Biomedical subjects

T Iwata

Publications and source records attributed to T Iwata.

At least 73 records · Page 4Linked to original sources

PubMed2000

Water structural changes involved in the activation process of photoactive yellow protein.

Fourier transform infrared (FTIR) spectroscopy was applied to the blue-light photoreceptor photoactive yellow protein (PYP) to investigate water structural changes possibly involved in the photocycle of PYP. Photointermediates were stabilized at low temperature, and difference IR spectra were obtained between intermediate states and the original state of PYP (pG). Water structural changes were never observed in the >3570 cm(-)(1) region for the intermediates stabilized at 77-250 K, such as the red-shifted pR and blue-shifted pB intermediates. In contrast, a negative band was observed at 3658 cm(-)(1) in the pB minus pG spectrum at 295 K, which shifts to 3648 cm(-)(1) upon hydration with H(2)(18)O. The high frequency of the O-H stretch of water indicates that the water O-H group does not form hydrogen bonds in pG, and newly forms these upon pB formation at 295 K, but not at 250 K. Among 92 water molecules in the crystal structure of PYP, only 1 water molecule, water-200, is present in a hydrophobic core inside the protein. The amide N-H of Gly-7 and the imidazole nitrogen atom of His-108 are its possible hydrogen-bonding partners, indicating that one O-H group of water-200 is free to form an additional hydrogen bond. The water band at 3658 cm(-)(1) was indeed diminished in the H108F protein, which strongly suggests that the water band originates from water-200. Structural changes of amide bands in pB were much greater in the wild-type protein at 295 K than at 250 K or in the H108F protein at 295 K. The position of water-200 is >15 A remote from the chromophore. Virtually no structural changes were reported for regions larger than a few angstroms away from the chromophore, in the time-resolved X-ray crystallography experiments on pB. On the basis of the present results, as well as other spectroscopic observations, we conclude that water-200 (buried in a hydrophobic core in pG) is exposed to the aqueous phase upon formation of pB in solution. In neither crystalline PYP nor at low temperature is this structural transition observed, presumably because of the restrictions on global structural changes in the protein under these conditions.

Bacterial Proteins↗

PubMed2000

A neonatal lethal mutation in FGFR3 uncouples proliferation and differentiation of growth plate chondrocytes in embryos.

We have generated the first mouse model of fibro-blast growth factor receptor 3 (Fgfr3) with the K644E mutation, which accurately reflects the embryonic onset of a neonatal lethal dwarfism, thanatophoric dysplasia type II (TDII). Long-bone abnormalities were identified as early as embryonic day 14, during initiation of endochondral ossification. Increased expression of PATCHED: (PTC:) was observed, independent of unaltered expression of parathyroid hormone-related peptide (PTHrP) receptor and Indian Hedgehog (IHH:), suggesting a new regulatory role for Fgfr3 in embryos. We demonstrate that the mutation enhances chondrocyte proliferation during the early embryonic skeletal development, in contrast to previous reports that showed decreased proliferation in postnatal-onset dwarf mice with activating Fgfr3 mutations. This suggests that signaling through Fgfr3 both promotes and inhibits chondrocyte proliferation, depending on the time during development. In contrast, suppressed chondrocyte differentiation was observed throughout the embryonic stages, defining decreased differentiation as the primary cause of retarded longitudinal bone growth in TDII. This model was successfully crossed with a cartilage-specific CRE: transgenic strain, excluding the lung as the primary cause of lethality.

Amino Acid Substitution↗

PubMed2000

Helicity dependence of gammap --> Npi below 450 MeV and contribution to the Gerasimov-Drell-Hearn sum rule.

The helicity dependence of the single pion photoproduction on the proton has been measured in the energy range from 200 to 450 MeV for the first time. The experiment, performed at the Mainz microtron MAMI, used a 4pi-detector system, a circularly polarized, tagged photon beam, and a frozen-spin target. The data obtained provide new information for multipole analyses of pion photoproduction and determine the main contributions to the Gerasimov-Drell-Hearn sum rule and the forward spin polarizability gamma(0).

Journal Article↗

PubMed2000

Association between elevated brain natriuretic peptide levels and the development of left ventricular hypertrophy in patients with hypertension.

PURPOSE: To examine whether plasma levels of brain natriuretic peptide identify hypertensive patients at risk for progressive cardiac hypertrophy. SUBJECTS AND METHODS: We examined the association between plasma brain natriuretic peptide levels and left ventricular structural changes in 54 hypertensive patients and 28 normotensive control subjects. Patients were divided into those with elevated (n = 14) or normal (n = 40) levels of brain natriuretic peptide, based on a cutoff level of 41 pg/mL (2 SD above the mean in the control subjects). Left ventricular function and geometry were assessed echocardiographically at baseline and follow-up. RESULTS: At baseline, initial left ventricular chamber size, wall thickness, and systolic function did not differ between the hypertensive patients and normotensive subjects. After a mean (+/- SD) follow-up of 9 +/- 3 months, blood pressure was relatively unchanged in the hypertensive patients with normal brain natriuretic peptide levels, whereas there were significant increases in systolic blood pressure and pulse pressure (both P <0.05 versus baseline) in patients with elevated brain natriuretic peptide levels. Moreover, left ventricular midwall systolic function had decreased significantly at follow-up in those with elevated levels (P <0.05 versus baseline). At follow-up, the hypertensive patients with elevated brain natriuretic peptide levels had a significantly greater left ventricular mass index and relative wall thickness than those with normal levels. Multiple regression analyses determined that only initial plasma brain natriuretic peptide was significantly (P <0.01) associated with subsequent left ventricular hypertrophy. CONCLUSION: Plasma brain natriuretic peptide levels may identify hypertensive patients who are likely to have progressive cardiac hypertrophy.

Aged↗

PubMed2000

Silefrin, a sodefrin-like pheromone in the abdominal gland of the sword-tailed newt, Cynops ensicauda.

Sodefrin-like female-attracting pheromone was purified from the abdominal glands of male sword-tailed newts, Cynops ensicauda, by gel-filtration chromatography and reversed-phase high-performance liquid chromatography. The final product comprises 10 amino acid residues with the sequence SILSKDAQLK which coincided with the sequence deduced from its precursor cDNA. This peptide was designated silefrin. The sequence of silefrin was different from that of sodefrin by two amino acid residues, with substitutions Leu for Pro and Gln for Leu at positions 3 and 8, respectively. Both native and synthetic silefrin exerted an equipotent activity in attracting conspecific females.

Animals↗

PubMed2000

Differential gene expression in male and female rat lenses undergoing cataract induction by transforming growth factor-beta (TGF-beta).

Epidemiologic studies in humans as well as immunohistologic studies in animals have demonstrated significant sex differences in the propensity to develop cataract. Several studies suggest that estrogen may play a protective role against cataractogenesis. Indeed, male and ovariectomized female rat lenses have a greater susceptibility to cataract induced by transforming growth factor-beta (TGF-beta) than do normal female lenses. However, in spite of the current evidence that estrogen may play a pivotal role in cataractogenesis, the molecular mechanisms behind this phenomenon are largely undetermined. Our study utilized the differential display procedure to examine gene up- and down-regulation in male, normal female and ovariectomized female rat lenses exposed to TGF-beta. Male and normal female rat lenses were cultured with or without 0.15 ng ml(-1)TGF-beta. Lenses were then harvested, and total RNA was isolated for analysis by reverse-transcriptase differential display. Differentially expressed mRNAs were subcloned, sequenced and identified through GenBank database searches. The original experiment was repeated with the addition of ovariectomized female TGF-beta(+/-) conditions, and all differential patterns of gene expression were verified using Northern blot and RT-PCR analysis. Screening of approximately 12% of the mRNA population led to the identification of 27 differentially expressed cDNAs. Notably, strong gender differences were found in expression levels of gammaB-crystallin. In addition, proteasome Z subunit was up-regulated in TGF-beta-treated male and ovariectomized female lenses, but was down-regulated in TGF-beta-treated normal female lenses. This pattern of expression is consistent with the increased susceptibility of male and ovariectomized lenses to TGF-beta-induced cataract. We conclude that differential display is a useful and expedient method for analysing changes in gene expression in the lens. Structural and functional studies of the genes identified in this study may further elucidate mechanisms underlying the TGF-beta-induced cataract formation and differential rates of cataractogenesis in males vs females. In particular, our data suggest that the role of proteasome Z subunit in TGF-beta-induced anterior subcapsular cataract warrants further investigation.

Animals↗

PubMed2000

Production of a recombinant newt growth hormone and its application for the development of a radioimmunoassay.

Complementary DNA (cDNA) encoding newt (Cynops pyrrhogaster) growth hormone (nGH) was cloned from a cDNA library constructed from mRNAs of newt pituitary glands and was expressed in Escherichia coli. Based on Northern blot analysis using the cDNA as a probe, the nGH mRNA was estimated to be 940 bases in length. The recombinant nGH (nGHr) had a molecular mass of 22 kDa as determined by SDS-PAGE and possessed considerable bioactivity as determined in a Xenopus cartilage assay. Using the nGHr, we produced a polyclonal antibody against nGHr. Western blot analysis of newt anterior pituitary gland homogenates revealed that this antiserum specifically detected a single 22-kDa band, and histological studies of newt pituitary gland sections showed that the cells that reacted immunologically by the anti-nGHr antiserum corresponded to those stained by an antiserum against rat GH. A radioimmunoassay (RIA) that is specific and sensitive for nGH was developed, employing the antiserum thus produced. The sensitivity of the RIA was 57 +/- 7 pg/100 microl assay buffer. Interassay and intraassay coefficients of variation were 1.22 and 2.70%, respectively. Serial dilutions of plasma and pituitary homogenate of C. pyrrhogaster yielded dose-response curves that were parallel to the standard curve. Plasma from hypophysectomized newts showed no cross-reactivity. Moreover, displacement curves obtained using pituitary homogenates of the sword-tailed newt (C. ensicauda) and the crested newt (Triturus carnifex) were also parallel to the standard curve. Mammalian and frog GHs and prolactins (PRLs), as well as newt PRL, showed no inhibition of binding, even at relatively high doses, in this RIA. The RIA was used to measure GH released from newt pituitaries in vitro. Enhancement of GH release by 10(-7) M thyrotropin-releasing hormone was observed in cultures of newt pituitaries.

Amino Acid Sequence↗

PubMed2000

Cloning of bullfrog thyroid-stimulating hormone (TSH) beta subunit cDNA: expression of TSHbeta mRNA during metamorphosis.

A thyroid-stimulating hormone beta subunit (TSHbeta) cDNA encoding both signal peptide and mature TSHbeta molecule was cloned from a cDNA library constructed from total RNA of the bullfrog (Rana catesbeiana) adenohypophysis. The bullfrog TSHbeta mRNA was estimated by Northern blot analysis to be approximately 1 kb. The deduced amino acid sequence showed 40-61% homologies with the sequences of TSH beta subunits of other vertebrates. Using the cDNA as a probe, we measured changes in mRNA expression in metamorphosing tadpoles of R. catesbeiana. The TSH beta subunit mRNA level increased progressively throughout prometamorphic stages, reaching its maximum at the end of prometamorphosis. The maximum level was maintained throughout early and mid climax, declining at late climax. These results, together with previously obtained data on plasma prolactin and pituitary prolactin mRNA levels, as well as thyroid hormone levels, are discussed in relation to metamorphic changes occurring in the bullfrog larvae.

Amino Acid Sequence↗

PubMed2000

Optic disk pallor and retinal atrophy in Sotos syndrome (cerebral gigantism).

PURPOSE: To report a case of Sotos syndrome (cerebral gigantism) with megalophthalmos, optic disk pallor, and retinal atrophy. METHODS: Case report. RESULTS: A 22-year-old man was diagnosed with Sotos syndrome with optic atrophy. His corneal diameters were 13 mm horizontally, and his optic disks were pale. His retinal vessels were attenuate in diameter, and small white spots were found on the retina. The results of a Goldmann visual field test were normal. His visually evoked potential to flash stimuli showed extended latent times and normal amplitudes. Keratometry was 40.5 diopters (8.33 mm) in both eyes. The axial length was 25.9 mm in the right eye and 25.4 mm in the left eye. CONCLUSION: Sotos syndrome may be associated with optic disk pallor and retinal atrophy.

Abnormalities, Multiple↗

PubMed2000

Hormonal control of urodele reproductive behavior.

Hormonal control of expression of courtship behavior and of development of structures related to the reproductive behavior in two species of Japanese newts, Cynops pyrrhogaster and Cynops ensicauda, was described. Prolactin (PRL) and androgen were essential factors for eliciting courtship behavior. In addition, arginine vasotocin markedly enhanced the expression of courtship behavior. PRL induced migration to water, in which courtship and oviposition take place, and converted the integument from the terrestrial type to the aquatic one. PRL also stimulated the growth of the tail fin, which was blocked by estrogen. Cellular and nuclear size and number of synapses on the somata of Mauthner cells, which are involved in tail movement, were also increased by PRL and androgen. Synthesis of sodefrin, a female-attracting pheromone, in the abdominal gland as well as that of mucopolysaccharides constituting the sac of sperm in the lateral gland was enhanced by PRL and androgen. Structural development of oviducts was elicited by estrogen or PRL to a certain extent, and full oviducal development by the combination of these two hormones, PRL being indispensable for the oviducal jelly secretion.

Animals↗

PubMed2000

Changes in vascular reactivity induced by acute hyperthyroidism in isolated rat aortae.

Hyperthyroidism was induced by subcutaneous injections of L-thyroxine (T(4)) (500 mg/kg/day) for 3 days in order to study whether adrenergic and muscarinic receptor-mediated vascular responses alter at an early stage of the disease. T(4) treatment was sufficient to induce a significant degree of thyroid weight loss, tachycardia, cardiac hypertrophy, and an elevation in serum T(4) levels. The tension of aortic ring preparations isolated from rats was measured isometrically to investigate the influence of acute hyperthyroidism. The contractions induced by norepinephrine (NE) were significantly suppressed in aortic rings from rats treated with T(4) compared with control rats. N(G)-nitro-L-arginine (L-NOARG), an inhibitor of nitric oxide synthase (NOS), significantly enhanced NE-induced contraction in aortic rings from both control and T(4)-treated rats, and the enhancement was greater in rats treated with T(4) than control rats. The relaxations induced by either acetylcholine (ACh) or sodium nitroprusside (SNP) were also significantly enhanced by T(4) treatment. L-NOARG abolished the relaxation induced by ACh in aortic rings from both control and T(4)-treated rats. L-NOARG shifted SNP-induced relaxation curves of aortic rings from those of control rats to the left, but not with rats treated with T(4). T(4) treatment showed no influence on the amount of endothelial NOS (eNOS) protein. These results suggest that vascular responses alter at an early stage of hyperthyroidism and that it may be due to a modification in the NO system which is independent from the amount of eNOS protein.

Acetylcholine↗

PubMed2000

Detection of Bruton's tyrosine kinase mutations in hypogammaglobulinaemic males registered as common variable immunodeficiency (CVID) in the Japanese Immunodeficiency Registry.

CVID is frequently diagnosed in male and female individuals with hypogammaglobulinaemia of unknown aetiology. To examine the possibility that sporadic male cases with X-linked agammaglobulinaemia (XLA), which is caused by mutations in the Bruton's tyrosine kinase (Btk) gene, might be misregistered as having CVID, we employed a flow cytometric test to identify XLA in hypogammaglobulinaemic males registered as CVID in the Japanese Immunodeficiency Registry. From 30 male cases registered as having CVID between 1992 and 1998, we selected 21 males with low or unreported peripheral B cell counts. Blood samples could be obtained from 11 patients and their mothers. Using flow cytometric analysis, the Btk-deficient status in monocytes was demonstrated in seven out of nine cases with decreased numbers of peripheral B cells. The diagnosis of XLA was confirmed in each of the seven patients by demonstration of Btk gene mutations in the patients or cellular mosaicism in the mother. This study demonstrates misregistration of XLA as CVID.

Agammaglobulinaemia Tyrosine Kinase↗

PubMed2000

Monoclonal antibodies against topoisomerase I suppressed DNA relaxation and HIV-1 cDNA synthesis.

Human immunodeficiency virus type 1 (HIV-1) virion is known to carry a number of cellular components including cellular topoisomerase I. Previously, we have demonstrated that topoisomerase I enhances HIV-1 cDNA synthesis in reverse transcription (RT) assays in vitro. In the present study, we have produced six monoclonal antibodies (MAbs) against human topoisomerase I. The MAbs suppressed nicking/closing of supercoiled DNA and cDNA synthesis in an endogenous reverse transcription (ERT) assay using a detergent-disrupted HIV-1 virion. Thus, the results suggest that topoisomerase I plays an important role in RNA-directed DNA polymerization.

Animals↗

PubMed2000

Effect of prolactin and androgen on the expression of the female-attracting pheromone silefrin in the abdominal gland of the newt, Cynops ensicauda.

Silefrin is a sodefrin-like, female-attracting pheromone comprising 10 amino acids that was isolated from the abdominal gland of the sword-tailed newt, Cynops ensicauda. Hormonal effects on the silefrin precursor mRNA expression and silefrin content in the abdominal gland were investigated in the present study by using Northern blot analysis and radioimmunoassay, respectively. In the abdominal gland of newts treated with prolactin (PRL) plus testosterone propionate (TP), silefrin precursor mRNA expression was markedly enhanced as compared with that in the newts injected with saline, PRL, or TP. Values for radioimmunoassayable silefrin content in the abdominal gland paralleled those for the silefrin precursor mRNA levels. Moreover, silefrin precursor mRNA signals, as revealed by in situ hybridization, as well as stainability of immunoreactive silefrin were much more intense in the epithelial cells of the abdominal gland of the PRL-plus-TP-treated animals than in those of controls. We thus conclude that PRL and androgen are important factors for enhancing silefrin synthesis.

Androgens↗

PubMed2000

Anaerobic bacterial extracts influence production of matrix metalloproteinases and their inhibitors by human dental pulp cells.

The role of human dental pulp (HDP) cells in extracellular matrix degradation in pulpitis is still unclear. In this study, the effects of sonicated bacterial extracts (SBEs) from Prevotella intermedia, Fusobacterium nucleatum, Porphyromonas endodontalis, and Porphyromonas gingivalis on the balance between the production of matrix metalloproteinases (MMPs) and that of their inhibitors [tissue inhibitors of metalloproteinases (TIMPs)] by HDP cells were examined. HDP cells were treated with SBEs, and their culture media were later harvested. MMP activities and TIMP concentrations were determined by use of independent measurement strategies and sensitive ELISAs. The production of MMP-1 and MMP-2 was accelerated by all SBE. On the other hand, TIMP-1 production was slightly elevated; and TIMP-2 production was markedly inhibited by all of the extracts. SBEs derived from these anaerobic bacteria seemed to affect the acceleration of extracellular matrix degradation activity by HDP cells. These findings suggest that HDP cells stimulated by bacterial byproducts may be involved in the pathogenesis of pulpitis.

Bacteria, Anaerobic↗

PubMed2000

Gonadotropin-releasing hormone analogue therapy for peritoneal inclusion cysts after gynecological surgery.

OBJECTIVE: The purpose of this report is to estimate the effectiveness of gonadotropin-releasing hormone analogue (Gn-RHa) therapy for peritoneal inclusion cysts. METHODS: Patients who had cystic masses that developed after gynecological surgery and were strongly suspected of being peritoneal inclusion cysts based on the results of ultrasound, magnetic resonance imaging (MRI), and tumor marker studies, were treated with a Gn-RHa (buserelin acetate or leuprorelin acetate). Buserelin acetate was administered at a dose of 900 microg/day, and leuprorelin acetate at a dose of 3.75 mg/month. Ultrasonography was performed in the outpatient clinic every 2 weeks after the start of administration to measure the diameter of cysts. RESULTS: We treated 8 patients with peritoneal inclusion cysts conservatively with a Gn-RHa. The cysts resolved in 7 of the patients. Two of them developed a recurrence, but further Gn-RHa therapy was followed by complete resolution of the cysts in one patient and we resume Gn-RHa therapy to the other patient. The peritoneal inclusion cysts failed to shrink in only one patient. CONCLUSION: It is suggested that Gn-RHa therapy is effective in some cases of peritoneal inclusion cysts.

Adult↗

PubMed2000

Bradykinin B(2) receptor gene polymorphism is associated with angiotensin-converting enzyme inhibitor-related cough.

The appearance of cough in association with angiotensin-converting enzyme (ACE) inhibitors is thought to be related to bradykinin, and it has been speculated that the elicitation of adverse effects is genetically predetermined. Several polymorphisms of the human bradykinin B(2) receptor gene may be involved in ACE inhibitor-related cough. To investigate this possibility, we identified the -58 thymine (T)/cytosine (C) polymorphism in subjects with ACE inhibitor-related cough. We classified the study population into 4 groups: subjects with and without cough that were treated with ACE inhibitors (n=30/30), nontreated essential hypertensive subjects (n=100), and normotensive subjects (n=100). The -58T/C was genotyped by the polymerase chain reaction single-strand conformation polymorphism method. The frequencies of the CC genotype and C allele of -58T/C were significantly higher in the nontreated hypertensive subjects than in the normotensive subjects. Conversely, the frequencies of the TT genotype and T allele were significantly higher in the subjects with cough than in the subjects without cough. These tendencies were more pronounced in females. Among the promoter assays of the human bradykinin B(2) receptor, -58T was found to have a higher transcription rate than that of -58C. This finding seems to suggest that the transcriptional activity of promoter might be involved in the appearance of ACE inhibitor-related cough. A genetic variant of the bradykinin receptor is involved in the elicitation of ACE inhibitor-related cough. It may be possible to predict the side effects of ACE inhibitors in advance.

Adult↗

PubMed2000

Involvement of the renal kallikrein-kinin system in furosemide-induced natriuresis in rats.

This study examined whether the renal kallikrein-kinin system (KKS) is involved with furosemide-induced natriuresis in rats. Intravenous administration of furosemide (10 mg/kg) to anesthetized rats infused with physiological saline (saline) increased renal KK excretion as well as urine volume and urinary excretions of sodium, chloride and potassium. The change in the increase of renal KK excretion by furosemide at a dose of 1.0 mg/kg relative to the control was larger than that of urine volume. Pretreatment with a B2-receptor antagonist, 8-[3-[N-[(E)-3-(6-acetamidopyridin-3-yl)acryloylglycyl]-N-methylamino]-2,6-dichlorobenzyloxy]-2-methylquinoline (FR173657, 100 mg/kg), significantly inhibited the furosemide-induced natriuresis by 58.6%. The effect of FR173657 on the furosemide-induced natriuresis was also examined in hypotonic saline-loading rats. Similar to the saline-loading rats, urinary excretion of sodium collected during the first 8 h in metabolic cages significantly reduced by 22.4% when FR173657 (100 mg/kg) was given concurrently with furosemide (100 mg/kg) and hypotonic saline (5% of body wt.). These results indicate that furosemide increased renal KK excretion through a mechanism different from a washout mechanism and induced natriuresis partly through an augmentation of the renal KKS following the increase in renal KK excretion in both the saline- and hypotonic saline-loading rats.

Animals↗