Biomedical subjects

H Kobayashi

Publications and source records attributed to H Kobayashi.

At least 145 records · Page 8Linked to original sources

PubMed2001

New epidermal keratin genes from Xenopus laevis: hormonal and regional regulation of their expression during anuran skin metamorphosis.

Xenopus larval keratin (XLK) was isolated by gel electrophoresis of proteins of tadpole skin. Screening of an expression cDNA library of tail tissues by specific polyclonal antibodies against XLK produced XLK cDNA (xlk). Its complete nucleotide and predicted amino acid sequences revealed that XLK was a new member of type II keratin. Screening of a cDNA library of adult Xenopus skin using an oligonucleotide probe which had been designed from well-conserved N-terminal amino acid sequences of the rod domain of type I keratin produced two cDNAs, xak-a and xak-b, which were found to be new members of type I keratin gene. Northern blot analysis showed that xlk was expressed exclusively in the larval skin whereas xak-a and xak-b were expressed exclusively in the adult skin. Their expression level was regulated in a region- and metamorphic stage- dependent manner during larval skin development. mRNA in situ hybridization experiments identified the cells that expressed xlk, and xak-a and xak-b as larva- specific epidermal cells (skein cells and basal cells), and adult suprabasal epidermal cells, respectively. These three genes were found to be late responsive to thyroid hormone. Phylogenetic relationships of these keratins with known ones are discussed.

Amino Acid Sequence↗

PubMed2001

The N-terminal helix of Xenopus cyclins A and B contributes to binding specificity of the cyclin-CDK complex.

Mitotic cyclins A and B contain a conserved N-terminal helix upstream of the cyclin box fold that contributes to a significant interface between cyclin and cyclin-dependent kinase (CDK). To address its contribution on cyclin-CDK interaction, we have constructed mutants in conserved residues of the N-terminal helix of Xenopus cyclins B2 and A1. The mutants showed altered binding affinities to Cdc2 and/or Cdk2. We also screened for mutations in the C-terminal lobe of CDK that exhibited different binding affinities for the cyclin-CDK complex. These mutations were at residues that interact with the cyclin N-terminal helix motif. The cyclin N-terminal helix mutations have a significant effect on the interaction between the cyclin-CDK complex and specific substrates, Xenopus Cdc6 and Cdc25C. These results suggest that the N-terminal helix of mitotic cyclins is required for specific interactions with CDKs and that to interact with CDK, specific substrates Cdc6 and Cdc25C require the CDK to be associated with a cyclin. The interaction between the cyclin N-terminal helix and the CDK C-terminal lobe may contribute to binding specificity of the cyclin-CDK complex.

Amino Acid Sequence↗

PubMed2001

Transplantation of infantile bladder in rats: an alternative procedure for bladder augmentation.

BACKGROUND: Our purpose was to evaluate whether bladder transplantation (BTx) can be used for bladder augmentation (BA). METHODS: Bladders from infantile Brown-Norway rats (less than 21 days old) were excised and each transplanted into a pouch created in the distal omentum of a 6-week-old Lewis rat (fully allogeneic BTx). No immunosuppressant was used in group I (n=12). Intramuscular FK506 was used daily from the day of BTx in group II (n=16; 0.2 mg/kg), group III (n=22; 0.6 mg/kg), and group IV (n=16; 1.2 mg/kg) until harvesting 3, 4, 5, or 6 weeks after BTx. FK506 was used for only 2 weeks in group V (n=12; 0.6 mg/kg/day) and group VI (n=12; 1.2 mg/kg/day). Syngeneic bladder transplants acted as controls (n=16). Hematoxylin and eosin staining was used to examine all grafts. In six rats from group III, BA was performed by anastomosing the graft to the recipient bladder 10 days after BTx. RESULTS: Each successfully transplanted graft appeared macroscopically as a thin-walled cyst. Rejection was seen in all grafts from groups I, II, V, and VI, and was minimal or absent in groups III and IV. On medium to long-term follow-up the only side effect of FK506 observed was reduced weight gain. Graft survival in the control group was 100%. BA was successful in all six cases, and the mucosa was normal throughout each augmented bladder. CONCLUSIONS: This is the first report of the successful transplantation of infantile tissue without vascular anastomosis. Because of the efficient, safe immunosuppression possible with FK506, our BTx technique could find clinical application for creating viable vesical tissue that could be used for BA.

Animals↗

PubMed2001

Characterization of binding properties of urinary trypsin inhibitor to cell-associated binding sites on human chondrosarcoma cell line HCS-2/8.

Urinary trypsin inhibitor (UTI) forms membrane complexes with UTI-binding proteins (UTI-BPs) and initiates modulation of urokinase-type plasminogen activator (uPA) expression, which results in UTI-mediated suppression of cell invasiveness. It has been established that suppression of uPA expression and invasiveness by UTI is mediated through inhibition of protein kinase C-dependent signaling pathways and that human chondrosarcoma cell line HCS-2/8 expresses two types of UTI-BPs; a 40-kDa UTI-BP (UTI-BP(40)), which is identical to link protein (LP), and a 45-kDa UTI-BP (UTI-BP(45)). Here we characterize binding properties of UTI-BPs.UTI complexes in the cells. In vitro ligand blot, cell binding and competition assays, and Scatchard analyses demonstrate that both UTI-BP(40) and UTI-BP(45) bind (125)I-UTI. A deglycosylated form of UTI (NG-UTI), from which the chondroitin-sulfate side chain has been removed, binds only to UTI-BP(40). Additional experiments, using various reagents to block binding of (125)I-UTI and NG-UTI to the UTI-BP(40) and UTI-BP(45) confirm that the chondroitin sulfate side chain of UTI is required for its binding to UTI-BP(45). Analysis of binding of (125)I-UTI and NG-UTI to the cells suggests that low affinity binding sites are the UTI-BP(40) (which can bind NG-UTI), and the high affinity sites are the UTI-BP(45). In addition, UTI-induced suppression of phorbol ester stimulated up-regulation of uPA is inhibited by reagents that were shown to prevent binding of UTI to the 40- and 45-kDa proteins. We conclude that UTI must bind to both of the UTI-BPs to suppress uPA up-regulation.

Binding Sites↗

PubMed2001

Aquaporin subtypes in rat cerebral microvessels.

We investigated the expression of aquaporin (AQP) subtypes in the rat cerebral microvessels by reverse transcription-polymerase chain reaction, immunoblotting and immunohistochemistry. mRNA for AQP4, but not for AQP1, 2, 3 or 5, was detected in the microvessels. Immunoblot analysis showed that AQP4 protein was detected as a 30 kDa band with higher molecular weight bands. Immunohistochemical staining showed that AQP4 was located on cell surface of the cerebral microvessels. These results suggest that AQP4 in the cerebral microvessels is involved in the regulation of water transport between blood and brain.

Animals↗

PubMed2001

Adrenomedullin receptors in rat choroid plexus.

To characterize transmembrane signaling of adrenomedullin (AM) in the choroid plexus, we studied the effects of AM on cyclic AMP (cAMP) and cyclic GMP (cGMP) levels as well as expression of mRNA for AM receptor in the rat choroid plexus slices. AM or calcitonin gene-related peptide (CGRP) increased cAMP (but not cGMP) level in a concentration-dependent manner, with AM being much more potently than CGRP. AM mRNA as well as calcitonin-receptor-like receptor mRNA and receptor-activity-modifying protein 2 mRNA, were highly expressed in the choroid plexus. Our biochemical and pharmacological studies may raise the possibility that choroid plexus secretes AM into the cerebrospinal fluid, and AM regulates choroid plexus function in an autocrine/paracrine manner via acting on AM-specific receptors.

Adrenomedullin↗

PubMed2001

A novel antiferromagnetic organic superconductor kappa-(BETS)(2)FeBr(4) [where BETS = bis(ethylenedithio)tetraselenafulvalene].

The electrical and magnetic properties of kappa-(BETS)(2)FeBr(4) salt [where BETS = bis(ethylenedithio)tetraselenafulvalene] showed that this system is the first antiferromagnetic organic metal at ambient pressure (T(N) = 2.5 K). The characteristic field dependence of the magnetization at 2.0 K indicates a clear metamagnetic behavior. The small resistivity drop observed at T(N) clearly shows the existence of the interaction between pi metal electrons and localized magnetic moments of Fe(3+) ions. In addition, this system underwent a superconducting transition at 1.1 K. That is, kappa-(BETS)(2)FeBr(4) is the first antiferromagnetic organic metal exhibiting a superconducting transition below Néel temperature. The magnetic field dependence of the superconducting critical temperature indicated that the superconductivity in this system is strongly anisotropic also in the conduction plane because of the existence of the metamagnetically induced internal field based on the antiferromagnetic ordering of the Fe(3+) 3d spins in contrast to the cases of the other conventional organic superconductors. Furthermore, the specific heat measurement exhibited a lambda-type large peak of zero-field specific heat corresponding to the three-dimensional antiferromagnetic ordering of high-spin Fe(3+) ions. The lack of distinct anomaly in the C(p) vs T curve at T(c) suggests the coexistence of the superconductivity and the antiferromagnetic order below T(c).

Journal Article↗

PubMed2001

A three-dimensional synthetic metallic crystal composed of single-component molecules.

Molecular metals normally require charge transfer between two different chemical species. We prepared crystals of [Ni(tmdt)2] (tmdt, trimethylenetetrathiafulvalenedithiolate) and carried out crystal structure analyses and resistivity measurements. The analyses and measurements revealed that these single-component molecular crystals are metallic from room temperature down to 0.6 kelvin. Ab initio molecular orbital calculations suggested that pi molecular orbitals form conduction bands. The compact molecular arrangement, intermolecular overlap integrals of the highest occupied and lowest unoccupied molecular orbitals, and tight-binding electronic band structure calculation revealed that [Ni(tmdt)2] is a three-dimensional synthetic metal composed of planar molecules.

Journal Article↗

PubMed2001

3D-micro-MR angiography of mice using macromolecular MR contrast agents with polyamidoamine dendrimer core with reference to their pharmacokinetic properties.

Four novel macromolecular MRI contrast agents, all of which had the same chemical composition but different molecular weights, were prepared using generation-3, -4, -5, and -6 polyamidoamine (PAMAM) dendrimers conjugated with a bifunctional diethylenetriaminepentaacetic acid derivative to change the blood retention, tissue perfusion, and excretion. Size-dependent changes in the pharmacokinetics were observed in the biodistribution study. (153)Gd-labeled generation-6 PAMAM-conjugates remained in the blood significantly longer than all of the other preparations (P < 0.001). The increase in blood-to-organ ratio of the preparations was found to correlate with increasing molecular size (P < 0.001). Additionally, 3D-micro MR images and angiography of mice of high quality and detail were obtained using PAMAM-(1B4M-Gd)x as a macro-molecular MRI contrast agent with a 1.5-T clinical MRI instrument. Numerous fine vessels of approximately 200 microm diameter were visualized on subtracted 3D-MR angiographms with G6D-(1B4M-Gd)(192). The quality of the images was sufficient to estimate the microvasculature of cancerous tissue for anti-angiogenesis therapy and to investigate knockout mice. Magn Reson Med 45:454-460, 2001.

Animals↗

PubMed2001

In vivo evaluation of a new sealant material on a rat lung air leak model.

The ability of an albumin-based hydrogel sealant (ABHS) to prevent air leakage through the suture line after pulmonary surgery was evaluated by comparison with that of a fibrin glue (FG). As an air-leak model, a rat lung was used in which a standard incision was made and the burst pressure for ABHS and FG was measured. The average burst pressures at time 0 for the FG and ABHS groups were 30.8+/-15.2 and 77.5 +/-19.1 mmHg, respectively. At Day 3, the value of ABHS (76.3 +/- 15.8 mmHg) was still significantly higher (P<0.05) than that of FG (60.0 +/- 21.9 mmHg). At Day 7, no statistical difference was observed between the FG group(71.2 +/- 18.6 mmHg) and the ABHS group(88.8 +/- 11.7 mmHg). Histological examination of the incision at Day 14 revealed that neither sealant was not visible at the incision site, and there was no evidence of adverse tissue reaction. It was concluded that ABHS had good sealing properties and is an alternative to FG for air leakage treatment in pulmonary surgery.

Air↗

PubMed2001

Micro-MR angiography of normal and intratumoral vessels in mice using dedicated intravascular MR contrast agents with high generation of polyamidoamine dendrimer core: reference to pharmacokinetic properties of dendrimer-based MR contrast agents.

Pharmacokinetic characteristics of intravascular macromolecular magnetic resonance imaging (MRI) contrast agents with polyamidoamine dendrimer cores smaller than generation-7 were previously studied in the literature. To evaluate the effects of greater hepatic uptake on the pharmacokinetics of the larger generation dendrimers, the MRI contrast agents GxD-(1B4M-Gd)(2(x+2)) were synthesized with generation-7, -8, and -9 polyamidoamine dendrimers and 2-(p-isothiocyanatobenzyl)-6-methyl-diethylenetriaminepentaacetic acid (1B4M). Their pharmacokinetic characteristics in mice were compared with that of G6D-(1B4M-Gd)(256). In biodistribution and dynamic micro-MRI studies, significantly less renal accumulation of G7D-(1B4M-Gd)(512), G8D-(1B4M-Gd)(1024), and G9D-(1B4M-Gd)(2048) was shown compared to G6D-(1B4M-Gd)(256) (P < 0.01). There was a significantly greater accumulation of G8D-(1B4M-Gd)(1024) and G9D-(1B4M-Gd)(2048) in the liver compared to G6D-(1B4M-Gd)(256) and G7D-(1B4M-Gd)(512) (P < 0.01). The highest blood retention of all dendrimer-based MRI contrast agents was exhibited by G7D-(1B4M-Gd)(512) (P < 0.01). The normal and intratumoral fine vessels of approximately 100 microm diameter were visualized in normal or tumor-bearing mice by high resolution three-dimensional-micro-MR angiographs with G7D-(1B4M-Gd)(512) and G8D-(1B4M-Gd)(1024) with good vessel-to-soft tissue contrast. In summary, increased accumulation in the liver with concomitant decreased uptake in the kidney was caused by increased molecular sizes of the dendrimer-based MRI contrast agents.

Analysis of Variance↗

PubMed2001

Improvement of macromolecular clearance via lymph flow in hamster gingiva by low-power carbon dioxide laser-irradiation.

BACKGROUND AND OBJECTIVE: Although therapeutic effects of low-power laser-irradiation on periodontal disease have been reported, little is known about the biological effects of laser-irradiation in the gingiva. Recently we reported that topical warming stimulated macromolecular clearance via lymph flow in hamster gingiva. This study was conducted to investigate whether low-power laser-irradiation affects macromolecular clearance via the lymph flow in the gingiva. STUDY DESIGN/MATERIALS AND METHODS: We injected 14C-methylated albumin into the mandibular gingiva of anesthetized hamsters followed by topical carbon dioxide (CO2) laser-irradiation (30 seconds, 0.5-1.5 W). We measured the clearance of radiolabeled albumin from the gingiva and its drainage into submandibular lymph nodes during 10 minutes. RESULTS: The clearance of the radiolabeled albumin from the gingiva and the influx into the submandibular lymph nodes were increased by CO2 laser-irradiation. CONCLUSION: Low-power CO2 laser-irradiation improves macromolecular clearance via the lymph flow in hamster gingiva.

Albumins↗

PubMed2001

Novel intravascular macromolecular MRI contrast agent with generation-4 polyamidoamine dendrimer core: accelerated renal excretion with coinjection of lysine.

One of the major limitations to macromolecular MRI contrast agents (MRI-CAs) is their slow clearance and associated decreased excretion of gadolinium (Gd(III)). The effect of coinjecting lysine to accelerate renal excretion of a macromolecular MRI-CA (generation-4 PAMAM dendrimer (G4D-(1B4M-Gd)64)) was investigated. The biodistribution and urine and fecal excretion in athymic mice was evaluated with and without lysine coinjection. 3D-dynamic-micro-MRI with G4D-(1B4M-Gd)64 was obtained with and without lysine coinjection, and the serial signal intensity (SI) change in the blood and organs was evaluated. When lysine was coinjected, urinary excretion of G4D-(1B4M-Gd)64 increased 5.4-fold compared to that without lysine, resulting in decreased renal accumulation of G4D-(1B4M-Gd)64 from 150% to 40% injected dose per gram (P < 0.001). On dynamic MRI with G4D-(1B4M-Gd)64, when lysine was coinjected, the kidney-to-blood SI ratio was significantly lower than that obtained without lysine (P < 0.001). When lysine was coinjected, the G4D-(1B4M-Gd)64 was excreted from the kidney intact.

Animals↗

PubMed2001

3D MR angiography of intratumoral vasculature using a novel macromolecular MR contrast agent.

Noninvasive methods to visualize blood flow in the intratumoral vasculature have not previously been studied. In the present study, the use of a novel intravascular MR contrast agent with a generation-6 polyamidoamine dendrimer core (G6-(1B4M-Gd)192; MW: 175kD) was investigated, and the vasculature in experimental tumors was visualized using 3D MR angiography (MRA). Xenografted tumors in nude mice of two different histologies-KT005 (human osteogenic sarcoma) and LS180 (human colon carcinoma)-were used to obtain 3D MRA using G6-(1B4M-Gd)192 and Gd-DTPA. The contrast MR sectional images were correlated with the corresponding histological sections. The intratumoral vasculature in the KT005 tumor was clearly visualized by 3D MRA, which became more evident with the growth of the tumor xenograft. In contrast, the intratumoral vasculature in the LS180 tumor was sparser and much less developed than that in KT005 tumors. Blood vessels with a diameter as small as 100 microm based on histology were visualized using 0.033 mmol Gd/kg of G6-(1B4M-Gd)192. In conclusion, intratumoral vasculature with a 100-microm diameter was visualized better using 3D MRA with G6-(1B4M-Gd)192 than with Gd-DTPA.

Animals↗

PubMed2001

Positive effects of polyethylene glycol conjugation to generation-4 polyamidoamine dendrimers as macromolecular MR contrast agents.

Macromolecules conjugated with polyethylene glycol (PEG) acquire more hydrophilicity, resulting in a longer half-life in circulation and lower immunogenicity. Two novel conjugates for MRI contrast agents were synthesized from a generation-4 polyamidoamine dendrimer (G4D), 2-(p-isothiocyanatobenzyl)-6-methyl-diethylenetriaminepentaacetic acid (1B4M), and one or two PEG molecules with a molecular weight of 20000 Da (PEG(2)-G4D-(1B4M-Gd)(62) (MW: 96 kD), PEG(1)-G4D-(1B4M-Gd)(63) (MW: 77 kD)). Their pharmacokinetics, excretion, and properties as vascular MRI contrast agents were evaluated and compared with those of G4D-(1B4M-Gd)(64) (MW: 57 kD). PEG(2)-G4D-(1B4M-Gd)(62) remained in the blood significantly longer and accumulated significantly less in the liver and kidney than the other two preparations (P < 0.01). Although the blood clearance was slower, PEG(2)-G4D-(1B4M-Gd)(62) was excreted more readily without renal retention than the other two preparations. In conclusion, the positive effects of PEG conjugation on a macromolecular MRI contrast agent were found to be prolonged retention in the circulation, increased excretion, and decreased accumulation in the organs.

Animals↗

PubMed2001

Novel liver macromolecular MR contrast agent with a polypropylenimine diaminobutyl dendrimer core: comparison to the vascular MR contrast agent with the polyamidoamine dendrimer core.

As MRI contrast agents, more hydrophobic molecules reportedly accumulate in the liver and thus are potentially useful as liver MRI contrast agents. In this study, a generation-4 polypropylenimine diaminobutane dendrimer (DAB-Am64), which is expected to be more hydrophobic than the generation-4 polyamidoamine dendrimer (PAMAM-G4D), was used to synthesize a conjugate with 2-(p-isothiocyanatobenzyl)-6-methyl-diethylenetriaminepentaacetic acid (1B4M) [DAB-Am64-(1B4M-Gd)(64)] for complexing Gd(III) ions. This DAB conjugate quickly accumulated in the liver and its characteristics were studied and compared with those of a PAMAM conjugate [PAMAM-G4D-(1B4M-Gd)(64)], which is known to be a useful vascular MRI contrast agent, in regard to its availability as a liver MRI contrast agent. DAB-Am64-(1B4M-Gd)(64) accumulated significantly more in the liver and less in blood than PAMAM-G4D-(1B4M-Gd)(64) (P < 0.001). Contrast-enhanced MRI with DAB-Am64-(1B4M-Gd)(64) was able to homogeneously enhance liver parenchyma and visualize both portal and hepatic veins of 0.5 mm diameter in mice. In conclusion, DAB-Am64-(1B4M-Gd)(64) is a good candidate for a liver MRI contrast agent.

Animals↗

PubMed2001

Pharmacokinetics and enhancement patterns of macromolecular MR contrast agents with various sizes of polyamidoamine dendrimer cores.

Four macromolecular contrast agents are synthesized to visualize small vessels by MRI using generation-3 (G3D), -4 (G4D), -5 (G5D), and -6 (G6D) polyamidoamine dendrimers conjugated to chelated gadolinium (Gd). The pharmacokinetics, enhancement patterns, and the ability of these constructs to visualize fine vessels is evaluated by dynamic MRI in relationship to their size. Gd-G6D and -G5D exhibit a prolonged high vascular (ventricular) signal intensity (SI) with high ventricle-to-organ SI ratios. The initial high vascular SI with Gd-G4D decreases to a value as low as that obtained with Gd-G3D and Gd-dimeglumine-diethylenetriaminepentaacetic acid (Gd-DTPA). Gd-G5D, -G4D, and -G3D show high renal SIs, and Gd-DTPA prominently enhances the skin. Gd-G6D and -G5D present fine vasculature significantly more clearly than Gd-G3D and -DTPA (P < 0.005). As the molecular size increases, the excretion of the 153Gd-conjugates is retarded. In conclusion, Gd-G6D and -G5D are retained in the blood and present fine vessels with high quality and detail, and should be adequate for visualizing small tumor vasculature.

Animals↗

PubMed2001

Unique morphology of the human eye and its adaptive meaning: comparative studies on external morphology of the primate eye.

In order to clarify the morphological uniqueness of the human eye and to obtain cues to understanding its adaptive significance, we compared the external morphology of the primate eye by measuring nearly half of all extant primate species. The results clearly showed exceptional features of the human eye: (1) the exposed white sclera is void of any pigmentation, (2) humans possess the largest ratio of exposed sclera in the eye outline, and (3) the eye outline is extraordinarily elongated in the horizontal direction. The close correlation of the parameters reflecting (2) and (3) with habitat type or body size of the species examined suggested that these two features are adaptations for extending the visual field by eyeball movement, especially in the horizontal direction. Comparison of eye coloration and facial coloration around the eye suggested that the dark coloration of exposed sclera of nonhuman primates is an adaptation to camouflage the gaze direction against other individuals and/or predators, and that the white sclera of the human eye is an adaptation to enhance the gaze signal. The uniqueness of human eye morphology among primates illustrates the remarkable difference between human and other primates in the ability to communicate using gaze signals.

Adaptation, Physiological↗