bluuebunny/crossref_metadata_2025 huggingface dataset:
Reproductive Cycle Regulation of Nuclear Import, Euchromatic Localization, and Association with Components of Pol II Mediator of a Mammalian Double-Bromodomain Protein
Authors: Crowley Thomas E. , Kaine Emily M. , Yoshida Manabu , Nandi Anindita , Wolgemuth Debra J.
Published: October 2002

Fsrg1 (female sterile homeotic-related gene 1) is the mouse homolog of the human RING3 protein, which has been shown to associate with the E2 promoter binding factor (E2F) transcription factor and to have a possible role in cell cycle-linked transcriptional regulation. The Fsrg1 protein is 60% identical in sequence to the RNA polymerase II mediator subunit Fsrg4, another member of this subfamily of double bromodomain-containing proteins that are homologs of Drosophila female sterile homeotic. Antibodies against murine Fsrg1 were generated and used in immunoblot and immunoprecipitation experiments to identify proteins interacting with Fsrg1 and RING3. In the presence of acetylated but not nonacetylated histone H3 and H4 peptides, RING3 was shown to interact with E2F, mediator components cyclin-dependent kinase 8 and thyroid receptor-associated protein 220, and the RNA polymerase II large subunit. Fsrg1 mRNA had been previously shown to be expressed at high levels in the epithelium of the adult mouse mammary gland. To determine the physiological relevance of these potential associations, we examined the patterns of expression of Fsrg1 mRNA and protein in the adult mammary epithelia during the reproductive cycle as the tissue is responding to estrogen, progesterone, and prolactin. Changes in the nuclear vs. cytoplasmic localization of Fsrg1 were observed and correlated with physiological changes in mammary gland function. The observations suggested that Fsrg1 may be involved in the transcriptional activities of genes involved in proliferation of the mammary epithelia during pregnancy and in orchestrating postlactation involution and apoptosis. Localization of Fsrg1 on euchromatin, the transcribed portion of the chromosomes, is consistent with its hypothesized function as a transcription regulator.
Evaluation of the Pharmacological Actions and Pharmacokinetics of BOF-4272, a Xanthine Oxidase Inhibitor, in Mouse Liver
Authors: Naito Shinsaku , Nishimura Masuhiro , Tamao Yumi
Published: October 2002

BOF-4272 (±)-8-(3-methoxy-4-phenylsulphinylphenyl) pyrazolo[1,5-a]-1,3,5-triazine-4-(1H)-one, a new synthetic anti-hyperuricaemic drug, which has a chiral centre and exists as racemates, is a potent inhibitor of xanthine oxidase/dehydrogenase in the purine catabolism pathways. The present studies using mice demonstrated that BOF-4272 was specifically distributed in the liver, which is the main organ of uric acid production. Therefore, a decrease in uric acid concentration in the liver, rather than the plasma, was identified as a pharmacological action of BOF-4272.

The ratio of liver to plasma concentrations of BOF-4272 increased from 2.5 to 6.3 over time, up to 8 h after oral administration. The elimination half-life of BOF-4272 in the liver was 5.1-fold longer than that in the plasma. High concentrations of BOF-4272 were observed in the liver up to 8 h after oral administration. Furthermore, the influx of BOF-4272 into hepatocytes occurred in a temperature-dependent manner.

The liver concentrations of uric acid from 1 h to 8 h after the oral administration of BOF-4272 (0.34–0.75 μg (g tissue)−1) were significantly lower than those in control animals (5.03–10.96 μg (g tissue)−1). BOF-4269 (the sulphide metabolite of BOF-4272) was the only metabolite detected in plasma or faeces after intravenous or oral administration. BOF-4269, which has no inhibitory action on the uric acid biosynthesis system, is generated by the metabolism of BOF-4272 in the intestinal tract.

In conclusion, this work using the liver as the target organ has allowed us to identify the pharmacological actions of BOF-4272 in mice. The long-lasting effect of BOF-4272 in reducing levels of hepatic uric acid was consistent with the prolonged high BOF-4272 concentrations in the liver. These results also demonstrate that the mouse is a suitable animal species for evaluating the clinical pharmacology and pharmacokinetics of BOF-4272.
Rapid Detection of CYP2C18 Genotypes by Real-time Fluorescence Polymerase Chain Reaction
Authors: Mizugaki Michinao , Hiratsuka Masahiro , Agatsuma Yasuyuki , Matsubara Yoichi , Fujii Kunihiro , Kure Shigeo , Narisawa Kuniaki
Published: October 2002

In man, CYP2C19, a liver enzyme, plays an important role in the metabolism of several drugs. Mutation of the CYP2C19 gene results in a poor metaboliser phenotype. S-Mephenytoin hydroxylation genetic polymorphism is due to two mutations of the CYP2C19 gene, namely CYP2C19*2, located in exon 5, and CYP2C19*3, located in exon 4. CYP2C18 is also polymorphically expressed. The mutant alleles of this enzyme are CYP2C18m1, located in exon 2 and CYP2C18m2, located in the 5′-flanking region. We have developed an allele-specific TaqMan polymerase chain reaction (PCR) assay with which to detect CYP2C18 mutant alleles. This assay combines hybridization of the TaqMan probe and allele-specific amplification primers to the target DNA. The TaqMan probe is labelled with 6-carboxyfluorescein at the 5′ end and 6-carboxytetramethylrhodamine together with a phosphate at the 3′ end. Genotypes are separated according to the different threshold cycles of the wild type and mutant primers.

We applied this procedure to DNA extracted from the blood or saliva of 144 healthy Japanese volunteers. The wt/wt, wt/m1, wt/m2, m1/m1, m1/m2 and m2/m2 genotypes of the CYP2C18 alleles detected by the assay were consistent with the results obtained from restriction enzyme cleavage. In accordance with a previous report, the genotypes of CYP2C18m1 and CYP2C18m2 coincided with those of CYP2C19*3 and CYP2C19*2, respectively. Therefore, detection of CYP2C18 mutant alleles also allows that of CYP2C19 mutant alleles. Among 19 poor metabolisers, eight showed the homozygous CYP2C19*2/CYP2C19*2, two the homozygous CYP2C19*3/CYP2C19*3 and nine the compound heterozygous CYP2C19*2/CYP2C19*3 genotype.

We found the allele-specific TaqMan PCR assay rapid, simple and cost-effective, as well as suitable for high-throughput applications in a routine laboratory. This assay allows the fast and reliable detection of inherited disorders that might influence diagnosis and treatment.
Antiplatelet Effect of Marchantinquinone, Isolated from Reboulia hemisphaerica, in Rabbit Washed Platelets
Authors: Liao Chang-Hui , Ko Feng-Nien , Wu Chia-Li , Teng Che-Ming
Published: October 2002

Platelet activation is involved in serious pathological situations, including atherosclerosis and restenosis. It is important to find efficient antiplatelet medicines to prevent fatal thrombous formation during the course of these diseases.

Marchantinquinone, a natural compound isolated from Reboulia hemisphaerica, inhibited platelet aggregation and ATP release stimulated by thrombin (0.1 units mL−1), platelet-activating factor (PAF; 2 ng mL−1), collagen (10 μg mL−1), arachidonic acid (100 μm), or U46619 (1 μm) in rabbit washed platelets. The IC50 values of marchantinquinone on the inhibition of platelet aggregation induced by these five agonists were 62.0 ± 9.0, 86.0 ± 7.8, 13.6 ± 4.7, 20.9 ± 3.1 and 13.4 ± 5.3 μm, respectively. Marchantinquinone inhibited thromboxane B2 (TxB2) formation induced by thrombin, PAF or collagen. However, marchantinquinone did not inhibit TxB2 formation induced by arachidonic acid, indicating that marchantinquinone did not affect the activity of cyclooxygenase and thromboxane synthase. Marchantinquinone did inhibit the rising intracellular Ca2+ concentration stimulated by the five platelet-aggregation inducers. The formation of inositol monophosphate induced by thrombin was inhibited by marchantinquinone. Platelet cAMP and cGMP levels were unchanged by marchantinquinone.

The results indicate that marchantinquinone exerts antiplatelet effects by inhibiting phosphoinositide turnover.
In-vivo and In-vitro Evidence of a Carrier-mediated Efflux Transport System for Oestrone-3-sulphate across the Blood-Cerebrospinal Fluid Barrier
Authors: Kitazawa Takeo , Hosoya Ken-Ichi , Takahashi Takeo , Sugiyama Yuichi , Terasaki Tetsuya
Published: October 2002

The efflux transport of oestrone-3-sulphate, a steroid hormone sulphate, across the blood-cerebrospinal fluid barrier has been examined following its intracerebroventricular administration. [3H]Oestrone-3-sulphate was eliminated from cerebrospinal fluid (CSF) with an apparent efflux clearance of 205 μL min−1 per rat. There was 25% of unmetabolized [3H]oestrone-3-sulphate in the plasma 5 min after intracerebroventricular administration, indicating that at least a part of [3H]oestrone-3-sulphate is transported from CSF to the circulating blood across the blood-CSF barrier. This efflux transport was inhibited by co-administration of excess oestrone-3-sulphate (25 mm 10 μL = 0.25 μmol) into rat cerebral ventricle.

To characterize the oestrone-3-sulphate transport process, an in-vitro uptake experiment was performed using isolated rat choroid plexus. Oestrone-3-sulphate uptake by isolated rat choroid plexus was found to be a saturable process with a Michaelis-Menten constant (Km) of 18.1 ± 6.3 μm, and a maximum uptake rate (Vmax) of 48.0± 15.1 pmol min−1 μL−1 of tissue. The oestrone-3-sulphate transport process was temperature dependent and was inhibited by metabolic inhibitors such as 2,4-dinitrophenol and rotenone, suggesting an energy dependence. This uptake process was also inhibited by steroid hormone sulphates (1 mm dehydroepiandrosterone sulphate and 1 mm oestrone sulphate), bile acids (1 mm taurocholic acid and 1 mm cholic acid) and organic anions (1 mm sulphobromophthalein and 1 mm phenolsulphonphthalein), whereas 1 mm  p-aminohippuric acid, 1 mm  p-nitrophenol sulphate, 0.1 mm methotrexate and the cardiac glycoside, 2.5 μm digoxin, had little effect.

In conclusion, these results provide evidence that oestrone-3-sulphate is transported from CSF to the circulating blood across the blood-CSF barrier via a carrier-mediated efflux transport system.
Effects of Lithium on the Pharmacokinetics of Valproate in Rats
Authors: Yoshioka Hiroshi , Ida Seiko , Yokota Masayuki , Nishimoto Ayako , Shibata Sachiko , Sugawara Akiko , Takiguchi Yoshiharu
Published: October 2002

Combined treatment with lithium and valproate has been used for bipolar disorder. However, the studied interaction between these two drugs has not been fully investigated. We therefore examined the effects of lithium on the pharmacokinetics (plasma disappearance, metabolism and urinary excretion) of valproate in rats.

Lithium (2 mEq kg−1) was administered intraperitoneally twice a day for ten days. Plasma disappearance curves of valproate (50 mg kg−1, i.v.), valproate-metabolizing activities of UDP-glucuronosyltransferase (UGT) and cytochrome P450 (CYP) in liver microsomes and urinary excretion of free valproate and valproate-glucuronide were examined. The metabolizing activity of UGT and CYP were determined by enzyme assays and a fluorescence polarization immunoassay system. Urinary valproate-glucuronide was obtained using this system by subtracting the free level from total level, which was determined after deconjugating the sample with heat and NaOH.

The half-life of plasma disappearance of valproate was 25% reduced by lithium pretreatment (0428 ± 0.031 h with repeated lithium pretreatment vs 0.578 ± 0.062 h for controls). The valproate-metabolizing activity of UGT and CYP were not altered by lithium although lithium increased the urinary excretion of valproate-glucuronide.

In conclusion, lithium pretreatment causes a decrease in plasma valproate levels and an increase in urinary excretion of valproate-glucuronide in rats.
Intestinal Transport of β-Thioglycosides by Na+/Glucose Cotransporter
Authors: Mizuma Takashi , Hagi Katsura , Awazu Shoji
Published: October 2002

Intestinal metabolism and transport of p-nitrophenyl β-d-thioglucoside (p-NPβSglc) and p-nitrophenyl β-d-thiogalactoside (p-NPβSgal) by the Na+/glucose cotransporter were studied in excised small intestine of the rat.

p-NPβSglc and p-NPβSgal were stable enough on the mucosal side to be transported to the serosal side. Transport of p-NPβSglc was inhibited in the presence of phloridzin (a Na+/glucose cotransporter inhibitor), glucose, or 3-O-methylglucose (3-OMG). p-NPβSglc transport was dependent on Na+ concentration in a sigmoidal manner. The activation energy for transport was 19.4 kcal mol−1. The distribution of transport activity of p-NPβSglc in each region of the small intestine correlated well with that of 3-OMG. These results indicate that p-NPβSglc is transported by the Na+/glucose cotransporter in small intestine. The order of turnover rate for glycoside transport by Na+/glucose cotransporter was 3-OMG > p-nitrophenyl β-O-glucoside > p-NPβSglc > p-NPβSgal, indicating that the presence of a galactose moiety and a sulphur between the monosaccharide moiety and aglycone decreases the turnover rate of the Na+/glucose cotransporter in the transport of glycosides. In an inhibition study using stable p-NPβSglc as a Na+/glucose cotransporter-transportable marker glucoside, it was also shown that the Na+/glucose cotransporter recognized several types of glycosides.

In conclusion, displacement of the oxygen at carbon C-l of glucose by sulphur in thioglycosides decreases the turnover rate of the Na+/glucose cotransporter, but thioglycosides are stable in the small intestine and are transported by the Na+/glucose cotransporter.
Effect of Trimethoprim on the Renal Clearance of Lamivudine in Rats
Authors: Takubo Takatoshi , Kato Toshihiro , Kinami Junji , Hanada Kazuhiko , Ogata Hiroyasu
Published: October 2002

Lamivudine undergoes minimal metabolism and renal clearance of the unchanged drug is the predominant mechanism of clearance. The effect of trimethoprim on the renal clearance of lamivudine was investigated in rats in-vivo.

Total renal clearance of lamivudine was about three times higher than the glomerular filtration rate in rats receiving an infusion of tritium-labelled lamivudine. Concomitant infusion of trimethoprim reduced the renal clearance of lamivudine to about half, but did not affect the level of radioactivity in the renal cortex. When rats received an infusion of lamivudine with probenecid, cimetidine or quinidine, the renal clearance of lamivudine was only significantly reduced by co-administration of cimetidine.

These findings suggest that secretion in the renal proximal tubule takes an active part in the total renal clearance of lamivudine, and that cationic drugs such as trimethoprim and cimetidine may inhibit the secretion of lamivudine without greatly affecting the concentration of lamivudine in the renal cortex.
Characterization of Histamine Release Induced by Fluoroquinolone Antibacterial Agents In-vivo and In-vitro
Authors: Mori Kazuhiko , Maru Chikako , Takasuna Kiyoshi
Published: October 2002

Characterization of histamine release induced by fluoroquinolone antibacterial agents, levofloxacin and ciprofloxacin, was investigated in-vivo and in-vitro.

Intravenous injection of levofloxacin and ciprofloxacin at 1–10 mg kg−1 produced dose-related elevations in plasma histamine level in anaesthetized dogs. In contrast, levofloxacin was devoid of plasma histamine increment in anaesthetized rats at 100 mg kg−1, whereas ciprofloxacin at the same dose caused endogenous histamine release. Levofloxacin and ciprofloxacin induced non-cytotoxic secretion of histamine from all mast cells tested in a concentration-dependent manner, whereas rat skin and peritoneal mast cells were thirty- to one-hundred-times less sensitive to the effect of fluoroquinolones as compared with the canine skin mast cells.

These results suggest that the functional heterogeneity of mast cells from different species in histamine releasing activity of fluoroquinolones may exist, and that mast cells from the dog appear to be particularly sensitive to the effect of the fluoroquinolones.
Anti-leukaemic and Anti-mutagenic Effects of Di(2-ethylhexyl)phthalate Isolated from Aloe vera Linne
Authors: Lee Keyong Ho , Kim Jeong Hwan , Lim Dae Seog , Kim Chang Han
Published: October 2002

Extracts of Aloe vera Linne have been found to exhibit cytotoxicity against human tumour cell lines. This study examines the anti-tumour effects of di(2-ethylhexyl)phthalate (DEHP) isolated from Aloe vera Linne, in human and animal cell lines. Its anti-mutagenic effects were examined using Salmonella typhimurium TA98 and TA100 strains.

Growth inhibition was specifically exerted by DEHP against three leukaemic cell lines at concentrations below 100 μg mL−1. At 100 μg mL−1 DEHP, K562, HL60 and U937 leukaemic cell lines showed growth inhibition of 95, 97 and 95%, respectively. DEHP exhibited an inhibitory activity of 74, 83 and 81%, respectively, in K562, HL60 and U937 cell lines at a concentration of 10 μg mL−1. At a concentration of 1 μg mL−1, DEHP exerted an inhibitory activity of 50, 51 and 52%, respectively, in K562, HL60 and U937. In a normal cell line, MDBK, DEHP exerted 30% growth inhibition at a concentration of 100 μg mL−1, and showed no inhibitory activity at concentrations below 50 μg mL−1.

It was found that DEHP exerted anti-mutagenic activity in the Salmonella mutation assay. The number of mutant colonies of Salmonella typhimurium strain TA98 upon exposure to AF-2 (0.2 μg/plate) decreased in a concentration-dependent manner in the presence of different DEHP concentrations (decreasing to 90.4, 83.9, 75.4, 69.6 and 46.9%, respectively, for DEHP concentrations of 100, 50, 10, 5 and 1 μg/plate). In the case of Salmonella typhimurium strain TA100, DEHP reduced AF-2-induced mutagenicity at 1, 5, 10, 50 and 100 μg/plate to 57.4, 77.5, 80.0, 89.0 and 91.5%, respectively.

The isolated compound from Aloe vera Linne, DEHP, was considered to be the active principle responsible for anti-leukaemic and anti-mutagenic effects in-vitro.
Investigation of the Absorption of Hypericin into the Skin of Hairless Mice
Authors: Kamuhabwa Appolinary R , Geboes Karel R , De Witte Peter A
Published: October 2002

The skin absorption of hypericin was evaluated in hairless mice to develop an optimised hypericin topical formulation that could be used in the clinical study of psoriasis.

Hypericin (0.01–1.0%) in Beeler basis, polyethylene glycol ointment, carbopol gel, cetomacrogol cream, petrolatum or emulsifying ointment, with and without skin-absorption enhancers (isopropylidene glycerol and diethylene glycol monoethyl ether), was tested in-vivo on hairless mice skin. Using a skin-stripping technique and the intrinsic fluorescence of hypericin under standardised UV365 irradiation, it was demonstrated that the absorption of hypericin very much depended on the vehicle used. The concentrations of hypericin in the skin were then estimated by HPLC analysis. For this purpose, two vehicles were employed, with which hypericin penetrated the skin of hairless mice well (emulsifying ointment with isopropylidene glycerol) or very poorly (polyethylene glycol ointment). In the case of emulsifying ointment with isopropylidene glycerol (0.05% hypericin), a substantial concentration of hypericin (8.6±3.2 μg g−1) (mean ± s.d., n = 5) was found in the skin. With polyethylene glycol ointment, however, only a limited hypericin skin concentration (0.38 ± 0.34 μg g−1, n = 5) was achieved.

These results show that emulsifying ointment with polyethylene glycol holds promise as an effective topical vehicle for the treatment of skin diseases, such as psoriasis, with hypericin.
The Upstream Regulation of p38 Mitogen-activated Protein Kinase Phosphorylation by Arachidonic Acid in Rat Neutrophils
Authors: Chang Ling C , Wang Jih P
Published: October 2002

The signal transduction pathways activated by arachidonic acid that lead to p38 mitogen-activated protein kinase (MAPK) activation in neutrophils remains unclear. In this study, selective inhibitors of several signalling pathways were utilized to investigate the mechanisms of activation of p38 MAPK by arachidonic acid in rat neutrophils.

Stimulation of p38 MAPK phosphorylation by arachidonic acid and its trifluoromethyl ketone analogue AACOCF3 was transient, peaking at 1 min, and was concentration-dependent. Arachidonic acid-stimulated p38 MAPK phosphorylation was attenuated in cells pretreated with the Gi/o inhibitor (pertussis toxin), but not with the dual cyclooxygenase/lipoxygenase inhibitor (BW755C) or the leukotriene biosynthesis inhibitor (MK886). Tyrosine kinase inhibitor (genistein), but not the extracellular signal-regulated kinase kinase inhibitors (PD98059 and U0126), attenuated the phosphorylation of p38 MAPK by arachidonic acid. Phosphoinositide 3-kinase inhibitors (wortmannin and LY294002) did not affect the arachidonic acid-induced response. After pretreatment of the cells with protein kinase C inhibitors (Gö6976, Gö6983 and GF109203X), only Gö6976 significantly attenuated the phosphorylation of p38 MAPK by arachidonic acid. In addition, phosphorylation of p38 MAPK by arachidonic acid was greatly attenuated by the phospholipase C inhibitor (U73122) and the Ca2+ chelator BAPTA ((1,2-bis-o-amino-phenoxy)-ethane-N,N,N′,N′-tetraacetic acid), but not altered by the nitric oxide synthase inhibitor, N-nitro-l-arginine methyl ester. Arachidonic acid did not cause an increase in cellular cyclic GMP level.

This study revealed the involvement of pertussis toxin-sensitive G protein, non-receptor tyrosine kinase, phospholipase C/Ca2+, and probably Ca2+-dependent protein kinase C in arachidonic acid-stimulated p38 MAPK activation.
Zidovudine, Diclofenac and Ketoprofen Pharmacokinetic Interactions in Rats
Authors: Radwan Mahasen A
Published: October 2002

Zidovudine (AZT) is widely used for the management of human immunodeficiency virus (HIV) infections. Non-steroidal anti-inflammatory drugs (NSAIDs) are often used for relief of non-specific fever and musculoskeletal pain in patients with HIV including those with AZT-induced myopathy. The effects of single oral doses of diclofenac and ketoprofen on AZT pharmacokinetics were studied in rats. The influence of AZT on the pharmacokinetics of diclofenac or ketoprofen was also investigated.

The administration of diclofenac (3 mg kg−1) or ketoprofen (1 mg kg−1) did not significantly alter AZT (1.5 mg kg−1) pharmacokinetic parameters compared with administering AZT alone. There was no significant difference between the pharmacokinetics of ketoprofen given alone or in combination with AZT. However, the co-administration of AZT with diclofenac affected the pharmacokinetics of diclofenac. The Cmax of diclofenac was significantly (P < 0.05) increased by approximately threefold within a shorter time (0.6 ± 0.2 h). The mean AUC value for diclofenac was increased from 2.29 to 5.04 μg mL−1 h in the presence of AZT. AZT decreased the mean apparent clearance of diclofenac by 54%. The increase in diclofenac concentrations could be attributed to a decrease in its clearance or delay in its metabolite formation due to a competitive effect.

The results show that diclofenac and AZT should be given with caution because of the possible increase of diclofenac toxicity, in anticipation of follow-up clinical studies to examine this finding in man. AZT and ketoprofen could be a safe combination since no pharmacokinetic interaction was detected.
Dammarane Derivatives Protect Cultured Rat Cortical Cells from Glutamate-induced Neurotoxicity
Authors: Kim So Ra , Sung Sang Hyun , Kwon Sung Won , Park Jeong Hill , Huh Hoon , Kim Young Choong
Published: October 2002

We previously reported that ginsenosides Rb1 and Rg3, dammarane glycosides, of Panax ginseng C. A. Meyer (Araliaceae), significantly attenuated glutamate-induced neurotoxicity in primary cultures of rat cortical cells. To seek more potent neuroprotective compounds, we attempted to modify the chemical structure of dammarane glycosides and obtained six derivatives, MA-11, PT-11, PT-111, POA-101, POA-111 and N-001. The neuroprotective activity of these dammarane derivatives were evaluated employing primary cultures of rat corticoid cells.

The glutamate-induced neuronal cell damage was significantly reduced by a pre-treatment with protopanaxadiol, MA-11 or PT-11 at concentrations ranging from 100nM to 10 μM. Both MA-11 and PT-11, preserved the levels of catalase and inhibited decreases in glutathione reductase in glutamate-injured cells. Furthermore, the dammarane derivatives reduced the content of intracellular peroxide in glutamate-intoxicated cells. Finally, they inhibited the formation of malondialdehyde, a compound produced during lipid peroxidation, in glutamate-insulted cells.

These results show that the dammarane derivatives, MA-11 and PT-11, exert significant neuroprotective effects on cultured cortical cells by a mechanism seemingly distinct from that afforded by ginsenosides Rb1 and Rg3. As such, the dammarane derivatives may be efficacious in protecting neurons from oxidative damage caused by exposure to excess glutamate.
Effect of Dimethylglycine on Gastric Ulcers in Rats
Authors: Hariganesh K , Prathiba J
Published: October 2002

Dimethylglycine is an anti-stress nutrient with antioxidant properties. Recently, studies have implicated the generation of oxygen-derived free radicals and lipid peroxidation as one of the mechanisms in the pathogenesis of gastric ulcer. Hence, we evaluated the antiulcer activity of dimethylglycine in various rat models of ulcer and also investigated the probable antioxidant mechanism of the anti-ulcer effect.

The results suggest that the gastroprotective effect of dimethylglycine could be mediated by its free radical scavenging activity and cytoprotection of gastric mucosa.

Dimethylglycine at a dose of 25 and 35 mg kg−1 significantly reduced ulcer number, ulcer size and ulcer index in pyloric-ligation-, ibuprofen- and stress-induced ulcers. The 35 mg kg−1 dose was more effective than 25 mg kg−1 and was comparable to famotidine. Dimethylglycine did not produce any significant change in acid secretion, unlike famotidine. There was a significant increase in plasma and tissue malondialdehyde levels in pyloric-ligated rats but these levels fell following dimethylglycine treatment. Also, there was a significant reduction in glutathione levels after dimethylglycine treatment.
Clonidine Attenuates the Carbachol-induced Contractile and Phosphatidylinositol Responses of Rat Trachea
Authors: Shibata Osamu , Saito Masataka , Hashimoto Shigeru , Sakai Kazuyuki , Makita Tetsuji , Sumikawa Koji
Published: October 2002

Although Clonidine is known to affect vascular smooth muscle, its effects on airway smooth muscle are not fully understood. This study was designed to examine the effects of Clonidine on carbachol-induced contractile and phosphatidylinositol responses of rat trachea.

Clonidine, at a dose of 100 μM or greater, attenuated carbachol-induced contraction and the accumulation of carbachol-induced inositol monophosphate (IP1). Clonidine also attenuated the accumulation of aluminium fluoride-induced IP1. The concentration-effect relationship of IP1 accumulation was similar to that of carbachol-induced contraction; r = 0.797, P< 0001.

These results suggest that Clonidine attenuates contractile responses, at least in part, through the inhibition of phospholipase C (coupled with G-proteins) in phosphatidylinositol responses.
AL-3138 Antagonizes FP Prostanoid Receptor-mediated Inositol Phosphates Generation: Comparison with Some Purported FP Antagonists
Authors: Sharif N A , Crider J Y , Davis T L
Published: October 2002

The aim of this study was to pharmacologically characterize the antagonist properties of a novel prostaglandin F2α (PGF2α) analogue (11-deoxy-16-fluoro PGF2α; AL-3138) using a variety of second-messenger assays of prostaglandin receptor subtypes. A detailed comparison was made between AL-3138 and some purported FP receptor antagonists such as PGF2α dimethylamine, PGF2α dimethylamide, glibenclamide and phloretin using the FP receptor-mediated phosphoinositide turnover assay in A7r5 rat thoracic aorta smooth muscle cells and mouse Swiss 3T3 fibroblasts.

The potency and efficacy of AL-3138 as an FP receptor agonist were: EC50 = 72.2 ± 17.9 nM (Emax = 37%) (n = 3) in A7r5 cells and EC50 = 20.5 ± 2.8 nM (Emax = 33%) (n = 5) in 3T3 cells. Being a partial agonist, the antagonist potency of AL-3138 against fluprostenol in A7r5 cells was determined to be: Ki = 296 ± 17 nM (n = 3) and Kb = 182 ± 44nM (n = 5) (-log Kb = 6.79 ± 0.1). AL-3138 exhibited very minimal or no antagonistic effects at EP2, EP4, DP and TP prostaglandin receptors. Both PGF2α dimethylamide and PGF2α dimethylamine were inactive as FP receptor antagonists, whereas phloretin and glibenclamide were very weak and had -log Kb values of 5.28 ± 0.09 (n = 3) and 3.58 ± 0.32 (n = 3), respectively. However, phloretin antagonized functional responses of EP2 and DP prostanoid receptors, and also the V1 — vasopressin receptor. AL-3138 competed for [3H]PGF2α binding to FP receptors with a relatively high affinity (IC50high = 312 ± 95nM) matching its functional antagonist potency.

In conclusion, AL-3138 is a more potent and selective FP receptor antagonist than glibenclamide, phloretin, PGF2α dimethylamide and PGF2α dimethylamine and is therefore a unique and novel pharmacological tool to help characterize FP receptor-mediated functions.
Pharmacokinetics and Bioavailability of Tacrolimus in Rats with Experimental Renal Dysfunction
Authors: Okabe Hiromi , Hashimoto Yukiya , Inui Ken-Ichi
Published: October 2002

The effects of renal failure on the pharmacokinetics and bioavailability of tacrolimus were investigated in rats. Experimental renal dysfunction was induced by intraperitoneal injection of cisplatin (5 mg kg−1) into rats. The blood concentration of tacrolimus was measured after intravenous and intra-intestinal administration of the drug.

The blood concentration of tacrolimus after intravenous administration (1 mg kg−1) was slightly increased (up to 1.3 fold) by induction of renal dysfunction. In contrast, the peak tacrolimus concentration after intra-intestinal administration (1 mg kg−1 or 3 mg kg−1) in rats with renal failure was about 2-fold higher than that in normal controls. The bioavailability was increased by about 35% in rats with impaired renal function as compared with normal controls. These results suggested that the bioavailability of tacrolimus, which is mainly metabolized in the liver and intestine after oral administration, is also influenced by renal function.
The Action of 5-Hydroxytryptamine on Normal and Cystic Fibrosis Mouse Colon: Effects on Secretion and Intracellular Calcium
Authors: Hardcastle J , Hardcastle P T , Klaren P H M , Taylor C J , White S J
Published: October 2002

The ability of mouse colon to generate a secretory response to stimulation by 5-hydroxytryptamine (5-HT) was investigated in intact colonic sheets mounted in Ussing chambers. A preparation of intact isolated crypts was used to determine whether 5-HT action was associated with an elevation of cytosolic calcium levels, measured using the calcium-sensitive fluorescent dye, fura-2.

5-HT increased the short-circuit current, an effect that was inhibited by 55% in the absence of chloride and by 83% in the presence of serosal frusemide, consistent with the stimulation of electrogenic chloride secretion. This was confirmed by the observation that colonic tissue from transgenic cystic fibrosis mice (n = 4) failed to respond to 5-HT, although wild-type tissues generated an increased short-circuit current of 52.4 ± 11.1 μA cm−2 (n = 9). The electrical response to 5-HT was calcium-dependent. 5-HT action was unaffected by tetrodotoxin and was not mimicked by the 5-HT3 agonist 1-phenylbiguanide, indicating that neural mechanisms are not involved. The cyclooxygenase inhibitor indomethacin, however, reduced the 5-HT-induced rise in short-circuit current by 73%, suggesting that prostaglandin production contributes to the response. Stimulation of crypts with acetylcholine elicited an increase in cytosolic calcium levels, but no such response was detected on application of 5-HT (10−6 to 10−4 M), suggesting that 5-HT does not directly modulate intracellular calcium in colonic crypt cells.

It is concluded that mouse colon responds to 5-HT challenge with a stimulation of electrogenic chloride secretion and that this effect is mediated by indirect mechanisms that might involve immune elements within the colonic wall.
The Long-lasting Effect of TU-199, a Novel H+,K+-ATPase Inhibitor, on Gastric Acid Secretion in Dogs
Authors: Uchiyama Kazuyuki , Wakatsuki Daisuke , Kakinoki Bunpei , Takeuchi Yoshishige , Araki Tsutomu , Morinaka Yasuhiro
Published: October 2002

We have used Heidenhain-pouch dogs to investigate the effects of (±)-5-methoxy-2-{[(4-methoxy-3,5-dimethylpyrid-2-yl)methyl]sulphinyl}-1H-imidazo[4,5-b]pyridine (TU-199), an imidazopyridine derivative, on gastric acid secretion stimulated by histamine, carbachol and tetragastrin. We have also investigated the duration of the antisecretory effect of TU-199 using a measurement of intragastric pH for 24 h in gastric fistula dogs whose gastric acid secretion was stimulated by histamine.

Single oral administration of TU-199 (0.1, 0.2 and 0.4 mg kg−1) dose-dependently suppressed gastric acid secretion stimulated by histamine infusion. Oral treatment with TU-199 (0.2, 0.4 and 0.8 mg kg−1) also dose-dependently inhibited acid secretion induced by carbachol and tetragastrin. The inhibitory effect of TU-199 on stimulated gastric acid secretion was more potent than that of omeprazole, a well-known H+,K+-ATPase inhibitor in dogs. Repeated oral treatment with TU-199 at a dose of 0.2 mg kg−1 once a day for seven days markedly suppressed histamine-stimulated gastric acid secretion in dogs. This inhibitory effect of TU-199 reached a maximum level after three or four doses and was more pronounced than that of omeprazole or lansoprazole. In gastric fistula dogs, the duration of intragastric pH-elevation by administration of TU-199 (0.3 mg kg−1) was much longer than that of omeprazole (0.6 mg kg−1) or lansoprazole (0.9 mg kg−1). The IC50 values (doses resulting in 50% inhibition) of TU-199, omeprazole and lansoprazole with regard to H+,K+-ATPase activity in dog gastric mucosal microsomes were 8.6, 8.8 and 9.9 μM, respectively.

These results indicate that TU-199 inhibits gastric acid secretion via suppression of a H+,K+-ATPase activity. Our findings also suggest that TU-199 might have potent and long-lasting effects on gastric acid secretion.
In-vivo and In-vitro Metabolic Clearance of Midazolam, a Cytochrome P450 3A Substrate, by the Liver under Normal and Increased Enzyme Activity in Rats
Authors: Higashikawa Fumiko , Murakami Teruo , Kaneda Tatsuya , Takano Mikihisa
Published: October 2002

The metabolic clearance of midazolam, a cytochrome P450 (CYP) 3A substrate, by the liver under normal and increased enzyme activity in rats was determined in-vivo and in-vitro to elucidate the reproducibility of the in-vivo hepatic extraction ratio of midazolam from the in-vitro study. The hepatic enzyme activity was modified by pretreating rats with a CYP inducer such as dexamethasone and clotrimazole.

The in-vivo hepatic extraction ratio (ERh,obs) of midazolam under a steady-state plasma concentration (approx. 3 nmol mL−1) in untreated (control) rats was 0.864. This value increased to 0.984 in dexamethasone-pretreated rats and to 0.964 in clotrimazole-pretreated rats. The in-vitro hepatic intrinsic clearance (CLint,in-vitro), expressed as mL min−1 (mg microsomal protein)−1, of midazolam was estimated as Vmax (Km)−1 by in-vitro metabolism studies using liver microsomes. The CLint,in-vitro value was converted to the CLint,cal value, expressed as mL min−1 kg−1, by considering the microsomal protein content (g liver)−1 and the microsomal protein content (g liver)−1 kg−1. The estimated CLint,cal value was then converted to the ERh value (ERh,cal) according to the well-stirred, the parallel-tube and the dispersion models. The ERh,cal values obtained by the parallel-tube model were in good agreement with corresponding in-vivo ERh,obs values.

In conclusion, it was demonstrated that high hepatic clearances of midazolam under normal and increased CYP3A activity were reasonably predicted from in-vitro metabolism studies using liver microsomes.
Characteristics of L-Carnitine Transport in Cultured Human Hepatoma HLF Cells
Authors: Yokogawa Koichi , Miya Kazuhiro , Tamai Ikumi , Higashi Yasuhiko , Nomura Masaaki , Miyamoto Ken-Ichi , Tsuji Akira
Published: October 2002

The recently cloned organic cation transporter, OCTN2, isolated as a homologue of OCTN1, has been shown to be of physiological importance in the renal tubular re-absorption of filtered L-carnitine as a high-affinity Na+ carnitine transporter in man. Although the mutation of the OCTN2 gene has been proved to be directly related to primary carnitine deficiency, there is little information about the L-carnitine transport system in the liver. In this study, the characteristics of L-carnitine transport into hepatocytes were studied by use of cultured human hepatoma HLF cells, which expressed OCTN2 mRNA to a greater extent than OCTN1 mRNA.

The uptake of L-carnitine into HLF cells was saturable and the Eadie—Hofstee plot showed two distinct components. The apparent Michaelis constant and the maximum transport rate were 6.59± 1.85 (mean ± s.d.) and 78.5 ± 21.4 pmol/5 min/106 cells, respectively, for high-affinity uptake, and 590 ± 134 μM and 1507 ± 142 pmol/5 min/106 cells, respectively, for low-affinity uptake. The high affinity L-carnitine transporter was significantly inhibited by metabolic inhibitors (sodium azide, dinitrophenol, iodoacetic acid) and at low temperature (4°C). Uptake of [3H]L-carnitine also required the presence of Na+ ions in the external medium. The uptake activity was highest at pH 7.4, and was significantly lower at acidic or basic pH. L-Carnitine analogues (D-carnitine, L-acetylcarnitine and γ-butyrobetaine) strongly inhibited uptake of [3H] L-carnitine, whereas β-alanine, glycine, choline, acetylcholine and an organic anion and cation had little or no inhibitory effect.

In conclusion, L-carnitine is absorbed by hepatocytes from man by an active carrier-mediated transport system which is Na+-, energy- and pH-dependent and has properties very similar to those of the carnitine transporter OCTN2.
Protamine Induces Elevation of Cytosolic Free Ca2+ in Cultured Porcine Aortic Endothelial Cells
Authors: Sato Nobuyoshi , Az-Ma Toshiharu , Fujii Kohyu , Yuge Osafumi
Published: October 2002

To test the hypothesis that protamine influences calcium movement in endothelial cells, we measured the concentration of intracellular free calcium ([Ca2+]i) in cultured porcine aortic endothelial (PAE) cells in Krebs solution (2-5mM Ca2+, pH7.4) at 37°C, by fura-2 fluorimetry.

The basal [Ca2+]i of PAE cells was 113 ± 18 nM (n − 6). Protamine increased [Ca2+]iin a concentration-dependent manner (EC50, the concentration having 50% of the maximum effect, 1.4 ± 0.3 μg mL−1, n = 6). The response of PAE cells to 100 μg mL−1 protamine (330 ± 80 nM, n = 6) was blocked by a Ca2+ chelator, 5 mM glycoletherdiaminetetraacetic acid (EGTA; 131 ± 16 nM, n = 6), and by a non-selective Ca2+ channel blocker, 3 mM Co2+ (134 ± 14 nM, n = 6).

These results suggest that Ca2+ influx through cell-membrane Ca2+ channels is mainly responsible for the protamine-induced Ca2+ elevation.
Pharmacokinetics of a New Proton-pump Inhibitor, YJA-20379-8, after Intravenous and Oral Administration to Rats with Streptozotocin-induced Diabetes Mellitus
Authors: Chung Su Y , Han Kye S , Shon Sang K , Chang Man S , Lee Myung G
Published: October 2002

Because physiological changes occurring in diabetes mellitus patients could alter the pharmacokinetics of the drugs used to treat the disease, the pharmacokinetics of a new proton pump inhibitor, YJA-20379-8, were investigated after intravenous and oral administration of the drug (50 mg kg−1) to control rats and to rats with streptozotocin-induced diabetes mellitus (SIDM).

After intravenous administration of YJA-20379-8 to SIDM rats, plasma concentrations of the drug were significantly higher and this resulted in a significantly greater AUC (area under the concentration-time curve; 2520 ± 366 compared with 1870 ± 272 min mL−1). This was because of significantly slower clearance (CL; 19.5 ± 2.88 compared with 27.2 ± 3.93 mL min−1 kg−1) in SIDM rats. The significantly slower metabolism of YJA-20379-8 in SIDM rats was confirmed by an in-vitro tissue metabolism study; the amounts of YJA-20379-8 remaining in the liver (27.1 ± 5.19 compared with 18.9 ± 8.24 μg (g tissue)−1) were significantly greater after 30-min incubation of the drug (50 μg) with supernatant fractions obtained from the tissues by centrifugation at 9000 g. After oral administration of YJA-20379-8 to SIDM rats the plasma concentrations of the drug were significantly lower and this resulted in significantly smaller AUC (128 ± 31.0 compared with 219 ± 45.6 μg min mL−1). This was because of reduced gastrointestinal absorption of YJA-20379-8 in SIDM rats; the amounts of the oral dose recovered as unchanged drug from the entire gastrointestinal tract after 24 h were significantly greater (32.9 compared with 19.2%) in SIDM rats.

The tissue distribution of YJA-20379-8 was not affected by SIDM.
Relative Blood-Brain Barrier Permeabilities of the Cholecystokinin Receptor Antagonists Devazepide and A-65186 in Rats
Authors: Woltman Todd A , Hulce Martin , Reidelberger Roger D
Published: October 2002

The blood-brain barrier permeabilities of the type-A cholecystokinin receptor antagonists devazepide and A-65186 (Nα-3-quinolinoyl-D-Glu-N,N-dipentylamide) have been compared with those of the reference compounds iodoantipyrine, which readily penetrates the blood-brain barrier, and mannitol, which does not.

Anaesthetized rats received a bolus injection into the left carotid artery of [14C]iodoantipyrine (0.25 μCi) combined with [3H]mannitol, [3H]devazepide or [3H]A-65186 (1 μCi each). Rats were decapitated 12 s after injection and the brains were removed. Four samples of left cerebrum (ca 100 mg each) were solubilized overnight and 14C and 3H activity were measured. The brain-uptake index for each test compound was determined as [(3H/14C for sample)]/[(3H/14C for injectate)] × 100, with a value of 100 representing blood-brain barrier permeability equal to that for iodoantipyrine. The brain-uptake index (mean ± s.e.m.) was 1.6 ± 0.3 for [3H]mannitol (n = 5), 90.6 ± 4.1 for [3H]devazepide (n = 7, P < 0.001 compared with mannitol) and 3.5 ± 0.7 for [3H]A-65186 (n = 4, P > 0.05 compared with mannitol, P < 0.001 compared with devazepide). Thus, devazepide readily penetrated the blood-brain barrier whereas A-65186 did not.

It is concluded that devazepide and A-65186 are likely to be useful pharmacological tools for determining whether cholecystokinin is acting peripherally or at brain sites beyond the blood—brain barrier to produce satiety or any other function mediated by the type A cholecystokinin receptor.
Rabbit Corneal and Conjunctival Permeability of the Novel Aldose Reductase Inhibitors: N-{[4-(Benzoylamino)phenyl] sulphonyl}glycines and N-Benzoyl-N-phenylglycines
Authors: Kompella Uday B , Sunkara Gangadhar , Thomas Erica , Clark C Randall , Deruiter Jack
Published: October 2002

Corneal and conjunctival permeability has been investigated for novel aldose reductase inhibitors (ARIs) of the N{[4-(benzoylamino)phenyl]sulphonyl}glycine (benzoylaminophenylsulphonylglycine) and N-benzoyl-N-phenylglycine (benzoylphenylglycine) series, compounds developed for prevention of cataract formation in diabetic subjects.

Six benzoylaminophenylsulphonylglycines were synthesized with modifications either of the phenyl group or of the glycine structure and three benzoylphenylglycines were synthesized with modification in the phenyl group of the benzoyl moiety. Transport of ARIs in the mucosal to serosal direction was evaluated across rabbit cornea and conjunctiva bathed in glutathione-bicarbonate Ringer's solution maintained at pH 7.4 and 37°C. The permeability coefficients of the novel ARIs across cornea and conjunctiva ranged from 1.87 to 8.95 times 10−6 cms−1 and from 4.6to19.15 times 10−6cms−1, respectively. The ratio of corneal to conjunctival permeability ranged from 0.12 to 0.79. The calculated log partition coefficient (log P) values for the ARIs were in the range 0.84 to 2.78. The log distribution coefficients (log D) were in the range −2.87 to −0.89. There was no apparent relationship between log P or log D and the permeability coefficients of the ARIs for either tissue. Cornea was more resistant to ARI transport than was conjunctiva. Substitution of a phenyl group for hydrogen in the glycine methylene group reduced the permeability coefficient. Permeability coefficients were different for different stereoisomers. Compared with the permeability coefficient of benzoylaminophenylsulphonylglycine, that of 4-fluorobenzoylaminophenylsulphonylglycine was lower in the cornea but similar in the conjunctiva. In both tissues, the permeability coefficient of 2-nitrobenzoylaminophenylsulphonylglycine was less than that of 4-nitrobenzoylaminophenylsulphonyl-glycine. There was no significant difference between the permeability coefficients of 3-nitro- and 4-nitrobenzoylphenylglycines through either tissue and the permeability coefficients of these compounds were greater than that of the more lipophilic 4-methylbenzoylphenylglycine.

The lack of dependence of the permeability coefficients on log P or log D and the different permeabilities of stereoisomers imply the existence of specialized transport processes for the ARIs tested in this study.
Inhibition of Na+ Channel or Na+/H+ Exchanger Attenuate the Hydrogen Peroxide-induced Derangements in Isolated Perfused Rat Heart
Authors: Hara Akiyoshi , Arakawa Johji , Xiao Chun-Yang , Hashizume Hiroko , Ushikubi Fumitaka , Abiko Yasushi
Published: October 2002

The effect of tetrodotoxin, a specific inhibitor of the Na+ channel, and 5-(N,N-dimethyl)-amiloride, a specific inhibitor of the Na+/H+ exchanger, on the mechanical and metabolic derangements induced by hydrogen peroxide (H2O2) was studied in the isolated perfused rat heart. The isolated rat heart was perfused aerobically at a constant flow rate and driven electrically.

H2O2 (600 μM) decreased the left ventricular developed pressure and increased the left ventricular end-diastolic pressure (i.e. mechanical dysfunction), decreased the tissue levels of adenosine triphosphate and adenosine diphosphate (i.e. metabolic derangement), and increased the tissue level of malondialdehyde (i.e. lipid peroxidation). These mechanical and metabolic derangements induced by H2O2 were significantly attenuated by tetrodotoxin (3 μM) or 5-(N,N-dimethyl)-amiloride (15 μM). Neither tetrodotoxin nor 5-(N,N-dimethyl)-amiloride modified the tissue malondialdehyde level, which was increased by H2O2. In the normal (H2O2-untreated) heart, neither tetrodotoxin nor 5-(N,N-dimethyl)-amiloride affected the mechanical function and energy metabolism.

These results suggested that inhibition of the Na+ channel or Na+/H+ exchanger was effective in attenuating the H2O2-induced mechanical dysfunction and metabolic derangements in the isolated perfused rat heart.
Antioxidant Activity of Tannin Components from Vaccinium vitis-idaea L.
Authors: Ho K Y , Huang J S , Tsai C C , Lin T C , Hsu Y F , Lin C C
Published: October 2002

Reactive oxygen molecules have been implicated as important pathological mediators in many clinical disorders and periodontal disease. To provide possible alternative treatment of periodontal disease, six tannins isolated from Vaccinium vitis-idaea L. were evaluated for anti-lipid peroxidation, anti-superoxide formation and free radical scavenging activity.

The results showed that cinnamtannin B1 displayed the strongest anti-lipid peroxidation activity, proanthocyanidin A-1 displayed the strongest superoxide scavenging activity, and epicatechin-(4βr̊ 6)-epicatechin-(4β 8, 2r̊ O r̊ 7)-catechin had the strongest anti-superoxide formation effect.

We conclude that tannins isolated from V. vitis-idaea L. exhibited multiple antioxidant activity, and could be used for the treatment of periodontal disease.