Hanging drop monoculture for selection of optimal antioxidants during in vitro maturation of porcine oocytes.

We analysed the effect of three antioxidants that have different functional mechanisms on the in vitro maturation (IVM) of porcine oocytes. Single oocyte monoculture using the hanging drop (HD) system has some advantages such as improving analysis efficiency brought by the smaller number of samples than the number of oocytes cultured in one drop. Direct effects of ligands on single oocytes could also be detected without considering the effects of paracrine factors from other oocytes. After 22 h of pre-culture, denuded oocytes were cultured for 22 h with 0.01 and 0.1 µg/ml of L-carnitine (LC), lactoferrin (LF) or sulforaphane (SF) in the presence/non-presence of oxidant stress induced by H2O2 supplementation to evaluate the reducing effects against oxidative stress on nuclear maturation. As a result, compared with LC and SF, LF showed effective reduction in oxidative stress at a lower concentration (0.01 µg/ml), suggesting that LF is a more effective antioxidant in porcine oocyte IVM. Additionally, LF also increased maturation rate even in culture without H2O2. Our results clearly suggest that the HD monoculture system is useful for screening the substances that affect porcine oocyte culture.

Coesite and stishovite in a shocked lunar meteorite, Asuka-881757, and impact events in lunar surface.

Microcrystals of coesite and stishovite were discovered as inclusions in amorphous silica grains in shocked melt pockets of a lunar meteorite Asuka-881757 by micro-Raman spectrometry, scanning electron microscopy, electron back-scatter diffraction, and transmission electron microscopy. These high-pressure polymorphs of SiO(2) in amorphous silica indicate that the meteorite experienced an equilibrium shock-pressure of at least 8-30 GPa. Secondary quartz grains are also observed in separate amorphous silica grains in the meteorite. The estimated age reported by the (39)Ar/(40)Ar chronology indicates that the source basalt of this meteorite was impacted at 3,800 Ma ago, time of lunar cataclysm; i.e., the heavy bombardment in the lunar surface. Observation of coesite and stishovite formed in the lunar breccias suggests that high-pressure impact metamorphism and formation of high-pressure minerals are common phenomena in brecciated lunar surface altered by the heavy meteoritic bombardment.

Disrupted galectin-3 causes non-alcoholic fatty liver disease in male mice.

Galectin-3, a beta-galactoside-binding animal lectin, is a multifunctional protein. Previous studies have suggested that galectin-3 may play an important role in inflammatory responses. Non-alcoholic fatty liver disease (NAFLD) is increasingly recognized as a liver condition that may progress to end-stage liver disease and based on the known functions of galectin-3, it was hypothesized that galectin-3 might play a role in the development of NAFLD. Thus, this study investigated the role of galectin-3 in NAFLD by comparing galectin-3 knockout (gal3(-/-)) mice and wild-type (gal3(+/+)) mice. The livers of gal3(-/-) male mice at 6 months of age histologically displayed mild to severe fatty change. The liver weight per body weight ratio, serum alanine aminotransferase levels, liver triglyceride levels, and liver lipid peroxide in gal3(-/-) mice were significantly increased compared with those in gal3(+/+) mice. Furthermore, the hepatic protein levels of advanced glycation end-products (AGE), receptor for AGE (RAGE), and peroxisome proliferator-activated receptor gamma (PPARgamma) were increased in gal3(-/-) mice relative to gal3(+/+) mice. In conclusion, this study suggests that the absence of gal3 can cause clinico-pathological features in male mice similar to those of NAFLD.

[Cardiac conduction abnormalities after neoadjuvant chemotherapy with doxorubicin and docetaxel for primary breast carcinoma: a case report].

A 65-year-old woman with primary breast carcinoma (T 3 N 0 M 0) received neoadjuvant chemotherapy consisting of 4 cycles of 50 mg.m-2 doxorubicin (ADM) and 60 mg.m-2 docetaxel (TXT). The patient received 50 mg.m-2 ADM and 60 mg.m-2 TXT intravenously on day 1 of each cycle every three weeks. The patient underwent transthoracic echocardiography and electrocardiography (ECG) before surgery and these results were normal. ECG showed bigeminy, trigeminy after intubation and incomplete AV block (Mobitz type II) during and after surgery. The patient recovered from these arrhythmias on the first postoperative day. Our data indicate one possibility that the management of anesthetic condition might easily induce the cardiac conduction abnormalities of this patient using ADM as neoadjuvant chemotherapy.

A high-strain-rate superplastic ceramic.

High-strain-rate superplasticity describes the ability of a material to sustain large plastic deformation in tension at high strain rates of the order of 10-2 to 10-1 s-1 and is of great technological interest for the shape-forming of engineering materials. High-strain-rate superplasticity has been observed in aluminium-based and magnesium-based alloys. But for ceramic materials, superplastic deformation has been restricted to low strain rates of the order of 10-5 to 10-4 s-1 for most oxides and nitrides with the presence of intergranular cavities leading to premature failure. Here we show that a composite ceramic material consisting of tetragonal zirconium oxide, magnesium aluminate spinel and alpha-alumina phases exhibits superplasticity at strain rates up to 1 s-1. The composite also exhibits a large tensile elongation, exceeding 1,050 per cent for a strain rate of 0.4 s-1. The tensile flow behaviour and deformed microstructure of the material indicate that superplasticity is due to a combination of limited grain growth in the constitutive phases and the intervention of dislocation-induced plasticity in the zirconium oxide phase. We suggest that the present results hold promise for the application of shape-forming technologies to ceramic materials.

A novel screening for inhibitors of a pleiotropic drug resistant pump, Pdr5, in Saccharomyces cerevisiae.

Yeast is an excellent model system of eukaryotes for the study of molecular mechanisms of ATP-binding cassette transporters. Pdr5 protein is a yeast Saccharomyces cerevisiae ATP-binding cassette transporter conferring resistance to several unrelated drugs. Here, we described a novel drug screening system designated to detect compounds that inhibit the function of Pdr5. An indicator strain with increased drug sensitivity was constructed with an ergosterol-deficient background (delta syr1/erg3 null mutation). The sensitivity of the indicator strain (delta syr1/erg3 delta pdr5 delta snq2) to the Pdr5 substrates, cycloheximide and cerulenin, was increased 16-fold and 4-fold against wild type, respectively. The screening system is mainly based on the growth inhibition of the PDR5-overexpressed indicator strain with the combination of a sample and cycloheximide or cerulenin. The effect of an mdr inhibitor, FK506 on the screening system was clearly detected even at a low concentration (approximately 0.5 microg/ml). In addition, accumulation of rhodamine 6G in the cells was detected as a result of Pdr5 inhibition by FK506. These results indicated that the screening system is useful for a sensitive screening of Pdr5-specific inhibitors with low toxicity.

Direct imaging of Guinier-Preston zones by high-angle annular detector dark-field scanning transmission electron microscopy.

We report atomic resolution imaging of Cu-planar precipitates in aged Al-Cu alloys, known as Guinier-Preston (GP) zones, by high-angle annular detector dark-field scanning transmission electron microscopy. Single layered GP-I zones as small as 2 nm in length were resolved among densely populated GP-I zones, whereas double layered GP zones were clearly identified. The images of GP-II zones showed not only the commonly accepted structure, in which single Cu layers are separated by three Al layers, but also a variant, in which double Cu layers are separated by a single Al layer.

[A comparison of the effectiveness of transtracheal heating and humidification system in maintaining body temperature during general anesthesia with low flow gases].

We evaluated the effectiveness of transtracheal heating and humidification system in maintaining body temperature during general anesthesia with low flow gases in 12 gastric cancer patients. Patients were divided into two group; Control group A in which a hot-water circulating system was used and group B in which a transtracheal heating and humidification system by ANAMED HUMITUBE was used, during gastric cancer operation. Compared to the hot-water circulating system, the transtracheal heating and humidification system is more effective for maintaining body temperature and humidification after abdominal lavage by warm saline water. But there was no difference between the two groups about awakening from general anesthesia. We concluded that transtracheal heating and humidification system by ANAMED HUMITUBE is effective in maintaining body temperature under general anesthesia with low flow gases.

Simultaneous induction of galectin-3 phosphorylated on tyrosine residue, p21(WAF1/Cip1/Sdi1), and the proliferating cell nuclear antigen at a distinctive period of repair of hepatocytes injured by CCl4.

The period of repair of hepatocytes injured by CCl4 and signaling proteins intrinsic to this period were examined. A 30 kDa polypeptide detected by immunoblot analysis using anti-phosphotyrosine antibody in livers from rats 48 to 72 h after administration of a single dose of CCl4 was identified as galectin-3 induced in cytoplasm of periportal hepatocytes and phosphorylated on tyrosine residue(s). Simultaneously, these hepatocytes induced p21(WAF1/Cip1/Sdi1) in the nucleus and the proliferating cell nuclear antigen in both the nucleus and the cytoplasm, suggesting that hepatocytes during this distinctive period are quiescent and repair cellular damage. Trabecular architecture of hepatocytes with the proliferating cell nuclear antigen only in the nucleus was found at 96 h. These findings indicate that galectin-3 is a novel member of signaling proteins downstream of tyrosine kinase, and suggest that it plays roles in supporting repair or survival of the injured hepatocytes rather than their proliferation that is likely to be initiated later than 72 h.

NMR structure of Streptomyces killer toxin-like protein, SKLP: further evidence for the wide distribution of single-domain betagamma-crystallin superfamily proteins.

A protein isolated from the culture supernatant of the soil bacterium, Streptomyces sp. F-287, exhibits cytocidal effects for both budding and fission yeasts, and causes morphological changes of yeasts and filamentous fungi. This protein, which was the first killer toxin-like protein for yeasts identified in the Streptomyces microorganism, was named SKLP (Streptomyces killer toxin-like protein). Since the amino acid sequence of the protein, as determined by sequential Edman degradations, seemed to be unique, we determined the structure by NMR spectroscopy. Although the actual target of SKLP in yeasts has not been determined yet, the structure might give us a clue to characterize the targets. The solution structure of SKLP determined by NMR, however, turned out to be a single-domain crystallin-like protein, with two Greek key motifs and a short extra beta-strand at the N terminus. The final ensemble of 20 NMR structures overlaid onto their mean coordinate with rmsd values of 0.32(+/-0.06) A for the backbone atoms involved in the secondary structure elements. As a yeast killer toxin, WmKT, isolated from the yeast strain Williopsis mrakii also has a Greek key beta-barrel fold, we have made a detailed comparison of the structural features of SKLP with the other crystallin superfamily proteins. It is very interesting that SKLP has a unique electrostatic potential distribution on the molecular surface. Namely, one surface of the beta-barrel fold in SKLP has a large negatively charged region, with an isolated positive charge of the Arg62 side-chain at the center. The edge of this surface is surrounded by positively charged residues, including Arg31, Arg65 and Arg74. The salient features of the charge distribution on this surface and the cluster of Arg residues might be related to the target binding of SKLP.

Usefulness of the evidence-based medicine-supported cancer pain management guideline.

The Japanese guidelines for the clinical practice of cancer pain management supported by evidence-based medicine were established by the Japanese Society for Palliative Medicine in 1999 [as their Evidence-based Medicine-supported Cancer Pain Management Guideline ]. To evaluate usefulness of the Guideline for the management of cancer pain, the same questionnaires were addressed to nurses and physicians of enrolled institutions twice. The first survey was conducted before the distribution of the Guideline in July, 1999 and the second was done after the distribution in January, 2000. Usefulness of the Guideline was examined by comparing the results of two surveys. Cancer patients were divided into two groups depending on their stages (conservative or terminal). (1) Morbidity of pain in cancer patients at each stage having some analgesics did not change at each survey period. (2) At the first survey the rate of pain relief in each stage of cancer patients was essentially unchanged from a previous result obtained in 1998. (3) The rate of pain relief shown in the second survey tended to be higher than that shown in the first in both groups of patients. (4) The rate of pain relief with per os morphine was shown to be significantly higher in the second survey than in the first for each group of patients at conservative or terminal stage. (5) The rate of pain relief of patients staying in the ward where the guidance for dosing of morphine had been carried out was 37.5% at the first survey versus 47.9% at the second. (6) The answers from physicians to questions about treatment of cancer pain remained unchanged between the first and the second survey. The usefulness of the Guideline for cancer pain management is partly confirmed by these results. The significance of the Guideline will be totally discussed by comparing its effects on nurses, pharmacists and physicians.

[Severe laryngeal edema after tracheal extubation--report of a case].

An 80-year old female underwent subtotal esophagectomy with neck lymphnode dissection for esophageal cancer. The tracheal intubation was done repeatedly. Anesthesia was maintained with O2-N2O-isoflurane. No complications were observed during the operation. About thirty minutes after tracheal extubation, she developed airway obstruction with tracheal tug and stridor. Severe laryngeal edema was found by bronchofiberscopy. Percutaneous minitracheostomy was performed under ventilation support by laryngeal mask airway. Seven days later, bronchoscopy examination revealed that the laryngeal edema had disappeared. It should be kept in mind that severe laryngeal edema could develop immediately after extubation.

Purification and some properties of an inhibitor for a yeast pleiotropic drug resistant pump from Kitasatospora sp. E-420.

A strain producing an inhibitor for pleiotropic drug resistant 5 (Pdr5) was isolated using our original screening system in yeast. The strain was classified and named as Kitasatospora sp. E-420. The purified inhibitor (molecular weight = 1,193 by FAB-MS) inhibited a Pdr5-mediated efflux of cycloheximide and cerulenin. The intracellular accumulation of a fluorescent dye, rhodamine 123, by the inhibitor was also confirmed. Some physicochemical data suggested that the Pdr5-specific inhibitor was different from an immunosuppressant, FK506, reported as the only inhibitor of Pdr5 in vivo.

An HREM study of channel structures in mesoporous silica SBA-15 and platinum wires produced in the channels.

The hyperbolic geometry of the pores in SBA-15 are drawn into a three-dimensional structure by small lateral pores, distributed in a random way, as shown in the picture, which interconnect the main channels. This construction was established by forming a "negative" of the structure with platinum precipitated in the channel and dissolving the SBA-15 material.

Transcription modulation by a rat nuclear scaffold protein, P130, and a rat highly repetitive DNA component or various types of animal and plant matrix or scaffold attachment regions.

The XmnI fragment, a highly repetitive DNA component, and animal and plant matrix or scaffold attachment region (MAR/SAR) were examined for similarity in interaction with nuclear scaffold. As the XmnI fragment bound a 130 kDa scaffold protein (P130) in vitro, various types of MAR/SAR fragments could bind 130 and 123 kDa scaffold proteins. The native XmnI and MAR/SAR fragments clearly augmented SV40 promoter-mediated luciferase gene transcription following transient transfection of recombinant plasmids into various types of recipient cells. In contrast, the XmnI fragment methylated at the cytosine base of the unique HindIII site, and a synthetic variant DNA deficient in base unpairing characteristic of MAR/SAR, could neither bind P130 nor augment this transcription. These two types of genomic regions appeared to have similar properties of interaction with nuclear scaffold, by which the activity of appropriately positioned promoter can be modulated.

[Effect of gargling the throat with Panaxylocaine as a lubricant for the laryngeal mask airway].

We used Panaxylocaine as a suitable lubricant with gargling before insertion of laryngeal mask for twenty preoperative breast cancer patients. Patients were divided into two groups; Control group (N = 10) received water-based lidocaine gel just before insertion of laryngeal mask, and Panaxylocaine group (N = 10) received Panaxylocaine as a premedication for breast cancer operation. Compared to water-based lidocaine gel, Panaxylocaine was more effective for oral discomfort after insertion of laryngeal mask. But there was no difference between two groups about oral paresthesia. We conclude that Panaxylocaine is a good premedication before insertion of laryngeal mask preventing oral and pharyngeal complications.

Pressure effect on the conformational fluctuation of apomyoglobin in the native state.

We have investigated the effect of pressure on fluctuations of the native state of sperm whale apomyoglobin (apoMb) by H/D exchange, fluorescence, and limited proteolysis. The results from intrinsic fluorescence showed that a large fraction of apoMb molecules is in the native conformation in the pressure range from 0.1 to 150 MPa at 293 K and pH 6.0. The H/D exchange of protons of the individual backbone amino acids in this pressure range was monitored by NMR. The rate of H/D exchange was enhanced at high pressure, with the protection factors for some residues decreasing by factors of more than 100 compared to the values at 0.1 MPa. The amplitude of the decrease of the protection factor varied among the individual amino acids on the same secondary structure unit. This result suggests that H/D exchange in apoMb is explained best by the penetration model, in which solvent penetrates into the protein matrix via small motions. The result from limited proteolysis under high pressure showed that a pressure increase does not induce local unfolding of the secondary structure units of apoMb. Conformational fluctuations much smaller than local unfolding evidently provide pathways for water to diffuse into the protein interior, and are enhanced by an increase of pressure.

Interaction of albumin mRNA with proteins from rat liver with CCl(4)-induced injury.

Acute phase responses to intragastric administration of a single dose of CCl(4) were examined with albumin mRNA turnover as a marker. Based on the combination of the changes in stability of albumin mRNA and activity of transcription of its gene, the entire course of liver injury was classified into three stages, the first stage for aggravation of injury until 9 h, the second from 9 to 24 h, and the third for repair of injury or regeneration of liver after 48 h. Liver S100 fractions from normal and CCl(4)-treated rats contained, in total, 11 polypeptides cross-linked with part of albumin mRNA, although they did not appear to be specific to this mRNA. Their profiles were altered together with the changes in stability of albumin mRNA in different stages. These findings suggest that the polypeptides with distinct properties play roles in physiologically significant processes involved in utilization and turnover of albumin mRNA, apparently altering its stability.