CDK1-PP2A-B55 interplay ensures cell cycle oscillation via Apc1-loop300.

Cell cycle control relies on a delicate balance of phosphorylation with CDK1 and phosphatases like PP1 and PP2A-B55. Yet, identifying the primary substrate responsible for cell cycle oscillations remains a challenge. We uncover the pivotal role of phospho-regulation in the anaphase-promoting complex/cyclosome (APC/C), particularly through the Apc1-loop300 domain (Apc1-300L), orchestrated by CDK1 and PP2A-B55. Premature activation of PP2A-B55 during mitosis, induced by Greatwall kinase depletion, leads to Apc1-300L dephosphorylation, stalling APC/C activity and delaying Cyclin B degradation. This effect can be counteracted using the B55-specific inhibitor pEnsa or by removing Apc1-300L. We also show Cdc20's dynamic APC/C interaction across cell cycle stages, but dephosphorylation of Apc1-300L specifically inhibits further Cdc20 recruitment. Our study underscores APC/C's central role in cell cycle oscillation, identifying it as a primary substrate regulated by the CDK-PP2A partnership.

A case of arteriovenous malformation in the inferior mesenteric artery region resected surgically under intraoperative indocyanine green fluorescence imaging.

INTRODUCTION AND IMPORTANCE: An arteriovenous malformation (AVM) is defined as a vascular malformation with a short, non-capillary communication between the arteries and veins. Most gastrointestinal AVMs are solitary, occurring predominantly in the stomach, small intestine and right colon, and rarely in the inferior mesenteric artery (IMA) region. CASE PRESENTATION: A 70-year-old man was first diagnosed with ischemic enteritis two years earlier, and was hospitalized several times with the same diagnosis. He visited our hospital because of left lower abdominal pain and melena. Colonoscopy showed findings suggestive of ischemic enteritis, and contrast-enhanced computed tomography (CT) and IMA angiography showed hyperplasia and dilation of blood vessels from the sigmoid-descending colon junction to the upper rectum. We performed conventional laparoscopic low anterior resection using intraoperative intravenous injection of indocyanine green (ICG). The final diagnosis was arteriovenous malformation in the IMA region. The patient had an uneventful postoperative course and was discharged on the 13th day after the operation. CLINICAL DISCUSSION: Cases of AVM in the IMA region are relatively rare. This is the first reported case of AVM in the IMA region that was resected under intraoperative ICG fluorescence imaging (FI), which provided useful information on the extent of intestinal resection and mesenteric dissection required, and confirmed the adequacy of intestinal blood flow during and after mesenteric dissection and anastomosis. CONCLUSION: It is advisable to use ICG FI intraoperatively during resection of AVMs in the IMA region, as with colorectal cancer surgery.

The association of tobacco use with prescription of muscle relaxants, benzodiazepines, and opioid analgesics for non-cancer pain.

BACKGROUND AND OBJECTIVES: Tobacco use and co-prescription of sedative hypnotics are risk factors for misuse of prescribed opioids among patients with non-cancer pain. However, the association between tobacco use and these co-prescriptions has not been clarified. We aimed to assess differences in the prescription and co-prescription rates of opioid analgesics with muscle relaxants and/or benzodiazepines between tobacco users and non-users. METHODS: Visit data were obtained from the 2006 to 2009 National Ambulatory Medical Care Survey, an annual cross-sectional survey of visits to office-based physicians in outpatient settings in the United States. Our sample patients were aged ≥18 years and diagnosed with non-cancer back and neck pain. The χ2 test and multiple logistic regression analysis were used to assess bivariate and multivariate associations between prescription or co-prescription rates and tobacco use status. RESULTS: We analyzed a total of 114,199,536 weighted visits (unweighted number: 3,521). Significant odds ratios (ORs) of tobacco users (vs non-users) for medical prescriptions were as follows: opioid analgesics, OR 2.14, 95% confidence interval (CI) 1.64-2.80; muscle relaxants and opioid analgesics, OR 2.57, 95%CI 1.76-3.74; benzodiazepines and opioid analgesics, OR 3.66, 95%CI 2.11-6.35, and muscle relaxants, benzodiazepines, and opioid analgesics, OR 7.02, 95%CI 2.98-16.57. CONCLUSIONS AND SCIENTIFIC SIGNIFICANCE: Tobacco users were more likely to receive prescriptions for opioid analgesics with muscle relaxants and/or benzodiazepines than non-users. Healthcare professionals need to limit co-prescription of opioid analgesics with muscle relaxants and/or benzodiazepines among tobacco users and provide a comprehensive approach to pain management. (Am J Addict 2019;XX:1-8).

Migraine-preventive prescription patterns by physician specialty in ambulatory care settings in the United States.

Many adults with migraine who require preventive therapy are often not prescribed the proper medications. The most likely reason is that primary care physicians are unacquainted with preventive medications for migraine. The present study assessed the migraine-preventive prescription patterns in office visits using data from the National Ambulatory Medical Care Survey from 2006 to 2009 in the United States. Patients who were 18 years or older and diagnosed with migraine were included in the analysis. In accordance with the recommendations of the headache guidelines, we included beta-blockers, antidepressants, triptans for short-term prevention of menstrual migraine, and other triptans for acute treatment. Weighted visits of adults with migraine prescribed with preventive medication ranged from 32.8% in 2006 to 38.6% in 2009. Visits to primary care physicians accounted for 72.6% of the analyzed adult migraine visits. Anticonvulsants (odds ratio [OR] 0.29, 95% confidence interval [CI] 0.14-0.57, p < 0.001) and triptans for menstrual migraine (OR 0.50, 95% CI 0.28-0.91, p = 0.025) were less frequently prescribed by primary care physicians compared with specialty care physicians, such as neurologists and psychiatrists. There were no significant differences in the prescription patterns of antidepressants and beta-blockers between primary and specialty care physicians. Beta-blockers were prescribed to patients with comorbidity of hypertension, and antidepressants were used by patients with comorbidity of depression. There are differences in the prescription patterns of certain type of preventive medications between primary care physicians and specialty care physicians.

Hepatitis C Virus Infection Increases c-Jun N-Terminal Kinase (JNK) Phosphorylation and Accentuates Hepatocyte Lipoapoptosis.

BACKGROUND Hepatitis C virus (HCV) infection and metabolic diseases including nonalcoholic steatohepatitis (NASH) exhibit a complex interplay. Although free fatty acid-mediated apoptosis is a prominent feature of NASH, the impact of HCV infection on hepatocyte lipotoxicity has remained largely unexplored. The study aimed at identifying whether infection by HCV affected the apoptotic pathway in hepatocytes during fatty acid assault. MATERIAL AND METHODS OR6 cells, which are derived from human hepatocellular carcinoma Huh-7 cells and harbor a full-length HCV RNA genome replication system, were treated with palmitate. Apoptosis was examined by 4',6-diamidino-2-phenylindole staining. Activation and expression of JNK, Bim, cIAP-1, and Mcl-1 were examined by immunoblotting. mRNA expression of CHOP, a major player in endoplasmic reticulum stress-mediated apoptosis, was assessed by real-time PCR. RESULTS Palmitate-induced hepatocyte apoptosis was significantly enhanced in OR6 cells compared to cured cells, in which the HCV genome had been eradicated by treatment with interferon-α. Although basal expression of CHOP mRNA was enhanced in OR6 cells compared to cured cells, it was similarly upregulated in both cell lines following palmitate treatment. Notably, palmitate-induced JNK phosphorylation was accentuated in OR6 cells compared to cured cells. Inhibition of JNK with SP600125 attenuated palmitate-induced apoptosis. Palmitate-mediated upregulation of BH3-only protein Bim, which acts downstream of JNK, was also enhanced in OR6 cells compared to cured cells. In contrast, Mcl-1 and cIAP-1 were equally reduced in OR6 cells and cured cells following palmitate treatment. CONCLUSIONS These findings suggest that during lipoapoptosis, HCV infection may enhance hepatocyte toxicity by increasing JNK phosphorylation.

Physicians' and pharmacists' information provision and patients' psychological distress.

Providing information related to medication has many benefits for patients. However, patients' conflicting perceptions about medical information provided by physicians and pharmacists may be associated with their psychological distress regarding treatment and medication. This study investigated associations between patients' perceptions of agreement between physicians and pharmacists about medical information and improvements in their psychological distress. It also clarified the specific relationships of their perceptions with psychological distress. A cross-sectional survey was conducted in Japanese community pharmacy settings. Pharmacists approached 1,500 patients visiting community pharmacies and provided them with questionnaire packages. Patients completed the questionnaires at home and returned them to the researchers by mail. Multivariate logistic regression analysis and signal detection analysis were conducted to examine associations of patients' perceptions of information agreement with improvement in psychological distress. Measures of improvement in worry and anxiety about disease, improvement in worry and anxiety about medication, and improvement in depressive mood were used to assess alleviation of psychological distress. A total of 645 patients returned the questionnaires; 628 contributed to the data. Multivariate logistic regression analyses clarified that patients' perceptions of agreement in information regarding need for medication, methods for adverse drug reaction reduction, adverse drug reaction symptoms, coping with forgetting to take medication, and advice for daily life were significantly associated with improvements in psychological distress. Furthermore, signal detection analysis showed that several combinations of patients' perceptions of agreement between physicians and pharmacists about specific medical information were also significantly associated with improvement in psychological distress. Consistent information provision by physicians and pharmacists could contribute to decreased psychological distress in patients, and consequently to adherence to treatment and taking medication.

Perceptions of pharmacists and patients on information provision and their influence on patient satisfaction in Japanese community pharmacies.

RATIONALE, AIMS AND OBJECTIVES: The provision of information is now considered a major area in pharmacist-patient interactions. However, few reports have simultaneously evaluated patient and pharmacist perceptions with regard to the pharmacist's information provision. The aims were to clarify the perceptions of pharmacists and patients regarding information provision and the level of influence of those perceptions on patient satisfaction. METHODS: A cross-sectional survey with respect to information provision was conducted for patients and pharmacists in community pharmacies in Fukuoka Prefecture, Japan. In total, 407 patient-pharmacist pairs were included in a t-test and multilevel analysis. RESULTS: The levels of patient perception regarding information provision were significantly higher than the levels of pharmacist perception in all variables. The pharmacists' perceived level of information provision concerning medication effects had a negative and significant association with patient satisfaction, while the patients' perceived level of information provision by the pharmacist had a positive and significant association with patient satisfaction. Higher patient expectations regarding the level of information provision concerning medication side effects and older age of the pharmacist were adversely related to patient satisfaction. Both pharmacist and patient perceptions of the information provision by pharmacists personalized to the patient had positive associations with patient satisfaction. CONCLUSIONS: Pharmacist perceptions related to the information provision were not associated with patient satisfaction. The present study highlights accurate information provision, building good patient-pharmacist relationships, and improving pharmaceutical care in community pharmacy settings.

Sense of coherence, depression, and anger among adults with atopic dermatitis.

The prevalence of atopic disease (AD) in adults has significantly increased in industrialized countries. Psychological traits and lack of skills to cope with stress could be maintaining factors of AD. The first aim of the present study was to compare sense of coherence (SOC), depression, and anger among adult patients with AD, adults with remission from AD, and adults without AD. The second aim was to explore whether depression and anger have significant impacts on SOC. A cross-sectional survey was carried out from May to September of 2007. In total, 43 adult patients with AD, 32 adults with remission from AD, and 63 adults without AD participated. A one-way analysis of variance showed that adult patients with AD have significantly higher levels of depression than adults with remission from AD and adults without AD. Furthermore, structural equation modeling indicated a significant impact of depression on SOC across the three groups, along with a significant impact of anger suppression on depression, particularly in adult patients with AD. The findings of this study suggest that psychological traits have a negative impact on skills for coping with stress, and that this relationship might be a contributory factor for maintenance of AD.

Prevalence of loss-of-function FTO mutations in lean and obese individuals.

OBJECTIVE: Single nucleotide polymorphisms (SNPs) in intron 1 of fat mass- and obesity-associated gene (FTO) are strongly associated with human adiposity, whereas Fto(-/-) mice are lean and Fto(+/-) mice are resistant to diet-induced obesity. We aimed to determine whether FTO mutations are disproportionately represented in lean or obese humans and to use these mutations to understand structure-function relationships within FTO. RESEARCH DESIGN AND METHODS: We sequenced all coding exons of FTO in 1,433 severely obese and 1,433 lean individuals. We studied the enzymatic activity of selected nonsynonymous variants. RESULTS: We identified 33 heterozygous nonsynonymous variants in lean (2.3%) and 35 in obese (2.4%) individuals, with 8 mutations unique to the obese and 11 unique to the lean. Two novel mutations replace absolutely conserved residues: R322Q in the catalytic domain and R96H in the predicted substrate recognition lid. R322Q was unable to catalyze the conversion of 2-oxoglutarate to succinate in the presence or absence of 3-methylthymidine. R96H retained some basal activity, which was not enhanced by 3-methylthymidine. However, both were found in lean and obese individuals. CONCLUSIONS: Heterozygous, loss-of-function mutations in FTO exist but are found in both lean and obese subjects. Although intron 1 SNPs are unequivocally associated with obesity in multiple populations and murine studies strongly suggest that FTO has a role in energy balance, it appears that loss of one functional copy of FTO in humans is compatible with being either lean or obese. Functional analyses of FTO mutations have given novel insights into structure-function relationships in this enzyme.

Loss-of-function mutation in the dioxygenase-encoding FTO gene causes severe growth retardation and multiple malformations.

FTO is a nuclear protein belonging to the AlkB-related non-haem iron- and 2-oxoglutarate-dependent dioxygenase family. Although polymorphisms within the first intron of the FTO gene have been associated with obesity, the physiological role of FTO remains unknown. Here we show that a R316Q mutation, inactivating FTO enzymatic activity, is responsible for an autosomal-recessive lethal syndrome. Cultured skin fibroblasts from affected subjects showed impaired proliferation and accelerated senescence. These findings indicate that FTO is essential for normal development of the central nervous and cardiovascular systems in human and establish that a mutation in a human member of the AlkB-related dioxygenase family results in a severe polymalformation syndrome.

Correlation of receptor occupancy of metabotropic glutamate receptor subtype 1 (mGluR1) in mouse brain with in vivo activity of allosteric mGluR1 antagonists.

The aim of this study was to clarify the relationship between receptor occupancy and in vivo pharmacological activity of mGluR1 antagonists. The tritiated mGluR1-selective allosteric antagonist [(3)H]FTIDC (4-[1-(2-fluoropyridin-3-yl)-5-methyl-1H-1,2,3-triazol-4-yl]-N-isopropyl-N-methyl-3,6-dihydropyridine-1(2H)-carboxamide) was identified as a radioligand having high affinity for mGluR1-expressing CHO cells (K(D) = 2.1 nM) and mouse cerebellum (K(D) = 3.7 nM). [(3)H]FTIDC bound to mGluR1 was displaced by structurally unrelated allosteric antagonists, suggesting there is a mutual binding pocket shared with different allosteric antagonists. The binding specificity of [(3)H]FTIDC for mGluR1 in brain sections was demonstrated by the lack of significant binding to brain sections prepared from mGluR1-knockout mice. Ex vivo receptor occupancy with [(3)H]FTIDC revealed that the receptor occupancy level by FTIDC correlated well with FTIDC dosage and plasma concentration. Intracerebroventricular administration of (S)-3,5-dihydroxyphenylglycine is known to elicit face washing behavior that is mainly mediated by mGluR1. Inhibition of this behavioral change by FTIDC correlated with the receptor occupancy level of mGluR1 in the brain. A linear relationship between the receptor occupancy and in vivo activity was also demonstrated using structurally diverse mGluR1 antagonists. The receptor occupancy assays could help provide guidelines for selecting appropriate doses of allosteric mGluR1 antagonist for examining the function of mGluR1 in vivo.

Unique antipsychotic activities of the selective metabotropic glutamate receptor 1 allosteric antagonist 2-cyclopropyl-5-[1-(2-fluoro-3-pyridinyl)-5-methyl-1H-1,2,3-triazol-4-yl]-2,3-dihydro-1H-isoindol-1-one.

A newly discovered metabotropic glutamate receptor (mGluR) 1 allosteric antagonist, 2-cyclopropyl-5-[1-(2-fluoro-3-pyridinyl)-5-methyl-1H-1,2,3-triazol-4-yl]-2,3-dihydro-1H-isoindol-1-one (CFMTI), was tested both in vitro and in vivo for its pharmacological effects. CFMTI demonstrated potent and selective antagonistic activity on mGluR1 in vitro and in vivo after oral administration. CFMTI inhibited L-glutamate-induced intracellular Ca(2+) mobilization in Chinese hamster ovary cells expressing human and rat mGluR1a, with IC(50) values of 2.6 and 2.3 nM, respectively. The selectivity of CFMTI to mGluR1 over mGluR5 was >2000-fold, and CFMTI at 10 microM showed no agonistic or antagonistic activities toward other mGluR subtypes and other receptors. It antagonized face-washing behavior in mice induced by (S)-3,5-dihidroxyphenylglycine at a dose range of 3 to 30 mg/kg, for which receptor occupancy was 73 to 94%. As with the classical neuroleptic haloperidol and an atypical antipsychotic, clozapine, orally administered CFMTI reduced methamphetamine-induced hyperlocomotion and disruption of prepulse inhibition (PPI) at the same dose range as required to antagonize the face-washing behavior. CFMTI and clozapine improved ketamine-induced hyperlocomotion, PPI disruption and (5S,10R)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801)-induced social withdrawal without any cataleptogenic activities, whereas haloperidol only improved ketamine-induced hyperlocomotion. CFMTI, unlike clozapine, caused neither hypolocomotion nor motor incoordination at therapeutic doses. In c-fos expression studies, CFMTI and clozapine increased the number of fos-positive neurons in the nucleus accumbens and medial prefrontal cortex but not in the dorsolateral striatum. These results suggest that the antipsychotic activities of mGluR1 antagonists are more similar to those of atypical antipsychotics than those of typical antipsychotics.

Pharmacological characterization of a new, orally active and potent allosteric metabotropic glutamate receptor 1 antagonist, 4-[1-(2-fluoropyridin-3-yl)-5-methyl-1H-1,2,3-triazol-4-yl]-N-isopropyl-N-methyl-3,6-dihydropyridine-1(2H)-carboxamide (FTIDC).

A highly potent and selective metabotropic glutamate receptor (mGluR) 1 antagonist, 4-[1-(2-fluoropyridin-3-yl)-5-methyl-1H-1,2, 3-triazol-4-yl]-N-isopropyl-N-methyl-3,6-dihydropyridine-1(2H)-carboxamide (FTIDC), is described. FTIDC inhibits, with equal potency, l-glutamate-induced intracellular Ca(2+) mobilization in Chinese hamster ovary cells expressing human, rat, or mouse mGluR1a. The IC(50) value of FTIDC is 5.8 nM for human mGluR1a and 6200 nM for human mGluR5. The maximal response in agonist concentration-response curves was reduced in the presence of higher concentrations of FTIDC, suggesting the inhibition in a noncompetitive manner. FTIDC at 10 microM showed no agonistic, antagonistic, or positive allosteric modulatory activity toward mGluR2, mGluR4, mGluR6, mGluR7, or mGluR8. FTIDC did not displace [(3)H]l-quisqualate binding to human mGluR1a, indicating FTIDC is an allosteric antagonist. Studies using chimeric and mutant receptors of mGluR1 showed that transmembrane (TM) domains 4 to 7, especially Phe801 in TM6 and Thr815 in TM7, play pivotal roles in the antagonism of FTIDC. FTIDC inhibited the constitutive activity of mGluR1a, suggesting that FTIDC acts as an inverse agonist of mGluR1a. Intraperitoneally administered FTIDC inhibited face-washing behavior elicited by a group I mGluR agonist, (S)-3,5-dihydroxyphenylglycine in mice at doses that did not produce motor impairment. Oral administration of FTIDC also inhibited the face-washing behavior. FTIDC is a highly potent and selective allosteric mGluR1 antagonist and a compound having oral activity without species differences in its antagonistic activity on recombinant human, mouse, and rat mGluR1. FTIDC could therefore be a valuable tool for elucidating the functions of mGluR1 not only in rodents but also in humans.

Probing the secondary structure of salmon SmaI SINE RNA.

SmaI is a short interspersed element (SINE) of the salmon genome, and is derived from tRNA(Lys). We probed the secondary structure of SmaI SINE RNA by enzymatic cleavage and found that the RNA structure comprises three separate domains. The 5'-terminal region (the 5' domain) forms a tRNA-like cloverleaf structure, whereas the 3'-terminal region (the 3' domain) forms an extended stem-loop. The loop region is thought to be recognized by the reverse transcriptase (RT) encoded by the long interspersed element (LINE). The two structural domains are linked by a single-stranded region (the linker domain). Our melting profile analyses indicated the presence of two structural domains having different thermal stabilities, thus supporting the domain composition described above. Based on these results, we discuss the structural generality and evolutionary advantage of the domain composition of SINE RNA.

Inhibitory effect of norditerpenes on LPS-induced TNF-alpha production from the Okinawan soft coral, Sinularia sp.

An ethanol (EtOH) extract of the soft coral (Sinularia sp.), collected in Okinawa, demonstrated a potent inhibitory effect on lipopolysaccharide (LPS)-induced tumor necrosis factor-alpha (TNF-alpha) production by murine macrophage-like cells (RAW264.7). The activity-guided purification of the EtOH extract resulted in the isolation of two norditerpenes, norcembrenolide (1) and sinuleptolide (2). These structures were identified from the spectroscopic data. Norcembrenolide (1) and sinuleptolide (2) inhibited TNF-alpha production in a dose-dependent manner, and showed a more potent effect than prednisolone at the concentration of 33 microg/ml. They also exhibited an inhibitory effect on nitric oxide (NO) production not influenced by cytotoxicity.