We previously reported that 8-oxoguanine (8-oxoG) accumulates in the cytoplasm of dopamine neurons in the substantia nigra of patients with Parkinson's disease and the expression of MTH1 carrying an oxidized purine nucleoside triphosphatase activity increases in these neurons, thus suggesting that oxidative damage in nucleic acids is involved in dopamine neuron loss. In the present study, we found that levels of 8-oxoG in cellular DNA and RNA increased in the mouse nigrostriatal system during the tyrosine hydroxylase (TH)-positive dopamine neuron loss induced by the administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). MTH1-null mice exhibited a greater accumulation of 8-oxoG in mitochondrial DNA accompanied by a more significant decrease in TH and dopamine transporter immunoreactivities in the striatum after MPTP administration, than in wild-type mice. We thus demonstrated that MTH1 protects the dopamine neurons from oxidative damage in the nucleic acids, especially in the mitochondrial DNA of striatal nerve terminals of dopamine neurons.

Enhanced oxidative stress has been implicated in the excitotoxicity of the CNS, and 8-oxo-7,8-dihydro-guanine (8-oxoG), a major type of oxidative damage in nucleic acids, was reported to be accumulated in the rat hippocampus after kainate administration. We herein showed that the 8-oxoG levels in mitochondrial DNA and cellular RNA increased significantly in the CA3 subregion of the mouse hippocampus 6 -12 h after kainate administration but returned to basal levels within a few days. Laser-scanning confocal microscopy revealed the 8-oxoG accumulation in mitochondrial DNA to be remarkable in CA3 microglia, whereas that in nuclear DNA or cellular RNA was also detected in the CA3 pyramidal cells and astrocytes. 8-oxoG accumulation in cellular DNA or RNA should be suppressed by MutT homolog 1 (MTH1) with 8-oxo-dGTPase (8-oxo-7,8-dihydro-2Ј-deoxyguanosine triphosphatase) activity and 8-oxoG-DNA glycosylase 1 (OGG1) with 8-oxoG DNA glycosylase activity. We thus examined the expression level of MTH1 and OGG1 in the mouse hippocampus after kainate administration. The Mth1 mRNA level decreased soon after kainate administration and then quickly recovered beyond the basal level, and a continuously increased MTH1 protein level was observed, whereas the Ogg1 mRNA level remained constant. MTH1-null and wild-type mice exhibited a similar degree of CA3 neuron loss after kainate administration; however, the 8-oxoG levels that accumulated in mitochondrial DNA and cellular RNA in the CA3 microglia significantly increased in the MTH1-null mice in comparison with wild-type mice, thus demonstrating that MTH1 efficiently suppresses the accumulation of 8-oxoG in both cellular DNA and RNA in the hippocampus, especially in microglia, caused by excitotoxicity.

We examined the expression of galectin-1, an endogenous lectin with one carbohydrate-binding domain, in the adult mouse hippocampus after systemic kainate administration. We found that the expression of galectin-1 was remarkably increased in activated astrocytes of the CA3 subregion and dentate gyrus of the hippocampus, and in nestin-positive neural progenitors in the dentate gyrus. Quantitative reverse transcription PCR (RT-PCR) analysis revealed that the galectin-1 mRNA level in hippocampus began to increase 1 day after kainate administration and that a 13-fold increase was attained within 3 days. Western blotting analysis confirmed that the level of galectin-1 protein increased to more than three-fold a week after the exposure. We showed that isolated astrocytes express and secrete galectin-1. To clarify the significance of the increased expression of galectin-1 in hippocampus, we compared the levels of hippocampal cell proliferation in galectin-1 knockout and wild-type mice after saline or kainate administration. The number of 5-bromo-2 0 -deoxyuridine (BrdU)-positive cells detected in the subgranular zone (SGZ) of galectin-1 knockout mice decreased to 62% with saline, and to 52% with kainate, as compared with the number seen in the wild-type mice. Most of the BrdU-positive cells in SGZ expressed doublecortin and neuron-specific nuclear protein, indicating that they are immature neurons. We therefore concluded that galectin-1 promotes basal and kainateinduced proliferation of neural progenitors in the hippocampus.

ABSTRACT:The aim of this study was to develop a pharmacokinetic model to describe the transplacental transfer of drugs, based on the human placental perfusion study. The maternal and fetal sides of human placentas were perfused with salicylic acid together with antipyrine, a passive diffusion marker. The drug concentration in the placental tissue was determined at the end of perfusion. A compartment model consisting of maternal space, fetal intravascular space, and placental tissue was fitted to the observed concentration profiles of salicylic acid in the maternal and fetal effluents. The developed model could adequately explain the concentration profiles of salicylic acid in the effluents with influx clearances from maternal and fetal perfusates to placental tissue of 0.0407 and 0.0813 ml/min/g cotyledon and efflux rate constants from placental tissue to maternal and fetal perfusates (k 2 and k 3 ) of 0.0238 and 0.176 min ؊1 , respectively. The kinetics of antipyrine was adequately described by assuming rapid equilibrium between fetal perfusate and placental tissue compartments. The influx plasma clearance from the maternal side (K؆ 1 ) in humans was estimated by taking into account the protein binding. The K؆ 1 /k 2 value of salicylic acid was 1.07 ml/g cotyledon and was larger than that of antipyrine (0.642 ml/g cotyledon). We evaluated the transplacental transfer kinetics of salicylic acid by human placental perfusion study with various perfusion protocols. Based on the data obtained, we developed a pharmacokinetic model, which should enable us to estimate the influx profile of drugs into umbilical arterial blood from the maternal plasma concentration profile.Exchange of materials between mother and fetus occurs across the placenta (Robertson and Karp, 1976), where fetal blood perfuses the villi and maternal blood fills the interstitial space. The blood-placental barrier consists of trophoblasts, which face the interstitial space, and fetal capillary endothelium (Stulc, 1989). To estimate the distribution of a drug into the fetus, it is essential to investigate the transplacental transfer process.Human placental perfusion, first designed by Schneider et al. (1972), is a useful technique to investigate drug transfer from the maternal to the fetal circulation in humans, because the influx and efflux profiles of the drug can be directly observed. In vivo distribution studies with pregnant animals cannot provide such information and also suffer from the problem of interspecies differences. The placental perfusion technique is also preferable to in vitro methods using cultured human cell lines, such as BeWo or Jar, or membrane vesicles prepared from human placenta (Martel and Keating, 2003;Manley et al., 2005), because physiological metabolism is quite well retained (Nanovskaya et al., 2002). However, many studies using human placental perfusion have provided only simple information, i.e., the ratio of drug concentration in fetal effluent to that in maternal effluent (Heikkinen et al., 2000;Nekhayeva et al., 2005...

OATP8, a member of the organic anion-transporting polypeptide family, is expressed on the sinusoidal membrane of hepatocytes, and transports endogenous organic anions, such as 17beta-glucuronosyl estradiol, and xenobiotic substances, such as digoxin. The objective of this study is to search for polymorphisms of the OATP8 gene and to assess the allele frequency of the polymorphisms in the Japanese population. Analysis of the OATP8 gene in 79 subjects revealed complete linkage of two deletion polymorphisms in the 5' regulatory region, deletion from position -28 to -11 and deletion from position -7 to -4, with an allele frequency of 0.196 for the deletion allele. The polymorphisms T334G (Ser112Ala) and G699A (Met233Ile) were also shown to be in complete linkage disequilibrium, with an allele frequency of 0.728 for the variant (112Ala/233Ile) allele. Interestingly, linkage disequilibrium was identified between the ins/del polymorphism and SNPs at 112 and 233. The predicted major haplotype was the insert-variant type with a haplotype frequency of 0.60.