Rikkunshito simultaneously improves dyspepsia correlated with anxiety in patients with functional dyspepsia: A randomized clinical trial (the DREAM study).

BACKGROUND: Functional dyspepsia (FD), a heterogeneous disorder, involves multiple pathogenetic mechanisms. Developing treatments for FD has been challenging. We performed a randomized, placebo-controlled, double-blind clinical trial to determine the efficacy of rikkunshito, a Japanese herbal medicine, in FD patients. METHODS: FD patients (n = 192) who met the Rome III criteria without Helicobacter pylori infection, predominant heartburn, and depression were enrolled at 56 hospitals in Japan. After 2 weeks of single-blind placebo treatment, 128 patients with continuous symptoms were randomly assigned to 8 weeks of rikkunshito (n = 64) or placebo (n = 61). The primary efficacy endpoint was global assessment of overall treatment efficacy (OTE). The secondary efficacy endpoints were improvements in upper gastrointestinal symptoms evaluated by the Patient Assessment of Upper Gastrointestinal Disorders-Symptom Severity Index (PAGI-SYM), the Global Overall Symptom scale (GOS), and the modified Frequency Scale for the Symptoms of Gastroesophageal Reflux Disease (m-FSSG), and psychological symptoms evaluated by the Hospital Anxiety and Depression Scale (HADS). KEY RESULTS: Rikkunshito increased OTE compared to placebo at 8 weeks (P = .019). Rikkunshito improved upper gastrointestinal symptoms (PAGI-SYM, GOS, and m-FSSG) at 8 weeks, especially postprandial fullness/early satiety (P = .015 and P = .001) and bloating (P = .007 and P = .002) of the PAGI-SYM subscales at 4 weeks and 8 weeks. Improvement of HADS at 8 weeks (P = .027) correlated with those of PAGI-SYM (r = .302, P = .001), GOS (r = .186, P = .044), and m-FSSG (r = .462, P < .001), postprandial fullness/early satiety (r = .226, P = .014), dyspepsia (r = .215, P = .019), and PDS (r = .221, P = .016). CONCLUSION & INFERENCES: Rikkunshito may be beneficial for FD patients to simultaneously treat gastrointestinal and psychological symptoms.

Impairment of gastric accommodation induced by water-avoidance stress is mediated by 5-HT2B receptors.

BACKGROUND: Psychological stress has been shown to impair gastric accommodation (GA), but its mechanism has not been elucidated. This study was conducted to clarify the role of 5-HT2B receptors in a guinea pig model of stress-induced impairment of GA. METHODS: Gastric accommodation was evaluated by measuring the intrabag pressure in the proximal stomach after administration of a liquid meal. The guinea pigs were subjected to water-avoidance stress. The role of 5-HT2B receptors in impairment of GA was investigated by administering a 5-HT2B receptor agonist (BW723C86) or antagonist (SB215505), the traditional Japanese medicine rikkunshito (RKT), a muscarinic M3 receptor antagonist (1,1-dimethyl-4-diphenylacetoxypiperidium iodide [4-DAMP]), or a nitric oxide synthase inhibitor (Nω -nitro-L-arginine [L-NNA]). KEY RESULTS: In normal animals, liquid meal-induced GA was inhibited by BW723C86, but was not affected by SB215505. The inhibition of GA by BW723C86 was reversed by co-administration of 4-DAMP. Compared to normal animals, GA in stressed animals was significantly inhibited. SB215505 and RKT significantly suppressed stress-induced impairment of GA. After meal administration, the level of cyclic guanosine monophosphate in gastric fundus tissue increased by approximately twofold in normal animals, but did not change in stressed animals. The inhibition of GA by L-NNA was suppressed by SB215505 or RKT. At a dose that did not affect GA in normal animals, BW723C86 exacerbated the impairment of GA in stressed animals. CONCLUSIONS AND INFERENCES: Stress-induced impairment of GA may be mediated by an increased responsiveness of 5-HT2B receptors, and activation of the 5-HT2B receptor signaling pathway may have an inhibitory effect on nitric oxide function.

Effect of rikkunshito on the expression of substance P and CGRP in dorsal root ganglion neurons and voluntary movement in rats with experimental reflux esophagitis.

BACKGROUND: While there are reports that the herbal medicine rikkunshito (RKT) relieves upper gastrointestinal disease symptoms, the effect of RKT on primary afferent neurons is unknown. METHODS: A model of reflux esophagitis (RE) was implemented using male Wistar rats aged 6-7 weeks. Ten days after surgery, the total area of esophageal mucosal erosion sites was determined. Th8-10 dorsal root ganglia (DRG) were dissected out and the expression of substance P (SP), calcitonin gene-related peptide (CGRP), and phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2) was determined in DRG using immunohistochemistry. RKT (0.6%/WV) or omeprazole (OME) (10 mg/kg) was administered for 10 days beginning on the day after surgery. Voluntary movement was measured with an infrared sensor for 22 h each day. KEY RESULTS: RE rats showed esophageal mucosal erosion and significantly increased number of SP/CGRP- and p-ERK1/2-immunoreactive neurons in DRG. Treatment with OME improved the size of erosive lesions in the esophageal mucosa of RE rats, while RKT did not. Treatment with RKT or OME significantly reduced the expression of SP/CGRP and p-ERK1/2 in DRG, and significantly increased voluntary movement in RE rats. CONCLUSIONS & INFERENCES: RKT inhibited the activation of ERK1/2 and decreased the expression of SP and CGRP in DRG of RE rats, which may be associated with the observed amelioration of voluntary movement.

An experimental attenuation plate to improve the dose distribution in intraoperative electron beam radiotherapy for breast cancer.

Intraoperative electron beam radiotherapy (IOERT) is a technique in which a single-fraction high dose is intraoperatively delivered to subclinical tumour cells using an electron beam after breast-conserving surgery. In IOERT, an attenuation plate consisting of a pair of metal disks is commonly used to protect the normal tissues posterior to the breast. However, the dose in front of the plate is affected by backscatter, resulting in an unpredictable delivered dose to the tumour cells. In this study, an experimental attenuation plate, termed a shielding plate, was designed using Monte Carlo simulation, which significantly diminished the electron beam without introducing any backscatter radiation. The plate's performance was verified by measurements. It was made of two layers, a first layer (source side) of polymethyl methacrylate (PMMA) and a second layer of copper, which was selected from among other metals (aluminium, copper and lead) after testing for shielding capability and the range and magnitude of backscatter. The optimal thicknesses of the PMMA (0.71 cm) and copper (0.3 cm) layers were determined by changing their thicknesses during simulations. On the basis of these results, a shielding plate was prototyped and depth doses with and without the plate were measured by radiophotoluminescence glass dosimeters using a conventional stationary linear accelerator and a mobile linear accelerator dedicated for IOERT. The trial shielding plate functioned as intended, indicating its applicability in clinical practice.

Potential immunosuppressive and antiinflammatory activities of Malaysian medicinal plants characterized by reduced cell surface expression of cell adhesion molecules.

In the search for agents effective against immune-mediated disorders and inflammation, we have screened Malaysian medicinal plants for the ability to inhibit the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) on the surface of murine endothelial cells (F-2), and mouse myeloid leukaemia cells (M1), respectively. Of 41 kinds (29 species, 24 genera, 16 families) of Malaysian plants tested, 10 and 19 plant samples significantly downregulated the expression of ICAM-1 and VCAM-1, respectively. Bioassay-directed fractionation of an extract prepared from the bark of Goniothalamus andersonii showed that its ingredients, goniothalamin (1) and goniodiol (2) inhibited the cell surface expression of both ICAM-1 and VCAM-1. The present results suggest that Malaysian medicinal plants may be abundant natural resources for immunosuppressive and antiinflammatory substances.

Influence of natural and synthetic compounds on cell surface expression of cell adhesion molecules, ICAM-1 and VCAM-1.

Various natural and synthetic compounds including alkaloids, terpenoids and phenolics were tested for inhibition of the cell surface expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), both of which are crucial in the regulation of immune response and inflammation. Of 40 compounds tested, two compounds significantly downregulated the expression of VCAM-1 on murine endothelial cells (F-2) and ten compounds that of ICAM-1 on mouse myeloid leukemia cells (M1). Sanguinarine chloride (5) and isoliquiritigenin (13) were capable of lowering the levels of both ICAM-1 and VCAM-1. The structure-activity relationships study on chalcone and flavone derivatives related to 13 suggested that the inhibitory activity of the chalcone derivatives is attributable to the 4-hydroxy group as well as the possible coplanarity between the phenyl ring and the adjacent conjugated ketone.

Expression of connexin 30 in the developing mouse cochlea.

Mutations in the GJB6 gene encoding connexin 30 (Cx30) can cause dominant forms of nonsyndromic deafness. By studying immunohistochemical localization of Cx30 in the mouse cochlea at different ages from 0 to 30 days after birth, we found that the expression of Cx30 is nearly the same as that of Cx26. These findings suggest that as well as Cx26, Cx30 may also contribute to the generation and maturation of endocochlear potential.

Combined treatment with TNP-470 and 5-fluorouracil effectively inhibits growth of murine colon cancer cells in vitro and liver metastasis in vivo.

The combined effects of TNP-470 (TNP), a semisynthetic analogue of fumagillin, and 5-fluorouracil (5FU), a representative chemotherapeutic agent for colorectal cancer, were investigated using murine colon 26 adenocarcinoma (CT 26) cells. In a cell-proliferation study in vitro, 50% inhibitory concentrations (IC50) were determined to be 5.2 microg/ml and 240 ng/ml for TNP and 5FU, respectively. When CT 26 cells were treated with TNP and 5FU in combination, a remarkable cytotoxic effect was obtained. Isobologram analysis revealed synergism of these two agents in inhibition of the cell growth. In vivo, using a dorsal air sac assay, we found that TNP significantly inhibited the CT 26-induced angiogenesis. In addition, the combination of TNP and 5FU exerted a synergistic anti-tumor effect in a model of hepatic metastasis by portal injection of CT 26 cells. Since TNP is known to exert inhibitory effects on tumor cell growth through suppression of cell cycle progress from the G1 to S phases as well as neovascularization, it is speculated that the treatment with TNP enhanced the anti-tumor effect of 5FU through suppression of the cell cycle and tumor-derived angiogenesis. Taken together, these results suggest that combined treatment with TNP and 5FU is potentially useful for inhibition of tumor cell growth and liver metastasis of colorectal cancer.

Effectiveness of 1,25-dihydroxyvitamin D supplementation on blood pressure reduction in a pseudohypoparathyroidism patient with high renin activity.

A 42-year-old man had biochemical and somatic abnormalities compatible with pseudohypoparathyroidism type I (PsHP) and also had high plasma renin activity (PRA). After 1,25-dihydroxyvitamin D (calcitriol) supplementation the systolic/diastolic blood pressure, assessed by 24-hour non-invasive ambulatory blood pressure monitoring, was reduced from 145/96 mm Hg to 128/85 mm Hg with normalization of the serum calcium level and its related hormones, as well as decreased PRA. Calcitriol supplementation successfully reduced the blood pressure in this patient with PsHP and a high PRA, suggesting that calcium-related hormones and/or the renin-angiotensin system were involved in lowering the blood pressure.

Alternatively spliced isoforms of the Na+/Ca2+ exchanger in the guinea pig cochlea.

The cochlea has been suggested to express some Na+/ Ca2+ exchangers (NCX), since efficient acoustic transduction requires cytosolic calcium homeostasis. The present study revealed that several spliced isoforms of NCX are expressed in the guinea pig cochlea. Moreover, to determine their localization in the cochlea, microdissected RT-PCR was performed. The guinea pig cochlea was microdissected into three parts (lateral wall, the organ of Corti and modiolus). The cochlear lateral wall and the organ of Corti expressed only a single isoform of NCX1. On the other hand, five isoforms of NCX1 and four isoforms of NCX3 were detected in the cochlear modiolus. The alternative splicing may provide diverse functions for NCX in the cochlea.

The crystal structure of zinc-containing ferredoxin from the thermoacidophilic archaeon Sulfolobus sp. strain 7.

The crystal structure of ferredoxin from the thermoacidophilic archaeon Sulfolobus sp. strain 7 was determined by multiple isomorphous replacement supplemented with anomalous scattering effects of iron atoms in the Fe-S clusters, and refined at 2.0 A resolution to a crystallographic R value of 0.173. The structural model contains a polypeptide chain of 103 amino acid residues, 2 [3Fe-4S] clusters, and 31 water molecules; in this model, the cluster corresponding to cluster II in bacterial dicluster ferredoxins loses the fourth iron atom although it may originally be a [4Fe-4S] cluster. The structure of the archaeal ferredoxin consists of two parts: the core fold part (residues 37-103) and the N-terminal extension part (residues 1-36). The "core fold" part has an overall main-chain folding common to bacterial dicluster ferredoxins, containing two clusters as the active center, two alpha-helices near the clusters, and two sheets of two-stranded antiparallel beta-sheet (the terminal and central beta-sheets). The "N-terminal extension" part is mainly formed by a one-turn alpha-helix and a three-stranded antiparallel beta-sheet. The beta-sheet in the N-terminal extension is hydrogen-bonded with the terminal beta-sheet in the core fold to form a larger beta-sheet. The distinct structural feature of this archaeal ferredoxin lies in the zinc-binding center where the zinc ion is tetrahedrally ligated by four amino acid residues (His 16, His 19, and His 34 from the N-terminal extension, and Asp 76 from the core fold). The zinc ion in the zinc-binding center is located at the interface between the core fold and the N-terminal extension, and connects the beta-sheet in the N-terminal extension and the central beta-sheet in the core fold through the zinc ligation. Thus, the zinc ion plays an important role in stabilizing the structure of the present archaeal ferredoxin by connecting the N-terminal extension and the core fold, which may be common to thermoacidophilic archaeal ferredoxins.

Laser-assisted uvulopalatoplasty for habitual snoring without sleep apnea: outcome and complications.

Thirty patients suffering from habitual snoring were subjected to laser-assisted uvulopalatoplasty with a KTP/532 laser under local anesthesia. The patients selected for the present study had no complaints of severe sleep apnea. The surgical procedure included bilateral vertical incision through the palate at the base of the uvula with or without removal of the lower half of the uvula. Ninety-three percent of the patients showed apparent improvement of snoring following the operation. However, other sleep-related symptoms such as sleep quality and daytime sleepiness were not significantly improved. No major or critical complications such as massive bleeding and asphyxia occurred. Post-operative pain on deglutition disappeared in most patients 2 weeks after the operation. This procedure is safe, minimally invasive and effective for habitual snoring without apnea.

Expression of voltage-dependent chloride channels in the rat cochlea.

Voltage-dependent chloride (ClC) channels have not yet been identified in the cochlea. In this study, an approach utilizing the reverse transcription-polymerase chain reaction (RT-PCR) was devised to clone the cDNA of ClC channels. PCR was performed using degenerate primers corresponding to two highly conserved regions of the ClC channels. By Southern hybridization and sequencing studies, the sequences corresponding to ClC-2 and ClC-3 were found in the cochlear lateral wall, while ClC-1 was not detected. These results suggest that ClC-2 and ClC-3 might be involved in Cl- transport in the cochlear lateral wall.

The Na-K-Cl cotransporters in the rat cochlea: RT-PCR and partial sequence analysis.

Nonsensory epithelial cells of the mammalian cochlea contribute to the production of the endolymph. The presence as well as the function of the Na-K-Cl cotransporter has been suggested, but not yet been proved. To identify the Na-K-Cl cotransporter isoforms expressed in the cochlea, mRNA was extracted from the cochlear lateral wall. After reverse transcription, resulting cDNA was amplified by polymerase chain reaction (PCR) with primers specific for Na-K-Cl cotransporter isoforms. In the rat cochlear lateral wall, mRNA homologous to the mouse bumetanide-sensitive Na-K-Cl cotransporter (mBSC2) was detected. These results suggest that the basolateral localization of the cotransporter in the marginal cell is involved in the secretory process of the endolymph.

Systemic and cellular calcium metabolism and hypertension.

The calcium status of humans with essential hypertension and genetic animal models of hypertension is characterized by low serum ionized calcium concentration, increased urinary calcium excretion, and increased parathyroid hormone (PTH) concentration. Calcitriol metabolism and bone mineralization are also altered in hypertension. These alterations in systemic calcium metabolism may be linked to factors responsible for the elevated blood pressure. Cytosolic free calcium tends to be increased in most cells that have been studied from hypertensive humans and animals. Changes in cellular calcium metabolism may be partly mediated by calcium-regulating hormones that tend to be elevated in essential hypertension such as PTH and calcitriol. Administration of supplemental dietary calcium tends to suppress PTH, calcitriol, and intracellular free calcium. Further research is need concerning the observed association among systemic markers of calcium metabolism, cellular calcium metabolism, and arterial blood pressure regulation.

Cutaneous iontophoretic application of condensed lidocaine.

The purpose of this study was to determine whether iontophoretic application of high concentrations of lidocaine, with the same current, would produce cutaneous local anaesthesia rapidly enough for clinical practice. Twenty healthy volunteers, 17 male and three female, were selected for study. After five-minute or ten-minute iontophoresis using lidocaine 4, 10, 20, 30, 50%, we assessed the response to pin prick with a 27-gauge sterile needle inserted to the depth of 2 mm at five random locations in the iontophoretically-stimulated area. Also, plasma lidocaine concentrations were measured in the venous blood samples which had been taken from three male subjects, at 3, 10, and 30 min after iontophoresis with lidocaine 50%. The pain score after five-minute iontophoresis was higher than that after ten-minute iontophoresis, using each concentration of lidocaine (P < 0.001), whereas the pain scores had no correlation with lidocaine concentration within five-minute and ten-minute iontophoresis groups, respectively (P: NS). On the other hand, plasma lidocaine concentration was < 1.0 micrograms.ml-1 in all samples. No side effects other than erythema were observed after iontophoresis using high concentrations of lidocaine up to 50%. These results showed that by increasing the lidocaine concentration of the applied solution up to 50%, the application time of iontophoresis cannot be reduced from ten to five minutes without losing analgesic effect, although iontophoresis itself can be performed with safety.

[Hydralazine-induced enhancement of hyperthermia treatment in vivo].

Hydralazine (Hyd) is a vaso-active drug that significantly affects the nature of blood flow in tumors. As a result, Hyd reduces blood flow and oxygen tension in tumors, causing an increase in the toxic effect of hyperthermia treatment. We investigated enhancement of the anti-tumor effect of hyperthermia by Hyd on SCC-VII tumors in C3H mice. Hyd was administered by intraperitoneal injection, and tumors were heated by water bath. We measured the tumor temperature in animals receiving Hyd by thermocouple. We found no significant change in tumor temperature with Hyd treatment. The effect of Hyd (2.5 mg/kg, 5.0 mg/kg, 7.5 mg/kg) on tumors was evaluated in terms of a growth delay value at which tumor volume reached four-fold. The growth delay values obtained were 6.25 +/- 0.82, 7.14 +/- 0.90, 8.50 +/- 0.98, 9.72 +/- 0.92, and 9.84 +/- 1.3 days for: hyperthermia alone, Hyd of 1.0 mg/kg, 2.5 mg/kg, 5.0 mg/kg, respectively. This effect was independent of the time course of administration of Hyd. These results indicate that Hyd can increase the therapeutic efficacy of hyperthermia treatment. Changes in the microenvironment, such as low pH and tumor hypoxia, induced by arterial embolization may have increased the sensitivity of tumors to heat.

[Evaluation of thermal damage after hyperthermia on murine experimental tumor by 31P-NMR spectroscopy--correlation between ATP and growth curve].

The possibility of using 31P-NMR spectroscopy (31P-MRS) to estimate the effect of hyperthermic treatment on mouse FM3A tumor was investigated. 1 x 10(6) cells, suspended in saline, were subcutaneously inoculated to the right thigh of C3H mice. For hyperthermic treatment, the tumor-bearing leg was heated by immersing it in a water bath at 44 degrees C for 10, 20 or 30 min. The signal intensities of ATP and Pi of the tumor were measured utilizing the 31P-MRS technique to calculate the ATP/Pi ratio. Immediately after heating, the ATP/Pi ratio decreased markedly. Eighteen hours after hyperthermic treatment, the ratio recovered but was still smaller than the control value, then became almost constant by 24 hours after heating. The ATP/Pi ratio at 24 hours after heating decreased with increased length of heating and was inversely related to tumor regrowth after hyperthermic treatment. We concluded that non-invasive monitoring with 31P-MRS might provide a good indication of the effect of hyperthermic treatment.

Recombinant human erythropoietin, but not iron supplementation, improves anemia in rats with adjuvant-induced arthritis.

Pathophysiological and therapeutic properties of anemia in rats with adjuvant-induced arthritis (AA) were investigated. Both anemia and chronic inflammation were induced in rats by a single injection of Freund's complete adjuvant. This study confirmed other earlier data that these anemic rats with AA had reduced serum iron levels and that the anemia was characterized as mild, non-progressive, hypochromic, microcytic. In addition, our studies showed that these anemic rats had slightly but significantly enhanced erythropoietin titers, but not renal failure; there was no significant difference in blood urea nitrogen and creatinine levels in anemic and normal groups. The anemia in rats with AA was improved by recombinant human erythropoietin (r-HuEPO) at 30 and 100 U/kg/day, given i.v. for 5 days. In contrast, iron-chondroitin-sulfate colloid (10 mg/kg/day, i.v. for 5 days) failed to improve the anemia and to enhance the effects of r-HuEPO. These data suggest that anemia in rats with adjuvant-induced arthritis is distinguished, pathophysiologically and therapeutically, from iron deficiency anemia, hemolytic anemia, and renal anemia.

Application of ion-pair high performance liquid chromatography for analysis of hyoscyamine and scopolamine in solanaceous crude drugs.

A new ion-pair high-performance liquid chromatographic method for the analysis of hyoscyamine and scopolamine in various kinds of crude drugs derived from solanaceous plants was evaluated using sodium dodecyl sulfate as a counter ion. A reversed-phase chromatographic system consisting of a chemically bonded ODS silica gel column with phosphate buffer (pH 2.5)-acetonitrile (65:35) containing 17.5 mM sodium dodecyl sulfate as the mobile phase was used. Hyoscyamine and scopolamine in crude drugs derived from Scopolia, Atropa, Hyoscyamus and Datura species were separated from other compounds in the crude drugs and determined within 20 min after direct injection of the solution extracted with the mobile phase. The results for various kinds of samples are presented.