Effectiveness of Artificial Neural Networks for Solving Inverse Problems in Magnetic Field-Based Localization.

Recently, indoor localization has become an active area of research. Although there are various approaches to indoor localization, methods that utilize artificially generated magnetic fields from a target device are considered to be the best in terms of localization accuracy under non-line-of-sight conditions. In magnetic field-based localization, the target position must be calculated based on the magnetic field information detected by multiple sensors. The calculation process is equivalent to solving a nonlinear inverse problem. Recently, a machine-learning approach has been proposed to solve the inverse problem. Reportedly, adopting the k-nearest neighbor algorithm (k-NN) enabled the machine-learning approach to achieve fairly good performance in terms of both localization accuracy and computational speed. Moreover, it has been suggested that the localization accuracy can be further improved by adopting artificial neural networks (ANNs) instead of k-NN. However, the effectiveness of ANNs has not yet been demonstrated. In this study, we thoroughly investigated the effectiveness of ANNs for solving the inverse problem of magnetic field-based localization in comparison with k-NN. We demonstrate that despite taking longer to train, ANNs are superior to k-NN in terms of localization accuracy. The k-NN is still valid for predicting fairly accurate target positions within limited training times.

X-ray structures of catalytic intermediates of cytochrome c oxidase provide insights into its O2 activation and unidirectional proton-pump mechanisms.

Cytochrome c oxidase (CcO) reduces O2 to water, coupled with a proton-pumping process. The structure of the O2-reduction site of CcO contains two reducing equivalents, Fe a32+ and CuB1+, and suggests that a peroxide-bound state (Fe a33+-O--O--CuB2+) rather than an O2-bound state (Fe a32+-O2) is the initial catalytic intermediate. Unexpectedly, however, resonance Raman spectroscopy results have shown that the initial intermediate is Fe a32+-O2, whereas Fe a33+-O--O--CuB2+ is undetectable. Based on X-ray structures of static noncatalytic CcO forms and mutation analyses for bovine CcO, a proton-pumping mechanism has been proposed. It involves a proton-conducting pathway (the H-pathway) comprising a tandem hydrogen-bond network and a water channel located between the N- and P-side surfaces. However, a system for unidirectional proton-transport has not been experimentally identified. Here, an essentially identical X-ray structure for the two catalytic intermediates (P and F) of bovine CcO was determined at 1.8 Šresolution. A 1.70 ŠFe-O distance of the ferryl center could best be described as Fe a34+ = O2-, not as Fe a34+-OH- The distance suggests an ∼800-cm-1 Raman stretching band. We found an interstitial water molecule that could trigger a rapid proton-coupled electron transfer from tyrosine-OH to the slowly forming Fe a33+-O--O--CuB2+ state, preventing its detection, consistent with the unexpected Raman results. The H-pathway structures of both intermediates indicated that during proton-pumping from the hydrogen-bond network to the P-side, a transmembrane helix closes the water channel connecting the N-side with the hydrogen-bond network, facilitating unidirectional proton-pumping during the P-to-F transition.

Determination of ω-6 and ω-3 PUFA metabolites in human urine samples using UPLC/MS/MS.

The ω-6 and ω-3 polyunsaturated fatty acids (PUFAs) such as arachidonic acid (AA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA) are the precursors of various bioactive lipid mediators including prostaglandins, thromboxanes, leukotrienes, hydroxyeicosatetraenoic acid, isoprostanes, lipoxins, and resolvins (Rvs). These lipid mediators play important roles in various physiological and pathological processes. The quantitative determination of PUFA metabolites seems necessary for disease research and for developing biomarkers. However, there is a paucity of analytical methods for the quantification of ω-6 and ω-3 PUFA metabolites­the specialized pro-resolving mediators (SPMs) present in the human urine. We developed a method for the quantification of ω-6 and ω-3 PUFA metabolites present in human urine using ultra-performance liquid chromatography/tandem mass spectrometry (UPLC/MS/MS). The developed method shows good linearity, with a correlation coefficient >0.99 for all of the analytes. The validation results indicate that our method is adequately reliable, accurate, and precise. The method was successfully used to examine urine samples obtained from 43 healthy volunteers. We could identify 20 PUFA metabolites, and this is the first report of the quantitative determination of RvD1, 17(R)-RvD1, 11-dehydro thromboxane B3, RvE2, and 5(S)-HETE in human urine. The urinary 8-iso PGF(2α) and PGE2 levels were significantly higher in the men smokers than in the men nonsmokers (p < 0.05). In this study, we developed an accurate, precise, and novel analytical method for estimating the ω-6 and ω-3 PUFA metabolites, and this is the first report that the SPMs derived from EPA and DHA are present in human urine.

Knowledge and Attitude towards Cervical Cancer and Human Papillomavirus among Pharmacists in Japan.

OBJECTIVE: The objective of this study was to assess the knowledge and attitude towards cervical cancer and human papillomavirus (HPV) among pharmacists in Japan. METHODS: Questionnaires were disseminated to 788 pharmacists employed by the Tsuruha Holdings Inc. A total of 617 pharmacists responded, generating a response rate of 78.3%. RESULT: Of the 362 females and 255 males, vaccination rates were 14.4% and 0.8%, respectively. In terms of cervical cytology, 35.1% of females received it once every two years, and 26.2% received it irregularly. As for HPV testing, 12.2% of females received it once every two years, and 16.6% received it irregularly. The rate of "school curriculum" as an information source was significantly higher among younger pharmacists; while "internet", "media", "training seminar for pharmacist", "advertisement in medical institution", "internal manual", and "others" were significantly higher among older pharmacists. The proportion of pharmacists with knowledge on general questions, except for those about HPV testing, was significantly higher among females than males. The vaccination rates of younger pharmacists were significantly higher than those of older pharmacists. The screening rates of cervical cytology were significantly higher among older than younger pharmacists, and also among those with at least 10 years of experience than those with less. There were no differences in the screening rates of HPV testing according to age or pharmacist experience. CONCLUSION: The proportion of pharmacists with knowledge about cervical cancer and HPV significantly varied depending on sex, age, and experience as a pharmacist. This study suggested that spreading the knowledge about cervical cancer and HPV might be effective for increasing the rates of cervical cancer screening.

Organic anion-transporting polypeptide (OATP) 2B1 contributes to the cellular uptake of theaflavin.

Organic anion-transporting polypeptide (OATP) 2B1 has been reported in the apical membranes of the human small intestinal epithelium, where it contributes to the intestinal absorption of pharmacologically active drugs. To investigate the potential for OATP2B1-mediated drug-food interactions, the effects of several polyphenolic compounds on OATP2B1-mediated estrone-3-sulfate (E3S) transport were studied by using OATP2B1-expressing HEK293 cells. Our results showed that some compounds, especially theaflavin, were strong inhibitors of OATP2B1-mediated E3S uptake. Theaflavin showed a significantly higher uptake into the OATP2B1-expressing HEK293 cells than the control cells. The concentration dependence of the uptake of theaflavin was determined over a range of concentrations (0.5-100 µM) and the kinetic parameters (Km and Vmax) of theaflavin uptake were found to be 5.12 ± 0.67 µM and 41.6 ± 1.3 pmol/mg protein/min, respectively. The OATP2B1-mediated theaflavin uptake was inhibited by known OATP2B1 substrates such as E3S, bromsulphthalein (BSP), dehydroepiandrosterone-3-sulfate (DHEAS), and fluvastatin. Our results indicate that theaflavin is a novel substrate of OATP2B1. The results of this study might be helpful to predict the potential OATP2B1-mediated drug-theaflavin interactions and to avoid undesirable clinical consequences.

Spectroscopic and DNA-binding characterization of the isolated heme-bound basic helix-loop-helix-PAS-A domain of neuronal PAS protein 2 (NPAS2), a transcription activator protein associated with circadian rhythms.

Neuronal PAS domain protein 2 (NPAS2) is a circadian rhythm-associated transcription factor with two heme-binding sites on two PAS domains. In the present study, we compared the optical absorption spectra, resonance Raman spectra, heme-binding kinetics and DNA-binding characteristics of the isolated fragment containing the N-terminal basic helix-loop-helix (bHLH) of the first PAS (PAS-A) domain of NPAS2 with those of the PAS-A domain alone. We found that the heme-bound bHLH-PAS-A domain mainly exists as a dimer in solution. The Soret absorption peak of the Fe(III) complex for bHLH-PAS-A (421 nm) was located at a wavelength 9 nm higher than for isolated PAS-A (412 nm). The axial ligand trans to CO in bHLH-PAS-A appears to be His, based on the resonance Raman spectra. In addition, the rate constant for heme association with apo-bHLH-PAS (3.3 x 10(7) mol(-1) x s(-1)) was more than two orders of magnitude higher than for association with apo-PAS-A (< 10(5) mol(-1) x s(-1)). These results suggest that the bHLH domain assists in stable heme binding to NPAS2. Both optical and resonance Raman spectra indicated that the Fe(II)-NO heme complex is five-coordinated. Using the quartz-crystal microbalance method, we found that the bHLH-PAS-A domain binds specifically to the E-box DNA sequence in the presence, but not in the absence, of heme. On the basis of these results, we discuss the mode of heme binding by bHLH-PAS-A and its potential role in regulating DNA binding.

Differentiation of induced pluripotent stem cells into dental mesenchymal cells.

Similar to embryonic stem cells, induced pluripotent stem (iPS) cells can differentiate into various cell types upon appropriate induction, and thus, may be valuable cell sources for regenerative medicine. However, iPS cells have not been reported to differentiate into odontogenic cells for tooth regeneration. Here we demonstrated that neural crest-like cells (NCLC) derived from mouse iPS cells have the potential to differentiate into odontogenic mesenchymal cells. We developed an efficient culture protocol to induce the differentiation of mouse iPS cells into NCLC. We confirmed that the cells exhibited neural crest (NC) cell markers as evidenced by immunocytochemistry, flow cytometry, and real-time reverse transcription-polymerase chain reaction. Further, in recombination cultures of NCLC and mouse dental epithelium, NCLC exhibited a gene expression pattern involving dental mesenchymal cells. Some NCLC also expressed dentin sialoprotein. Conditioned medium of mouse dental epithelium cultures further enhanced the differentiation of NCLC into odontoblasts. These results suggest that iPS cells are useful cell sources for tooth regeneration and tooth development studies.

Endomorphin 2 analogues containing Dmp residue as an aromatic amino acid surrogate with high mu-opioid receptor affinity and selectivity.

To investigate the effectiveness of a 2',6'-dimethylphenylalanine (Dmp) residue as an aromatic amino acid surrogate, endomorphin 2 (EM(2): Tyr-Pro-Phe-Phe-NH(2)) analogues were prepared, in which the constitutive aromatic amino acids (Tyr(1), Phe(3), or Phe(4)) were replaced by Dmp or its isomer, D-Dmp. Replacement of Phe(3) by Dmp increased the affinity over 10-fold for both mu- and delta-opioid receptors, without affecting receptor selectivity. In contrast, replacement of Phe(4) considerably reduced the mu-receptor affinity and selectivity. These data indicated that the Dmp-substitution of Phe(3), but not Phe(4), in EM(2) is favorable for improving mu-receptor specificity. Inversion of the chirality of the substituted Dmp residue resulted in marked decrease in the mu-receptor affinity. Replacement of Tyr(1) by Dmp yielded an analogue that exhibited only a limited decrease in mu-receptor affinity and GPI potency, despite the lack of a phenolic hydroxyl group at the N-terminal residue. In contrast, D-Dmp(1)- or Phe(1)-substitution of Tyr(1) resulted in a significant decrease in mu-receptor affinity and GPI potency. These results suggested that the Dmp residue can mimic Tyr(1), which is one of the critical structural elements of opioid peptides.

Dermorphin tetrapeptide analogues with 2',6'-dimethylphenylalanine (Dmp) substituted for aromatic amino acids have high mu opioid receptor binding and biological activities.

To investigate the value of the 2',6'-dimethylphenylalanine (Dmp) residue as an aromatic amino acid substitution, we prepared analogues of the mu opioid receptor-selective dermorphin tetrapeptide Tyr-D-Arg-Phe-betaAla-NH(2) (YRFB) in which Dmp or its D-isomer replaced Tyr(1) or Phe(3). Replacing Phe(3) with Dmp essentially tripled mu receptor affinity and the receptor's in vitro biological activities as determined with the guinea pig ileum (GPI) assay but did not change delta receptor affinity. Despite an inversion of the D configuration at this position, mu receptor affinity and selectivity remained comparable with those of the L-isomer. Replacing the N-terminal Tyr residue with Dmp produced a slightly improved mu receptor affinity and a potent GPI activity, even though the substituted compound lacks the side chain phenolic hydroxyl group at the N-terminal residue. Dual substitution of Dmp for Tyr(1) and Phe(3) produced significantly improved mu receptor affinity and selectivity compared with the singly substituted analogues. Subcutaneous injection of the two analogues, [Dmp(3)]YRFB and [Dmp(1)]YRFB, in mice produced potent analgesic activities that were greater than morphine in the formalin test. These lines of evidence suggest that the Dmp residue would be an effective aromatic amino acid surrogate for both Tyr and Phe in the design and development of novel opioid mimetics.

2',6'-dimethylphenylalanine (Dmp) can mimic the N-terminal Tyr in opioid peptides.

Substitution of 2',6'-dimethyltyrosine (Dmt) for the N-terminal Tyr in opioid peptides has recently been shown to be a promising tool for improving opioid receptor affinity and biological activity. We have also demonstrated that another unnatural amino acid, 2',6'-dimethylphenylalanine (Dmp), is not only an excellent substitute for Phe at position 3 but also can mimic the aromatic N-terminal Tyr residue in a micro opioid receptor-selective dermorphin analogue (YRFB: Tyr-D-Arg-Phe-betaAla-NH(2)). To further evaluate the value of Dmp in opioid peptides, we investigated Dmp(1)-substituted analogues of the delta receptor ligands, deltorphin II (DLT: Tyr-D-Ala-Phe-Glu-Val-Val-Gly-NH(2)) and enkephalin (ENK: Tyr-Gly-Gly-Phe-Leu). In the receptor binding assay, both [Dmp(1)]DLT and [Dmp(1)]ENK bound to the delta-receptor with high affinity and selectivity, and were nearly as effective as the parent peptides. The potency of the Dmp(1)-peptides on the MVD and GPI assays correlated well with the receptor binding affinity data. These results are in contrast to the tendency of corresponding Dmt(1)-analogues to have poor receptor selectivity. Taken together with the results with YRFB, we conclude that the Dmp(1)-peptide is superior to the corresponding Dmt(1)-peptide in its receptor selectivity. [Dmp(1)]DLT and [Dmp(1)]YRFB may serve as pharmacological tools for the studies of ligand recognition and opioid receptor signal transduction.