Biomedical subjects

Masahiro Iwaki

Publications and source records attributed to Masahiro Iwaki.

9 recordsLinked to original sources

PubMed2006

Effects of alterations in CAR on bilirubin detoxification in mouse collagen-induced arthritis.

Nuclear receptors such as constitutive androstane receptor (CAR) and pregnane X receptor (PXR) regulate the transcription of cytochromes P450 and transporters. We investigated whether quantitative and functional changes in CAR and PXR could affect bilirubin detoxification in chronic arthritis. The CAR mRNA level was significantly decreased in the liver of mice with collagen-induced arthritis (CIA) compared with control mice. In normal mice treated with CAR agonists, relatively rapid elimination of bilirubin was observed after its intravenous injection. Next, we investigated the effects of CAR on bilirubin-detoxifying enzymes and transporters in arthritis. The mRNA levels of organic anion transporter peptide (OATP) 2, glutathione S-transferase (GST) A1, and GSTA2 were decreased in CIA mice, whereas the mRNA levels of OATP4, UDP-glucuronosyl-transferase 1A1, and multidrug resistance-associated protein 2 remained unchanged. The protein levels and transport activities of OATP2 were also decreased in CIA mice. Furthermore, the CIA mice actually exhibited retarded elimination of bilirubin after its intravenous injection. These results indicate that alterations to CAR during arthritis affect the elimination of bilirubin because of changes in multiple bilirubin-detoxifying enzymes and transporters.

Animals↗

PubMed2005

Nitrogen-containing bisphosphonate, YM529/ONO-5920 (a novel minodronic acid), inhibits RANKL expression in a cultured bone marrow stromal cell line ST2.

Increase in bone resorption by osteoclasts can cause metabolic bone diseases, such as osteoporosis. Recent attention has been paid to the receptor activator of the NF-kappaB ligand (RANKL), an accelerator of osteoclast differentiation. RANKL is expressed on the bone marrow-derived stromal cell membrane and induces the differentiation of osteoclasts by binding to RANK expressed on the osteoclast precursor cell membrane. Since the inhibition of RANKL expression can lead to the inhibition of osteoclastic bone resorption, the clinical application of RANKL inhibition could be expected to have a major effect on metabolic bone disease therapy. In this study, we investigated whether or not YM529/ONO-5920, a nitrogen-containing bisphosphonate (a novel minodronic acid), inhibits RANKL expression in a bone marrow-derived stromal cell line (ST2 cells). Reverse transcription-polymerase chain reaction revealed that the administration of YM529/ONO-5920 to ST2 cells inhibited RANKL mRNA expression and reduced RANKL proteins as assessed by Western blot analysis. The inhibition of RANKL mRNA expression was reversed when geranylgeranyl pyrophosphate (GGPP), an intermediate in the mevalonate pathway, was used in combination. Furthermore, YM529/ONO-5920 reduced phosphorylated extracellular signal-regulated kinase 1/2 (ERK1/2), and similarly, U0126, a mitogen-activated protein kinase kinase 1/2 inhibitor, inhibited RANKL expression. Pretreatment with GGPP reversed the YM529/ONO-5920-induced decrease in phosphorylation of ERK. Furthermore, YM529/ONO-5920 decreased TRAP-positive cells in co-culture of ST2 cells and an osteoclast cell line, C7 cells, and this decrease was inhibited by pretreatment with GGPP. This indicates that YM529/ONO-5920 inhibits GGPP biosynthesis in the mevalonate pathway and then signal transduction in the Ras-mitogen-activated protein kinase pathway, thereby inhibiting RANKL expression on ST2 cells. These results suggest a newly elucidated action of bisphosphonates in the inhibition of bone resorption.

Animals↗

PubMed2005

Nonlinear pharmacokinetics of propafenone in rats and humans: application of a substrate depletion assay using hepatocytes for assessment of nonlinearity.

Linear pharmacokinetic profiles of propafenone in female Wistar rats were found after oral administration of up to 20 mg/kg. These profiles differed from nonlinear pharmacokinetics in a dose-dependent manner with increasing plasma concentrations in humans (Hollmann M, Brode E, Hotz D, Kaumeier S, and Kehrhahn OH (1983) Arzneim-Forsch 33:763-770). We investigated the species differences in pharmacokinetics of propafenone between rats and humans. In rats, after intravenous administration, clearance was constant at all doses examined (0.2-10 mg/kg), whereas the distribution volume at a steady state increased and the resultant elimination half-life was prolonged with increasing doses. In a substrate depletion assay without plasma, rat and human hepatocytes showed a concentration-dependent elimination of propafenone with low Km values (<0.4 microM). However, in the depletion assay with plasma incubation, the profiles were altered to a concentration-independent profile in rat but not human hepatocytes. The differing effect of adding plasma in rat and human hepatocytes can be explained by species differences in plasma binding (unbound fraction, 0.0071 versus 0.0754 for rats and humans, respectively, at 0.1 microg/ml). In rat plasma, the unbound fraction increased with concentrations of 0.1 to 1.0 microg/ml, whereas it was constant in human plasma. Accordingly, the in vivo nonlinear disposition in humans can be ascribed to the saturation of hepatic metabolism due to the low Km values. In contrast, the influence of saturable metabolism is canceled out with nonlinear plasma binding in rats leading to the apparent linear pharmacokinetic behavior. The newly developed depletion assay with plasma incubation gave insights into the nonlinear pharmacokinetics of propafenone.

Animals↗

PubMed2005

Pharmacokinetics and metabolism of metoprolol and propranolol in the female DA and female Wistar rat: the female DA rat is not always an animal model for poor metabolizers of CYP2D6.

The purpose of this study was to clarify the pharmacokinetics of CYP2D6 substrates in female DA and Wistar rats, which are regarded as animal models of poor metabolizers and extensive metabolizers, respectively. In vivo pharmacokinetic and in vitro metabolic studies were conducted using metoprolol and propranolol, which show substantial and marginal polymorphisms in humans, respectively. After oral administration, the areas under the plasma concentration curves (AUC) for metoprolol and propranolol in DA rats were ca. 5- and 35-fold higher, respectively, than those in Wistar rats. There were no strain differences for serum protein binding or metabolism inhibition by quinine between the two compounds. Using a substrate depletion assay, the intrinsic clearances estimated for the two strains differed by 7.2-fold for metoprolol and 4.5-fold for propranolol. The discrepancy between the in vitro and in vivo profiles observed for propranolol, but not metoprolol, would be due to nonlinearity between the normalized AUC and the oral doses in DA rats, being associated with lower K(m) values. The larger strain difference in the AUCs of propranolol was proved by the in vitro kinetic parameters, implying that DA rats do not always reflect the polymorphic profiles in humans.

Animals↗

PubMed2005

Application of substrate depletion assay for early prediction of nonlinear pharmacokinetics in drug discovery: assessment of nonlinearity of metoprolol, timolol, and propranolol.

The purpose of this study was to investigate the advantages of the substrate depletion assay for evaluating linearity of pharmacokinetics compared with the metabolite formation assay. For propranolol, metoprolol, and nisoldipine with multiple and/or sequential metabolisms, the Michaelis constant (Km) and maximum metabolic intrinsic clearance obtained from the depletion assay using rat and human liver microsomes showed a good correlation with relevant parameters with the formation assay. In vitro kinetics and in vivo pharmacokinetic profiles after oral administration of timolol, metoprolol, and propranolol, were investigated in rats using the depletion assay. The same rank order was found between nonlinearities based on dose-normalized areas under the plasma concentration curve (AUC/Dose) and Km values. Using the kinetic parameters of these compounds, AUC was predicted based on a physiological based pharmacokinetic model incorporated saturable metabolism. The AUCs predicted for propranolol and metoprolol had a good relationship with those observed in the in vivo studies, implying that the depletion assay could be useful for assessing linearity of pharmacokinetics.

Adrenergic beta-Antagonists↗

PubMed2005

Mevastatin induces apoptosis in HL60 cells dependently on decrease in phosphorylated ERK.

Mevastatin which is an inhibitor of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, the rate-limiting enzyme in cholesterol synthesis, suppress cell proliferation and induce apoptosis. However, the molecular mechanism of apoptosis induction is not well understood. So, in the present study, we attempted to clarify the mechanism by which mevastatin induces apoptosis in HL60 cells. It was found that mevastatin induced apoptosis. At that time, we observed an increase in caspase-3 activity and morphological fragmentation of the nuclei. The apoptosis induced by mevastatin was not inhibited by the addition of farnesyl pyrophosphate (FPP), squalene, ubiquinone, and isopentenyladenine, but was inhibited by the addition of geranylgeranyl pyrophosphate (GGPP). When we examined the survival signals at the time of apoptotic induction, we also observed that the administration of mevastatin had caused a remarkable decrease in the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2). However, other survival signals, such as nuclear factor kappa B (NF-kappaB), protein kinase B (Akt), and p38 mitogen-activated protein kinase (p38), exhibited no change. In addition, no quantitative change was observed in Bcl-2, which was an anti-apoptosis protein. It was also observed that apoptosis was induced when U0126, an MEK inhibitor, was added to the cells to inhibit ERK. These results suggested that mevastatin induced apoptosis when it inhibited GGPP biosynthesis and consequently decreased the level of phosphorylated ERK, which was a survival signal; moreover, at that time, there was no influence on NF-kappaB, Akt, p38, and Bcl-2. The results of this study also suggested that mevastatin could be used as an anticancer agent.

Antineoplastic Agents↗

PubMed2004

Potent and metabolically stable agonists for protease-activated receptor-2: evaluation of activity in multiple assay systems in vitro and in vivo.

To develop potent and metabolically stable agonists for protease-activated receptor-2 (PAR-2), we prepared 2-furoylated (2f) derivatives of native PAR-2-activating peptides, 2f-LIGKV-OH, 2f-LIGRL-OH, 2f-LIGKV-NH(2), and 2f-LIGRL-NH(2), and systematically evaluated their activity in PAR-2-responsive cell lines and tissues. In both HCT-15 cells and NCTC2544 cells overexpressing PAR-2, all furoylated peptides increased cytosolic Ca(2+) levels with a greater potency than the corresponding native peptides, although a similar maximum response was recorded. The absolute potency of each peptide was greater in NCTC2544, possibly due to a higher level of receptor expression. Furthermore, the difference in potency between the 2-furoylated peptides and the native peptides was enhanced when evaluated in the rat superior mesenteric artery and further increased when measuring PAR-2-mediated salivation in ddY mice in vivo. The potency of 2f-LIGRL-NH(2), the most powerful peptide, relative to SLIGKV-OH, was about 100 in the cultured cell Ca(2+) signaling assays, 517 in the vasorelaxation assay, and 1100 in the salivation assay. Amastatin, an aminopeptidase inhibitor, augmented salivation caused by native peptides, but not furoylated peptides. The PAR-2-activating peptides, including the furoylated derivatives, also produced salivation in the wild-type C57BL/6 mice, but not the PAR-2-deficient mice. Our data thus demonstrate that substitution of the N-terminal serine with a furoyl group in native PAR-2-activating peptides dramatically enhances the agonistic activity and decreases degradation by aminopeptidase, leading to development of 2f-LIGRL-NH(2), the most potent peptide. Furthermore, the data from PAR-2-deficient mice provide ultimate evidence for involvement of PAR-2 in salivation and the selective nature of the 2-furoylated peptides.

Aminopeptidases↗

PubMed2003

Stereoselective pharmacokinetics of flurbiprofen and formation of covalent adducts with plasma protein in adjuvant-induced arthritic rats.

To determine the effect of arthritis on the disposition of flurbiprofen (FP) and its acyl glucuronide (FPG) as well as formation of covalent adducts with plasma protein, a pharmacokinetic study was carried out in adjuvant-induced arthritic (AA) rats. In control animals the pharmacokinetics of FP were stereoselective following intravenous bolus injection of rac-FP: (-)-(R)-FP showed higher plasma clearance (CL(tot)) and shorter mean residence time (MRT) compared to (+)-(S)-FP. The CL(tot) and clearance for the glucuronide formation (CL(glu)) of both enantiomers in AA rats were extremely increased compared to those in control rats. Increased total clearance in AA rats was due, at least in part, to a remarkable increase in the plasma unbound fraction of FP, consistent with a decrease in the plasma albumin level. The yield of covalent binding of FP to plasma protein in AA rats was less than that in controls, being consistent with the decrease in the plasma acyl glucuronide level.

Animals↗

PubMed2002

Transfollicular drug delivery: penetration of drugs through human scalp skin and comparison of penetration between scalp and abdominal skins in vitro.

In order to quantitatively investigate the importance of transfollicular pathway for drug delivery, drug penetration through human scalp skin was investigated using liquid formulations containing lipophilic and hydrophilic drugs in vitro. The penetration pathway for drugs through the scalp skin was examined using fluorescent probes. Additionally, the drug penetration through the scalp skin was compared with that via human abdominal skin to clarify the usefulness of intrafollicular delivery. Lipophilic melatonin (MT) and ketoprofen (KP) showed high permeabilities through the scalp skin, although the flux of KP was much higher. Absorption enhancers, N-methyl-2-pyrrolidone and isopropylmyristate, only slightly increased the fluxes. Hydrophilic fluorouracil (5FU) and acyclovir (ACV) penetrated through the scalp skin with relatively large fluxes. However, there was large variability in the fluxes of these drugs across scalp skin from different sources. When the relationship between the flux and hair follicle density was estimated, there was good correlation between the two (r = 0.651 for MT and r = 0.666 for ACV, P < 0.05). The histologic examination of the scalp skin, following application of the formulation with nile red or sodium fluorescein, indicated that the probes permeated into the junction of the internal and external root sheath of follicles and diffused into the dermis via the outer root sheath at the initial times. The penetration of nile red, a lipophilic probe, via the stratum corneum of scalp skin was later than that via the follicles. The permeation of MT and 5FU through the scalp skin was much higher than that via the abdominal skin, being 27 and 48 times as high as the abdominal skin, respectively. These results indicate that the drug delivery through the scalp skin will offer an available delivery means for drugs, particularly for hydrophilic drugs.

Abdomen↗