Search PubMed⌕ Search

Biomedical subjects

H Okamoto

Publications and source records attributed to H Okamoto.

At least 181 records · Page 10Linked to original sources
PubMed2000

Two zebrafish (Danio rerio) antizymes with different expression and activities.

Cellular polyamines are regulated by a unique feedback mechanism involving ornithine decarboxylase (ODC) antizyme. The synthesis of mammalian antizyme requires a programmed translational frameshift event induced by polyamines. Antizyme represses ODC, a key enzyme for polyamine synthesis, through accelerating enzyme degradation by the 26 S proteasome. Antizyme also inhibits the cellular uptake of polyamines. In the present study we isolated two distinct zebrafish (Danio rerio) antizyme cDNA clones (AZS and AZL) from an embryonic library. Their sequences revealed that both clones required translational frameshifting for expression. Taking account of +1 frameshifting, AZS and AZL products were 214 and 218 residues long respectively and shared 51.8% amino acid identity. In rabbit reticulocyte lysates, both mRNA species were translated through spermidine-induced frameshifting. The presence of the two antizyme mRNA species in embryos, adult fish and a cultured cell line was confirmed by Northern blot analysis. The ratio of AZS mRNA to AZL mRNA in the adult fish was 1.8-fold higher than in the embryos. Whole-mount hybridization in situ demonstrated that both mRNA species are expressed in every tissue in embryo, but predominantly in the central nervous system and the eyes. Bacterial expression products of both cDNA species inhibited ODC activity, but only the AZS product accelerated ODC degradation in vitro. These results show that both zebrafish antizymes are induced by polyamines but their mRNA species are expressed differently during development. The difference in activities on ODC degradation suggests their functional divergence.

Amino Acid Sequence↗
PubMed2000

Visualization of cranial motor neurons in live transgenic zebrafish expressing green fluorescent protein under the control of the islet-1 promoter/enhancer.

We generated germ line-transmitting transgenic zebrafish that express green fluorescent protein (GFP) in the cranial motor neurons. This was accomplished by fusing GFP sequences to Islet-1 promoter/enhancer sequences that were sufficient for neural-specific expression. The expression of GFP by the motor neurons in the transgenic fish enabled visualization of the cell bodies, main axons, and the peripheral branches within the muscles. GFP-labeled motor neurons could be followed at high resolution for at least up to day four, when most larval neural circuits become functional, and larvae begin to swim and capture prey. Using this line, we analyzed axonal outgrowth by the cranial motor neurons. Furthermore, by selective application of DiI to specific GFP-positive nerve branches, we showed that the two clusters of trigeminal motor neurons in rhombomeres 2 and 3 innervate different peripheral targets. This finding suggests that the trigeminal motor neurons in the two clusters adopt distinct fates. In future experiments, this transgenic line of zebrafish will allow for a genetic analysis of cranial motor neuron development.

Animals↗
PubMed2000

A novel apparatus for rat in vivo evaluation of dry powder formulations for pulmonary administration.

The lungs have attracted increasing attention as a site for administration of drugs, including macromolecules that are poorly absorbed from the intestine. There have been a number of basic studies in which peptide solutions were administered to experimental animals via the lungs. Although there have been several studies of pulmonary peptide absorption from dry powder formulations, a simpler and more inexpensive apparatus for administration of dry powders would enhance rapid screening of the formulations. In this study, we developed a simple apparatus to disperse dry powders. The apparatus has two 3-way stopcocks; one allows dispersal of powders at a constant pressure and airflow, and the other allows rats to breathe before and after administration. Dry powders of fluorescein (FL) and FITC-dextran (FD4) were manufactured by the spray-drying technique. The effects of operating conditions on the absorption of these model drugs were examined in rats. The C(max) for FL from dry powder was lower than that from solution and mean residence time was extended, suggesting that dissolution was the rate-determining step for FL absorption from dry powder. For FD4, the rate of absorption may not be regulated by dissolution but by epithelial transport. Absorption of insulin from spray-dried powder via the rat trachea was investigated using this apparatus. Intratracheally administered spray-dried insulin powder decreased plasma glucose level to a greater extent than spray-dried insulin solution administered via the same route. Thus, the apparatus is simple, inexpensive, and useful for rapid screening of dry powder formulations.

Absorption↗
PubMed2000

Switching characteristics of a model for biochemical-reaction networks describing autophosphorylation versus dephosphorylation of Ca2+/calmodulin-dependent protein kinase II.

Ca2+/calmodulin-dependent protein kinase II (CaMKII) has been suggested to participate in various cellular phenomena triggered by Ca2+ signalling. In the present study, we addressed the functional role of CaMKII in molecular-signal transduction in cells by mathematical modelling of putative biochemical-reaction networks thought to represent an essential part of molecular events responsible for CaMKII-related cellular phenomena. These networks include Ca2+/calmodulin-dependent threonine-286/287 (Thr286/287) autophosphorylation of CaMKII versus dephosphorylation of the enzyme. Computer simulation of the model was performed to examine the relation between the Ca2+-signalling pattern as an input and the resulting degree of Thr286/287 autophosphorylation (m) as an output. Under the simplified condition that the Ca2+ concentration during Ca2+ signalling was set to remain constant with time, the biochemical-reaction networks were shown to function as a switch. There is a threshold for gamma, a parameter representing the probability that the Thr286/287-dephosphorylated CaMKII subunit binds with the Ca2+/calmodulin complex; if gamma is above this threshold, m increases with time to a large degree (switch-on); otherwise, it remains near zero (switch-off). Mathematically, this sharp onset of m at the threshold can be accounted for by a change in the structure of the dynamic system describing the model, from bistability to monostability; this is analogous to the first-order phase transition in statistical physics. For the oscillatory time course of [Ca2+], switching characteristics were also shown with respect to the frequency and the maximum amplitude of the oscillation. These results suggest that graded information mediated by Ca2+ signalling is digitized into all-or-non information mediated by Thr286/287 autophosphorylation of CaMKII.

Animals↗
PubMed2000

Changes in left ventricular end-diastolic area, end-systolic wall stress, and fractional area change during anesthetic induction with propofol or thiamylal.

PURPOSE: To elucidate the mechanisms of the more profound hypotensive effects of propofol relative to thiamylal, we monitored changes in left ventricular (LV) preload, afterload, and contractility during the course of anesthetic induction with propofol and thiamylal. METHODS: Thirty-two patients (ASA I) were randomly assigned into two groups and injected with propofol (2 mg.kg(-1)) or thiamylal (4 mg.kg(-1)) as anesthetic induction agents. Transthoracic echocardiography (TTE) was used to assess LV performance before and during induction by the two anesthetics. The LV end-diastolic area (EDA) and LV end-systolic wall stress (ESWS) were used as indices of LV preload and LV afterload, respectively, while LV contractility was assessed by the fractional area change (FAC). RESULTS: Both propofol and thiamylal significantly reduced EDA and ESWS without significant change in FAC. Propofol-induced reductions in EDA and ESWS were significantly greater than those of thiamylal. CONCLUSION: The more profound hypotension observed during induction of anesthesia with propofol is due to the greater decrease in preload and afterload than with thiamylal, but not to a decrease in LV contractility.

Clinical Trial↗
PubMed2000

Thyroid lymphoma of mucosa-associated lymphoid tissue with monoclonal gammopathy occurring in an atomic bomb survivor: report of a case.

We describe herein a case of thyroid lymphoma of the mucosa-associated lymphoid tissue (MALT) with monoclonal gammopathy (IgG kappa), occurring in a woman who was exposed to atomic-bomb (A-bomb) radiation 1.5 km from the explosion of the hypocenter when she was 31 years old. Her estimated radiation dose was 0.2 Gy. A total thyroidectomy resulted in the disappearance of monoclonal gammopathy, thus indicating extramedullary monoclonal gammopathy of the thyroid. Three patterns of protein electrophoresis, i.e., normal, the appearance of monoclonal immunoglobulins, and their disappearance after the total thyroidectomy, were consecutively recorded. The utility of checking of the monoclonal immunoglobulin levels after a thyroidectomy due to lymphoma of the MALT to monitor recurrence is therefore suggested. This lymphoma is most likely not related to the patient's A-bomb radiation exposure.

Aged↗
PubMed2000

The entire nucleotide sequences of two distinct TT virus (TTV) isolates (TJN01 and TJN02) remotely related to the original TTV isolates.

TT virus (TTV) has a wide range of sequence divergence by which it is classified into at least 16 genotypes. A TTV isolate of genotype 12 (TJNO1) and another of genotype 13 (TJN02) were sequenced in the entire genome, and compared with the reported TTV isolates. TJN01 and TJN02 had genomic lengths of 3787 and 3794 nucleotides (nt), respectively, which were shorter by 66 and 59 nt than the prototype TTV isolate of genotype 1 (TA278). TJN01 and TJN02 shared the nucleotide sequence with TA278 merely in 53.9% and 55.2%, respectively. They possessed two major open reading frames (ORFs) and the noncoding region with a GC-rich region forming stem-loop structures, which are characteristic of TTV. However, their amino acid sequences in ORF1 were similar to that of TA278 in only 35.4 and 34.0%, respectively; TJN01 was 45.4% similar to TJN02. Comparison with TTV isolates of the same genotype identified hypervariable regions in ORF1 of TJN01 and TJN02, as in the prototype TTV of genotype 1. However, quasispecies were barely observed in them. Furthermore, sequences of hypervariable regions scarcely changed during 2-5.5 years in both TJN01 and TJN02. These results indicate that TTV of genotypes 12 and 13 are much different from the prototype TTV of genotype 1.

Adolescent↗
PubMed2000

Theoretical analysis of right gastroepiploic artery grafting to right coronary artery.

BACKGROUND: The right gastroepiploic artery (GEA) has been used as the second reliable arterial graft for coronary artery bypass grafting (CABG). However, concern regarding the flow competition with the recipient coronary artery has remained. METHODS: An application of in situ GEA grafting to the right coronary artery (RCA) was studied by using a theoretical model. The theoretical model of CABG was given variables; ie, the diameters and the lengths of both in situ GEA and proximal segment of the RCA, and the degree of proximal stenosis in the RCA. According to the range of these variables obtained from clinical data, the ratio of the GEA flow to the flow of the RCA distal to the anastomosis was calculated. RESULTS: Main factors to determine the flows in the two parallel paths were the inner diameters of both vessels, and the degree of the proximal stenosis. When the inner diameters of the GEA were 0.5 mm larger than that of the RCA, the GEA carried more than 50% of the total flow of the RCA distal to the anastomosis despite a moderate stenosis in the RCA. When the inner diameter of the GEA was equal to, or 0.5 mm smaller than, that of the RCA, the GEA flow was dominated by the native RCA flow unless the proximal stenosis was critical. CONCLUSIONS: If the inner diameter of the GEA is 0.5 mm larger than that of the RCA, CABG with the GEA can be applied more widely. If not, the application would basically be limited.

Coronary Artery Bypass↗
PubMed2000

Differentiation of hepatitis B virus genotypes D and E by ELISA using monoclonal antibodies to epitopes on the preS2-region product.

An enzyme-linked immunosorbent assay (ELISA) has been described for serological determination of hepatitis B virus genotypes, using monoclonal antibodies (mAb) against seven distinct epitopes (b, m, k, s, u, f and g) on the preS2-region products of hepatitis B surface antigen (HBsAg). The usefulness of this method for serological detection of genotype E, however, was theoretical, because no HBsAg samples of this genotype were included in the original test panel. Moreover, the predicted serotype of genotype E (bksufg) closely resembled that of genotype D (bksu, bksuf or bksug). Four HBsAg samples of genotype E were tested by the original described ELISA. The epitope g, predicted to be present in these samples by amino acid sequences, was not detected when HBsAg of genotype E was captured on a solid phase by mAb to the common determinant 'a' of HBsAg and then reacted with mAb to g (5156) labeled with horseradish peroxidase. However, the four examples of HBsAg of genotype E were captured by mAb 5156 to g on a solid phase; they were then detected by labeled mAb to the common determinant 'a'. Since epitopes f and g co-occurred on HBsAg of genotype E, HBsAg samples of this genotype were also detected, by 'sandwiching' them between immobilized mAb to g and labeled mAb to f. By contrast, HBsAg of genotype D in 90 sera was not reactive when sandwiched between mAb to f and g. Thus, this modified ELISA enables the serological determination of all six genotypes of HBsAg and, by inference, of hepatitis B virus.

Africa↗
PubMed2000

A model for neural representation of temporal duration.

To address how temporal duration is encoded in neural systems, we put forward a simple model for recurrent neural networks. Particular assumptions are only the following two: (1) neuronal bistability and; (2) environmental effects described by a heat bath. The results of Monte Carlo simulation show that population activity triggered at an initial time continues for a prolonged duration, followed by an abrupt self-termination. This time course seems highly suitable for neural representation of temporal duration. The time scale of this prolonged duration is much longer than the time scale of neuronal firing which is of the order of ms. The former time scale implies that of interval timing in cognition and behaviour. Thus, the model provides a possible explanation for a link between these two separated time scales. The Weber law, a hallmark of humans and animals' interval timing, can also be reproduced in our model.

Animals↗
PubMed2000

A model for molecular mechanisms of synaptic competition for a finite resource.

Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) undergoes Ca(2+)/calmodulin-dependent autophosphorylation of threonine-286/287 (Thr(286/287)). Extremely high concentration of CaMKII in the postsynaptic spine indicates that increase in the Ca(2+) concentration in the spine induced by synaptic activation results in Thr(286/287) autophosphorylation of this enzyme. It has recently been shown that the K(d) value of CaMKII for Ca(2+)/calmodulin (Ca(2+)/CaM) drastically decreases after Thr(286/287) autophosphorylation. Therefore, Ca(2+)/CaM associated with CaMKII becomes tightly bound to this kinase after Thr(286/287) autophosphorylation. This has been called 'Ca(2+)/CaM trapping'. We discussed the functional significance of Ca(2+)/CaM trapping in the neuronal system by a mathematical-modelling approach. We considered neighbouring synapses formed on the same dendrite and different increase in the Ca(2+) concentration in each spine. CaMKII undergoing Thr(286/287) autophosphorylation in each spine is eager to recruit nearby calmodulin in the dendrite for Ca(2+)/CaM trapping. Since the amount of calmodulin is limited, recruiting calmodulin to each spine causes competition among synapses for this finite resource. The results of our computer simulation show that this competition leads to 'winner-take-all': almost all calmodulin is taken by a few synapses to which relatively large increases in the Ca(2+) concentration are assigned. These results suggest a novel form of synaptic encoding of information.

Calcium↗
PubMed2000

Midline signals regulate retinal neurogenesis in zebrafish.

In zebrafish, neuronal differentiation progresses across the retina in a pattern that is reminiscent of the neurogenic wave that sweeps across the developing eye in Drosophila. We show that expression of a zebrafish homolog of Drosophila atonal, ath5, sweeps across the eye predicting the wave of neuronal differentiation. By analyzing the regulation of ath5 expression, we have elucidated the mechanisms that regulate initiation and spread of neurogenesis in the retina. ath5 expression is lost in Nodal pathway mutant embryos lacking axial tissues that include the prechordal plate. A likely role for axial tissue is to induce optic stalk cells that subsequently regulate ath5 expression. Our results suggest that a series of inductive events, initiated from the prechordal plate and progressing from the optic stalks, regulates the spread of neuronal differentiation across the zebrafish retina.

Animals↗
PubMed2000

Inhibitory influences of tranilast on multinucleated giant cell formation from monocytes by supernatant of concanavalin A-stimulated mononuclear cells.

Tranilast is an anti-allergic drug that inhibits the release of chemical mediators from mast cells. There have been cases-reports showing that tranilast is effective for the treatment of granulomatous diseases such as granuloma annulare and cutaneous sarcoidosis. Here we examined the in vitro effects of tranilast on the formation of multinucleated giant cells (MGCs) from human peripheral monocytes. Supernatant of concanavalin A (Con A)-stimulated mononuclear cells induced Langhans-type and foreign body-type MGCs and the addition of 10 or 100 microg/ml tranilast inhibited the formation of total MGCs and foreign body-type MGCs. Tranilast decreased the number of MGCs with 16<nuclei and increased that of MGCs with three to five nuclei. Fluorescence-activated cell sorting analysis showed that tranilast-treated monocytes had lower expressions of intercellular adhesion molecule-1 (ICAM-1). These findings suggest that tranilast is effective for cutaneous lesions in some cases of granulomatous disorders partly through a direct effect on monocyte/macrophage-lineage cells.

Anti-Allergic Agents↗
PubMed2000

Anti-metastatic efficacy and safety of MMI-166, a selective matrix metalloproteinase inhibitor.

The anti-metastatic efficacy and safety of a newly-developed matrix metalloproteinase (MMP) inhibitor were examined. MMI-166, a N-sulfonylamino acid derivative, inhibited the enzyme activity of MMP-2, 9, and 14 but not MMP-1, 3 or 7. Daily oral administration of MMI-166 resulted in potent inhibition of metastatic lung colonization of Lewis lung carcinoma injected via the tail vein and liver metastasis of C-1H human colon cancer implanted into the spleen at inhibition levels of 43% and 63%, respectively. Daily administration of MMI-166 also resulted in prolonged survival of mice given intraperitoneal implantation of Ma44 human lung cancer cells. The anti-metastatic activity of MMI-166 was as effective as that of other MMP inhibitors with broad inhibitory spectrum. MMI-166 did not affect in vitro tumor cell growth. Neither body weight losses nor hematotoxicity was observed during long-term treatment, indicating the safety of MMI-166 in mice. These results indicate that the selective MMP inhibitor MMI-166 has therapeutic potential as an anti-metastasis agent.

Animals↗
PubMed2000

Immunohistochemical expression of chromogranins A and B, prohormone convertases 2 and 3, and amidating enzyme in carcinoid tumors and pancreatic endocrine tumors.

Although chromogranin A (CgA) is widely distributed in neuroendocrine tumors, the distribution of chromogranin B (CgB) has not been elucidated. Hormones produced by tumors are sometimes prohormones and not necessarily bioactive hormones. Prohormones have to be processed into bioactive peptides by prohormone convertases (PCs), and some of them have to be amidated by peptidylglycine a-amidating monooxygenase (PGM). Whether PCs and PGM are present or not in tumors may explain why some tumors are functioning and some are nonfunctioning. We investigated 45 carcinoids and 16 pancreatic endocrine tumors. Of the carcinoids, CgA was expressed in most of the tumors, except for the rectal and ovarian carcinoids, which expressed CgB strongly. The expressions of PC2, PC3, and PGM were 31%, 100%, and 87%, respectively. In the pancreatic tumors, CgA was expressed in all tumors, whereas CgB was not expressed in any tumor. The expressions of PC2, PC3, and PGM were 63%, 88%, and 63%, respectively. PC3 was expressed in all of the functioning tumors but not in two of the four nonfunctioning tumors. PC2 and PGM were not expressed in three of the four nonfunctioning tumors. In conclusion, expression of CgA and CgB was different depending on the tumor location. High frequency of PCs and PGM may explain why even nonfunctioning tumors produce some inconspicuous peptides.

Adenoma, Islet Cell↗