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Biomedical subjects

H Murakami

Publications and source records attributed to H Murakami.

At least 127 records · Page 7Linked to original sources
PubMed2000

Analysis of segmental renal gene expression by laser capture microdissection.

UNLABELLED: Analysis of segmental renal gene expression by laser capture microdissection. BACKGROUND: The study of normal renal physiology has been greatly aided by microdissection techniques that have delineated the exceptional functional and cellular heterogeneity both along the nephron and between different nephron populations. These techniques are not widely used to study renal injury as microdissection is difficult because of tissue necrosis or fibrosis. We developed a procedure to detect specific gene expression in specific locations of the kidney in histologic sections. METHODS: The anatomic specificity of laser capture microdissection (LCM) was employed with the sensitivity of reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: LCM/RT-PCR detected mRNA for podoplanin in 2% of a single glomerulus, rat basic amino acid transporter in 6% of a single cross-section of proximal straight tubule, and renin in eight proximal convoluted tubule cross-sections. LCM/RT-PCR could isolate pure populations of proximal convoluted tubules, proximal straight tubules, and thick ascending limbs from renal histologic sections, although pure collecting ducts could not be isolated. LCM/RT-PCR localized ischemia-reperfusion-induced induction of KC/interleukin-8 primarily to the medullary thick ascending limb, and detected transforming growth factor-beta (TGF-beta) mRNA in glomeruli of a patient with membranous glomerulonephropathy. CONCLUSIONS: When used with an appropriate laser spot size, LCM/RT-PCR can measure gene expression in glomeruli or specific parts of the nephron and can study alterations in steady-state mRNA levels in animal models of renal disease. The applications, limitations, and refinements of this approach are discussed.

Animals↗
PubMed2000

Low power laser protects human erythrocytes In an In vitro model of artificial heart-lung machines.

The protective effect of the low power helium-neon (He-Ne) laser against the damage of human erythrocytes in whole blood was examined in a perfusion model using an artificial heart-lung machine. Preserved human whole blood was diluted and perfused in 2 closed circuits with a double roller pump. The laser irradiated one of the circuits (laser group), and none the other (control group). In the laser group, erythrocyte deformability and erythrocyte adenosine triphosphate (ATP) levels were significantly higher, and free hemoglobin levels were significantly lower than those in the control group. Subsequent morphological findings by means of scanning electron microscope were consistent with these results. Low power He-Ne laser protected human erythrocytes in the preserved diluted whole blood from the damage caused by experimental artificial heart-lung machines. The clinical application of low power laser treatment for extracorporeal circulation is suggested.

Adenosine Triphosphate↗
PubMed2000

High incidence of SV40-like sequences detection in tumour and peripheral blood cells of Japanese osteosarcoma patients.

Recent studies have revealed the evidence for the significance of SV40 genome in human malignancies. In this paper, the presence of SV40-like sequences was investigated in 54 Japanese osteosarcomas in which mutations of the retinoblastoma (Rb), p53, MDM2, and CDK4 genes had been already analysed. Using polymerase chain reaction and Southern hybridization, SV40-like sequences were detected in 25 cases (46.3%). In most cases, only a part of SV40 genome was detected, and the regulatory region containing enhancer sequences was most frequently found (21/54, 38.9%). There was no apparent relationship between the presence of SV40-like sequences and tumour suppressor genes mutations in each tumour. The SV40-like sequences were also detected in peripheral blood cells of substantial proportion of the patients (43.3%), whereas the incidence was much lower (4.7%) in normal healthy controls. This difference is statistically highly significant (P < 0.0001), suggesting that the presence of SV40-like sequences, even if only a part, may play some roles to predispose individuals to osteosarcoma.

Adolescent↗
PubMed2000

Relationship between time after intake of grapefruit juice and the effect on pharmacokinetics and pharmacodynamics of nisoldipine in healthy subjects.

A clinical study was performed in eight healthy volunteers to investigate the effect of various timing of grapefruit juice intake on nisoldipine pharmacokinetics and pharmacodynamics, and to validate our pharmacokinetic model. The subjects were given 10 mg oral nisoldipine with water (control), or 5 mg oral nisoldipine with 200 mL grapefruit juice (G0) or with water at 14 (G14), 38 (G38), 72 (G72) or 96 hours (G96) after a 7-day period of thrice-daily intake of grapefruit juice. Grapefruit juice ingestion did not affect heart rate or the effect area during the first 8 hours of heart rate after nisoldipine administration, although significant decreases of systolic and diastolic blood pressure were caused in G0 by coadministration of grapefruit juice with nisoldipine. Headaches were reported by 3, 2, and 1 persons in G0, G14, and G38, respectively, but no subjects in G72 and G96 reported headaches. Compared with the control group, the maximum plasma concentration of nisoldipine was significantly increased after grapefruit juice intake in G0 and G14, and the plasma concentration was significantly increased at each time in G0 to G72. Therefore the effect of grapefruit juice decreased time dependently and lasted for at least 3 days after intake. Furthermore, our model gave predicted values in good agreement with the observed values. It is therefore necessary to withhold grapefruit juice for at least 3 days before administration of the drug to prevent grapefruit juice-nisoldipine interaction.

Adult↗
PubMed2000

Acetabular augmentation using a glass-ceramic block: 3 patients followed 3-4 years.

We have developed a block of glass-ceramic to augment the dysplastic acetabulum. 3 patients with acetabular dysplasia underwent implantation of a block of glass-ceramic on the lateral surface of the ilium just above the hip joint. The patients did not require immobilization and returned to their daily lives, walking without a cane 4 weeks after the operation. The mean Harris hip score was 47 points preoperatively and 94 points 3 years postoperatively.

Acetabulum↗
PubMed2000

True aneurysm of the digital artery. Case report and review.

We report a case of true aneurysm of the ulnar digital artery, the aetiology of which was related to gripping a golf club. It was diagnosed preoperatively by magnetic resonance imaging. The patient made a good recovery from excision, and the aneurysm had not recurred two years later.

Adult↗
PubMed2000

Association between p53 immunostaining and cigarette smoking in squamous cell carcinoma of the esophagus.

BACKGROUND: It is generally accepted that cigarette smoking is closely associated with esophageal squamous cell carcinoma. This study investigated the molecular targets of cigarette smoke in carcinogenesis of the esophagus. METHODS: Seventy-four patients with esophageal squamous cell carcinoma (SCC) were grouped according to daily cigarette consumption: heavy smoking group (group H) (n = 26), moderate smoking group (group M) (n = 39) and non-smoking group (group N) (n = 9). We compared p53 and retinoblastoma (RB) expression among the three groups by immunohistochemistry. In addition, fresh tumor tissues from 30 smokers with esophageal SCC were tested for p53 mutations in exons 5-8 by direct sequencing. RESULTS: Staining for the p53 product was positive in 65.4% of group H, 38.5% of group M and 44.4% of group N. The frequency of positive staining in the group H was significantly higher than in group M (p = 0.033) and in group M + group N (p = 0.034). The difference with respect to the frequency of overexpression of RB was not significant. The patterns of p53 base-pair mutations in direct sequencing study were of five types, most commonly G:C to T:A transversion (35.3%). CONCLUSIONS: Our study suggests that one of the molecular targets of cigarette smoke is the p53 gene. The pattern of p53 point mutations involved a wide range of base-pair changes.

Aged↗
PubMed2000

Random insertion and deletion mutagenesis for construction of protein library containing nonnatural amino acids.

A new method was developed for the generation of a library of mutant proteins that contained nonnatural amino acids. The method, "random insertion and deletion (RID) mutagenesis", is based on the deletion of an arbitrary number of bases at random positions and, at the same time, the insertion of an arbitrary sequence into the same position. By using this method, randomly selected three consecutive bases in the gene of green fluorescence protein (GFP) were replaced by a CGGT 4-base codon. When this DNA library was expressed in E. coli, about 80% of colonies lost the fluorescence. The non-fluorescent colonies were picked up and the genes were sequenced. Replacement of three consecutive bases by CGGT 4-base codon was found in two of the four mutated genes.

Amino Acids↗
PubMed2000

The contribution of skeletal muscle tumor necrosis factor-alpha to insulin resistance and hypertension in fructose-fed rats.

OBJECTIVE: The aim of this study was to determine the role of tumor necrosis factor-alpha (TNF-alpha) in skeletal muscle tissue in insulin resistance and hypertension and the effect of anti-hypertensive medicine on skeletal muscle TNF-alpha in fructose-induced insulin-resistant and hypertensive rats (fructose-fed rats: FFR). DESIGN AND METHODS: Six-week-old male Sprague-Dawley rats were fed either normal rat chow or fructose-rich chow. For the last 2 weeks of a 6-week period of either diet, the rats were treated with a vehicle (control or FFR); temocapril, an angiotensin converting enzyme inhibitor (ACEI); or CS-866, an angiotensin II type 1 receptor blocker (ARB). The euglycemic hyperinsulinemic glucose clamp technique was performed to evaluate insulin sensitivity (M value). TNF-alpha levels in soleus and extensor digitorum longus (EDL) muscles and epididymal fat pads were measured. We also measured the TNF-alpha concentration in an incubated medium secreted from soleus muscle strips with or without angiotensin II. RESULTS: TNF-alpha levels were significantly higher in the soleus and EDL muscles, but not in the epididymal fat, in the FFRs compared with the control rats. Temocapril and CS-866 lowered systolic blood pressure, improved insulin resistance, and reduced TNF-alpha in both skeletal muscles. There were significant negative correlations between M values and TNF-alpha levels in both soleus and EDL muscles. Also, the soleus muscle strip incubation with 10(-7) mol/l angiotensin II increased TNF-alpha secreted into the incubation medium compared to the incubation without angiotensin II. These results suggest that skeletal muscle TNF-alpha is linked to insulin resistance and hypertension and that angiotensin II may be one of the factors that regulate skeletal muscle TNF-alpha.

Adipose Tissue↗
PubMed2000

Contribution of a thickened cell wall and its glutamine nonamidated component to the vancomycin resistance expressed by Staphylococcus aureus Mu50.

Staphylococcus aureus Mu50, which has reduced susceptibility to vancomycin, has a remarkably thickened cell wall with an increased proportion of glutamine nonamidated muropeptides. In addition, Mu50 had enhanced glutamine synthetase and L-glutamine D-fructose-6-phosphate aminotransferase activities, which are involved in the cell-wall peptidoglycan synthesis pathway. Furthermore, significantly increased levels of incorporation of (14)C-labeled D-glucose into the cell wall was observed in Mu50. Unlike a femC mutant S. aureus strain, increased levels of production of nonamidated muropeptides in Mu50 was not caused by lower levels of glutamine synthetase activity but was considered to be due to the glutamine depletion caused by increased glucose utilization by the cell to biosynthesize increased amounts of peptidoglycan. After the cells were allowed to synthesize cell wall in the absence or presence of glucose and glutamine, cells with different cell-wall thicknesses and with cell walls with different levels of cross-linking were prepared, and susceptibility testing of these cells demonstrated a strong correlation between the cell-wall thickness and the degree of vancomycin resistance. Affinity trapping of vancomycin molecules by the cell wall and clogging of the outer layers of peptidoglycan by bound vancomycin molecules were considered to be the mechanism of vancomycin resistance of Mu50. The reduced cross-linking and the increased affinity of binding to vancomycin of the Mu50 cell wall presumably caused by the increased proportion of nonamidated muropeptides may also contribute to the resistance to some extent.

Bacterial Proteins↗
PubMed2000

Genetic studies with the fission yeast Schizosaccharomyces pombe suggest involvement of wee1, ppa2, and rad24 in induction of cell cycle arrest by human immunodeficiency virus type 1 Vpr.

Accessory protein Vpr of human immunodeficiency virus type 1 (HIV-1) arrests cell cycling at G(2)/M phase in human and simian cells. Recently, it has been shown that Vpr also causes cell cycle arrest in the fission yeast Schizosaccharomyces pombe, which shares the cell cycle regulatory mechanisms with higher eukaryotes including humans. In this study, in order to identify host cellular factors involved in Vpr-induced cell cycle arrest, the ability of Vpr to cause elongated cellular morphology (cdc phenotype) typical of G(2)/M cell cycle arrest in wild-type and various mutant strains of S. pombe was examined. Our results indicated that Vpr caused the cdc phenotype in wild-type S. pombe as well as in strains carrying mutations, such as the cdc2-3w, Deltacdc25, rad1-1, Deltachk1, Deltamik1, and Deltappa1 strains. However, other mutants, such as the cdc2-1w, Deltawee1, Deltappa2, and Deltarad24 strains, failed to show a distinct cdc phenotype in response to Vpr expression. Results of these genetic studies suggested that Wee1, Ppa2, and Rad24 might be required for induction of cell cycle arrest by HIV-1 Vpr. Cell proliferation was inhibited by Vpr expression in all of the strains examined including the ones that did not show the cdc phenotype. The results supported the previously suggested possibility that Vpr affects the cell cycle and cell proliferation through different pathways.

Cell Cycle↗
PubMed2000

Fission yeast Eso1p is required for establishing sister chromatid cohesion during S phase.

Sister chromatid cohesion is essential for cell viability. We have isolated a novel temperature-sensitive lethal mutant named eso1-H17 that displays spindle assembly checkpoint-dependent mitotic delay and abnormal chromosome segregation. At the permissive temperature, the eso1-H17 mutant shows mild sensitivity to UV irradiation and DNA-damaging chemicals. At the nonpermissive temperature, the mutant is arrested in M phase with a viability loss due to a failure to establish sister chromatid cohesion during S phase. The lethal M-phase arrest phenotype, however, is suppressed by inactivation of a spindle checkpoint. The eso1(+) gene is not essential for the onset and progression of DNA replication but has remarkable genetic interactions with those genes regulating the G(1)-S transition and DNA replication. The N-terminal two-thirds of Eso1p is highly homologous to DNA polymerase eta of budding yeast and humans, and the C-terminal one-third is homologous to budding yeast Eco1p (also called Ctf7p), which is required for the establishment of sister chromatid cohesion. Deletion analysis and determination of the mutation site reveal that the function of the Eco1p/Ctf7p-homologous domain is necessary and sufficient for sister chromatid cohesion. On the other hand, deletion of the DNA polymerase eta domain in Eso1p increases sensitivity to UV irradiation. These results indicate that Eso1p plays a dual role during DNA replication. The C-terminal region acts to establish sister chromatid cohesion, and the N-terminal region presumably catalyzes translesion DNA synthesis when template DNA contains lesions that block regular DNA replication.

Acetyltransferases↗
PubMed2000

Comparison of blood-brain barrier permeability in mice and rats using in situ brain perfusion technique.

Here we present a method for measuring the permeability coefficient-surface area product (PS) values at the blood-brain barrier in mice, using the in situ brain perfusion technique originally developed for rats by Takasato et al. (Am J Physiol Heart Circ Physiol 247: H484-H493, 1984). Retrograde infusion into the right external carotid artery increased the carotid perfusion pressure in proportion to the perfusion rate. Intravascular volume and cerebral perfusion fluid flow at a perfusion rate of 1.0 ml/min in mice were similar to those in rats. In addition, the contribution of systemic blood to total flow in the hemisphere was small (only 3. 2%). These findings indicated that this perfusion rate is suitable for mice. The PS values of more than 20 different compounds were determined in mice by using the in situ brain perfusion technique, and comparisons were made with data from rats. There was a close relationship (1:1) between the PS values in mice and rats, indicating that brain capillary permeabilities are similar in mice and rats.

Animals↗
PubMed2000

Electrophysiologic analysis of antidepressant drug effects on the GABA(A) receptor complex based upon antagonist-induced encephalographic power spectrum changes.

To better understand antidepressant drug effects on the GABA(A) receptor complex (the GABA(A) receptor, chloride ionophore and benzodiazepine receptor), we investigated how antidepressants influenced power spectrum changes induced by pentylenetetrazol (PTZ), a chloride ionophore antagonist, in the rat hippocampal electroencephalogram (EEG). In control recording, PTZ (27.5 mg/kg i. p.) increased EEG power at frequencies under 12 Hz up to five times. After rats were pretreated with imipramine, fluoxetine or trazodone for 7 days (10 mg/kg i.p., twice a day), PTZ could not increase EEG power to more than three times the power before injection; this effect was not observed after pretreatment for 3 days. These three antidepressants inhibit serotonin uptake, while two other antidepressants, desipramine and nortriptyline, that inhibit norepinephrine uptake failed to counter the PTZ effect. We concluded that antidepressants with serotonergic effects enhanced the function of the GABA(A) receptor complex.

Animals↗
PubMed2000

Meniscal tearing after ACL reconstruction.

The knees of 72 patients with unilateral anterior-cruciate- ligament (ACL) injury were analyzed before ACL reconstruction as well as by follow-up arthroscopy on the day of staple removal. At ACL reconstruction 31 lateral menisci and 40 medial menisci were found to be normal. 28 lateral menisci and 24 medial menisci were treated surgically, while 13 lateral menisci and 8 medial menisci with small or incomplete meniscal tearing were not treated. At follow-up arthroscopy there were 3 new cases of lateral meniscal tearing and 3 new cases of medial meniscal tearing in the groups diagnosed as normal prior to surgery. Two of the 13 cases with small or incomplete lateral meniscal tearing required resection, 8 healed and the other 3 demonstrated no progressive change. Four of the 8 cases with small or incomplete medial meniscal tears healed, 3 exhibited no progressive change and one required surgical treatment. There was no correlation between meniscal tearing and knee instability as indicated by a positive Lachman test or a positive pivot shift sign. The results of the present study indicate that ACL reconstruction prevents progressive changes in meniscal tears and will prevent secondary osteoarthritis, and that some small tears of the lateral meniscus require no surgical treatment.

Journal Article↗
PubMed2000

Evidence for the AT1 subtype of the angiotensin II receptor in the rat submandibular gland.

We have characterized angiotensin II (Ang II) receptor subtypes on rat submandibular gland membranes using a radioligand binding assay. [3H]Ang II binding to the membrane fractions exhibited both high (Kd =0.08 nm, Bmax =2.19 fmol/mg protein) and low (Kd =4.19 nm, Bmax = 13.7 fmol/mg protein) affinity. Ang 11, Ang III and saralasin completely displaced the [3H]Ang II binding, whereas CV-11974, an AT1 receptor antagonist and PD123319, an AT2 receptor antagonist maximally displaced up to approximately 87 and 13% of the total binding, respectively. [3H]DuP753 binding to the membrane fractions exhibited a single population of binding site with a Kd of 4.22 nM and Bmax of 3.77 pmol/mg protein. Ang II, Ang III and CV-11974 completely displaced the [3H]DuP753 binding with slope factors near unity, but PD123319 did not. These findings suggest that rat submandibular gland membranes contain predominantly the AT1 receptor subtype.

Angiotensin Receptor Antagonists↗
PubMed2000

Synthesis of novel heterobranched beta-cyclodextrins from 4(2)-O-beta-D-galactosyl-maltose and beta-cyclodextrin by the reverse action of pullulanase, and isolation and characterization of the products.

From the mixture of 4(2)-O-beta-D-galactosyl-maltose (Gal-G2) and beta-cyclodextrin (betaCD), novel heterobranched betaCDs, (Gal-G2)-betaCD and (Gal-G2)2-betaCDs, were synthesized by the reverse action of debranching enzyme. The optimum conditions for the production of (Gal-G2)2-betaCDs were examined. A mixture of (Gal-G2)2-betaCDs was produced in about 4% yield when Aerobacter aerogenes pullulanase (64 units per 1 g of Gal-G2) was incubated with 1.6 M Gal-G2 and 0.16 M betaCD at 50 degrees C for 4 days. The reaction products, (Gal-G2)2-betaCDs, were separated into three peaks by HPLC analysis on a Hypercarb S column. Their structures were analyzed by fast atom bombardment mass spectroscopy and NMR spectroscopies, and confirmed by comparison of their hydrolyzates by beta-galactosidase with the authentic (G2)2 -betaCDs. The structures of (Gal-G2)-betaCD and three components of (Gal-G2)2-betaCDs were identified as 6-O-(GalG2)-betaCD, 6(1),6(2)-, 6(1),6(3)- and 6(1),6(4)-di-O-(Gal-G2)2-betaCD, respectively.

Carbohydrate Sequence↗