Effects of Rice Wine Lees on Cognitive Function in Community-Dwelling Physically Active Older Adults: A Pilot Randomized Controlled Trial.

BACKGROUND: Rice wine lees (RWL), a Japanese traditional fermented product, is a rich source of one-carbon metabolism-related nutrients, which may have beneficial effects on cognitive function. OBJECTIVES: We aimed to examine the effect of the RWL on cognitive function in community-dwelling physically active older adults. DESIGN: Double-blind, randomized, placebo-controlled study (clinical trial number: UMIN 000027158). SETTING: Community-based intervention including assessments conducted at the University of Hyogo and a public liberal arts school in Himeji City, Japan. PARTICIPANTS: A total of 35 community-dwelling older adults (68-80 years) who performed mild exercise before and during the trial were assigned to either the RWL (n=17) or the placebo group (n=18). INTERVENTION: Daily consumption of 50 g RWL powder, which contained one-carbon metabolism-related nutrients, or the placebo powder (made from soy protein and dextrin) for 12 weeks. Both supplements included equivalent amounts of energy and protein. MEASUREMENTS: Montreal Cognitive Assessment, computerized cognitive function test, and measurements of serum predictive biomarkers (transthyretin, apolipoprotein A1, and complement C3) were conducted at baseline and follow-up. RESULTS: Visual selective attention and serum transthyretin significantly improved in the RWL group, whereas there was no significant change in the placebo group. No significant group difference was observed in the remaining cognitive performance tests. CONCLUSIONS: RWL supplements seem to have a few effects on cognitive function in community-dwelling physically active older adults. However, the impact was limited; therefore, further studies with sufficient sample size are warranted to elucidate this issue.

Developmental ability of trophoblast stem cells in uniparental mouse embryos.

Neither parthenogenetic (PG) nor androgenetic (AG) mouse embryos survive after day 9.5 of pregnancy, owing to the inadequate growth of extraembryonic tissues, including the placenta. At day 9.5 of pregnancy, the placental structures are poorly developed in PG embryos, while trophoblast giant cells are abundant at the implantation site in AG embryos. These findings suggest that both parental genomes are required for placental development. To gain further insight into the trophoblast lineage in PG and AG embryos, we attempted to derive trophoblast stem (TS)-like cell lines from uniparental embryos. Furthermore, we sought to assess their ability to differentiate into cells of the trophoblast lineage by using gene expression analysis. Three cell lines that expressed marker genes for undifferentiated TS cells (Cdx2 and Errbeta) were derived from AG embryos. Under differentiation conditions, these cells expressed the trophoblast giant cell-specific genes, but did not express the spongiotrophoblast-specific genes. In contrast, none of the four cell lines from PG embryos expressed marker genes for undifferentiated TS cells, but they expressed Oct3/4, a marker gene for embryonic stem cells. Immunohistochemical analysis indicated that PG blastocysts expressed Oct3/4 and Cdx2 specifically in inner cell mass and the trophectoderm respectively. These results suggest that PG embryos do not possess TS cells, because of the lack of the developmental ability of trophoblast cells.

Understanding pluripotency--how embryonic stem cells keep their options open.

Embryonic stem (ES) cells have the capacity to proliferate indefinitely in culture while maintaining the ability to differentiate to form any of the cells of the body. This unique combination of functions suggests that these cells could provide a potentially unlimited source of differentiated cells for the treatment of disease and aging. Understanding the molecular processes that underpin these functions in ES cells will allow us to harness their potential and develop strategies that control their differentiation. Combination of controlled differentiation with ground-breaking technologies for the reversal of somatic cells to an ES cell-like state promise the generation of patient-derived pluripotent cell lines for the treatment of disease in the future.

Public health measures implemented during the SARS outbreak in Singapore, 2003.

The SARS outbreak hit Singapore between March and May 2003. Public health control measures were applied along three fronts; prevention and control within healthcare settings, community and at the borders. Nosocomial spread composed majority of SARS cases in Singapore. To prevent infection within healthcare facilities, cases were centralized in a SARS-designated hospital, a no-visitors rule was applied and movement of patients and healthcare staff were restricted. For triaging purposes, fever clinics were established. A dedicated ambulance service was used to transport possible cases to the SARS-designated hospital. Hospitals were surveyed for fever clusters. The challenge was to identify cases with atypical presentation. Effective and safe discharge criteria were established from the lessons learnt. To prevent community spread, contacts of cases were stringently traced, quarantined in their homes and monitored daily. For prompt identification of a case and to reduce the time between onset of symptoms and isolation, the Infectious Diseases Act was amended. A large wholesale market closure resulted in massive quarantine thereby limiting the spread of infection. A mass education campaign was implemented in order to educate and raise awareness of the public. At all air, sea and land points-of-entry, exit and entry screening took place that resulted in zero importation and exportation of SARS cases after implementation of screening. Coordinated effort of the cross sectional inter-ministerial collaboration and strong coordination by the Task Force and commitment from different professionals made it possible to conquer the disease.

[Placement of the inferior vena cava filter proximal to the renal vein in a patient with left-sided inferior vena cava complicated by pulmonary embolism: a case report].

A 55-year-old man was admitted to our hospital complaining of dyspnea and chest pain. Transthoracic echocardiography showed dilation of the right ventricle. Chest computed tomography with contrast medium showed multiple emboli in the pulmonary arteries. Venography of the lower extremities showed multiple thrombi in the right popliteal vein and the presence of left-sided vena cava. This unusual case of left-sided vena cava was complicated by deep vein thrombosis due to hemostasis. A Greenfield filter was placed in the vena cava proximal to the right renal vein in a right internal jugular vein approach.

Biological characterization and chemokine receptor usage of HIV type 1 isolates prevalent in Brazil.

The human immunodeficiency virus type 1 (HIV-1), the etiological agent of the acquired immunodeficiency syndrome (AIDS), shows a variety of biological properties, which may constitute an obstacle to development of effective vaccines or antiretroviral therapy. To characterize Brazilian strains of HIV-1, we studied 24 viruses isolated from blood samples of HIV-1-positive patients from different regions of the country. To examine the cell tropism and the virus ability to form syncytia, primary macrophages and the CD4+ T cell line MT-2 were infected with these viruses. We found that 22 isolates replicated well in macrophages (macrophage-tropic isolates), 2 infected only MT-2 cells (T cell line tropic variants), while 6 of them grew in both cells. We found 8 syncytium-inducing (SI) and 16 non-SI (NSI) isolates. Continuous cultures of 18 isolates were established in the CCR5+/CXCR4+ cell line PM-1, and SI/NSI features of these viruses were confirmed by cell fusion assay with uninfected CD4+ T cell lines (PM-1, MT-2, H9, and SUP-T1). The coreceptor usage of 18 isolates was investigated by infecting U87 cells transfected with CD4 and chemokine receptors, and we found that 11 isolates infected only CCR5+ cells, 3 only CXCR4+ cells, whereas 4 used both coreceptors. We also observed that X4 isolates were more sensitive to neutralization by dextran sulfate than R5 or R5X4 viruses. Our findings show that the Brazilian isolates are phenotypically similar to those prevalent in other regions, which could mean that therapeutic strategies based on HIV-1 phenotypic properties would be efficient in Brazil, as in other countries.

One-step subcellular fractionation of rat liver tissue using a Nycodenz density gradient prepared by freezing-thawing and two-dimensional sodium dodecyl sulfate electrophoresis profiles of the main fraction of organelles.

In the present study, we describe a new procedure using freezing-thawing to density gradient solution of Nycodenz for one-step separation of organelles from the rat liver and subsequent proteome analysis of subcellular fractions. To prepare two-dimensional electrophoresis (2-D PAGE) profiles of tissue organelles, we performed one-step subcellular fractionation of rat liver homogenate using a density gradient of Nycodenz solution, which resulted in the separation of the cytosolic fraction from the postnuclear supernatant. The density gradient of Nycodenz was prepared from a 20% solution in a centrifuge tube by freezing-thawing overnight at -20 degrees C and at room temperature for a few hours without the initial centrifugation procedure. The shape of the gradient density curve was dependent on Nycodenz concentration and tube size. After fractionation, the protein profiles were examined using one-dimensional sodium dodecyl sulfate (SDS)-PAGE. The organelles were confirmed using Western blotting. Our results indicate that our procedure provides a simple method for the separation of organelle fractions from the rat liver tissue.

Evaluation of the Japanese school health surveillance system for influenza.

In order to evaluate the Japanese nationwide school absenteeism surveillance system for pediatric influenza in comparison with the national sentinel surveillance for influenza, we used surveillance guidelines (Centers for Disease Control and Prevention, 1998) to determine the efficacy of the school health surveillance system (SHSS). Data regarding school absenteeism (age 4-15 years old) was compared with data regarding influenza-like illness (ILI) per sentinel sites during the second to the 11th weeks of 1998 and 1999. Despite the system's high simplicity and acceptability, telecommunication costs were estimated at US$ 490,000 (1998). Representativeness of schoolchildren was very accurate, but ILI for pre-school children (4-6 years) remained uncountable. Sensitivity, specificity, and positive predictive value of the SHSS compared to sentinel surveillance were calculated as 80%, 100%, and 100%, respectively (P=0.004). Although the SHSS was found to provide accurate surveillance data during periods of high influenza activity, non-influenza virus infections (e.g., adenovirus, rotavirus, and Norwalk virus, etc.) may become mixed in the SHSS data. Evaluation using this system should be continued employing a new case definition excluding gastrointestinal symptoms.

Characterization of native and recombinant peptidyl prolyl cis-trans isomerases derived from Methanococcus thermolithotrophicus based on cDNA sequence.

It is important to establish whether a recombinant protein is an authentic copy of the predicted cDNA sequence. In this study, recombinant protein for native peptidyl prolyl cis-trans isomerase (N-PPIase) and double-labeled (13C- and 15N-) protein (DL-PPIase) appeared on the sodium dodecyl sulfate (SDS) electropherograms as two bands for N-PPIase and four bands for DL-PPIase. Since the N-terminal amino acid residues of all bands were the same, we characterized these bands using the peptide mapping method and amino acid composition analysis. Peptide mapping of the proteins seemed to be almost identical but they could not reflect the whole amino acid sequences of the protein. The bands on the polyvinylidene difluoride (PVDF) membrane, electroblotted after SDS-polyacrylamide gel electrophoresis (SDS-PAGE), were hydrolyzed and their amino acid composition was analyzed using a highly sensitive 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) amino acid analysis and compared with the cDNA sequences for proteins. The matching score (sigma(T%-E%)2) for similarity of proteins was calculated by summation of the square difference between the theoretical (T%) and the experimental (E%) amino acid composition of the recombinant protein. The amino acid composition of all bands of both proteins showed more than 93% of the theoretical values. The major molecular weights of both proteins were 16812 and 17694 by electrospray ionization (ESI)-mass spectrometry. However, the purified proteins also contained minor compounds with Mr of 3721 for N-PPIase and 5285 for DL-PPIase. These compounds were considered to be nonpeptidyl products that comigrated with the protein. Similarities of the amino acid composition of the four bands were more than 98%. Our results indicate that AQC amino acid analysis is the most suitable method for characterization of a recombinant protein.

Rapid determination of parvalbumin amino acid sequence from Rana catesbeiana (pI 4.78) by combination of ESI mass spectrometry, protein sequencing, and amino acid analysis.

The complete amino acid sequence of beta-type parvalbumin (PA) from bullfrog Rana catesbeiana (pI 4.78) was determined by tandem mass spectrometry in combination with amino acid analysis and peptide sequencing following Arg-C and V(8) protease digestion. The primary structure of the protein was compared with that of beta-type PA from R. esculenta (pI 4.50), with which it is highly homologous. Compared with R. esculenta beta-type PA4.50, R. catesbeiana beta-type parvalbumin (PA 4.78) differed in 15 out of 108 amino acid residues (14% displacement), PA4.78 had Cys at residue 64 and was acetylated at the amino terminus, but 25 residues of the carboxyl terminus were completely conserved. Several amino acid displacements were found between residues 51 and 80 (30% displacement), although the functionally important sequence of PA was completely conserved. The amino acids residues of putative calcium-binding sites were Asp-51, Asp-53, Ser-55, Phe-57, Glu-59, Glu-62, Asp-90, Asp-92, Asp-94, Lys-96, and Glu-101, which were conserved in all a and b-types of R. catesbeiana as well as other parvalbumins. In addition, Arg-75 and Glu-81, which are thought to form a salt bridge located in the interior of the molecule [Coffee, C.J. et al. (1976) Biochim. Biophys. Acta 453, 67-80], were also conserved in PA4.78.

Stage-specific isoforms of Ascaris suum complex. II: The fumarate reductase of the parasitic adult and the succinate dehydrogenase of free-living larvae share a common iron-sulfur subunit.

Complex II of adult Ascaris suum muscle exhibits high fumarate reductase (FRD) activity and plays a key role in anaerobic electron-transport during adaptation to their microaerobic habitat. In contrast, larval (L2) complex II shows a much lower FRD activity than the adult enzyme, and functions as succinate dehydrogenase (SDH) in aerobic respiration. We have reported the stage-specific isoforms of complex II in A. suum mitochondria, and showed that at least the flavoprotein subunit (Fp) and the small subunit of cytochrome b (cybS) of the larval complex II differ from those of adult. In the present study, complete cDNAs for the iron-sulfur subunit (Ip) of complex II, which with Fp forms the catalytic portion of complex II, have been cloned and sequenced from anaerobic adult A. suum, and the free-living nematode, Caenorhabditis elegans. The amino acid sequences of the Ip subunits of these two nematodes are similar, particularly around the three cysteine-rich regions that are thought to comprise the iron-sulfur clusters of the enzyme. The Ip from A. suum larvae was also characterized because Northern hybridization showed that the adult Ip is also expressed in L2. The Ip of larval complex II was recognized by the antibody against adult Ip, and was indistinguishable from the adult Ip by peptide mapping. The N-terminal 42 amino acid sequence of Ip in the larval complex II purified by DEAE-cellulofine column chromatography was identical to that of the mature form of the adult Ip. Furthermore, the amino acid composition of larval Ip determined by micro-analysis on a PVDF membrane is almost the same as that of adult Ip. These results, together with the fact, that homology probing by RT-PCR, using degenerated primers, failed to find a larval-specific Ip, suggest that the two different stage-specific forms of the A. suum complex II share a common Ip subunit, even though the adult enzyme functions as a FRD, while larval enzyme acts as an SDH.

[Morbidity and mortality of influenza in Japan].

Influenza is reportable in Japan since 1947. The current sentinel surveillance system for influenza started in 1987. Influenza morbidity is clearly correlated with increased influenza and pneumonia mortality and mortality from other major underlying diseases, which results in an increased overall mortality. The expected mortality was calculated from a seasonal ARIMA model for influenza and pneumonia. Excess mortality is defined as the number of deaths exceeding the upper confidence limit that was calculated from the model. During the 1998/1999 season, the estimated number of excess deaths from pneumonia and influenza was about 10,000 and from all causes about 40,000. More than 90% of these deaths occurred among elderly.

Identification of multidrug resistant protein 1 of mouse leukemia P388 cells on a PVDF membrane using 6-aminoquinolyl-carbamyl (AQC)-amino acid analysis and World Wide Web (WWW)-accessible tools.

Multidrug resistant protein 1 (MDR1) in a doxorubicin-resistant mouse leukemia cell line (P388/DOX) was identified using its amino acid composition combined with protein database searching (ExPASy and EMBL PROPSEARCH) via the World Wide Web. The proteins were separated by one-dimensional SDS-polyacrylamide gel electrophoresis, blotted onto a polyvinylidene fluoride membrane, and stained with Coomassie brilliant blue. A 160-kDa protein band was acid-hydrolyzed in the vapor phase (6 N HC1) and converted to 6-aminoquinolyl-carbamyl (AQC)-amino acids without extraction of the amino acids from the membrane. The amino acid composition of the protein was determined using the sensitive AQC-amino acid analysis method, improving our previously described method. The improved method involved using a Cosmosil 5C8-MS column instead of a Pegasil C8; replacement of the mobile phase A, constituent, 75 mM ammonium phosphate (pH 7.5), with 30 mM sodium phosphate buffer (pH 7.2); and slight modification of the separation program (9). All manipulations for protein hydrolysis and AQC derivatization were carried out in a hood using clean tools. This minimized contamination of amino acids at the low femtomolar level. A database search was carried out with bovine serum albumin as a calibration protein. MDR1 in P388/DOX was ranked first by both databases with high reliability (score 14 for ExPASy, distance 1.34 for EMBL).

[A clinical investigation of bacteremia for the past ten years at the Second Department of Internal Medicine, Jikei University Hospital].

We clinically investigated a total of 288 cases of bacteremia for the past ten years, from January 1986 to December 1995, at the Second Department of Internal Medicine in the jikei University Hospital. All of the subjects who had a positive reaction to blood culture or catheter tip culture were investigated for their basic disease, complications, and detected bacteria. Malignant tumors, chronic renal failure, diabetes mellitus, and hematologic disease were frequent by noted. The cases due to primary infection were mainly respiratory organ infection or urinary tract infection, which were 47.8% of the total. In 31.3% of the total, catheter tip cultures were positive. Except for catheter related infection, Gram-positive coccus were detected in 40.3%, which was most frequent. Methicillin resistant Staphylococcus aureus (MRSA) were 8.1% and Staphylococcus epidermidis were 11.2%. In catheter related infection, Gram-positive coccus were detected in 59.9%, which was most frequent amongst them, MRSA was 17.2%, S. epidermidis was 16.2%. The mortality of bacteremia was 12.5%, mainly from hematologic diseases, immunodeficiency due to long term steroid administration etc. Accordingly, the more the advance of chemotherapy, the better the prognosis of septicemia is. Appearance of catheter related infection was unexpected frequent. Increase of immunocompromised host is thought to be one of the main factors in the outbreak of bacteremia.

Separation of 18 6-aminoquinolyl-carbamyl-amino acids by ion-pair chromatography.

Eighteen 6-aminoquinolyl-carbamyl (AQC)-amino acids were separated by ion-pair chromatography, using tetrabutylammonium (TBA) as a counterion. Optimum separation was obtained on a C8 reverse-phase column using gradient elution with two mobile phases, A (5 mM TBA, 75 mM ammonium acetate, pH 7.5) and B (80% acetonitrile). The AQC-amino acids were detected by fluorescence with excitation at 250 nm and emission at 395 nm, and the analysis time was 65 min. The response factors of individual AQC-amino acids to AQC-phenylalanine ranged from 0.42 to 1.08 (except for tryptophan at 0.01), with an average of 0.8. Detection limits by fluorescence ranged from 11.8 fmol (threonine) to 51.7 fmol (methionine), except for tryptophan (1.8 pmol).

The structure of Silurus asotus (catfish) roe lectin (SAL): identification of a noncovalent trimer by mass spectrometry and analytical ultracentrifugation.

We identified a noncovalent trimer of Sirurus asotus roe lectin (SAL) at Mr 95,362 along with its monomer at M(r) 31,750 by electrospray ionization mass spectrometry when SAL was dissolved in 0.5% acetic acid, sprayed into the ion source with methanol as a sheath liquid, and desolvated at 75 degrees C in a heated capillary column. The molecular weight of SAL, determined by the sedimentation equilibrium method, was 95,200 and the sedimentation coefficient (S20,w) of SAL in water was 5.58. SAL existed as a noncovalent trimer in solution and showed the ability to agglutinate rabbit erythrocytes. SAL showed three peaks (sal 1, sal 2, and sal 3) by C8 reverse-phase HPLC, and these appeared to be a monomer, a dimer, and a trimer, respectively, by matrix-assisted laser desorption ionization-time of flight mass spectrometry, sal 1 and sal 2 were shown to have a structure interchangeable with that of sal 3 in water.

[Application of heparin-coated percutaneous cardiopulmonary support system to elective cardiac and aortic operations].

Non emergency use of heparin-coated percutaneous cardiopulmonary support (PCPS) in cardiovascular surgery was applied in 4 patients (3 patients with descending aortic aneurysm, and one with aortic valvular stenosis and impaired left ventricular contractility (LVEF = 0.18). In aortic surgery, femoral inflow and outflow cannulae were inserted after introduction of anesthesia, and low flow F F bypass was established with the patient in supine position. Subsequently lateral posture was taken for aortic grafting. Activated clotting time was kept around 200 sec during the whole operative period and F F bypass was continued after the grafting procedure, and the cannulae were removed when the patient posture was returned to supine. With this method cannulation and decannulation were easy, and almost prone position, which was sometimes necessary for dissection of the adhesion of lung to the posterior aspect of the aneurysmal wall, was easily performed. Because of the percutaneous insertion, there was no local hemorrhage in the groin. Heparin coating circuit permitted only low grade systemic anticoagulation during the operative period, which could lead to decreased amount of hemorrhage. Long F F bypass without full heparinization made no harmful effect on patients' coagulability nor their hemodynamic status. For the patient with extremely impaired cardiac contractility, "scheduled PCPS and IABP" was proved to be safe, reliable, and cost-effective: F F bypass was established before sternotomy, and intracardiac procedure with aortic crossclamping was performed under total cardiopulmonary bypass using PCPS. After declamping of the aorta, weaning from the bypass was not tried and total anticoagulation was reversed with protamine. Then the sternum was closed and patient was transferred to ICU with the PCPS and IABP on. After more than 10 hours, when the cardiac contractility recovered from the intraoperative ischemic insult to a considerable degree, weaning from PCPS was successfully accomplished.