Differential scanning fluorimetric analysis of the amino-acid binding to taste receptor using a model receptor protein, the ligand-binding domain of fish T1r2a/T1r3

Taste receptor type 1 (T1r) is responsible for the perception of essential nutrients, such as sugars and amino acids, and evoking sweet and umami (savory) taste sensations. T1r receptors recognize many of the taste substances at their extracellular ligand-binding domains (LBDs). In order to detect a wide array of taste substances in the environment, T1r receptors often possess broad ligand specificities. However, the entire ranges of chemical spaces and their binding characteristics to any T1rLBDs have not been extensively analyzed. In this study, we exploited the differential scanning fluorimetry (DSF) to medaka T1r2a/T1r3LBD, a current sole T1rLBD heterodimer amenable for recombinant preparation, and analyzed their thermal stabilization by adding various amino acids. The assay showed that the agonist amino acids induced thermal stabilization and shifted the melting temperatures (T-m) of the protein. An agreement between the DSF results and the previous biophysical assay was observed, suggesting that DSF can detect ligand binding at the orthostericbinding site in T1r2a/T1r3LBD. The assay further demonstrated that most of the tested Lamino acids, but no D-amino acid, induced T-m shifts of T1r2a/T1r3LBD, indicating the broad L-amino acid specificities of the proteins probably with several different manners of recognition. The T-m shifts by each amino acid also showed a fair correlation with the responses exhibited by the full-length receptor, verifying the broad amino-acid binding profiles at the orthosteric site in LBD observed by DSF

Circumstances of Mountain Accidents in Japan : Problems of Senior-aged Persons

Fatal mountain accidents in Japan in the year 1994 was the worst record in years, because a total of 206 persons in 774 incidents lost their lives. Our concern of "Record Number of Accidents" is entirely due to a large increase of accidents that involve senior-aged persons. In comparison with the year 1979, the proportion of senior-aged persons as victims increased from 23% to 67%. This is because a mountaineering boom came with the decrease in labor hours from 1988 and with the increased interest in nature. A number of papers and reports for researching the mountain accidents, however, are very poor except for a lot of mountaineering guide-books and monthly magazines. Moreover, the authors of guide-books who believe in their own evidence have sometimes made comments on the mountain accidents without hard data of accidents. In this study, the data by the police headquarters ,which were collected from all branches in each prefecture in Japan, helped to analyze the circumstances of mountain accidents. Main items used in the police data were cause, activity, terrain, victim identities, and ages. The increment rate of incidents of senior-aged persons was 3.2% each year. So more increase of the senior aged victims persons will be expected in the near future because the baby-boom generation is getting older. The most common causes in mountain accidents were falls and slips for those under the ,age of 55, while the cause for people over 55 was getting lost. The parameters of getting lost will be a useful index to explain the spatial recognition ability in the mountain accidents from childhood to old age. The statistics analyzed for mountain accidents could pointed out the problems involved in considering a group of people over the age of 40 as only one group and will be a indicator how to consider the mountain accidents and aging process of hikers and climbers

17β-Hydroxysteroid Dehydrogenase Type 2 Expression Is Induced by Androgen Signaling in Endometrial Cancer.

Endometrial cancer is one of the most common female pelvic cancers and has been considered an androgen-related malignancy. Several studies have demonstrated the anti-cell proliferative effect of androgen on endometrial cancer cells; however, the mechanisms of the anti-cancer effect of androgen remain largely unclear. 17β-hydroxysteroid dehydrogenase type 2 (17β-HSD2), which catalyzes the conversion of E2 to E1, is known to be upregulated by androgen treatment in breast cancer cells. In this study, we therefore focused on the role of androgen on estrogen dependence in endometrial cancer. Dihydrotestosterone (DHT) was found to induce 17β-HSD2 mRNA and protein expression in HEC-1B endometrial cancer cells. DHT could also inhibit cell proliferation of HEC-1B when induced by estradiol treatment. In 19 endometrioid endometrial adenocarcinoma (EEA) tissues, intratumoral DHT concentration was measured by liquid chromatography/electrospray tandem mass spectrometry and was found to be significantly correlated with 17β-HSD2 immunohistochemical status. We further examined the correlations between 17β-HSD2 immunoreactivity and clinicopathological parameters in 53 EEA tissues. 17β-HSD2 status was inversely associated with the histological grade, clinical stage, and cell proliferation marker Ki-67, and positively correlated with progesterone receptor expression. 17β-HSD2 status tended to be positively associated with androgen receptor status. In 53 EEA cases, the 17β-HSD2-positive group tended to have better prognosis than that for the negative group with respect to progression-free survival and endometrial cancer-specific survival. These findings suggest that androgen suppresses the estrogen dependence of endometrial cancer through the induction of 17β-HSD2 in endometrial cancer

STAT3 gene mutations and their association with pure red cell aplasia in large granular lymphocyte leukemia

Large granular lymphocyte leukemia (LGLL) has been morphologically characterized as a group of lymphoproliferative diseases that include T-cell large granular lymphocytic leukemia (T-LGLL) and chronic lymphoproliferative disorders of natural killer cells (CLPD-NK). We investigated mutations in the Src homology 2 (SH2) domain of the signal transducer and activator of transcription 3 (STAT3) gene in Asian cohorts of T-LGLL and CLPD-NK (n=42 and 11, respectively). Two mutations, Y640F and D661Y, were identified using direct sequencing or allele-specific (AS) PCR. Y640F and D661Y mutations were found in seven and 18 patients, respectively. Two patients were positive for both mutations. Frequencies of STAT3 mutations in T-LGLL and CLPD-NK were 47.6% and 27.2%, respectively. Pure red cell aplasia (PRCA) was associated with the mutations (P=0.005). The mutations were persistently found at stable levels in some patients after more than 5years using AS-quantitative PCR. The results of the present study indicate that the SH2 domain of the STAT3 gene is frequently mutated in Asian T-LGLL and CLPD-NK, and that PRCA is closely correlated with the mutations. SH2 domain of the STAT3 gene is frequently mutated in Asian T cell large granular lymphocyte leukemia and chronic lymphoproliferative disorders of NK cells. Pure red cell aplasia is closely associated with the mutations.ArticleCANCER SCIENCE. 105(3):342-346 (2014)journal articl

Pillar[6]arene acts as a biosensor for quantitative detection of a vitamin metabolite in crude biological samples

ビタミン代謝物を迅速定量できる超分子バイオセンサーを開発. 京都大学プレスリリース. 2020-12-09.Metabolic syndrome is associated with obesity, hypertension, and dyslipidemia, and increased cardiovascular risk. Therefore, quick and accurate measurements of specific metabolites are critical for diagnosis; however, detection methods are limited. Here we describe the synthesis of pillar[n]arenes to target 1-methylnicotinamide (1-MNA), which is one metabolite of vitamin B3 (nicotinamide) produced by the cancer-associated nicotinamide N-methyltransferase (NNMT). We found that water-soluble pillar[5]arene (P5A) forms host–guest complexes with both 1-MNA and nicotinamide, and water-soluble pillar[6]arene (P6A) selectively binds to 1-MNA at the micromolar level. P6A can be used as a “turn-off sensor” by photoinduced electron transfer (detection limit is 4.38 × 10−6 M). In our cell-free reaction, P6A is used to quantitatively monitor the activity of NNMT. Moreover, studies using NNMT-deficient mice reveal that P6A exclusively binds to 1-MNA in crude urinary samples. Our findings demonstrate that P6A can be used as a biosensor to quantify 1-MNA in crude biological samples

Ａ大学附属病院救命救急センターにおける心不全患者の心臓リハビリテーションの実施状況

急性期心臓リハビリテーション（以下，心リハ）導入に向け，入院から退院までの心不全患者の状態や実施されたリハビリテーションの状況を明らかにすることを目的に，Ａ大学附属病院救命救急センター（以下，センター）に入院した心不全患者の心リハの実施状況を調査した．調査は，平成26年４月から28年８月にセンターに入院した心不全患者の診療録から，基本属性，治療経過，心リハの実施状況について情報を収集し分析した．医師による心リハ処方の有無および心リハ処方までの期間を中央値で２群に分け，各調査項目について比較検討し，より詳細にセンターに入院した心不全患者の心リハの実施状況を分析した．結果，対象となった患者は164名であり，うち心リハ処方ありは123名（75％）であった．心リハ処方ありの群はなしの群に比べ，入院期間，センター在院日数が有意に長かった．また，処方ありの群の転帰は死亡退院が少なかった．心リハ処方までの期間の中央値は６日であり，心リハ処方６日以上群は，６日未満群に比べ，入院期間，センター在院日数が有意に長かった．また，心リハ処方６日以上群はBNP が高値であり，昇圧剤使用，CHDFおよびIABPの機器使用が多かった．心リハ開始の判断は，重症度から安静臥床を余儀なくされたものと推察され，重症度に応じた心リハプログラムの開発が必要であると考える．The aim of this study was to clarify the current status of cardiac rehabilitation practice in preparation for the introduction of cardiac rehabilitation in the acute phase. Subjects were patients with heart failure who were admitted to the Center for Emergency and Critical Care (hereafter the Center) of University Hospital A from April 2014 to August 2016. The medical records of 164 such patients were reviewed and patient characteristics, course of treatment, and implementation of cardiac rehabilitation were examined. The patients were then divided into 2 groups based on the median time to prescription of cardiac rehabilitation as cutoff, and the examined parameters were compared between the groups. Cardiac rehabilitation was prescribed for 123 of the 164 patients (75%). Median time to prescription of cardiac rehabilitation was 6 days. Length of hospital stay and stay in the Center were significantly longer when the time to prescription of cardiac rehabilitation was ≥ 6 days compared with when it was ≥ 6 days. When the time to prescription of cardiac rehabilitation was < 6 days, brain natriuretic peptide level was elevated and a higher proportion of patients were treated with pressor agents, continuous hemodiafiltration, and intra-aortic balloon pump devices. It is important to establish an interprofessional system and cardiac rehabilitation program that together facilitate early and safe commencement of rehabilitation from the acute stage

Display of both N- and C-terminal target fusion proteins on the Aspergillus oryzae cell surface using a chitin-binding module

A novel cell surface display system in Aspergillus oryzae was established by using a chitin-binding module (CBM) from Saccharomyces cerevisiae as an anchor protein. CBM was fused to the N or C terminus of green fluorescent protein (GFP) and the fusion proteins (GFP-CBM and CBM-GFP) were expressed using A. oryzae as a host. Western blotting and fluorescence microscopy analysis showed that both GFP-CBM and CBM-GFP were successfully expressed on the cell surface. In addition, cell surface display of triacylglycerol lipase from A. oryzae (tglA), while retaining its activity, was also successfully demonstrated using CBM as an anchor protein. The activity of tglA was significantly higher when tglA was fused to the C terminus than N terminus of CBM. Together, these results show that CBM used as a first anchor protein enables the fusion of both the N and/or C terminus of a target protein

Emerin plays a crucial role in nuclear invagination and in the nuclear calcium transient.

Alteration of the nuclear Ca2+ transient is an early event in cardiac remodeling. Regulation of the nuclear Ca2+ transient is partly independent of the cytosolic Ca2+ transient in cardiomyocytes. One nuclear membrane protein, emerin, is encoded by EMD, and an EMD mutation causes Emery-Dreifuss muscular dystrophy (EDMD). It remains unclear whether emerin is involved in nuclear Ca2+ homeostasis. The aim of this study is to elucidate the role of emerin in rat cardiomyocytes by means of hypertrophic stimuli and in EDMD induced pluripotent stem (iPS) cell-derived cardiomyocytes in terms of nuclear structure and the Ca2+ transient. The cardiac hypertrophic stimuli increased the nuclear area, decreased nuclear invagination, and increased the half-decay time of the nuclear Ca2+ transient in cardiomyocytes. Emd knockdown cardiomyocytes showed similar properties after hypertrophic stimuli. The EDMD-iPS cell-derived cardiomyocytes showed increased nuclear area, decreased nuclear invagination, and increased half-decay time of the nuclear Ca2+ transient. An autopsied heart from a patient with EDMD also showed increased nuclear area and decreased nuclear invagination. These data suggest that Emerin plays a crucial role in nuclear structure and in the nuclear Ca2+ transient. Thus, emerin and the nuclear Ca2+ transient are possible therapeutic targets in heart failure and EDMD. © The Author(s) 2017