PubMed Health⌕ Search

Biomedical subjects

K Ikebukuro

Publications and source records attributed to K Ikebukuro.

At least 37 records · Page 2Linked to original sources
PubMed2000

A new method for bone marrow cell harvesting.

To minimize contamination of bone marrow cells (BMCs) with T cells from the peripheral blood, a new "perfusion method" for collecting BMCs is proposed using cynomolgus monkeys. Two BM puncture needles are inserted into a long bone such as the humerus, femur, or tibia. One needle is connected to an extension tube and the end of the tube is inserted into a culture flask to collect the BM fluid. The other needle is connected to a syringe containing 30 ml of phosphate-buffered saline. The solution is pushed gently from the syringe into the medullary cavity, and the medium containing the BM fluid is collected into the culture flask. There is significantly less contamination with peripheral blood, determined from the frequencies of CD4(+) and CD8(+) T cells, when using this method (<6%) than when using the conventional method (>20%) consisting of multiple BM aspirations from the iliac crest. Furthermore, the number and progenitor activities of the cells harvested using this "perfusion method" are greater than those harvested using the conventional aspiration method. This perfusion method was carried out 42 times using 15 cynomolgus monkeys, and no complications such as pulmonary infarction or paralysis were observed. These findings suggest that the "perfusion method" is safe and simple and would be of great advantage in obtaining pure BMCs, resulting in a less frequent occurrence of acute graft-versus-host-disease in allogeneic BM transplantation.

Animals↗
PubMed1999

Application of peptide nucleic acid to the direct detection of deoxyribonucleic acid amplified by polymerase chain reaction.

Double-stranded DNA amplified by polymerase chain reaction (PCR) was detected by peptide nucleic acid (PNA) using a BIAcore 2000 biosensor based on surface plasmon resonance (SPR). PNA is an artificial oligo amide that is capable of forming highly stable complexes with complementary oligonucleotides. We succeeded in the direct detection of double-stranded DNA, amplified by PCR with high-sequence specificity. It was shown that the target DNA was available for detection over the range of 40-160 nM. Therefore, the detection limit was 7.5 pmol of the target DNA (143 bases, applied volume 30 microliters). Our DNA detection system, the combination of BIAcore and the probe PNA, could detect the target DNA with good reproducibility. In this report, we show that our system is a powerful tool for the diagnosis of pathologically significant DNA.

Bacterial Toxins↗
PubMed1999

Detection of PCR products in solution using surface plasmon resonance.

Polymerase chain reaction (PCR) products were detected using a flow injection-type sensor based on surface plasmon resonance. Asymmetric PCR was used to amplify the target DNA sequence, and two products with different length were produced. The novelty of our DNA detection system was that our target DNA was double stranded but the probe binding site, located in the 3'-terminus, was single stranded. This avoids the formation of intra- and intermolecular complexes. This novel design permitted us not only to detect PCR product but also to develop a rapid detection system for the detection of the verotoxin 2 gene of Escherichia coli O157:H7.

Bacterial Toxins↗
PubMed1999

Age-dependent abnormalities of hematopoietic stem cells in (NZW x BXSB)F1 mice.

The (NZW x BXSB)F1 (W/BF1) mouse is known as an autoimmune-prone strain which develops lupus nephritis, thrombocytopenia due to platelet-specific autoantibodies, leukocytosis, and myocardial infarction. In this experiment, we investigated the age-dependent abnormalities of the hematopoietic stem cells (HSCs) and hematopoiesis in this mouse. White blood cell counts (especially Mac-1- or Gr-1-positive cells) in the peripheral blood of 12-week-old W/BF1 mice increased in comparison with those of four-week-old W/BF1 or normal mice. To investigate whether the abnormal hematopoiesis can be attributed to the HSCs of W/BF1 mice, colony-forming unit in spleen (CFU-S) and colony-forming unit in culture (CFU-C) assays were performed. Day 12 CFU-S counts of 12-week-old W/BF1 mice significantly increased in comparison with those of four-week-old W/BF1 mice or normal mice. In the CFU-C assay, CFU-GEMM and CFU-GM counts in 12-week-old W/BF1 mice increased in comparison with those of four-week-old W/BF1 or control mice. The bone marrow cells (BMCs) from 12-week-old W/BF1 mice showed a high level of G-CSF and a low level of GM-CSF in mRNA expression. To examine the effect of HSCs from 12-week-old W/BF1 mice on the onset of autoimmune diseases and the abnormal hematopoiesis, T- and B-cell-depleted BMCs of four-week-old or 12-week-old W/BF1 mice were transplanted to C3H mice. Recipient C3H mice that had received the BMCs from 12-week-old W/BF1 mice showed an earlier onset of autoimmune diseases and a shorter survival rate than those that had received the BMCs from four-week-old W/BF1 mice. These data suggest that the HSCs from 12-week-old W/BF1 mice showing the symptoms of autoimmune diseases have the capacity to induce autoimmune diseases earlier than the HSCs from four-week-old W/BF1 mice.

Age of Onset↗
PubMed1999

Prenatally diagnosed cystic neuroblastoma.

An exceedingly rare case of prenatally diagnosed cystic neuroblastoma (NB) is described. Prenatal ultrasonography at 28 weeks' gestation revealed a cystic right suprarenal mass that measured 15 x 17 mm in diameter. The mass increased parallel to fetal size until it was 41 x 44 mm in diameter at 39 weeks' gestation. Magnetic resonance imaging performed prenatally and after delivery showed two different intracystic intensities with fluid levels, suggesting intracystic hemorrhage. The patient underwent a right adrenalectomy at 20 days of age, and the resected tumor was diagnosed as a poorly-differentiated ganglioneuroblastoma. Eight months after surgery, the patient is generally healthy with no evidence of recurrence. We reviewed 25 additional cases of prenatally diagnosed adrenal cystic NB and examined the clinical features of this rare entity.

Adrenal Gland Neoplasms↗
PubMed1999

Extensive epidural teratoma in early infancy treated by multi-stage surgery.

We report a rare case of extensive extradural teratoma successfully treated by multi-stage laminotomy and thoracotomy. A 34-day-old, dyspneic infant had a large posterior mediastinal mass identified on a chest X-ray radiograph. Imaging studies disclosed that the mass originated from the extradural space at the level of the lower thoracic spine, extending cephalad to C4 and caudad to L4 and severely compressing the spinal cord anteriorly, causing paraplegia. The tumor expanded bilaterally through the intraspinal foramina, coalescing to form a huge mediastinal mass. The upper half of the teratoma was removed utilizing a laminotomy from T3 through T9; 2 months later the lower half was excised via a laminotomy from T11 to L3. An additional procedure was required to resect recurrent tumor through a laminotomy from T8 to T12. The reconstructed vertebral arches were well-preserved in shape, with an almost normal spinal canal.

Epidural Neoplasms↗
PubMed1999

Apoptosis of colorectal adenocarcinoma induced by 5-FU and/or IFN-gamma through caspase 3 and caspase 8.

The cytotoxic effect of 5-FU has been shown by the induction of apoptosis in cancer cells, and reported to be enhanced by IFN-gamma. We examined the role of caspases on the apoptosis induction of 5-FU and IFN-gamma using a colorectal adenocarcinoma cell line. The activities of caspase 3 and caspase 8 increased when apoptosis was induced by 5-FU and/or IFN-gamma. Moreover, all apoptotic cells showed high caspase 3 activity in these conditions. Although the inhibitors of caspase 3 and caspase 8 inhibit apoptosis, anti-Fas ligand antibody does not affect the apoptosis induced by 5-FU. Thus, caspase 3 and caspase 8 play crucial roles in apoptosis induced by 5-FU and/or IFN-gamma, regardless of the Fas-Fas ligand system.

Adenocarcinoma↗
PubMed1998

Biosensors for environmental monitoring.

Environmental monitoring is of great importance for its protection. Conventional monitoring methods are often slow and complex and require expensive equipment, making them unsuitable for in situ, real-time monitoring of pollutants. Biosensors based on a combination of a biological sensing element and an electronic signal-transducing element are alternative methods to conventional ones. Biosensors have a number of advantages, such as high selectivity, high stability, and short response time. Various kinds of biosensors have been developed and employed for detection of pollutants such as phosphate, cyanide, and herbicides. Some of these have already been exploited as real-time monitoring in situ. In this article, some of the applications of biosensors for environmental control are described.

Biosensing Techniques↗
PubMed1998

Application of a linear alkylbenzene sulfonate biosensor to river water monitoring.

A novel whole cell biosensor was constructed for the detection of anionic surfactants in aquatic environments. The analysis was rapid, convenient and did not require organic reagents. In this report, the application of this sensor to river water samples was investigated when applied to environmental samples; other organic substances present in river water may affect the measurement of linear alkylbenzene sulfonates. In order to deal with this problem, a correction system was developed using whole cells of Trichosporon cutaneum. This system was applied to in situ 24 h continuous monitoring in the Saka river.

Benzenesulfonates↗
PubMed1998

Quantitative analysis of polymerase chain reaction using anisotropy ratio and relative hydrodynamic volume of fluorescence polarization method.

The method based on the combination of polymerase chain reaction (PCR) and fluorescence polarization is presented. A targeted DNA was amplified with a 5'-fluorescein labeled primer, using a 256 bp DNA fragment of stx2 gene in Escherichia coli O157:H7 (188-443 bp) as a template. The fluorescence anisotropy of the 5'-fluorescein labeled primer increased upon the polymerization through Taq polymerase. The conversion of primer to PCR product was quantitatively monitored by anisotropy ratio and relative hydrodynamic volume. This system was also applied to the determination of E.coli O157:H7.

Bacterial Toxins↗
PubMed1998

Investigation of the potential active site of a cyanide dihydratase using site-directed mutagenesis.

Cyanide dihydratase has conserved residues in the amino acid sequence with nitrilase, and cyanide hydratase. The conserved amino acid residues in the cyanide dihydratase from Pseudomonas stutzeri AK61 were altered by site-directed mutagenesis. The enzyme completely lost its activity of the cyanide hydrolysis by the replacement of cysteine-163 to serine. The replacement of tyrosine-53 to phenylalanine caused an increase of the K(m) value of the enzyme for cyanide. Substitution of nine other residues seemed to affect the structure of the enzyme.

Amino Acid Sequence↗
PubMed1998

Cloning and expression of a gene encoding cyanidase from Pseudomonas stutzeri AK61.

The gene coding for cyanidase, which catalyzes the hydrolysis of cyanide to formate and ammonia, was cloned from chromosomal DNA of Pseudomonas stutzeri AK61 into Escherichia coli. The cyanidase gene consisted of an open reading frame of 1004 bp, and it was predicted that cyanidase was composed of 334 amino acids with a calculated molecular mass of 37,518 Da. The amino acid sequence of cyanidase showed a 35.1% and 26.4% homology to aliphatic nitrilase from Rhodococcus rhodochrous K22 and cyanide hydratase from Fusarium lateritium, respectively. A unique cysteine residue of aliphatic nitrilase, which was suggested to play an essential role in the catalytic activity, was conserved in cyanidase. The active form of cyanidase was successfully expressed by a DNA clone containing the cyanidase gene in E. coli. Its productivity was approximately 230 times larger than that of P. stutzeri AK61. The characteristics of the expressed cyanidase, including optimum pH, optimum temperature. Michaelis constant (K(m)) for cyanide and specific activity, were similar to those of the native enzyme from P. stutzeri AK61.

Amino Acid Sequence↗
PubMed1998

Complete resection is not required in patients with neuroblastoma under 1 year of age.

BACKGROUND/PURPOSE: The prognosis of neuroblastoma (NB) patients when detected before 1 year of age is excellent, particularly in patients whose disease is detected by screening. In some institutions patients with stage I or II NB detected by screening have been observed closely without any treatment. Most showed tumor regression or maturation and are surviving without tumor resection. Resection of infantile NB sometimes is complicated by vascular accidents resulting in vanishing kidney or intestinal infarction. The role of surgery in the treatment of infantile NB therefore is becoming controversial. The authors have treated infants who have easily resectable primary tumors with resection and sampling for lymph node metastases. If the resection of the primary tumor seemed difficult, only biopsy was performed initially followed by chemotherapy with or without resection. The authors analyzed the clinical outcome of the infantile NB in their institution to elucidate the role of surgery in this particular group of patients. METHODS: Thirty-six patients less than 12 months of age were treated between 1982 and 1997. Twenty-five patients who showed no symptoms had NB detected by screening at 6 months. Five patients had stage IVS disease in early infancy. No patients had bone or remote lymph node metastases. N-mycamplification was not detected in any of the 28 tumors examined. There were four diploid and 12 near-triploid tumors. RESULTS: Complete resection was possible in 13 stage I and II patients without any complications. Obvious lymph node metastases were not resected in 13 patients with stage II or III NB. Partial resection leaving gross residual disease was accomplished in three patients with stage III NB. Surgical resection was not attempted in four patients because of unresectability in three patients with stage III or IVS disease and because of marked tumor shrinkage after a short course of chemotherapy in one patient with stage III NB involving the celiac axis. A neonatal cervicomediastinal stage III NB recurred very rapidly after partial resection but started to regress spontaneously 2 months postoperatively. Another neonatal stage II cervical NB progressed to stage IVS with hepatomegaly causing respiratory distress but regressed spontaneously. All the patients are alive and disease free without surgical morbidity. CONCLUSION: All of the 21 patients who underwent partial tumor resection with or without chemotherapy are alive and doing well without tumor, indicating that complete resection is not required in this particular group of patients.

Age Factors↗
PubMed1997

The conserved residue tyrosine 34 is essential for maximal activity of iron-superoxide dismutase from Escherichia coli.

We have expressed, purified, and analyzed the iron-containing superoxide dismutase (FeSOD) of Escherichia coli with mutations directed at tyrosine position 34 to introduce phenylalanine (SODY34F), serine (SODY34S), or cysteine (SODY34C). FeSOD and mutant enzymes were purified from SOD-deficient cells using a GST-FeSOD fusion protein intermediate which was subsequently cleaved with thrombin and repurified. Specific activities were measured using the xanthine-xanthine oxidase method and gave 3148 u/mg for wild-type FeSOD. The SODY34S mutation virtually inactivates the enzyme (42 u/mg); mutation to cysteine greatly reduces activity (563 u/mg), but the SODY34F mutant retains nearly 40% of the activity of wild type (1205 u/mg). Fusion protein intermediates were also shown to be active and were demonstrated to protect SOD-deficient E. coli cells from the induced effects of oxidative stress, with growth rates directly proportional to the specific activities of the expressed mutant enzymes. SODY34F exhibited decreased thermal stability, reduced activity at high pH, and a pronounced increase in sensitivity to the inhibitor sodium azide compared with wild-type FeSOD. These results suggest that tyrosine at position 34 is multifunctional and plays a structural role (probably through hydrogen bonding to glutamine at position 69) in maintaining the integrity of the active site, a stabilizing role at high pH, and a steric role in obstructing access to the active site of both substrate and inhibitor molecules.

Amino Acid Sequence↗
PubMed1997

A chemiluminescent FIA biosensor for phosphate ion monitoring using pyruvate oxidase.

We have constructed an automatic phosphate ion sensing system for the quality control of drinking water. The analyte was detected using the phosphate ion-dependent pyruvate oxidase reaction and the hydrogen peroxide produced was detected by luminol chemiluminescence catalyzed by Arthromyces ramosus peroxidase. We obtained a detection limit of 0.16 microM phosphate ion (5 ppb phosphorus) and it was possible to detect 0.32 microM phosphate ion for 48 days using pyruvate oxidase immobilized on Chitopearl BCW-2601 beads. An excellent correlation (r2 = 1.00) was obtained between the results obtained using our phosphate ion sensor and those using a modified Molybdenum Blue method.

Automation↗
PubMed1997

Novel detection system of flow injection analysis (1). The existence of significant relation between secondary structure of DNA and sensitivity in signal detection.

Polymerase Chain Reaction (PCR) products was detected quantitatively using a flow injection type sensor, based on Surface Plasmon Resonance (SPR). We used asymmetric PCR to amplify the two kinds of products; their DNA lengths are different. This novel design permitted us not only to detect PCR products with high-sensitivity, but also to develop a rapid DNA detection system for the sense of the genetic pathogen.

Bacterial Toxins↗
PubMed1997

Thrombin-binding properties of thrombin aptamer derivatives.

We synthesized a single-stranded DNA with triple G-quartet structure and found that its thrombin-binding activity at 37 degrees C is as much as that of thrombin aptamer with double G-quartet structure which is a product of in vitro selection and amplification. This result suggests that the sequence motifs obtained by in vitro selection and amplification method might not be the only answer.

Base Sequence↗