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Biomedical subjects

S Okuno

Publications and source records attributed to S Okuno.

At least 73 records · Page 4Linked to original sources
PubMed2000

Macrophage-mediated activation of camptothecin analogue T-2513-carboxymethyl dextran conjugate (T-0128): possible cellular mechanism for antitumor activity.

Camptothecin (CPT) analogue T-2513-carboxymethyl (CM) dextran conjugate (T-0128) suppressed human tumor xenografts that were refractory to CPTs. This improvement was explained by its altered pharmacokinetics, but the cellular mechanism of action is still not clear. For this reason, in the present study we examined the determinants of T-0128 action at the cellular level. In vitro tests showed that T-0128 was inactive, indicating that the requirement for its activity lies in the release of linked T-2513, accompanied by the cellular uptake of the conjugate. The accumulation varied between cell lines: tumor cells, including Walker-256 carcinoma and B16 melanoma, showed only a marginal uptake and an undetectable drug release in the medium. In contrast, macrophage-like cells, such as J774.1, internalized T-0128 very efficiently, and liberated T-2513. With regard to the mode of accumulation, fluid-phase pinocytosis seems to be a key factor based on the followings: a similar cell-specificity existed in the uptake of FITC dextran, a marker of fluid-phase pinocytosis. Also, the macrophage uptake of T-0128 increased almost linearly with its medium concentration and was insensitive to dextran sulfate, a ligand for macrophage scavenger receptor. Comparative efficacy studies of T-0128 in the presence and absence of macrophages demonstrated that macrophages increased the efficacy of T-0128. The enhancement could be explained in terms of increases in the amount of released T-2513. Overall, these results lead us to the conclusion that T-0128 acts like a Trojan horse with the help of macrophages: T-0128 is taken up by macrophages in tumor tissues, and the liberated T-2513 kills tumor cells.

Animals↗
PubMed2000

Determinants for the drug release from T-0128, camptothecin analogue-carboxymethyl dextran conjugate.

To improve pharmacological profiles of camptothecins (CPTs), a new macromolecular prodrug, denoted T-0128, was synthesized. This prodrug comprises a novel CPT analog (T-2513: 7-ethyl-10-aminopropyloxy-CPT) bound to carboxymethyl (CM) dextran through a Gly-Gly-Gly linker, with a molecular weight of 130 kDa. The present study was designed to elucidate the mechanisms that promote the release of linked T-2513. First, we compared the abilities of a rat liver homogenate, a cocktail of its lysosomal enzymes, and different types of pure enzymes, to liberate T-2513 from the conjugate. The releasing rate in the homogenate was very slow, but was accelerated with the lysosomes. Lysosomal cysteine proteinases, such as cathepsin B, were responsible, coupled with the results of in vitro and in vivo inhibition studies using proteinase inhibitors. The pH optimum for the cathepsin B-mediated drug release was approximately 4. This corresponds to the pH in lysosomes, suggesting lysosomotropic release. Second, to assess the effect of the length and composition of the peptidyl linker, we synthesized the conjugates with a different linker and compared the drug-releasing rates. We found that the insertion of Phe into Gly-Gly-Gly allowed various kinds of enzymes to produce a rapid cleavage, and the Gly-chain lengthening enhanced the lysosome-mediated drug release. The released T-2513 levels in the liver and tumor of the tumor-bearing rats dosed with each conjugate increased with the length of Gly linker, suggesting a good in vitro to in vivo relationship. Comparative efficacy studies of the conjugates with a different linker demonstrated that T-0128 showed the maximum efficacy against MX-1 human mammary xenograft tumors. Thus the Gly-Gly-Gly linker exploits lysosomal cathepsin B to liberate T-2513 slowly and steadily, resulting in improved therapeutic efficacy.

Animals↗
PubMed2000

Complete regression of xenografted human carcinomas by camptothecin analogue-carboxymethyl dextran conjugate (T-0128).

Clinically available camptothecins (CPTs), such as irinotecan (CPT-11) and topotecan, represent one of the most promising classes of antitumor agents, despite their toxicity. To improve their pharmacological profiles, a new macromolecular prodrug, denoted T-0128, was synthesized. This prodrug is a novel CPT analogue (T-2513)-carboxymethyl (CM) dextran conjugate via a triglycine spacer, with a molecular weight of Mr 130,000. This study was designed to test the concept that the rational design of a CPT-polymer conjugate would increase the efficacy of the parent drug. The in vivo antitumor study against Walker-256 carcinoma demonstrated that T-0128 was 10 times as active as T-2513, supporting this concept. Additionally, comparative efficacy studies of T-0128, T-2513, CPT-11, and topotecan were performed using a panel of human tumor xenografts in nude mice, showing the advantage of T-0128 over these CPTs. The maximal tolerated doses (MTDs) of T-0128, T-2513, and CPT-11 were comparable. Even a single i.v. injection of T-0128 at 6 mg/kg (based on the amount of T-2513 bound to CM dextran) induced complete regression of MX-1 mammary carcinoma. T-0128 at 10 mg/kg weekly for 3 weeks (one-tenth of its MTD) cured LX-1 lung carcinoma. Also, T-0128 below its MTD consistently cured or regressed St-4 gastric and HT-29 colorectal tumor xenografts that are highly refractory to CPTs. These demonstrate the broad range of therapeutic doses achieved with T-0128. Pharmacokinetic studies were performed to correlate the efficacy results obtained for T-0128 with plasma and tissue drug concentrations using Walker-256 tumor-bearing rats. Results showed that after i.v. administration of T-0128, the conjugate continued to circulate at a high concentration for an extended period, resulting in tumor accumulation. In the tumor, the sustained release of T-2513 occurred. In contrast, T-2513 disappeared rapidly from the body. The significant increases in the amount and exposure time of released T-2513 in the tumor explain well the enhanced efficacy of T-0128. In conclusion, this study indicated that T-0128 improved the potency of T-2513, demonstrating the proof of the above concept.

Animals↗
PubMed2000

Immunohistochemical study of the distribution of Ca(2+)/calmodulin-dependent protein kinase phosphatase in the rat central nervous system.

Distribution of Ca(2+)/calmodulin-dependent protein kinase phosphatase (CaM-K Pase) which dephosphorylate multifunctional Ca(2+)/calmodulin-dependent protein kinases (CaM-kinases) in the rat brain and spinal cord were examined immunohistochemically by using an antibody against this enzyme. CaM-K Pase was localized only in the cytoplasm as has been investigated in PC 12 cells, and was never observed in the nucleus. Immunostainability varied from cell group to cell group. Mitral cells in the olfactory bulb, pyramidal neurons in the fifth layer of the cerebral cortex, hippocampal and striatal interneurons, dorsal and ventral pallidal, entopeduncular, and the reticular part of the substantia nigra neurons were intensely immunolabeled. Motoneurons in all the cranial nerve nuclei and the anterior horn of the spinal cord also revealed intense immunolabeling. On the contrary, pyramidal neurons in the Ammon's horn of the hippocampal formation, granule cells in the olfactory bulb, dentate gyrus and cerebellar cortex, Purkinje cells, neurons in the medial habenular nucleus and the inferior olivary nucleus have not shown immunoreactivity. Axons in the white matter or nerve root of the cranial nerve nuclei were immunolabeled. Glial cells in the white matter also showed immunostaining. Because the substrate of CaM-K Pase is multifunctional CaM-kinase II, I and IV, localization of each CaM-kinase was compared with that of CaM-K Pase. The distribution of CaM-K Pase and these CaM-kinases was found to overlap in various regions in the brain and spinal cord. It was concluded, therefore, that CaM-K Pase could regulate the activity of these CaM-kinases by dephosphorylation, when they existed together in neurons.

Animals↗
PubMed2000

Characterization of newly developed SSLP markers for the rat.

We have isolated more than 12,000 clones containing microsatellite sequences, mainly consisting of (CA)n dinucleotide repeats, using genomic DNA from the BN strain of laboratory rat. Data trimming yielded 9636 non-redundant microsatellite sequences, and we designed oligonucleotide primer pairs to amplify 8189 of these. PCR amplification of genomic DNA from five different rat strains yielded clean amplification products for 7040 of these simple-sequence-length-polymorphism (SSLP) markers; 3019 markers had been mapped previously by radiation hybrid (RH) mapping methods (Nat Genet 22, 27-36, 1998). Here we report the characterization of these newly developed microsatellite markers as well as the release of previously unpublished microsatellite marker information. In addition, we have constructed a genome-wide linkage map of 515 markers, 204 of which are derived from our new collection, by genotyping 48 F2 progeny of (OLETFxBN)F2 crosses. This map spans 1830.9 cM, with an average spacing of 3.56 cM. Together with our ongoing project of preparing a whole-genome radiation hybrid map for the rat, this dense linkage map should provide a valuable resource for genetic studies in this model species.

Animals↗
PubMed2000

Synthesis and cytokinin activity of fluorescent 7-Phenylethynylimidazo[4,5-b]pyridine and its riboside.

7-Phenylethynylimidazo[4,5-b]pyridine and its riboside have been newly developed as fluorescent carbon-substituted cytokinin analogues. Palladium-catalyzed coupling of 7-iodo-3-(tri-O-acetyl-beta-D-ribofuranosyl)imidazo[4,5-b]pyridine with phenylacetylene followed by ammonolysis afforded the 7-phenylethynyl riboside via its tri-O-acetate. Acid hydrolysis of the riboside provided its free base, which showed a marked enhancement in fluorescence intensity in an aqueous alkaline solution. The free base and its riboside were more active than the corresponding 6-phenylethynylpurine and its riboside, respectively, in Amaranthus betacyanin and tobacco callus bioassays. Surprisingly, the imidazo[4,5-b]pyridine base exhibited strong cytokinin activity comparable to that of N(6)-benzyladenine in the tobacco callus bioassay. This compound would be useful for studying localization and transport of cytokinins in cells or tissues of plants.

Cytokinins↗
PubMed2000

Quantitative trait loci for lipid metabolism in the study of OLETF x (OLETF x Fischer 344) backcross rats.

1. The Otsuka Long-Evans Tokushima Fatty (OLETF) rat is a model of type II diabetes with accompanying dyslipidaemia and obesity. 2. To define chromosomal intervals associated with obesity (abdominal fat weight and plasma leptin levels), dyslipidaemia (plasma triglyceride, cholesterol and free fatty acids) and hyperglycaemia (plasma glucose levels), we have performed genome-wide quantitative traits loci (QTL) analyses of 115 male OLETF x (OLETF x Fischer 344) backcross animals at 16 weeks of age. 3. The Diabetes Mellitus OLETF type I (Dmo1) locus on rat chromosome 1 showed statistically significant involvement in elevations of plasma levels of triglycerides (P = 4.87 x 10(-6) at D1Rat90) and total cholesterol (P = 1.16 x 10(-5) at D1Rat306). 4. No other loci produced significant linkage to these observed phenotypes. 5. These analyses have confirmed the importance of Dmo1 in lipid homeostasis at younger ages as well as during overt diabetes, which appears later. Thus, alterations at the Dmo1 locus are a major risk factor for pathogenesis in the strain, a finding that agrees with physiological studies that indicate a role for dyslipidaemia in the type II diabetic syndrome of OLETF rats.

Animals↗
PubMed2000

Molecular cloning and expression of human xCT, the light chain of amino acid transport system xc-.

Transport of system xc- is an exchange agency with high specificity for anionic form of cystine and glutamate. The protein mediating this transport is a disulfide-linked heterodimer of a light chain named xCT and a heavy chain previously known as 4F2hc. We have isolated two cDNAs encoding xCT from the human cDNA library. One clone coded for a protein of 501 amino acids with 12 putative transmembrane domains. When functionally expressed in Xenopus oocytes together with the human 4F2hc, human xCT induced the transport activity whose characteristics are similar to those of system xc-. Another clone seemed to contain a partial human xCT and a long 3' untranslated region. The human xCT gene was localized at chromosome 4q28-31. Analysis of the 5'-flanking region of the human xCT gene revealed several sites for potentially binding of transcriptional factors, including NF-E2 and AP-1. Transport of cystine via system xc- has been known as a regulatory factor for the intracellular glutathione level, and its transport activity is induced in response to the oxygen tension in culture. Northern blot analysis demonstrated that the expression of both xCT and 4F2hc was significantly enhanced by oxygen. The results suggest that oxygen regulates the activity of system xc- by modulating the expression of both xCT and 4F2hc mRNAs.

Amino Acid Sequence↗
PubMed2000

Studies on the phosphorylation of protein kinase B by Ca(2+)/calmodulin-dependent protein kinases.

Protein kinase B (PKB) was recently reported to be activated on the phosphorylation of Thr(308) by Ca(2+)/calmodulin-dependent protein kinase kinase alpha (CaM-kinase kinase alpha), suggesting that PKB was regulated through not only the phosphoinositide 3-kinase pathway but also the Ca(2+)/calmodulin protein kinase pathway. The activation of PKB by CaM-kinase kinase alpha was as high as 300-fold after incubation for 30 min under the phosphorylation conditions, and still increased thereafter, suggesting that the maximal activation of PKB on phosphorylation of the Thr(308) residue is several hundred fold. On the other hand, the V(max) value of CaM-kinase kinase alpha for the phosphorylation of PKB was more than two orders of magnitude lower than that for CaM-kinase IV, although the K(m) values for PKB and CaM-kinase IV were not significantly different, raising the question of whether or not PKB is a physiological substrate of CaM-kinase kinase alpha. Besides CaM-kinase kinase alpha, CaM-kinase II also remarkably activated PKB. However, the specific activities of CaM-kinase kinase alpha and CaM-kinase II as to the activation of PKB were more than three orders of magnitude lower than that of 3-phosphoinositide-dependent protein kinase 1 (PDK1).

Animals↗
PubMed2000

Sex differences in fluorouracil-induced stomatitis.

PURPOSE: A meta-analysis of six North Central Cancer Treatment Group (NCCTG) trials involving patients receiving their first ever fluorouracil (5-FU)-based chemotherapy was undertaken to explore the association of sex with reports of the incidence and severity of stomatitis. PATIENTS AND METHODS: Data were obtained on a total of 731 patients (402 men and 329 women). Comparisons of incidence and severity rates and average stomatitis across sex were performed using standard binomial testing and t tests, respectively. Logistic regression analysis and a weighted analysis using data summarized to study level served as evidence of cross-validation. RESULTS: Women reported stomatitis both more often and with greater severity than did men. The incidence of any stomatitis for women was 63% versus 52% for men (P =.002). The incidence of severe or very severe stomatitis for men and women was 22% and 12%, respectively (P =. 0006). On average, women reported stomatitis of roughly 0.4 points higher than men on a 0 to 4 ordinal scale (P <.00001). Comparison of results across treatment and placebo arms was carried out to validate the initial findings. Logistic regression modelling further confirmed the results conditional on the presence of a number of potentially confounding covariates. Women were also 11% more likely than men to experience leukopenia of common toxicity criteria grade >/= 1, (70% v 59%, respectively; P <.00001) and grade 3+ (18% v 11%, respectively; P =.004). CONCLUSION: More women than men reported 5-FU-induced stomatitis. The precise mechanism resulting in different degrees of stomatitis across sex is not evident.

Adult↗
PubMed2000

Distribution characteristics of carboxymethylpullulan-peptide-doxorubicin conjugates in tumor-bearing rats: different sequence of peptide spacers and doxorubicin contents.

Plasma and tissue distribution of conjugates of carboxymethylpullulan (CMPul) and doxorubicin (DXR), either bound directly or through three types of tetrapeptide spacers, was studied after intravenous injection to rats bearing Walker 256 carcinosarcoma and compared with that of DXR. In contrast to DXR, each conjugate retained high levels of DXR in the conjugated form in plasma and displayed high accumulation in the tumor at 6 h after the administration. Disposition characteristics of [3H]CMPul in rats bearing Walker 256 carcinosarcoma indicate that pullulan, which had molecular weight over 50 kDa, is a suitable macromolecular carrier for tumor targeting in cancer chemotherapy by carboxymethylation. We find that the in vivo antitumor effect of the conjugates depends on the tumor AUC of free DXR released from the conjugates. CMPul-DXR conjugates were also distributed in the reticuloendothelial organs, such as liver, spleen and bone marrow; however, the tissue concentrations of the conjugates in the heart, lung and muscle were lower than those of DXR. We next investigated the effect of the DXR contents of CMPul-DXR conjugates on their body distribution in rats bearing Walker 256. The half life of CMPul-DXR conjugates in plasma were shorter and the conjugates had greater accumulation in the reticuloendothelial system, while they showed lower concentrations in the tumor with increasing DXR contents. Antitumor activity of CMPul-DXR conjugates were reduced and the lethal toxicities of CMPul-DXR conjugates were amplified with increasing DXR contents.

Animals↗
PubMed2000

Relationship between home blood pressure measurement and medication compliance and name recognition of antihypertensive drugs.

This study examined the relationship of home blood pressure measurement to medication compliance and name recognition of antihypertensive drugs in outpatients with hypertension. A total of 1,452 consecutive outpatients (842 males, 610 females; mean age 65+/-11 yr) seeking care at our institute answered questions at our cardiovascular outpatient clinic such as whether they had a sphygmomanometer at home, how often they measured their blood pressure at home, and how often they missed taking their medication. Among a total of 777 patients on antihypertensive drugs who had a sphygmomanometer at home, 16 of the 242 patients (6.5%) who measured their home blood pressure every day occasionally missed taking their medication, whereas this number was 22 for the 216 patients (10.1%) who measured their home blood pressure several times a week, 16 for the 146 patients (11.0%) who measured their home blood pressure several times a month, and 25 for the 173 patients (14.5%) who never measured their home blood pressure (p< 0.01 between patients who measured their home blood pressure every day and those who did not measure their home blood pressure). Among a total of 271 patients taking one or two antihypertensive drugs, the number of patients who could name their antihypertensive drugs was 47 of the 86 patients (55%) who measured their home blood pressure every day, 43 of the 78 patients (55%) who measured their home blood pressure several times a week, 24 of the 41 patients (58%) who measured their home blood pressure several times a month, and 22 of the 66 patients (33%) who never measured their home blood pressure (p< 0.02). In conclusion, medication compliance and antihypertensive drug name recognition were better in patients who measured their home blood pressure than in patients who did not measure their home blood pressure. From these results, we conclude that physicians should recommend home blood pressure measurement to patients being treated with antihypertensive drugs, because there is a possibility that home blood pressure measurement might improve medication compliance.

Aged↗
PubMed2000

Distinct increase in hematocrit associated with paroxysm of atrial fibrillation.

In a previous study we found that hemoconcentration, which was identified by an increase in hematocrit, occured during a paroxysm of atrial fibrillation. In the present study we investigated the changes in hematocrit from sinus rhythm to paroxysm in 10 patients who had multiple paroxysms of atrial fibrillation in order to assess the ranges of the changes in hematocrit among the paroxysms. In these patients hematocrit was measured simultaneously with electrocardiographic recording during 3 or more paroxysms and sinus rhythm just before each paroxysm. The changes in hematocrit varied among the paroxysms. The maximum increase in hematocrit in each patient ranged from 3.5 to 8.0 points with an average of 5.1 points. Such a distinct increase in hematocrit which abruptly develops with a paroxysm of atrial fibrillation may be a potential risk for thrombus formation.

Aged↗
PubMed2000

Effect of direct arterial bypass on the prevention of future stroke in patients with the hemorrhagic variety of moyamoya disease.

OBJECT: The authors evaluated the effects of superficial temporal artery-middle cerebral artery (STA-MCA) bypass in the prevention of future stroke, including rebleeding or an ischemic event, in patients suffering from hemorrhagic moyamoya disease by comparing this method with indirect bypass and conservative treatment. METHODS: Twenty-two patients who had hemorrhagic moyamoya disease but no aneurysm comprised the study group. These patients' clinical charts were examined with respect to their treatment and clinical course after an initial hemorrhagic episode. The mean age of the patients was 43 years and the follow-up period ranged from 0.8 to 15.1 years, with a mean of 8 years. Eleven patients (50%) were conservatively treated. Among the 11 patients who were surgically treated, STA-MCA bypass was performed in six patients (27%) and encephaloduroarteriosynangiosis (EDAS) in the other five patients (23%). Nine patients (41%) presented with an ischemic or rebleeding event during the follow-up period. The incidence of future stroke events in patients who had undergone an STA-MCA bypass was significantly lower (p<0.05) than that in patients who had been treated conservatively or with EDAS. Kaplan-Meier plots comparing stroke-free times in patients treated with direct bypass and those in patients who conservatively or with indirect bypass showed a significant difference (p<0.05) in favor of direct bypass. CONCLUSIONS: The effect of STA-MCA bypass on the prevention of recurrent hemorrhage or an ischemic event in patients with hemorrhagic moyamoya disease has been statistically confirmed in this study.

Adult↗
PubMed2000

Transient mitral regurgitation following vagal stimulation-induced ventricular arrhythmia in rabbits: a color Doppler echocardiographic study.

BACKGROUND AND AIM OF THE STUDY: Cervical vagal stimulation in rabbits frequently causes systolic murmur with bigeminy due to premature ventricular contractions. The bigeminy disappears in a few minutes, but the systolic murmur persists for a few days. Peculiar lesions of the mitral valves, mitral annulus and papillary muscles, and an increase in left atrial weight, frequently develop in a week. In this study, color Doppler echocardiography was used to examine whether the systolic murmur was due to mitral regurgitation. METHODS: Echocardiographic monitoring was carried out in anesthetized rabbits restrained in the supine position. RESULTS: Doppler echocardiography and phonocardiography showed systolic murmur at 6 h, three days, and at one, two, three and four weeks after vagal stimulation. At 6 h after stimulation, phonocardiography showed systolic click and late systolic murmur; Doppler echocardiography showed marked mitral regurgitation. The systolic murmur and mitral regurgitation were attenuated and the papillary muscle was swollen three days after vagal stimulation. Following stimulation, mitral regurgitation disappeared within one week, and papillary muscle swelling improved after three weeks. CONCLUSION: Doppler echocardiography confirmed that systolic murmur caused by vagal stimulation in rabbits was due to mitral regurgitation.

Animals↗
PubMed2000

[A surgical case of growing cavernous angioma at the pontomedullary junction].

We described a surgical case of growing cavernous angioma located at the pontomedullary junction. This 52-year-old woman presented with symptoms caused by a small hemorrhage in the right cerebellopontine angle. Magnetic resonance images (MRI) suggested cavernous angioma as the underlying pathology. 9 months after the first episode, the second hemorrhage occurred with a deteriorated neurological state that disappeared under conservative treatment except for right facial paresis and hearing disturbance. During careful observation for 1 year, the size of the lesion gradually increased on MRI and additional neurological deficits including left hemiparesis and right abducent nerve palsy were diagnosed. The first operation was carried out through the right lateral suboccipital approach, but only partial removal of the cavernous angioma was accomplished due to the overlying seventh and lower cranial nerves. After more than 4 months, a third hemorrhagic episode was presented with a sudden onset of right cerebellar signs and facial numbness. The cavernous angioma grew in size to reach the ventrolateral corner of the 4th ventricle with dense hemosiderin deposition around the core lesion on MRI. An enhancement inside the lesion was also demonstrated after gadolinium-diethylenetriaminepenta-acetic acid administration. The second operation through the midline suboccipital approach was selected for the complete resection of the residual cavernous angioma. The lesion was too hard to resect without internal decompression. The pontine part of the lesion was almost totally resected, but manipulation for the medullary part to create a discrete layer between the lesion and surrounding neural tissues was unsuccessful and generated severe bradycardia, so this part of the cavernous angioma had to be left. The problems for the management of cavernous angioma in the brain stem should be discussed, especially focussing on the surgical indication in reference to our experience and previous literatures.

Brain Stem Neoplasms↗