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BACKGROUND Postoperative prolonged air leakage is a frequent complication after lung resection. We have developed a new sealant based on a hydrophobically modified Alaska pollock-derived gelatin (ApGltn) sealant. The purpose of this study was to evaluate the adhesive strength of the ApGltn sealant in comparison with a fibrin sealant using a new spray system in ex vivo and in vivo models. METHODS Pleural defects in ex vivo and in vivo porcine models were created, to which the ApGltn sealant or the fibrin sealant was applied. The pressure resistance was assessed with a stepwise increase in airway pressure to confirm air leakage from the sealing site. Tissue samples covered with each sealant were obtained for histologic assessment. RESULTS In the ex vivo experiment, the leak pressures of the ApGltn sealant were significantly greater than those of the fibrin sealant (102.94 ± 15.6 cm H2O and 28.37 ± 5.1 cm H2O, respectively) (P < .01). In the in vivo experiment, the leak pressures of the ApGltn sealant were also significantly greater than those of the fibrin sealant (68.82 ± 18.04 cm H2O and 43.33 ± 7.13 cm H2O, respectively) (P = .043). The histologic examination confirmed that the ApGltn sealant adhered tightly to both the pleura and the surface of the pleural defect. CONCLUSIONS The ApGltn sealant has sufficiently high adhesive quality in ex vivo and in vivo porcine lungs, which could be considered suitable and effective for use in the prevention of air leakage from the lungs.

Giardia lamblia is a pathogenic unicellular eukaryotic parasite that causes giardiasis. Its genome encodes the canonical histones H2A, H2B, H3, and H4, which share low amino acid sequence identity with their human orthologues. We determined the structure of the G. lamblia nucleosome core particle (NCP) at 3.6 Å resolution by cryo-electron microscopy. G. lamblia histones form a characteristic NCP, in which the visible 125 base-pair region of the DNA is wrapped in a left-handed supercoil. The acidic patch on the G. lamblia octamer is deeper, due to an insertion extending the H2B α1 helix and L1 loop, and thus cannot bind the LANA acidic patch binding peptide. The DNA and histone regions near the DNA entry-exit sites could not be assigned, suggesting that these regions are asymmetrically flexible in the G. lamblia NCP. Characterization by thermal unfolding in solution revealed that both the H2A–H2B and DNA association with the G. lamblia H3–H4 were weaker than those for human H3–H4. These results demonstrate the uniformity of the histone octamer as the organizing platform for eukaryotic chromatin, but also illustrate the unrecognized capability for large scale sequence variations that enable the adaptability of histone octamer surfaces and confer internal stability., Graphical Abstract Graphical AbstractG. lamblia has an equivalent nucleosome structure, despite low histone sequence identity to metazoan nucleosomes. The DNA entry-exit and acidic patch regions are unique characteristics of G. lamblia nucleosomes.

ObjectiveTo determine whether a minimal intervention based on the data envelopment analysis (DEA)-identified efficiency score effectively prevents hypertension.DesignRandomised controlled trial.SettingTakahata town (Yamagata, Japan).ParticipantsResidents aged 40–74 years belonged to the information provision group for specific health guidance. Participants with a blood pressure ≥140/90 mm Hg, those taking antihypertensive medication, or those with a history of cardiac diseases were excluded. Participants were consecutively assigned based on their health check-up visit at a single centre from September 2019 to November 2020 and were followed up at the check-up in the following year, until 3 December 2021.InterventionA targeted approach using minimal intervention. Target was identified using DEA and 50% of participants with higher risk were targeted. The intervention was notifying the results of their risk of hypertension according to the efficiency score obtained by the DEA.Primary outcome measuresA reduction in the proportion of participants who developed hypertension (≥140/90 mm Hg or taking antihypertensive medication).ResultsA total of 495 eligible participants were randomised, and follow-up data were available for 218 and 227 participants in the intervention and control groups, respectively. The risk difference for the primary outcome was 0.2% (95% CI −7.3 to 6.9) with 38/218 (17.4%) and 40/227 (17.6%) events in the intervention and control group, respectively (Pearson’s χ2test, p=0.880). The adjusted OR of the effect of the intervention was 0.95 (95% CI 0.56 to 1.61, p=0.843), and that of the efficiency score (10-rank increase) was 0.81 (95% CI 0.74 to 0.89, pConclusionsMinimal intervention to a high-risk population stratified by DEA was not effective in reducing the onset of hypertension in 1 year. The efficiency score could predict the risk of hypertension.Trial registration numberUMIN000037883

Nucleosome positioning and stability affect gene regulation in eukaryotic chromatin. Histone H2A.Z is an evolutionally conserved histone variant that forms mobile and unstable nucleosomes in vivo and in vitro. In the present study, we reconstituted nucleosomes containing human H2A.Z.1 mutants, in which the N‐terminal or C‐terminal half of H2A.Z.1 was replaced by the corresponding canonical H2A region. We found that the N‐terminal portion of H2A.Z.1 is involved in flexible nucleosome positioning, whereas the C‐terminal portion leads to weak H2A.Z.1‐H2B association in the nucleosome. These results indicate that the N‐terminal and C‐terminal portions are independently responsible for the H2A.Z.1 nucleosome characteristics., The N‐terminal and C‐terminal portions are independently responsible for the H2A.Z.1 nucleosome characteristics. The N‐terminal portion of H2A.Z.1 is involved in multiple positioning of the nucleosome. The C‐terminal portion of H2A.Z.1 leads to weak H2A.Z.1‐H2B association in the nucleosome.

Recent advances in genome‐wide technologies have enabled analyses using small cell numbers of even single cells. However, obtaining tissue epigenomes with cell‐type resolution from large organs and tissues still remains challenging, especially when the available material is limited. Here, we present a ChIL‐based approach for analyzing the diverse cellular dynamics at the tissue level using high‐depth epigenomic data. “ChIL for tissues” allows the analysis of a single tissue section and can reproducibly generate epigenomic profiles from several tissue types, based on the distribution of target epigenomic states, tissue morphology, and number of cells. The proposed method enabled the independent evaluation of changes in cell populations and gene activation in cells from regenerating skeletal muscle tissues, using a statistical model of RNA polymerase II distribution on gene loci. Thus, the integrative analyses performed using ChIL can elucidate in vivo cell‐type dynamics of tissues., “ChIL for tissues” allows obtaining epigenome information from a single‐thin tissue section with high sensitivity. A statistical model of RNApolII distribution is combined with ChIL to analyze cell‐type‐specific transcriptional states in a tissue.

During neocortical development, tight regulation of neurogenesis-to-astrogenesis switching of neural precursor cells (NPCs) is critical to generate a balanced number of each neural cell type for proper brain functions. Accumulating evidence indicates that a complex array of epigenetic modifications and the availability of extracellular factors control the timing of neuronal and astrocytic differentiation. However, our understanding of NPC fate regulation is still far from complete. Bone morphogenetic proteins (BMPs) are renowned as cytokines that induce astrogenesis of gliogenic late-gestational NPCs. They also promote neurogenesis of mid-gestational NPCs, although the underlying mechanisms remain elusive. By performing multiple genome-wide analyses, we demonstrate that Smads, transcription factors that act downstream from BMP signaling, target dramatically different genomic regions in neurogenic and gliogenic NPCs. We found that histone H3K27 trimethylation and DNA methylation around Smad-binding sites change rapidly as gestation proceeds, strongly associated with the alteration of accessibility of Smads to their target binding sites. Furthermore, we identified two lineage-specific Smad-interacting partners—Sox11 for neurogenic and Sox8 for astrocytic differentiation—that further ensure Smad-regulated fate-specific gene induction. Our findings illuminate an exquisite regulation of NPC property change mediated by the interplay between cell-extrinsic cues and -intrinsic epigenetic programs during cortical development.

The pregnane X receptor (PXR) is the key regulator of our defense mechanism against foreign substances such as drugs, dietary nutrients, or environmental pollutants. Because of increased health consciousness, the use of dietary supplements has gradually increased, and most of them can activate PXR. Therefore, an analysis of the interaction between drugs and nutrients is important because altered levels of drug-metabolizing enzymes or transporters can remarkably affect the efficiency of a co-administered drug. In the present study, we analyzed the effect of vitamin K-mediated PXR activation on drug metabolism-related gene expression in intestine-derived LS180 cells via gene expression studies and western blotting analyses. We demonstrated that menaquinone 4 (MK-4), along with other vitamin Ks, including vitamin K1, has the potential to induce MDR1 and CYP3A4 gene expression. We showed that PXR knockdown reversed MK-4-mediated stimulation of these genes, indicating the involvement of PXR in this effect. In addition, we showed that the expression of MDR1 and CYP3A4 genes increased synergistically after 24 h of rifampicin and MK-4 co-treatment. Our study thus elucidates the importance of drug–nutrient interaction mediated via PXR.

Recent advances in omics studies have enabled analysis at the single-cell level; however, methods for analyzing the whole cell of large organs and tissues remain challenging. Here, we developed a method named tsChIL to understand the diverse cellular dynamics at the tissue level using high-depth epigenomic data. tsChIL allowed the analysis of a single tissue section and could reproducibly acquire epigenomic profiles from several types of tissues, based on the distribution of target epigenomic states, tissue morphology, and number of cells. The proposed method enabled the independent evaluation of changes in cell populations and gene activation of cells in regenerating skeletal muscle tissues, using a statistical model of RNA polymerase II distribution on gene loci. Thus, the integrative analysis by tsChIL can elucidate in vivo cell-type dynamics of tissues.

Cardiopulmonary bypass (CPB) recovery is complicated by lung inflammation from bone marrow (BM)-derived polymorphonuclear leukocytes (PMNs) and monocytes (MO). Although Sivelestat reduces inflammatory mediators and Rolipram inhibits PMN and MO activation, any kinetic effects to improve CPB recovery in vivo are unknown. We hypothesized that intraoperative co-administration of these compounds would reduce CPB-induced lung inflammation through downregulation of PMN and MO recruitment. A 2-h CPB was surgically established in cynomolgus monkeys (n = 13), and BM leukocyte release and lung recruitment were monitored postoperatively by flow cytometry with 5'-bromo-2'-deoxyuridine (BrdU) and cytokine ELISA. Either Sivelestat, Sivelestat plus Rolipram, or saline (control) was administered intraoperatively and both peripheral and perfusion sampling courses revealed BrdU-labeled cells representative of activated leukocyte infiltration. Levels of cytokines CD11b and CD18 were leukocytic activation markers. Sivelestat plus Rolipram attenuated increases in CPB-associated circulating band cells, prolonged BM-transit time (PMN: 121.0 ± 3.7 to 96.2 ± 4.3 h [control], p = 0.012; MO: 84.4 ± 4.1 to 61.4 ± 3.0 h [control], p = 0.003), and reduced their alveolar appearance. CD11b-mediated PMN and MO changes during CPB and the post-surgical increases of Interleukin (IL)-6 and IL-8 in the bronchoalveolar lavage fluid were suppressed. Sivelestat alone increased PMN transit time to 115.8 ± 6.6 h, but monocytes were unaffected. Therefore, Rolipram has additive inhibitory effects with Sivelestat on the CPB-induced activation and release of BM-derived PMNs and MO and their recruitment to the lungs. Co-administration of these compounds could, therefore, hold value for preventing CPB-induced lung injury.

The N-methyl-D-aspartate (NMDA) receptor has been implicated in several neurodegenerative diseases, including stroke. Low-density lipoprotein receptor-related protein 1 (LRP1) plays pivotal roles in endocytosis and signaling in the cell. Immature LRP1 is processed by furin in the trans-Golgi network (TGN) and transported to the cell surface as its mature form. Activation of mature LRP1 exerts a protective effect against glutamate-induced degeneration of the rat retinal ganglion cells, as was shown in our previous study. However, the roles of LRP1 in the pathogenesis of excitotoxic neuronal injuries remain to be determined. The aim of this present study was to achieve further insight into the pathophysiologic roles of LRP1 after excitotoxic neuronal injuries. Our findings are the first to demonstrate that LRP1 was significantly cleaved by furin after cerebral ischemia in rats as well as after exposure of cultured cortical neurons to NMDA. It was noteworthy that the intracellular domain (ICD) of LRP1 was co-localized with TGN and furin. Furthermore, a furin inhibitor inhibited the cleavage of LRP1 and co-localization of LRP1-ICD with TGN or furin. Our findings suggest that furin-mediated cleavage of LRP1 and changes in the localization of LRP1-ICD were involved in the excitotoxic neuronal injury.

Geranylgeraniol (GGOH), a natural isoprenoid found in plants, has anti-inflammatory effects via inhibiting the activation of nuclear factor-kappa B (NF&kappa, B). However, its detailed mechanism has not yet been elucidated. Recent studies have revealed that isoprenoids can modulate signaling molecules in innate immune responses. We found that GGOH decreased the expression of lipopolysaccharide (LPS)-induced inflammatory genes in human macrophage-like THP-1 cells. Furthermore, we observed that the suppression of NF&kappa, B signaling proteins, in particular interleukin-1 receptor-associated kinase 1 (IRAK1) and tumor necrosis factor receptor-associated factor 6 (TRAF6), occurred in GGOH-treated cells prior to LPS stimulation, suggesting an immunomodulatory effect. These results indicate that GGOH may modulate and help prevent excessive NF&kappa, B activation that can lead to numerous diseases.

Sialic acid-binding lectin from Rana catesbeiana eggs (cSBL) is a multifunctional protein that has lectin and ribonuclease activity. In this study, the anti-tumor activities of cSBL were assessed using a panel of breast cancer cell lines. cSBL suppressed the cell growth of all cancer cell lines tested here at a concentration that is less toxic, or not toxic at all, to normal cells. The growth suppressive effect was attributed to the cancer-selective induction of apoptosis. We assessed the expressions of several key molecules associated with the breast cancer phenotype after cSBL treatment by western blotting. cSBL decreased the expression level of estrogen receptor (ER) &alpha, while it increased the phosphorylation level of p38 mitogen-activated protein kinase (MAPK). cSBL also suppressed the expression of the progesterone receptor (PgR) and human epidermal growth factor receptor type 2 (HER2). Furthermore, it was revealed that cSBL decreases the expression of the epidermal growth factor receptor (EGFR/HER1) in triple-negative breast cancer cells. These results indicate that cSBL induces apoptosis with decreasing ErbB family proteins and may have great potential for breast cancer chemotherapy, particularly in triple-negative phenotype cells.

Background Air leakage still remains a major problem in lung resection despite the introduction of surgical sealants. We have developed a novel sealant based on hydrophobically modified Alaska pollock–derived gelatin (ApGltn), which showed high adhesive quality in vitro. In this study, we evaluated the adhesive quality and conformability of our ApGltn sealant compared with a fibrin sealant. Methods The adhesive quality of the sealants was evaluated using excised porcine lungs with ventilation. Pleural defects were created, to which the ApGltn sealant or fibrin sealant was applied. Pressure resistance was assessed using a stepwise increase of airway pressure. Conformability was evaluated by measuring the area of the sealant for its maximum conformity on the gradually inflated lung surface. Results Leak and burst pressures of the ApGltn sealant were significantly higher than those of the fibrin sealant (47.1 ± 10.5 cm H2O and 52.3 ± 9.4 cm H2O versus 33.9 ± 6.0 cm H2O and 37.5 ± 5.9 cm H2O, respectively). Maximum expansion areas of the ApGltn sealant and fibrin sealant were 2652.4 ± 324.6 mm2 and 1276.6 ± 323.5 mm2, respectively. The ApGltn sealant also showed higher adhesive quality and conformability compared with the fibrin sealant. Histological examination confirmed that the ApGltn sealant showed tight adhesion to the pleural surface, while a gap was observed with the fibrin sealant. Conclusions The ApGltn sealant showed higher adhesive quality and conformability than the fibrin sealant in an excised porcine lung model. We expect that, in the future, the ApGltn sealant will be used for lung resections in clinical settings.

Background and Aim Infliximab is an established therapy for ulcerative colitis (UC). The aim of this study was to examine various serum cytokine levels and to identify possible markers predictive of therapeutic efficacy of infliximab for UC patients. Methods Twenty-one patients with moderately active UC were given intravenous infliximab (5 mg/kg) at 0, 2, and 6 weeks as induction therapy. The serum levels of 17 cytokines were determined using a Bio-Plex suspension array system before and 8 weeks after induction therapy. Partial Mayo score (PMS) and serum C-reactive protein levels were used for the determination of clinical activities at 0 and 8 weeks after the treatment. The overall therapeutic effect was determined at 26 weeks according to the PMS. Results The median value of the PMS decreased significantly 8 weeks after the treatment (from 6 to 1.5, P

Brain Aβ(1–42) accumulation is considered an upstream event in pathogenesis of Alzheimer's disease. However, accumulating evidence indicates that other neurochemical changes potentiate the toxicity of this constitutively generated peptide. Here we report that the interaction of Aβ(1–42) with extracellular Zn(2+) is essential for in vivo rapid uptake of Aβ(1–42) and Zn(2+) into dentate granule cells in the normal rat hippocampus. The uptake of both Aβ(1–42) and Zn(2+) was blocked by CaEDTA, an extracellular Zn(2+) chelator, and by Cd(2+), a metal that displaces Zn(2+) for Aβ(1–42) binding. In vivo perforant pathway LTP was unaffected by perfusion with 1000 nm Aβ(1–42) in ACSF without Zn(2+). However, LTP was attenuated under preperfusion with 5 nm Aβ(1–42) in ACSF containing 10 nm Zn(2+), recapitulating the concentration of extracellular Zn(2+), but not with 5 nm Aβ(1–40) in ACSF containing 10 nm Zn(2+). Aβ(1–40) and Zn(2+) were not taken up into dentate granule cells under these conditions, consistent with lower affinity of Aβ(1–40) for Zn(2+) than Aβ(1–42). Aβ(1–42)-induced attenuation of LTP was rescued by both CaEDTA and CdCl(2), and was observed even with 500 pm Aβ(1–42). Aβ(1–42) injected into the dentate granule cell layer of rats induced a rapid memory disturbance that was also rescued by coinjection of CdCl(2). The present study supports blocking the formation of Zn-Aβ(1–42) in the extracellular compartment as an effective preventive strategy for Alzheimer's disease. SIGNIFICANCE STATEMENT Short-term memory loss occurs in normal elderly and increases in the predementia stage of Alzheimer's disease (AD). Amyloid-β(1–42) (Aβ(1–42)), a possible causing peptide in AD, is bound to Zn(2+) in the extracellular compartment in the hippocampus induced short-term memory loss in the normal rat brain, suggesting that extracellular Zn(2+) is essential for Aβ(1–42)-induced short-term memory loss. The evidence is important to find an effective preventive strategy for AD, which is blocking the formation of Zn-Aβ(1–42) in the extracellular compartment.

The aim of this study was to clarify the relationship between the severity of aortic valve calcification (AVC) and stiffness of the proximal thoracic ascending aorta (TAA), and to examine their influence on left ventricular (LV) function and renal function.A total of 138 hypertensive patients including 32 with diabetes mellitus and 60 with dyslipidemia were divided into four groups based on the severity of AVC. We analyzed the elastic properties of the proximal TAA from the following strain-rate indices based on tissue Doppler imaging: maximum strain rate [SR(+)], minimum SR [SR(-)], and the time between the QRS peak and the peak SR(-) of the proximal TAA (SRT).SR(+) and SRT were significantly greater in patients with moderate AVC than in patients with mild AVC. SRT and SR(-) were well correlated with age, peak velocity across AV, TAA wall thickness (IMC), LV diastolic function, and renal function. SRT was independently related to IMC, dyslipidemia, and LV diastolic function.The severity of AVC was correlated with the elastic properties of the proximal TAA. The SR indices are useful for assessing the relation of TAA stiffness to LV function and renal function in patients with AVC.

Background We tested the hypothesis that heparin administration prior to the emergence of tissue factor (TF) would increase plasma TF pathway inhibitor (TFPI) and attenuate TF-mediated thrombin generation during simulated cardiopulmonary bypass (CPB). Methods Human blood was recirculated for 120 minutes using an oxygenator and roller pump. Four groups were examined: control group (heparin 3.75 U/mL, in donor blood, n = 7), rTF group (heparin + recombinant TF 1000 pg/mL, in donor blood, n = 7), TFPI boost group (heparin, in preheparinized donor blood, n = 8), and rTF + TFPI boost group (heparin + rTF, in preheparinized blood, n = 7). In the two TFPI boost groups, 50 U/kg of heparin was given to the donors intravenously five minutes before donation to boost plasma TFPI levels. Total plasma TFPI, thrombin–antithrombin complex, and prothrombin fragment F1+2 levels were measured before and during CPB. Results Preheparinization increased total plasma TFPI levels by a factor of 8.0. Administration of rTF significantly enhanced the generation of F1+2 (p = 0.0002). The heparin-induced TFPI elevation reduced both thrombin–antithrombin complex and F1+2 to control levels in rTF + TFPI boost group (p = 0.0158 for thrombin–antithrombin complex, p

Ulcerative colitis is the typical progression of chronic inflammatory bowel disease. Amino acids, particularly tryptophan, have been reported to exert a protective effect against colitis induced by dextran sodium sulfate (DSS), but the precise underlying mechanisms remain incompletely clarified. Tryptophan metabolites are recognized to function as endogenous ligands for aryl hydrocarbon receptor (Ahr), which is a critical regulator of inflammation and immunity. Thus, we conducted this study to investigate whether dietary tryptophan supplementation protects against DSS-induced colitis by acting through Ahr. Female wild-type (WT) and Ahr-deficient (knockout; KO) mice (10–12 weeks old) were divided into four groups and fed either a control or 0.5% tryptophan diet. The tryptophan diet ameliorated DSS-induced colitis symptoms and severity in WT mice but not in KO mice, and the diet reduced the mRNA expression of Il-6, Tnfα, Il-1β and the chemokines Ccl2, Cxcl1 and Cxcl2 in the WT groups. Furthermore, Il-22 and Stat3 mRNA expression in the colon was elevated in WT mice fed with the tryptophan diet, which mainly protected epithelial layer integrity, and Ahr also modulated immune homeostasis by regulating Foxp3 and Il-17 mRNA expression. These data suggest that tryptophan-containing diet might ameliorate DSS-induced acute colitis and regulate epithelial homeostasis through Ahr. Thus, tryptophan could serve as a promising preventive agent in the treatment of ulcerative colitis.

Serum endonucleases are essential for degrading the chromatin released from dead cells and preventing autoimmune diseases such as systemic lupus erythematosus. Serum DNase I is known as the major endonuclease, but recently, another endonuclease, DNase γ/DNase I-like 3, gained attention. However, the precise role of each endonuclease, especially that of DNase γ, remains unclear. In this study, we distinguished the activities of DNase γ from those of DNase I in mouse serum and concluded that both cooperated in degrading DNA during necrosis: DNase γ functions as the primary chromatolytic activity, causing internucleosomal DNA fragmentation, and DNase I as the secondary one, causing random DNA digestion for its complete degradation. These results were confirmed by two in vivo experimental mouse models, in which necrosis was induced, acetaminophen-induced hepatic injury and streptozotocin-induced β-cell necrosis models. We also determined that DNase γ functions as a backup endonuclease for caspase-activated DNase (CAD) in the secondary necrosis phase after γ-ray-induced apoptosis in vivo.

We report the design, synthesis, and physicochemical/biological evaluation of novel silanol derivative 6 (sila-T) as a silanol analog of multi-target nuclear receptor modulator T0901317 (5). Compound 6 showed intermediate hydrophobicity between the corresponding alcohol 13 and perfluoroalcohol 5. While 5 exhibited potent activities toward liver X receptor α and β, farnesoid X receptor, pregnane X receptor (PXR) and retinoic acid receptor-related orphan receptor (ROR)γ, silanol 6 exhibited activity only toward PXR and RORs. Incorporation of silanol instead of perfluoroalcohol is a promising option for developing novel target-selective, biologically active compounds.

Supplementation to an AIN93G-based diet of tocotrienol (T3) for 13 weeks administered to Fischer 344/slc rats showed a safety profile with no side effects. Dose-dependent T3 levels were detected in many tissues. Under the present experimental conditions, a continuous intake of the T3 concentrate would be safe in the rats as long as the T3 content was less than 0.20% of the dietary intake.

We investigated whether postweaning protein malnutrition (PM) affects serotonergic systems. Mice were fed a PM diet or normal protein (control) diet from weaning (21 d of age). Twenty days later, we tested for behavioral effects of the selective serotonin (5-HT)(1A) receptor agonist 8-hydroxy-N,N-dipropyl-2-aminoteralin (8-OH-DPAT) and the 5-HT releaser d-fenfluramine. The number of head weaving responses induced by 8-OH-DPAT or d-fenfluramine in the PM mice was significantly increased compared with the control diet group. The effects of 8-OH-DPAT and d-fenfluramine were blocked by pretreatment with the selective 5-HT(1A) receptor antagonist WAY-100635 (0.01 mg/kg). However, postpubertal (56 d of age) mice fed with the PM diet did not show an enhancement of the 8-OH-DPAT-induced head weaving response. These results indicate the occurence of a supersensitivity of postsynaptic 5-HT(1A) receptor in the postweaning PM group. Moreover, they highlight the postweaning stage as a vulnerable period to malnutrition-induced alterations in central serotonergic systems.

Members of the genus Nepenthes are carnivorous plants that use the pitfall method of insect capture as a supplementary nutritional source. We extracted metagenomic DNA from the microbial community found in the pitcher fluid of Nepenthes and constructed a plasmid-based metagenomic library. An activity-based screening method enabled the isolation of two lipase genes, lip1 and lip2. Both Lip1 and Lip2 belong to a novel family or subfamily of lipases and show lipase activities in acidic conditions, such as those found in pitcher fluid. This study was conducted under the assumption that the secreted Lip1 and Lip2 were capable of enzymatic activity in the acidic pitcher fluid.

Since the introduction of milled zirconia copings for ceramic crowns, a variety of techniques have been introduced to reduce the incidence of chipping or fracturing of the porcelain veneer. These include methods of improving the interface between the coping and the veneer, reducing thermal incompatibility between the two, and optimizing the coping design to minimize tensile loading on the porcelain. Another method of reducing porcelain chipping and fracture is to limit or eliminate the porcelain coverage of zirconia copings and frameworks. Even though patients often demand tooth colored or nonmetallic restorations, they tend to be less concerned with the optimal esthetics of their posterior teeth. This article describes 4 representative clinical situations where efforts were made to minimize or eliminate porcelain coverage on posterior zirconia crowns and a fixed dental prosthesis, while still achieving acceptable, but not optimal, esthetics.

Summary Paroxysmal nocturnal haemoglobinuria (PNH) is a haemolytic disease characterized by complement-sensitive red blood cells (RBC). PNH-affected RBC (PNH-RBC) should have a shortened mean lifespan (MLS); however, direct measurement is difficult. We have recently developed a sensitive flow cytometric assay to analyse PNH-affected reticulocytes that may closely correspond to the PNH clone-derived erythropoiesis. Naturally, the CD59-negative populations in reticulocytes were larger than those in whole RBC in PNH. We estimated the MLS of PNH-RBC in six PNH patients from the differences in the ratios of CD59-negative populations between reticulocytes and whole RBC. The MLS of PNH-RBC was calculated using the following formula: W/100 = R × M/[(100 − R) × 120 + R × M], where W, percentage CD59-negative whole RBC; R, percentage CD59-negative reticulocytes; M, MLS (days) of CD59-negative RBC. The MLS of PNH-RBC, estimated as 16–45 days in the PNH patients, showed a weak positive and a weak negative relation with RBCs and percentage reticulocytes, respectively, among the patients. The MLS, in individual patients, altered irrespective of RBC and percentage reticulocytes. The MLS calculated from our methods may be a parameter that evaluates the haemolytic conditions in PNH.

In vitro protein phosphorylation in soluble fractions (85,000×g supernatant) prepared from carrot hairy roots, which were induced by inoculation with Agrobacterium rhizogenes harboring the Ri plasmid, was analyzed by SDS-PAGE and autoradiography. The autoradiograms indicated that at least four proteins, with molecular masses of 65, 57.5, 52, and 38 kDa, were phosphorylated. The presence of Ca2+ enhanced the phosphorylation of the 65, 57.5, and 52 kDa proteins, but the presence of both Ca2+ and calmodulin did not enhance phosphorylation as much as Ca2+ alone. W-7 strongly inhibited the phosphorylation of the 65, 57.5, and 52 kDa proteins while W-5 mildly inhibited the phosphorylation of these proteins. The extent of phosphorylation of these proteins and the growth rate of the hairy roots became higher with increasing incubation temperature. W-7 had a considerable inhibitory effect on the growth of hairy roots, W-5 had a weak inhibitory effect on the growth of the roots, and a high concentration of TMB-8 inhibited growth. These results suggest that the 65, 57.5, and 52 kDa proteins are phosphorylated by CDPK and that the phosphorylation is connected with the increase in growth of carrot hairy roots.

A 30-year-old woman with corrected transposition of great arteries (c-TGA) was scheduled for elective cesarean section at 37 weeks of gestation. At previous cesarean section, she received general anesthesia for dyspnea and lower cardiac function by severe mitral regurgitation, with a pulmonary catheter inserted. In the current pregnancy, she had tricuspid regurgitation, but she had no signs of heart failure. Cardiac index (CI) and stroke volume variation (SVV) were monitored by the FloTrack, before induction of anesthesia. Because the CI was 3.6 l x min(-1) x m(-2), and the SVV was 18%, we decided to perform combined spinal epidural anesthesia. Epidural anesthesia was performed at L1-2, and spinal anesthesia was performed at L3-4. Hyperbaric 0.5% bupivacaine 2.0 ml with fentanyl 10 μg was given to the subarachnoid space. The total dose of phenylephrine administered was 150 μg, and the CI as well as the SVV were stable during surgery. Her postpartum couse was uneventful. Anesthetic management of c-TGA is discussed, and we should select anesthetic method carefully.

Infliximab is an established therapy for ulcerative colitis (UC). The aim of this study was to examine various serum cytokine levels and to identify possible markers predictive of therapeutic efficacy of infliximab for UC patients.Twenty-one patients with moderately active UC were given intravenous infliximab (5 mg/kg) at 0, 2, and 6 weeks as induction therapy. The serum levels of 17 cytokines were determined using a Bio-Plex suspension array system before and 8 weeks after induction therapy. Partial Mayo score (PMS) and serum C-reactive protein levels were used for the determination of clinical activities at 0 and 8 weeks after the treatment. The overall therapeutic effect was determined at 26 weeks according to the PMS.The median value of the PMS decreased significantly 8 weeks after the treatment (from 6 to 1.5, P 0.05). However, C-reactive protein levels did not change significantly. Levels of serum interleukin (IL)-8 (P 0.05) and macrophage inflammatory protein-1β (P 0.005) significantly decreased 8 weeks after the induction. Serum levels of the other 15 cytokines did not change significantly. At 26 weeks, 13 of 20 patients (65%) were responders while 7 patients were non-responders. Levels of serum IL-6 at 8 weeks were significantly lower in responders than in non-responders (P 0.05).Serum IL-8 and macrophage inflammatory protein-1β seem to be sensitive markers for UC patients treated with infliximab, while IL-6 at 8 weeks after induction therapy may be predictive of subsequent response to infliximab.

Paroxysmal nocturnal hemoglobinuria (PNH) is a hematopoietic stem cell disorder characterized by the deficiency of glycosyl phosphatidylinositol (GPI)-anchored proteins in the affected blood cell membranes. Analysis of blood cells by flow cytometry is useful to identify the affected blood cells with PNH-specific phenotypes. Because PNH-affected red blood cells (RBC) have shortened life-spans in the circulation, ratios of PNH-affected populations analyzed by flow cytometry in whole RBC are lower than those in PNH-affected erythropoiesis. We developed a reticulocyte-gated 2-color flow cytometry of RBC and revealed that the percentages of PNH-affected, CD59--populations in reticulocytes were higher than in whole RBC in patients with PNH. A serial analysis of a patient with PNH who underwent peripheral blood stem cell transplantation confirmed the usefulness of this method to evaluate PNH-affected RBC populations with high sensitivity; ie, the presence of CD59- reticulocytes in the circulation could be a sensitive marker for PNH-affected erythropoiesis.

We have found that protein malnutrition (PM) causes a significant impairment of memory-related behavior on the 15th and 20th day after the start of PM (5% casein) feeding in prepubertal mice but not in postpubertal mice, as measured by a passive-avoidance task. This impairment was almost completely reversed by merely switching to a standard protein (20% casein) diet on the 10th day after the start of PM. However, the reversal was not observed when the switching to a standard protein regimen was done on the 15th day of the PM diet. Interestingly, the impairment of memory-related behavior on the 20th day was improved by the chronic administration of physostigmine (0.1 mg/kg/day × last 10 days, i.p.), a cholinesterase inhibitor. To correlate brain cholinergic neuron function with the memory-related behavior impairment induced by PM, microphotometry was used to determine the histological distribution of the imunofluorescence intensity for choline acetyltransferase (ChAT), a functional marker of presynapse in cholinergic neurons. The change in the intensity of fluorescence indicated that ChAT protein was decreased in the hippocampus (CA1, CA3 and dentate gyrus) on the 20th day after PM feeding in comparison with controls. These results suggest the possibility that the memory-related behavior deficits observed in prepubertal mice with PM are caused by a dysfunction of the cholinergic neurons in the hippocampus.

Neutrophil is a major focus in efforts to ameliorate the systemic inflammatory response associated with cardiopulmonary bypass. Neutrophil elastase is a powerful proteolytic enzyme, and plays a pivotal role in the development of the inflammatory response. This study assesses the inhibitory effects of sivelestat, a highly specific neutrophil elastase inhibitor, on elastase levels, cytokine production, and the functional changes of neutrophils in a simulated extracorporeal circulation model.Simulated recirculation was established by recirculating heparinized (3.75 U/mL) human blood for 120 minutes in an oxygenator and a roller pump circuit with and without 100 micromol/L of sivelestat (n = 7 for each group). Neutrophil elastase and interleukin-8 were measured with an enzyme immunoassay. Neutrophil deformability was evaluated by simulated microcapillaries. The neutrophil F-actin and the expression of CD11b and L-selectin were measured by flow cytometry.Sivelestat reduced both neutrophil elastase levels (p = 0.0006) and interleukin-8 production (p0.0001) at 120 minutes of recirculation. Sivelestat also significantly preserved neutrophil deformability (p = 0.017) and reduced F-actin expression (p = 0.0037). The drug did not modulate the changes of CD11b or L-selectin.This study suggests that specific elastase inhibition with sivelestat could be a feasible therapeutic strategy for patients undergoing cardiopulmonary bypass to attenuate neutrophil-derived inflammatory response and organ injuries.

近年, 世界各地において適切な処置をされずに投棄・埋設された廃棄物による土壌汚染が問題となっている. こうした背景から本研究では, 原位置で汚染物質を長期にわたり封じ込めるバリアシステムの開発を目指している. 本報告では, 定水位透水試験を応用した実大規模のバリアシステムの止水性能評価法, ラジオアイソトープ (RI法) によるバリア継手部の漏水箇所推定手法の適用に関する検討結果を述べている. 検討の結果, いずれの手法とも実用可能であることが確認された. さらに, 実大規模で施工されたバリアの透水係数はk=1.0×10-7cm/sec以下で, 世界で最も厳しい基準である米国環境保護庁 (EPA) の要求性能を満たしており, 本バリアシステムの有効性も確認できた.

A 67-year-old woman with rheumatoid arthritis was scheduled for lumbar anterior fusion (L5-S1). The patient had undergone several major operations on the cervical to the lumbar spine. Cervical spine movement was severely restricted, the mouth opening was limited (inter-incisor distance 3 cm), and the jaw was small (thyro-mental distance 2 cm). During previous anesthesia tracheal intubation was always difficult. Fiberoptic nasotracheal intubation while the patient was sedated was planned. After bilateral superior laryngeal nerves had been blocked using 1% lidocaine, sedation was achieved using midazolam 1.4 mg and fentanyl 0.025 mg. Fiberscopy showed an edematous larynx, due probably to rheumatoid arthritis and to a long-term steroid therapy. It was possible to insert a fiberscope into the trachea, but it was difficult to pass a reinforced tube (6.0 mmID) and the procedure led to airway obstruction with a decreased arterial hemoglobin oxygen saturation. At the second attempt at fiberoptic intubation a rapidly swollen larynx was observed and awake intubation was abandoned. Fiberoptic intubation could be perfomed after induction of general anesthesia. This case indicates that, although awake fiberoptic intubation is regarded as the safest and the most reliable method, this may also be associated with severe airway obstruction.