Identification of a novel target of sulforaphane: Sulforaphane binds to acyl-protein thioesterase 2 (APT2) and attenuates its palmitoylation.

Sulforaphane (SFaN) is a food-derived compound with several bioactive properties, including atherosclerosis, diabetes, and obesity treatment. However, the mechanisms by which SFaN exerts its various effects are still unclear. To elucidate the mechanisms of the various effects of SFaN, we explored novel SFaN-binding proteins using SFaN beads and identified acyl protein thioesterase 2 (APT2). We also found that SFaN binds to the APT2 via C56 residue and attenuates the palmitoylation of APT2, thereby reducing plasma membrane localization of APT2. This study reveals a novel bioactivity of SFaN as a regulator of APT2 protein palmitoylation.

HNF4α is required for Tkfc promoter activation by ChREBP.

Triokinase/FMN cyclase (Tkfc) is involved in fructose metabolism and is responsible for the phosphorylation of glyceraldehyde to glyceraldehyde-3-phosphate. In this study, we show that refeeding induced hepatic expression of Tkfc in mice. Luciferase reporter gene assays using the Tkfc promoter revealed the existence of two hepatocyte nuclear factor 4α (HNF4α)-responsive elements (HNF4RE1 and HNF4RE2) and one carbohydrate-responsive element-binding protein (ChREBP)-responsive element (ChoRE1). Deletion and mutation of HNF4RE1 and HNF4RE2 or ChoRE1 abolished HNF4α and ChREBP responsiveness, respectively. HNF4α and ChREBP synergistically stimulated Tkfc promoter activity. ChoRE1 mutation attenuated but maintained HNF4α responsiveness, whereas HNF4RE1 and HNF4RE2 mutations abolished ChREBP responsiveness. Moreover, Tkfc promoter activity stimulation by ChREBP was attenuated upon HNF4α knockdown. Furthermore, Tkfc expression was decreased in livers of ChREBP-/- and liver-specific HNF4-/- (Hnf4αΔHep) mice. Altogether, our data indicate that Tkfc is a target gene of ChREBP and HNF4α, and Tkfc promoter activity stimulation by ChREBP requires HNF4α.

Chrysin 7-O-ß-d-glucopyranoside increases hepatic low-density lipoprotein receptor expression through AMP-activated protein kinase activation.

Elevated plasma low-density lipoprotein (LDL) cholesterol level is a risk factor for developing atherosclerosis. Increased LDL receptor (LDLR) expression is expected to reduce the risk of atherosclerotic disease since hepatic LDLR is essential for clearing plasma LDL cholesterol. Here, we screened human LDLR promoter effectors and observed that extracts from peduncles of sweet cherry (Prunus avium) 'Sato-Nishiki' induce LDLR gene promoter activity. We used several analytical and chemical methods to show that chrysin 7-O-ß-d-glucopyranoside (chrysin-7G) is one of the compounds that stimulate LDLR gene promoter activity in cherry peduncle extracts. Furthermore, synthetic chrysin-7G increased the expression and activity of LDLR. The chrysin-7G-mediated increase in LDLR expression and activity was completely abolished by treatment with an AMP-activated protein kinase (AMPK) inhibitor, compound C. These results indicate that chrysin-7G increases LDLR expression through AMPK activation and may be a useful compound that can be recycled from waste parts of agricultural products.

Sulforaphane suppresses the activity of sterol regulatory element-binding proteins (SREBPs) by promoting SREBP precursor degradation.

Sterol regulatory element-binding proteins (SREBPs) are transcription factors that regulate various genes involved in cholesterol and fatty acid synthesis. In this study, we describe that naturally occurring isothiocyanate sulforaphane (SFaN) impairs fatty acid synthase promoter activity and reduces SREBP target gene (e.g., fatty acid synthase and acetyl-CoA carboxylase 1) expression in human hepatoma Huh-7 cells. SFaN reduced SREBP proteins by promoting the degradation of the SREBP precursor. Amino acids 595-784 of SREBP-1a were essential for SFaN-mediated SREBP-1a degradation. We also found that such SREBP-1 degradation occurs independently of the SREBP cleavage-activating protein and the Keap1-Nrf2 pathway. This study identifies SFaN as an SREBP inhibitor and provides evidence that SFaN could have major potential as a pharmaceutical preparation against hepatic steatosis and obesity.

MIG12 is involved in the LXR activation-mediated induction of the polymerization of mammalian acetyl-CoA carboxylase.

Liver X receptors (LXR) α and ß are a family of nuclear receptors that regulate lipogenesis by controlling the expression of the genes involved in the synthesis of fatty acids. MID1IP1, which encodes MIG12, is a target gene of LXR. MIG12 induces fatty acid synthesis by stimulating the polymerization-mediated activation of acetyl-CoA carboxylase (ACC). Here, we show that LXR's activation stimulates ACC polymerization in HepG2 cells by increasing the expression of MIG12. A knockdown of MID1IP1 abrogated the stimulation completely. The mutations of MIG12's leucine-zipper domain reduced the interaction between MIG12 and ACC, thus decreasing the MIG12's capacity to stimulate ACC polymerization. These results indicate that LXR's activation stimulates lipogenesis not only through the induction of the genes encoding lipogenic enzymes but also through MIG12's stimulation of ACC polymerization.

Characterization of a novel compound that promotes myogenesis via Akt and transcriptional co-activator with PDZ-binding motif (TAZ) in mouse C2C12 cells.

Transcriptional co-activator with PDZ-binding motif (TAZ) plays versatile roles in the regulation of cell proliferation and differentiation. TAZ activity changes in response to the cellular environment such as mechanic and nutritional stimuli, osmolarity, and hypoxia. To understand the physiological roles of TAZ, chemical compounds that activate TAZ in cells are useful as experimental reagents. Kaempferol, TM-25659, and ethacridine are reported as TAZ activators. However, as each TAZ activator has a distinct property in cellular functions, additional TAZ activators are awaiting. We screened for TAZ activators and previously reported IB008738 as a TAZ activator that promotes myogenesis in C2C12 cells. In this study, we have characterized IBS004735 that was obtained in the same screening. IBS004735 also promotes myogenesis in C2C12 cells, but is not similar to IBS008738 in the structure. IBS004735 activates TAZ via Akt and has no effect on TAZ phosphorylation, which is the well-described key modification to regulate TAZ activity. Thus, we introduce IBS004735 as a novel TAZ activator that regulates TAZ in a yet unidentified mechanism.

Application of Split-GFP Reassembly Assay to Study Myogenesis and Myofusion In Vitro.

Green fluorescent protein (GFP) is composed of 11 ß-strands, and loses GFP signals, when divided into the N-terminal ten ß-strands (GFP1-10) and the C-terminal last ß-strand (GFP11). However, when GFP1-10 and GFP11 encounter, they reassemble into the fluorescent GFP. We expressed GFP1-10 and blasticidin resistance gene product-fused GFP11 (BSR-GFP11) in C2C12 cells. Both the cell lines do not show GFP but when they undergo myogenesis and myofusion, GFP1-10 and BSR-GFP11 form the fluorescent complex in multi-nuclear myotubes, so that GFP signals reflect myogenesis and myofusion.

Exploring suicide risk factors among Japanese individuals: The largest case-control psychological autopsy study in Japan.

This case-control psychological autopsy study explored suicide risk factors among Japanese individuals. We targeted 102 suicide completers over 20 years of age and 334 living controls matched to suicide cases by gender, age, and residential municipality in Japan. After controlling for other factors, we found significant associations between suicide and mental disorders, sleeping problems, and verbal communication of one's own death. We discuss the importance of sensitive responses to verbal expressions of suicidal ideation.

Loop diuretics affect skeletal myoblast differentiation and exercise-induced muscle hypertrophy.

Muscle wasting or sarcopenia contributes to morbidity and mortality in patients with cancer, renal failure, or heart failure, and in elderly individuals. Na+-K+-2Cl- cotransporter 1 (NKCC1) is highly expressed in mammalian skeletal muscle, where it contributes to the generation of membrane ion currents and potential. However, the physiologic function of NKCC1 in myogenesis is unclear. We investigated this issue using the NKCC1 inhibitors bumetanide and furosemide, which are commonly used loop diuretics. NKCC1 protein levels increased during C2C12 murine skeletal myoblast differentiation, similarly to those of the myogenic markers myogenin and myosin heavy chain (MHC). NKCC1 inhibitors markedly suppressed myoblast fusion into myotubes and the expression of myogenin and MHC. Furthermore, phosphorylated and total NKCC1 levels were elevated in mouse skeletal muscles after 6 weeks' voluntary wheel running. Immunofluorescence analyses of myofiber cross-sections revealed more large myofibers after exercise, but this was impaired by daily intraperitoneal bumetanide injections (0.2 or 10 mg/kg/day). NKCC1 plays an essential role in myogenesis and exercise-induced skeletal muscle hypertrophy, and sarcopenia in patients with renal or heart failure may be attributable to treatment with loop diuretics.

Female suicides: Psychosocial and psychiatric characteristics identified by a psychological autopsy study in Japan.

AIM: Although the female suicide rate in Japan is one of the highest among OECD countries, little has been done to assess the psychosocial and psychiatric characteristics of Japanese female suicide completers. This study aimed to examine sex differences in psychosocial and psychiatric characteristics of suicide completers using a psychological autopsy study method, and to identify female suicide factors and intervention points to prevent female suicides. METHODS: A semi-structured interview was conducted with close family members of adult suicide completers. The interview included questions regarding sociodemographic factors, suicide characteristics, previous suicidal behaviors and a family history of suicidal behaviors, financial problems, and physical/psychiatric problems. Fisher's exact test and the Student's t-test were used to explore sex differences in these survey items, and individual descriptive information of female suicide cases was also examined. RESULTS: Of the 92 suicide completers, 28 were female and 64 were male. Females had a significantly higher prevalence of a history of self-harm/suicide attempts (P < 0.001). The prevalence of eating disorders was significantly higher among females than males (P < 0.01). CONCLUSION: The findings of this study highlight the importance of providing psychological and social support to caregivers of those who repeatedly attempt suicide and express suicidal thoughts, and to suggest the need to improve community care systems to be aware of suicide risk factors among female suicide attempters.

Birthdays are associated with an increased risk of suicide in Japan: Evidence from 27,007 deaths in Tokyo in 2001-2010.

BACKGROUND: Previous research has produced conflicting findings concerning whether birthdays are associated with an increased risk of suicide. This study examined the association in Tokyo, Japan. METHODS: Suicide data (ICD-10 codes X60-X84) for the period 2001-2010 were obtained from the Japanese Ministry of Health, Labour and Welfare. A time-stratified case-crossover design was used with conditional logistic regression analysis being performed to estimate within-subject 'birthday exposures' while controlling for meteorological conditions and public holidays. RESULTS: There were 27,007 suicides in the study period. For males the 5 days before the birthday and the week after the birthday were associated with significantly higher odds for suicide with the odds ratio being highest on the actual birthday (OR =1.677, 95% CI: 1.294, 2.172). For females, significantly higher odds for completed suicide were observed 7-11 days before the birthday. Stratified analyses showed different at risk time patterns among men from different age groups, and that married men had higher odds for suicide on, and for the 4 days before and in the 2 weeks after their birthday. LIMITATIONS: We lacked detailed information on suicides which would have enabled a better understanding of the observed associations. CONCLUSIONS: Birthdays are associated with an increased risk for suicide in Tokyo, Japan. Health professionals who work with individuals at risk of suicide should be made aware that birthdays are associated with an elevated suicide risk. This information should also be communicated in wider suicide prevention campaigns.

A new cell-based assay to evaluate myogenesis in mouse myoblast C2C12 cells.

The development of the efficient screening system of detecting compounds that promote myogenesis and prevent muscle atrophy is important. Mouse C2C12 cells are widely used to evaluate myogenesis but the procedures of the assay are not simple and the quantification is not easy. We established C2C12 cells expressing the N-terminal green fluorescence protein (GFP) and the C-terminal GFP (GFP1-10 and GFP11 cells). GFP1-10 and GFP11 cells do not exhibit GFP signals until they are fused. The signal intensity correlates with the expression of myogenic markers and myofusion. Myogenesis-promoting reagents, such as insulin-like growth factor-1 (IGF1) and ß-guanidinopropionic acid (GPA), enhance the signals, whereas the poly-caspase inhibitor, z-VAD-FMK, suppresses it. GFP signals are observed when myotubes formed by GFP1-10 cells are fused with single nuclear GFP11 cells, and enhanced by IGF1, GPA, and IBS008738, a recently-reported myogenesis-promoting reagent. Fusion between myotubes formed by GFP1-10 and GFP11 cells is associated with the appearance of GFP signals. IGF1 and GPA augment these signals, whereas NSC23766, Rac inhibitor, decreases them. The conditioned medium of cancer cells suppresses GFP signals during myogenesis and reduces the width of GFP-positive myotubes after differentiation. Thus the novel split GFP-based assay will provide the useful method for the study of myogenesis, myofusion, and atrophy.

MAGI2/S-SCAM outside brain.

Membrane-associated guanylate kinase with an inverted arrangement of protein-protein interaction domains (MAGI)2 (also called synaptic scaffolding molecule (S-SCAM), atrophin-1-interacting protein 1, activin receptor-interacting protein 1) is a scaffold protein that binds a wide variety of receptors, cell adhesion molecules and signalling molecules. It also interacts with other scaffold proteins and adaptors, and forms a protein network that supports cell junctions. As it is highly expressed in brain, the study on its roles in synaptic organization initially preceded. However, mounting evidence indicates that MAGI2/S-SCAM functions as a tumour suppressor and plays essential roles to maintain the integrity of cell structures in non-neuronal tissues. We review the articles regarding to MAGI2/S-SCAM outside brain and discuss future perspectives for the research of MAGI family proteins.

The mammalian Hippo pathway: regulation and function of YAP1 and TAZ.

The Hippo pathway was originally identified as the signaling that controls organ size in Drosophila, with the core architecture conserved in mammals. In the mammalian Hippo pathway, mammalian Ste20-like kinases (MST1/2) and large tumor suppressor kinases (LATS1/2) regulate transcriptional co-activators, Yes-associated protein (YAP1) and Transcriptional co-activator with a PDZ-binding motif (TAZ). The Hippo pathway was initially thought to be quite straightforward; however, the identification of additional components has revealed its inherent complexity. Regulation of YAP1 and TAZ is not always dependent on MST1/2 and LATS1/2. MST1/2 and LATS1/2 play various YAP1/TAZ-independent roles, while YAP1 and TAZ cross-talk with other signaling pathways. In this review we focus on YAP1 and TAZ and discuss their regulation, function, and the consequences of their dysregulation.

Suicide prevention strategies in Japan: a 15-year review (1998-2013).

Suicide is a global public health problem and solutions to it can be found only through a global dialog. The suicide rate in Japan has been alarming, but Japan has made substantial efforts to reduce this rate, making prevention a high priority. This report reviews the developmental stages of a comprehensive policy of suicide prevention in Japan from 1998 to 2013. Our review suggests that suicide prevention activities were facilitated by the 2006 Basic Act for Suicide Prevention and the 2007 General Principles of Suicide Prevention Policy. Along with the establishment of a Special Fund program for local governments, the Basic Act and General Principles led to the development of a comprehensive and multi-sector approach to suicide prevention. Suicide rates in Japan, especially among middle-aged men, decreased consistently after 2009, suggesting that the initiatives were effective. Continuous monitoring is needed to evaluate Japan's suicide prevention policy.

Suicide risk among individuals with sleep disturbances in Japan: a case-control psychological autopsy study.

OBJECTIVE: This case-control psychological autopsy study aimed to explore a relationship between sleep disturbances and suicide among Japanese, as well as determine the importance and usability of screening for sleep disturbances in suicide prevention. METHODS: A semi-structured interview was conducted with the close family members of 49 adult suicide completers and 145 gender-, age-, and residential municipality-matched living controls. The survey included sections on demographics, sleep disturbances, and mental disorders. Conditional logistic regression analyses were performed to compare sleep disturbance prevalence between the two groups. RESULTS: A significantly higher prevalence of sleep disturbances was found among the suicide group (75.5%) compared to the controls (11.0%) (odds ratio [OR]=21.6, p<0.001). The association remained significant after adjusting for mental disorders (OR=12.7, p<0.001). The population attributable risk percent of suicide associated with sleep disturbances and mental disorders was estimated to be 56.4% and 35.3%, respectively. CONCLUSIONS: The study confirmed that sleep disturbances are an important risk factor of suicide, independent of mental disorders. Sleep disturbances accounted for a greater proportion of suicide cases than did mental disorders in the Japanese population given the higher prevalence, and could thus be considered an important target in suicide prevention in Japan.

Factors associated with attitudes toward suicide: among Japanese pharmacists participating in the Board Certified Psychiatric Pharmacy Specialist Seminar.

BACKGROUND: Pharmacists could play a significant role in providing care for suicidal individuals. Previous studies reported that gatekeeper attitudes toward suicide could affect caregiving behaviors toward people at risk for suicide. Nevertheless, only few studies have examined the attitudes of pharmacists toward suicide. AIMS: This study explores the factors associated with the attitudes of pharmacists toward suicide. METHODS: A self-administered questionnaire and written explanation of the study were mailed to 541 pharmacists who had registered for the Board Certified Psychiatry Pharmacy Specialist Seminar in Tokyo, Japan. A Japanese version of the Attitudes Toward Suicide questionnaire was used to assess participant attitudes toward suicide. Multivariate analysis of covariance was used to assess relationships between participant attitudes toward suicide as well as demographics, occupational, and personal factors. RESULTS: Participants who had previously received suicide-related education were more likely to have positive attitudes toward suicide prevention. Conversely, those with a lifetime history of suicidal thoughts had more permissive attitudes toward one's right to commit suicide. CONCLUSIONS: Our findings suggest that pharmacist suicide prevention training programs should aim to alter permissiveness attitudes of trainees toward suicide.

Exploration of factors associated with social worker attitudes toward suicide.

BACKGROUND: Social workers are expected to play important roles in suicide intervention. Caregiving behaviours of medical personnel to suicidal individuals have been reported to be influenced by their own attitudes toward suicide. In this context, only a limited number of studies have examined social workers' attitudes toward suicide. AIM: The purpose of this study was to explore associations between personal or occupational factors of social workers and their attitudes toward suicide. METHODS: A self-administered questionnaire was mailed to 2,999 study participants registered with the Tokyo chapter of the Japanese Association of Certified Social Workers. We adopted the Attitudes Toward Suicide Scale (ATTS) to measure attitudes toward suicide. MANCOVA was used to test for the effects of demographic, personal and occupational factors on ATTS sub-scale scores. RESULTS: Participants with a history of suicidal thoughts had stronger attitudes regarding the right to suicide than those with no history; these attitudes were not affected by a history of participating in suicide-prevention training. CONCLUSIONS: Our findings suggest that suicide education should incorporate programmes directed at altering permissive attitudes toward suicide.

Factors associated with perceived feasibility and willingness of non-psychiatric doctors in Japan to treat depressed patients.

OBJECTIVES: We previously reported that many non-psychiatric doctors in Japan believe that treating depression was not part of their duties. Educational interventions must address motivation of physicians to play a role in depression care. In this study, we explored factors associated with perceived feasibility and willingness of non-psychiatric doctors in Japan to treat depression. METHODS: The study population included non-psychiatric doctors of the General Physician-Psychiatrist (G-P) Network group in Japan. We explored perceived feasibility and willingness to treat depressed patients, and examined preliminary associations with attitudes toward depression (the Depression Attitude Questionnaire: DAQ) and current depression treatment in routine medical practice. RESULTS: Responses were obtained from 56 non-psychiatric doctors (response rate: 35.4%). The doctors who scored high on the "Professional" and "Pessimism" subscale of the DAQ believed that treating depressed patients was not feasible (chi2 = 13.6, p < 0.01; chi2 = 7.3, p < 0.05, respectively) and were not willing to treat depressed patients (chi2 = 9.4, p < 0.01; chi2 = 6.6, p < 0.05, respectively) as part of their routine medical practice. The doctors who scored high on the "Professional" subscale referred fewer depressed patients to psychiatrists (r = -0.33, p < 0.05), and those who scored high on the "Pessimism" subscale recognized fewer depressed patients (r = -0.39, p < 0.01). CONCLUSIONS: The present study showed that attitudes toward depression were associated with perceived feasibility and willingness to treat depressed patients and with under-diagnosis of depression. Educational interventions optimized for these attitudes should be developed to improve recognition and treatment of depression in Japan.

Attitudes toward depression among Japanese non-psychiatric medical doctors: a cross-sectional study.

BACKGROUND: Under-recognition of depression is common in many countries. Education of medical staff, focusing on their attitudes towards depression, may be necessary to change their behavior and enhance recognition of depression. Several studies have previously reported on attitudes toward depression among general physicians. However, little is known about attitudes of non-psychiatric doctors in Japan. In the present study, we surveyed non-psychiatric doctors' attitude toward depression. METHODS: The inclusion criteria of participants in the present study were as follows: 1) Japanese non-psychiatric doctors and 2) attendees in educational opportunities regarding depression care. We conveniently approached two populations: 1) a workshop to depression care for non-psychiatric doctors and 2) a general physician-psychiatrist (G-P) network group. We contacted 367 subjects. Attitudes toward depression were measured using the Depression Attitude Questionnaire (DAQ), a 20-item self-report questionnaire developed for general physicians. We report scores of each DAQ item and factors derived from exploratory factor analysis. RESULTS: We received responses from 230 subjects, and we used DAQ data from 187 non-psychiatric doctors who met the inclusion criteria. All non-psychiatric doctors (n = 187) disagreed with "I feel comfortable in dealing with depressed patients' needs," while 60 % (n = 112) agreed with "Working with depressed patients is heavy going." Factor analysis indicated these items comprised a factor termed "Depression should be treated by psychiatrists" - to which 54 % of doctors (n = 101) agreed. Meanwhile, 67 % of doctors (n = 126) thought that nurses could be useful in depressed patient support. The three factors derived from the Japanese DAQ differed from models previously derived from British GP samples. The attitude of Japanese non-psychiatric doctors concerning whether depression should be treated by psychiatrists was markedly different to that of British GPs. CONCLUSIONS: Japanese non-psychiatric doctors believe that depression care is beyond the scope of their duties. It is suggested that educational programs or guidelines for depression care developed in other countries such as the UK are not directly adaptable for Japanese non-psychiatric doctors. Developing a focused educational program that motivates non-psychiatric doctors to play a role in depression care is necessary to enhance recognition and treatment of depression in Japan.