Biomedical subjects

Yuji Ishida

Publications and source records attributed to Yuji Ishida.

14 recordsLinked to original sources

PubMed2006

Binary vectors and super-binary vectors.

A binary vector is a standard tool in the transformation of higher plants mediated by Agrobacterium tumefaciens. It is composed of the borders of T-DNA, multiple cloning sites, replication functions for Escherichia coli and A. tumefaciens, selectable marker genes, reporter genes, and other accessory elements that can improve the efficiency of and/or give further capability to the system. A super-binary vector carries additional virulence genes from a Ti plasmid, and exhibits very high frequency of transformation, which is valuable for recalcitrant plants such as cereals. A number of useful vectors are widely circulated. Whereas vectors with compatible selectable markers and convenient cloning sites are usually the top criteria when inserting gene fragments shorter than 15 kb, the capability of maintaining a large DNA piece is more important for consideration when introducing DNA fragments larger than 15 kb. Because no vector is perfect for every project, it is recommended that modification or construction of vectors should be made according to the objective of the experiments. Existing vectors serve as good sources of components.

Agrobacterium tumefaciens↗

PubMed2005

Differential expression pattern of C4 bundle sheath expression genes in rice, a C3 plant.

NADP-malic enzyme (NADP-ME) and phosphoenolpyruvate carboxykinase (PCK) are specifically expressed in bundle sheath cells (BSCs) in NADP-ME-type and PCK-type C4 plants, respectively. Unlike the high activities of these enzymes in the green leaves of C4 plants, their low activities have been detected in the leaves of C3 plants. In order to elucidate the differences in the gene expression system between C3 and C4 plants, we have produced chimeric constructs with the beta-glucuronidase (GUS) reporter gene under the control of the maize NADP-Me (ZmMe) or Zoysia japonica Pck (ZjPck) promoter and introduced these constructs into rice. In leaves of transgenic rice, the ZmMe promoter directed GUS expression not only in mesophyll cells (MCs) but also in BSCs and vascular cells, whereas the ZjPck promoter directed GUS expression only in BSCs and vascular cells. Neither the ZjPck nor ZmMe promoters induced GUS expression due to light. In rice leaves, the endogenous NADP-Me (OsMe1) was expressed in MCs, BSCs and vascular cells, whereas the rice Pck (OsPck1) was expressed only in BSCs and vascular cells. Taken together, the results obtained from transgenic rice demonstrate that the expression pattern of ZmMe or ZjPck in transgenic rice was reflected by that of its counterpart gene in rice.

Chimera↗

PubMed2005

The influence of dialysate calcium on the therapeutic effects of sevelamer hydrochloride in hemodialysis patients with secondary hyperparathyroidism under treatment of intravenous vitamin d metabolites.

The management of hyperphosphatemia is essential to treat secondary hyperparathyroidism and to prevent ectopic calcification. Sevelamer hydrochloride (sevelamer), a new phosphate binder that contains neither aluminum nor calcium, which could be theoretically beneficial for the management of hyperphosphatemia in dialysis patients with secondary hyperparathyroidism who are receiving intravenous vitamin D metabolites (maxacalcitol or calcitriol). To reduce calcium loads, a dialysate calcium concentration of 2.5 mEq/L is recommended by Kidney Disease Outcome Quality Initiative (K/DOQI) guidelines. In Japan, a dialysate calcium concentration of 3.0 mEq/L prevails. We investigated the influence of dialysate calcium on the therapeutic effect of sevelamer in 40 hemodialysis patients who are under treatment of intravenous vitamin D metabolites for secondary hyperparathyroidism (VD(+)) and compared the results with those of 41 patients who had not received vitamin D metabolites (VD(-)). Serum phosphorus and calcium-phosphorus products showed no significant change by sevelamer in either the VD(+) subgroup of patients receiving hemodialysis with dialysate calcium of 2.5 mEq/L (DCa2.5) or those receiving hemodialysis with dialysate calcium of 3.0 mEq/L (DCa3.0), while serum phosphorus and calcium-phosphorus products decreased in both the VD(-) subgroups. Serum calcium decreased in the DCa2.5 subgroup and did not change in the DCa3.0 subgroup in both the VD(+) and the VD(-) subjects. Parathyroid hormone and alkaline phosphatase increased in the DCa2.5 subgroup and did not change in the Ca 3.0 subgroup in the VD(+) subjects. Serum calcium decreased in both subgroups in the VD(-) subjects. Parathyroid hormone obtained after sevelamer administration in the VD(-) group was within the target range of the K/DOQI guidelines. In conclusion, the concomitant use of sevelamer as a phosphate binder and the dialysate of calcium concentration of 2.5 mEq/L have possibilities for worsening secondary hyperparathyroidism in patients receiving intravenous vitamin D.

Calcitriol↗

PubMed2004

Expression of cold-tolerant pyruvate, orthophosphate dikinase cDNA, and heterotetramer formation in transgenic maize plants.

Maize is a typical C4 plant of the NADP-malic enzyme type, and its high productivity is supported by the C4 photosynthetic cycle, which concentrates atmospheric CO2 in the leaves. The plant exhibits superior photosynthetic ability under high light and high temperature, but under cold conditions the photosynthetic rate is significantly reduced. Pyruvate orthophosphate dikinase (PPDK), a key enzyme of the C4 pathway in maize, loses its activity below about 12 degrees C by dissociation of the tetramer and it is considered as one possible cause of the reduction in the photosynthetic rate of maize at low temperatures. To improve the cold stability of the enzyme, we introduced a cold-tolerant PPDK cDNA isolated from Flaveria brownii into maize by Agrobacterium-mediated transformation. We obtained higher levels of expression by using a double intron cassette and a chimeric cDNA made from F. bidentis and F. brownii with a maximum content of I mg/g fresh weight. In leaves of transgenic maize, PPDK molecules produced from the transgene were detected in cold-tolerant homotetramers or in heterotetramers of intermediate cold susceptibility formed with the internal PPDK. Simultaneous introduction of an antisense gene for maize PPDK generated plants in which the ratio of heterolologous and endogenous PPDK was greatly improved. Arrhenius plot analysis of the enzyme extracted from one such plant revealed that the break point was shifted about 3 degrees C lower than that of the wild type.

Cold Temperature↗

PubMed2003

Effects of eplerenone on heart and kidney in two-kidney, one-clip rats.

BACKGROUND: The role of aldosterone has been less investigated compared to the renin-angiotensin-aldosterone system in renovascular hypertension. The purpose of the present study was to compare the effects of a selective aldosterone receptor blocker, eplerenone (EP), and an angiotensin II receptor type 1 antagonist (AT1RA), losartan (LO) on cardiac and renal damage produced by two-kidney, one-clip (2K-1C) renovascular hypertension in rats. METHOD: Wistar rats (n = 48) were placed on one of six groups. Group 1 received sham operation. From group 2 to 6, all rats were made as 2K-1C renovascular hypertension. Group 2 received vehicle. Group 3 orally received 100 mg/kg/day of EP from the initiation of the study. Group 4 received 100 mg/kg/day of LO, from the initiation of the study. Groups 5 and 6 received EP and LO from the 4th week after the clipping respectively. Systolic blood pressure (SBP) and urinary protein excretion (UPE) were measured before and every 2 weeks. The remnant kidney was obtained for histopathological analysis and for measurement of endothelial cell nitric oxide synthase (ecNOS) gene expression (GE). RESULTS: SBP increased in the placebo group (132.1 +/- 2.4 vs. 115.0 +/- 0.6 mm Hg in sham group at week 10, p = 0.019). Treatment with LO or EP from the beginning of the study decreased SBP significantly as measured in the sham group at week 10. The placebo group developed significant UPE (21.7 +/- 1.9 mg/day) compared with the sham group (13.4 +/- 0.8 mg/day, p < 0.05). Treatment with both LO (12.5 +/- 1.5 mg/day, p < 0.01 vs. placebo) and EP (14.8 +/- 1.0 mg/ day, p < 0.05 vs. placebo) significantly decreased UPE. On the other hand, the late start of treatment with EP failed to decrease the increased UPE. UPEs were not significantly different between the LO- and EP-treated groups throughout the study. There was no significant pathological change in heart and kidney in all groups. In heart, ecNOS GE was significantly increased in the EP-treated (from the beginning of the study) rats compared with placebo group (0.47 +/- 0.01 vs. 0.43 +/- 0.01, p < 0.05). LO did not have an effect on ecNOS GE in heart. In aorta, ecNOS GE was significantly increased in the two EP-treated groups compared with the placebo group (0.22 +/- 0.01, 0.22 +/- 0.02 vs. 0.15 +/- 0.01, p < 0.05, respectively). LO also did not have an effect on ecNOS GE in aorta. In kidney, ecNOS GE was significantly increased in the LO group (from the beginning of the study) and two EP-treated groups compared with placebo. CONCLUSION: This study demonstrated that EP treatment significantly reduced SBP and UPE compared with placebo in both development and established 2K-1C renovascular hypertension. EP was as effective as LO in lowering the blood pressure of this renin-dependent animal model.

Angiotensin II Type 1 Receptor Blockers↗

PubMed2003

Hyperbilirubinemia protects against focal ischemia in rats.

Heme oxygenase-1 (HO1) catalyzes oxidation of the heme molecule in concert with NADPH-cytochrome P450 reductase following the specific cleavage of heme into carbon monoxide, iron, and biliverdin, which is rapidly metabolized to bilirubin. HO1 is a stress-inducible protein that protects cells against oxidative injury, but its protective mechanism is not fully understood. The Eizai hyperbilirubinemic rat (EHBR), a mutant strain derived from the Sprague-Dawley rat (SDR), has a mutation in the gene for the canalicular multispecific organic anion transporter, which results in a phenotype of hyperbilirubinemia, and thus is a model of Dubin-Johnson syndrome in humans. In this study, we compared EHBR and SDR with regard to neuronal death induced by 2 hr of occlusion of the middle cerebral artery and reperfusion. In EHBR, the area that was immunoreactive for microtubule-associated protein-2 was significantly reduced, and the HO1-immunoreactive area was smaller than that in SDR. These results suggest that bilirubin has essentially a neuroprotective effect against focal ischemia and may participate in HO1-induced neuroprotection.

Animals↗

PubMed2003

Significance of aberrant (cytoplasmic/nuclear) expression of beta-catenin in pancreatoblastoma.

This study concerns the significance of aberrant (nuclear/cytoplasmic) expression of beta-catenin in pancreatoblastoma (PBL). On immunohistochemistry, all seven PBLs examined showed nuclear/cytoplasmic expression of beta-catenin, predominantly in the squamoid corpuscles (SCs). In areas with acinar/ductular differentiation, few tumour cells displayed nuclear/cytoplasmic expression of beta-catenin and more than half of the tumour cells showed membranous expression. Two out of five (40%) tumours examined showed missense mutations in codons 33 and 37 of exon 3 of the beta-catenin gene. No mutation of the adenomatous polyposis coli (APC) gene was detected in two of the remaining three tumours. Amplifiable DNA for APC analysis was not obtained from the one other tumour. Immunoreactivity for cyclin D1, one of the nuclear targets of beta-catenin, was found predominantly in the SCs of the seven tumours. In contrast, the Ki-67 labelling index was 2-4% (median 3%) in the SCs and 8-18% (median 12%) in the other areas, indicating a negative correlation with nuclear cyclin D1 reactivity. These results imply that in PBLs, nuclear/cytoplasmic accumulation of beta-catenin and overexpression of its target gene cyclin D1 are not associated with the induction of tumour cell proliferation. Nuclear/cytoplasmic accumulation of beta-catenin may be related to the morphogenesis of the SCs that are considered most characteristic for PBL.

Child↗

PubMed2003

Effects of antihypertensive drugs on peritoneal vessels in hypertensive dogs with mild renal insufficiency.

The transport capacity of any membrane depends on its surface area and permeability. In addition, peritoneal capillaries are probably barriers to solute transport. Although no decisive use of antihypertensive drugs has been reported in continuous ambulatory peritoneal dialysis (CAPD) patients with hypertension, those drugs are known to have various effects on vessels. In the present study, we used a charge-coupled-device (CCD) camera in renovascular hypertensive dogs with mild renal insufficiency to investigate the effects of various antihypertensive drugs on the peritoneal capillaries. Renovascular hypertension was induced in the dogs by placing silver clips on both renal arteries to create 90% occlusion. After confirmation of elevation of blood pressure (usually 20 days after the operation), each dog's abdomen was opened while the animal was under general anesthesia. Using a CCD camera, the diameters of the small arteries of the peritoneum were measured after 3 days' oral administration of a placebo (n = 5); or of 8 mg CS866, a selective angiotensin II type 1 receptor blocker (n = 5); or of 10 mg benazepril, an angiotensin-converting enzyme inhibitor (n = 5); or of 10 mg amlodipine, a calcium antagonist (n = 5). In dogs receiving CS866, blood pressure decreased to 128 +/- 6 mmHg from 160 +/- 6 mmHg (p < 0.01). A similar decrease in blood pressure was observed with the use of the other drugs. The diameter of the small vessels increased by 28% +/- 6% in dogs receiving CS866 and by 24% +/- 5% in dogs receiving benazepril, as compared with 3% +/- 3% in dogs receiving the calcium antagonist. These data clearly demonstrate that blockade of the renin-angiotensin system produces an increase in solute clearance in hypertensive dogs with mild renal insufficiency and that such blockade may be applicable as therapy for hypertensive patients on CAPD.

Amlodipine↗

PubMed2003

Gastric angiodysplasia in patients undergoing maintenance dialysis.

We successfully used argon plasma coagulation (APC) to treat two cases of dialysis patients with hemorrhagic gastric angiodysplasia. Gastric angiodysplasia is recognized as an important cause of gastrointestinal bleeding. Angiodysplastic lesion confined to the gastric antrum was first described in 1953 and named gastric antral vascular ectasia (GAVE). The condition is characterized as submucosal capillary dilatation and fibromuscular hyperplasia. The typical finding of GAVE is the so-called watermelon stomach, attributable to vasodilatation. In case 1, a 69-year-old man was introduced continuous ambulatory peritoneal dialysis (CAPD) in July 1997 because of chronic renal failure due to nephrosclerosis. He was hospitalized for severe anemia in December 1997. Gastrointestinal fiberscopy (GIF) showed oozing in the antrum, and gastritis and esophagitis with sliding hernia. Famotidine was started and recombinant human erythropoietin (rHuEPO) was used for anemia. However, the severe anemia did not improve. The patient was hospitalized again for severe anemia and hematemesis. Another GIF showed typical watermelon stomach, which corresponded with GAVE. An APC was performed without complications. Three months later, the anemia was improved, and the dose of rHuEPO was reduced. In case 2, a 57-year-old woman was introduced to hemodialysis in 1998 for uremia due to nephrosclerosis. In October 2000, she was hospitalized for rHuEPO-resistant anemia. A GIF showed oozing in the antrum with diffuse vasodilation in the antrum; GAVE was diagnosed. An APC was carried out without complications. Three months later, anemia was improved. Recently, gastric angiodysplasia was reported to be an important complication in dialysis patients and was recognized as an important cause of rHuEPO-resistant anemia. Argon plasma coagulation is an effective treatment for gastric angiodysplasia in patients on dialysis.

Aged↗

PubMed2003

Role of adhesion molecules in the progression of peritoneal sclerosis.

To investigate the role of adhesion molecules [intracellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), and integrin alpha 5 beta 1] in the progression of encapsulating peritoneal sclerosis (EPS) under peritoneal dialysis, we examined changes in the expression of those adhesion molecules in Wistar-Kyoto (WKY) rats treated with acidic dialysis solution with or without angiotensin II type 1a receptor blocker (ARB). We divided 48 WKY rats into 4 groups and dialyzed them with various solutions as follows: (1) pH 7 1.5% glucose dialysis solution (control group, n = 12); (2) pH 3.5 1.5% glucose dialysis solution (EPS group, n = 12); (3) pH 3.5 1.5% glucose dialysis solution, plus oral administration of CS866 5 mg/kg daily (ARB group, n = 12); and (4) pH 3.5 1.5% glucose dialysis solution, plus oral administration of amlodipine (CA group, n = 12). We injected the dialysis solutions into the abdominal cavity and administered the ARB and CA daily for 42 days. On days 3, 7, 14, and 42, three rats in each group were humanely killed by decapitation, and we studied the expression of adhesion molecules in peritoneum by the immunofluorescence method. In the EPS rats, expression of adhesion molecules was observed in peritoneum on day 3 after start of acidic solution treatment, in conjunction with an increment of interleukin 6 (IL-6) in the dialysate. The peritoneum of EPS rats showed peritoneal fibrosis with interstitial cell infiltration. Treatment with ARB significantly suppressed expression of adhesion molecules in the peritoneum and suppressed peritoneal fibrosis. Treatment with a neutral solution induced no peritoneal fibrosis nor expression of adhesion molecules in the peritoneum. Our results suggest that adhesion molecules play an important role in the progression of peritoneal fibrosis and resultant EPS. Treatment with ARB prevents the progression of peritoneal fibrosis and suppresses expression of adhesion molecules in the peritoneum.

Angiotensin II Type 1 Receptor Blockers↗

PubMed2002

Accumulation of maize response regulator proteins in mesophyll cells after cytokinin treatment.

The maize response regulator genes ZmRR1 and ZmRR2 respond to cytokinin, and the translated products seem to be involved in nitrogen signal transduction mediated by cytokinin through the His-Asp phosphorelay. To elucidate the physiological function of the proteins, we examined the temporal and spatial distribution in maize leaves by immunochemical analysis and use of transgenic plants. ZmRR1 and ZmRR2 polypeptides could be distinctively detected by western blotting. The polypeptides accumulated in leaves within 5 h of the supply of nitrate to nitrogen-depleted maize, and the accumulation was transient. The extent of induction was larger in the leaf tip, which is rich in photosynthetically matured cells, than elsewhere. In leaves, the polypeptides accumulated mostly in mesophyll cells. Histochemical analyses of transgenic maize harboring a ZmRR1 promoter-beta-glucuronidase fusion gene also showed most of the expression to be in these cells. These results suggest that ZmRR1 and ZmRR2 are induced in mesophyll cells and function in nitrogen signal transduction mediated by cytokinin.

Cytokinins↗

PubMed2002

Glucocorticoid protects against the development of encapsulating peritoneal sclerosis on peritoneal dialysis.

Encapsulating peritoneal sclerosis (EPS) is an important complication of peritoneal dialysis in which all or part of the intestine is enveloped in a fibrous ball resulting in a cocoon. Previously, we reported that acid dialysis solution (DS) induces peritoneal fibrosis. In the present study, we examined the effect of treatment with glucocorticoid (GC) in a model of EPS in rats. We divided 20 male Wistar-Kyoto rats into four groups and dialyzed them with various solutions for 40 days as follows: (1) pH 3.5 DS, 10 mL (pH 3.5, containing 1.35% glucose, n = 5); (2) pH 3.5 DS, 10 mL + GC (0.1 mg dexamethasone daily, n = 5); (3) pH 7.0 DS, 10 mL (n = 5); and (4) pH 7.0 DS, 10 mL + GC (n = 5). At the end of 40 days, all rats were humanely killed by decapitation. Expression of mRNA of aquaporins (AQPs) and glucose transporters (GLUTs) were studied by reverse-transcriptase polymerase chain reaction. In rats treated with pH 3.5 DS, necropsy findings showed evidence of EPS. The typical appearance was multiple surfaces covered with granulation tissue or fibrotic tissue or both. Multiple adhesions were present. Microscopic findings revealed that low-pH DS induced peritoneal fibrosis and loss of mesothelium. In the dialyzed rats, mRNA of AQP-1, AQP-4, GLUT-1, GLUT-4, and GLUT-5 was expressed in peritoneum. In rats treated with pH 3.5 DS, expression of AQPs was significantly suppressed and expression of GLUTs was significantly enhanced. However, glucocorticoid treatment prevented the progression of peritoneal fibrosis and adhesion of peritoneum. In rats treated with pH 7.0 DS, no signs of EPS were seen. Our study suggests that low-pH DS induced the development of EPS. Glucocorticoid protects against the development of EPS on peritoneal dialysis.

Animals↗

PubMedDate not supplied

Molecular identification of the skin transformation center of anuran larval skin using genes of Rana adult keratin (RAK) and SPARC as probes.

Anuran larval skin undergoes a process of metamorphosis into pre-adult and adult skin. Basal skein, larval basal and adult basal cells are basement membrane-attaching cells in the larval, pre-adult and adult epidermis, respectively, and are identified as cells expressing genes of RLK (Rana larval keratin), both RLK and RAK (Rana adult keratin), and RAK. Larval to pre-adult skin conversion takes place in the histological entity called the skin transformation center (STC). The present study performed a cDNA subtractive gene screening on cDNA of the larval and the pre-adult skin, and cloned the secreted protein acidic and rich in cysteine (SPARC) gene as an upregulated gene in the larva to pre-adult skin conversion. RAK gene-positive basal skein cells and fibroblasts in and around the STC were weakly and strongly sparc-positive, respectively. Using sparc and rak, we redefined the STC and visualized it on a histological section as an approximately 150 microm-long region that contained about 20 rak-negative and weakly sparc-positive basal cells. Intense sparc expression was observed in basal skein cells, but not in larval basal cells, suggesting that SPARC acts as a suppressor of rak during epidermal differentiation. This suggestion was tested by investigating the effect of SPARC on cultured larval basal cells. We observed that SPARC suppressed the expression of rak, but not rlk.

Amino Acid Sequence↗

PubMedDate not supplied

Pancreatoblastoma. A case report with special emphasis on squamoid corpuscles with optically clear nuclei rich in biotin.

BACKGROUND: Pancreatoblastoma (PBL) is a rare neoplasm that generally occurs in the pediatric age group and shows unique histopathology, including squamoid corpuscles that may contain tumor cells with optically clear nuclei (OCN) rich in biotin. In the English-language literature there have been two reports on the cytology of PBL, but neither of them refers to the cytologic features of squamoid corpuscles. CASE: A 3-year-old boy with nausea and general fatigue was referred to our center. Imaging studies showed an approximately 7.5-cm, left-sided abdominal mass and multiple metastases in the lung. The abdominal mass was biopsied, and its histology showed solid cellular nests with occasional acinar differentiation and squamoid corpuscles. Imprint cytology of the biopsied sample displayed cellular epithelial nests with focal acinar structures and foci composed of larger cells with a low nuclear/cytoplasmic ratio. These foci contained a few tumor cells with biotin-rich OCN and were determined to be squamoid corpuscles. CONCLUSION: Detection of occasional squamoid corpuscles with biotin-rich OCN can be useful in making a diagnosis of PBL on cytologic samples.

Biomarkers, Tumor↗