The EspB protein of enterohaemorrhagic Escherichia coli interacts directly with alpha-catenin

Enterohaemorrhagic Escherichia coli (EHEC) belongs to a family of pathogens that cause attaching and effacing (A/E) lesion on target cells. The EspB protein of EHEC is translocated both to the host cell cytoplasm and to the membrane, and is essential for the signalling events leading to A/E lesion. To determine the actual role of EspB in this process, we tried to identify the EspB binding partner of the host cell protein, using a yeast two-hybrid assay, and obtained a cytoskeletal-associated protein, α -catenin. The α -catenin bound directly to the N-terminal region of EspB, both in solid (overlay assay) and solution (pull-down assay) phases, and it was recruited to the EHEC adherence site, dependent on EspB. Expression of the N-terminal region of EspB, as well as the whole EspB in host cells, inhibited F-actin accumulation on the adherence site. We conclude that EspB recruits α -catenin at the EHEC adherence site by direct interaction, and that the recruitment of α -catenin is essential for EHEC-induced A/E lesion formation.     

Diterpenoid from Salvia greggii

The structure of a diterpenoid, designated salvigresin, that was isolated from the aerial parts of Salvia greggii, has been confirmed by spectroscopic investigation and X-ray analysis.     

Involvement of sulfoquinovosyl diacylglycerol in the structural integrity and heat-tolerance of photosystem II

To examine the role of sulfoquinovosyl diacylglycerol (SQDG) in thylakoid membranes, we compared the structural and functional properties of photosystem II (PSII) between a mutant of Chlamydomonas reinhardtii defective in SQDG ( hf-2) and the wild type. The PSII core complex of hf-2, as compared with that of the wild type, showed structural fragility when solubilized with a detergent, dodecyl beta- d-maltoside, suggesting that the physical properties of the PSII complex were altered by the loss of SQDG. On the other hand, exposure of the cells to 41 degrees C for 120 min in the dark decreased the PSII activity to 70% and 50% of the initial levels in the wild type and hf-2, respectively, which implies that the PSII activity, in the absence of SQDG, becomes less stable under heat-stress conditions. PSII inactivated to 60% of the initial level by dark incubation at 41 degrees C was reactivated by following illumination even at 41 degrees C to more than 90% in the wild type, but only to 70% in hf-2. These results suggest that PSII inactivated by heat recovers through some mechanism dependent on light, and that SQDG participates in functioning of the mechanism. The conformational disorder of PSII caused by the defect in SQDG might be correlated with the increased susceptibility of its activity to heat-stress.     

Adrenomedullin expression in coronary circulation after stent implantation as a prognostic factor for restenosis

The authors determined the changes in adrenomedullin (AM) expression in the coronary circulation of patients with ischemic heart disease who underwent coronary stent implantation. Blood samples were drawn from the coronary sinus before, immediately after and four hours after coronary stent implantation, and plasma levels of AM were measured. AM levels in the coronary sinus blood were significantly increased four hours after stent implantation. On the other hand, in the femoral arterial blood, no significant changes in AM levels were observed. A significant positive correlation was found between AM level in the coronary sinus blood four hours after stent implantation and late loss index six months after the procedure. These results suggest that inflammation after coronary stent implantation induces AM expression, which might play an important role in restenosis after stenting.     

Inhibitory effects of resveratrol derivatives from dipterocarpaceae plants on tyrosinase activity

Stilbene derivatives, which are resveratrol (3,4',5-trihydroxy-trans-stilbene) oligomers ranging from monomer to tetramer, isolated from Dipterocarpaceae plants were tested for their inhibitory effects against murine tyrosinase activity. The structure-activity relationships obtained in this study suggest that the double bond in the stilbene skeleton is critical for the inhibition, and also that molecular size is important for inhibitory potency.     

Gnetol as a potent tyrosinase inhibitor from genus Gnetum

Gnetol (2,3',5',6-tetrahydroxy-trans-stilbene), a naturally occurring compound particularly found in the genus Gnetum, had a strong inhibitory effect on murine tyrosinase activity. Gnetol (IC50, 4.5 microM) was stronger than kojic acid (IC50, 139 microM) as a standard inhibitor for murine tyrosinase activity. Moreover, gnetol significantly suppressed, melanin biosynthesis in murine B16 melanoma cells.     

Clarithromycin inhibits overproduction of muc5ac core protein in murine model of diffuse panbronchiolitis

Long-term treatment of macrolide antibiotics is considered an effective treatment for diffuse panbronchiolitis (DPB). Although hypersecretion is a common feature of this disease, and it is known that macrolides inhibit mucin production, the mechanism of the effect on mucin production is unclear. The aim of our study was to determine the production of muc5ac core protein, a major core protein of mucin in airway secretion, and the effect of clarithromycin treatment on such production in a mouse model mimicking DPB. Alcian blue-periodic acid-Schiff-positive cells were detected in the lungs of Pseudomonas aeruginosa-infected mice. Western blots of these mice showed muc5ac glycoprotein at day 1 and increased progressively from day 4 to day 14 after inoculation of bacteria. Clarithromycin (10 mg. kg-1. day-1 for 7 days) significantly reduced the muc5ac expression at both the mRNA and protein levels. To investigate the role of molecules upstream in muc5ac regulation, we examined the role of mitogen-activated protein kinase. Extracellular signal-regulated kinase 1/2 phosphorylation increased in the infected lung and decreased after treatment. Our results suggest that overproduction of muc5ac plays an important role in the pathogenesis of DPB and that clinical improvement following macrolide therapy seems to involve, at least in part, its inhibition of mucin overproduction, through modulation of intracellular signal transduction.     

Evaluation of the use of an integration-type laser-Doppler flowmeter with a temperature-loading instrument for measuring skin blood flow in elderly subjects during cooling load: comparison with younger subjects

An integration-type laser-Doppler flowmeter, equipped with a temperature-load instrument, for measuring skin blood flow (ILD-T), and analytical parameters developed in a previous study were used to compare changes in the skin blood flow in the forehead and cheek in elderly subjects (in their 60s and 70s) with those in younger subjects (in their teens to 50s). Age-related differences in skin blood flow in the forehead and cheek in response to cooling were evaluated in 90 healthy women in their teens to 70s (mean age: 17.2 +/- 0.33 years for teenagers; 24.3 +/- 0.76 years for those aged 20-29 years; 34.8 +/- 1.12 years for those aged 30-39 years; 43.3 +/- 0.78 years for those aged 40-49 years; 53.8 +/- 1.13 years for those aged 50-59 years; 63.5 +/- 0.55 years for those aged 60-69 years; 72.2 +/- 0.70 years for those aged 70-79 years). The measurement was performed continuously for 5 min: for 1 min at a sensor temperature of 30 degrees C, for 2 min after the setting of the sensor temperature had been changed to 10 degrees C, and for 2 min after the temperature setting had been cancelled. The parameters analyzed were (1) skin temperature in a resting state before measurement ( T(rest)), (2) mean skin blood flow in 1 min at a sensor temperature of 30 degrees C ( F(30 degrees C)), (3) minimum skin blood flow at a sensor temperature of 10 degrees C ( F(min)), (4) slope of the blood flow plot during the period from the beginning of cooling at 10 degrees C to F(min) ( S(fall)), (5) time required for the sensor temperature to reach 10 degrees C (Delta t(s)), (6) maximum skin blood flow during the period from the end of cooling to the end of measurement ( F(max)), (7) slope of the blood flow plot during the period from F(min) to F(max) ( S(rise)), (8) rate of decrease of the skin blood flow during cooling: FDR = ( F(min)/ F(30 degrees C))x100, (9) recovery rate of the skin blood flow after the end of cooling: FRR = ( F(max)/ F(30 degrees C))x100. When correlations among the above nine parameters were evaluated by combining all age groups, significant correlations ( P < 0.01) were observed between F(30 degrees C) and F(min), F(30 degrees C) and F(max), F(30 degrees C) and S(fall), F(min) and F(max), and F(max) and S(rise) in the forehead. In the cheek, significant correlations ( P < 0.01) were observed in all these combinations except between F(max) and S(rise). When these analytical parameters were compared among the age groups, F(30 degrees C), T(rest), F(max), and S(rise) decreased significantly ( P < 0.02 for F(30 degrees C) and T(rest), P < 0.01 for F(max) and S(rise)) and S(fall) increased significantly ( P < 0.03) in the forehead with aging. However, no significant change with aging was observed in FDR, Delta t(s), F(min), and FRR. In the cheek, FDR increased significantly ( P < 0.03), and S(rise) decreased significantly ( P < 0.01) with aging. However, no significant change with aging was observed in F(30 degrees C), T(rest), F(max), S(fall), Delta t(s), F(min), and FRR. Thus, the decrease in the skin blood flow during cooling showed no marked quantitative change with age, but, with aging, the rate of this decrease was clearly reduced in the forehead. In the cheek, on the other hand, the skin blood flow decreased markedly with aging, but no clear change was observed in the rate of this decrease. By using ILD-T and examining various parameters obtained, the skin hemodynamics in the forehead and cheek during cooling from 30 degrees C to 10 degrees C could be analyzed, and differences in the hemodynamics between the forehead and cheek and between elderly and younger individuals were clarified. This instrument is expected to be clinically useful.     

Reactivation of lytic replication from B cells latently infected with Epstein-Barr virus occurs with high S-phase cyclin-dependent kinase activity while inhibiting cellular DNA replication

Productive infection and replication of herpesviruses usually occurs in growth-arrested cells, but there has been no direct evidence in the case of Epstein-Barr virus (EBV), since an efficient lytic replication system without external stimuli does not exist for the virus. Expression of the EBV lytic-switch transactivator BZLF1 protein in EBV-negative epithelial tumor cell lines, however, is known to arrest the cell cycle in G(0)/G(1) by induction of the tumor suppressor protein p53 and the cyclin-dependent kinase (CDK) inhibitors p21(WAF-1/CIP-1) and p27(KIP-1), followed by the accumulation of a hypophosphorylated form of the Rb protein. In order to determine the effect of the onset of lytic viral replication on cellular events in latently EBV-infected B LCLs, a tightly controlled induction system of the EBV lytic-replication program by inducible BZLF1 protein expression was established in B95-8 cells. The induction of lytic replication completely arrested cell cycle progression and cellular DNA replication. Surprisingly, the levels of p53, p21(WAF-1/CIP-1), and p27(KIP-1) were constant before and after induction of the lytic program, indicating that the cell cycle arrest induced by the lytic program is not mediated through p53 and the CDK inhibitors. Furthermore, although cellular DNA replication was blocked, elevation of cyclin E/A expression and accumulation of hyperphosphorylated forms of Rb protein were observed, a post-G(1)/S phase characteristic of cells. Thus, while the EBV lytic program promoted specific cell cycle-associated activities involved in the progression from G(1) to S phase, it inhibited cellular DNA synthesis. Such cellular conditions appear to especially favor viral lytic replication.     

SM-31900, a novel NMDA receptor glycine-binding site antagonist,reduces infarct volume induced by permanent middle cerebral artery occlusion in spontaneously hypertensive rats

The purpose of this study was to investigate the effect of (3S)-7-chloro-3-[2-((1R)-1-carboxyethoxy)-4-aminomethylphenyl]aminocarbonylmethyl-1,3,4,5-tetrahydrobenz[c,d]indole-2-carboxylic acid hydrochloride (SM-31900), an antagonist with high selectivity and affinity for the NMDA receptor glycine-binding site, on the cerebral infarct volume in a permanent middle cerebral artery occlusion (MCAo) model, which was constructed by electrocoagulation of a unilateral middle cerebral artery distal to the olfactory tract using spontaneously hypertensive rats (SHRs). To investigate the dose-response characteristics and the therapeutic time window of SM-31900 in this MCAo model, we conducted three experiments, in which the administration of SM-31900 was started 5min (experiment I), 30min (experiment II), or 60min (experiment III) after MCAo, respectively. In all the studies, SM-31900 was administered by intravenous bolus injection followed by continuous intravenous infusion to obtain a steady-state level of this compound in blood immediately after its administration. The treatment with SM-31900 was continued until 24h after MCAo, at which time the cerebral infarct volume was measured. In experiment I, SM-31900 significantly reduced the infarct volume by 37% at a dosage of 0.38mg/kg bolus followed by 1.5mg/kg/h continuous infusion (0.38mg/kg+1.5mg/kg/h). In experiment II, the neuroprotective effect of SM-31900 was also significant, with a 25% reduction in infarct volume at a dosage of 0.38mg/kg+1.5mg/kg/h, and a 40% reduction at 1.5mg/kg+6.0mg/kg/h. Furthermore, even in experiment III, SM-31900 exerted a significant neuroprotective effect, with a 20% reduction at 1.5mg/kg+6.0mg/kg/h. These studies revealed that SM-31900 can exert a neuroprotective effect when it is administered up to at least 60min after the onset of ischemia in the MCAo model, an animal model of stroke, indicating that SM-31900 is a good candidate for treating acute brain ischemia.