Factors that Influence Attitudes toward Advance Directives among Hemodialysis Patients.

Purpose: Advance directives (ADs) are legal documents that outline a person's preferences or decisions regarding end-of-life care ahead of time. In Korea, there is insufficient awareness and knowledge about ADs among patients undergoing hemodialysis. This study explored the relationship between perceptions of a good death, knowledge about ADs, and attitudes toward ADs in this patient population. Methods: This cross-sectional survey enrolled 119 hemodialysis patients from a secondary hospital in 2021. The participants completed a self-administered questionnaire, and the data were analyzed using the t-test, analysis of variance, Pearson correlation coefficients, Spearman rank correlation coefficients, and multiple regression analysis. Results: The average score for perceptions of a good death among hemodialysis patients was 2.80 out of 4, with clinical symptoms identified as the most critical factor. The average scores for knowledge about ADs and attitudes toward ADs were 5.69 out of 9 and 2.79 out of 4, respectively. There was a positive correlation between perceptions of a good death and attitudes toward ADs (r=0.34, P<0.001), as well as between knowledge about Ads and attitudes toward ADs (r=0.19, P=0.037). Factors influencing attitudes toward Ads included employment status (ß=0.22, P=0.011), education level (ß=0.22, P=0.013), and perceptions of a good death (ß=0.29, P=0.001), which accounted for 24.8% of the variance in attitudes toward ADs. Conclusion: A positive perception of a good death among patients undergoing hemodialysis was associated with a positive attitude toward ADs. Educational programs are needed to improve individuals' understanding of a good death and encourage the development of end-of-life care plans.

Response surface analyses of antihypertensive effects of angiotensin receptor blockers and amlodipine or hydrochlorothiazide combination therapy in patients with essential hypertension.

While previous studies have examined the dose-response characteristics of certain antihypertensive drugs alone or in combination, response surface analysis for combination therapies involving angiotensin receptor blockers (ARBs) and either amlodipine (AML) or hydrochlorothiazide (HCT) has not been explored, particularly in the context of low-dose combinations. The objectives of present study were to generate useful dose-response information for the combination of ARB/AML or ARB/HCT and to predict the blood pressure lowering effects of combination therapies compared to monotherapies. We reviewed the New Drug Application data of combination drugs of ARB/AML and ARB/HCT. Data on systolic blood pressure (SBP), from studies conducted using a factorial dose-response design over a period of 8-12 weeks, were used. The placebo-subtracted SBP change was used for analysis. Response surface analyses of the collected data were conducted using a polynomial regression model. For ARB/AML combination, the quadratic polynomial regression model containing two linear terms, two quadratic terms, and one interaction term was best fitted to the naïve pooled data. Meanwhile, for ARB/HCT combination, the best-fitted model was a quadratic model that included two linear terms and two quadratic terms. The 1/2-dose combination of these medications, compared to each monotherapy, resulted in predicted SBP reductions that were 8-30% greater. The ratio of the estimated antihypertensive effects of the combination to the expected additive effects of each component ranged from 82% to 100% of the expected effect. These results can provide a rationale for developing lower-dose combinations of ARB/AML or ARB/HCT and assist in designing clinical trials.

Performance comparison of platelet function analyzers in cardiology patients: VerifyNow and Anysis-200 aspirin assays.

BACKGROUND: Analysis of responsiveness to antiplatelet therapy is crucial in the management of patients with cardiovascular diseases. OBJECTIVE: This study aimed to evaluate a new platelet function analysis system (Anysis-200) and to compare it with VerifyNow (Accumetrics, San Diego, CA, USA) in cardiology patients. METHODS: Overall, 125 citrated blood samples were collected from 85 cardiology patients referred for platelet function testing. In Anysis-200, platelet function was measured as blood migration distance (MD) until clogging of flow passage, which is comparable to aspirin resistance units obtained using VerifyNow. The two devices were simultaneously used and compared. RESULTS: The MDs before and after taking aspirin were 175±51 and 247±27 mm, respectively (p < 0.0001). Compared with VerifyNow (reference), the sensitivity and specificity of Anysis-200 was 91.5% and 75.5%, respectively (area under the curve, 0.829). Further, the true positive rate in patients newly taking aspirin was 85% for VerifyNow and 92.5% for Anysis-200, respectively. The Cohen's kappa coefficient between the two devices was 0.682, indicating a relatively high agreement. CONCLUSIONS: Anysis-200, a novel system for assessing platelet aggregation, has accuracy and precision equivalent to that of, and significant agreement with, VerifyNow. Anysis-200 may be useful in screening patients with abnormal platelet reactivity and aspirin nonresponsiveness.

Direct interaction with 14-3-3γ promotes surface expression of Best1 channel in astrocyte.

BACKGROUND: Bestrophin-1 (Best1) is a calcium-activated anion channel (CAAC) that is expressed broadly in mammalian tissues including the brain. We have previously reported that Best1 is expressed in hippocampal astrocytes at the distal peri-synaptic regions, called microdomains, right next to synaptic junctions, and that it disappears from the microdomains in Alzheimer's disease mouse model. Although Best1 appears to be dynamically regulated, the mechanism of its regulation and modulation is poorly understood. It has been reported that a regulatory protein, 14-3-3 affects the surface expression of numerous membrane proteins in mammalian cells. METHODS: The protein-protein interaction between Best1 and 14-3-3γ was confirmed by yeast-two hybrid assay and BiFC method. The effect of 14-3-3γ on Best1-mediated current was measured by whole-cell patch clamp technique. RESULTS: We identified 14-3-3γ as novel binding partner of Best1 in astrocytes: among 7 isoforms of 14-3-3 protein, only 14-3-3γ was found to bind specifically. We determined a binding domain on the C-terminus of Best1 which is critical for an interaction with 14-3-3γ. We also revealed that interaction between Best1 and 14-3-3γ was mediated by phosphorylation of S358 in the C-terminus of Best1. We confirmed that surface expression of Best1 and Best1-mediated whole-cell current were significantly decreased after a gene-silencingof 14-3-3γ without a significant change in total Best1 expression in cultured astrocytes. Furthermore, we discovered that 14-3-3γ-shRNA reduced Best1-mediated glutamate release from hippocampal astrocyte by recording a PAR1 receptor-induced NMDA receptor-mediated current from CA1 pyramidal neurons in hippocampal slices injected with adenovirus carrying 14-3-3γ-shRNA. Finally, through a structural modeling, we found critical amino acid residues containing S358 of Best1 exhibiting binding affinities to 14-3-3γ. CONCLUSIONS: 14-3-3γ promotes surface expression of Best1 channel in astrocytes through direct interaction.

Circulating Microparticles and Coronary Plaque Components Assessed by Virtual Histology Intravascular Ultrasound of the Target Lesion in Patients with Stable Angina.

High levels of microparticles (MPs) circulate in the blood of patients with atherosclerotic diseases where they can serve as potential biomarkers of vascular injury and cardiovascular outcome. We used virtual histology intravascular ultrasound (VH-IVUS) to evaluate the relationship between the levels of circulating MPs and the coronary plaque composition in patients with stable angina. We included 35 patients with stable angina (22 men, age 64 ± 9 years) and a de novo target lesion. Preintervention gray-scale and VH-IVUS analysis was performed across the target lesion. Volumetric analysis was performed over a 10-mm-long segment centered at the minimum luminal site. Blood samples were obtained from the femoral artery before coronary angioplasty. MPs were measured using a solid-phase capture assay from a commercial kit. We divided participants into either a low MPs group or high MPs group based on the median value of MPs. There was no significant difference in baseline characteristics between the groups. The plaque burden and remodeling index were similar between the groups. The presence of VH-IVUS-derived thin-cap fibroatheroma was not different between the groups. The percentage of the necrotic core (NC) was significantly higher in the high MPs group than in the low MPs group, both in planar (17.0 ± 8.8% vs. 24.1 ± 6.9%, p = 0.012) and volumetric analyses (17.0 ± 4.8% vs. 22.1 ± 4.3%, p = 0.002). Circulating MPs were positively correlated with the percentage of the NC area at the minimal luminal site (r = 0.491, p = 0.003) and the percentage of the NC volume (r = 0.496, p = 0.002). Elevated levels of circulating MPs were associated with the amount of NC in the target lesion in those with stable angina, suggesting a potential role of circulating MPs as a biomarker for detecting unstable plaque in patients with stable angina.

Clinicopathologic Features and Molecular Characteristics of Glucose Metabolism Contributing to ¹8F-fluorodeoxyglucose Uptake in Gastrointestinal Stromal Tumors.

Fluorine-18 fluorodeoxyglucose (18F-FDG) positron emission tomography-computed tomography (PET/CT) is useful in the preoperative diagnosis of gastrointestinal stromal tumors (GISTs); however, the molecular characteristics of glucose metabolism of GIST are unknown. We evaluated 18F-FDG uptake on preoperative PET/CT of 40 patients and analyzed the expression of glycolytic enzymes in resected GIST tissues by qRT-PCR, western blotting, and immunohistochemistry. Results of receiver operating characteristic curve analysis showed that the maximum standardized uptake value (SUVmax) cut-off value of 4.99 had a sensitivity of 89.5%, specificity was 76.2%, and accuracy of 82.5% for identifying tumors with a high risk of malignancy. We found that 18F-FDG uptake correlated positively with tumor size, risk grade, and expression levels of glucose transporter 1 (GLUT1), hexokinase 1 (HK1), and lactate dehydrogenase A (LDHA). Elevated HK and LDH activity was found in high-risk tumors. Among the isoforms of GLUT and HK, GLUT1 and HK1 expression increased with higher tumor risk grade. In addition, overexpression of glycolytic enzymes M2 isoform of pyruvate kinase (PKM2) and LDHA was observed in GISTs, especially in high-risk tumors. These results suggest that upregulation of GLUT1, HK1, PKM2, and LDHA may play an important role in GIST tumorigenesis and may be useful in the preoperative prediction of malignant potential.

The Poly-γ-d-Glutamic Acid Capsule Surrogate of the Bacillus anthracis Capsule Is a Novel Toll-Like Receptor 2 Agonist.

Bacillus anthracis is a pathogenic Gram-positive bacterium that causes a highly lethal infectious disease, anthrax. The poly-γ-d-glutamic acid (PGA) capsule is one of the major virulence factors of B. anthracis, along with exotoxins. PGA enables B. anthracis to escape phagocytosis and immune surveillance. Our previous study showed that PGA activates the human macrophage cell line THP-1 and human dendritic cells, resulting in the production of the proinflammatory cytokine interleukin-1ß (IL-1ß) (M. H. Cho et al., Infect Immun 78:387-392, 2010, http://dx.doi.org/10.1128/IAI.00956-09). Here, we investigated PGA-induced cytokine responses and related signaling pathways in mouse bone marrow-derived macrophages (BMDMs) using Bacillus licheniformis PGA as a surrogate for B. anthracis PGA. Upon exposure to PGA, BMDMs produced proinflammatory mediators, including tumor necrosis factor alpha (TNF-α), IL-6, IL-12p40, and monocyte chemoattractant protein 1 (MCP-1), in a concentration-dependent manner. PGA stimulated Toll-like receptor 2 (TLR2) but not TLR4 in Chinese hamster ovary cells expressing either TLR2 or TLR4. The ability of PGA to induce TNF-α and IL-6 was retained in TLR4(-/-) but not TLR2(-/-) BMDMs. Blocking experiments with specific neutralizing antibodies for TLR1, TLR6, and CD14 showed that TLR6 and CD14 also were necessary for PGA-induced inflammatory responses. Furthermore, PGA enhanced activation of mitogen-activated protein (MAP) kinases and nuclear factor-kappa B (NF-κB), which are responsible for expression of proinflammatory cytokines. Additionally, PGA-induced TNF-α production was abrogated not only in MyD88(-/-) BMDMs but also in BMDMs pretreated with inhibitors of MAP kinases and NF-κB. These results suggest that immune responses induced by PGA occur via TLR2, TLR6, CD14, and MyD88 through activation of MAP kinase and NF-κB pathways.

Effect of clinical and RBC hemorheological parameters on myocardial perfusion in patients with type 2 diabetes mellitus.

Myocardial ischemia may be present even when there is no significant stenosis of the epicardial coronary artery, or after coronary angioplasty for significant coronary artery disease. This phenomenon is related to disturbance of the coronary microcirculation or vasomotor tone. The aim of this study was to determine the influence of clinical and RBC hemorheological factors, such as RBC deformability and aggregation, on myocardial perfusion in patients with type 2 diabetes mellitus (DM) when compared to patients without DM, presenting with stable angina or acute coronary syndrome. Myocardial perfusion was graded using the myocardial blush grade (MBG) which describes the relative "blush" or intensity of the radio-opacity of myocardial tissue observed after an epicardial coronary injection of contrast medium during coronary angiography. MBG was counted before any medical or mechanical intervention, and in the myocardial territory without anatomical flow limitation (<50% of luminal narrowing on coronary angiogram), in order to remove the direct influence of anatomical stenosis. Myocardial perfusion in this region was associated with diabetes, renal function, LV diastolic function, inflammatory biomarkers such as hs-CRP, fibrinogen and ESR, but not with the clinical presentation. Among the hemorheological parameters, reduced myocardial perfusion was linked to increased RBC aggregation, but not to variation in RBC deformability. In conclusion, myocardial perfusion was affected by diabetes, left ventricular diastolic function, and inflammatory activity indicated by clinical parameters, and by the hemorheological factor RBC aggregation.

Effect of abciximab on the levels of circulating microparticles in patients with acute myocardial infarction treated by primary angioplasty.

BACKGROUND AND OBJECTIVES: We investigated the effect of the additional use of abciximab during percutaneous coronary intervention (PCI) on the level of procoagulant microparticles (MPs) in patients with ST-segment elevation myocardial infarction (STEMI) who had undergone primary PCI. SUBJECTS AND METHODS: In this study, we studied 86 patients with STEMI (72 men, age 58±13) who had undergone primary PCI. The decision to administer abciximab immediately prior to PCI was left to the discretion of the operator. Blood samples for analysis of MPs were obtained from the femoral artery before and after PCI. MPs with procoagulant potential were measured using a commercial kit. The cellular origins of MPs were determined by antigenic capture with specific antibodies. RESULTS: Procoagulant MPs captured onto annexin V were not changed significantly after PCI {13.4±13.2 nM vs. 13.2±16.1 nM phosphatidylserine equivalent (PS eq), p=0.479}. Abciximab was used in 30 of 86 patients (35%) immediately prior to PCI. In patients who had undergone PCI without abciximab, no significant change in the level of MPs was observed after PCI. However, in the abciximab group, the level of circulating MPs was significantly decreased after PCI (12.0±10.7 nM vs. 7.8±11.7 nM PS eq, p=0.018). Levels of endothelial- and platelet-derived MPs also showed a significant reduction after PCI in the abciximab group. CONCLUSION: Primary PCI with additional abciximab significantly reduced the level of procoagulant MPs regardless of their cellular origins in patients with STEMI.

Alveolar distraction osteogenesis versus autogenous onlay bone graft for vertical augmentation of severely atrophied alveolar ridges after 12 years of long-term follow-up.

OBJECTIVE: The aim of this study was to evaluate alveolar distraction osteogenesis (ADO) and autogenous onlay bone graft (AOBG) procedures by comparing their long-term results and their ability to correct severe vertical alveolar defects for further rehabilitation. STUDY DESIGN: Fourteen patients treated with ADO and 28 patients treated with AOBG were included in this study. Radiographic measurements of alveolar bone heights over time and the implant survival and success were compared for the 2 groups. RESULTS: The vertical bone gain was 8.4 ± 2.6 mm for ADO and 6.5 ± 2.3 mm for AOBG. After a mean follow-up period of 7.1 years, there was no significant difference between the 2 groups regarding alveolar bone heights over time and implant survival and success (P > .05). The cumulative survival and success rates were 97.3%, 92.7% in the ADO group and 94.1%, 90.2% in the AOBG group, respectively. CONCLUSION: Both ADO and AOBG may be reliable and effective for ridge augmentation procedures of severe vertical alveolar bone defects and subsequent dental rehabilitation using implants.

The role of critical shear stress on acute coronary syndrome.

Increased aggregation of RBC is associated with many vascular diseases, including acute coronary syndrome (ACS). Critical shear-stress (CSS) as in index of red cell aggregation is defined as either the minimum shear-stress required dispersing the aggregates. The objective of this study is to access the role of CSS in ACS comparing to SA, and to evaluate the correlation with usual biomarkers for atherosclerosis such as fibrinogen, hs-CRP. 169 SA and 223 ACS patients were finally enrolled. A detailed medical history and laboratory data were obtained for each participant from clinical records. CSS is measured by simultaneous measurement of shear stress and light backscattering using a small disposable kit with a microfluidic hemorheometer. We hypothesized that higher value of CSS might be associated increased thrombosis in ACS. As results, relatively younger age was shown and more male in ACS patients, and inflammatory markers (WBC, hs-CRP) were higher in ACS. Whole blood viscosities were significantly higher in ACS than SA along at all shear rates. CSS was 25.7% higher in ACS (333.8 ± 147.8) than in SA (265.4 ± 149.9 mPa) (p < 0.001). CSS was highly correlated white blood cell counts, hs-CRP, fibrinogen, and erythrocyte sedimentation rate (ESR). Among those variables, fibrinogen, and ESR were strongly correlated with CSS. We may suggest that CSS could be used as a novel risk marker for ACS.

Blood characteristics effect on pulse wave velocity.

PWV, a surrogate marker for vascular stiffness, can be also expressed by the Bramwell-Hill equation. The effect of blood density to PWV has been ignored, because variation of blood density is assumed to be negligible. In some clinical situation, blood density could be changed, and blood density as a mechanical property of blood flow might affect to PWV. While the elastic property plays an important role in determining the wave propagation in an elastic tube, our assumption is that there might be some relation between blood flow and vascular wall, and that the characteristics of blood flow might influence PWV. This study was objected to investigate the role of mechanical and hemorheologic parameters on PWV in subjects with cardiovascular disease. We have measured and analyzed the PWV, hemorheologic parameters, and other clinical parameters in 814 patients with coronary arterial disease scheduled for coronary angiography. There is no commercial method for measuring whole blood density. So, we defined the density score, which is sum of hemoglobin and total protein. And the hemorheologic parameters were measured within 4 hours after sampling by automated microfluidic hemorheometer. And the effect of all the clinical and hemorheologic parameter on PWV was analyzed by multiple linear regression analysis. Many clinical parameters including age and blood pressure, high shear WBV and ESR as hemorheologic parameters, and density score were correlated well with ba-PWV. However, many clinical variables, high shear WBV and ESR lost the independent significance on multivariable regression analysis. Only age, SBP, and density score were independent variables (p < 0.001). In conclusion, density score as a mechanical property of blood might be suggested as an independent variable influencing PWV in addition to age and blood pressure, but hemorheologic parameters, such as RBC deformability, aggregation, and whole blood viscosity do not affect PWV independently.

Local increase in microparticles from the aspirate of culprit coronary arteries in patients with ST-segment elevation myocardial infarction.

OBJECTIVE: It has been reported that the levels of procoagulant microparticles (MPs) are increased in patients with acute coronary syndromes and this may contribute to the formation of intracoronary thrombi. In the current study, we investigated the presence of locally elevated MPs within the culprit coronary arteries of patients with ST-segment elevation myocardial infarction (STEMI). METHODS: The study population consisted of 45 patients with STEMI who underwent primary percutaneous coronary intervention (PCI), and 16 control patients. Before and after PCI, blood samples were collected from the femoral artery and from the culprit coronary arteries. In controls, only peripheral blood was obtained. MPs were measured by a solid-phase capture assay using a commercial kit. The cell origins of MPs were determined by antigenic capture with specific antibodies. RESULTS: Baseline levels of MPs in patients with STEMI were higher than in controls. Before PCI, the levels of MPs were significantly higher in culprit coronary arteries than in peripheral arteries in STEMI patients (20.7 ± 15.5 vs. 14.6 ± 15.4 nM phosphatidylserine (PS) equivalent, p = 0.027). MPs from the culprit coronary artery were significantly reduced after PCI (20.7 ± 15.5 vs. 14.3 ± 14.9 nM PS equivalent, p = 0.010). Similarly, the locally increased levels of endothelial- and platelet-derived MPs within the culprit coronary arteries were significantly decreased after PCI. CONCLUSION: Locally increased levels of MPs in culprit coronary arteries and their significant reduction after successful PCI suggest a potential role in coronary atherothrombosis in the early period of STEMI.

Transconjunctival 25-gauge pars plana vitrectomy and internal limiting membrane peeling for chronic macular edema.

BACKGROUND: The purpose of this study was to investigate the visual and anatomic outcomes in patients with chronic macular edema who underwent 25-gauge pars plana vitrectomy with internal limiting membrane peeling. METHODS: This study was a retrospective chart review of 24 eyes from 21 patients who underwent 25-gauge pars plana vitrectomy and indocyanine green-assisted internal limiting membrane peeling for chronic macular edema. Preoperative and postoperative spectral-domain optical coherence tomography (OCT) was examined for macular thickness and macular volume. Outcomes and variables were analyzed using the two-tailed t-test and Spearman's rank correlation coefficient. RESULTS: Twenty-four eyes from 11 men and 10 women of mean age 69 (range 55-84) years were included. Four patients (17%) had chronic macular edema from uveitis, four (17%) from retinal vein occlusion, and 16 (67%) from diabetes. Mean visual acuity was 20/103 preoperatively and 20/87 postoperatively (P = 0.55). Sixty-three percent of the eyes had improved vision (47% better than 20/40), 21% maintained the same vision, and 17% had worse vision. Forty-seven percent of improved eyes and 30% of total eyes gained more than two lines of visual acuity (range -9 to +7 lines). Mean macular thickness was 455 µm preoperatively and 396 µm postoperatively (P = 0.29). Mean macular volume was 7.9 mm(3) preoperatively and 7.5 mm(3) postoperatively (P = 0.51). The strongest predictor of postoperative visual acuity was initial visual acuity (r = 0.673, P = 0.0003). CONCLUSION: Even though a majority of patients had improved vision and decreased macular thickening after 25-gauge pars plana vitrectomy with internal limiting membrane peeling for chronic macular edema of various etiologies, the difference in visual acuity or macular thickening did not reach statistical significance.

Complete genome sequence of Bacillus anthracis H9401, an isolate from a Korean patient with anthrax.

Bacillus anthracis H9401 (NCCP 12889) is an isolate from a Korean patient with gastrointestinal anthrax. The whole genome of H9401 was sequenced. It is a circular chromosome containing 5,480 open reading frames (ORFs) and two plasmids, pXO1 containing 202 ORFs and pXO2 containing 110 ORFs. H9401 shows high pathogenicity and genome sequence similarity to Ames Ancestor.

Optical coherence tomography predicts visual outcome in macula-involving rhegmatogenous retinal detachment.

PURPOSE: Visual recovery after rhegmatogenous retinal detachment (RRD) repair depends upon various anatomical factors. We investigated spectral-domain optical coherence tomography (SD-OCT) abnormalities, pre- and postoperatively, in patients with nontraumatic RRD and correlated these findings with visual outcome. METHODS: The medical records of all patients presenting to Weill Cornell Medical College with nontraumatic macula-involving RRD from August 2010 to September 2011 were retrospectively reviewed in this single-center, consecutive case series. All patients underwent pre- and postoperative visual acuity (VA) testing, slit-lamp biomicroscopy, and dilated fundus examination. Spectral domain optical coherence tomography was obtained preoperatively in twelve patients and postoperatively in ten patients. RESULTS: Twelve patients (12 eyes) were included in the final analysis. Preoperative optical coherence tomography revealed that the inner segment/outer segment (IS/OS) junction was disrupted in 10/12 eyes (83%), the external limiting membrane (ELM) was disrupted in 9/12 (75%) eyes, cystoid macular edema (CME) was present in 10/12 (83%) eyes, an epiretinal membrane (ERM) was present in 2/12 eyes (17%) and outer retinal corrugation was present in 7/12 (58%) eyes. In postoperative imaging of 10 eyes, the IS/OS junction was disrupted in 4/10 (40%), the ELM was disrupted in 3/10 (30%) eyes, CME was present in 2/10 (20%), and an ERM in 1/10 (10%). All retinas were attached postoperatively. Outer retinal corrugation was the most predictive of worse preoperative (P = 0.0016) and 1-month postoperative visual acuity (P = 0.05). CONCLUSION: Preoperative SD-OCT demonstrating outer retinal corrugation in macula involving RRD predicts poor visual acuity outcome in nontraumatic RRD. Such findings may have implications for the urgency for these eyes to undergo surgical repair.

GFP-tagged E. coli shows bacterial distribution in mouse organs: pathogen tracking using fluorescence signal.

PURPOSE: In vaccine efficacy evaluation, visualization of pathogens in whole organism at each time point would be able to reduce the consuming animals and provide the in vivo information within consistent background with identical organism. MATERIALS AND METHODS: Using IVIS spectrum whole live-animal imaging system, fluorescent intensity was optimized and visualized proportionately by concentrating Escherichia coli MC1061 strain which expresses GFP (E. coli-GFP) in BALB/C mice after injection. RESULTS: Local distribution of disseminated E. coli-GFP was traced in each organ by fluorescence. Detached organ showed more obvious fluorescent signal, and intestine showed strongest fluorescent signal. CONCLUSION: This in vivo imaging method using GFP-tagged pathogen strain suggest quantified infected pathogens by fluorescence intensity in whole animals can provide the information about the localization and distribution after infection.

Expression and characterization of an iron-containing superoxide dismutase from Burkholderia pseudomallei.

A superoxide dismutase (SOD) gene from Burkholderia pseudomallei, the causative agent of melioidosis, was cloned and expressed in Escherichia coli, and its product was functionally and physically characterized. The gene has an open-reading frame of 579 bp. The deduced amino acid sequence has 192 residues with a calculated molecular mass of ~22 kDa. Sequence comparison with other bacterial SODs showed that the protein contains typical metal-binding motifs and other Fe-SOD-conserved residues. The sequence has substantial similarity with other bacterial Fe-SOD sequences. The enzymatic activity of the expressed protein was inhibited by hydrogen peroxide but not by sodium azide or potassium cyanide, attributes that indeed are characteristic of typical bacterial Fe-SODs. Western blotting with antiserum against the recombinant Fe-SOD revealed that it is expressed in B. pseudomallei. Transformed E. coli that expressed the Fe-SOD had significantly increased SOD activity and was highly tolerant to paraquat-mediated replication inhibition, compared to transformed cells carrying an empty vector. Our results provide a basis for further biochemical characterization of the enzyme and elucidation of its role in the pathogenesis of B. pseudomallei.

Ultra-wide-field angiography improves the detection and classification of diabetic retinopathy.

PURPOSE: To evaluate patients with diabetic retinopathy using ultra-wide-field fluorescein angiography and to compare the visualized retinal pathology with that seen on an overly of conventional 7 standard field (7SF) imaging. METHODS: Two hundred and eighteen eyes of 118 diabetic patients who underwent diagnostic fluorescein angiography using the Optos Optomap Panoramic 200A imaging system were included. The visualized area of the retina, retinal nonperfusion, retinal neovascularization, and panretinal photocoagulation were quantified by two independent masked graders. The respective areas identified on the ultra-wide-field fluorescein angiography image were compared with an overly of a modified 7SF image as outlined in the Early Treatment Diabetic Retinopathy Study. RESULTS: Ultra-wide-field fluorescein angiograms imaging, on average, demonstrated 3.2 times more total retinal surface area than 7SF. When compared with 7SF, ultra-wide-field fluorescein angiography showed 3.9 times more nonperfusion (P < 0.001), 1.9 times more neovascularization (P = 0.036), and 3.8 times more panretinal photocoagulation (P < 0.001). In 22 eyes (10%), ultra-wide-field fluorescein angiography demonstrated retinal pathology (including nonperfusion and neovascularization) not evident in an 7SF overly. CONCLUSION: Compared with conventional 7SF imaging, ultra-wide-field fluorescein angiography reveals significantly more retinal vascular pathology in patients with diabetic retinopathy. Improved retinal visualization may alter the classification of diabetic retinopathy and may therefore influence follow-up and treatment of these patients.

Serological Correlate of Protection in Guinea Pigs for a Recombinant Protective Antigen Anthrax Vaccine Produced from Bacillus brevis.

OBJECTIVE: Recombinant protective antigen (rPA) is the active pharmaceutical ingredient of a second generation anthrax vaccine undergoing clinical trials both in Korea and the USA. By using the rPA produced from Bacillus brevis pNU212 expression system, correlations of serological immune response to anthrax protection efficacy were analyzed in a guinea pig model. METHODS: Serological responses of rPA anthrax vaccine were investigated in guinea pigs that were given single or two injections (interval of 4 weeks) of various amounts of rPA combined with aluminumhydroxide adjuvant. Guinea pigs were subsequently challenged by the intramuscular injection with 30 half-lethal doses (30LD50) of virulent Bacillus anthracis spores. Serumantibody titerswere determined by anti-PA IgGELISA and the ability of antibodies to neutralize the cytotoxicity of lethal toxin on J774A.1 cell was measured through the toxin neutralizing antibody (TNA) assay. RESULTS: To examine correlations between survival rate and antibody titers, correlation between neutralizing antibody titers and the extent of protection was determined. Toxin neutralization titers of at least 1176 were sufficient to confer protection against a dose of 30LD50 of virulent anthrax spores of the H9401 strain. Such consistency in the correlation was not observed from those antibody titers determined by ELISA. CONCLUSION: Neutralizing-antibody titers can be used as a surrogate marker.