Details of a Researcher - Tomi, Masatoshi

Papers - Tomi, Masatoshi

Division display 41 - 60 of about 86 ／ All the affair displays >>

Hypoxic adaptation engages the CBP/CREST-induced coactivator complex of Creb-HIF-1α in transactivating murine neuroblastic glucose transporter

Thamotharan S., Raychaudhuri N., Tomi M., Shin B., Devaskar S.

American Journal of Physiology - Endocrinology and Metabolism (American Journal of Physiology - Endocrinology and Metabolism) 304 ( 6 ) E583 - E598 2013.03

Research paper (scientific journal), Accepted, ISSN 0193-1849

We have shown in vitro a hypoxia-induced time-dependent increase in facilitative glucose transporter isoform 3 (GLUT3) expression in N2A murine neuroblasts. This increase in GLUT3 expression is partially reliant on a transcriptional increase noted in actinomycin D and cycloheximide pretreatment experiments. Transient transfection assays in N2A neuroblasts using murine glut3-luciferase reporter constructs mapped the hypoxia-induced enhancer activities to -857- to -573-bp and -203- to -177-bp regions. Hypoxia-exposed N2A nuclear extracts demonstrated an increase in HIF-1α and p-Creb binding to HRE (-828 to -824 bp) and AP-1 (-187 to -180 bp) cis-elements, respectively, in electromobility shift and supershift assays, which was confirmed by chromatin immunoprecipitation assays. In addition, the interaction of CBP with Creb and HIF-1α and CREST with CBP in hypoxia was detected by coimmunoprecipitation. Furthermore, small interference (si)RNA targeting Creb in these cells decreased endogenous Creb concentrations that reduced by twofold hypoxia-induced glut3 gene transcription. Thus, in N2A neuroblasts, phosphorylated HIF-1α and Creb mediated the hypoxia-induced increase in glut3 transcription. Coactivation by the Ca⁺⁺-dependent CREST and CBP proteins may enhance cross-talk between p-Creb-AP-1 and HIF-1α/HRE of the glut3 gene. Collectively, these processes can facilitate an adaptive response to hypoxic energy depletion targeted at enhancing glucose transport and minimizing injury while fueling the proliferative potential of neuroblasts. © 2013 the American Physiological Society.

Access to Document (DOI)

Early life nutrient restriction impairs blood-brain metabolic profile and neurobehavior predisposing to Alzheimer's disease with aging

Tomi M., Zhao Y., Thamotharan S., Shin B.C., Devaskar S.U.

Brain Research (Brain Research) 1495 61 - 75 2013.02

Research paper (scientific journal), Accepted, ISSN 0006-8993

Prenatal nutrient restriction (NR) culminating in intra-uterine growth restriction (IUGR) with postnatal catch up growth leads to diabesity. In contrast, postnatal NR with growth restriction (PNGR) superimposed on IUGR (IPGR) protects young and aging adults from this phenotype. We hypothesized that PNGR/IPGR will compromise the blood-brain metabolic profile impairing neurobehavior and predisposing to Alzheimer's disease (AD). NR (50%) in late gestation followed by cross-fostering of rat pups to either ad lib fed (CON) or NR (50%) lactating mothers generated CON, IUGR, PNGR and IPGR male (M) and female (F) offspring that were examined through the life span. In PNGR/IPGR plasma/CSF glucose and lactate decreased while ketones increased in (M) and (F) (PN21, PN50). In addition increased brain glucose transporters, Glut1 & Glut3, greater brain derived neurotrophic factor (BDNF), reduced Glut4, with unchanged serotonin transporter concentrations were noted in (F) (PN50-60). While (F) displayed more hyperactivity, both (F) and (M) exhibited anxiety although socially and cognitively unimpaired (PN25-28&50). Aging (15-17 m) (F) not (M), expressed low plasma insulin, reduced brain IRS-2, pAkt, and pGSK-3β^Ser9, unchanged pPDK1, pTau or lipoprotein receptor related protein 1 (LRP1), higher glial fibrillary acidic protein (GFAP) and spinophilin but a 10-fold increased amyloid-β42. We conclude that therapeutically superimposing PNGR on IUGR (IPGR) should be carefully weighed in light of unintended consequences related to perturbed neurobehavior and potential predilection for AD. © 2012 Elsevier B.V.

Access to Document (DOI)

Mechanism of Nucleoside Uptake in Rat Placenta and Induction of Placental CNT2 in Experimental Diabetes

NISHIMURA Tomohiro, CHISHU Takuya, TOMI Masatoshi, NAKAMURA Ryo, SATO Kazuko, KOSE Noriko, SAI Yoshimichi, NAKASHIMA Emi

Drug Metabolism and Pharmacokinetics (The Japanese Society for the Study of Xenobiotics) 27 ( 4 ) 439 - 446 2012.08

Research paper (scientific journal), Accepted, ISSN 1347-4367

The purpose of this study was to clarify the transport characteristics of nucleosides in rat placenta and the changes of functional expression of nucleoside transporters in rat placenta with experimental diabetes mellitus. Placental uptake clearances of [³H]adenosine and [³H]zidovudine from maternal blood was much higher than that of [¹⁴C]mannitol. Xenopus oocytes injected with rat ENT1 and ENT2 cRNA took up [³H]adenosine with K_m values of 6.1 and 26 μM, respectively. [³H]Adenosine transport by rat placental brush-border membrane vesicles (BBMV) was saturable and was inhibited by nitrobenzylthioinosine (NBMPR), a specific ENT inhibitor, in a manner consistent with involvement of both rat ENT1 and ENT2. [³H]Didanosine was modestly taken up by placenta, and the inhibitory effect of 100 μM NBMPR on [³H]ddI uptake by BBMV suggested a role of ENT2-mediated transport. Expression of ENT1, ENT2, ENT3, CNT2, and CNT3 mRNAs was detected in placenta of control and streptozotocin (STZ)- induced diabetic pregnant rats, and CNT2 (SLC28A2) expression was significantly increased in STZinduced diabetic rats. Consistently, Na⁺-dependent adenosine uptake by BBMV from STZ-induced diabetic pregnant rats was higher than that from control rats. These results suggest the involvement of placental ENT2 as well as ENT1 in nucleoside uptake from maternal blood, and the induction of CNT2 in experimental diabetes mellitus. © 2012 by the Japanese Society for the Study of Xenobiotics (JSSX).

Up-regulation of L-Type Amino Acid Transporter 1 (LAT1) in Cultured Rat Retinal Capillary Endothelial Cells in Response to Glucose Deprivation

MATSUYAMA Ryo, TOMI Masatoshi, AKANUMA Shin-ichi, TABUCHI Akiko, KUBO Yoshiyuki, TACHIKAWA Masanori, HOSOYA Ken-ichi

Drug Metabolism and Pharmacokinetics (The Japanese Society for the Study of Xenobiotics) 27 ( 3 ) 317 - 324 2012.06

Research paper (scientific journal), Accepted, ISSN 1347-4367

We investigated the regulation of L-type amino acid transporter 1 (LAT1) in a conditionally immortalized rat retinal capillary endothelial cell line (TR-iBRB2 cells) in response to glucose deprivation. The amounts of LAT1 and 4F2 heavy chain (4F2hc) mRNA in TR-iBRB2 cells exposed to glucose-free culture medium for 8 to 24 h were significantly elevated compared with those in control medium. Concomitantly, [³H]L-leucine uptake activity was increased, suggesting that LAT1 transport activity is induced under glucose-deprivation. To determine the transcriptional activity of the LAT1 gene under glucose-free conditions, the promoter activity of the LAT1 gene of approximately 2 kbp (-1958 bp to +70 bp) in TR-iBRB2 cells was assayed using a dual-luciferase reporter assay system. The transcriptional activity of the 2 kbp LAT1 promoter under the glucose-free conditions was 1.7-fold greater than that under normal glucose conditions. The presence of an activator site(s) between -162 bp and -155 bp was indicated by the low activities exhibited by the construct spanning this region and mutagenesis. These results suggest that the glucose deprivation sensitivity of LAT1 expression is transcriptionally regulated, and cis-elements within the LAT1 promoter region from -162 bp to -155 bp mediate this regulation. © 2012 by the Japanese Society for the Study of Xenobiotics (JSSX).

Effectiveness of Aromatherapy Evaluated with Subjective and Objective Cognitive Indicators: A Clinical Trial on Young Adults Using Near-infrared Spectroscopy

Shinno Akemi, Isawa Minae, Itoh Hiroshi, Kose Noriko, Ishino Haruyuki, Nishimura Tomohiro, Tomi Masatoshi, Tsujii Takeo, Shimada Hideyo, Saito Hidetsugu, Nakashima Emi

Japanese Journal of Pharmaceutical Health Care and Sciences (Japanese Society of Pharmaceutical Health Care and Sciences) 38 ( 4 ) 265 - 271 2012.04

Research paper (scientific journal), Joint Work, Accepted, ISSN 1346-342X

The purpose of this research was to noninvasively evaluate the influence of aromatherapy on work performance in 14 healthy young adults. Participants in a room filled with the fragrance of lavender completed a self-reported questionnaire and multi-dimensional fatigue inventory-20 (MFI-20). Further, as an objective measure, blood flow in the inferior frontal cortex was evaluated via near-infrared spectroscopy (NIRS) as a parameter of working memory capacity. Compared to the control stage (no aromatherapy), exposure to aromatherapy achieved a significant reduction in general and mental fatigue according to MFI-20. Self-reported questionnaires also indicated improvement, but the differences were not statistically significant. NIRS measurement during the task performance of an N-back program indicated that regional blood flow in the inferior frontal cortex was significantly increased through exposure to aromatherapy, compared to without. The prefrontal area in the brain is involved in working memory, attention concentration and judgment. These results suggest that lavender may improve both cognitive ability and mood. A larger study with more aroma oils seems warranted.

Strategies for therapy of retinal diseases using systemic drug delivery: Relevance of transporters at the blood retinal barrier

Hosoya K., Tomi M., Tachikawa M.

Expert Opinion on Drug Delivery (Expert Opinion on Drug Delivery) 8 ( 12 ) 1571 - 1587 2011.12

Research paper (scientific journal), Joint Work, ISSN 1742-5247

Introduction: There is an increasing need for managing rapidly progressing retinal diseases because of the potential loss of vision. Although systemic drug administration is one possible route for treating retinal diseases, retinal transfer of therapeutic drugs from the circulating blood is strictly regulated by the bloodretinal barrier (BRB). Areas covered: This review discusses the constraints and challenges of drug delivery to the retina. In addition, this article discusses the properties of drugs and the conditions of the BRB that affect drug permeability. The reader will gain insights into the strategies for developing therapeutic drugs that are able to cross the BRB for treating retinal diseases. Further, the reader will gain insights into the role of BRB physiology including barrier functions, and the effect of influx and efflux transporters on retinal drug delivery. Expert opinion: When designing and selecting optimal drug candidates, it's important to consider the fact that they should be recognized by influx transporters and that efflux transporters at the BRB should be avoided. Although lipophilic cationic drugs are known to be transported to the brain across the bloodbrain barrier, verapamil transport to the retina is substantially higher than to the brain. Therefore, lipophilic cationic drugs do have a great ability to increase influx transport across the BRB. © 2011 Informa UK, Ltd.

Access to Document (DOI)

Enhancement of zidovudine transfer to molt-4 cells, a human t-cell model, by dehydroepiandrosterone sulfate

Nishimura T., Tanaka J., Tomi M., Seki Y., Kose N., Sai Y., Nakashima E.

Journal of Pharmaceutical Sciences (Journal of Pharmaceutical Sciences) 100 ( 9 ) 3959 - 3967 2011.09

Research paper (scientific journal), Accepted, ISSN 0022-3549

A possible approach to improve antiretroviral therapy with nucleoside reverse transcriptase inhibitors is to enhance inhibitor delivery to CD4-positive T cells. We previously showed that dehydroepiandrosterone sulfate (DHEAS) enhances zidovudine (AZT) transfer into syncytiotrophoblast. Here, we investigated whether DHEAS also enhances AZT transfer into a cellular model of human T lymphocytes, and whether AZT is taken up by a specific transport system. The effects of DHEAS and related compounds on the uptake of [3H]AZT and other nucleosides by Molt-4 cells (a model of human CD4-positive T cells) were measured. [3H]AZT uptake by Molt-4 cells was nitrobenzylthioinosine insensitive and pH dependent, and the uptake was significantly inhibited by 1 mM ethylisopropylamiloride. [3H]AZT uptake by Molt-4 cells was increased in the presence of DHEAS, whereas uptake of other nucleosides was reduced. Kinetic study revealed that the maximum uptake velocity (up to 30 min) was increased in the presence of DHEAS. The structural requirements for AZT uptake-enhancing activity were studied using structural analogues of DHEAS. Estrone-3-sulfate and 16α-hydroxy DHEAS also enhanced AZT uptake into Molt-4 cells. The use of uptake enhancers may be a good strategy to improve the efficacy of antiretroviral therapy. © 2011 Wiley-Liss, Inc. and the American Pharmacists Association.

Access to Document (DOI)

Mother-to-fetus transfer of antiviral drugs and the involvement of transporters at the placental barrier

Tomi M., Nishimura T., Nakashima E.

Journal of Pharmaceutical Sciences (Journal of Pharmaceutical Sciences) 100 ( 9 ) 3708 - 3718 2011.09

Research paper (scientific journal), Accepted, ISSN 0022-3549

Administration of antiviral drugs to pregnant women with influenza, human immunodeficiency virus (HIV), and herpes simplex virus (HSV) infections is widely accepted as an effective treatment to safeguard the health of the mother and fetus. This review deals with the transfer of antiviral drugs to the fetus across the placental barrier, which is formed by an epithelial layer of syncytiotrophoblasts. First, the structure, function, and developmental change of the placenta and the placental barrier are briefly presented. We then review the transplacental permeability of antiviral drugs, such as oseltamivir for influenza, antiretrovirals (e.g., zidovudine, didanosine, and saquinavir) for HIV, and acyclovir for HSV, focusing on the involvement of ATP-binding cassette, organic anion/cation, and nucleoside transporters. The increasing evidence that is becoming available about transport mechanisms operating at the placental barrier is expected to be useful in the development of techniques to control fetal and placental transfer of antiviral drugs, and thereby to obtain maximum therapeutic benefit while minimizing potential fetal toxicity. © 2011 Wiley-Liss, Inc. and the American Pharmacists Association.

Access to Document (DOI)

Effectiveness of Repeated Administration of Energy Drinks Based on Cognitively Subjective and Objective Indicators

Itoh Hiroshi, Isawa Minae, Nishimura Tomohiro, Tomi Masatoshi, Shimada Hideyo, Mochizuki Mayumi, Nakashima Emi

JAPANESE JOURNAL OF PHARMACEUTICAL HEALTH CARE AND SCIENCES (Japanese Society of Pharmaceutical Health Care and Sciences) 36 ( 12 ) 847 - 854 2010.12

Research paper (scientific journal), Joint Work, Accepted, ISSN 1346-342X

The purpose of this research was to clarify the effect of repeated administration of 2 kinds of energy drink,1 containing vitamins,taurine and caffeine (simple-type),and the other containing vitamins,taurine,caffeine and several botanical herb extracts (herb-type),on work performance using cognitively subjective and objective indicators. The simple-type and herb-type drinks were given to 14 healthy young adults in a consecutive double-blind,randomized,2-period cross-over clinical trial.Participants took each energy drink once a day for five days during separate periods.Motivation,feeling of drowsiness,and general fatigue were evaluated on a subjective scale using a self-reported questionnaire both before and after administration.Other subjective evaluations were those based on the fatigue-visual analog scale (Fatigue-VAS) and multi-dimensional fatigue inventory-20 (MFI-20).As an objective measure,work performance was examined using the Kreapelin mental work test. Compared to the simple-type drink,the herb-type drink achieved a significant reduction in drowsiness and significantly increased motivation based on the self-reported questionnaire results for the post-administration assessment.In the Kreapelin mental work test,the mean amount of work was significantly increased by both drinks.MFI-20 revealed significant decreases in 2 categories of physical fatigue and general fatigue due to the 2 drinks.The fatigue-VAS also indicated decreases but they were not statistically significant. Both types of energy drink appeared to be effective in terms of subjective and objective indicators.Though the herb-type drink was found to be superior to the simple-type drink regarding subjective measures,there was no significant difference between the 2 regarding the objective measure.

Roles of TauT and system A in cytoprotection of rat syncytiotrophoblast cell line exposed to hypertonic stress

Nishimura T., Sai Y., Fujii J., Muta M., Iizasa H., Tomi M., Deureh M., Kose N., Nakashima E.

Placenta (Placenta) 31 ( 11 ) 1003-1009 - 1009 2010.11

Research paper (scientific journal), Joint Work, Accepted, ISSN 0143-4004

The purpose of this study was to clarify the cytoprotective mechanism(s) induced in a conditionally immortalized syncytiotrophoblast cell line (TR-TBT 18d-1) exposed to hypertonic conditions. Hypertonicity-induced apoptosis of TR-TBT 18d-1 cells, but this was blocked by addition of 1 mM taurine to the culture medium. TauT-knockdown using siRNA revealed that TauT is a major contributor to taurine uptake by TR-TBT 18d-1 cells, at least under normal conditions. Cellular uptake of [3H]taurine and [14C] betaine by TR-TBT 18d-1 cells cultured under hypertonic conditions was increased compared to that under normal conditions. TauT, BGT-1, ATA2 and HSP70 mRNAs were upregulated by hypertonicity, while OCTN2, ENT1 and CNT1 mRNAs were downregulated. [3H]Taurine uptake was strongly inhibited by TauT inhibitors such as hypotaurine and β-alanine. MeAIB, a system A specific substrate, inhibited hypertonic stress-induced [14C]betaine uptake. These results suggest that TauT and system A play cytoprotective roles in syncytiotrophoblasts exposed to hypertonic stress. © 2010 Elsevier Ltd. All rights reserved.

Access to Document (DOI)

Proton-coupled erythromycin antiport at rat blood-placenta barrier

Sai Y., Nishimura T., Ochi K., Tanaka N., Takagi A., Tomi M., Kose N., Kobayashi Y., Miyakoshi N., Kitagaki S., Mukai C., Nakashima E.

Drug Metabolism and Disposition (Drug Metabolism and Disposition) 38 ( 9 ) 1576 - 1581 2010.09

Research paper (scientific journal), Accepted, ISSN 0090-9556

The aim of the present study was to characterize the mechanism of erythromycin transport at the blood-placenta barrier, using TR-TBT 18d-1 cells as a model of rat syncytiotrophoblasts. [14C]Erythromycin was taken up by TR-TBT 18d-1 cells with a Michaelis constant of 466 μM. Although the uptake was not dependent on extracellular Na+ or Cl-, it was increased at weakly alkaline pH. Significant overshoot of [ 14C]erythromycin uptake by placental brush-border membrane vesicles was observed in the presence of an outwardly directed proton gradient. These results indicate that erythromycin is transferred by the H+-coupled transport system in syncytiotrophoblasts. To address the physiological transport of erythromycin in rat placenta, fetal-to-maternal transport clearance was estimated by means of the single placental perfusion technique. Clearance of [14C]erythromycin was higher than that of [14C]inulin, a paracellular pathway marker, and was decreased by the addition of 5 mM erythromycin, indicating that saturable efflux system from fetus to mother is involved. The effect of various transporter inhibitors on [14C] erythromycin efflux from TR-TBT 18d-1 cells was evaluated. cyclosporin A, fumitremorgin C, and probenecid had no effect, whereas ethylisopropylamiloride, a specific inhibitor of Na+/H+ exchangers (NHEs), was significantly inhibitory. These results suggest that erythromycin efflux transport at the rat blood-placenta barrier is mediated by an erythromycin/H+ antiport system, driven by H+ supplied by NHEs. Copyright © 2010 by The American Society for Pharmacology and Experimental Therapeutics.

Access to Document (DOI)

The role of blood-ocular barrier transporters in retinal drug disposition: An overview

TOMI Masatoshi

Expert Opinion on Drug Metabolism and Toxicology (Expert Opinion on Drug Metabolism and Toxicology) 6 ( 9 ) 1111 - 1124 2010.09

Research paper (scientific journal), Accepted, ISSN 1742-5255

Importance of the field: Delivery of drugs to the retina remains a major challenge which needs to be addressed urgently because retinal disorders are leading causes of visual impairment and significantly affect a patient's quality of life. Systemic drug administration is one possible route for treating retinal disorders; however, retinal transfer of drugs from the circulating blood is strictly regulated by two bloodocular barrier systems, the bloodaqueous barrier and the bloodretinal barrier. Areas covered in this review: This review summarizes the latest biological research regarding bloodocular barrier drug transporters. What the reader will gain: The bloodocular barrier sites and their respective roles in aqueous humor dynamics and retinal homeostasis are briefly presented. The potential impact of ATP-binding cassette (ABC) and solute carrier (SLC) drug transporters, such as ABCB, ABCC, ABCG, SLC7, SLC16, SLC19, SLCO/SLC21A, SLC22A and SLC29 transporters, on the permeability of drugs across the bloodocular barriers is then illustrated. Take home message: As more information becomes available regarding the bloodocular barrier transporters, we may be able to design simpler and more effective routes for drug delivery to the retina and, consequently, improve the treatment of retinal diseases. © 2010 Informa UK, Ltd.

Involvement of system A in the retina-to-blood transport of l-proline across the inner blood-retinal barrier

Yoneyama D., Shinozaki Y., Lu W., Tomi M., Tachikawa M., Hosoya K.

Experimental Eye Research (Experimental Eye Research) 90 ( 4 ) 507 - 513 2010.04

Research paper (scientific journal), Accepted, ISSN 0014-4835

The purpose of the present study was to elucidate the mechanisms of retina-to-blood transport of l-proline across the blood-retinal barrier (BRB) in vivo and in vitro, and to identify the responsible transporter(s). The vitreous humor/retina-to-blood transport of [3H]l-proline across the BRB was evaluated by microdialysis. Transport mechanisms of [3H]l-proline were investigated by cellular uptake using an in vitro model of the inner BRB (TR-iBRB2 cells). The mRNA level of system A was determined by quantitative real-time PCR analysis with specific primers. [3H]l-Proline and [14C]d-mannitol, which is a bulk flow marker, were bi-exponentially eliminated from the vitreous humor after vitreous bolus injection. The elimination rate constant of [3H]l-proline during the terminal phase was 1.6-fold greater than that of [14C]d-mannitol. The terminal elimination rate constant difference between [3H]l-proline and [14C]d-mannitol was reduced in the retinal presence of 3 mM l-proline and 5 mM α-methylaminoisobutyric acid, suggesting that l-proline is transported via a carrier-mediated retina-to-blood transport process across the BRB. [3H]l-Proline uptake by TR-iBRB2 cells appeared to be mediated through a saturable and Na+-dependent process. The corresponding Michaelis-Menten constant was 392 μM. This process was reduced by substrates for system A, suggesting that system A is involved in l-proline uptake. Of the isoforms of system A, ATA1, ATA2, and ATA3, ATA2 mRNA is predominantly expressed in TR-iBRB2 cells and isolated rat retinal endothelial cells. In conclusion, system A, most likely ATA2, is responsible for the retina-to-blood transport of l-proline across the inner BRB and may play a role in maintaining the concentration of small neutral amino acids in the retina. © 2010 Elsevier Ltd.

Access to Document (DOI)

Involvement of OCTN2 in the transport of acetyl-lcarnitine across the inner blood-retinal barrier

TOMI Masatoshi

Investigative Ophthalmology and Visual Science (Investigative Ophthalmology and Visual Science) 51 ( 1 ) 430 - 436 2010.01

Research paper (scientific journal), Accepted, ISSN 0146-0404

PURPOSE. To elucidate the mechanisms of acetyl-L-carnitine transport across the inner blood-retinal barrier (inner BRB). METHODS. In vivo integration plot and retinal uptake index (RUI) analyses were used to examine acetyl-L-[3H]carnitine transport in the retina across the inner BRB in rats. RUI was determined from the ratio of acetyl-L-[3H]carnitine and [14C]n-butanol, a freely diffusible internal reference, in the retina divided by the same ratio in the solution injected in the carotid artery. The transport mechanism was characterized in a conditionally immortalized rat retinal capillary endothelial cell line (TR-iBRB2 cells), as an in vitro inner BRB model. RESULTS. The apparent influx permeability clearance (Kin) per gram retina of acetyl-L-[3H]carnitine was found to be 2.31 μL/(minute·g retina). The Kin of acetyl-L-[3H]carnitine was 3.7-fold greater than that of [3H]D-mannitol, a nonpermeable paracellular marker. Acetyl-L-[3H]carnitine uptake by the retina was found to be significantly inhibited by L-carnitine and acetyl- L-carnitine, supporting a carrier-mediated influx transport of acetyl-L-carnitine at the inner BRB. L-[3H]carnitine and acetyl-L- [3H]carnitine uptake by TR-iBRB2 cells was Na+- and concentration- dependent, with a Km of 29 and 26 μM, respectively. These forms of transport were significantly inhibited by organic cation/carnitine transporter (OCTN) substrates and inhibitors such as L-carnitine and acetyl-L-carnitine, tetraethylammonium, quinidine, and betaine. These transport properties are consistent with those of carnitine transport by OCTN2. OCTN2 was predominantly expressed in TR-iBRB2 cells and isolated rat retinal vascular endothelial cells. CONCLUSIONS. The findings suggest that OCTN2 is involved in the transport of acetyl-L-carnitine from the circulating blood to the retina across the inner BRB. © Association for Research in Vision and Ophthalmology.

Cationic amino acid transporter 1-mediated l-arginine transport at the inner blood-retinal barrier

Tomi M., Kitade N., Hirose S., Yokota N., Akanuma S., Tachikawa M., Hosoya K.

Journal of Neurochemistry (Journal of Neurochemistry) 111 ( 3 ) 716 - 725 2009.11

Research paper (scientific journal), Accepted, ISSN 0022-3042

The purpose of this study was to identify the transporter mediating l-arginine transport at the inner blood-retinal barrier (BRB). The apparent uptake clearance of [3H]l-arginine into the rat retina was found to be 118 μL/(min·g retina), supporting a carrier-mediated influx transport of l-arginine at the BRB. [3H]l-Arginine uptake by a conditionally immortalized rat retinal capillary endothelial cell line (TR-iBRB2 cells), used as an in vitro model of the inner BRB, was primarily an Na +-independent and saturable process with Michaelis-Menten constants of 11.2 μM and 530 μM. This process was inhibited by rat cationic amino acid transporter (CAT) 1-specific small interfering RNA as well as substrates of CATs, l-arginine, l-lysine, and l-ornithine. The expression of cationic amino acid transporter (CAT) 1 mRNA was 25.9- and 796-fold greater than that of CAT3 in TR-iBRB2 and magnetically isolated rat retinal vascular endothelial cells, respectively. The expression of CAT1 protein was detected in TR-iBRB2 cells and immunostaining of CAT1 was observed along the rat retinal capillaries. In conclusion, CAT1 is localized in retinal capillary endothelial cells and at least in part mediates l-arginine transport at the inner BRB. This process seems to be closely involved in visual functions by supplying precursors of biologically important molecules like nitric oxide in the neural retina. © 2009 International Society for Neurochemistry.

Access to Document (DOI)

Roles of inner blood-retinal barrier organic anion transporter 3 in the vitreous/retina-to-blood efflux transport of p-aminohippuric acid, benzylpenicillin, and 6-mercaptopurine

Hosoya K., Makihara A., Tsujikawa Y., Yoneyama D., Mori S., Terasaki T., Akanuma S., Tomi M., Tachikawa M.

Journal of Pharmacology and Experimental Therapeutics (Journal of Pharmacology and Experimental Therapeutics) 329 ( 1 ) 87 - 93 2009.04

Research paper (scientific journal), Accepted, ISSN 0022-3565

The purpose of the present study was to characterize rat organic anion transporter (Oat) 3 (Oat3, Slc22a8) in the efflux transport at the inner blood-retinal barrier (BRB). Reverse transcription-polymerase chain reaction analysis showed that rat (r) Oat3 mRNA is expressed in retinal vascular endothelial cells (RVECs), but not rOat1 and rOat2 mRNA. The expression of Oat3 in the retina and human cultured retinal endothelial cells was further confirmed by Western blot analysis. Immunohisto-chemical staining in RVECs showed that rOat3 is colocalized with glucose transporter 1, but not P-glycoprotein, suggesting that rOat3 is possibly located at the abluminal membrane of the RVEC. The contribution of rOat3 to the efflux of [3H]p-aminohippuric acid ([3H]PAH), [3H]benzylpenicillin ([3H]PCG), and [14C]6-mercaptopurine ([14C]6-MP), substrates of rOat3, from the vitreous humor/retina to the circulating blood across the inner BRB was evaluated using the microdialysis method. [3H]PAH, [ 3H]PCG, [14C]6-MP, and [14C] or [ 3H]D-mannitol, a bulk flow marker, were biexponentially eliminated from the vitreous humor after vitreous bolus injection. The elimination rate constant of [3H]PAH, [3H]PCG, and [14C]6-MP during the terminal phase was approximately 2-fold greater than that of D-mannitol. This efflux transport was reduced in the retinal presence of probenecid, PAH, and PCG, whereas it was not inhibited by digoxin. In conclusion, rOat3 is expressed at the inner BRB and involved in the vitreous humor/retina-to-blood transport of PAH, PCG, and 6-MP. This transport system is one mechanism to limit the retinal distribution of PAH, PCG, and 6-MP. Copyright © 2009 by The American Society for Pharmacology and Experimental Therapeutics.

Access to Document (DOI)

Modulation of retinal capillary endothelial cells by Müller glial cell-derived factors.

Abukawa H., Tomi M., Kiyokawa J., Hori S., Kondo T., Terasaki T., Hosoya K.

Molecular vision (Molecular vision) 15 451 - 457 2009.02

PURPOSE: The inner blood-retinal barrier (BRB) is a gliovascular unit in which macroglial cells surround capillary endothelial cells and regulate retinal capillaries by paracrine interactions. The purpose of the present study was to identify genes of retinal capillary endothelial cells whose expression is modulated by Müller glial cell-derived factors. METHODS: Conditionally immortalized rat retinal capillary endothelial (TR-iBRB2) and Müller (TR-MUL5) cell lines were chosen as an in vitro model. TR-iBRB2 cells were incubated with conditioned medium of TR-MUL5 (MUL-CM) for 24 h and subjected to microarray and quantitative real-time PCR analysis. RESULTS: TR-MUL5 cell-derived factors increased alkaline phosphatase activity in TR-iBRB2 cells, indicating that paracrine interactions occurred between TR-iBRB2 and TR-MUL5 cells. Microarray analysis demonstrated that MUL-CM treatment leads to a modulation of several genes including an induction of plasminogen activator inhibitor 1 (PAI-1) and a suppression of an inhibitor of DNA binding 2 (Id2) in TR-iBRB2 cells. Treatment with TGF-beta1, which is incorporated in MUL-CM, also resulted in an induction of PAI-1 and a suppression of Id2 in TR-iBRB2 cells. CONCLUSIONS: In vitro inner BRB model study revealed that Müller glial cell-derived factors modulate endothelial cell functions including the induction of anti-angiogenic PAI-1 and the suppression of pro-angiogenic Id2. Therefore, Müller cells appear to be one of the modulators of retinal angiogenesis.

Expression and possible role of creatine transporter in the brain and at the blood-cerebrospinal fluid barrier as a transporting protein of guanidinoacetate, an endogenous convulsant

Tachikawa M., Fujinawa J., Takahashi M., Kasai Y., Fukaya M., Sakai K., Yamazaki M., Tomi M., Watanabe M., Sakimura K., Terasaki T., Hosoya K.I.

Journal of Neurochemistry (Journal of Neurochemistry) 107 ( 3 ) 768 - 778 2008.11

Research paper (scientific journal), Accepted, ISSN 0022-3042

Little is known about the cerebral distribution and clearance of guanidinoacetate (GAA), the accumulation of which induces convulsions. The purpose of the present study was to identify creatine transporter (CRT)-mediated GAA transport and to clarify its cerebral expression and role in GAA efflux transport at the blood-cerebrospinal fluid barrier (BCSFB). CRT mediated GAA transport with a Km value of 269 μM/412 μM which was approximately 10-fold greater than that of CRT for creatine. There was wide and distinct cerebral expression of CRT and localization of CRT on the brush-border membrane of choroid plexus epithelial cells. The in vivo elimination clearance of GAA from the CSF was 13-fold greater than that of d-mannitol reflecting bulk flow of the CSF. This process was partially inhibited by creatine. The characteristics of GAA uptake by isolated choroid plexus and an immortalized rat choroid plexus epithelial cell line (TR-CSFB cells) used as an in vitro model of BCSFB are partially consistent with those of CRT. These results suggest that CRT plays a role in the cerebral distribution of GAA and GAA uptake by the choroid plexus. However, in the presence of endogenous creatine in the CSF, CRT may make only a limited contribution to the GAA efflux transport at the BCSFB. © 2008 The Authors.

Access to Document (DOI)

Evaluation of amino acid-mustard transport as L-type amino acid transporter 1 (LAT1)-mediated alkylating agents

Hosoya K., Kyoko H., Toyooka N., Kato A., Orihashi M., Tomi M., Tachikawa M.

Biological and Pharmaceutical Bulletin (Biological and Pharmaceutical Bulletin) 31 ( 11 ) 2126 - 2130 2008.11

Research paper (scientific journal), Accepted, ISSN 0918-6158

The L-type amino acid transporter 1 (LAT1, SLC7A5) is an Na +-independent neutral amino acid transporter the expression of which is located in retinal endothelial cells. Due to its broad substrate selectivity, LAT1 has been proposed to mediate the transport of amino acid-related drugs across the blood-tissue barriers. Here, we have investigated the transport screening of amino acid-mustards using a conditionally immortalized rat retinal capillary endothelial cell line (TR-iBRB2) which expresses LAT1. We synthesized 5 amino acid-mustards: tyrosine-mustard, phenylglycine-mustard, alanine-mustard, ornithine-mustard, and lysine-mustard. LAT1-mediated [3H]L- phenylalanine (Phe) uptake by TR-iBRB2 cells was inhibited in a competitive manner by tyrosine-mustard and phenylglycine-mustard as well as melphalan (phenylalanine-mustard). Phenylglycine-mustard was able to induce the efflux of [3H]Phe preloaded into the TR-iBRB2 cells expressing LAT1 through the obligatory exchange mechanism, although tyrosine-mustard, alanine-mustard, ornithine-mustard, lysine-mustard, and melphalan did not induce any significant efflux. These findings suggest that phenylglycine-mustard is a better substrate for LAT1 than melphalan and other amino acid-mustards. © 2008 Pharmaceutical Society of Japan.

Access to Document (DOI)

Function of taurine transporter (Slc6a6/TauT) as a GABA transporting protein and its relevance to GABA transport in rat retinal capillary endothelial cells

Tomi M., Tajima A., Tachikawa M., Hosoya K.

Biochimica et Biophysica Acta - Biomembranes (Biochimica et Biophysica Acta - Biomembranes) 1778 ( 10 ) 2138 - 2142 2008.10

Research paper (scientific journal), Accepted, ISSN 0005-2736

The purpose of this study was to identify the uptake mechanism of γ-aminobutyric acid (GABA) via taurine transporter (Slc6a6/TauT) and its relationship with GABA transport at the inner BRB. Rat Slc6a6/TauT-transfected HeLa cells exhibited Na+-, Cl--, and concentration-dependent [3H]GABA uptake with a Km of 1.5 mM. Taurine, β-alanine, and GABA markedly inhibited Slc6a6/TauT-mediated uptake of [3H]GABA. The uptake of [3H]GABA by a conditionally immortalized rat retinal capillary endothelial cell line (TR-iBRB2) was Na+-, Cl--, and concentration-dependent with a Km of 2.0 mM. This process was more potently inhibited by substrates of Slc6a6/TauT, taurine and β-alanine, than those of GABA transporters, GABA and betaine. In the presence of taurine, there was competitive inhibition with a Ki of 74 μM. [3H]Taurine also exhibited competitive inhibition with a Ki of 1.8 mM in the presence of GABA. In conclusion, rat Slc6a6/TauT has the ability to use GABA as a substrate and Slc6a6/TauT-mediated GABA transport appears to be present at the inner BRB. © 2008 Elsevier B.V. All rights reserved.

Access to Document (DOI)

　 Previous page - Next page 　