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Imaging plays an indispensable role in the diagnosis and treatment of breast cancer. Several new imaging tools are currently being developed for clinical use to improve diagnostic performance and tumor response evaluation. Abbreviated magnetic resonance imaging (MRI) allows shortening scanning time without penalizing diagnostic performances. Ultrafast dynamic contrast-enhanced MRI is a new approach that provides kinetic information and helps identify breast lesions developed in breast with marked background parenchymal enhancement. Intravoxel incoherent motion and kurtosis are parameters of diffusion-weighted imaging (DWI) that bring useful information in breast cancer. High-resolution DWI, allows morphological assessment of lesions without using a contrast agent. High-resolution breast positron emission tomography provides detailed metabolic information on small breast cancer. Photoacoustic imaging utilizes hemoglobin as an intrinsic contrast agent and depicts fine -branching structure of the tumor vessels related to cancer. It is important that experts in breast imaging understand the advantages and limitations of these new techniques in order to offer optimal imaging to each patient., (Copyright © 2022 Société française de radiologie. Published by Elsevier Masson SAS. All rights reserved.)

Pentazocine (PTZ) is a widely used drug for postoperative pain. It should be administered at appropriate dosing intervals not only because of its morphine-like side effects but also because frequent inappropriate dosing can lead to dependence. Although perioperative patients reportedly have nonnegligible effects on placebo drugs and postoperative wound healing, no pharmacokinetic (PK)/pharmacodynamic (PD) model has been established and simulated using real-world data for the perioperative period. This study aimed to perform PTZ modeling and simulation and to establish an indicator of the timing of drug efficacy evaluation in clinical practice. Participants were in-hospital orthopedic surgery patients who received 15 mg of PTZ within 48 h postoperatively. Pain severity was assessed using the numerical rating scale (NRS). A two-compartment model was selected for the population PK model and an indirect response model for the PK/PD model. Using these models, a virtual population of 1000 patients with Pain base NRS of 5 and 6 and body weights of 40, 80, and 120 kg were treated with single and multiple PTZ administrations (4, 8, and 24 h apart) of 15 mg. Simulation results indicate that its analgesic efficacy should be evaluated within 1 h after administration of 15 mg of PTZ. Additional doses should be considered every 8-12 h in postoperative patients with Pain base NRS of 5 weighing 40-80 kg. Simulation using the PK/PD model developed in this study may provide useful information for determining the analgesic effects and timing of the dosing interval after PTZ administration in perioperative patients.

Warfarin is a representative anticoagulant with large interindividual variability. The published kinetic-pharmacodynamic (K-PD) model allows the prediction of warfarin dose requirement in Swedish patients; however, its applicability in Japanese patients is not known. We evaluated the model's predictive performance in Japanese patients with various backgrounds and relationships using Bayesian parameter estimation and sampling times. A single-center retrospective observational study was conducted at Tokyo Women's Medical University, Medical Center East. The study population consisted of adult patients aged >20 years who commenced warfarin with a prothrombin time-international normalized ratio (PT-INR) from June 2015 to June 2019. The published K-PD model modified by Wright and Duffull was assessed using prediction-corrected visual predictive checks, focusing on clinical characteristics, including age, renal function, and individual prediction error. The external dataset included 232 patients who received an initial warfarin daily dose of 3.2 ± 1.28 mg with 2278 PT-INR points (median [range] follow-up period of 23 d [7-28]). Prediction-corrected visual predictive checks carried a propensity for underprediction. Additionally, age >60 years, body mass index ≤25 kg/m 2 , and estimated glomerular filtration rate ≤60 mL/min/1.73 m 2 had a pronounced tendency to underpredict PT-INR. However, Bayesian prediction using four prior observations reduced underprediction. To improve the prediction performance of these special populations, further studies are required to construct a model to predict warfarin dose requirements in Japanese patients.

Predicting pathogenic germline variants (PGVs) in breast cancer patients is important for selecting optimal therapeutics and implementing risk reduction strategies. However, PGV risk factors and the performance of prediction methods in the Japanese population remain unclear. We investigated clinicopathological risk factors using the Tyrer-Cuzick (TC) breast cancer risk evaluation tool to predict BRCA PGVs in unselected Japanese breast cancer patients (n = 1,995). Eleven breast cancer susceptibility genes were analyzed using target-capture sequencing in a previous study; the PGV prevalence in BRCA1, BRCA2, and PALB2 was 0.75%, 3.1%, and 0.45%, respectively. Significant associations were found between the presence of BRCA PGVs and early disease onset, number of familial cancer cases (up to third-degree relatives), triple-negative breast cancer patients under the age of 60, and ovarian cancer history (all P < .0001). In total, 816 patients (40.9%) satisfied the National Comprehensive Cancer Network (NCCN) guidelines for recommending multigene testing. The sensitivity and specificity of the NCCN criteria for discriminating PGV carriers from noncarriers were 71.3% and 60.7%, respectively. The TC model showed good discrimination for predicting BRCA PGVs (area under the curve, 0.75; 95% confidence interval, 0.69-0.81). Furthermore, use of the TC model with an optimized cutoff of TC score ≥0.16% in addition to the NCCN guidelines improved the predictive efficiency for high-risk groups (sensitivity, 77.2%; specificity, 54.8%; about 11 genes). Given the influence of ethnic differences on prediction, we consider that further studies are warranted to elucidate the role of environmental and genetic factors for realizing precise prediction., (© 2021 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

Background: The Medical Imaging Projection System (MIPS) projects indocyanine green (ICG) fluorescence images directly on the surgical field using a projection mapping technique. We conducted an observational study of sentinel lymph node (SLN) biopsy using the prototype MIPS; we found a high identification rate. However, the number of SLN-positive cases was small, and the sensitivity could not be evaluated. The aim of this study was to investigate the clinical usefulness of the MIPS assisted ICG fluorescence method using commercially available equipment., Methods: This was a retrospective observational study. Patients with primary breast cancer who underwent SLN biopsy using the MIPS at Kyoto University Hospital from April to December 2020 were included in the study. The primary endpoints were the identification rate of SLNs and detection of positive SLNs by the MIPS. The secondary endpoint was the number of SLNs excised using the MIPS per patient. We also conducted a questionnaire survey focused on the utility of the MIPS; it involved doctors with an experience in using the MIPS., Results: Seventy-nine patients (84 procedures) were included in the study. In 60 (71%) procedures, both the radioisotope (RI) method and MIPS were used. At least one SLN could be detected by the MIPS in all the procedures, with an identification rate of 100% (95% confidence interval 95.6-100%). A total of 19 (7%) positive SLNs were removed, which were identifiable by the MIPS. Among 57 patients in whom the MIPS and RI methods were used, there was no positive SLN only identified by the RI method. The results of the questionnaire survey showed that the MIPS enabled the operator and assistant to share the ICG fluorescence image in the surgical field and to communicate with each other easily., Conclusion: The current study demonstrated that the identification rate of SLNs using the MIPS was high, and the MIPS can be used for detecting positive SLNs. It was suggested that the MIPS will be useful in learning SLN biopsy procedures., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Nakamura, Takada, Imamura, Higami, Jiaxi, Fujino, Nakagawa, Inagaki, Matsumoto, Kawaguchi, Kawashima, Suzuki and Toi.)

Background: There is a diurnal variation in the blood pressure fluctuation of hypertension, and blood pressure fluctuation abnormality is considered to be an independent risk factor for organ damage including cardiovascular complications. In the current study, we tried to identify molecules responsible for blood pressure circadian rhythm formation under the control of the kidney biological clock in hypertension., Methods: DNA microarray analysis was performed in kidneys from 5-week-old spontaneously hypertensive rats (SHRs)/Izm, stroke-prone SHR rats (SHRSP)/Izm, and Wistar Kyoto (WKY)/Izm rats. To detect variation, mouse tubular epithelial cells (TCMK-1) were stimulated with dexamethasone. We performed immunostaining and western blot analysis in the renal medulla of kidney from 5-week-old WKY rats and SHRs., Results: We extracted 1,032 genes with E-box, a binding sequence for BMAL1 and CLOCK using a Gene Set Enrichment Analysis. In a microarray analysis, we identified 12 genes increased as more than 2-fold in the kidneys of SHRs and SHRSP in comparison to WKY rats. In a periodic regression analysis, phosphoribosyl pyrophosphate amidotransferase (Ppat) and fragile X mental retardation, autosomal homolog 1 (Fxr1) showed circadian rhythm. Immunocytochemistry revealed PPAT-positivity in nuclei and cytoplasm in the tubules, and FXR1-positivity in the cytoplasm of TCMK-1. In 5-week-old WKY rat and SHR kidneys, PPAT was localized in the nucleus and cytoplasm of the proximal and distal tubules, and FXR1 was localized to the cytoplasm of the proximal and distal tubules., Conclusions: PPAT and FXR1 are pivotal molecules in the control of blood pressure circadian rhythm by the kidney in hypertension., (© The Author(s) 2020. Published by Oxford University Press on behalf of American Journal of Hypertension, Ltd.)

Hemodialysis (HD) is a method used to remove biogenic substances or blood components that cause disease and some drugs used by patients to treat their diseases. Therefore, dosing schedule must be planned according to HD clearance (CL HD ) when medical treatment is provided to patients receiving HD. We aimed to clarify the physical properties (eg, octanol-water partition coefficient and molecular electronegativity) or pharmacokinetic parameters (eg, volume of distribution) of compounds affecting CL HD and to construct a mathematical model to predict CL HD . The analysis covered individual CL HD data for nine compounds from the literature. The molecular descriptors which are physical properties or pharmacokinetic parameters were calculated using the structural formula of each compound, and searched for factors related to CL HD among the calculated 148 molecular descriptors. Nonlinear mixed-effects model analysis with CL HD as objective variable and molecular descriptors as explanatory variable was conducted to examine the factor affecting CL HD and develop a model for predicting CL HD . The logarithm of the brain/blood partition coefficient was detected as a factor affecting CL HD . The predictive accuracy of CL HD using the constructed mathematical model with the logarithm of the brain/blood partition coefficient as explanatory variable was adequate., (© 2020 International Society for Apheresis, Japanese Society for Apheresis, and Japanese Society for Dialysis Therapy.)

Background: Solid papillary carcinoma (SPC) of the breast is a rare breast cancer that accounts for less than 1% of all breast cancers. The optimal clinical management of SPC remains controversial. Here, we report a case of invasive SPC with neuroendocrine differentiation in addition to review of the current literature., Case Presentation: A premenopausal 46-year-old female presented with a mass in her left breast that tended to increase in size over a 10-month period. Mammography and ultrasonography revealed a mass in the left upper-inner quadrant. The resulting images suggested a category 3 breast tumor according to the Breast Imaging Reporting and Data System (BI-RADS). A core needle biopsy (CNB) was performed, and the pathological findings showed a solid papillary pattern and atypical cells suggestive of noninvasive SPC. After a left partial mastectomy and sentinel lymph node biopsy (SLNB), the specimens were sent for histopathological analysis for further investigation. Postoperative pathological findings suggested invasive SPC. Whole-breast radiation therapy and adjuvant hormonal therapy were performed as postoperative treatments. Three years after surgery, multiple lung metastases were detected, and the patient was treated with a gonadotropin-releasing hormone agonist and an aromatase inhibitor. Five months later, multiple liver metastases and bone metastases appeared, and oral 5-fluorouracil was chosen for the subsequent treatment. The patient has been treated for 5 years to date, and she is continuing to take oral 5-fluorouracil and is alive without any further disease progression., Conclusions: We report a rare case of premenopausal invasive SPC with multiple metastases. Further study is needed to clarify the molecular characteristics and clinical behaviors of SPC and to explore the optimal treatment strategy.

Background: Photoacoustic tomography (PAT) is a noninvasive vascular imaging modality that uses near-infrared pulse laser beams and ultrasound (US) to visualize vessels. We previously demonstrated the utility of PAT for visualizing anterolateral thigh (ALT) perforators in a clinical study of 10 thighs in 5 healthy adults. Evaluation of the correlation between PAT and US findings showed that PAT had comparable diagnostic potential but was superior in visualizing subcutaneous microvessels; however, there was no comparison with intraoperative findings. In this study, we used a newly developed technique to transfer a PAT image to a body-attachable transparent sheet to compare PAT and intraoperative findings., Methods: Eight patients were recruited in this prospective study. Patient age ranged from 32 to 79 years (average 60). Seven ALT flaps were applied in head and neck reconstruction. One flap was elevated in chest wall reconstruction. Each PAT scan of an 18 cm × 13.5 cm region took approximately 5 min. Acquired data were processed three-dimensionally using a novel imaging software program. Perforator vessel data from PAT imaging were traced and corrected for projection onto medical film sheets. The correlation between the perforator stem portions predicted by PAT and the intraoperative findings at the level of the fascia-penetrating points was evaluated, and distal branching patterns were analyzed., Results: PAT imaging showed 16 perforators in 8 thighs. Intraoperative surgical findings revealed that all the perforator penetrating points at the deep fascia level matched the PAT findings within 10 mm. None of the eight ALT flaps demonstrated postoperative complications. The perforator complexes were classified as type I in three cases (19%), type II in eight cases (50%), and type III in five cases (31%)., Conclusions: PAT imaging matched the intraoperative findings within 10 mm. Preoperative vascular evaluation allows for the creation of a vascular map for facilitating ALT flap surgeries., (© 2019 Wiley Periodicals, Inc.)

Mammary ductal dysplasia is a phenotype observed in precancerous lesions and early-stage breast cancer. However, the mechanism of dysplasia formation remains elusive. Here we show, by establishing a novel dysplasia model system, that estrogen, a female hormone, has the potential to cause mammary ductal dysplasia. We injected estradiol (E2), the most active form of estrogen, daily into scid mice with a defect in non-homologous end joining repair and observed dysplasia formation with cell proliferation at day 30. The protooncogene Myc is a downstream target of estrogen signaling, and we found that its expression is augmented in mammary epithelial cells in this dysplasia model. Treatment with a Myc inhibitor reduced E2-induced dysplasia formation. Moreover, we found that isoflavones inhibited E2-induced dysplasia formation. Our dysplasia model system provides insights into the mechanistic understanding of breast tumorigenesis and the development of breast cancer prevention., Competing Interests: Declaration of Interests J.I. was an employee of Kyoto University's Sponsored Research Program funded by Taiho Pharmaceutical. R.T., H.S., M. Tsuda, S.M., Y.M., T.I., F.S., S.T., and M. Toi have no conflict of interest. The funding source had no role in the study design, experiment, analysis, interpretation, or writing the manuscript., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

To improve the efficiency of drug-discovery research on pyrrole-imidazole polyamides (PIs), a more rapid method for quantitative and qualitative measurement of PI in rat plasma samples was developed here using ultra-fast liquid chromatography-ultraviolet spectrometry (UFLC-UV) in order to shorten the measurement time. A measurement method of PIs by HPLC developed until now takes 45 min for one sample measurement. This method was inefficient to investigate extraction conditions from biological samples and measurement of animal experimental samples. In the developed method of this study, PI and phenacetin (internal standard, IS) were separated with an ACQUITY UPLC HSS T3 (1.8 µm, 2.1 × 50 mm; Nihon Waters K.K., Japan) column using a mobile phase of 0.1% acetic acid (mobile phase A) and acetonitrile (mobile phase B) at a flow rate of 0.3 mL/min with a linear gradient. The detection wavelength was 310 nm. The calibration curve was linear in the range of 0.225-4.5 µg/mL (correlation coefficients ≥0.9995, n = 5). The intra- and inter-day accuracies were in the range of -6.04 to 12.2%, and the precision was less than 2.99%. The measurement time of this method (7 min per injection) was markedly shortened to about one-sixth of the previous measurement time (45 min per injection). This is the first report describing the quantitative and qualitative measurement of PI in plasma using UFLC-UV. The present method will be very useful for the drug-discovery research of PIs.

Background :  A breast-specific photoacoustic imaging (PAI) system prototype equipped with a hemispherical detector array (HDA) has been reported as a promising system configuration for providing high morphological reproducibility for vascular structures in living bodies. Methods :  To image the vasculature of human limbs, a newly designed PAI system prototype (PAI-05) with an HDA with a higher density sensor arrangement was developed. The basic device configuration mimicked that of a previously reported breast-specific PAI system. A new imaging table and a holding tray for imaging a subject's limb were adopted. Results :  The device's performance was verified using a phantom. Contrast of 8.5 was obtained at a depth of 2 cm, and the viewing angle reached up to 70 degrees, showing sufficient performance for limb imaging. An arbitrary wavelength was set, and a reasonable PA signal intensity dependent on the wavelength was obtained. To prove the concept of imaging human limbs, various parts of the subject were scanned. High-quality still images of a living human with a wider size than that previously reported were obtained by scanning within the horizontal plane and averaging the images. The maximum field of view (FOV) was 270 mm × 180 mm. Even in movie mode, one-shot 3D volumetric data were obtained in an FOV range of 20 mm in diameter, which is larger than values in previous reports. By continuously acquiring these images, we were able to produce motion pictures. Conclusion :  We developed a PAI prototype system equipped with an HDA suitable for imaging limbs. As a result, the subject could be scanned over a wide range while in a more comfortable position, and high-quality still images and motion pictures could be obtained., Competing Interests: Competing interests: Ken-ichi Nagae, Yoshiaki Sudo, Yasufumi Asao and Takayuki Yagi are employees of Canon Inc., Japan. Katsumi Ohira and Yuusuke Tanaka are employees of Japan Probe, Co. Ltd., Japan. Naoyuki Murayama is an employee of Hitachi, Ltd., Japan. YA and TY temporarily transferred to the Japan Science and Technology Agency. Canon Inc. designed and invented the photoacoustic imaging system used in this study. The other authors have no conflict of interest.

Background: Inability to visualize indocyanine green fluorescence images in the surgical field limits the application of current near-infrared fluorescence imaging (NIR) systems for real-time navigation during sentinel lymph node (SLN) biopsy in breast cancer patients. The aim of this study was to evaluate the usefulness of the Medical Imaging Projection System (MIPS), which uses active projection mapping, for SLN biopsy., Methods: A total of 56 patients (59 procedures) underwent SLN biopsy using the MIPS between March 2016 and November 2017. After SLN biopsy using the MIPS, residual SLNs were removed using a conventional NIR camera and/or radioisotope method. The primary endpoint of this study was identification rate of SLNs using the MIPS., Results: In all procedures, at least one SLN was detected by the MIPS, giving an SLN identification rate of 100% [95% confidence interval (CI) 94-100%]. SLN biopsy was successfully performed without operating lights in all procedures. In total, 3 positive SLNs were excised using MIPS, but were not included in the additional SLNs excised by other methods. The median number of SLNs excised using the MIPS was 3 (range 1-7). Of procedures performed after preoperative systemic therapy, the median number of SLNs excised using the MIPS was 3 (range 2-6)., Conclusions: The MIPS is effective in detecting SLNs in patients with breast cancer, providing continuous and accurate projection of fluorescence signals in the surgical field, without need for operating lights, and could be useful in real-time navigation surgery for SLN biopsy.

This study aimed to identify the characteristics of the vascular network in the superficial subcutaneous layer of the breast and to analyze differences between breasts with cancer and contralateral unaffected breasts using vessel branching points (VBPs) detected by three-dimensional photoacoustic imaging with a hemispherical detector array. In 22 patients with unilateral breast cancer, the average VBP counts to a depth of 7 mm below the skin surface were significantly greater in breasts with cancer than in the contralateral unaffected breasts (p < 0.01). The ratio of the VBP count in the breasts with cancer to that in the contralateral breasts was significantly increased in patients with a high histologic grade (p = 0.03), those with estrogen receptor-negative disease (p < 0.01), and those with highly proliferative disease (p < 0.01). These preliminary findings indicate that a higher number of VBPs in the superficial subcutaneous layer of the breast might be a biomarker for primary breast cancer.

The distal branching pattern of perforators is associated with thin anterolateral thigh flap failure. The purpose of this study was to investigate the feasibility of using photoacoustic tomography as a diagnostic imaging modality to identify anterolateral thigh perforators and their branching patterns in the subcutaneous layer. Ten thighs in five healthy men were studied. The anterolateral aspect of the midthigh was examined using photoacoustic tomography. The correlation between photoacoustic tomography and ultrasound findings was evaluated. To determine the detectability of photoacoustic tomography by depth, the depth of vessels in the stem portion was compared to the depth of the deep fascia measured by ultrasound. Branching patterns of vessels in the adipose and suprafascial layers were evaluated by three-dimensional observation. A total of 18 perforators were visualized by photoacoustic tomography. Photoacoustic tomography and ultrasound had comparable diagnostic potential for the detection of perforators. Photoacoustic tomography visualized microvessels in the subcutaneous layer, especially those in oblique or horizontal orientations. The estimated mean depth of visualized vessels was 9 mm; the maximum depth was 13 mm. There was a strong correlation between the depth of visualized vessels in the stem portion and the depth of the deep fascia. Three-dimensional observation of photoacoustic tomographic images showed the branching morphology of perforators. This study showed the applicability of photoacoustic tomography to identification of the branching patterns of anterolateral thigh perforators in vivo, although limited visualization of subfascial vessels is a technical issue. The authors believe that photoacoustic tomography has the potential to be a new imaging modality for thin anterolateral thigh flap surgery., Clinical Question/level of Evidence: Diagnostic, IV.

Sal-like 4 (SALL4) is a transcription factor that enhances proliferation and migration in breast cancer cells. SALL4 expression therefore has the potential to promote cancer malignancy. However, the regulatory mechanisms involved in SALL4 protein expression have not been thoroughly elucidated. In this study, we observed that treating MCF-7 and SUM159 breast cancer cell lines with a proteasome inhibitor increases SALL4 protein levels, suggesting that SALL4 is degraded by the ubiquitin-proteasome system. Using immunoprecipitation to uncover SALL4-binding proteins, we identified an E3 ubiquitin-protein ligase, tripartite motif-containing 21 (TRIM21). Using an EGFP reporter probe of the major SALL4 isoform SALL4B, we observed that shRNA-mediated knockdown of TRIM21 increases cellular SALL4B levels. Immunostaining experiments revealed that TRIM21 localizes to the nucleus, and a K64R substitution in the nuclear localization motif in SALL4B increased SALL4B levels in the cytoplasm. These results suggested that TRIM21 is involved in nuclear SALL4 degradation. To identify the amino acid residue that is targeted by TRIM21, we fragmented the SALL4B sequence, fused it to EGFP, and identified Lys-190 in SALL4B as TRIM21's target residue. Amino acid sequence alignments of SALL family members indicated that the region around SALL4 Lys-190 is conserved in both SALL1 and SALL3. Because SALL1 and SALL4 have similar functions, we constructed a SALL1-EGFP probe and found that the TRIM21 knockdown increases SALL1 levels, indicating that TRIM21 degrades both SALL1 and SALL4. Our findings extend our understanding of SALL4 and SALL1 regulation and may contribute to the development of SALL4-targeting therapies., (© 2018 by The American Society for Biochemistry and Molecular Biology, Inc.)

Understanding the mechanism by which cancer cells enhance stemness facilitates cancer therapies. Here, we revealed that a stem cell transcription factor, SALL4, functions to enhance stemness in basal-like breast cancer cells. We used shRNA-mediated knockdown and gene overexpression systems to analyze gene functions. To evaluate stemness, we performed a sphere formation assay. In SALL4 knockdown cells, the sphere formation ability was reduced, indicating that SALL4 enhances stemness. CD44 is a membrane protein and is known as a stemness factor in cancer. CD44 splicing variants are involved in cancer stemness. We discovered that SALL4 modulates CD44 alternative splicing through the upregulation of KHDRBS3, a splicing factor for CD44. We cloned the KHDRBS3-regulated CD44 splicing isoform (CD44v), which lacks exons 8 and 9. CD44v overexpression prevented a reduction in the sphere formation ability by KHDRBS3 knockdown, indicating that CD44v is positively involved in cancer stemness. In addition, CD44v enhanced anoikis resistance under the control of the SALL4 - KHDRBS3 network. Basal-like breast cancer is an aggressive subtype among breast cancers, and there is no effective therapy so far. Our findings provide molecular targets for basal-like breast cancer therapy. In the future, this study may contribute to the establishment of drugs targeting cancer stemness., (© 2018 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

A high-performance liquid chromatography-ultraviolet spectrophotometry (HPLC-UV) method for the determination of meloxicam (MEL) and meloxicam metabolites (5'-hydroxy meloxicam (5-HMEL) and 5'-carboxy meloxicam (5-CMEL)) has been developed. After extraction of MEL, 5-HMEL, and 5-CMEL from rat plasma using Oasis HLB cartridges, the extracts were separated with a Luna C18 (2) 100 A column (5 µm, 4.6×150 mm, Phenomenex) using a mobile phase of 50 mM phosphate buffer (pH 2.15, solvent A) and acetonitrile (solvent B) at a flow rate of 0.8 mL/min in a linear gradient. The detection wavelength was 360 nm, and the internal standard (IS) was piroxicam. Each calibration curve was linear in the range of 40 to 1000 ng/mL (r 2 >0.999). The extraction rates of MEL, 5-HMEL, and 5-CMEL were greater than 86.9%. The intra- and inter-day accuracies were in the range of 95.0 to 119.0%, and the precision was 0.2 to 17.0%. To the best of our knowledge, this is the first report of the quantitative and qualitative measurement of meloxicam and each metabolite using an HPLC-UV method.

A method for evaluating hydroxyl radical (·OH) scavenging activities using sequential injection analysis (SIA) with chemiluminescence (CL) detection was developed. In this system, CL was produced by the reaction of luminol with ·OH generated from the Fenton reaction. The scavenging activity was expressed as a diminution rate of the CL due to the scavenging of ·OH by a sample. The SIA system allows the automation of a series of experimental procedures including Fenton's reaction, scavenging of ·OH, and luminol CL reaction. The evaluation of scavenging activities in one sample (n = 3) was completed within 3.0 min. Relative standard deviations (n = 3) of scavenging activity with 700 μM L-ascorbic acid were 2.6% (intraday) and 3.7% (interday). The SIA-CL system was applied to measure ·OH scavenging activities of several antioxidants and pharmaceuticals.

A simple and rapid ultra-fast liquid chromatography-ultraviolet spectrophotometry (UFLC-UV) method combined with modified 2-nitrophenylhydrazine (2-NPH) derivatization was developed for determining phytanic acid (Phy) in rat serum. Serum Phy and heptadecanoic acid (the internal standard) were derivatized by 2-NPH at ambient temperature for 20 min and extracted in n-hexane. After extracting derivatized Phy (D-Phy) and derivatized IS from the reaction mixture, the extracts were separated with a YMC-Pack C8 column (150 × 3.0 mm i.d., S-3 μm) using an isocratic mobile phase comprised of acetonitrile:H 2 O (90:10; pH 4.4) at 0.5 mL/min. The detection wavelength was 228 nm. Linearity was observed over 1 - 20 μg/mL (r = 0.9997). The intra- and inter-day reproducibilities of D-Phy measurements were ≤13.0%. To our knowledge, this is the first report of the quantitative and qualitative measurement of serum Phy using 2-NPH derivatization and UFLC-UV. This method can be performed rapidly under mild conditions.

Objective: Azithromycin (AZM) is widely used as a first-line treatment option for children with mycoplasma pneumonia. Although pharmacists perform medication counseling in the pediatric ward, children often experience vomiting as a result of oral AZM administration. Drugs that are administered rectally are generally considered to enter the circulation system without passing through the liver first. The aim of our study was to prepare an AZM suppository and investigate the pharmaceutical properties and well as pharmacokinetics of the rectal administration route in humans., Materials and Methods: Five healthy volunteers were enrolled in the study. All subjects provided written informed consent before participating in the study. Subjects were randomly assigned to either oral administration of oral AZM 500-mg tablet or rectal administration of 125-mg, 250-mg, or 500-mg AZM suppository. Blood samples for preparation of serum were collected predose as well as at 1, 2, 3, 4, 6, 12, and 24 hours following the first rectal dose. Serum concentrations of AZM were determined by high-performance liquid chromatography (HPLC) with electrochemical detection., Results and Conclusion: The bioavailability of the AZM suppository through rectal administration was 20.3% compared to oral administration. We hypothesize that the surface area where AZM is absorbed also affects the absorption by rectal administration. Although further investigation is necessary to improve the absorption of AZM by the rectum and to ensure safety in children, the AZM suppository may be an effective preparation in cases where oral administration is not tolerated.

Three-dimensional (3D) cell culture is a powerful tool to study cell growth under 3D condition. To perform a simple test for anti-cancer drugs in 3D culture, visualization of non-proliferated cells is required. We propose a fluorescent imaging-based assay to analyze cancer cell proliferation in 3D culture. We used a pulse-labeling technique with a photoconvertible fluorescent protein Kaede to identify non-proliferated cells. This assay allows us to observe change in cell proliferation in 3D culture by simple imaging. Using this assay, we obtained the data of the effects of anti-cancer drugs, 5-fluorouracil and PD0332991 in a breast cancer cell line, MCF-7.

Pyrrole-imidazole (PI) polyamide is a novel gene regulating agent that competitively inhibits transcription factor binding to the promoter of the specific target gene. Liver fibrosis is an integral stage in the development of chronic liver disease, and transforming growth factor β (TGFβ) is known to play a central role in the progression of this entity. The aim of this study was to evaluate the effect of PI polyamide targeting TGFβ1 on rat liver fibrosis. PI polyamide was designed to inhibit activator protein 1 (AP-1) transcription factor binding to the TGFβ1 gene promoter. The effect of PI polyamide on hepatic stellate cells was evaluated by real time polymerase chain reaction (PCR) in RI-T cells. To determine the effect of PI polyamide in vivo, PI polyamide was intravenously administered at a dose of 3 mg/kg/week in dimethylnitrosamine (DMN)-induced rat model of liver fibrosis. Treatment of RI-T cells with 1.0 µM PI polyamide targeting TGFβ1 significantly inhibited TGFβ1 mRNA expression. Azan staining showed that DMN treatment significantly increased areas of fibrous materials compared with controls. PI polyamide targeting TGFβ1 significantly decreased the fibrous area compared with DMN group. mRNA expression levels of α-smooth muscle actin and matrix metalloproteinase-2 were significantly increased in DMN-treated group compared with control. Treatment with TGFβ1 PI polyamide significantly decreased mRNA expression of these genes compared with DMN group. The novel gene regulator PI polyamide targeting TGFβ1 may be a feasible therapeutic agent for the treatment of chronic liver disease.

Background: Chronic kidney disease (CKD) is associated with a high risk of cardiovascular disease complications. Therefore, medical institutions conduct educational hospitalization for early treatment and education of CKD patients. However, patients who have been discharged after achieving educational targets can end up showing poor symptoms at home. There also have been several cases of rehospitalization or disease aggravation. In this study, we analyzed rehospitalized patients who were discharged from the hospital after CKD educational hospitalization and investigated the purpose of analyzing rehospitalization factors., Materials and Methods: This was an observational case-control study conducted at Yokosuka Kyousai Hospital. We performed univariate analysis using patient background features and laboratory data between a rehospitalization group and a no-rehospitalization group. Next, we performed multiple logistic regression analysis using the results of the univariate analysis., Results: From the results of this study, we identified independent risk factors, such as serum albumin level, heart-failure complications, and estimated glomerular filtration rate (eGFR). Moreover, the serum Alb level was identified as the most important risk factor for rehospitalization. Therefore, we considered that it is important to live a life that makes it possible to maintain CKD stage G3b for a long time after discharge, because the cutoff level of eGFR is 31 mL/minute/1.73 m(2)., Conclusion: We believe that it is important to educate patients, their families, and medical staff on the importance of early detection and treatment, and we consider that this approach is important to inclusively protect the kidney.

Hyperuricemia and hyperlipidemia have attracted attention as progression factors for chronic kidney disease (CKD). In the drug treatment of hyperuricemia and hyperlipidemia complications, Atorvastatin (ATV), which inhibits urinary protein, increases glomerular filtration rate (GFR) and has renal protective effects, and Rosuvastatin (ROS) were found be suitable because they promote serum uric acid (SUA) excretion. However, these drugs were administered at very high doses in previous studies. In this study, we have investigated the effects of ATV or ROS on renal protective effects and their SUA levels before and three months after each drug administration in CKD patients. We retrospectively investigated outpatients presenting with CKD (stages 3) on the basis of their electronic medical records as subjects. Estimated GFR (eGFR) was significantly increased after ATV administration, whereas no change in eGFR was observed following ROS administration. Furthermore, SUA levels significantly decreased after ATV administration, whereas no changes were observed following ROS administration. Therefore, it may be not necessary to administer drugs that lower the SUA levels to patients presenting with hyperuricemia and hyperlipidemia complications associated with moderate renal failure, such as patients with at least stage 3 CKD. We consider that, by selecting ATV, the renal protective effects and SUA-lowering effect would be sufficient.

In cell cultures, the dispersed phenotype is indicative of the migratory ability. Here we characterized Sal-like 4 (SALL4) as a dispersion factor in basal-like breast cancer. Our shRNA-mediated SALL4 knockdown system and SALL4 overexpression system revealed that SALL4 suppresses the expression of adhesion gene CDH1, and positively regulates the CDH1 suppressor ZEB1. Cell behavior analyses showed that SALL4 suppresses intercellular adhesion and maintains cell motility after cell-cell interaction and cell division, which results in the dispersed phenotype. Our findings indicate that SALL4 functions to suppress CDH1 expression and to maintain cell dispersion in basal-like breast cancer., (Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

Fcγ receptors I and III are thought to be involved in the development of lupus nephritis. Expression of Fc receptor common gamma chain (FcRγ) is necessary for the stable expression of Fcγ receptors I and III. The aim of this study was to develop a novel agent for the treatment of immune complex related renal disease using a gene regulator, pyrrole(Py)-imidazole(Im) (PI) polyamide, targeting the mouse FcRγ gene promoter. Two PI polyamides targeting FcRγ promoters were designed and synthesized. The effect of the PI polyamides on FcRγ mRNA expression was evaluated in J774.A cells by real-time polymerase chain reaction (PCR), and CD16/32 protein expression was determined by immunocytochemical analysis and flow cytometry. The effects of these polyamides on FcRγ gene expression and CD16/32 protein expression were evaluated in mouse peripheral blood mononuclear cells (PBMCs). One milligram per kilogram body weight of PI polyamide was injected via the tail vein every 2 d for 1 week and PBMCs were collected and analyzed. PI polyamide showed a specific binding to the target DNA in a gel mobility shift assay. Treatment of J774.A cells with 1.0 µM PI polyamide 1 significantly reduced FcRγ mRNA expression and CD16/32 surface protein expression in J774.A cells. Similarly, PI polyamide significantly decreased expression of FcRγ mRNA and CD16/32 in the PBMCs of C57B6 mice. PI polyamide designed to bind the FcRγ promoter decreased FcRγ gene and CD16/32 protein expression. PI polyamide targeting the FcRγ gene may be a novel gene regulator for the prevention of lupus nephritis or other immune complex-related disease.

Introduction: The aim of this study was to investigate the association between the variation in expression profile of clock genes and obesity using peripheral blood mononuclear (PMN) cells., Material and Methods: The subjects comprised 10 obese patients and 10 healthy volunteers. Blood was collected at different time-points during the day and levels of blood sugar, IRI, adiponectin and leptin were determined. Peripheral blood mononuclear cells were sampled, and expression levels of brain and muscle Arnt-like protein-1 (BMAL1), Period (PER)1, PER2, Cryptochrome (CRY)1, CRY2, and REV-ERBα mRNA were quantified., Results: During the day, the expression levels of BMAL1, CRY1, CRY2 and PER2 genes in PMN cells of the obese group were all significantly higher compared to those in the non-obese group. In addition, expression of BMAL1, CRY1, CRY2 and PER2 genes in PMN cells increased between 12:00 and 21:00 in the obese group. In PMN cells of both groups, PER1 gene expression showed a bimodal pattern, with high expression at 9:00 and 18:00., Conclusions: Differences were observed in the expression profile variation of clock genes between the obese and non-obese groups. This study reveals the differences in clock gene expression profiles between obese and non-obese subjects, with evidence for two distinct chronotypes, and suggests a contribution of these chronotypes to fat accumulation in humans.

Germinated barley foodstuff contains prebiotics which are reported to have anti-cancerous effects in colorectal cancer model, but the detailed mechanism remains unclear. Recent studies revealed that the role of microbiota was strongly related to the regulation of incidence and progression of colorectal cancer. The aim of this study was to examine the anti-neoplastic mechanism by prebiotics. Azoxymethane treated F344 rats were used as the sporadic cancerous model. After azoxymethane injection, either a control or germinated barley foodstuff diet was administered to the rats for another 5 weeks, and the number of abberant crypt foci, toll like receptor 4, Kirsten rat sarcoma viral oncogene homolog, adenomatous polyposis coli tumor suppressor gene and cyclooxygenase 2 mRNA expression of colonic mucosa and cecal short chain fatty acids were examined. The germinated barley food stuff significantly attenuated the number of abberant crypt focis and the expression of toll like receptor 4 and cyclooxygenase 2 mRNA, compared to the control group. In addition, the cecal butyrate production in the germinated barley foodstuff group was significantly higher than that in the control. In conclusion, this prebiotic treatment for colorectal cancer may be useful without causing the adverse effects seen in either anti-cancer drugs or anti-inflammatory drugs.

Background/aims: Gastric mucus protects the gastric mucosa. Plaunotol, a gastroprotective agent, has been shown to increase mucus production in animal models. However, it is unclear whether plaunotol benefits human gastric mucus secretion., Methodology: Twenty-five patients with atrophic gastritis were studied. All patients underwent gastroendoscopy and gastric juice was collected before and after plaunotol treatment for 3 months. Gastric juice mucin was examined by gel filtration as well as anion-exchange chromatography. The identification of each fraction was examined by enzyme-linked immunosorbent assay (ELISA) with the use of HGM75 and HIK1083, antibodies against mucin from surface mucus cells and from gastric glandular mucus cells, respectively., Results: Plaunotol significantly increased the total gastric juice volume (7.8mL before vs. 10.7mL, after administration; p=0.03). By anion exchange chromatography, we detected three mucin fractions (Fr I-III). Fr I strongly reacted with HGM75 but did not react with HIK1083. The other fractions (Fr II, III) reacted with HIK1083 but weakly reacted with HGM75. After administration of plaunotol, a significant increase in Fr III (acidic mucin) was observed (p=0.02)., Conclusions: Long-term administration of plaunotol changes the composition of gastric juice mucin, including a significant increase in the proportion of acidic mucin fraction.

Pyrrole-imidazole (PI) polyamide can bind to specific sequences in the minor groove of double-helical DNA and inhibit transcription of the genes. We designed and synthesized a PI polyamide to target the human connective tissue growth factor (hCTGF) promoter region adjacent to the Smads binding site. Among coupling activators that yield PI polyamides, 1-[bis(dimethylamino)methylene]-5-chloro-1H-benzotriazolium 3-oxide hexafluorophosphate (HCTU) was most effective in total yields of PI polyamides. A gel shift assay showed that a PI polyamide designed specifically for hCTGF (PI polyamide to hCTGF) bound the appropriate double-stranded oligonucleotide. A fluorescein isothiocyanate (FITC)-conjugated PI polyamide to CTGF permeated cell membranes and accumulated in the nuclei of cultured human mesangial cells (HMCs) and remained there for 48 h. The PI polyamide to hCTGF significantly decreased phorbol 12-myristate acetate (PMA)- or transforming growth factor-β1 (TGF-β1)-stimulated luciferase activity of the hCTGF promoter in cultured HMCs. The PI polyamide to hCTGF significantly decreased PMA- or TGF-β1-stimulated expression of hCTGF mRNA in a dose-dependent manner. The PI polyamide to hCTGF significantly decreased PMA- or TGF-β1-stimulated levels of hCTGF protein in HMCs. These results indicate that the developed synthetic PI polyamide to hCTGF could be a novel gene silencer for fibrotic diseases.

Pharmacokinetic/pharmacodynamic (PK/PD) modeling and simulation enable the prediction of the effect of a medication in various situations in clinical practice. The aims of this study were to predict the relationships between the effect of rosuvastatin and various factors such as poor compliance, and morning and evening dosages, as well as the change in the pharmacokinetics of rosuvastatin. We characterized the PK/PD model of plasma mevalonic acid (MVA) profiles after rosuvastatin administration and simulated the plasma MVA concentration in various dosage regimens. The plasma rosuvastatin and MVA concentrations reported by Martin et al. were used as the source of PK/PD modeling data. For each simulation, a summary parameter, the area under the plasma MVA concentration-time curves for 24 h in the steady state (AUEC(24)), was used to characterize the time course of each endpoint. To estimate the influence of PK parameters on rosuvastatin effects, the AUEC(24) reduction ratio of baseline levels was simulated from the 0.33-3.0-fold value of each PK parameter estimate. The AUEC(24) reduction ratio was 7.7% lower after morning administration than after evening administration. The changes in the PK parameters more prominently affected the AUEC(24) reduction ratio after morning administration than after evening administration. The simulated plasma MVA concentrations almost reached their baseline levels in the case of patients who forgot to take rosuvastatin. These results suggest that the parameters can be used to determine the effective rosuvastatin dosage regimen.

The use of urinary and/or biliary excretion data was considered as an alternative approach if the bioanalytical method lacked the appropriate sensitivity to adequately characterize the serum or plasma concentration-time profile. This approach is used for the analysis of plasma concentration-time profile under the lower limit of quantification (LLOQ) of various analytical instruments. The objective of this study was to develop a pharmacokinetic (PK) model that describes the plasma concentration-time profiles under LLOQ of HPLC using urinary and biliary excretion data. As model compounds, pyrrole (Py)-imidazole (Im) polyamides 1035 (MW, 1035.12) and 1666 (MW, 1665.78) were used. The cumulative urinary excretions of Py-Im polyamides 1035 and 1666 were 72.4+/-11.6 and 4.8+/-0.5% of the administered dose, respectively. The cumulative biliary excretion of Py-Im polyamide 1035 was 4.3+/-0.4% of the administered dose, and Py-Im polyamide 1666 was not detected. The plasma concentration-time profiles of Py-Im polyamide 1035 were adequately described using linear and non-linear output compartments. The developed PK model could be used to describe the plasma concentration profiles using the linear output compartment interpreted as the urine compartment and the non-linear output compartment interpreted as the bile compartment. This PK model will be able to provide a more accurate prediction of the plasma concentration profiles under LLOQ.

Background/aims: Helicobacter pylori (H. pylori) eradication therapy increases acid secretion and promotes the development of gastroesophageal reflux disease (GERD) and reflux esophagitis (RE). Rebound acid hypersecretion develops after the use of proton pump inhibitors (PPI). We examined the clinical necessity of acid inhibitors to prevent GERD or RE caused by PPI rebound phenomenon and prior H. pylori eradication therapy., Methodology: We enrolled 39 patients who underwent successful H. pylori eradication therapy prior to endoscopic mucosal resection of gastric cancer. After 8-week rabeprazole treatment for iatrogenic ulcer, they were randomly divided into two groups (who took nizatidine (group N) and sofalcone (group S)), and took each for 16 weeks, we compared RE/GERD symptoms with the baseline by endoscopy and QUEST score., Results: All patients had corpus atrophy in which there was no difference between the two groups. Only 1 patient in group S (5.9%) developed symptomatic GERD, and 1 patient in group N (4.5%) developed RE., Conclusions: In severe atrophic gastritis patients, there is little clinical necessity of acid inhibitors to prevent GERD/RE caused by PPI rebound and prior H. pylori eradication therapy.

In this study, we aimed to determine an index of anti-MRSA drugs for long-term treatment. We examined adult patients to whom the anti-MRSA drugs arbekacin sulfate (ABK), vancomycin hydrochloride (VCM), and teicoplanine (TEIC) had been administered in the St. Marianna University School of Medicine, Yokohama City Seibu Hospital, for 1, year. The number of patients treated for>or==14 days was 22 (31%) among 71 patients. Immunosuppressive agent positivity (p=0.07), albumin (ALB) level of or==10 mg/dl (p=0.01),%STAB >or==15% (p=0.11), and the period until the blood drug level is measured (p=0.06) were analyzed with respect to the differences between both groups by univariate analysis. An ALB level of

Aims: Although the use of drug-eluting stents (DESs) has been shown to limit neointima hyperplasia, currently available DESs may adversely affect re-endothelialization. To evaluate whether a novel gene silencer pyrrole-imidazole (PI) polyamide targeting transforming growth factor (TGF)-beta1 is a candidate agent for the DESs, we examined the effects of PI polyamide targeting the TGF-beta1 promoter on neointimal formation in rat carotid artery after balloon injury., Methods and Results: PI polyamide was designed to span the boundary of the AP-1 binding site of the TGF-beta1 promoter. After inducing balloon injury to arteries, incubation with PI polyamide was carried out for 10 min. Neointimal thickening and re-endothelialization were evaluated at 21 days after injury. Fluoresceinisothiocyanate-labelled PI polyamide was distributed into most of the nuclei in the injured artery without any delivery reagents. PI polyamide (100 microg) significantly inhibited neointimal thickening at 21 days after injury by 57%. PI polyamide targeting TGF-beta1 significantly decreased the expression of TGF-beta1 mRNA and protein in the artery at 3 days after injury and also suppressed the expression of connective tissue growth factor (CTGF), fibronectin, collagen type 1, and lectin-like ox-LDL receptor-1 mRNAs. A morphometric analysis showed that PI polyamide targeting TGF-beta1 accelerated re-endothelialization in the injured artery., Conclusion: These findings suggest that the synthetic PI polyamide targeting the TGF-beta1 promoter may have the potential to suppress neointimal hyperplasia after arterial injury by the down-regulation of TGF-beta1 and CTGF and the reduction of the extracellular matrix. As a result, PI polyamide targeting TGF-beta1 may therefore be a potentially effective agent for the treatment of in-stent restenosis, as a candidate agent for the next-generation DES.

The pharmacokinetics of pyrrole (Py)-imidazole (Im) polyamides was studied in rats after the intravenous administration of these compounds. Py-Im polyamide (A) was composed of Ac-ImPyPy-ImPyPy-beta-Dp (beta: beta-alanine, Dp: N,N-dimethylaminopropylamide). Py-Im polyamide (B) was composed of Ac-PyIm-beta-ImIm-PyPy-beta-PyPy-beta-Dp. Py-Im polyamide (C) was composed of Ac-PyPy-beta-PyImPy-PyPyPy-beta-ImPy-beta-Dp. The molecular weight of Py-Im polyamide (A) was 1035.12, that of Py-Im polyamide (B) was 1422.51 and that of Py-Im polyamide (C) was 1665.78. After the intravenous injection of Py-Im polyamide (A) at 1.3, 2.0, 7.5 and 15.0 mg/kg, Py-Im polyamides (B) and (C) at 1.0, 2.0, 3.0 and 5.0 mg/kg, the average systemic clearance and the volume of distribution at the steady state obtained by a non-compartmental method were in the ranges of 4.6-6.4 ml/min/kg and 244-412 ml/kg, 8.9-10.3 ml/min/kg and 1990-4567 ml/kg, and 7.3-11.9 ml/min/kg and 407-667 ml/kg, respectively. Dose linearity of Py-Im polyamides was observed. The plasma concentration-time profiles after the intravenous administration of Py-Im polyamides (A) and (B) were fitted well by a two-compartment model. Py-Im polyamide (C) was observed at high concentrations in the lungs. The plasma concentration-time profiles after the intravenous administration of Py-Im polyamide (C) were described using a catenary two-compartment model. This model is useful for describing the time course after the administration of high-molecular-weight Py-Im polyamides., (2009 John Wiley & Sons, Ltd.)

Three types of medication, Arbekacin, Vancomycin, and Teicoplanin, are used primarily to treat MRSA infections. These medications differ in their respective anti-bacterial actions, antibacterial spectrums, and pharmacokinetics. Proper use and dosage is required, and is based on patient background and the conditions of infection, among other factors. This study was conducted for a period of over one year at St. Marianna University School of Medicine, Yokohama City Seibu Hospital. It was designed to compare the conditions as they related to why doctors ordered a certain drug, the background, and their clinical examination values. The tendency to avoid selection of Arbekacin Sulfate (ABK) for patients who had kidney dysfunction was recognized, although there were a few exceptions made. Other than that, there were not any standard criteria set in selecting which medication to prescribe. Therefore, it is necessary to examine the appropriateness of the selection since ordering anti-MRSA medication seemed to depend on each doctor's own experience. Serum concentration was measured in order to avoid any side effects. Moreover, cases of young people, normal renal function and malignant tumor patients were recognized in which serum concentration of the anti-MRSA medications was not within the therapeutic range of therapeutic drug monitoring (TDM). This was to show that there is a possibility the medications involved were not sufficiently effective. Therefore, in the future it will be necessary to ensure that proper dosing instructions are followed.

Aim: To examine the effect of Eubacterium limosum (E. limosum) on colonic epithelial cell line in vitro, and to evaluate the effect of E. limosum on experimental colitis., Methods: E. limosum was inoculated anaerobically and its metabolites were obtained. The growth stimulatory effect of the E. limosum metabolites on T84 cells was evaluated by SUDH activity, and the anti-inflammatory effect by IL-6 production. The change in mRNA of toll like receptor 4 (TLR4) was evaluated by real time PCR. Colitis was induced by feeding BALB/C mice with 2.0% dextran sodium sulfate. These mice received either 5% lyophilized E. limosum (n = 7) or control diet (n = 7). Seven days after colitis induction, clinical and histological scores, colon length, and cecal organic acid levels were determined., Results: The E. limosum produced butyrate, acetate, propionate, and lactate at 0.25, 1.0, 0.025 and 0.07 mmol/L, respectively in medium. At this concentration, each acid had no growth stimulating activity on T84 cells; however, when these acids were mixed together at the above levels, it showed significantly high activity than control. Except for lactate, these acids significantly attenuated IL-6 production at just 0.1 mmol/L. In addition, under TNF-alpha stimulation, butyrate attenuated the production of TLR4 mRNA. The treatment with E. limosum significantly attenuated clinical and histological scores of colitis with an increase of cecal butyrate levels, compared with the control group., Conclusion: E. limosum can ameliorate experimental colonic inflammation. In part, the metabolite of E. limosum, butyrate, increases mucosal integrity and shows anti-inflammatory action modulation of mucosal defense system via TLR4.

We studied the effects of 8-hydroxy-2-(di-n-propylamino)tetralin (8-OHDPAT), a 5-HT1A receptor agonist, on both delta9-tetrahydrocannabinol-induced spatial memory impairment in an 8-arm radial maze, and the reduction of acetylcholine release in the dorsal hippocampus as assessed by in vivo microdialysis in rats. A 6 mg/kg i.p. dose of delta9-tetrahydrocannabinol impaired spatial memory in the 8-arm radial maze and decreased the acetylcholine release in the dorsal hippocampus. 8-OHDPAT, at very low doses of 0.1-0.3 microg/kg, reversed both the impairment of spatial memory and the decrease in acetylcholine release induced by delta9-tetrahydrocannabinol. These findings suggest that low doses of 8-OHDPAT may improve delta9-tetrahydrocannabinol-induced impairment of spatial memory by enhancing acetylcholine release in the dorsal hippocampus.

Midazolam and its active metabolites have a depressant effect on respiration and consciousness level, and therefore their effects should be considered in all patients for whom brain death testing is contemplated. The concentrations of midazolam and its active metabolites were measured in critically ill patients on a ventilator during and after continuous intravenous infusion of midazolam. Three days after cessation of midazolam infusion, the concentrations of midazolam and 1-hydroxymidazolam decreased to below the therapeutic range (100-1000 ng/ml) in all patients, although the concentrations of 1-hydroxymidazolam glucuronide remained extremely high in a patient who showed deteriorating renal function. The concentrations of 1-hydroxymidazolam glucuronide (19,497-29,761 ng/ml) were measured in this patient. When it is impossible to confirm factors consistent with irreversible brain death, such as the lack of cerebral blood flow, until 3 days after cessation of midazolam infusion, monitoring of the concentration of these substances should be carried out in all patients in whom suspicion exists prior to the evaluation of brain death. It is particularly imperative that monitoring of the 1-hydroxymidazolam glucuronide concentration be carried out in patients with poor renal function.

In recent years, a pharmacists' work has changed in terms of therapeutic management and counseling of patients. Generally, a pharmacist has to provide pharmaceutical care in cooperation with a doctor. In the present study, a questionnaire survey on what a doctor expects of a pharmacist was conducted, and directors of hospital were interviewed on what they expect of a pharmacist. Eighty-two doctors participated in the survey. As results, doctors consider that a pharmacist's work is dispensing of drugs (41%), and highly evaluated the works of a pharmacist in advising doctors regarding pharmacotherapy (29%), checking the prescription (28%), providing patient compliance instructions (27%) and providing drug information (26%). Five directors of hospital expect the pharmacists to carry out risk managements and prescription checks, provide drug information, and consultation on drug costs. In addition, doctors want pharmacists to have more knowledge on pharmacotherapy. These results clearly show what doctors expect of a pharmacist's work.

To clarify the mechanism by which Delta9-tetrahydrocannabinol, a major psychoactive component of marijuana, impairs spatial memory in the 8-arm radial maze in rats via the cholinergic system, we used two acetylcholinesterase inhibitors, physostigmine and tetrahydroaminoacridine. Moreover, we examined the effect of Delta9-tetrahydrocannabinol on acetylcholine release in the frontal cortex and dorsal and ventral hippocampus using in vivo microdialysis. Physostigmine (0.01-0.05 mg/kg, i.p.) and tetrahydroaminoacridine (1-5 mg/kg, p.o.) improved the impairment of spatial memory induced by Delta9-tetrahydrocannabinol (6 mg/kg, i.p.) in the 8-arm radial maze. Delta9-tetrahydrocannabinol (6 mg/kg, i.p.) produced a significant decrease in acetylcholine release in the dorsal hippocampus as assessed by microdialysis. Moreover, tetrahydroaminoacridine at a dose of 1 mg/kg, which improved the impairment of spatial memory, reversed the decrease in acetylcholine release induced by Delta9-tetrahydrocannabinol in the dorsal hippocampus during 60-120 min after the Delta9-tetrahydrocannabinol injection. These findings suggest that inhibition of the cholinergic pathway by reduced acetylcholine release is one of the means by which Delta9-tetrahydrocannabinol impairs spatial memory in the 8-arm radial maze.

The present study investigated the involvement of the serotonin (5-hydroxytryptamine, 5-HT) neuronal system in the Delta(9)-tetrahydrocannabinol-induced impairment of spatial memory in the eight-arm radial maze in rats. Delta(9)-Tetrahydrocannabinol (6 mg/kg, i.p.), which impairs spatial memory, significantly increased the 5-HT content in the ventral hippocampus. A microdialysis study showed that Delta(9)-tetrahydrocannabinol (6 mg/kg, i.p.) decreased 5-HT release in the ventral hippocampus. The 5-HT precursor, 5-hydroxy-L-tryptophan (5-HTP; 50 mg/kg, i.p.), the 5-HT re-uptake inhibitor, clomipramine (0.01 and 0.1 mg/kg, i.p.), the 5-HT receptor agonist, 5-methoxy-N,N-dimethyltryptamine (5-MeODMT; 0.01 and 0.03 mg/kg, i.p.), and the 5-HT(2) receptor agonist, 1-(2,5-dimethoxy 4-iodophenyl)-2-amino propane (DOI; 10 microg/kg, i.p.), significantly attenuated the Delta(9)-tetrahydrocannabinol-induced impairment of spatial memory. These results suggest that the 5-HT neuronal system may be involved in the Delta(9)-tetrahydrocannabinol-induced impairment of spatial memory.