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ABSTRACT: Ginseng, the root of Panax ginseng C.A. Meyer, is used as a general tonic. Recently, we isolated a novel ginsengderived lysophosphatidic acid (LPA) receptor ligand, gintonin. Gintonin activates G protein-coupled LPA receptors with high affinity in cells endogenously expressing LPA receptors, e.g., Xenopus oocytes. P2X receptors are ligandgated ion channels activated by extracellular ATP, and 7 receptor subtypes (P2X-P2X) have been identified. Most of the P2X receptors are expressed in the smooth muscles of genitourinary organs involved in reproduction. A main characteristic of the P2X receptor is rapid desensitization after repeated ATP treatment of cells or tissues expressing P2X receptors. In the present study, we examined the effect of gintonin on P2X receptor channel activity. P2X receptors were heterologously expressed in Xenopus oocytes. ATP treatment of oocytes expressing P2X receptors induced large inward currents (I ), but repetitive ATP treatments induced a rapid desensitization of I . Gintonin treatment after P2X receptor desensitization potentiated I in a concentration-dependent manner. We further examined the signaling transduction pathways involved in gintonin-mediated potentiation of I . Gintoninmediated I potentiation was blocked by Ki16425, an LPA1/3 receptor antagonist, a PKC inhibitor, a PLC inhibitor, and a PI4-Kinase inhibitor but not by a calcium chelator. In addition, mutations of the phosphoinositide binding site of the P2X receptor greatly attenuated the gintonin-mediated I potentiation. These results indicate that G protein-coupled LPA receptor activation by gintonin is coupled to the potentiation of the desensitized P2X receptor through a phosphoinositide-dependent pathway.
Molecules and Cells 02/2013; 35(2):142-50. · 2.18 Impact Factor
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ABSTRACT: Recently, cognitive therapy in people at ultra-high risk (UHR) for psychosis has been reported to show modest treatment benefits. The aim of this study was to assess the feasibility and efficacy of cognitive therapy in reducing psychiatric symptoms in UHR people in Korea.
We developed cognitive therapy for people at UHR for psychosis inspired by Morrison in 2004. Twenty-two UHR subjects were assigned to cognitive therapy, and 18 subjects completed the 10-session therapy. Psychopathology scores were assessed at baseline and post-treatment.
Cognitive therapy significantly reduced the severity of psychopathology including positive, negative and depressive symptoms. The within-group effect sizes indicated large treatment benefits for these psychopathologies.
These findings suggest that cognitive therapy can be administered to people at UHR for psychosis in non-western culture.
Early Intervention in Psychiatry 05/2011; 5(2):174-8. · 0.92 Impact Factor
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Byung-Hwan Lee,
Sun-Hye Choi,
Tae-Joon Shin,
Mi Kyung Pyo,
Sung-Hee Hwang, Bo-Ra Kim,
Sang-Mok Lee,
Jun-Ho Lee,
Hyoung-Chun Kim,
Hye-Young Park,
Hyewhon Rhim,
Seung-Yeol Nah
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ABSTRACT: The flavonoid quercetin is a low molecular weight substance found in fruits and vegetables. Aside from its anti-oxidative effect, quercetin, like other flavonoids, has a wide range of neuropharmacological actions. The α7 nicotinic acetylcholine receptor (α7 nAChR) has a Ca(2+)-binding site, is highly permeable to the Ca(2+) ion, and plays important roles in Ca(2+)-related normal brain functions. Dysfunctions of α7 nAChR are associated with a variety of neurological disorders. In the present study, we investigated the effects of quercetin on the ACh-induced inward peak current (I(ACh)) in Xenopus oocytes that heterologously express human α7 nAChR. I(ACh) was measured with the two-electrode voltage clamp technique. In oocytes injected with α7 nAChR cRNA, the effects of the co-application of quercetin on I(ACh) were concentration-dependent and reversible. The ED(50) was 36.1 + 6.1 μM. Quercetin-mediated enhancement of I(ACh) caused more potentiation when quercetin was pre-applied. The degree of I(ACh) potentiation by quercetin pre-application was time-dependent and saturated after 1 min. Quercetin-mediated I(ACh) enhancement was not affected by ACh concentration and was voltage-independent. However, quercetin-mediated I(ACh) enhancement was dependent on extracellular Ca(2+) concentrations and was specific to the Ca(2+) ion, since the removal of extracellular Ca(2+) or the addition of Ba(2+) instead of Ca(2+) greatly diminished quercetin enhancement of I(ACh). The mutation of Glu195 to Gln195, in the Ca(2+)-binding site, almost completely diminished quercetin-mediated I(ACh) enhancement. These results indicate that quercetin-mediated I(ACh) enhancement human α7 nAChR heterologously expressed in Xenopus oocytes could be achieved through interactions with the Ca(2+)-binding site of the receptor.
Molecules and Cells 09/2010; 30(3):245-53. · 2.18 Impact Factor
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Sun-Hye Choi,
Tae-Joon Shin,
Byung-Hwan Lee,
Dae-hyun Chu,
Han Choe,
Mi-Kyung Pyo,
Sung-Hee Hwang, Bo-Ra Kim,
Sang-Mok Lee,
Jun-Ho Lee,
Dong-Hyun Kim,
Hyoung-Chun Kim,
Hye-whon Rhim,
Seung-Yeol Nah
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ABSTRACT: The slowly activating delayed rectifier K(+) channels (I(Ks)) are one of the main pharmacological targets for development of drugs against cardiovascular diseases. Cardiac I(Ks) consists of KCNQ1 plus KCNE1 subunits. Ginsenoside, one of the active ingredient of Panax ginseng, enhances cardiac I(Ks) currents. However, little is known about the molecular mechanisms of how ginsenoside interacts with channel proteins to enhance cardiac I(Ks). In the present study, we investigated ginsenoside Rg(3) (Rg(3)) effects on human I(Ks) by co-expressing human KCNQ1 plus KCNE1 subunits in Xenopus oocytes. Rg(3) enhanced I(Ks) currents in concentration- and voltage-dependent manners. The EC(50) was 15.2+/-8.7 microM. However, in oocytes expressing KCNQ1 alone, Rg(3) inhibited the currents with concentration- and voltage-dependent manners. The IC(50) was 4.8+/-0.6 microM. Since Rg(3) acts opposite ways in oocytes expressing KCNQ1 alone or KCNQ1 plus KCNE1 subunits, we examined Rg(3) effects after co-expression of different ratios of KCNE1 and KCNQ1. The increase of KCNE1/KCNQ1 ratio converted I(Ks) inhibition to I(Ks) activations. One to ten ratio of KCNE1 and KCNQ1 subunit is required for Rg(3) activation of I(Ks). Mutations of K318 and V319 into K318Y and V319Y of KCNQ1 channel abolished Rg(3) effects on KCNQ1 or KCNQ1 plus KCNE1 channel currents. The docked modeling revealed that K318 residue plays a key role in stabilization between Rg(3) and KCNQ1 plus KCNE1 or KCNQ1 subunit. These results indicate that Rg(3)-induced activation of I(Ks) requires co-assembly of KCNQ1 and KCNE1 subunits and achieves this through interaction with residues K318 and V319 of KCNQ1 subunit.
European journal of pharmacology 07/2010; 637(1-3):138-47. · 2.59 Impact Factor
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Tae-Joon Shin,
Sun-Hye Choi,
Byung-Hwan Lee,
Mi Kyung Pyo,
Sung-Hee Hwang, Bo-Ra Kim,
Sang-Mok Lee,
Ye Sun Han,
Jun-Ho Lee,
Ji-Ho Park,
Hyoung-Chun Kim,
Hyewhon Rhim,
Seung-Yeol Nah
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ABSTRACT: Quercetin is a low molecular weight flavonoid found in dietary fruits and vegetables. Quercetin, like other flavonoids, has demonstrated neuroprotective effects in vitro and in vivo. However, relatively little is known about how quercetin achieves its neuroprotective abilities. The alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor is one of several excitatory receptors, which play an important role in postsynaptic neurotransmission. Over-stimulation of ionotropic glutamate receptor including AMPA receptors is closely associated with excitatory neurotoxicities. In the present study, we investigated the effects of quercetin on the glutamate-induced inward current (IGlu) in Xenopus oocytes that heterologously express human AMPA receptor and stargazin, an auxiliary subunit of AMPA receptor. IGlu was measured using the two-electrode voltage clamp technique. In oocytes injected with cRNAs coding AMPA receptor (GluR1) and stargazin, quercetin inhibited IGlu in a reversible and concentration-dependent manner. The IC50 was 84.9+/-15.0 microM. Quercetin action on IGlu was attenuated by increasing glutamate concentration, and was membrane holding potential-dependent. These results show a possibility that quercetin interacts with AMPA receptor, which was heterologously expressed in Xenopus oocytes and that quercetin action on IGlu of AMPA receptor could be one of contributions of quercetin-mediated neuroprotections.
Biological & Pharmaceutical Bulletin 01/2010; 33(9):1615-9. · 1.66 Impact Factor
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Sun-Hye Choi,
Jun-Ho Lee,
Mi Kyung Pyo,
Byung-Hwan Lee,
Tae-Joon Shin,
Sung-Hee Hwang, Bo-Ra Kim,
Sang-Mok Lee,
Jae-Wook Oh,
Hyoung-Chun Kim,
Chun Sik Bae,
Hyewhon Rhim,
Seung-Yeol Nah
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ABSTRACT: Many lines of evidences have shown that Panax ginseng exhibits beneficial effects on cardiovascular systems. We previously demonstrated that ginsenoside Rg(3) (Rg(3)), one of active ingredients of Panax ginseng, inhibits Ca(2+) channel currents in a stereospecific manner and affects the steady-state activation but not inactivation. This points a possibility that Rg(3) regulates Ca(2+) channels through specific interaction site(s) for Ca(2+) influx inhibition through Ca(2+) channels. However, it was not known how Rg(3) interacts with Ca(2+) channel proteins. In the current study, we sought to identify these site(s) in Xenopus oocytes expressing cardiac wild-type and mutant L(alpha(1C))-type Ca(2+) channels using the two-microelectrode voltage-clamp technique. To this end, we assessed how various point mutations of the L-type Ca(2+) channel affected the Rg(3) action. Mutations of L427R, N428R and L431K in transmembrane domain-I-segment 6 (IS6) of the channel significantly attenuated the Rg(3) action and caused rightward shifts in dose-response curves. Rg(3) treatment produced a negative shift in the inactivation voltage but did not alter the steady-state activation voltage, and none of the mutant channels affected the Rg(3)-induced negative shift of inactivation voltage. Rg(3) had no effects on inactivation time constant in wild-type and mutant channels. These results indicate that Rg(3) inhibition of L-type Ca(2+) channel currents is attenuated by mutations of Leu427, Asn428 and Leu431 in transmembrane IS6 residues. Leu427, Asn428 and Leu431 residues of the L-type Ca(2+) channel play important roles in the Rg(3) effect on channel properties.
Biological & Pharmaceutical Bulletin 08/2009; 32(7):1224-30. · 1.66 Impact Factor
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ABSTRACT: Previous reports have shown that diltiazem and TMB, calcium channel antagonists, inhibit 5-hydroxytryptamine type 3A (5-HT(3A)) receptor-mediated currents (I(5-HT)) in cell lines and in heterologously expressed Xenopus oocytes. In the present study, we sought to elucidate the molecular mechanisms underlying diltiazem- and TMB-induced 5-HT(3A) receptor regulations. We used the two-microelectrode voltage clamp technique to investigate the effect of diltiazem and TMB on 5-HT-mediated ion currents in Xenopus oocytes expressing wild-type or 5-HT(3A) receptors harboring mutations in the gating pore region of transmembrane domain 2 (TM2). In oocytes expressing wild-type 5-HT(3A) receptors, diltiazem and TMB dose-dependently inhibited peak I(5-HT) with an IC(50) of 71.4+/-4.9 and 4.5+/-0.3 microM, respectively. Among various mutants of TM2, mutation V291A greatly attenuated and abolished the TMB- and diltiazem-induced inhibition of peak I(5-HT), respectively. Mutation V291A also induced constitutively active ion currents in the absence of 5-HT. Diltiazem and TMB inhibited constitutively active ion currents in a dose-dependent manner. The IC(50) values of constitutively active ion currents in V291A receptors were 165.3+/-11.1 and 6.6+/-0.5 microM for diltiazem and 3,4,5-trimethoxybenzoic acid 8-(diethylamino)octyl ester (TMB-8), respectively. Results of site-directed mutagenesis experiments suggest that the Val291 residue could be a candidate for common interaction site for diltiazem- and TMB-8-mediated 5-HT(3A) receptor regulations.
Biological & Pharmaceutical Bulletin 06/2009; 32(5):861-7. · 1.66 Impact Factor
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Byung-Hwan Lee,
Sun-Hye Choi,
Mi Kyung Pyo,
Tae-Joon Shin,
Sung-Hee Hwang, Bo-Ra Kim,
Sang-Mok Lee,
Jun-Ho Lee,
Joon-Hee Lee,
Hui Sun Lee,
Han Choe,
Kyou-Hoon Han,
Hyoung-Chun Kim,
Hyewhon Rhim,
Joon-Hwan Yong,
Seung-Yeol Nah
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ABSTRACT: Nicotinic acetylcholine receptors (nAChRs) play important roles in nervous system functions and are involved in a variety of diseases. We previously demonstrated that ginsenosides, the active ingredients of Panax ginseng, inhibit subsets of nAChR channel currents, but not alpha7, expressed in Xenopus laevis oocytes. Mutation of the highly conserved Leu247 to Thr247 in the transmembrane domain 2 (TM2) channel pore region of alpha7 nAChR induces alterations in channel gating properties and converts alpha7 nAChR antagonists into agonists. In the present study, we assessed how point mutations in the Leu247 residue leading to various amino acids affect 20(S)-ginsenoside Rg(3) (Rg(3)) activity against the alpha7 nAChR. Mutation of L247 to L247A, L247D, L247E, L247I, L247S, and L247T, but not L247K, rendered mutant receptors sensitive to Rg(3). We further characterized Rg(3) regulation of L247T receptors. We found that Rg(3) inhibition of mutant alpha7 nAChR channel currents was reversible and concentration-dependent. Rg(3) inhibition was strongly voltage-dependent and noncompetitive manner. These results indicate that the interaction between Rg(3) and mutant receptors might differ from its interaction with the wild-type receptor. To identify differences in Rg(3) interactions between wild-type and L247T receptors, we utilized docked modeling. This modeling revealed that Rg(3) forms hydrogen bonds with amino acids, such as Ser240 of subunit I and Thr244 of subunit II and V at the channel pore, whereas Rg(3) localizes at the interface of the two wild-type receptor subunits. These results indicate that mutation of Leu247 to Thr247 induces conformational changes in the wild-type receptor and provides a binding pocket for Rg(3) at the channel pore.
Molecules and Cells 06/2009; 27(5):591-9. · 2.18 Impact Factor
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Jun-Ho Lee,
Sun-Hye Choi,
Byung-Hwan Lee,
Tae-Joon Shin,
Mi Kyung Pyo,
Sung-Hee Hwang, Bo-Ra Kim,
Sang-Mok Lee,
Dong-Ho Bae,
Hyewhon Rhim,
Seung-Yeol Nah
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ABSTRACT: Kv1.4 channel belongs to the family of voltage-gated potassium channels that mediate transient and rapidly inactivating A-type currents and N-type inactivation. This N-type inactivation can be removed by the deletion of N-terminal domains, which exhibit non-inactivating currents and C-type inactivation. In our previous report, we demonstrated that 20(S)-ginsenoside Rg(3) (Rg(3)), one of the active ingredients of ginseng saponins, inhibits human Kv1.4 (hKv1.4) channel currents through the interaction with amino acids, including Lys (K) residue, which is known as K(+) activation and the extracellular tetraethylammonium (TEA) binding site. In the present study, we examined the effects of Rg(3) on hKv1.4 channel currents without the N-terminal rapid inactivation domain. We constructed hKv1.4Delta2-61 channels by N-terminal deletion of 2-61 amino acid residues. We investigated the effect of Rg(3) on hKv1.4Delta2-61 channel currents. We found that Rg(3) preferentially inhibited non-inactivating outward currents rather than peak outward currents of hKv1.4Delta2-61 channels. The mutation of K531 hKv1.4Delta2-61 to K531Y hKv1.4Delta2-61 and raising of extracellular [K(+)](o) abolished Rg(3) inhibitions on non-inactivating outward currents. Rg(3) treatment increased the C-type inactivation rate, but raising the extracellular [K(+)](o) reversed Rg(3) action. These results provide additional evidence that K531 residue also plays an important role in the Rg(3)-mediated non-inactivating current blockages and in Rg(3)-mediated increase of the C-type inactivation rate in hKv1.4Delta2-61 channels.
Biological & Pharmaceutical Bulletin 05/2009; 32(4):614-8. · 1.66 Impact Factor
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ABSTRACT: Two-photon fluorescent probes for the cellular membrane, derived from 6-acyl-2-aminonaphthalene as the fluorophore and hexanoyl (CH), lauryl (CL), and stearyl (CS) groups as the receptor, have been synthesized. Their photophysical properties and utility as membrane probes were also studied. Whereas CH cannot be used as a membrane probe due to its high water solubility, CL and CS are useful two-photon probes for membrane lateral heterogeneity, as they can easily stain cells, emit fluorescence with high sensitivity to the environment polarity, and are capable of imaging the membrane lateral heterogeneity in live cells. Moreover, CS is more likely to be located in the plasma membrane due to its negligible water solubility. Our results show that the liquid ordered-like domain covers 31-35% of the cellular surface.
ChemBioChem 12/2008; 9(17):2830-8. · 3.94 Impact Factor
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ChemBioChem 10/2008; 9(17):2830 - 2838. · 3.94 Impact Factor
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ABSTRACT: 2-Acetyl-6-(dimethylamino)naphthalene-derived two-photon fluorescent Ca2+ probes (ACa1-ACa3) are reported. They can be excited by a 780 nm laser beam, show 23-50-fold enhancement in one- and two-photon excited fluorescence in response to Ca2+, emit fourfold stronger two-photon excited fluorescence than Oregon Green 488 BAPTA-1 upon complexation with Ca2+, and can selectively detect intracellular free Ca2+ ions in live cells and living tissues with minimum interference from other metal ions and membrane-bound probes. Moreover, these probes are capable of monitoring calcium waves at a depth of 120-170 microm in live tissues for 1100-4000 s using two-photon microscopy with no artifacts of photobleaching.
Chemistry 02/2008; 14(7):2075-83. · 5.93 Impact Factor
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Angewandte Chemie International Edition 02/2007; 46(39):7445-8. · 13.45 Impact Factor
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Angewandte Chemie International Edition 01/2007; 46(19):3460-3. · 13.45 Impact Factor
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ABSTRACT: Relative transcript levels of eight rice diterpene cyclases at the branch points of gibberellins and phytoalexins biosynthesis pathway were measured by reverse transcription quantitative PCR. Metabolic flux analysis by the distribution ratio of common substrate showed that UV-irradiation of etiolated rice seedlings decreased the flux for primary metabolism of gibberellins biosynthesis by half (from 62 to 27%) and 41% of geranylgeranyl pyrophosphate was used for induction of pimaradiene intermediate as the major phytoalexin. In comparison, light-illumination used almost all geranylgeranyl pyrophosphate (96%) for gibberellin biosynthesis to stimulate the plant growth and strongly repressed the metabolic flux for phytoalexins biosynthesis.
Biotechnology Letters 10/2005; 27(18):1375-80. · 1.68 Impact Factor
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ABSTRACT: 1-Deoxy-D-: xylulose-5-phosphate synthase (DXS) encoded by a multigene family in plants, catalyzes the first step in the methylerythritol 4-phosphate (MEP) pathway. Three rice DXS-related sequences (OsDXS) were identified from available rice databases. The open reading frame of three OsDXS genes (dxs1, dxs2, and dxs3) were amplified against cDNA template. Ratio of their transcript levels in etiolated rice leaf was 9:181:1. While the expression levels were not changed along the growth stages of etiolated culture, UV-irradiation of the etiolated rice induced the expression of dxs3 up to nine-fold compared with that of unirradiated control. In the case of light-illumination, the relative expression of dxs1 based on unilluminated control increased two-fold. The differential expression of three OsDXS genes suggested their distinct and complementary roles in the control of the first step of the MEP pathway in response to environmental stimuli.
Biotechnology Letters 08/2005; 27(14):997-1001. · 1.68 Impact Factor
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ABSTRACT: Reverse transcription followed by RT Q-PCR is useful for the systematic measurement of changes in gene expression. RT Q-PCR with two pairs of primers for each gene was used for relative expression of three genes with high homology encoding 3-hydroxy-methylglutaryl-CoA reductase (HMGR) in rice. At various growth stages of etiolated seedling and various times after UV-irradiation treatment, RT Q-PCR of each HMGR gene showed a consistent pattern of relative expression with the RT Q-PCR data, using two pairs of primers, giving a high degree of accuracy. Furthermore, the different expression levels of three HMGR genes in a sample were determined by diluting the cDNA concentration. These results indicate that RT Q-PCR with only one pair of primers for a gene can quantify the relative expression and that the high expression level of HMGR2 could be quantified in comparison to the low level of HMGR1 expression.
Biotechnology Letters 07/2004; 26(12):985-8. · 1.68 Impact Factor
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ABSTRACT: Reverse transcription followed by real-time quantitative polymerase chain reaction (RT Q-PCR) is useful for the systematic measurement of plant physiological changes in gene expression. The validity of using 18S rRNA and three housekeeping genes, glyceraldehyde-3-phosphate dehydrogenase, actin, and tubulin, was tested as a reference of RT Q-PCR. Under various growth stages of etiolated seedlings, different cultivars, and various times after UV-irradiation treatment, expression level of 18S rRNA correlated with total RNA suggesting the uniformity of RT Q-PCR efficiencies among samples. Relative expressions of housekeeping genes varied among samples and independently of experimental conditions, up to two-fold, signifying generally constant fraction of mRNA in total RNA. Results indicate 18S rRNA was the most reliable reference gene for RT Q-PCR of total RNA.
Biotechnology Letters 12/2003; 25(21):1869-72. · 1.68 Impact Factor
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ABSTRACT: The use of X-ray computed microtomography (PCT) has increased in biomedical research and industrial applications. The inherited high quality of synchrotron radiation (SR) X-rays including high flux, collimation, and coherence, has been used recently to produce radiographic images with high spatial resolution and contrast. A simple and stable imaging system using an unmonochromatized SR source based on the principle of phase contrast X-ray imaging consists of a charge-coupled device (CCD) detector coupled with an optical lens system at the Pohang Light Source (PLS) 5C1 beamline. The spatial resolution of the imaging system was determined using the modulation transfer function (MTF), which was measured by step-by step calculations obtained from sharp edge images. Projection image data were obtained at 250 steps over 180 degrees of rotation with an acquisition time, depending on the imaged object materials, of 30 to 150 ms per projection image. The tomographic images were reconstructed using a simple filtered backprojection algorithm to reconstruct two-dimensional (2-D) images using projection data which may include characteristics of beam collimation and phase contrast. Although the use of a monochromatic X-ray beam has previously demonstrated to provide high resolution and enhanced contrast, our approach uses an unmonochromatized SR X-ray beam and shows similar image capability, without the needs for sophisticated X-ray optics, in an exposure time which is significantly less, by two orders of magnitude, than that for the monochromatic SR system. The current PLS 5C1 SR imaging system can produce projection images at a spatial resolution of 8.3 μm over a field of view of about 5 mm at an exposure time of 30 ms per projection image for 1.5 × optical magnification. This study presents the results of SR μCT images of cancerous human breast tissue containing microcalcifications, mouse lumbar vertebra, and mouse coccygeal vertebra. The unmonochromatized SR μCT imaging system provides an effective means of evaluating microstructures, not only in biomedical specimens but also in inorganic samples.
IEEE Transactions on Nuclear Science 11/2002; · 1.45 Impact Factor
