Significant association of urokinase plasminogen activator Pro141Leu with serum lipid profiles in a Japanese population

Urokinase plasminogen activator (uPA) plays important physiological and pathological roles in fibrinolysis, cancer metastasis, and atherosclerosis. One study suggested that uPA also has a major role in cholesterol biosynthesis in humans via its receptor uPAR. Thus, we investigated the associations of functional uPA polymorphism (plasminogen activator, urokinase; PLAU Pro141Leu, rs2227564) with serum lipid profiles in a Japanese cohort. The study included 5152 participants (1465 male, 3687 female; age range, 35–69 years) of the Daiko Study, a part of the Japan Multi-Institutional Collaborative Cohort Study (J-MICC Study). Subjects were enrolled at the Daiko Medical Center from June 2008 to May 2010. Low-density lipoprotein cholesterol (LDL-C) and non-HDL-C (subtraction of high-density lipoprotein cholesterol from total cholesterol) in fasting blood of participants were each classified into two groups, < or ≥ 140 mg/dL, and < or ≥ 170 mg/dL, respectively. Genotype frequencies of PLAU Pro141Leu (rs2227564) were 59.1% for ProPro, 35.6% for ProLeu, and 5.3% for LeuLeu, and were in Hardy–Weinberg equilibrium (p = 0.789). The allele frequencies were 0.769 for Pro and 0.231 for Leu. The multivariate-adjusted odd ratios (ORs) and 95% confidence intervals (CIs) for high LDL-C and non-HDL-C were 1.11 (95%CI; 1.00–1.23) and 1.16 (95%CI; 1.03–1.30) for those with Leu allele relative to ProPro. This study suggested that PLAU Pro141Leu (rs2227564) is significantly associated with serum lipid levels in a Japanese population.     

Validity and reliability of a visual scoring method for masticatory ability using test gummy jelly

Background: For quantitative evaluation of masticatory ability of the elderly patients, there should be a simple and reliable method without special techniques and instruments. Objective: The purpose of this study was to examine the validity and reliability of a visual scoring method for assessing masticatory performance. Materials and Methods: A 10‐stage scale for visually scoring was rated based on the range of the glucose concentration dissolved from comminuted jelly. Photographic images of comminuted jellies were produced as a standard material for each score. Fifty subjects were recruited as raters who graded the visual score for 50 photographic images of comminuted jellies on the screen of a lap‐top three times in random order. Results: There were strong correlations (rs = 0.911– 0.981, Spearman’s rank coefficient) between the actual scores determined from the glucose concentration and the visual scores graded by subjects in all three measurements. The intraclass correlation coefficients (ICCs) of the inter‐rater reliability and the ICCs of the intra‐rater reliability of the visual scoring ranged from 0.946 to 0.947 and from 0.860 to 0.987 in three measurements, respectively. Conclusions: These results indicated that the visual scoring method was valid and reliable for evaluation of masticatory performance.     

Abrogation of neutral cholesterol ester hydrolytic activity causes adrenal enlargement

We have previously demonstrated that neutral cholesterol ester hydrolase 1 (Nceh1) regulates foam cell formation and atherogenesis through the catalytic activity of cholesterol ester hydrolysis, and that Nceh1 and hormone-sensitive lipase (Lipe) are responsible for the majority of neutral cholesterol ester hydrolase activity in macrophages. There are several cholesterol ester-metabolizing tissues and cells other than macrophages, among which adrenocortical cells are also known to utilize the intracellular cholesterol for steroidogenesis. It has been believed that the mobilization of intracellular cholesterol ester in adrenal glands was facilitated solely by Lipe. We herein demonstrate that Nceh1 is also involved in cholesterol ester hydrolysis in adrenal glands. While Lipe deficiency remarkably reduced the neutral cholesterol ester hydrolase activity in adrenal glands as previously reported, additional inactivation of Nceh1 gene completely abrogated the activity. Adrenal glands were enlarged in proportion to the degree of reduced neutral cholesterol ester hydrolase activity, and the enlargement of adrenal glands and the accumulation of cholesterol esters were most pronounced in the Nceh1/Lipe double-deficient mice. Thus Nceh1 is involved in the adrenal cholesterol metabolism, and the cholesterol ester hydrolytic activity in adrenal glands is associated with the organ enlargement.     

Induction of antigen-specific immunity by pH-sensitive carbonate apatite as a potent vaccine carrier

The ability of carbonate apatite (CO₃Ap) to enhance antigen-specific immunity was examined in vitro and in vivo to investigate its utility as a vaccine carrier. Murine bone marrow-derived dendritic cells took up ovalbumin (OVA) containing CO₃Ap more effectively than free OVA. Interestingly, mice immunized with OVA-containing CO₃Ap produced OVA-specific antibodies more effectively than mice immunized with free OVA. Furthermore, immunization of C57BL/6 mice with OVA-containing CO₃Ap induced the proliferation and antigen-specific production of IFN-γ by splenocytes more strongly than immunization with free OVA. Moreover, no significant differences were detected in the induction of delayed-type hypersensitivity responses, an immune reaction involving an antigen-specific, cell-mediated immune response between OVA-containing CO₃Ap and OVA-containing alumina salt (Alum), suggesting that CO₃Ap induced cell-mediated immune response to the same degree as Alum, which is commonly used for clinical applications. This study is the first to demonstrate the induction of antigen-specific immune responses in vivo by CO₃Ap.     

Hes1 regulates the number and anterior-posterior patterning of mesencephalic dopaminergic neurons at the mid/hindbrain boundary (isthmus)

The lack of the Hes1 gene leads to the failure of cranial neurulation due to the premature onset of neural differentiation. Hes1 homozygous null mutant mice displayed a neural tube closure defect, and exencephaly was induced at the mid/hindbrain boundary. In the mutant mesencephalon, the roof plate was not formed and therefore the ventricular zone showing cell proliferation was displaced to the brain surface. Furthermore, the telencephalon and ventral diencephalon were defective. Despite the severe defects of neurogenesis in null mutants, the mesencephalic dopaminergic (mesDA) neurons were specified at the midline of the ventral mesencephalon in close proximity to two important signal centers — floor plate and mid/hindbrain boundary (i.e., the isthmic organizer). Using mesDA neuronal markers, tyrosine hydroxylase (TH) and Pitx3, the development of mesDA neurons was studied in Hes1 null mice and compared with that in the wild type. At early stages, between embryonic day (E) 11.5 and E12.5, mesDA neurons were more numerous in null mutants than in the wild type. From E13.5 onward, however, the cell number and fiber density of mesDA neurons were decreased in the mutants. Their distribution pattern was also different from that of the wild type. In particular, mesDA neurons grew dorsally and invaded the rostral hindbrain. 5-HT neurons were also ectopically located in the mutant midbrain. Thus, the loss of Hes1 resulted in disturbances in the inductive and repulsive activities of the isthmic organizer. It is proposed that Hes1 plays a role in regulating the location and density of mesDA neurons.     

Inhibitors of the Influenza A Virus M2 Proton Channel Discovered Using a High-Throughput Yeast Growth Restoration Assay

The M2 proton channel of the influenza A virus is the target of the anti-influenza drugs amantadine and rimantadine. The effectiveness of these drugs has been dramatically limited by the rapid spread of drug resistant mutations, mainly at sites S31N, V27A and L26F in the pore of the channel. Despite progress in designing inhibitors of V27A and L26F M2, there are currently no drugs targeting these mutated channels in clinical trials. Progress in developing new drugs has been hampered by the lack of a robust assay with sufficient throughput for discovery of new active chemotypes among chemical libraries and sufficient sensitivity to provide the SAR data essential for their improvement and development as drugs. In this study we adapted a yeast growth restoration assay, in which expression of the M2 channel inhibits yeast growth and exposure to an M2 channel inhibitor restores growth, into a robust and sensitive high-throughput screen for M2 channel inhibitors. A screen of over 250,000 pure chemicals and semi-purified fractions from natural extracts identified 21 active compounds comprising amantadine, rimantadine, 13 related adamantanes and 6 non-adamantanes. Of the non-adamantanes, hexamethylene amiloride and a triazine derivative represented new M2 inhibitory chemotypes that also showed antiviral activity in a plaque reduction assay. Of particular interest is the fact that the triazine derivative was not sufficiently potent for detection as an inhibitor in the traditional two electrode voltage clamp assay for M2 channel activity, but its discovery in the yeast assay led to testing of analogues of which one was as potent as amantadine.     

Isolation and Characterization of Antimicrobial Compounds in Plant Extracts against Multidrug-Resistant Acinetobacter baumannii

The number of fully active antibiotic options that treat nosocomial infections due to multidrug-resistant Acinetobacter baumannii (A. baumannii) is extremely limited. Magnolia officinalis, Mahonia bealei, Rabdosia rubescens, Rosa rugosa, Rubus chingii, Scutellaria baicalensis, and Terminalia chebula plant extracts were previously shown to have growth inhibitory activity against a multidrug-resistant clinical strain of A. baumannii. In this study, the compounds responsible for their antimicrobial activity were identified by fractionating each plant extract using high performance liquid chromatography, and determining the antimicrobial activity of each fraction against A. baumannii. The chemical structures of the fractions inhibiting >40% of the bacterial growth were elucidated by liquid chromatography/mass spectrometry analysis and nuclear magnetic resonance spectroscopy. The six most active compounds were identified as: ellagic acid in Rosa rugosa; norwogonin in Scutellaria baicalensis; and chebulagic acid, chebulinic acid, corilagin, and terchebulin in Terminalia chebula. The most potent compound was identified as norwogonin with a minimum inhibitory concentration of 128 µg/mL, and minimum bactericidal concentration of 256 µg/mL against clinically relevant strains of A. baumannii. Combination studies of norwogonin with ten anti-Gram negative bacterial agents demonstrated that norwogonin did not enhance the antimicrobial activity of the synthetic antibiotics chosen for this study. In conclusion, of all identified antimicrobial compounds, norwogonin was the most potent against multidrug-resistant A. baumannii strains. Further studies are warranted to ascertain the prophylactic and therapeutic potential of norwogonin for infections due to multidrug-resistant A. baumannii.     

PTIP Associated protein 1, PA1, Is an Independent Prognostic Factor for Lymphnode Negative Breast Cancer

Pax transactivation domain interacting protein (PTIP) associated protein 1, PA1, was a newly found protein participating in the modulation of transactivity of nuclear receptor super family members such as estrogen receptor (ER), androgen receptor (AR) and glucocorticoid receptor (GR). Breast cancer is one of the most life threatening diseases for women and has tight association with estrogen and ER. This study was performed to understand the function of PA1 in breast cancer. The expression of PA1 had been evaluated in a total of 344 primary invasive breast cancer samples and examined the relationship with clinical output, relapse free survival (RFS), breast cancer-specific survival (BCSS). PA1 expression was observed in both nucleus and cytoplasm, however, appeared mainly in nuclear. PA1 nuclear expression was correlated with postmenopausal (P = 0.0097), smaller tumor size (P = 0.0025), negative Ki67 (P = 0.02), positive AR (P = 0.049) and positive ERβ (P = 0.0020). Kaplan–Meier analysis demonstrated PA1 nuclear positive cases seemed to have a longer survival than negative ones for RFS (P = 0.023) but not for BCSS (P = 0.23). In the Cox hazards model, PA1 nuclear protein expression proved to be a significant prognostic univariate parameter for RFS (P = 0.03), but not for BCSS (P = 0.20). In addition, for those patients without lymphnode metastasis PA1 was found to be an independent prognostic factor for RFS (P = 0.025), which was verified by univariate and multivariate analyses. These investigations suggested PA1 expression could be a potential prognostic indicator for RFS in breast cancer.