Biomedical subjects

S Yamano

Publications and source records attributed to S Yamano.

At least 55 records · Page 3Linked to original sources

PubMed1999

Z-line structural diversity in frog single muscle fiber in the passive state.

The structural changes of the Z-line between small square net (ss) and basket weave (bw) cross-sectional patterns were examined using intact single fibers and mechanically skinned fibers in the passive state to determine if the pattern is related to the sarcomere length (SL) and if the pattern undergoes a reversible transition in low- and high-osmotic medium. Frog single fibers were isolated from the anterior tibial muscle in Ringer's solution. Entirely or partially skinned single fibers were prepared in relaxing solution (also called low-osmotic medium). The high osmotic medium contained 10% polyvinylpyrrolidone (PVP) in relaxing solution. The sarcomere length (SL) of each fiber was measured directly by use of a laser beam or indirectly from electron micrographs with use of a correction factor. The ss and bw forms in cross sections were quantified by analysis of electron micrographs. The results show that the structural change of Z-line occurs around bw << 2.3-2.4 microns << ss (n = 25) and bw << 3.1-3.2 microns << ss (n = 13) in intact single fibers and skinned fibers, respectively. With the quick freeze-freeze substitution method, an intact single fiber with a SL of 2.35 microns showed almost 100% of ss form. The structural transition in cross section was also confirmed in four partially skinned fibers, where patterns went from mostly ss form (intact portion) to mostly bw form (skinned portion) at the SL between 2.40 to 3.20 microns. The reversibility of the change between ss and bw was proved by using low- and high-osmotic medium. The transition and reversion of cross-sectional patterns both occur in the passive state.

Animals↗

PubMed1999

Purification and characterization of rat liver naloxone reductase that is identical to 3alpha-hydroxysteroid dehydrogenase.

1. Rat liver cytosol produced exclusively 6beta-naloxol from naloxone in the presence of either NADPH or NADH at pH 7.4. The amount of 6beta-naloxol formed with NADPH was about four times that with NADH. The enzyme responsible for this reaction, termed naloxone reductase, was purified to a homogeneous protein by various chromatographic techniques. 2. The purified enzyme is a monomeric protein with a molecular weight of 34000 and an isoelectric point of 5.9, and it has a dual co-factor specificity for NADPH and NADH. The enzyme catalysed the reduction of various carbonyl compounds as well as naloxone analogues, and the dehydrogenation of 3alpha-hydroxysteroids and alicyclic alcohols. Indomethacin, quercetin and sulphhydryl reagents potently inhibited the enzyme, but pyrazole and barbital had no effect on the enzyme activity. 3. Identity of naloxone reductase and 3alpha-hydroxysteroid dehydrogenase in rat liver was demonstrated by comparing the elution profiles of the two enzyme activities during purification, the ratios of the two enzyme activities at each purification steps, and thermal stability and susceptibility to inhibitors for the two enzyme activities. 4. Amino acid sequences of five peptides obtained by proteolytic digestion of the purified enzyme were completely identical to the corresponding regions of previously reported 3alpha-hydroxysteroid dehydrogenase.

3-Hydroxysteroid Dehydrogenases↗

PubMed1999

Expression and characterization of rat UDP-N-acetylglucosamine: alpha-3-D-mannoside beta-1,2-N-acetylglucosaminyltransferase I in Saccharomyces cerevisiae.

The yeast Saccharomyces cerevisiae is a useful host for the production of heterologous proteins through the secretory pathway. However, because of the potential antigenicity of mannan-type sugar chains in humans, yeast cannot be used as a host for the production of glycoprotein therapeutics. To overcome this problem, we are trying to breed a yeast which can produce hybrid- or complex-type carbohydrates. UDP- N- acetylglucosamine:alpha-3-d-mannoside beta-1, 2- N- acetylglucosaminyltransferase I (GnT-I) is essential for the conversion of high mannose-type N- glycans to hybrid- and complex-type ones. As yeast lacks this enzyme, we have introduced the rat GnT-I cDNA into yeast cells. The transformed yeast cells expressed GnT-I activity in vitro. The expressed GnT-I was localized in all organella, including the endoplasmic reticulum (ER), Golgi apparatus, and vacuole, suggesting that the mammalian Golgi retention signal of GnT-I did not function in yeast cells. Analysis of the GnT-I gene product with a c-Myc epitope tag at the C-terminus elucidates that the N - terminal region of GnT-I, including the mammalian Golgi retention signal, should be removed in the yeast ER.

Animals↗

PubMed1999

An azide-insensitive superoxide dismutase from a hyperthermophilic archaeon, Sulfolobus solfataricus.

The superoxide dismutase (SOD) gene of Sulfolobus solfataricus, a hyperthermophilic archaeon, was cloned and expressed in Escherichia coli, and its gene product was characterized. When the protein was expressed in E. coli, it formed a homodimer that contained both Mn and Fe. Metal reconstitution experiments of the SOD with Fe or Mn showed that only the Fe-reconstituted SOD was active. Substitution of Tyr88 to Phe did not affect the metal specificity of the enzyme. The Fe-reconstituted SOD was extremely resistant to thermal denaturation; e.g. 96% of the initial activity was retained after heating at 95 degreesC for 2 h. Fe-reconstituted SOD was not inhibited by azide, but fluoride inhibition was observed. This suggests that some steric hindrance in the substrate funnel of the enzyme prevents the access of N3- but allows O2- and F- access to the active site.

Amino Acid Sequence↗

PubMed1999

A cambialistic SOD in a strictly aerobic hyperthermophilic archaeon, Aeropyrum pernix.

The superoxide dismutase (SOD) gene of Aeropyrum pernix, a strictly aerobic hyperthermophilic archaeon, was cloned and expressed in Escherichia coli, and its gene product was characterized. The molecular mass of the protein, based on the deduced amino acid sequence, was 24.6 kDa. The sequence showed overall similarity to the sequences of known Mn- and Fe-SODs. The metal binding residues conserved in Mn- and Fe-SODs were also found in A. pernix SOD. When the SOD gene was expressed in E. coli cells, the product formed a homodimer, and contained both Mn and Fe. Metal reconstitution experiments showed that A. pernix SOD is cambialistic, i.e. active with either Fe or Mn. The specific activities were 906 U/mg with Mn and 175 U/mg with Fe. No loss of activity of Mn-reconstituted SOD was observed at 105 degrees C even after 5 h incubation. Sodium azide, an inhibitor of SODs, did not inhibit the Mn-reconstituted SOD from A. pernix even at concentrations up to 400 mM. This SOD from an aerobic hyperthermophilic archaeon, Aeropyrum pernix, was extremely thermostable and active with either Mn or Fe. With Mn as a metal cofactor, it was more thermostable, and less sensitive to sodium azide and sodium fluoride than with Fe.

Aerobiosis↗

PubMed1999

Production of lysophosphatidic acids by lysophospholipase D in human follicular fluids of In vitro fertilization patients.

Lysophosphatidic acids (LPAs) are known to be normal constituents of mammalian serum, and they mimic some biological effects of the serum. We previously reported that lysophospholipase D (LPLD) was involved in the accumulation of LPAs in incubated rat plasma and serum. In this study we detected, by gas-liquid chromatography, various molecular species of LPA in follicular fluids collected from women programmed for in vitro fertilization. When the follicular fluid was incubated at 37 degrees C for 48 h, persistent increases in the amounts of LPAs were observed concomitant with decreases in the amounts of the corresponding lysophosphatidylcholines (LPCs), although the concentrations of saturated LPCs increased in the first 6 h of incubation. These results suggest that human follicular fluid has LPLD activity, and this was confirmed by experiments with follicular fluids mixed with an exogenous radioactive LPC. The LPLD showed preference for unsaturated over saturated LPCs, similar to plasma LPLD, indicating that it originated from the circulation.

Adult↗

PubMed1999

Adenovirus type 5 E1A immortalizes primary rat cells expressing wild-type p53.

Adenovirus (Ad) E1A induces apoptosis in cells expressing wild-type p53, and stable transformation by Ad E1A requires the co-introduction of an anti-apoptotic gene such as Ad E1B 19K. Thus, cells immortalized by Ad E1A alone might have lost functional p53. In order to analyze the p53 in rat cells expressing Ad E1A, we established rat cell lines by transfecting primary rat embryo fibroblast (REF) and baby rat kidney (BRK) cells with cloned Ad5 E1A. By using a yeast functional assay, we analyzed p53 in six primary REF and three BRK cell lines immortalized by Ad5 E1A as well as five spontaneously immortalized rat cell lines (REF52, NRK, WFB, Rat-1 and 3Y1). The yeast functional assay revealed that all of the spontaneously and Ad5 ElA-immortalized rat cell lines except for 3Y1 expressed wild-type p53. All of the Ad5 E1A-immortalized rat cell lines contained p53 detectable by immunoprecipitation. Recombinant adenovirus expressing rat p53 cloned from a REF cell line immortalized by Ad5 E1A, as well as that expressing murine wild-type p53, induced apoptosis in p53-null cells in collaboration with E1A. Thus, it is suggested that the mutation of p53 appears to be not frequent in the spontaneous immortalization of primary rat cells, and that the functional loss of wild-type p53 is not a prerequisite of E1A-mediated immortalization.

Adenoviridae↗

PubMed1999

Induction of transformation and p53-dependent apoptosis by adenovirus type 5 E4orf6/7 cDNA.

Adenovirus (Ad) E4orf6/7, one of the early gene products of human Ads, forms a stable complex with the cellular transcription factor E2F to activate transcription from the Ad E2 promoter. E2F cDNAs have growth-promoting and apoptosis-inducing activities when overexpressed in cells. We cloned Ad5 E4orf6/7 cDNA in both simian virus 40- and human cytomegalovirus-based expression vectors to examine its transforming and apoptotic activities. The cloned E4orf6/7 collaborated with a retinoblastoma protein (RB)-nonbinding and therefore E2F-nonreleasing mutant of Ad5 E1A (dl922/947) to morphologically transform primary rat cells, suggesting that E2F is an important cellular protein functioning downstream of E1A for transformation. In a G418 colony formation assay, E4orf6/7 was shown to suppress growth of untransformed rat cells. Moreover, a recombinant Ad expressing Ad5 E4orf6/7 induced apoptosis in rat cells when coinfected with wild-type p53-expressing Ad. Mutational analysis of E4orf6/7 revealed that both of the domains required for growth inhibition and transformation by E4orf6/7 lay in the C-terminal region, which is essential for transactivation from the upstream sequence of an E2a promoter containing E2F-binding sites. However, the smallest mutant of E4orf6/7, encoding the C-terminal 59 amino acids, failed to complement the RB-nonbinding dl922/947 mutant despite showing growth inhibition and E2F transactivation activities. Thus, it is suggested that a subregion of E4orf6/7 which is required for growth inhibition and transformation in collaboration with dl922/947 overlaps the transactivation domain of E4orf6/7.

Adenovirus E1A Proteins↗

PubMed1999

Possible involvement of IL-12 expression by Epstein-Barr virus in Sjögren syndrome.

AIM: To determine the correlation between interleukin 12 (IL-12) expression and Epstein-Barr virus (EBV) in Sjögren syndrome. METHODS: Indirect immunohistochemical technique, enzyme linked immunosorbent assay (ELISA), and immunoblot analysis were used to investigate IL-12 expression by EBV activation, using 13 surgical specimens and four B cell lines. RESULTS: Marked expression of IL-12 was found in the epithelial cells and the infiltrating B cells of salivary gland tissues from patients with Sjögren syndrome (six of 10 cases), but not in those from normal individuals (none of three cases). A striking topographic correlation between IL-12 and EBV was found. In addition, levels of IL-12 production by B cell lines were clearly enhanced by EBV activation in vitro. CONCLUSIONS: IL-12 expression closely reflects the intracellular event of EBV activation in Sjögren syndrome, and may contribute to the T helper cell type 1 (Th1) cytokine overexpression seen in this disease.

Adult↗

PubMed1999

Purification and characterization of two major forms of naloxone reductase from rabbit liver cytosol, new members of aldo-keto reductase superfamily.

Rabbit liver cytosol produced approximately equal amounts of 6alpha-naloxol and 6beta-naloxol from naloxone in the presence of NADPH at pH 7.4, and contained at least four forms of naloxone reductase. The two major forms, NR1 and NR2, which catalyze the stereospecific reduction of naloxone to 6alpha-naloxol and 6beta-naloxol, respectively, were purified to apparent homogeneity by various chromatographic techniques. Both enzymes are monomeric proteins with similar molecular weights of 35000-36000, but NR1 is a basic protein with an isoelectric point (pI) of 9.3, while NR2 is an acidic protein (pI of 5.9). NR1 and NR2 gave the maximal activities at pH 8.0 and 6.1, respectively. NR1 exhibited considerable activity with NADH as well as with NADPH, whereas NR2 showed highly restricted specificity for NADPH. The Km and Vmax values of NR1 and NR2 for naloxone were 1.0 and 0.06 mM, and 76 and 162 munits/mg, respectively. In addition to naloxone, naltrexone and dihydromorphinone served as good substrates for NR2 but were poor substrates for NR1. Both enzymes reduced aliphatic and aromatic aldehydes, cyclic and aromatic ketones, and quinones at higher rates. The two enzymes catalyzed the dehydrogenation of 17beta-hydroxysteroids with low Km values, and NR2 showed an additional 3alpha-hydroxysteroid dehydrogenase activity. Amino acid sequence data of NRI (99% of whole protein) and NR2 (66%) showed that both enzymes belong to the aldo-keto reductase (AKR) superfamily and can be classified into the AKR1C subfamily. These findings therefore indicate that they are new members of the AKR superfamily and may be involved physiologically in the steroid metabolism as well as in the detoxification of xenobiotic carbonyl compounds.

Alcohol Oxidoreductases↗

PubMed1999

[Production of morphinone as a metabolite of morphine and its physiological role].

Morphine is a potent analgesic and is widely used in the clinical management of severe acute and chronic pain; however, its clinical usefulness is limited due to the development of both tolerance and dependence after repeated morphine administration. The morphine metabolism has been studied in order to elucidate its pharmacological actions as well as its adverse effects. Thus far several metabolites have been identified and their analgesic potency and toxicity have been also investigated. In the toxicological viewpoint, the production of reactive metabolites that can bind cellular glutathione and protein has been postulated. We found morphine 6-dehydrogenase, which catalyzes the dehydrogenation of 6-hydroxy group of morphine to produce morphinone, in the guinea pig liver. It was also found that morphinone antagonizes the morphine analgesia and binds with glutathione and protein. We here demonstrate the presence of a metabolic pathway of morphine to morphinone and subsequently to morphinone-glutathione adduct, and compare the property including primary structure among the guinea pig, rabbit, mouse and hamster liver morphine 6-dehydrogenases. We also describe the toxicological significance of morphinone.

Alcohol Oxidoreductases↗

PubMed1999

Relationship between brain atrophy estimated by a longitudinal computed tomography study and blood pressure control in patients with essential hypertension.

To evaluate the relationship between blood pressure control and the progression of brain atrophy in the elderly, patients with essential hypertension and brain atrophy were longitudinally evaluated using computerized tomography (CT). The study evaluated 48 patients with essential hypertension aged 46-78 years, and 30 sex- and age-matched normotensive control subjects. The extent of brain atrophy as determined by caudate head index (CHI), the inverse cella media index (iCMI), and Evans' ratio (ER) was estimated twice at an interval of 5-9 years (mean, 6.9 years). The mean annual increases in CHI (deltaCHI), iCMI (delta iCMI), and ER (deltaER) were evaluated. Mean blood volume in the common carotid artery (BF) and the decrease in BF per year (deltaBF) were also determined. The deltaCHI, delta iCMI, and deltaER increased with age in the hypertensive subjects as well as the control group across all age groups evaluated. The deltaCHI, delta iCMI, and deltaER were significantly greater in the patients with essential hypertension in their 50 s as compared with the controls. In patients with essential hypertension aged 65 years or older, the deltaCHI, delta iCMI, and deltaER were significantly lower in the group in whom the blood pressure was controlled within the range of borderline hypertension than the groups in which it was controlled in the range of normal or mild hypertension. In the younger patients under the age of 65 with essential hypertension, blood pressure control did not affect the deltaCHI, delta iCMI, and deltaER. The deltaCHI, delta iCMI, and deltaER were significantly correlated with deltaBF in both groups. These findings indicate that control of systolic blood pressure within the range of borderline hypertension may delay the progression of brain atrophy in elderly patients with essential hypertension.

Aged↗

PubMed1999

Asymptomatic cerebrovascular lesions detected by magnetic resonance imaging in patients with systemic lupus erythematosus lacking a history of neuropsychiatric events.

OBJECTIVE: To clarify the extent of asymptomatic cerebrovascular involvement in systemic lupus erythematosus (SLE). PATIENTS AND METHODS: Cerebral magnetic resonance imaging (MRI) findings and ultrasonography findings of 100 patients with SLE lacking present or past clinical neurologic deficits were compared with 66 age-matched volunteers to determine the combined intima-media thickness (IMT) of the common carotid artery, and tests for anti-cardiolipin antibodies (aCL). RESULTS: Thirty-eight patients, but only 2 controls, showed imaging abnormalities. Among 23 SLE patients with cerebrovascular lesions by MRI who underwent single-photon emission computed tomography (SPECT), 14 showed hypoperfusion of the lesion. The IMT value and prevalence of aCL did not differ between the 55 SLE patients tested and controls. SLE disease activity index (SLEDAI) as assessed by a quantitative clinical index was significantly greater in patients with brain lesions than in those without. CONCLUSION: The prevalence of asymptomatic brain lesions in SLE patients is highs and shows a relationship to disease activity.

Adolescent↗

PubMed1999

[An elderly case of triple-vessel coronary artery disease with alternating bundle branch blocks in serial electrocardiograms].

An 82-year-old woman was admitted with severe chest pain and orthopnea on January 17, 1997. Physical examination revealed bilateral leg edema and cyanosis at the periphery of the extremities. The serum CK level was 488 IU/l on admission and increased to a maximum value of 4,866 IU/l 8 hours after admission. An echocardiogram demonstrated diffuse severe hypokinesis in the left ventricle. Serial electrocardiograms showed transient right bundle branch block, left bundle branch block, and normal sinus rhythm. The patient was diagnosed as having congestive heart failure. Artificial ventilation was performed, and furosemide, isosorbide dinitrate and dopamine were administered. A right ventricular endomyocardial biopsy performed on the 13th hospital day demonstrated moderate hypertrophy and disparity of cardiac myocytes and fibrosis around the myocytes, and few inflammatory cells in the specimens. This biopsy finding was not compatible with acute myocarditis but with the chronic stage of myocarditis. The patient was discharged on the 45th hospital day, but returned because of a recurrence of congestive heart failure. After an improvement of the heart failure, a coronary angiography was performed on the 20th hospital day. The coronary angiography revealed significant stenosis in three vessels. This elderly patient had congestive heart failure and triple-vessel coronary artery disease with transient alternating bundle branch blocks on serial electrocardiograms. Alternating bundle branch blocks and diffuse left ventricular dysfunction was considered to be induced by the aging process, postmyocarditic change of myocytes, and triple-vessel coronary artery disease in this case.

Aged↗

PubMed1999

[A case of systemic lupus erythematosus discovered from left heart failure due to lupus induced mitral regurgitation].

A 50-year-old female was admitted to a local hospital because of dyspnea, and diagnosed as having left heart failure secondary to mitral regurgitation. After the improvement of congestive heart failure, polyarthralgia, fever, and positive anti-nuclear antibody were pointed out. She was referred to our hospital for the further evaluation. Serological test showed anti-double stranded DNA antibodies, anti-SS-A antibodies, anti-beta 2-GPI antibodies and biological false positive for syphilis. The diagnosis of SLE has been made from the clinical signs and the serology. Therefore mitral valvular lesion of this patient was considered to be one of the symptoms of SLE. We reported a rare case in which left heart failure was a initial clinical manifestation of SLE.

Female↗

PubMed1998

Biochemical and genetic characterization of an FK506-sensitive peptidyl prolyl cis-trans isomerase from a thermophilic archaeon, Methanococcus thermolithotrophicus.

A peptidyl prolyl cis-trans isomerase (PPIase) was purified from a thermophilic methanogen, Methanococcus thermolithotrophicus. The PPIase activity was inhibited by FK506 but not by cyclosporine. The molecular mass of the purified enzyme was estimated to be 16 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 42 kDa by gel filtration. The enzyme was thermostable, with the half-lives of its activity at 90 and 100 degrees C being 90 and 30 min, respectively. The catalytic efficiencies (k(cat)/Km) measured at 15 degrees C for the peptidyl substrates, N-succinyl-Ala-Leu-Pro-Phe-p-nitroanilide and N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide, were 0.35 and 0.20 microM(-1) s(-1), respectively, in chymotrypsin-coupled assays. The purified enzyme was sensitive to FK506 and therefore was called MTFK (M. thermolithotrophicus FK506-binding protein). The MTFK gene (462 bp) was cloned from an M. thermolithotrophicus genomic library. The comparison of the amino acid sequence of MTFK with those of other FK506-binding PPIases revealed that MTFK has a 13-amino-acid insertion in the N-terminal region that is unique to thermophilic archaea. The relationship between the thermostable nature of MTFK and its structure is discussed.

Amino Acid Sequence↗

PubMed1998

Sjögren's syndrome: a model for dental care in the 21st century.

The diagnosis and treatment of Sjögren's syndrome, which poses many severe complications, should be of critical interest to dentists, who are often the first practitioners to detect symptoms. Dentistry is an integral part of health care delivery for patients with this condition. Management of Sjögren's syndrome can be seen as a model for the expanded scope of dental care in the future.

Autoimmune Diseases↗

PubMed1998

[Hemodynamics of asymptomatic brain infarction determined by Doppler sonography].

We compared the usefulness of two methods for diagnosing asymptomatic brain infarction: an ultrasonic quantitative flow measurement system (QFM) and a transcranial Doppler arteriography (TCDA). A total of 137 patients (73 men and 64 women) who underwent magnetic resonance imaging of the brain, QFM, and TCDA were enrolled. Their ages ranged from 41 to 83 years (mean age, 63 years). The patients were divided into 3 groups: 45 without cerebrovascular disease (Group N); 40 with asymptomatic brain infarction (Group AS); and 52 with lacunar infarction (Group LI). The mean blood flow in the common carotid artery (CCA-BF) was measured by QFM. The mean blood velocity and Fourier pulsatility index in the middle cerebral artery (MCA-BV, MCA-PI) were measured by TCDA. In Group N, 28 patients were examined twice at a mean interval of 2 years; 19 remained without asymptomatic brain infarction (Group N1), and asymptomatic brain infarction developed in the remaining 9 (Group N2). The 3 groups differed significantly in MCA-PI (N < AS < LI), but not in CCA-BF or in MCA-BV. The MCA-PI in Group N2 was higher than that in Group N1. These results indicate that the Fourier pulsatility index determined by TCDA may be useful for detecting the onset of asymptomatic brain infarction.

Adult↗