The transcription factor ATF7 mediates lipopolysaccharide-induced epigenetic changes in macrophages involved in innate immunological memory.

Immunological memory is thought to be mediated exclusively by lymphocytes. However, enhanced innate immune responses caused by a previous infection increase protection against reinfection, which suggests the presence of innate immunological memory. Here we identified an important role for the stress-response transcription factor ATF7 in innate immunological memory. ATF7 suppressed a group of genes encoding factors involved in innate immunity in macrophages by recruiting the histone H3K9 dimethyltransferase G9a. Treatment with lipopolysaccharide, which mimics bacterial infection, induced phosphorylation of ATF7 via the kinase p38, which led to the release of ATF7 from chromatin and a decrease in repressive histone H3K9me2 marks. A partially disrupted chromatin structure and increased basal expression of target genes were maintained for long periods, which enhanced resistance to pathogens. ATF7 might therefore be important in controlling memory in cells of the innate immune system.

Dectin-1 Reactivity to Paramylon Derived from Euglena gracilis EOD-1.

Euglena gracilis is a microalga that has recently attracted attention because of its bioactivities. Paramylon (PM), a major ß-1,3-glucan, constitutes 70-80% of the cells of the E. gracilis EOD-1 strain. Dectin-1 is a pattern recognition receptor that recognizes ß-glucan. However, it is unclear whether PM binds to dectin-1. In this study, we investigated the reactivity of EOD1PM with dectin-1 by analyzing the binding of soluble murine and human dectin-1-Fc fusion protein (m dectin-1 Fc, h dectin-1 Fc) to EOD1PM using flow cytometry and enzyme-linked immunosorbent assay (ELISA). m Dectin-1 Fc bound to EOD1PM particles when m dectin-1-Fc is added. Furthermore, the binding specificity was examined in a competitive reaction following addition of a soluble antigen. It was found that the binding of m dectin-1-Fc to EOD1PM was not inhibited by the addition of dextran or ovalbumin but by the addition of solubilized EOD1PM or Candida cell wall- solubilized ß-glucan. In addition, the h dectin-1-Fc fusion protein was found to specifically bind to EOD1PM. These results suggest that dectin-1 recognizes and binds to the ß-glucan structure of EOD1PM. Dectin-1 is expressed in leukocytes as a ß-glucan receptor and is involved in the expression of various biological activities; therefore, the dectin-1 pathway may be involved in the biological activity of EOD1PM.

Factors Influencing Cardiac Sympathetic Nervous Function in Patients With Severe Aortic Stenosis: Assessment by 123I-Metaiodobenzylguanidine Myocardial Scintigraphy.

BACKGROUND: Numerous studies have shown that 123I-metaiodobenzylguanidine (MIBG) scintigraphy, an index of cardiac sympathetic nervous (CSN) activity, is useful for predicting prognosis in patients with heart failure. However, the factors influencing the CSN activity of patients with severe aortic stenosis (AS) remain unclear. METHODS: We enrolled 91 patients with severe AS who underwent 123I-MIBG scintigraphy, coronary computed tomography (CCT), and transthoracic echocardiography. When CCT angiography (CCTA) showed an obstructive epicardial artery, invasive coronary angiography was performed within 1 week of CCTA. RESULTS: There were 21 male and 70 female patients with a mean age of 84±5 years. Eighty-five (85) patients (93%) had hypertension and 13 patients (14%) had diabetes. Two (2) patients (2%) had previous myocardial infarction and eight (9%) had a previous coronary intervention. All patients had severe AS: aortic valve area was 0.63±0.18 cm2 and the mean pressure gradient was 56±19 mmHg. Regarding 123I-MIBG parameters, early heart-to-mediastinum (H/M) ratio was 3.1±0.5, delayed H/M ratio was 2.8±0.6, and the washout rate (WR) was 35%±13%. Multivariable linear regression analysis showed that coronary artery disease (ß=-0.30, p=0.002) was an independent predictor of delayed H/M ratio, and that aortic valve area (ß=-0.20, p=0.048) was an independent predictor of WR. CONCLUSIONS: Our findings suggest that coronary artery disease is an independent predictor of delayed H/M ratio, and aortic valve area is an independent predictor of WR in patients with severe AS.

Development of a Highly Sensitive ß-Glucan Detection System Using Scanning Single-Molecule Counting Method.

To overcome the limitations of the Limulus amebocyte lysate (LAL) assay method for the diagnosis of invasive fungal infection, we applied a reaction system combining recombinant ß-glucan binding proteins and a scanning single-molecule counting (SSMC) method. A novel (1→3)-ß-D-glucan recognition protein (S-BGRP) and a (1→6)-ß-glucanase mutant protein were prepared and tested for the binding of (1→6)-branched (1→3)-ß-D-glucan from fungi. S-BGRP and (1→6)-ß-glucanase mutant proteins reacted with ß-glucan from Candida and Aspergillus spp. Although LAL cross-reacted with plant-derived ß-glucans, the new detection system using the SSMC method showed low sensitivity to plant (1→3)-ß-D-glucan, which significantly improved the appearance of false positives, a recognized problem with the LAL method. Measurement of ß-glucan levels by the SSMC method using recombinant ß-glucan-binding proteins may be useful for the diagnosis of fungal infections. This study shows that this detection system could be a new alternative diagnostic method to the LAL method.

Possibility of Japanese Cedar Pollen Causing False Positives in the Deep Mycosis Test.

Because Japanese cedar pollen (JCP) contains beta-1,3-d-glucan (BG), there is concern that its lingering presence in the atmosphere, especially during its scattering period, may cause false positives in the factor-G-based Limulus amebocyte lysate (LAL) assay used to test for deep mycosis (i.e., G-test). Hence, we examined whether the LAL assay would react positively with substances contained in JCP by using the G-test to measure JCP particles and extracts. BG was purified from the JCP extract on a BG-specific affinity column, and the percentage extractability was measured using three different BG-specific quantitative methods. The G-test detected 0.4 pg BG in a single JCP particle and 10 fg from a single particle in the extract. The percentage extractability of JCP-derived BG was not significantly different among the three quantitative methods. As the JCP particles should technically have been removed during serum separation, they should be less likely to be a direct false-positive factor. However, given that the LAL-assay-positive substances in the JCP extract were not distinguishable by the three BG-specific quantitative methods, we conclude that they may cause the background to rise. Therefore, in Japan false positives arising from JCP contamination should be considered when testing patients for deep mycosis.

The Monocyte to High-Density Lipoprotein Cholesterol Ratio Is Associated with Left Ventricular Diastolic Function in Patients with No Significant Perfusion Abnormality.

The monocyte to high-density lipoprotein cholesterol (HDL-C) ratio has been considered to be a prognostic marker. Whether this ratio is associated with left ventricular (LV) diastolic function remains undetermined. We tested the hypothesis that the monocyte to HDL-C ratio is associated with LV diastolic parameters derived from gated myocardial perfusion single-photon emission computed tomography (SPECT) in patients with no significant perfusion abnormality.The study population included 196 patients with no significant perfusion abnormalities and preserved ejection fraction. The peak filling rate (PFR) and one-third mean filling rate (1/3 MFR) were obtained as LV diastolic parameters using gated SPECT. Monocyte counts and plasma HDL-C levels were also examined.Significant associations were observed between the monocyte to HDL-C ratio and PFR (r = -0.20; P = 0.005) and 1/3 MFR (r = -0.19; P = 0.009). Multivariate linear regression analysis was performed to determine factors associated with LV diastolic parameters. Age (ß = -0.27; P < 0.001), LV end-diastolic volume (ß = -0.19; P = 0.034), and monocyte to HDL-C ratio (ß = -0.15; P = 0.027) were determined to be significantly associated with PFR. Moreover, age (ß = -0.13; P = 0.007), LV mass index (ß = -0.18; P = 0.037), and the monocyte to HDL-C ratio (ß = -0.13; P = 0.045) were significantly associated with 1/3 MFR.These results demonstrated that the monocyte to HDL-C ratio is associated with LV diastolic function, as derived from gated SPECT in patients with no significant perfusion abnormality.

Latent 1,3-ß-D-glucan acts as an adjuvant for allergen-specific IgE production induced by Japanese cedar pollen exposure.

BACKGROUND: The pollen grains of several plant species contain 1,3-ß-D-glucan (BG). BG activates dendritic cells (DCs) and subsequently regulates the innate immune responses. Within Japan, the most common disease associated with type-I hypersensitivity is Japanese cedar pollinosis. However, the role of BG in Japanese cedar pollen (JCP) remains unclear. This study examined the localization and immunological effects of BG in JCP. METHODS: The localization of BG in JCP grain was determined by immunohistochemical staining using a soluble dectin-1 protein probe and a BG recognition protein (BGRP). The content of BG extracted from JCP was measured by a BGRP-based ELISA-like assay. The cytokine production by bone marrow-derived DCs (BMDCs) obtained from wild-type and BG receptor (dectin-1) knock-out mice was examined in vitro. The mice were intranasally administered JCP grains and the specific serum Ig levels were then quantified. RESULTS: BG was detected in the exine and cell wall of the generative cell and tube cell of the JCP grain. Moreover, BG in the exine stimulated production of TNF-α and IL-6 in the BMDCs via a dectin-1-dependent mechanism. Meanwhile, JCP-specific IgE and IgG were detected in the serum of wild-type mice that had been intranasally administered with JCP grains. These mice also exhibited significantly enhanced sneezing behavior. However, dectin-1 knock-out mice exhibited significantly lower JCP-specific IgE and IgG levels compared to wild-type mice. CONCLUSIONS: Latent BG in JCP can act as an adjuvant to induce JCP-specific antibody production via dectin-1.

Non-Invasive Central Venous Pressure Measurement Using Enclosed-Zone Central Venous Pressure (ezCVPTM).

BACKGROUND: Central venous pressure (CVP) is measured to assess intravascular fluid status. Although the clinical gold standard for evaluating CVP is invasive measurement using catheterization, the use of catheterization is limited in a clinical setting because of its invasiveness. We developed novel non-invasive technique, enclosed-zone (ezCVPTM) measurement for estimating CVP. The purpose of this study was to assess the feasibility of ezCVP and the relationship between ezCVP and CVP measured by a catheter.Methods and Results:We conducted 291 measurements in 97 patients. Linear regression analysis revealed that ezCVP was significantly correlated with CVP (r=0.65, P<0.0001). The Bland-Altman analysis showed that ezCVP had an underestimation bias of -2.5 mmHg with 95% limits of agreement of -14.1 mmHg and 9.6 mmHg for CVP (P<0.0001). The areas under the curves of receiver operating curve with ezCVP to detect the CVP ≥12 cmH2O (8.8 mmHg) and CVP >10 mmHg were 0.81 or 0.88, respectively. The sensitivity, specificity and positive likelihood ratio of ezCVP for the CVP ≥8.8 mmHg and CVP >10 mmHg were 0.59, 0.96 and 14.8 with a cut-off value of 11.9 and 0.79, 0.97 and 26.3 with a cut-off value of 12.7. CONCLUSIONS: These findings suggest that ezCVP measurement is feasible and useful for assessing CVP.

Coronary Vasculitis Induced in Mice by the Cell Wall Mannoprotein of Candida krusei.

Kawasaki disease (KD) is an inflammatory disease that was identified by Professor Tomisaku Kawasaki in 1961. Candida albicans-derived substances, such as the hot water extract of C. albicans (CADS) and Candida water-soluble fraction (CAWS), induced coronary vasculitis similar to KD in mice. An increasing proportion of deep-seated candidiasis cases are caused by non-albicans Candida and are often resistant to antifungal drugs. We herein investigated whether the hot water extract of C. krusei, inherently resistant to fluconazole, induces vasculitis in mice. Three strains of C. krusei, NBRC1395, NBRC1162, and NBRC10737, were cultured in natural (Y) and chemically defined (C) media and cell wall mannoprotein (MN) fractions were prepared by autoclaving cells (CKY1395MN, CKC1395MN, CKY1162MN, CKC1162MN, CKY10737MN, and CKC10737MN). All MN fractions reacted strongly with Concanavalin A (Con A) and dectin-2 and induced anaphylactoid shock in ICR mice. MNs induced severe coronary vasculitis in DBA/2 mice, resulting in cardiac hypertrophy. MNs also induced coronary vasculitis in C57Bl/6 mice. These results suggest that the MNs of non-albicans Candida, such as C. krusei, induce similar toxicity to those of C. albicans.

Frontal QRS-T angle and left ventricular diastolic function assessed by ECG-gated SPECT in the absence of significant perfusion abnormality.

The frontal QRS-T angle, defined as the angle between QRS and T-wave axes, has recently become an area of research interest. We tested the hypothesis that the frontal QRS-T angle is associated with left ventricular (LV) diastolic function in the absence of significant perfusion abnormality using ECG-gated SPECT. One hundred twenty eight patients with no significant perfusion abnormality and preserved LV ejection fraction were enrolled. The peak filling rate (PFR) and the one-third mean filling rate (1/3 MFR) were obtained as LV diastolic parameters on ECG-gated SPECT. There were 115 male and 13 female patients with a mean age of 70 ± 9 years. The PFR and 1/3 MFR were 2.1 ± 0.4/s and 1.2 ± 0.3/s, respectively. The frontal QRS-T angle was 33° ± 31°, ranging from 0° to 151°. There were significant associations of frontal QRS-T angle with PFR (r = - 0.29, p = 0.001) and 1/3 MFR (r = - 0.30, p < 0.001). Multivariate linear regression analysis showed that age (ß = - 0.25, p = 0.003), heart rate (ß = 0.26, p = 0.002), LV ejection fraction (ß = 0.43, p < 0.001) and frontal QRS-T angle (ß = - 0.16, p = 0.03) were significant factors associated with PFR. Also, heart rate (ß = - 0.32, p < 0.001), LV mass index (ß = - 0.19, p = 0.03), LV ejection fraction (ß = 0.30, p < 0.001) and frontal QRS-T angle (ß = - 0.26, p = 0.002) were significant factors associated with 1/3 MFR. Our data suggested that the frontal QRS-T angle was associated with LV diastolic function in the absence of significant perfusion abnormality.

Associations of left ventricular shape with left ventricular volumes and functions assessed by ECG-gated SPECT in patients without significant perfusion abnormality.

Left ventricular (LV) remodeling often results from conditions with an elevated LV hemodynamic load or after myocardial infarction. The present study was undertaken to investigate the associations of LV shape with LV volumes and functions in patients without significant perfusion abnormality. One hundred and sixty-seven patients without significant perfusion abnormality on ECG-gated SPECT were enrolled. LV ejection fraction (LVEF) was obtained for assessing LV systolic function. Peak filling rate (PFR) and one-third mean filling rate (1/3 MFR) were obtained for assessing LV diastolic function. LV shape index (LVSI) was defined as the ratio of the maximum three-dimensional short- and long-axis LV dimension, and varies from 0 (line) to 1 (sphere). There were 125 male and 42 female patients with a mean age of 70 ± 8 years. End-systolic LVSI was 0.49 ± 0.07 (0.34-0.65). End-systolic LVSI was associated with LV end-diastolic volume (r = 0.51, p < 0.001) and LV end-systolic volume (LVESV) (r = 0.64, p < 0.001), and was inversely associated with LVEF (r = - 0.69, p < 0.001), PFR (r = - 0.45, p < 0.001) and 1/3 MFR (r = - 0.26, p = 0.008). End-systolic LVSI was increased with increased LVESV, and was not any more with LVESV of > 40 ml. Multivariate liner regression analysis showed that age (ß = 0.16, p = 0.01), LVESV (ß = 0.20, p = 0.03) and LVEF (ß = - 0.53, p < 0.001) were significantly associated with end-systolic LVSI. Our data suggest that end-systolic LVSI, a measurement of LV shape, has close correlations with LV volumes and functions in patients without significant perfusion abnormality.

Effects of aging on coronary flow reserve in patients with no evidence of myocardial perfusion abnormality.

Coronary flow reserve (CFR) reflects the functional capacity of microcirculation to adapt to blood demand during increased cardiac work. We tested the hypothesis that aging had impacts on coronary flow velocities and CFR in patients with no evidence of myocardial perfusion abnormality on single photon emission computed tomography (SPECT). Seventy-six patients undergoing transthoracic Doppler echocardiography with no evidence of myocardial perfusion abnormality on SPECT were enrolled in this study. CFR was defined as the ratio of hyperemic to resting peak diastolic coronary flow velocity. Patients were divided into the three groups based on age: 17 patients aged less than 70 years (Group I), 38 patients aged 70-79 years (Group II), and 21 patients aged 80 years or more (Group III). Compared with Group I, CFR was significantly lower in Group II (p < 0.01) and Group III (p < 0.01). Multivariate linear regression analysis showed that female (ß = - 0.26, p = 0.03), cigarette smoking (ß = - 0.32, p = 0.004), hemoglobin level (ß = - 0.40, p = 0.001) and LV mass index (ß = 0.24, p = 0.03) were determinants for resting coronary flow velocity. On the other hand, age (ß = -0.30, p = 0.008), hemoglobin level (ß = -0.47, p < 0.001) and LV mass index (ß = 0.24, p = 0.04) were determinants for hyperemic coronary flow velocity. Age was only determinant for CFR (ß = -0.48, p < 0.001). Our data suggested that that aging had a decreased effect on hyperemic coronary flow velocity rather than resting coronary flow velocity, and was further associated with impaired CFR in patients with no evidence of myocardial perfusion abnormality.

Impact of Prosthesis-Patient Mismatch After Transcatheter Aortic Valve Replacement on Changes in Cardiac Sympathetic Nervous Function.

The impact of prosthesis-patient mismatch (PPM) after transcatheter aortic valve replacement (TAVR) on changes in cardiac sympathetic nervous (CSN) function remains unclear. Using 123I-metaiodobenzylguanidine (MIBG) myocardial scintigraphy, we investigated the impact of PPM after TAVR on CSN activity.We enrolled 44 of 117 patients with severe aortic stenosis who underwent TAVR for analysis in the present study. We conducted 123I-MIBG scintigraphy at baseline and at about 9 months after TAVR. Differences between baseline and post-TAVR 123I-MIBG parameters were compared between cases with and without PPM.There were 17 and 27 patients with and without PPM, respectively. Those without PPM exhibited significantly decreased left ventricular mass index (122 ± 36 g/m2 versus 108 ± 30 g/m2, P < 0.001) following TAVR, whereas those with PPM did not (117 ± 21 g/m2 versus 110 ± 17 g/m2, P = 0.09). Significant improvements in delayed heart-to-mediastinum (H/M) ratio (2.8 ± 0.4 versus 3.0 ± 0.4, P = 0.004) and washout rate (WR) (33% ± 10% versus 24% ± 12%, P < 0.001) were observed after TAVR in patients without PPM but not in those with PPM. Multivariable linear regression analysis revealed PPM to be a negative predictor of improvements in delayed H/M ratio and WR.Delayed H/M ratio and WR improve significantly after TAVR in the absence of PPM, whereas these improvements are not observed in patients with PPM. Hence, the presence of PPM is a negative predictor of improvements in delayed H/M ratio and WR in patients undergoing TAVR.

Effects of deep inspiration on QRS axis, T-wave axis and frontal QRS-T angle in the routine electrocardiogram.

The frontal QRS-T angle on the electrocardiogram has been described as a variable of ventricular repolarization. We evaluated how deep inspiration affected QRS axis, T-wave axis and frontal QRS-T angle. We also assessed the effects on left ventricular volume on the association using myocardial perfusion SPECT. Fifty patients undergoing ECGs both in resting state and in deep inspiration and subsequent SPECT were enrolled. Frontal QRS-T angle was defined as the absolute value of the difference between the frontal QRS axis and T-wave axis. Change in frontal QRS-T angle was calculated using (QRS-T angle in deep inspiration-QRS-T angle in resting state). In resting state, QRS axis and T-wave axis were 20.9° ± 30.0° and 40.9° ± 36.1°, respectively. Frontal QRS-T angle was 35.9° ± 36.1°. Deep inspiration caused rightward shifts of QRS axis (42.3° ± 29.5°, p < 0.001) and T-wave axis (49.5° ± 39.7°, p < 0.001). However, deep inspiration did not affect frontal QRS-T angle (33.9° ± 35.8°, p = 0.44). Frontal QRS-T angle in deep inspiration had good correlation (r = 0.87, p < 0.001) and agreement with that in resting state. Left ventricular (LV) end-diastolic volume had a significant association with change in frontal QRS-T angle (r = 0.29, p = 0.04). Our data suggest that frontal QRS-T angle in deep inspiration has a good correlation with that in resting state, and the agreement is acceptable. In patients with dilated LV, QRS-T angle in deep inspiration may be susceptible to the overestimation.

Myocardial perfusion defect assessed by single-photon emission computed tomography and frontal QRS-T angle in patients with prior anterior myocardial infarction.

The frontal QRS-T angle is one of the markers of ventricular repolarization. We sought to assess the effects of myocardial perfusion defect on QRS-T angle in patients with prior anterior myocardial infarction (MI). Seventy-one patients with prior anterior MI and 71 age- and sex-matched control subjects having no myocardial perfusion defect were selected. Frontal QRS-T angle was defined as the absolute value of the difference between the frontal plane QRS axis and T-wave axis. The extent of myocardial perfusion defect was determined using myocardial perfusion single-photon emission computed tomography (SPECT). The extent of myocardial perfusion defect of patients with prior anterior MI was 21.8 ± 13.7%. Frontal QRS-T angle was significantly larger in patients with prior anterior MI than control subjects (82° ± 49° vs 30° ± 26°, p < 0.001). Prevalence of abnormal frontal QRS-T angle defined as more than 90° was significantly higher in patients with prior anterior MI than control subjects (42% vs 4%, p < 0.001). Multivariate linear regression analysis showed that age (ß=0.18, p = 0.02) and myocardial perfusion defect (ß = 0.46, p = 0.02) were independent determinants of frontal QRS-T angle. Our results suggest that the extent of myocardial perfusion defect is an independent determinant of frontal QRS-T angle in patients with prior anterior MI.

N-Terminal (1→3)-ß-d-Glucan Recognition Proteins from Insects Recognize the Difference in Ultra-Structures of (1→3)-ß-d-Glucan.

Recognition of (1→3)-ß-d-glucans (BGs) by invertebrate ß-1,3-d-glucan recognition protein (BGRP) plays a significant role in the activation of Toll pathway and prophenoloxidase systems in insect host defense against fungal invasion. To examine the structure diversity of BGRPs for the recognition of physiochemically different BGs, the binding specificity of BGRPs cloned from four different insects to structure different BGs was characterized using ELISA. Recombinant BGRPs expressed as Fc-fusion proteins of human IgG1 bound to the solid phase of BGs. Based on the binding specificities, the BGRPs were categorized into two groups with different ultrastructures and binding characters; one group specifically binds BGs with triple-helical conformation, while the other group recognizes BGs with disordered conformations like single-helical or partially opened triple helix. The BGRPs from the silkworm and the Indian meal moth bound to the BGs with a triple-helical structure, whereas BGRPs from the red flour beetle and yellow mealworm beetle showed no binding to triple-helical BGs, but bound to alkaline-treated BGs that have a partially opened triple-helical conformation. This evidence suggests that the insect BGRPs can distinguish between different conformations of BGs and are equipped for determining the diversity of BG structures.

Coronary Artery Calcium Is Associated with Left Ventricular Diastolic Function Independent of Myocardial Ischemia.

It has been shown in several studies that coronary artery calcium (CAC) burden or CAC progression is associated with heart failure. We tested the hypothesis that the extent of CAC is associated with left ventricular (LV) diastolic parameters derived from gated myocardial perfusion single-photon emission computed tomography (SPECT) in patients with no evidence of myocardial ischemia.157 patients undergoing coronary computed tomography (CT), gated SPECT, and transthoracic echocardiography (TTE) were enrolled in this study. The CAC score was calculated according to the Agatston method. The peak filling rate (PFR) and the one-third mean filling rate (1/3MFR) were obtained as LV diastolic parameters.There were 139 patients with CAC and 18 patients without. The CAC score ranged from 0 to 4,976. There were no significant differences in the LV end-diastolic volume (LVEDV) (61 ± 21 mL versus 62 ± 22 mL, P = 0.79) and LV ejection fraction (LVEF) (66 ± 9% versus 68 ± 9%, P = 0.43). Patients with CAC had lower PFR than those without (2.2 ± 0.5 EDV/s versus 2.6 ± 0.7 EDV/s, P = 0.03). Multivariate linear regression analysis showed that ln (CAC score + 1) was significantly associated with PFR (ß = -0.20, P = 0.01) and 1/3MFR (ß = -0.18, P = 0.049).Our data suggest that the extent of CAC is inversely associated with LV diastolic parameters derived from gated SPECT independent of myocardial ischemia.

Activation of macrophages by a laccase-polymerized polyphenol is dependent on phosphorylation of Rac1.

Various physiologically active effects of polymerized polyphenols have been reported. In this study, we synthesized a polymerized polyphenol (mL2a-pCA) by polymerizing caffeic acid using mutant Agaricus brasiliensis laccase and analyzed its physiological activity and mechanism of action. We found that mL2a-pCA induced morphological changes and the production of cytokines and chemokines in C3H/HeN mouse-derived resident peritoneal macrophages in vitro. The mechanisms of action of polymerized polyphenols on in vitro mouse resident peritoneal cells have not been characterized in detail previously. Herein, we report that the mL2a-pCA-induced production of interleukin-6 (IL-6) and monocyte chemotactic protein-1 (MCP-1) in C3H/HeN mouse-derived resident peritoneal cells was inhibited by treatment with the Rac1 inhibitor NSC23766 trihydrochloride. In addition, we found that mL2a-pCA activated the phosphorylation Rac1. Taken together, the results show that mL2a-pCA induced macrophage activation via Rac1 phosphorylation-dependent pathways.

Amphiphobic Septa Enhance the Mechanical Stability of Free-Standing Bilayer Lipid Membranes.

Artificial bilayer lipid membranes (BLMs) provide well-defined systems for investigating the fundamental properties of membrane proteins, including ion channels, and for screening the effect of drugs that act on them. However, the application of this technique is limited due to the low stability and low reconstitution efficiency of the process. We previously reported on improving the stability of BLM based on the fabrication of microapertures having a tapered edge in SiO2/Si3N4 septa and efficient ion channel incorporation based on vesicle fusion accelerated by a centrifugal force. Although the BLM stability and incorporation probability were dramatically improved when these approaches were used, some BLMs were ruptured when subjected to a centrifugal force. To further improve the BLM stability, we investigated the effect of modifying the surface of the SiO2/Si3N4 septa on the stability of BLM suspended in the septa. The modified surfaces were characterized in terms of hydrophobicity, lipophobicity, and surface roughness. Diffusion coefficients of the lipid monolayers formed on the modified surfaces were also determined. Highly fluidic lipid monolayers were formed on the amphiphobic substrates that had been modified with long-chain perfluorocarbons. Free-standing BLMs formed in amphiphobic septa showed a much higher mechanical stability, including tolerance to water movement and applied centrifugal forces with and without proteoliposomes, than those formed in the septa that had been modified with a short alkyl chain. These results demonstrate that highly stable BLMs are formed when the surface of the septa has amphiphobic properties. Because highly fluidic lipid monolayers that are formed on the septa seamlessly connect with BLMs in a free-standing region, the high fluidity of the lipids contributes to decreasing potential damage to BLMs when mechanical stresses are applied. This approach to improve the BLM stability increases the experimental efficiency of the BLM systems and will contribute to the development of high-throughput platforms for functional assays of ion channel proteins.

Effects of aortic tortuosity on left ventricular diastolic parameters derived from gated myocardial perfusion single photon emission computed tomography in patients with normal myocardial perfusion.

Aortic tortuosity is often found on chest radiograph, especially in aged patients. We tested the hypothesis that aortic tortuosity was associated with LV diastolic parameters derived from gated SPECT in patients with normal myocardial perfusion. One-hundred and twenty-two patients with preserved LV ejection fraction and normal myocardial perfusion were enrolled. Descending aortic deviation was defined as the horizontal distance from the left line of the aortic knob to the most prominent left line of the descending aorta. This parameter was measured for the quantitative assessment of aortic tortuosity. Peak filling rate (PFR) and one-third mean filling rate (1/3 MFR) were obtained from redistribution images as LV diastolic parameters. Descending aortic deviation ranged from 0 to 22 mm with a mean distance of 4.5 ± 6.3 mm. Descending aortic deviation was significantly correlated with age (r = 0.38, p < 0.001) and estimated glomerular filtration rate (eGFR) (r = - 0.21, p = 0.02). Multivariate linear regression analysis revealed that eGFR (ß = 0.23, p = 0.02) and descending aortic deviation (ß = - 0.23, p = 0.01) were significantly associated with PFR, and that only descending aortic deviation (ß = - 0.21, p = 0.03) was significantly associated with 1/3 MFR. Our data suggest that aortic tortuosity is associated with LV diastolic parameters derived from gated SPECT in patients with normal myocardial perfusion.