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Emi Nakashima - One of the best experts on this subject based on the ideXlab platform.
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6 mercaptopurine transport by equilibrative nucleoside transporters in conditionally immortalized rat Syncytiotrophoblast Cell lines tr tbts
Journal of Pharmaceutical Sciences, 2011Co-Authors: Emi Nakashima, Sumio Ohtsuki, Young-sook KangAbstract:Abstract Recently, more women were provided with 6-mercaptopurine (6-MP) during pregnancy. Therefore, we attempted to clarify the transport mechanisms of 6-MP through blood–placenta barrier using rat conditionally immortalized Syncytiotrophoblast Cell lines (TR-TBTs). The uptake of 6-MP was time- and ATP dependent, but sodium independent in TR-TBTs. 6-MP was eliminated over 50% from the Cells within 30min. The uptake of 6-MP was saturable with Michaelis–Menten constant values of 198 μM and 250 μM in TR-TBT 18d-1 and TR-TBT 18d-2, respectively. 6-Thioguanine, azathioprine, and hypoxantine, structural analogues of 6-MP, strongly inhibited [ 14 C]6-MP uptake. Equilibrative nucleoside transporter (ENT) inhibitors, adenosine and uridine, significantly inhibited [ 14 C]6-MP uptake. However, several organic anions and cations had no effect on [ 14 C]6-MP uptake in TR-TBTs. These results suggest that sodium-independent transporters, ENTs, may be involved in 6-MP uptake at the placenta. In addition, multidrug resistance protein (MRP) inhibitors, methotrexate, probenecid, cefmetazole, and sulfinpyrazone, significantly increased the accumulation of [ 14 C]6-MP in the Cells. It is indicated that 6-MP may be eliminated across the blood–placental barrier via MRPs. TR-TBTs expressed mRNA of ENT1, ENT2, MRP4, and MRP5. These findings are important for the therapy of acute lymphoblastic leukemia and autoimmune diseases of pregnant women, and should be useful data in elucidating teratogenicity of 6-MP during pregnancy. © 2011 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 100:3773–3782, 2011
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Roles of TauT and system A in cytoprotection of rat Syncytiotrophoblast Cell line exposed to hypertonic stress
Placenta, 2010Co-Authors: Tomohiro Nishimura, Yoshimichi Sai, Noriko Kose, Hisashi Iizasa, J. Fujii, M. Muta, Masatoshi Tomi, M. Deureh, Emi NakashimaAbstract:Abstract 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 [ 3 H]taurine and [ 14 C]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. [ 3 H]Taurine uptake was strongly inhibited by TauT inhibitors such as hypotaurine and β-alanine. MeAIB, a system A specific substrate, inhibited hypertonic stress-induced [ 14 C]betaine uptake. These results suggest that TauT and system A play cytoprotective roles in Syncytiotrophoblasts exposed to hypertonic stress.
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influx mechanism of 2 3 dideoxyinosine and uridine at the blood placenta barrier
Placenta, 2009Co-Authors: Kazuko Sato, Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Noriko Kose, Emi NakashimaAbstract:Abstract The blood–placenta barrier (BPB) serves to protect the fetus from exposure to toxins, and to transport various nutrients, including nucleosides, and hormones from mother to fetus. It is known that nucleoside transporters contribute to the transfer of nucleosides and nucleoside analogues. 2′,3′-Dideoxyinosine (ddI) has a nucleoside structure, and crosses the BPB. Although ddI is a substrate of several transporters, including equilibrative nucleoside transporters (ENT1 and ENT2), the transport mechanism of ddI in the placenta has not yet been characterized. Therefore, the purpose of this study was to clarify the influx mechanisms of ddI from the maternal to the fetal side, and to examine the interaction between ddI and uridine transport at the BPB. We studied ddI and uridine uptakes using a conditionally immortalized rat Syncytiotrophoblast Cell line, TR-TBT 18d-1, as a BPB model. The ddI uptake was temperature-dependent, Na+-independent and saturable. Kinetic analysis yielded Km values for ddI and uridine of 6.51 mM and 23.4 μM, respectively. Uridine uptake was inhibited by ENT1 and ENT2 substrates, and ddI uptake was also inhibited by substrates or inhibitors at concentrations that inhibit ENT2. Uridine uptake in Xenopus laevis oocytes expressing rat ENT2 was inhibited by 5 mM ddI, in agreement with the results for TR-TBT 18d-1. Our results indicate that ddI and uridine are both taken up in part via ENT2 in TR-TBT 18d-1 Cells, and therefore that ENT2 may contribute to their uptake at the BPB.
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Influx Mechanism of 2′,3′-Dideoxyinosine and Uridine at the Blood–Placenta Barrier
Placenta, 2009Co-Authors: Kazuko Sato, Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Noriko Kose, Emi NakashimaAbstract:Abstract The blood–placenta barrier (BPB) serves to protect the fetus from exposure to toxins, and to transport various nutrients, including nucleosides, and hormones from mother to fetus. It is known that nucleoside transporters contribute to the transfer of nucleosides and nucleoside analogues. 2′,3′-Dideoxyinosine (ddI) has a nucleoside structure, and crosses the BPB. Although ddI is a substrate of several transporters, including equilibrative nucleoside transporters (ENT1 and ENT2), the transport mechanism of ddI in the placenta has not yet been characterized. Therefore, the purpose of this study was to clarify the influx mechanisms of ddI from the maternal to the fetal side, and to examine the interaction between ddI and uridine transport at the BPB. We studied ddI and uridine uptakes using a conditionally immortalized rat Syncytiotrophoblast Cell line, TR-TBT 18d-1, as a BPB model. The ddI uptake was temperature-dependent, Na+-independent and saturable. Kinetic analysis yielded Km values for ddI and uridine of 6.51 mM and 23.4 μM, respectively. Uridine uptake was inhibited by ENT1 and ENT2 substrates, and ddI uptake was also inhibited by substrates or inhibitors at concentrations that inhibit ENT2. Uridine uptake in Xenopus laevis oocytes expressing rat ENT2 was inhibited by 5 mM ddI, in agreement with the results for TR-TBT 18d-1. Our results indicate that ddI and uridine are both taken up in part via ENT2 in TR-TBT 18d-1 Cells, and therefore that ENT2 may contribute to their uptake at the BPB.
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Enhancement of Zidovudine Uptake by Dehydroepiandrosterone Sulfate in Rat Syncytiotrophoblast Cell Line TR-TBT 18d-1
Drug metabolism and disposition: the biological fate of chemicals, 2008Co-Authors: Tomohiro Nishimura, Yoshimichi Sai, Takuya Chishu, Kazuko Sato, Noriko Kose, Tetsuya Terasaki, Young-sook Kang, Yoshiaki Seki, Emi NakashimaAbstract:AZT (3'-azido-3'-deoxythymidine; zidovudine), which is used for the prevention of mother-to-child transmission of HIV-1, is transplacentally transferred to the fetus across the blood-placenta barrier, which is composed of Syncytiotrophoblasts. We recently showed that apical uptake of AZT by Syncytiotrophoblasts is mediated by saturable transport system(s) in the TR-TBT 18d-1 Cell line, and the Cellular accumulation of AZT was increased in the presence of dehydroepiandrosterone sulfate (DHEAS). Here, we aimed to clarify the mechanism of this effect of DHEAS. Inhibitors of efflux transporters, including breast cancer resistance protein, P-glycoprotein, and multidrug resistance proteins, had little effect on the Cellular accumulation of AZT in TR-TBT 18d-1. Kinetic study revealed that the rate constant for AZT uptake was greatly increased in the presence of 1 mM DHEAS. These results suggested that the effect of DHEAS was because of enhancement of the uptake process(es), rather than inhibition of efflux. When AZT uptake was analyzed according to the Michaelis-Menten equation, the estimated Michaelis constant, Km, for AZT uptake in the presence of 1 mM DHEAS was lower than that in its absence, whereas maximum uptake velocity, Vmax, and nonsaturable uptake clearance, kns, were similar in the presence and absence of DHEAS, indicating that DHEAS may change the recognition characteristics of the transporter for AZT in TR-TBT 18d-1. Thus, the increase of AZT uptake in TR-TBT 18d-1 Cells in the presence of DHEAS was concluded to be because of a DHEAS-induced change in the affinity of AZT uptake system, although the transporter responsible for AZT uptake has not been identified.
Yoshimichi Sai - One of the best experts on this subject based on the ideXlab platform.
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Roles of TauT and system A in cytoprotection of rat Syncytiotrophoblast Cell line exposed to hypertonic stress
Placenta, 2010Co-Authors: Tomohiro Nishimura, Yoshimichi Sai, Noriko Kose, Hisashi Iizasa, J. Fujii, M. Muta, Masatoshi Tomi, M. Deureh, Emi NakashimaAbstract:Abstract 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 [ 3 H]taurine and [ 14 C]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. [ 3 H]Taurine uptake was strongly inhibited by TauT inhibitors such as hypotaurine and β-alanine. MeAIB, a system A specific substrate, inhibited hypertonic stress-induced [ 14 C]betaine uptake. These results suggest that TauT and system A play cytoprotective roles in Syncytiotrophoblasts exposed to hypertonic stress.
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influx mechanism of 2 3 dideoxyinosine and uridine at the blood placenta barrier
Placenta, 2009Co-Authors: Kazuko Sato, Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Noriko Kose, Emi NakashimaAbstract:Abstract The blood–placenta barrier (BPB) serves to protect the fetus from exposure to toxins, and to transport various nutrients, including nucleosides, and hormones from mother to fetus. It is known that nucleoside transporters contribute to the transfer of nucleosides and nucleoside analogues. 2′,3′-Dideoxyinosine (ddI) has a nucleoside structure, and crosses the BPB. Although ddI is a substrate of several transporters, including equilibrative nucleoside transporters (ENT1 and ENT2), the transport mechanism of ddI in the placenta has not yet been characterized. Therefore, the purpose of this study was to clarify the influx mechanisms of ddI from the maternal to the fetal side, and to examine the interaction between ddI and uridine transport at the BPB. We studied ddI and uridine uptakes using a conditionally immortalized rat Syncytiotrophoblast Cell line, TR-TBT 18d-1, as a BPB model. The ddI uptake was temperature-dependent, Na+-independent and saturable. Kinetic analysis yielded Km values for ddI and uridine of 6.51 mM and 23.4 μM, respectively. Uridine uptake was inhibited by ENT1 and ENT2 substrates, and ddI uptake was also inhibited by substrates or inhibitors at concentrations that inhibit ENT2. Uridine uptake in Xenopus laevis oocytes expressing rat ENT2 was inhibited by 5 mM ddI, in agreement with the results for TR-TBT 18d-1. Our results indicate that ddI and uridine are both taken up in part via ENT2 in TR-TBT 18d-1 Cells, and therefore that ENT2 may contribute to their uptake at the BPB.
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Influx Mechanism of 2′,3′-Dideoxyinosine and Uridine at the Blood–Placenta Barrier
Placenta, 2009Co-Authors: Kazuko Sato, Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Noriko Kose, Emi NakashimaAbstract:Abstract The blood–placenta barrier (BPB) serves to protect the fetus from exposure to toxins, and to transport various nutrients, including nucleosides, and hormones from mother to fetus. It is known that nucleoside transporters contribute to the transfer of nucleosides and nucleoside analogues. 2′,3′-Dideoxyinosine (ddI) has a nucleoside structure, and crosses the BPB. Although ddI is a substrate of several transporters, including equilibrative nucleoside transporters (ENT1 and ENT2), the transport mechanism of ddI in the placenta has not yet been characterized. Therefore, the purpose of this study was to clarify the influx mechanisms of ddI from the maternal to the fetal side, and to examine the interaction between ddI and uridine transport at the BPB. We studied ddI and uridine uptakes using a conditionally immortalized rat Syncytiotrophoblast Cell line, TR-TBT 18d-1, as a BPB model. The ddI uptake was temperature-dependent, Na+-independent and saturable. Kinetic analysis yielded Km values for ddI and uridine of 6.51 mM and 23.4 μM, respectively. Uridine uptake was inhibited by ENT1 and ENT2 substrates, and ddI uptake was also inhibited by substrates or inhibitors at concentrations that inhibit ENT2. Uridine uptake in Xenopus laevis oocytes expressing rat ENT2 was inhibited by 5 mM ddI, in agreement with the results for TR-TBT 18d-1. Our results indicate that ddI and uridine are both taken up in part via ENT2 in TR-TBT 18d-1 Cells, and therefore that ENT2 may contribute to their uptake at the BPB.
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Enhancement of Zidovudine Uptake by Dehydroepiandrosterone Sulfate in Rat Syncytiotrophoblast Cell Line TR-TBT 18d-1
Drug metabolism and disposition: the biological fate of chemicals, 2008Co-Authors: Tomohiro Nishimura, Yoshimichi Sai, Takuya Chishu, Kazuko Sato, Noriko Kose, Tetsuya Terasaki, Young-sook Kang, Yoshiaki Seki, Emi NakashimaAbstract:AZT (3'-azido-3'-deoxythymidine; zidovudine), which is used for the prevention of mother-to-child transmission of HIV-1, is transplacentally transferred to the fetus across the blood-placenta barrier, which is composed of Syncytiotrophoblasts. We recently showed that apical uptake of AZT by Syncytiotrophoblasts is mediated by saturable transport system(s) in the TR-TBT 18d-1 Cell line, and the Cellular accumulation of AZT was increased in the presence of dehydroepiandrosterone sulfate (DHEAS). Here, we aimed to clarify the mechanism of this effect of DHEAS. Inhibitors of efflux transporters, including breast cancer resistance protein, P-glycoprotein, and multidrug resistance proteins, had little effect on the Cellular accumulation of AZT in TR-TBT 18d-1. Kinetic study revealed that the rate constant for AZT uptake was greatly increased in the presence of 1 mM DHEAS. These results suggested that the effect of DHEAS was because of enhancement of the uptake process(es), rather than inhibition of efflux. When AZT uptake was analyzed according to the Michaelis-Menten equation, the estimated Michaelis constant, Km, for AZT uptake in the presence of 1 mM DHEAS was lower than that in its absence, whereas maximum uptake velocity, Vmax, and nonsaturable uptake clearance, kns, were similar in the presence and absence of DHEAS, indicating that DHEAS may change the recognition characteristics of the transporter for AZT in TR-TBT 18d-1. Thus, the increase of AZT uptake in TR-TBT 18d-1 Cells in the presence of DHEAS was concluded to be because of a DHEAS-induced change in the affinity of AZT uptake system, although the transporter responsible for AZT uptake has not been identified.
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Characterization of the Mechanism of Zidovudine Uptake by Rat Conditionally Immortalized Syncytiotrophoblast Cell Line TR-TBT
Pharmaceutical Research, 2008Co-Authors: Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Kazuko Sato, Noriko Kose, Tetsuya Terasaki, Chisato Mukai, Shinji Kitagaki, Naoki MiyakoshiAbstract:Purpose To characterize the uptake mechanism of zidovudine (AZT), a nucleoside reverse transcriptase inhibitor, in Syncytiotrophoblast Cells using the TR-TBT 18d-1 Cell line previously established by our group. Materials and Methods The effects of several transporter inhibitors on the initial and steady-state apical uptake of AZT by TR-TBT 18d-1 were characterized, in order to identify the transporter(s) involved. Results Initial uptake of AZT was sodium-independent and saturable; the K _m value was about 16 μM. Nitrobenzylthioinosine (NBMPR), probenecid and cimetidine each had little effect on the saturable AZT uptake, indicating that well characterized transporters, such as organic anion transporters (OATs and OATPs), organic cation transporters (OCTs) and equilibrative nucleoside transporters (ENTs), are not involved. However, thymidine and 2′-deoxyuridine strongly inhibited AZT uptake. These results suggest that an unidentified nucleoside uptake transporter is responsible for the uptake of AZT. Cyclosporin A, Ko143 and probenecid had little effect on AZT accumulation by TR-TBT 18d-1 Cells, suggesting that transporter-mediated efflux of AZT is not substantial. Conclusion Our results indicate that saturable AZT uptake into TR-TBT 18d-1 is mediated by a so-far-unidentified transporter.
Tomohiro Nishimura - One of the best experts on this subject based on the ideXlab platform.
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Roles of TauT and system A in cytoprotection of rat Syncytiotrophoblast Cell line exposed to hypertonic stress
Placenta, 2010Co-Authors: Tomohiro Nishimura, Yoshimichi Sai, Noriko Kose, Hisashi Iizasa, J. Fujii, M. Muta, Masatoshi Tomi, M. Deureh, Emi NakashimaAbstract:Abstract 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 [ 3 H]taurine and [ 14 C]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. [ 3 H]Taurine uptake was strongly inhibited by TauT inhibitors such as hypotaurine and β-alanine. MeAIB, a system A specific substrate, inhibited hypertonic stress-induced [ 14 C]betaine uptake. These results suggest that TauT and system A play cytoprotective roles in Syncytiotrophoblasts exposed to hypertonic stress.
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influx mechanism of 2 3 dideoxyinosine and uridine at the blood placenta barrier
Placenta, 2009Co-Authors: Kazuko Sato, Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Noriko Kose, Emi NakashimaAbstract:Abstract The blood–placenta barrier (BPB) serves to protect the fetus from exposure to toxins, and to transport various nutrients, including nucleosides, and hormones from mother to fetus. It is known that nucleoside transporters contribute to the transfer of nucleosides and nucleoside analogues. 2′,3′-Dideoxyinosine (ddI) has a nucleoside structure, and crosses the BPB. Although ddI is a substrate of several transporters, including equilibrative nucleoside transporters (ENT1 and ENT2), the transport mechanism of ddI in the placenta has not yet been characterized. Therefore, the purpose of this study was to clarify the influx mechanisms of ddI from the maternal to the fetal side, and to examine the interaction between ddI and uridine transport at the BPB. We studied ddI and uridine uptakes using a conditionally immortalized rat Syncytiotrophoblast Cell line, TR-TBT 18d-1, as a BPB model. The ddI uptake was temperature-dependent, Na+-independent and saturable. Kinetic analysis yielded Km values for ddI and uridine of 6.51 mM and 23.4 μM, respectively. Uridine uptake was inhibited by ENT1 and ENT2 substrates, and ddI uptake was also inhibited by substrates or inhibitors at concentrations that inhibit ENT2. Uridine uptake in Xenopus laevis oocytes expressing rat ENT2 was inhibited by 5 mM ddI, in agreement with the results for TR-TBT 18d-1. Our results indicate that ddI and uridine are both taken up in part via ENT2 in TR-TBT 18d-1 Cells, and therefore that ENT2 may contribute to their uptake at the BPB.
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Influx Mechanism of 2′,3′-Dideoxyinosine and Uridine at the Blood–Placenta Barrier
Placenta, 2009Co-Authors: Kazuko Sato, Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Noriko Kose, Emi NakashimaAbstract:Abstract The blood–placenta barrier (BPB) serves to protect the fetus from exposure to toxins, and to transport various nutrients, including nucleosides, and hormones from mother to fetus. It is known that nucleoside transporters contribute to the transfer of nucleosides and nucleoside analogues. 2′,3′-Dideoxyinosine (ddI) has a nucleoside structure, and crosses the BPB. Although ddI is a substrate of several transporters, including equilibrative nucleoside transporters (ENT1 and ENT2), the transport mechanism of ddI in the placenta has not yet been characterized. Therefore, the purpose of this study was to clarify the influx mechanisms of ddI from the maternal to the fetal side, and to examine the interaction between ddI and uridine transport at the BPB. We studied ddI and uridine uptakes using a conditionally immortalized rat Syncytiotrophoblast Cell line, TR-TBT 18d-1, as a BPB model. The ddI uptake was temperature-dependent, Na+-independent and saturable. Kinetic analysis yielded Km values for ddI and uridine of 6.51 mM and 23.4 μM, respectively. Uridine uptake was inhibited by ENT1 and ENT2 substrates, and ddI uptake was also inhibited by substrates or inhibitors at concentrations that inhibit ENT2. Uridine uptake in Xenopus laevis oocytes expressing rat ENT2 was inhibited by 5 mM ddI, in agreement with the results for TR-TBT 18d-1. Our results indicate that ddI and uridine are both taken up in part via ENT2 in TR-TBT 18d-1 Cells, and therefore that ENT2 may contribute to their uptake at the BPB.
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Enhancement of Zidovudine Uptake by Dehydroepiandrosterone Sulfate in Rat Syncytiotrophoblast Cell Line TR-TBT 18d-1
Drug metabolism and disposition: the biological fate of chemicals, 2008Co-Authors: Tomohiro Nishimura, Yoshimichi Sai, Takuya Chishu, Kazuko Sato, Noriko Kose, Tetsuya Terasaki, Young-sook Kang, Yoshiaki Seki, Emi NakashimaAbstract:AZT (3'-azido-3'-deoxythymidine; zidovudine), which is used for the prevention of mother-to-child transmission of HIV-1, is transplacentally transferred to the fetus across the blood-placenta barrier, which is composed of Syncytiotrophoblasts. We recently showed that apical uptake of AZT by Syncytiotrophoblasts is mediated by saturable transport system(s) in the TR-TBT 18d-1 Cell line, and the Cellular accumulation of AZT was increased in the presence of dehydroepiandrosterone sulfate (DHEAS). Here, we aimed to clarify the mechanism of this effect of DHEAS. Inhibitors of efflux transporters, including breast cancer resistance protein, P-glycoprotein, and multidrug resistance proteins, had little effect on the Cellular accumulation of AZT in TR-TBT 18d-1. Kinetic study revealed that the rate constant for AZT uptake was greatly increased in the presence of 1 mM DHEAS. These results suggested that the effect of DHEAS was because of enhancement of the uptake process(es), rather than inhibition of efflux. When AZT uptake was analyzed according to the Michaelis-Menten equation, the estimated Michaelis constant, Km, for AZT uptake in the presence of 1 mM DHEAS was lower than that in its absence, whereas maximum uptake velocity, Vmax, and nonsaturable uptake clearance, kns, were similar in the presence and absence of DHEAS, indicating that DHEAS may change the recognition characteristics of the transporter for AZT in TR-TBT 18d-1. Thus, the increase of AZT uptake in TR-TBT 18d-1 Cells in the presence of DHEAS was concluded to be because of a DHEAS-induced change in the affinity of AZT uptake system, although the transporter responsible for AZT uptake has not been identified.
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Characterization of the Mechanism of Zidovudine Uptake by Rat Conditionally Immortalized Syncytiotrophoblast Cell Line TR-TBT
Pharmaceutical Research, 2008Co-Authors: Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Kazuko Sato, Noriko Kose, Tetsuya Terasaki, Chisato Mukai, Shinji Kitagaki, Naoki MiyakoshiAbstract:Purpose To characterize the uptake mechanism of zidovudine (AZT), a nucleoside reverse transcriptase inhibitor, in Syncytiotrophoblast Cells using the TR-TBT 18d-1 Cell line previously established by our group. Materials and Methods The effects of several transporter inhibitors on the initial and steady-state apical uptake of AZT by TR-TBT 18d-1 were characterized, in order to identify the transporter(s) involved. Results Initial uptake of AZT was sodium-independent and saturable; the K _m value was about 16 μM. Nitrobenzylthioinosine (NBMPR), probenecid and cimetidine each had little effect on the saturable AZT uptake, indicating that well characterized transporters, such as organic anion transporters (OATs and OATPs), organic cation transporters (OCTs) and equilibrative nucleoside transporters (ENTs), are not involved. However, thymidine and 2′-deoxyuridine strongly inhibited AZT uptake. These results suggest that an unidentified nucleoside uptake transporter is responsible for the uptake of AZT. Cyclosporin A, Ko143 and probenecid had little effect on AZT accumulation by TR-TBT 18d-1 Cells, suggesting that transporter-mediated efflux of AZT is not substantial. Conclusion Our results indicate that saturable AZT uptake into TR-TBT 18d-1 is mediated by a so-far-unidentified transporter.
Noriko Kose - One of the best experts on this subject based on the ideXlab platform.
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Roles of TauT and system A in cytoprotection of rat Syncytiotrophoblast Cell line exposed to hypertonic stress
Placenta, 2010Co-Authors: Tomohiro Nishimura, Yoshimichi Sai, Noriko Kose, Hisashi Iizasa, J. Fujii, M. Muta, Masatoshi Tomi, M. Deureh, Emi NakashimaAbstract:Abstract 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 [ 3 H]taurine and [ 14 C]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. [ 3 H]Taurine uptake was strongly inhibited by TauT inhibitors such as hypotaurine and β-alanine. MeAIB, a system A specific substrate, inhibited hypertonic stress-induced [ 14 C]betaine uptake. These results suggest that TauT and system A play cytoprotective roles in Syncytiotrophoblasts exposed to hypertonic stress.
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influx mechanism of 2 3 dideoxyinosine and uridine at the blood placenta barrier
Placenta, 2009Co-Authors: Kazuko Sato, Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Noriko Kose, Emi NakashimaAbstract:Abstract The blood–placenta barrier (BPB) serves to protect the fetus from exposure to toxins, and to transport various nutrients, including nucleosides, and hormones from mother to fetus. It is known that nucleoside transporters contribute to the transfer of nucleosides and nucleoside analogues. 2′,3′-Dideoxyinosine (ddI) has a nucleoside structure, and crosses the BPB. Although ddI is a substrate of several transporters, including equilibrative nucleoside transporters (ENT1 and ENT2), the transport mechanism of ddI in the placenta has not yet been characterized. Therefore, the purpose of this study was to clarify the influx mechanisms of ddI from the maternal to the fetal side, and to examine the interaction between ddI and uridine transport at the BPB. We studied ddI and uridine uptakes using a conditionally immortalized rat Syncytiotrophoblast Cell line, TR-TBT 18d-1, as a BPB model. The ddI uptake was temperature-dependent, Na+-independent and saturable. Kinetic analysis yielded Km values for ddI and uridine of 6.51 mM and 23.4 μM, respectively. Uridine uptake was inhibited by ENT1 and ENT2 substrates, and ddI uptake was also inhibited by substrates or inhibitors at concentrations that inhibit ENT2. Uridine uptake in Xenopus laevis oocytes expressing rat ENT2 was inhibited by 5 mM ddI, in agreement with the results for TR-TBT 18d-1. Our results indicate that ddI and uridine are both taken up in part via ENT2 in TR-TBT 18d-1 Cells, and therefore that ENT2 may contribute to their uptake at the BPB.
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Influx Mechanism of 2′,3′-Dideoxyinosine and Uridine at the Blood–Placenta Barrier
Placenta, 2009Co-Authors: Kazuko Sato, Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Noriko Kose, Emi NakashimaAbstract:Abstract The blood–placenta barrier (BPB) serves to protect the fetus from exposure to toxins, and to transport various nutrients, including nucleosides, and hormones from mother to fetus. It is known that nucleoside transporters contribute to the transfer of nucleosides and nucleoside analogues. 2′,3′-Dideoxyinosine (ddI) has a nucleoside structure, and crosses the BPB. Although ddI is a substrate of several transporters, including equilibrative nucleoside transporters (ENT1 and ENT2), the transport mechanism of ddI in the placenta has not yet been characterized. Therefore, the purpose of this study was to clarify the influx mechanisms of ddI from the maternal to the fetal side, and to examine the interaction between ddI and uridine transport at the BPB. We studied ddI and uridine uptakes using a conditionally immortalized rat Syncytiotrophoblast Cell line, TR-TBT 18d-1, as a BPB model. The ddI uptake was temperature-dependent, Na+-independent and saturable. Kinetic analysis yielded Km values for ddI and uridine of 6.51 mM and 23.4 μM, respectively. Uridine uptake was inhibited by ENT1 and ENT2 substrates, and ddI uptake was also inhibited by substrates or inhibitors at concentrations that inhibit ENT2. Uridine uptake in Xenopus laevis oocytes expressing rat ENT2 was inhibited by 5 mM ddI, in agreement with the results for TR-TBT 18d-1. Our results indicate that ddI and uridine are both taken up in part via ENT2 in TR-TBT 18d-1 Cells, and therefore that ENT2 may contribute to their uptake at the BPB.
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Enhancement of Zidovudine Uptake by Dehydroepiandrosterone Sulfate in Rat Syncytiotrophoblast Cell Line TR-TBT 18d-1
Drug metabolism and disposition: the biological fate of chemicals, 2008Co-Authors: Tomohiro Nishimura, Yoshimichi Sai, Takuya Chishu, Kazuko Sato, Noriko Kose, Tetsuya Terasaki, Young-sook Kang, Yoshiaki Seki, Emi NakashimaAbstract:AZT (3'-azido-3'-deoxythymidine; zidovudine), which is used for the prevention of mother-to-child transmission of HIV-1, is transplacentally transferred to the fetus across the blood-placenta barrier, which is composed of Syncytiotrophoblasts. We recently showed that apical uptake of AZT by Syncytiotrophoblasts is mediated by saturable transport system(s) in the TR-TBT 18d-1 Cell line, and the Cellular accumulation of AZT was increased in the presence of dehydroepiandrosterone sulfate (DHEAS). Here, we aimed to clarify the mechanism of this effect of DHEAS. Inhibitors of efflux transporters, including breast cancer resistance protein, P-glycoprotein, and multidrug resistance proteins, had little effect on the Cellular accumulation of AZT in TR-TBT 18d-1. Kinetic study revealed that the rate constant for AZT uptake was greatly increased in the presence of 1 mM DHEAS. These results suggested that the effect of DHEAS was because of enhancement of the uptake process(es), rather than inhibition of efflux. When AZT uptake was analyzed according to the Michaelis-Menten equation, the estimated Michaelis constant, Km, for AZT uptake in the presence of 1 mM DHEAS was lower than that in its absence, whereas maximum uptake velocity, Vmax, and nonsaturable uptake clearance, kns, were similar in the presence and absence of DHEAS, indicating that DHEAS may change the recognition characteristics of the transporter for AZT in TR-TBT 18d-1. Thus, the increase of AZT uptake in TR-TBT 18d-1 Cells in the presence of DHEAS was concluded to be because of a DHEAS-induced change in the affinity of AZT uptake system, although the transporter responsible for AZT uptake has not been identified.
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Characterization of the Mechanism of Zidovudine Uptake by Rat Conditionally Immortalized Syncytiotrophoblast Cell Line TR-TBT
Pharmaceutical Research, 2008Co-Authors: Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Kazuko Sato, Noriko Kose, Tetsuya Terasaki, Chisato Mukai, Shinji Kitagaki, Naoki MiyakoshiAbstract:Purpose To characterize the uptake mechanism of zidovudine (AZT), a nucleoside reverse transcriptase inhibitor, in Syncytiotrophoblast Cells using the TR-TBT 18d-1 Cell line previously established by our group. Materials and Methods The effects of several transporter inhibitors on the initial and steady-state apical uptake of AZT by TR-TBT 18d-1 were characterized, in order to identify the transporter(s) involved. Results Initial uptake of AZT was sodium-independent and saturable; the K _m value was about 16 μM. Nitrobenzylthioinosine (NBMPR), probenecid and cimetidine each had little effect on the saturable AZT uptake, indicating that well characterized transporters, such as organic anion transporters (OATs and OATPs), organic cation transporters (OCTs) and equilibrative nucleoside transporters (ENTs), are not involved. However, thymidine and 2′-deoxyuridine strongly inhibited AZT uptake. These results suggest that an unidentified nucleoside uptake transporter is responsible for the uptake of AZT. Cyclosporin A, Ko143 and probenecid had little effect on AZT accumulation by TR-TBT 18d-1 Cells, suggesting that transporter-mediated efflux of AZT is not substantial. Conclusion Our results indicate that saturable AZT uptake into TR-TBT 18d-1 is mediated by a so-far-unidentified transporter.
Takuya Chishu - One of the best experts on this subject based on the ideXlab platform.
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influx mechanism of 2 3 dideoxyinosine and uridine at the blood placenta barrier
Placenta, 2009Co-Authors: Kazuko Sato, Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Noriko Kose, Emi NakashimaAbstract:Abstract The blood–placenta barrier (BPB) serves to protect the fetus from exposure to toxins, and to transport various nutrients, including nucleosides, and hormones from mother to fetus. It is known that nucleoside transporters contribute to the transfer of nucleosides and nucleoside analogues. 2′,3′-Dideoxyinosine (ddI) has a nucleoside structure, and crosses the BPB. Although ddI is a substrate of several transporters, including equilibrative nucleoside transporters (ENT1 and ENT2), the transport mechanism of ddI in the placenta has not yet been characterized. Therefore, the purpose of this study was to clarify the influx mechanisms of ddI from the maternal to the fetal side, and to examine the interaction between ddI and uridine transport at the BPB. We studied ddI and uridine uptakes using a conditionally immortalized rat Syncytiotrophoblast Cell line, TR-TBT 18d-1, as a BPB model. The ddI uptake was temperature-dependent, Na+-independent and saturable. Kinetic analysis yielded Km values for ddI and uridine of 6.51 mM and 23.4 μM, respectively. Uridine uptake was inhibited by ENT1 and ENT2 substrates, and ddI uptake was also inhibited by substrates or inhibitors at concentrations that inhibit ENT2. Uridine uptake in Xenopus laevis oocytes expressing rat ENT2 was inhibited by 5 mM ddI, in agreement with the results for TR-TBT 18d-1. Our results indicate that ddI and uridine are both taken up in part via ENT2 in TR-TBT 18d-1 Cells, and therefore that ENT2 may contribute to their uptake at the BPB.
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Influx Mechanism of 2′,3′-Dideoxyinosine and Uridine at the Blood–Placenta Barrier
Placenta, 2009Co-Authors: Kazuko Sato, Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Noriko Kose, Emi NakashimaAbstract:Abstract The blood–placenta barrier (BPB) serves to protect the fetus from exposure to toxins, and to transport various nutrients, including nucleosides, and hormones from mother to fetus. It is known that nucleoside transporters contribute to the transfer of nucleosides and nucleoside analogues. 2′,3′-Dideoxyinosine (ddI) has a nucleoside structure, and crosses the BPB. Although ddI is a substrate of several transporters, including equilibrative nucleoside transporters (ENT1 and ENT2), the transport mechanism of ddI in the placenta has not yet been characterized. Therefore, the purpose of this study was to clarify the influx mechanisms of ddI from the maternal to the fetal side, and to examine the interaction between ddI and uridine transport at the BPB. We studied ddI and uridine uptakes using a conditionally immortalized rat Syncytiotrophoblast Cell line, TR-TBT 18d-1, as a BPB model. The ddI uptake was temperature-dependent, Na+-independent and saturable. Kinetic analysis yielded Km values for ddI and uridine of 6.51 mM and 23.4 μM, respectively. Uridine uptake was inhibited by ENT1 and ENT2 substrates, and ddI uptake was also inhibited by substrates or inhibitors at concentrations that inhibit ENT2. Uridine uptake in Xenopus laevis oocytes expressing rat ENT2 was inhibited by 5 mM ddI, in agreement with the results for TR-TBT 18d-1. Our results indicate that ddI and uridine are both taken up in part via ENT2 in TR-TBT 18d-1 Cells, and therefore that ENT2 may contribute to their uptake at the BPB.
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Enhancement of Zidovudine Uptake by Dehydroepiandrosterone Sulfate in Rat Syncytiotrophoblast Cell Line TR-TBT 18d-1
Drug metabolism and disposition: the biological fate of chemicals, 2008Co-Authors: Tomohiro Nishimura, Yoshimichi Sai, Takuya Chishu, Kazuko Sato, Noriko Kose, Tetsuya Terasaki, Young-sook Kang, Yoshiaki Seki, Emi NakashimaAbstract:AZT (3'-azido-3'-deoxythymidine; zidovudine), which is used for the prevention of mother-to-child transmission of HIV-1, is transplacentally transferred to the fetus across the blood-placenta barrier, which is composed of Syncytiotrophoblasts. We recently showed that apical uptake of AZT by Syncytiotrophoblasts is mediated by saturable transport system(s) in the TR-TBT 18d-1 Cell line, and the Cellular accumulation of AZT was increased in the presence of dehydroepiandrosterone sulfate (DHEAS). Here, we aimed to clarify the mechanism of this effect of DHEAS. Inhibitors of efflux transporters, including breast cancer resistance protein, P-glycoprotein, and multidrug resistance proteins, had little effect on the Cellular accumulation of AZT in TR-TBT 18d-1. Kinetic study revealed that the rate constant for AZT uptake was greatly increased in the presence of 1 mM DHEAS. These results suggested that the effect of DHEAS was because of enhancement of the uptake process(es), rather than inhibition of efflux. When AZT uptake was analyzed according to the Michaelis-Menten equation, the estimated Michaelis constant, Km, for AZT uptake in the presence of 1 mM DHEAS was lower than that in its absence, whereas maximum uptake velocity, Vmax, and nonsaturable uptake clearance, kns, were similar in the presence and absence of DHEAS, indicating that DHEAS may change the recognition characteristics of the transporter for AZT in TR-TBT 18d-1. Thus, the increase of AZT uptake in TR-TBT 18d-1 Cells in the presence of DHEAS was concluded to be because of a DHEAS-induced change in the affinity of AZT uptake system, although the transporter responsible for AZT uptake has not been identified.
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Characterization of the Mechanism of Zidovudine Uptake by Rat Conditionally Immortalized Syncytiotrophoblast Cell Line TR-TBT
Pharmaceutical Research, 2008Co-Authors: Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Kazuko Sato, Noriko Kose, Tetsuya Terasaki, Chisato Mukai, Shinji Kitagaki, Naoki MiyakoshiAbstract:Purpose To characterize the uptake mechanism of zidovudine (AZT), a nucleoside reverse transcriptase inhibitor, in Syncytiotrophoblast Cells using the TR-TBT 18d-1 Cell line previously established by our group. Materials and Methods The effects of several transporter inhibitors on the initial and steady-state apical uptake of AZT by TR-TBT 18d-1 were characterized, in order to identify the transporter(s) involved. Results Initial uptake of AZT was sodium-independent and saturable; the K _m value was about 16 μM. Nitrobenzylthioinosine (NBMPR), probenecid and cimetidine each had little effect on the saturable AZT uptake, indicating that well characterized transporters, such as organic anion transporters (OATs and OATPs), organic cation transporters (OCTs) and equilibrative nucleoside transporters (ENTs), are not involved. However, thymidine and 2′-deoxyuridine strongly inhibited AZT uptake. These results suggest that an unidentified nucleoside uptake transporter is responsible for the uptake of AZT. Cyclosporin A, Ko143 and probenecid had little effect on AZT accumulation by TR-TBT 18d-1 Cells, suggesting that transporter-mediated efflux of AZT is not substantial. Conclusion Our results indicate that saturable AZT uptake into TR-TBT 18d-1 is mediated by a so-far-unidentified transporter.
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characterization of the mechanism of zidovudine uptake by rat conditionally immortalized Syncytiotrophoblast Cell line tr tbt
Pharmaceutical Research, 2008Co-Authors: Yoshimichi Sai, Tomohiro Nishimura, S Shimpo, Takuya Chishu, Kazuko Sato, Noriko Kose, Tetsuya Terasaki, Chisato Mukai, Shinji Kitagaki, Naoki MiyakoshiAbstract:Purpose To characterize the uptake mechanism of zidovudine (AZT), a nucleoside reverse transcriptase inhibitor, in Syncytiotrophoblast Cells using the TR-TBT 18d-1 Cell line previously established by our group.
