknown, especially at the transcriptome scale. Here, we address this question using RNA-Seq. We generated Rx+ optic tissue using SFEBq, exposed these tissues to either Fgf or Wnt/β-catenin stimulation, and assayed their gene expression across multiple time points using RNA-Seq. This comparative dataset will help elucidate how Fgf and Wnt/β-catenin signaling affect gene expression during optic tissue differentiation and will help inform future efforts to optimize in vitro optic tissue culture technology.

The striatum is a major target of cerebral cortical output. The cortico-striatal projection has been well described, however, the neurochemical changes that occur in the striatum after prolonged cortical hyperactivation remain to be investigated. In this study, extracellular levels of glutamate, GABA, and alanine levels were measured in the dorsal striatum using microdialysis in anesthetized mice at resting condition and during 4-aminopyridine (4-AP)-induced cortical seizures. After topical application of 4-AP on the primary motor cortex that induced cortical seizures, the extracellular level of striatal GABA increased by 40% in 60 min. By contrast, the extracellular level of striatal glutamate decreased by 20%. Moreover, the surface amounts of striatal glutamate/aspartate transporter (GLAST) and glutamate transporter 1 (GLT-1), the major astrocytic high-affinity glutamate transporters, tended to increase by cortical seizures in 60 min, suggesting a recruitment of the glutamate transporters from internal stores. 4-AP also resulted in a steady increase of alanine levels which are thought to reflect glutamate and pyruvate metabolism in neurons and astrocytes. These observations possibly delineate adaptive changes of striatal metabolism by severe cortical seizures., (Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.)

An increase in the ratio of cellular excitation to inhibition (E/I ratio) has been proposed to underlie the pathogenesis of neuropsychiatric disorders, such as autism spectrum disorders (ASD), obsessive-compulsive disorder (OCD), and Tourette's syndrome (TS). A proper E/I ratio is achieved via factors expressed in neuron and glia. In astrocytes, the glutamate transporter GLT1 is critical for regulating an E/I ratio. However, the role of GLT1 dysfunction in the pathogenesis of neuropsychiatric disorders remains unknown because mice with a complete deficiency of GLT1 exhibited seizures and premature death. Here, we show that astrocyte-specific GLT1 inducible knockout (GLAST(CreERT2/+)/GLT1(flox/flox), iKO) mice exhibit pathological repetitive behaviors including excessive and injurious levels of self-grooming and tic-like head shakes. Electrophysiological studies reveal that excitatory transmission at corticostriatal synapse is normal in a basal state but is increased after repetitive stimulation. Furthermore, treatment with an N-methyl-D-aspartate (NMDA) receptor antagonist memantine ameliorated the pathological repetitive behaviors in iKO mice. These results suggest that astroglial GLT1 has a critical role in controlling the synaptic efficacy at corticostriatal synapses and its dysfunction causes pathological repetitive behaviors.

The development of gene-encoded indicators and actuators to observe and manipulate cellular functions is being advanced and investigated. Expressing these probe molecules in glial cells is expected to enable observation and manipulation of glial cell activity, leading to elucidate the behaviors and causal roles of glial cells. The first step toward understanding glial cell functions is to express the probes in sufficient amounts, and the Knockin-mediated ENhanced Gene Expression (KENGE)-tet system provides a strategy for achieving this. In the present article, three examples of KENGE-tet system application are reviewed: depolarization of oligodendrocytes, intracellular acidification of astrocytes, and observation of intracellular calcium levels in the fine processes of astrocytes.

A 50-year-old woman with chronic hepatitis associated with hepatitis B virus was found to have a hypoechoic liver tumor of 20 mm in diameter on ultrasonography. Dynamic computed tomography and magnetic resonance imaging revealed multiple tumors in the liver. The portal vein passed through the tumor, which is not a typical feature of hepatocellular carcinoma. We subsequently performed tumor biopsy and diagnosed mucosa-associated lymphoid tissue (MALT) lymphoma of hepatic origin. Here we report this rare case of hepatic MALT lymphoma in a patient with chronic hepatitis B, and present a review of the literature.

The authors report the results of an indirect comparison of the standards of absorbed dose to water in high-energy photon beams from a clinical linac and (60)Co radiation beam performed between the National Metrology Institute of Japan (NMIJ) and the Australian Radiation Protection and Nuclear Safety Agency (ARPANSA). Three ionisation chambers were calibrated by the NMIJ in April and June 2013 and by the ARPANSA in May 2013. The average ratios of the calibration coefficients for the three ionisation chambers obtained by the NMIJ to those obtained by the ARPANSA were 0.9994, 1.0040 and 1.0045 for 6-, 10- and 15-MV (18 MV at the ARPANSA) high-energy photon beams, respectively. The relative standard uncertainty of the value was 7.2 × 10(-3). The ratio for (60)Co radiation was 0.9986(66), which is consistent with the results published in the key comparison of BIPM.RI(I)-K4., (© The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The dorsal and ventral hippocampal regions (dHP and vHP) are proposed to have distinct functions. Electrophysiological studies have revealed intra-hippocampal variances along the dorsoventral axis. Nevertheless, the extra-hippocampal influences of dHP and vHP activities remain unclear. In this study, we compared the spatial distribution of brain-wide responses upon dHP or vHP activation and further estimate connection strengths between the dHP and the vHP with corresponding extra-hippocampal areas. To achieve this, we first investigated responses of local field potential (LFP) and multi unit activities (MUA) upon light stimulation in the hippocampus of an anesthetized transgenic mouse, whose CA1 pyramidal neurons expressed a step-function opsin variant of channelrhodopsin-2 (ChR2). Optogenetic stimulation increased hippocampal LFP power at theta, gamma, and ultra-fast frequency bands, and augmented MUA, indicating light-induced activation of CA1 pyramidal neurons. Brain-wide responses examined using fMRI revealed that optogenetic activation at the dHP or vHP caused blood oxygenation level-dependent (BOLD) fMRI signals in situ. Although activation at the dHP induced BOLD responses at the vHP, the opposite was not observed. Outside the hippocampal formation, activation at the dHP, but not the vHP, evoked BOLD responses at the retrosplenial cortex (RSP), which is in line with anatomical evidence. In contrast, BOLD responses at the lateral septum (LS) were induced only upon vHP activation, even though both dHP and vHP send axonal fibers to the LS. Our findings suggest that the primary targets of dHP and vHP activation are distinct, which concurs with attributed functions of the dHP and RSP in spatial memory, as well as of the vHP and LS in emotional responses.

Main Conclusion: Secondary cell wall-associated CesA genes in Populus have undergone a functional differentiation in expression pattern that may be attributable to evolutionary alteration of regulatory modules. Gene duplication is an important mechanism for functional divergence of genes. Secondary cell wall-associated cellulose synthase genes (CesA4, CesA7 and CesA8) are duplicated in Populus plants due to a recent whole genome duplication event. Here, we demonstrate that duplicate CesA genes show tissue-dependent expression divergence in Populus plants. Real-time PCR analysis of Populus CesA genes suggested that Pt × tCesA8-B was more highly expressed than Pt × tCesA8-A in phloem and secondary xylem tissue of mature stem. Histochemical and histological analyses of transformants expressing a GFP-GUS fusion gene driven by Populus CesA promoters revealed that the duplicate CesA genes showed different expression patterns in phloem fibers, secondary xylem, root cap and leaf trichomes. We predicted putative cis-regulatory motifs that regulate expression of secondary cell wall-associated CesA genes, and identified 19 motifs that are highly conserved in the CesA gene family of eudicotyledonous plants. Furthermore, a transient transactivation assay identified candidate transcription factors that affect levels and patterns of expression of Populus CesA genes. The present study reveals that secondary cell wall-associated CesA genes in Populus have undergone a functional differentiation in expression pattern that may be attributable to evolutionary alteration of regulatory modules.

The influence of astrocytes on synaptic function has been increasingly studied, owing to the discovery of both gliotransmission and morphological ensheathment of synapses. While astrocytes exhibit at best modest membrane potential fluctuations, activation of G-protein coupled receptors (GPCRs) leads to a prominent elevation of intracellular calcium which has been reported to correlate with gliotransmission. In this review, the possible role of astrocytic GPCR activation is discussed as a trigger to promote synaptic plasticity, by affecting synaptic receptors through gliotransmitters. Moreover, we suggest that volume transmission of neuromodulators could be a biological mechanism to activate astrocytic GPCRs and thereby to switch synaptic networks to the plastic mode during states of attention in cerebral cortical structures.

A large gap exists in our understanding of the course of differentiation from mesoderm to definitive hematopoietic stem cells (HSCs). Previously, we reported that Runx1(+) cells in embryonic day 7.5 (E7.5) embryos contribute to the hemogenic endothelium in the E10.5 aorta-gonad-mesonephros (AGM) region and HSCs in the adult bone marrow. Here, we show that two Runx1(+) populations subdivided by Gata1 expression exist in E7.5 embryos. The hemogenic endothelium and the HSCs are derived only from the Runx1(+)Gata1(-) population. A subset of this population moves from the extra- to intraembryonic region during E7.5-E8.0, where it contributes to the hemogenic endothelium of the dorsal aorta (DA). Migration occurs before the heartbeat is initiated, and it is independent of circulation. This suggests a developmental trajectory from Runx1(+) cells in the E7.5 extraembryonic region to definitive HSCs via the hemogenic endothelium., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

Recent articles have reported an association between fatty liver disease and systemic insulin resistance in humans, but the causal relationship remains unclear. The liver may contribute to muscle insulin resistance by releasing secretory proteins called hepatokines. Here we demonstrate that leukocyte cell-derived chemotaxin 2 (LECT2), an energy-sensing hepatokine, is a link between obesity and skeletal muscle insulin resistance. Circulating LECT2 positively correlated with the severity of both obesity and insulin resistance in humans. LECT2 expression was negatively regulated by starvation-sensing kinase adenosine monophosphate-activated protein kinase in H4IIEC hepatocytes. Genetic deletion of LECT2 in mice increased insulin sensitivity in the skeletal muscle. Treatment with recombinant LECT2 protein impaired insulin signaling via phosphorylation of Jun NH2-terminal kinase in C2C12 myocytes. These results demonstrate the involvement of LECT2 in glucose metabolism and suggest that LECT2 may be a therapeutic target for obesity-associated insulin resistance.

This study investigated the effect of interpersonal dependency on judgments of gaze direction of individuals with different facial expressions. Based on interpersonal dependency scores, 46 participants were divided into two groups (high interpersonal dependency and low interpersonal dependency). Participants judged the gaze direction of photographs of faces with angry, neutral or happy expressions. Relative to the low interpersonal dependency group, the high interpersonal dependency group was more accurate in the judgments of gaze direction. This tendency was more salient for the happy and neutral expressions than for the angry expressions. Since people with high interpersonal dependency are highly motivated to seek support from others, this result suggests that they are sensitive to signals with pro-social information such as the gaze direction of others with positive attitudes.

We recently discovered an unexpected phenomenon of somatic cell reprogramming into pluripotent cells by exposure to sublethal stimuli, which we call stimulus-triggered acquisition of pluripotency (STAP). This reprogramming does not require nuclear transfer or genetic manipulation. Here we report that reprogrammed STAP cells, unlike embryonic stem (ES) cells, can contribute to both embryonic and placental tissues, as seen in a blastocyst injection assay. Mouse STAP cells lose the ability to contribute to the placenta as well as trophoblast marker expression on converting into ES-like stem cells by treatment with adrenocorticotropic hormone (ACTH) and leukaemia inhibitory factor (LIF). In contrast, when cultured with Fgf4, STAP cells give rise to proliferative stem cells with enhanced trophoblastic characteristics. Notably, unlike conventional trophoblast stem cells, the Fgf4-induced stem cells from STAP cells contribute to both embryonic and placental tissues in vivo and transform into ES-like cells when cultured with LIF-containing medium. Taken together, the developmental potential of STAP cells, shown by chimaera formation and in vitro cell conversion, indicates that they represent a unique state of pluripotency.

Using a perennial model plant allows the study of reoccurring seasonal events in a way that is not possible using a fast-growing annual such as Arabidopsis thaliana (Arabidopsis). In this study, we present a hybrid aspen (Populus tremula × P. tremuloides) as our perennial model plant. These plants can be grown in growth chambers to shorten growth periods and manipulate day length and temperature in ways that would be impossible under natural conditions. In addition, the use of growth chambers allows easy monitoring of height and diameter expansion, accelerating the collection of data from new strategies that allow evaluation of promoters or inhibitors of growth. Here, we describe how to study and quantify responses to seasonal changes (mainly using P. tremula × P. tremuloides) by measuring growth rate and key events under different photoperiodic cycles.

Over the last several decades, changes in global temperatures have led to changes in local environments affecting the growth conditions for many species. This is a trend that makes it even more important to understand how plants respond to local variations and seasonal changes in climate. To detect daily and seasonal changes as well as acute stress factors such as cold and drought, plants rely on a circadian clock. This chapter introduces the current knowledge and literature about the setup and function of the circadian clock in various tree and perennial species, with a focus on the Populus genus.

Age-related androgen depletion is known to be a risk factor for various diseases, such as osteoporosis and sarcopenia. Furthermore, recent studies have demonstrated that age-related androgen depletion results in accumulation of β-amyloid protein and thereby acts as a risk factor for the development of Alzheimer's disease. Supplemental androgen therapy has been shown to be efficacious in treating osteoporosis and sarcopenia. In addition, studies in animals have demonstrated that androgens can play a protective role against Alzheimer's disease. However, androgen therapy is not used routinely for these indications, because of side effects. Selective androgen receptor modulators (SARMs) are a new class of compounds. SARMs maintain the beneficial effects of androgens on bone and muscle while reducing unwanted side effects. NEP28 is a new SARM exhibiting high selectivity for androgen receptor. To investigate the pharmacological effects of NEP28, we compared the effects on muscle, prostate, and brain with mice that were androgen depleted by orchidectomy and then treated with either placebo, NEP28, dihydrotestosterone, or methyltestosterone. We demonstrated that NEP28 showed tissue-selective effect equivalent to or higher than existing SARMs. In addition, the administration of NEP28 increased the activity of neprilysin, a known Aβ-degrading enzyme. These results indicate that SARM is efficacious for the treatment of not only osteoporosis and sarcopenia, but also Alzheimer's disease., (© 2013 Published by Elsevier B.V.)

In this study we investigated the changes in the sensitivity of cutaneous points and the oral mucosa that occur after intraoral vertical ramus osteotomy (IVRO). Additionally, postoperative changes in the sensitivity and the relationships between neurosensory disturbance and factors associated with IVRO operations were evaluated. An objective evaluation of the neurosensory status of cutaneous points and the oral mucosa of each patient was completed preoperatively and at 1, 2, 4, 8, 12, and 24 weeks postoperatively. The other variables studied for each patient included sex, age, magnitude of mandibular setback, and the amount of haemorrhage that occurred during surgery. In addition, the relationships between neurosensory disturbance and factors connected with IVRO operations were evaluated. We found that at cutaneous points, contributing factors such as sex, age, the magnitude of mandibular setback, and haemorrhage were associated with an increased risk of neurosensory disturbance after IVRO. However, these factors were not associated with that in the oral mucosa. In conclusion, we demonstrated the changes that occur in the sensitivity of cutaneous points and the oral mucosa after IVRO, the postoperative changes in sensitivity, and the relationships between neurosensory disturbance and factors connected with IVRO operations., (Copyright © 2013 International Association of Oral and Maxillofacial Surgeons. Published by Elsevier Ltd. All rights reserved.)

The signal current from an ionisation chamber with a PMMA build-up cap decreases with irradiation time due to electric fields produced by positive charges induced on the cap. In the present study, it was confirmed that the signal current decreases faster for irradiation using narrower (60)Co gamma-ray beams. This is because the number of secondary electrons that are emitted from surrounding materials and penetrate the build-up cap is smaller in a narrower gamma-ray beam, so that fewer positive charges are neutralised. The ionisation chamber was first subjected to continuous gamma-ray irradiation for 24 h, following which it was irradiated with shorter periodic gamma-ray bursts while measuring the current signal. This allowed the coefficients of positive charge accumulation and dissipation to be determined. It was found that the dissipation coefficient has a large constant value during gamma-ray irradiation and decreases asymptotically to a small value after irradiation is stopped. From the coefficients, the minimum signal current was calculated, which is the value when accumulation and dissipation balance each other under continuous irradiation. The time required for the signal current to recover following irradiation was also calculated.

We report a case of synchronous locally advanced bilateral breast cancer with different pathological responses to neoadjuvant chemotherapy with different biological character. The patient had presented bilateral breast cancer: the left breast cancer was hormone receptor negative, human epidermal growth factor receptor-2 (HER2) positive, and classified as T4bN1M0, stage IIIb, while the right was hormone receptor positive, HER2-negative, and classified as T4bN0M0, stage IIIb. We administered four cycles of anthracycline-based therapy followed by 12 weekly cycles of taxane with trastuzumab for neoadjuvant chemotherapy. We had achieved a significant left tumor reduction after each chemotherapy, but not right tumor. Bilateral modified radical mastectomies with axillary lymph-node dissection were performed. The therapeutic effect in the left was determined as a pathological complete response, in contrast to the right side. She has no recurrence for more than five years, though she had advanced cancer with oncologic emergency. This case could be an informative experience to understand the relation of tumor biology and response to systemic therapy.

We report that a brief electrical stimulation of the nucleus basalis of Meynert (NBM), the primary source of cholinergic projection to the cerebral cortex, induces a biphasic cerebral cortical blood flow (CBF) response in the somatosensory cortex of C57BL/6J mice. This CBF response, measured by laser Doppler flowmetry, was attenuated by the muscarinic type acetylcholine receptor antagonist atropine, suggesting a possible involvement of astrocytes in this type of CBF modulation. However, we find that IP3R2 knockout mice, which lack cytosolic Ca2+ surges in astrocytes, show similar CBF changes. Moreover, whisker stimulation resulted in similar degrees of CBF increase in IP3R2 knockout mice and the background strain C57BL/6J. Our results show that neural activity-driven CBF modulation could occur without large cytosolic increases of Ca2+ in astrocytes.

EPF1-EPF2 and EPFL9/Stomagen act antagonistically in regulating leaf stomatal density. The aim of this study was to elucidate the evolutionary functional divergence of EPF/EPFL family genes. Phylogenetic analyses showed that AtEPFL9/Stomagen-like genes are conserved only in vascular plants and are closely related to AtEPF1/EPF2-like genes. Modeling showed that EPF/EPFL peptides share a common 3D structure that is constituted of a scaffold and loop. Molecular dynamics simulation suggested that AtEPF1/EPF2-like peptides form an additional disulfide bond in their loop regions and show greater flexibility in these regions than AtEPFL9/Stomagen-like peptides. This study uncovered the evolutionary relationship and the conformational divergence of proteins encoded by the EPF/EPFL family genes.

We reported the results of a clinical pharmacological study of MC710 (a mixture of plasma-derived FVIIa and FX) in haemophilia patients with inhibitors during a non-haemorrhagic state. This report provides the results of a clot waveform analysis (CWA) and thrombin generation test (TGT) using blood samples obtained in this study. CWA and TGT were conducted using blood samples obtained from a pharmacokinetic and pharmacodynamic study in which MC710 (five dose rates: 20, 40, 80, 100 and 120 μg kg(-1)) was compared with NovoSeven (120 μg kg(-1)) and FEIBA (two dose rates: 50 and 75 U kg(-1)) as control drugs in 11 haemophilia patients with inhibitors without haemorrhagic symptoms. CWA showed that MC710 provided significantly greater improvement than the control drugs in activated partial thromboplastin time (APTT) at 80 μg kg(-1); maximum clot velocity and maximum clot acceleration were more enhanced by MC710 than by control drugs. TGT revealed that MC710 significantly shortened the initiation time of thrombin generation in comparison to FEIBA and induced greater thrombin generation potency than NovoSeven. It was not clear whether or not MC710 caused significant dose-dependent changes in the two measurements; however, differences between MC710 and the control drugs were clarified. MC710 was confirmed to have superior coagulation activity and thrombin productivity and is expected to have superior bypassing activity., (© 2012 Blackwell Publishing Ltd.)

Optogenetics, the use of channelrhodopsin: genetically-encodable light-activated proteins, allows the manipulation of specific neural circuit elements with millisecond precision. It has been 10 years since the discovery of channelrhodopsin, and optogenetics is now widely accepted as a powerful tool for addressing causal relationships between neural activity and behavior.

While pneumatosis intestinalis (PI) is a rare condition associated with a wide variety of underlying diseases, PI with intraperitoneal free air and ascites is extremely uncommon and is difficult to distinguish from diffuse peritonitis. We herein describe the case of an 87-year-old male who was admitted to our hospital with abdominal pain, distension and nausea. Abdominal plain radiography and computed tomography revealed intramural air collection in the entire intestine, intraperitoneal free air and ascites. Although we first suspected bowel necrosis and perforation, his physical findings and the properties of the diagnostic abdominal paracentesis did not support this diagnosis. Therefore, we selected conservative management, and the intramural air, intraperitoneal free air and ascites disappeared 1 week later. Recognition of the possible presence of non-surgical PI and intraperitoneal free air, although it is extremely rare, is important to avoid a misdiagnosis and the associated unnecessary surgical intervention.

Vertical distraction osteogenesis can extend not only to hard tissues but also to soft tissues. There is a tendency to cause progressive lingual inclination of the distracted segment. This study describes a method for preventing the lingual inclination of the transport segment in patients with vertical distraction osteogenesis in the anterior region of the mandible and reports the results of long-term follow-up. The subjects included 5 patients who had severely atrophic ridges in the anterior mandible. In all cases, a part of the mental protuberance was scraped out, and the distractor was placed suitably in a labioinclination beforehand. Therefore, the transport segments did not lean to the lingual side and had long-term stability.

The purpose of this retrospective study was to determine the outcome of Brånemark System TiUnite® implants (Nobel Biocare/Sweden), and to identify the risk factors associated with implant failure. A total of 151 patients (83 maxillae and 91 mandibles) received 619 implants from July 2003 until May 2010. The patients included 86 males and 65 females, with a median age of 51.6 years and an age range of 16 to 90 years at the time of implant surgery. Seventeen maxillae and 16 mandibles were completely edentulous, and 66 maxillae and 75 mandibles were partially edentulous. All the patients were followed until June 2011. Among the 619 implants, 9 maxillary implants and 8 mandibular implants were unsuccessful. The overall survival rate was 96.82%. A logistic regression analysis identified that a history of steroid treatment, application of a dento-maxillary prosthesis, a lack of mechanical coupling between the implants, and the length of the implants (≤8.5mm) were significant predictors of implant failure.

In this report, we demonstrate that an optic cup structure can form by self-organization in human ESC culture. The human ESC-derived optic cup is much larger than the mouse ESC-derived one, presumably reflecting the species differences. The neural retina in human ESC culture is thick and spontaneously curves in an apically convex manner, which is not seen in mouse ESC culture. In addition, human ESC-derived neural retina grows into multilayered tissue containing both rods and cones, whereas cone differentiation is rare in mouse ESC culture. The accumulation of photoreceptors in human ESC culture can be greatly accelerated by Notch inhibition. In addition, we show that an optimized vitrification method enables en bloc cryopreservation of stratified neural retina of human origin. This storage method at an intermediate step during the time-consuming differentiation process provides a versatile solution for quality control in large-scale preparation of clinical-grade retinal tissues., (Copyright © 2012 Elsevier Inc. All rights reserved.)