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Large primary tumor volume has been identified as a poor prognostic factor of esophageal squamous cell carcinoma (ESCC) treated with definitive concurrent chemoradiotherapy (CCRT). However, when neoadjuvant CCRT and surgery are adopted, the prognostic impact of primary tumor and lymph node (LN) volume on clinical outcomes in ESCC remains to be elucidated. This study included 107 patients who received neoadjuvant CCRT and surgery for ESCC. The volume of the primary tumor and LN was measured using radiotherapy planning computed tomography scans, and was correlated with overall survival (OS), disease-free survival (DFS), and cancer failure pattern. The median OS was 24.2 months (IQR, 11.1-93.9) after a median follow-up of 18.4 months (IQR, 8.1-40.7). The patients with a baseline LN volume > 7.7 ml had a significantly worse median OS compared to those with smaller LN volume (18.8 vs. 46.9 months, p = 0.049), as did those with tumor regression grade (TRG) 3-5 after CCRT (13.9 vs. 86.7 months, p < 0.001). However, there was no association between OS and esophageal tumor volume (p = 0.363). Multivariate analysis indicated that large LN volume (HR 1.753, 95% CI 1.015-3.029, p = 0.044) and high TRG (HR 3.276, 95% CI 1.556-6.898, p = 0.002) were negative prognostic factors for OS. Furthermore, large LN volume was linked to increased locoregional failure (p = 0.033) and decreased DFS (p = 0.041). In conclusion, this study demonstrated that large LN volume is correlated with poor OS, DFS, and locoregional control in ESCC treated with neoadjuvant CCRT and esophagectomy.

BackgroundIn the treatment of methicillin-resistant Staphylococcus aureus (MRSA) bloodstream infections (BSIs), vancomycin stands as the prevalent therapeutic agent. Daptomycin remains an alternative antibiotic to treat MRSA BSIs in cases where vancomycin proves ineffective. However, studies have conflicted on whether daptomycin is more effective than vancomycin among patients with MRSA BSI.ObjectiveTo compare the effectiveness of daptomycin and vancomycin for the prevention of mortality among adult patients with MRSA BSI.MethodsSystematic searches of databases were performed, including Embase, PubMed, Web of Science, and Cochrane Library. The Newcastle Ottawa Scale (NOS) and Revised Cochrane risk-of-bias tool for randomized trials (RoB 2) were used to assess the quality of individual observational and randomized control studies, respectively. Pooled odd ratios were calculated using random effects models.ResultsTwenty studies were included based on a priori set inclusion and exclusion criteria. Daptomycin treatment was associated with non-significant lower mortality odds, compared to vancomycin treatment (OR = 0.81; 95% CI, 0.62, 1.06). Sub-analyses based on the time patients were switched from another anti-MRSA treatment to daptomycin demonstrated that switching to daptomycin within 3 or 5 days was significantly associated with 55% and 45% decreased odds of all-cause mortality, respectively. However, switching to daptomycin any time after five days of treatment was not significantly associated with lower odds of mortality. Stratified analysis based on vancomycin minimum inhibitory concentration (MIC) revealed that daptomycin treatment among patients infected with MRSA strains with MIC≥1 mg/L was significantly associated with 40% lower odds of mortality compared to vancomycin treatment.ConclusionCompared with vancomycin, an early switch from vancomycin to daptomycin was significantly associated with lower odds of mortality. In contrast, switching to daptomycin at any time only showed a trend towards reduced mortality, with a non-significant association. Therefore, the efficacy of early daptomycin use over vancomycin against mortality among MRSA BSIs patients may add evidence to the existing literature in support of switching to daptomycin early over remaining on vancomycin. More randomized and prospective studies are needed to assess this association.

BackgroundThe literature regarding esophageal fistula after definitive concurrent chemotherapy and intensity modulated radiotherapy (IMRT) for esophageal squamous cell carcinoma (ESCC) remains lacking. We aimed to investigate the risk factors of esophageal fistula among ESCC patients undergoing definitive concurrent chemoradiotherapy (CCRT) via IMRT technique.MethodsA total of 129 consecutive ESCC patients receiving definitive CCRT with IMRT between 2008 and 2018 were reviewed. The cumulative incidence of esophageal fistula and survival of patients were estimated by the Kaplan-Meier method and compared between groups by the log-rank test. The risk factors of esophageal fistula were determined with multivariate Cox proportional hazards regression analysis.ResultsMedian follow-up was 14.9 months (IQR, 7.0-28.8). Esophageal perforation was identified in 20 (15.5%) patients, resulting in esophago-pleural fistula in nine, esophago-tracheal fistula in seven, broncho-esophageal fistula in two, and aorto-esophageal fistula in two patients. The median interval from IMRT to the occurrence of esophageal fistula was 4.4 months (IQR, 3.3-10.1). Patients with esophageal fistula had an inferior median overall survival (10.0 vs. 17.2 months, p = 0.0096). T4 (HR, 3.776; 95% CI, 1.383-10.308; p = 0.010) and esophageal stenosis (HR, 2.601; 95% CI, 1.053-6.428; p = 0.038) at baseline were the independent risk factors for esophageal fistula. The cumulative incidence of esophageal fistula was higher in patients with T4 (p = 0.018) and pre-treatment esophageal stenosis (p = 0.045). There was a trend toward better survival after esophageal fistula among patients receiving repair or stenting for the fistula than those only undergoing conservative treatments (median survival, 5.9 vs. 0.9 months, p = 0.058).ConclusionsT4 and esophageal stenosis at baseline independently increased the risk of esophageal fistula in ESCC treated by definitive CCRT with IMRT. There existed a trend toward improved survival after the fistula among patients receiving repair or stenting for esophageal perforation.

BackgroundThis study aimed to investigate the correlation between primary tumor volume and cancer failure patterns in esophageal squamous cell carcinoma (ESCC) treated with definitive concurrent chemoradiotherapy (CCRT) and examine whether increasing radiation dose can improve the outcome.MethodsWe retrospectively reviewed 124 patients with stage III ESCC treated by definitive CCRT. The primary tumor volume calculated from the radiotherapy planning computed tomography scans was correlated to treatment response, time to disease progression, and overall survival. We further analyzed whether a higher radiation dose correlated with better disease control and patient survival.ResultsPatients with poor CCRT response had a larger primary tumor volume than those with good response (97.9 vs 64.3 cm3, P = 0.032). The optimal cutoff value to predict CCRT response was 55.3 cm3. Large primary tumor volume (≥ 55.3 cm3) correlated with shorter time to tumor progression in the esophagus (13.6 vs 48.6 months, P = 0.033) compared with small tumor volume (< 55.3 cm3). For the large esophageal tumors (≥ 55.3 cm3), radiation dose > 60 gray significantly prolonged the time to tumor progression in esophagus (20.3 vs 10.1 months, P = 0.036) and overall survival (12.2 vs 8.0 months, P = 0.030), compared with dose ≤ 60 gray. In contrast, higher radiation dose did not benefit local disease control or overall survival in the small esophageal tumors (< 55.3 cm3).ConclusionLarge primary tumor volume correlates with poor local control and overall survival in ESCC treated with definitive CCRT. Radiation dose > 60 gray can improve the outcomes in patients with large primary tumor. Further prospective dose escalation trials are warranted.

Hyperuricemia (HU) is a marker for heart failure. There are relatively few data in the Asian population regarding the effects of hyperuricemia and gouty disorders on cardiac remodeling and diastolic dysfunction (DD), an intermediate stage in the development of heart failure. We consecutively recruited asymptomatic Asian individuals to undergo cardiovascular surveys. We categorized them into Non-HU, HU, and Gout groups. We measured cardiac structure and indices for diastolic function, including tissue Doppler (TDI)-derived LV e' and E/e'. Among 5525 participants, 1568 had HU and 347 had gout. The presence of gout and higher uric acid levels (SUA) ( = 10 mg/dL) were associated with greater LV wall thickness, greater LV mass/volumes, larger LA volume, lower LV e' and higher E/e'. Higher SUA was associated with greater LV mass index (adjusted coefficient: 0.37), greater mass/volume ratio (adjusted coefficient: 0.01) and larger LA volume index (adjusted coefficient: 0.39, all p

Serological methods are relatively convenient and simple for the detection of pathogens for front-line workers. On-site visualization of the test results plays a pivotal role in the process. However, an efficient, universal labeling agent for antibodies is needed for the development of efficient serological detection tools. In this study, a Bamboo mosaic virus (BaMV)-based viral vector was employed to express recombinant proteins, collectively designated GfED, consisting of Staphylococcus aureus Protein A domain ED (SpaED) fused to either the N- or C-terminal of an improved green florescent protein (GFP) with or without the coat protein (CP) of BaMV, efficiently in Chenopodium quinoa. The GfED in crude leaf extracts could specifically attach to IgG molecules of rabbits and mice, effectively labeling IgG with GFP, emitting green light at 506 nm when excited at 450 nm using simple, handheld equipment. To demonstrate the applicability of GfED in serological assays, we have developed a fluorescent dot blot assay for the rapid detection of Acidovorax citrulli (Ac), a bacterial pathogen of cucurbits, and BaMV, a viral pathogen of bamboos. By using the crude extracts of inoculated C. quinoa leaves expressing GfED as an IgG-labeling agent, the pathogens were easily and quickly detected through uncomplicated operations using simple equipment, with results observable by the naked eye. Examination using fluorescent microscopy and transmission electron microscopy revealed that the GfED subunits may assemble into virus-like particles, which were further involved in the formation of aggregates of GfED-antibody-antigen complexes with the potential for fluorescence signal enhancement. The results suggested that plant-expressed GfED may serve as a promising alternative of IgG-labeling agent for current serological assays.

OBJECTIVES:We retrospectively reviewed the evolution of segmentectomy for pulmonary tuberculosis (TB) and the feasibility of multi- and single-incision video-assisted thoracoscopic segmentectomy. METHODS:Of 348 patients undergoing surgery for TB, the medical records of 121 patients undergoing segmentectomy between January 1996 and November 2015 were reviewed. Clinical information and computed tomography (CT) image characteristics were investigated and analyzed. RESULTS:Eighteen patients underwent direct or intended thoracotomy. Sixty-four underwent video-assisted thoracoscopic segmentectomy (VATS), including 53 multi-incision thoracoscopic segmentectomy (MITS), and 11 single-incision thoracoscopic segmentectomy (SITS). Thirty-nine were converted to thoracotomy. The intended thoracotomy group had more operative blood loss (p = 0.005) and hospital stay (p = 0.001) than the VATS group although the VATS group had higher grade of cavity (p = 0.007). The intended thoracotomy group did not differ from converted thoracotomy in operative time, blood loss, or hospital stay, and the grade of pleural thickening was higher in the converted thoracotomy group (p = 0.001). The converted thoracotomy group had more operative blood loss, hospital stay, and complication rate than the MITS group (p = 0.001, p

Central arterial stiffness has been shown to play a key role in cardiovascular disease. However, evidence regarding such arterial stiffness from various arterial segments in relation to B-type natriuretic peptide (BNP) remains elusive.A total of 1255 participants (47.8% men; mean age: 62.6 ± 12.3 [SD] years) with preserved left ventricular function (ejection fraction ≥50%) and ≥1 risk factors were consecutively studied. Arterial pulse wave velocity (PWV) by automatic device (VP-2000; Omron Healthcare) for heart-femoral (hf-PWV), brachial-ankle (ba-PWV), and heart-carotid (hc-PWV) segments were obtained and related to BNP concentrations (Abbott Diagnostics, Abbott Park, IL, USA).Subjects in the highest hf-PWV quartile were older and had worse renal function and higher blood pressure (all P < 0.05). Elevated PWV (m/s) was correlated with elevated BNP (pg/ml) (beta coefficient = 19.3, 12.4, 5.9 for hf-PWV, ba-PWV, hc-PWV respectively, all p < 0.05). After accounting for clinical co-variates and left ventricle mass index (LVMI), both hf-PWV and ba-PWV were correlated with higher BNP (beta coefficient = 8.3, 6.4 respectively, P < 0.01 for each). Adding both hf-PWV and ba-PWV to LVMI significantly expanded ROC in predicting abnormal BNP>100 pg/ml (both P < 0.01), but only hf-PWV presented significant integrated discrimination improvement to predict risk for BNP concentrations (0.7%, P = 0.029).A significant segmental PWV associated with biomarker BNP concentrations suggests that arterial stiffness is associated with myocardial damage.

Granulomatous inflammation causes severe tissue damage in mycobacterial infection while redox status was reported to be crucial in the granulomatous inflammation. Here, we used a NADPH oxidase 2 (NOX2)-deficient mice (Ncf1-/-) to investigate the role of leukocyte-produced reactive oxygen species (ROS) in mycobacterium-induced granulomatous inflammation. We found poorly controlled mycobacterial proliferation, significant body weight loss, and a high mortality rate after M. marinum infection in Ncf1-/- mice. Moreover, we noticed loose and neutrophilic granulomas and higher levels of interleukin (IL)-1β and neutrophil chemokines in Ncf1-/- mice when compared with those in wild type mice. The lack of ROS led to reduced production of IL-1β in macrophages, whereas neutrophil elastase (NE), an abundant product of neutrophils, may potentially exert increased inflammasome-independent protease activity and lead to higher IL-1β production. Moreover, we showed that the abundant NE and IL-1β were present in the caseous granulomatous inflammation of human TB infection. Importantly, blocking of IL-1β with either a specific antibody or a recombinant IL-1 receptor ameliorated the pulmonary inflammation. These findings revealed a novel role of ROS in the early pathogenesis of neutrophilic granulomatous inflammation and suggested a potential role of IL-1 blocking in the treatment of mycobacterial infection in the lung.

Chronic obstructive pulmonary disease (COPD) is a low grade systemic inflammatory disease characterized by dyspnea and exercise intolerance even under standard therapy. Rhodiola crenulata (RC) has been shown to exert anti-inflammatory effects and to enhance exercise endurance, thereby having the potential to treat COPD. In this 12-week, randomized, double-blind, placebo-controlled clinical trial, 57 patients with stable moderate-to-severe COPD aged 70±8.8 years were given RC (250 mg twice/day) (n=38) or a placebo (250 mg twice/day) (n=19) in addition to their standard regimen. There were no significant differences in anthropometrics, quality of life, lung function, six-minute walk and incremental exercise tests between the two groups at enrollment. Over the 12 weeks, RC was well tolerated, significantly reduced triceps skin thickness (Δ=-1 mm, p=.04), change of FEV1 (4.5%, p=.03), and improved workload (Δ=10%, p=.01); although there were no significant differences in these factors between the two groups. However, there were significant between-group differences in tidal volume and ventilation-CO2-output ratio at peak exercise (both p=.05), which were significantly related to peak work rate (both p

BACKGROUND:Dioxin is an industrial pollutant related to various diseases, but epidemiological data on its effects on the kidney are limited. Therefore, we conducted a study to evaluate the association between dioxin exposure and chronic kidney disease (CKD) and identify the related factors. METHODS:We conducted a community-based cross-sectional study and recruited participants from an area where the residents were exposed to dioxin released from a factory. We defined a "high dioxin level" as polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) ≥ 20 pg WHO98-TEQDF/g lipid in the serum and defined CKD as having an estimated glomerular filtration rate (e-GFR) ≤ 60 mL/min/1.73m2 or a diagnosis of CKD by a physician. The renal function was assessed between 2005 and 2010, and we excluded those who had had kidney diseases before the study started. Comparisons between patients of CKD and those who did not have CKD were made to identify the risk factors for CKD. RESULTS:Of the 2898 participants, 1427 had high dioxin levels, and 156 had CKD. In the univariate analyses, CKD was associated with high dioxin levels, age, gender, metabolic syndrome, diabetes mellitus, hypertension, and high insulin and uric acid levels. After adjusting for other factors, we found high dioxin levels (adjusted odds ratio [AOR] = 1.76, 95% confidence interval [CI]: 1.04-2.99), female gender (AOR = 1.74, 95%CI: 1.20-2.53), hypertension (AOR = 1.68, 95%CI: 1.17-2.42), high insulin levels (AOR = 2.14, 95% CI: 1.26-3.61), high uric acid levels (AOR = 4.25, 95% CI: 2.92-6.20), and older age (AOR = 4.66, 95% CI: 1.87-11.62 for 40-64 year and AOR = 26.66, 95% CI: 10.51-67.62 for age ≥ 65 year) were independent predictors of CKD. CONCLUSION:A high dioxin level was associated with an increased prevalence of CKD. Therefore, the kidney function of populations with exposure to dioxin should be monitored.

BackgroundCoronary artery disease (CAD) may coexist with extracranial carotid artery stenosis (ECAS), but the influence of CAD on procedure-related complications after carotid artery stenting (CAS) has not been well investigated. The study aimed to determine the impact of CAD on the occurrence of peri-CAS cerebral ischemic lesions on diffusion-weighted imaging (DWI) scanning.MethodsCoronary angiography was performed within six months before CAS. DWI scanning was repetitively done within 1 week before and after CAS. Clinical outcome measures were stroke, angina, myocardial infarction and death within 30 days.ResultsAmong 126 patients (69.5±9.0 years) recruited for unilateral protected CAS, 33 (26%) patients had peri-CAS DWI-positive lesions. CAD was noted in 79% (26 in 33) and 48% (45 in 93) of patients with and without peri-CAS DWI-positive lesions (OR, 4.0; 95% CI, 1.6-10.0; P = .0018), and the number of concomitant CAD on coronary angiography was positively correlated with the risk for peri-CAS DWI-positive lesions (P = .0032). In patients with no CAD (n = 55), asymptomatic CAD (n = 41) and symptomatic CAD (n = 30), the occurrence rates of peri-CAS DWI-positive lesions were 13%, 41% and 30% (P = .0048), and the peri-CAS stroke rates were 2%, 7% and 0% (P = .2120).ConclusionsThe severity of morphological CAD and the presence of either symptomatic or asymptomatic CAD are associated with the occurrence of peri-CAS cerebral ischemic lesions.

Total mortality and sudden cardiac death is highly prevalent in patients with chronic kidney disease (CKD). In CKD patients, the protein-bound uremic retention solute indoxyl sulfate (IS) is independently associated with cardiovascular disease. However, the underlying mechanisms of this association have yet to be elucidated. The relationship between IS and cardiac electrocardiographic parameters was investigated in a prospective observational study among early CKD patients. IS arrhythmogenic effect was evaluated by in vitro cardiomyocyte electrophysiological study and mathematical computer simulation. In a cohort of 100 early CKD patients, patients with corrected QT (QTc) prolongation had higher IS levels. Furthermore, serum IS level was independently associated with prolonged QTc interval. In vitro, the delay rectifier potassium current (IK) was found to be significantly decreased after the treatment of IS in a dose-dependent manner. The modulation of IS to the IK was through the regulation of the major potassium ion channel protein Kv 2.1 phosphorylation. In a computer simulation, the decrease of IK by IS could prolong the action potential duration (APD) and induce early afterdepolarization, which is known to be a trigger mechanism of lethal ventricular arrhythmias. In conclusion, serum IS level is independently associated with the prolonged QTc interval in early CKD patients. IS down-regulated IK channel protein phosphorylation and the IK current activity that in turn increased the cardiomyocyte APD and QTc interval in vitro and in the computer ORd model. These findings suggest that IS may play a role in the development of arrhythmogenesis in CKD patients.

Tumor cells display a shift in energy metabolism from oxidative phosphorylation to aerobic glycolysis. A subset of papillary thyroid carcinoma (PTC) is refractory to surgery and radioactive iodine ablation. Doxorubicin and sorafenib are the drugs of choice for treating advanced thyroid cancer but both induce adverse effects. In this study, we assessed the anti-cancer activity of 2-deoxy-d-glucose (2-DG) alone and in combination with doxorubicin or sorafenib in PTC cell lines with (BCPAP) and without (CG3) the BRAFV600E mutation. BCPAP cells were more glycolytic than CG3 cells, as evidenced by their higher extracellular l-lactate production, lower intracellular ATP level, lower oxygen consumption rate (OCR), and lower ratio of OCR/extracellular acidification rate. However, dose-dependent reduction in cell viability, intracellular ATP depletion, and extracellular l-lactate production were observed after 2-DG treatment. Regression analysis showed that cell growth in both cell lines was dependent on ATP generation. 2-DG increased the chemosensitivity of BCPAP and CG3 cell lines to doxorubicin and sorafenib. These results demonstrate that the therapeutic effects of low combined doses of 2-DG and doxorubicin or sorafenib are similar to those of high doses of doxorubicin or sorafenib alone in PTC cell lines regardless of the BRAFV600E mutation.

Milkfish (Chanos chanos), an important marine aquaculture species in southern Taiwan, show considerable euryhalinity but have low tolerance to sudden drops in water temperatures in winter. Here, we used high throughput next-generation sequencing (NGS) to identify molecular and biological processes involved in the responses to environmental changes. Preliminary tests revealed that seawater (SW)-acclimated milkfish tolerated lower temperatures than the fresh water (FW)-acclimated group. Although FW- and SW-acclimated milkfish have different levels of tolerance for hypothermal stress, to date, the molecular physiological basis of this difference has not been elucidated. Here, we performed a next-generation sequence analysis of mRNAs from four groups of milkfish. We obtained 29669 unigenes with an average length of approximately 1936 base pairs. Gene ontology (GO) analysis was performed after gene annotation. A large number of genes for molecular regulation were identified through a transcriptomic comparison in a KEGG analysis. Basal metabolic pathways involved in hypothermal tolerance, such as glycolysis, fatty acid metabolism, amino acid catabolism and oxidative phosphorylation, were analyzed using PathVisio and Cytoscape software. Our results indicate that in response to hypothermal stress, genes for oxidative phosphorylation, e.g., succinate dehydrogenase, were more highly up-regulated in SW than FW fish. Moreover, SW and FW milkfish used different strategies when exposed to hypothermal stress: SW milkfish up-regulated oxidative phosphorylation and catabolism genes to produce more energy budget, whereas FW milkfish down-regulated genes related to basal metabolism to reduce energy loss.

Bone remodeling has been linked to glucose metabolism in animal studies, but the results of human trials were inconclusive. Bisphosphonates may play a role in glucose metabolism through their impacts on bone remodeling enzymes. In this study, we aimed to examine the influence of alendronate usage on the incidence of type 2 diabetes mellitus (DM) among osteoporotic patients.A retrospective cohort study was designed to include osteoporotic patients without DM from a population-based cohort containing 1,000,000 subjects. Patients treated with alendronate (exposed group, N=1,011) were compared with those who received no treatment (age and gender matched non-exposed group, N=3,033). Newly diagnosed DM was identified from medical records by International Classification of Diseases, Ninth Revision, Clinical Modification (ICD-9CM) code. The incidence of DM in both groups was calculated for comparison.The non-exposed group had a significantly higher incidence of DM (Odds ratio 1.21, 95% confidence interval 1.03~1.41) when compared with the exposed group. In subgroup analysis, the DM risk reduction in exposed group was only significant among those younger than 65 years and those without hypertension or dyslipidemia. Patients who were prescribed alendronate more than or equal to 3 times had demonstrated a significant reduction in DM risk.Our study showed alendronate might yield a protective effect for incident DM. This effect became insignificant in patients with older age, dyslipidemia or hypertension. The underlying mechanism needs further exploration with prospective data for confirmation of the observed findings.

Graves' ophthalmopathy (GO) is a disfiguring and sometimes blinding disease, which is characterized by inflammation and swelling of orbital tissues, with fibrosis and adipogenesis being predominant features. The aim of this study is to investigate whether the expression levels of fibrosis-related genes, especially that of connective tissue growth factor (CTGF), are altered in orbital fibroblasts of patients with GO. The role of oxidative stress in the regulation of CTGF expression in GO orbital fibroblasts is also examined. By a SYBR Green-based real time quantitative PCR (RT-QPCR), we demonstrated that the mRNA expression levels of fibronectin, apolipoprotein J, and CTGF in cultured orbital fibroblasts from patients with GO were significantly higher than those of age-matched normal controls (p = 0.007, 0.037, and 0.002, respectively). In addition, the protein expression levels of fibronectin, apolipoprotein J, and CTGF analyzed by Western blot were also significantly higher in GO orbital fibroblasts (p = 0.046, 0.032, and 0.008, respectively) as compared with the control. Furthermore, after treatment of orbital fibroblasts with a sub-lethal dose of hydrogen peroxide (200 μM H2O2), we found that the H2O2-induced increase of CTGF expression was more pronounced in the GO orbital fibroblasts as compared with those in normal controls (20% vs. 7%, p = 0.007). Importantly, pre-incubation with antioxidants including N-acetylcysteine (NAC) and vitamin C, respectively, resulted in significant attenuation of the induction of CTGF in GO orbital fibroblasts in response to H2O2 (p = 0.004 and 0.015, respectively). Taken together, we suggest that oxidative stress plays a role in the alteration of the expression of CTGF in GO orbital fibroblasts that may contribute to the pathogenesis and progression of GO. Antioxidants may be used in combination with the therapeutic agents for effective treatment of GO.

Dendritic cell (DC) vaccines are a newly emerging immunotherapeutic approach for the treatment and prevention of cancer, but major challenges still remain particularly with respect to clinical efficacy. Engineering and optimization of adjuvant formulations for DC-based vaccines is one strategy through which more efficacious treatments may be obtained. In this study, we developed a new ex vivo approach for DC vaccine preparation. We evaluated two highly purified mixed polysaccharide fractions from the root of Astragalus membranaceus and Codonopsis pilosulae, named Am and Cp, for their use in enhancing the efficiency of a DC-based cancer vaccine against metastasis of 4T1 mammary carcinoma in mice. Mixed lymphocyte reaction showed all Am-, Cp- and [Am+Cp]-treated DCs enhanced mouse CD4+ and CD8+ T-cell proliferation. [Am+Cp]-treated DCs exhibited the strongest anti-4T1 metastasis activity in test mice. Treatments with Am, Cp and [Am+Cp] also resulted in augmented expression of CD40, CD80 and CD86 markers in test DCs. Bioinformatics analysis of the cytokine array data from treated DCs identified that [Am+Cp] is efficacious in activation of specific immune functions via mediating the expression of cytokines/chemokines involved in the recruitment and differentiation of defined immune cells. Biochemical analysis revealed that Am and Cp are composed mainly of polysaccharides containing a high level (70-95%) glucose residues, but few or no (< 1%) mannose residues. In summary, our findings suggest that the specific plant polysaccharides Am and Cp extracted from traditional Chinese medicines can be effectively used instead of bacterial LPS as a potent adjuvant in the formulation of a DC-based vaccine for cancer immunotherapies.

Retinopathy of prematurity, formerly known as a retrolental fibroplasia, is a leading cause of infantile blindness worldwide. Retinopathy of prematurity is caused by the failure of central retinal vessels to reach the retinal periphery, creating a nonperfused peripheral retina, resulting in retinal hypoxia, neovascularization, vitreous hemorrhage, vitreoretinal fibrosis, and loss of vision. We established a potential retinopathy of prematurity model by using a green fluorescent vascular endothelium zebrafish transgenic line treated with cobalt chloride (a hypoxia-inducing agent), followed by GS4012 (a vascular endothelial growth factor inducer) at 24 hours postfertilization, and observed that the number of vascular branches and sprouts significantly increased in the central retinal vascular trunks 2-4 days after treatment. We created an angiography method by using tetramethylrhodamine dextran, which exhibited severe vascular leakage through the vessel wall into the surrounding retinal tissues. The quantification of mRNA extracted from the heads of the larvae by using real-time quantitative polymerase chain reaction revealed a twofold increase in vegfaa and vegfr2 expression compared with the control group, indicating increased vascular endothelial growth factor signaling in the hypoxic condition. In addition, we demonstrated that the hypoxic insult could be effectively rescued by several antivascular endothelial growth factor agents such as SU5416, bevacizumab, and ranibizumab. In conclusion, we provide a simple, highly reproducible, and clinically relevant retinopathy of prematurity model based on zebrafish embryos; this model may serve as a useful platform for clarifying the mechanisms of human retinopathy of prematurity and its progression.

To compare the neocortical amyloid loads among cognitively normal (CN), amnestic mild cognitive impairment (aMCI), and Alzheimer's disease (AD) subjects with [(18)F]AV-45 positron emission tomography (PET).[(18)F]AV-45 PET was performed in 11 CN, 13 aMCI, and 12 AD subjects to compare the cerebral cortex-to-whole cerebellum standard uptake value ratios (SUVRs) of global and individual volumes of interest (VOIs) cerebral cortex. The correlation between global cortical [(18)F]AV-45 SUVRs and Mini-Mental State Examination (MMSE) scores was analyzed.The global cortical [(18)F]AV-45 SUVRs were significantly different among the CN (1.08±0.08), aMCI (1.27±0.06), and AD groups (1.34±0.13) (p = 0.0003) with amyloidosis positivity rates of 9%, 62%, and 92% in the three groups respectively. Compared to CN subjects, AD subjects had higher SUVRs in the global cortical, precuneus, frontal, parietal, occipital, temporal, and posterior cingulate areas; while aMCI subjects had higher values in the global cortical, precuneus, frontal, occipital and posterior cingulate areas. There were negative correlations of MMSE scores with SUVRs in the global cortical, precuneus, frontal, parietal, occipital, temporal, posterior cingulate and anterior cingulate areas on a combined subject pool of the three groups after age and education attainment adjustment.Amyloid deposition occurs relatively early in precuneus, frontal and posterior cingulate in aMCI subjects. Higher [(18)F]AV-45 accumulation is present in parietal, occipital and temporal gyri in AD subjects compared to the aMCI group. Significant correlation between MMSE scores and [(18)F]AV-45 SUVRs can be observed among CN, aMCI and AD subjects.

Satellite RNAs (satRNAs) are subviral agents that depend on cognate helper viruses for genome replication and encapsidation. Their negative impacts on helper viruses have been exploited to control plant viral diseases. SatBaMV is a commonly found satRNA associated with Bamboo mosaic virus (BaMV) that infects diverse bamboo species in the field. To investigate the genetic diversity and evolution of satRNAs, we examined seven satBaMV populations derived from five bamboo species and cultivars from Taiwan, China, and India and one from the greenhouse. We found 3 distinct clades among the seven populations. Clade I is consisted of all satBaMV isolates, except for those from Dendrocalamus latiflorus in Taiwan and Bambusa vulgaris in India, which belong to Clades II and III, respectively. Interestingly, nucleotide diversity was lower for Clade I than II and III. However, the nucleotide diversity did not seem to depend on bamboo species or geographic location. Our population genetic analyses revealed the presence of excessive low-frequency polymorphic sites, which suggests that the satBaMV population was under purifying selection and/or population expansion. Further analysis of P20, the only satBaMV gene that encodes a non-structural protein involved in the long-distance movement of satBaMV, showed evidence of purifying selection. Taken together, our results suggest that purifying selection against defective P20 protein is responsible at least in part for the evolution of the satBaMV genome.

OBJECTIVES: Pericardial fat (PCF) may induce local inflammation and subsequent structural remodeling of the left atrium (LA). However, the adverse effects of PCF on LA are difficult to be evaluated and quantified. The atrial electromechanical interval determined by transthoracic echocardiogram was shown to be a convenient parameter which can reflect the process of LA remodeling. The goal of the present study was to investigate the association between the electromechanical interval and PCF. METHODS AND RESULTS: A total of 337 patients with mean age of 51.9 ± 9.0 years were enrolled. The electromechanical interval (PA-PDI) defined as the time interval from the initiation of the P wave deflection to the peak of the mitral inflow A wave on the pulse wave Doppler imaging was measured for every patient. The amount of PCF was determined by multi-detector computed tomography. The PA-PDI interval was significantly correlated with the amount of PCF (r = 0.641, p value

BACKGROUND: Three dimensional (3D) echocardiography-derived measurements of myocardial deformation and twist have recently advanced as novel clinical tools. However, with the exception of left ventricular ejection fraction and mass quantifications in hypertension and heart failure populations, the prognostic value of such imaging techniques remains largely unexplored. METHODS: We studied 200 subjects (mean age: 60.2±16 years, 54% female, female n = 107) with known hypertension (n = 51), diastolic heart failure (n = 61), or systolic heart failure (n = 30), recruited from heart failure outpatient clinics. Fifty-eight healthy volunteers were used as a control group. All participants underwent 3D-based myocardial deformation and twist analysis (Artida, Toshiba Medical Systems, Tokyo, Japan). We further investigated associations between these measures and brain natriuretic peptide levels and clinical outcomes. RESULTS: The global 3D strain measurements of the healthy, hypertension, diastolic heart failure, and systolic heart failure groups were 28.03%, 24.43%, 19.70%, and 11.95%, respectively (all p

A cryptic prokaryotic promoter, designated AV3 promoter, has been previously identified in certain begomovirus genus, including ageratum yellow vein virus isolate NT (AYVV-NT). In this study, we demonstrated that the core nucleotides in the putative -10 and -35 boxes are necessary but not sufficient for promoter activity in Escherichia coli, and showed that AYVV-NT AV3 promoter could specifically interact with single-stranded DNA-binding protein and sigma 70 of E. coli involved in transcription. Several AYVV-NT-encoded proteins were found to increase the activity of AV3 promoter. The transcription start sites downstream to AV3 promoter were mapped to nucleotide positions 803 or 805 in E. coli, and 856 in Nicotiana benthamiana. The eukaryotic activity of AV3 promoter and the translatability of a short downstream open reading frame were further confirmed by using a green fluorescent protein reporter construct in yeast (Saccharomyces cerevisiae) cells. These results suggested that AV3 promoter might be a remnant of evolution that retained cryptic activity at present.

OBJECTIVES: Epicardial fat was closely related to atrial fibrillation (AF). Transthoracic echocardiography (TTE) has been proposed to be a convenient imaging tool in assessing epicardial adipose tissue (EAT). The goal of the present study was to investigate whether the EAT thickness measured on TTE was a useful parameter in predicting procedural outcomes of AF ablations. METHODS AND RESULTS: A total of 227 paroxysmal AF (PAF) and 56 non-paroxysmal AF (non-PAF) patients receiving catheter ablations from 2008-2010 were enrolled. Echocardiography-derived regional EAT thickness from parasternal long-axis view was quantified for each patient. Free of recurrence was defined as the absence of atrial arrhythmias without using antiarrhythmic agents after ablations. The mean EAT thickness of the study population was 6.1 ± 0.8 mm. Non-PAF patients had a thicker EAT than that of PAF patients (7.0 ± 0.7 mm versus 5.9 ± 0.7 mm, p value

Functional endothelial-like cells (EC) have been successfully derived from different cell sources and potentially used for treatment of cardiovascular diseases; however, their relative therapeutic efficacy remains unclear. We differentiated functional EC from human bone marrow mononuclear cells (BM-EC), human embryonic stem cells (hESC-EC) and human induced pluripotent stem cells (hiPSC-EC), and compared their in-vitro tube formation, migration and cytokine expression profiles, and in-vivo capacity to attenuate hind-limb ischemia in mice. Successful differentiation of BM-EC was only achieved in 1/6 patient with severe coronary artery disease. Nevertheless, BM-EC, hESC-EC and hiPSC-EC exhibited typical cobblestone morphology, had the ability of uptaking DiI-labeled acetylated low-density-lipoprotein, and binding of Ulex europaeus lectin. In-vitro functional assay demonstrated that hiPSC-EC and hESC-EC had similar capacity for tube formation and migration as human umbilical cord endothelial cells (HUVEC) and BM-EC (P>0.05). While increased expression of major angiogenic factors including epidermal growth factor, hepatocyte growth factor, vascular endothelial growth factor, placental growth factor and stromal derived factor-1 were observed in all EC cultures during hypoxia compared with normoxia (P

A 70-kDa galactose-specific lectin was purified from the tubers of Dioscorea opposita cv. nagaimo. The purification involved three chromatographic steps: anion exchange chromatography on a Q-Sepharose column, FPLC-anion exchange chromatography on a Mono Q column, and FPLC-gel filtration on a Superdex 75 column. The purified nagaimo lectin presented as a single 35-kDa band in reducing SDS-PAGE while it exhibited a 70-kDa single band in non-reducing SDS-PAGE suggesting its dimeric nature. Nagaimo lectin displayed moderate thermostability, retaining full hemagglutinating activity after heating up to 62°C for 30 minutes. It also manifested stability over a wide pH range from pH 2 to 13. Nagaimo lectin was a galactose-specific lectin, as evidenced by binding with galactose and galactose-containing sugars such as lactose and raffinose. The minimum concentration of galactose, lactose and raffinose required to exert an inhibitory effect on hemagglutinating activity of nagaimo lectin was 20 mM, 5 mM and 40 mM, respectively. Nagaimo lectin inhibited the growth of some cancer cell lines including breast cancer MCF7 cells, hepatoma HepG2 cells and nasopharyngeal carcinoma CNE2 cells, with IC(50) values of 3.71 µM, 7.12 µM and 19.79 µM, respectively, after 24 hour treatment with nagaimo lectin. The induction of phosphatidylserine externalization and mitochondrial depolarization indicated that nagaimo lectin evoked apoptosis in MCF7 cells. However, the anti-proliferative activity of nagaimo lectin was not blocked by application of galactose, signifying that the activity was not related to the carbohydrate binding specificity of the lectin.

Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) have recently been recognized as a potential source for cell-based therapy in various preclinical animal models, such as Parkinson's disease, cerebral ischemia, spinal cord injury, and liver failure; however, the precise cellular and molecular mechanisms underlying the beneficial outcomes remain under investigation. There is a growing concern regarding rejection and alteration of genetic code using this xenotransplantation approach. In this study, a novel strain of murine MSCs derived from the umbilical cord of wild-type and green fluorescent protein (GFP) transgenic mice have been successfully isolated, expanded, and characterized. After 10 passages, the mUC-MSCs developed a rather homogeneous, triangular, spindle-shaped morphology, and were sub-cultured up to 7 months (over 50 passages) without overt changes in morphology and doubling time. Cell surface markers are quite similar to MSCs isolated from other tissue origins as well as hUC-MSCs. These mUC-MSCs can differentiate into osteoblasts, adipocytes, neurons, and astrocytes in vitro, as well as hematopoietic lineage cells in vivo. mUC-MSCs also possess therapeutic potential against two disease models, focal ischemic stroke induced by middle cerebral artery occlusion (MCAo) and acute hepatic failure. Subtle differences in the expression of cytokine-related genes exist between mUC-MSCs and hUC-MSCs, which may retard and jeopardize the advance of cell therapy. Allografts of these newly established mUC-MSCs into various mouse disease models may deepen our insights into the development of more effective cell therapy regimens.

To investigate the plant genes affected by Bamboo mosaic virus (BaMV) infection, we applied a cDNA-amplified fragment length polymorphism technique to screen genes with differential expression. A serine/threonine kinase-like (NbSTKL) gene of Nicotiana benthamiana is upregulated after BaMV infection. NbSTKL contains the homologous domain of Ser/Thr kinase. Knocking down the expression of NbSTKL by virus-induced gene silencing reduced the accumulation of BaMV in the inoculated leaves but not in the protoplasts. The spread of GFP-expressing BaMV in the inoculated leaves is also impeded by a reduced expression of NbSTKL. These data imply that NbSTKL facilitates the cell-to-cell movement of BaMV. The subcellular localization of NbSTKL is mainly on the cell membrane, which has been confirmed by mutagenesis and fractionation experiments. Combined with the results showing that active site mutation of NbSTKL does not change its subcellular localization but significantly affects BaMV accumulation, we conclude that NbSTKL may regulate BaMV movement on the cell membrane by its kinase-like activity. Moreover, the transient expression of NbSTKL does not significantly affect the accumulation of Cucumber mosaic virus (CMV) and Potato virus X (PVX); thus, NbSTKL might be a specific protein facilitating BaMV movement.

Geminiviruses are known to exhibit both prokaryotic and eukaryotic features in their genomes, with the ability to express their genes and even replicate in bacterial cells. We have demonstrated previously the existence of unit-length single-stranded circular DNAs of Ageratum yellow vein virus (AYVV, a species in the genus Begomovirus, family Geminiviridae) in Escherichia coli cells, which prompted our search for unknown prokaryotic functions in the begomovirus genomes. By using a promoter trapping strategy, we identified a novel prokaryotic promoter, designated AV3 promoter, in nts 762-831 of the AYVV genome. Activity assays revealed that the AV3 promoter is strong, unidirectional, and constitutive, with an endogenous downstream ribosome binding site and a translatable short open reading frame of eight amino acids. Sequence analyses suggested that the AV3 promoter might be a remnant of prokaryotic ancestors that could be related to certain promoters of bacteria from marine or freshwater environments. The discovery of the prokaryotic AV3 promoter provided further evidence for the prokaryotic origin in the evolutionary history of geminiviruses.

The literature shows an inconsistent relationship between lifestyle behaviors and metabolic syndrome (MetS), especially in the elderly. We designed this study to investigate the interrelationships among cigarette smoking, tea drinking and MetS, and to verify the factors associated with MetS in elderly males dwelling in rural community. In July 2010, with a whole community sampling method, 414 male subjects aged over 65 dwelling in Tianliao township were randomly sampled. The response rate was 60.8%. Each subject completed the structured questionnaires including sociodemographic characteristics, habitual behaviors (including cigarette smoking and tea drinking habits) and medical history. After an overnight fast, the laboratory and anthropometric data were obtained. MetS was confirmed according to the criteria defined by the modified NCEP ATP III for the male Chinese population. Subjects were split into either non-MetS or MetS groups for further analysis. Of the 361 subjects with complete data, 132 (36.6%) elderly men were classified as having MetS. Using binary logistic regression, body mass index, serum uric acid, high sensitivity C-reactive protein, HOMA index, current smokers (OR = 2.72, 95%CI: 1.03 ~ 7.19), total smoking amount > = 30 (OR = 2.78, 95%CI: 1.31 ~ 5.90) and more than 20 cigarettes daily (OR = 2.54, 95%CI: 1.24 ~ 5.18) were positively associated with MetS. Current un- or partial fermented tea drinker (OR = 0.42, 95%CI: 0.22 ~ 0.84), tea drinking habit for 1-9 years (OR = 0.36, 95%CI: 0.15 ~ 0.90) and more than 240 cc daily (OR = 0.35, 95%CI: 0.17 ~ 0.72) were negatively associated with MetS. In conclusion, this study suggests that smoking habit was positively associated with MetS, but tea drinking habit was negatively associated with MetS in elderly men dwelling in rural community.

BackgroundPPARγ and RBP4 are known to regulate lipid and glucose metabolism and insulin resistance. The influences of PPARγ (C1431T and Pro12Ala) and RBP4 (-803GA) polymorphisms on metabolic syndrome in HIV-infected patients receiving anti-retroviral therapy were examined in this study.Materials and methodsA cross-sectional study of HIV-1 infected adults with antiretroviral therapy for more than one year in the National Cheng Kung University Hospital was conducted. The gene polymorphisms were determined by quantitative PCR.ResultsNinety-one patients were included in the study. Eighty-two (90.1%) patients were males with a mean age of 44.4 years. For the C1431T polymorphism in PPARγ, while patients with the T allele (48.4%) had trends toward lower rate of hypertriglyceridemia, the borderline significance together with insignificant power did not support the protective effect of the T allele against development of hypertriglyceridemia. For the Pro12Ala polymorphism in PPARγ, although patients with the Pro/Ala genotype (8.8%) had a higher level of serum LDL (138.0 vs. 111.5 mg/dl, P = 0.04) and trends toward higher rates of hypercholesterolemia and serum LDL>110 mg/dl, these variables were found to be independent of the Pro/Ala genotype in the multivariate analysis. For the -803GA polymorphism in RBP4, patients with the A allele (23.1%) more often had insulin resistance (HOMA>3.8; 33.3 vs. 8.7%, P = 0.01) and more often received anti-hypoglycemic drugs (14.3 vs. 1.4%, P = 0.04). The detrimental effect of the A allele in RBP4 -803GA polymorphism on development of insulin resistance was supported by the multivariate analysis adjusting for covariates.ConclusionThe impacts of PPARγ C1431T and Pro12Ala polymorphisms on metabolism in HIV-infected patients are not significant. RBP4 -803GA polymorphism has increased risk of insulin resistance in HIV-infected patients with anti-retroviral therapy.

OBJECTIVE: Myosin binding protein C (MYBPC3) plays a role in ventricular relaxation. The aim of the study was to investigate the association between cardiac myosin binding protein C (MYBPC3) gene polymorphisms and diastolic heart failure (DHF) in a human case-control study. METHODS: A total of 352 participants of 1752 consecutive patients from the National Taiwan University Hospital and its affiliated hospital were enrolled. 176 patients diagnosed with DHF confirmed by echocardiography were recruited. Controls were matched 1-to-1 by age, sex, hypertension, diabetes, renal function and medication use. We genotyped 12 single nucleotide polymorphisms (SNPs) according to HapMap Han Chinese Beijing databank across a 40 kb genetic region containing the MYBPC3 gene and the neighboring DNA sequences to capture 100% of haplotype variance in all SNPs with minor allele frequencies ≥ 5%. We also analyzed associations of these tagging SNPs and haplotypes with DHF and linkage disequilibrium (LD) structure of the MYBPC3 gene. RESULTS: In a single locus analysis, SNP rs2290149 was associated with DHF (allele-specific p = 0.004; permuted p = 0.031). The SNP with a minor allele frequency of 9.4%, had an odds ratio 2.14 (95% CI 1.25-3.66; p = 0.004) for the additive model and 2.06 for the autosomal dominant model (GG+GA : AA, 95% CI 1.17-3.63; p = 0.013), corresponding to a population attributable risk fraction of 12.02%. The haplotypes in a LD block of rs2290149 (C-C-G-C) was also significantly associated with DHF (odds ratio 2.10 (1.53-2.89); permuted p = 0.029). CONCLUSIONS: We identified a SNP (rs2290149) among the tagging SNP set that was significantly associated with early DHF in a Chinese population.

Breast cancer ranks as a common and severe neoplasia in women with increasing incidence as well as high risk of metastasis and relapse. Translational and laboratory-based clinical investigations of new/novel drugs are in progress. Medicinal plants are rich sources of biologically active natural products for drug development. The 27-kDa trichosanthin (TCS) is a ribosome inactivating protein purified from tubers of the Chinese herbal plant Trichosanthes kirilowii Maximowicz (common name Tian Hua Fen). In this study, we extended the potential medicinal applications of TCS from HIV, ferticide, hydatidiform moles, invasive moles, to breast cancer. We found that TCS manifested anti-proliferative and apoptosis-inducing activities in both estrogen-dependent human MCF-7 cells and estrogen-independent MDA-MB-231 cells. Flow cytometric analysis disclosed that TCS induced cell cycle arrest. Further studies revealed that TCS-induced tumor cell apoptosis was attributed to activation of both caspase-8 and caspase-9 regulated pathways. The subsequent events including caspase-3 activation, and increased PARP cleavage. With regard to cell morphology, stereotypical apoptotic features were observed. Moreover, in comparison with control, TCS- treated nude mice bearing MDA-MB-231 xenograft tumors exhibited significantly reduced tumor volume and tumor weight, due to the potent effect of TCS on tumor cell apoptosis as determined by the increase of caspase-3 activation, PARP cleavage, and DNA fragmentation using immunohistochemistry. Considering the clinical efficacy and relative safety of TCS on other human diseases, this work opens up new therapeutic avenues for patients with estrogen-dependent and/or estrogen-independent breast cancers.

A dimeric 64-kDa glucosamine-specific lectin was purified from seeds of Phaseolus vulgaris cv. "brown kidney bean." The simple 2-step purification protocol involved affinity chromatography on Affi-gel blue gel and gel filtration by FPLC on Superdex 75. The lectin was absorbed on Affi-gel blue gel and desorbed using 1M NaCl in the starting buffer. Gel filtration on Superdex 75 yielded a major absorbance peak that gave a single 32-kDa band in SDS-PAGE. Hemagglutinating activity was completely preserved when the ambient temperature was in the range of 20 °C-60 °C. However, drastic reduction of the activity occurred at temperatures above 65 °C. Full hemagglutinating activity of the lectin was observed at an ambient pH of 3 to 12. About 50% activity remained at pH 0-2, and only residual activity was observed at pH 13-14. Hemagglutinating activity of the lectin was inhibited by glucosamine. The brown kidney bean lectin elicited maximum mitogenic activity toward murine splenocytes at 2.5 µM. The mitogenic activity was nearly completely eliminated in the presence of 250 mM glucosamine. The lectin also increased mRNA expression of the cytokines IL-2, TNF-α and IFN-γ. The lectin exhibited antiproliferative activity toward human breast cancer (MCF7) cells, hepatoma (HepG2) cells and nasopharyngeal carcinoma (CNE1 and CNE2) cells with IC(50) of 5.12 µM, 32.85 µM, 3.12 µM and 40.12 µM respectively after treatment for 24 hours. Flow cytometry with Annexin V and propidum iodide staining indicated apoptosis of MCF7 cells. Hoechst 33342 staining also indicated formation of apoptotic bodies in MCF7 cells after exposure to brown kidney bean lectin. Western blotting revealed that the lectin-induced apoptosis involved ER stress and unfolded protein response.

BACKGROUND:Twist1a and twist1b are the principal components of twists that negatively regulate a number of cellular signaling events. Expression of runx2 and downstream targets is essential for skeletal development and ventral organizer formation and specification in early vertebrate embryos, but what controls ventral activity of maternal runx2 and how twists function in zebrafish embryogenesis still remain unclear. METHODOLOGY/PRINCIPAL FINDINGS:By studying the loss of twist induced by injection of morpholino-oligonucleotide in zebrafish, we found that twist1a and twist1b, but not twist2 or twist3, were required for proper skeletal development and dorsoventral patterning in early embryos. Overexpression of twist1a or twist1b following mRNA injection resulted in deteriorated skeletal development and formation of typical dorsalized embryos, whereas knockdown of twist1a and twist1b led to the formation of abnormal embryos with enhanced skeletal formation and typical ventralized patterning. Overexpression of twist1a or twist1b decreased the expression of runx2b, whereas twist1a and twist1b knockdown increased runx2b expression. We have further demonstrated that phenotypes induced by twist1a and twist1b knockdown were rescued by runx2b knockdown. CONCLUSIONS/SIGNIFICANCE:Together, these results suggest that twist1a and twist1b control skeletal development and dorsoventral patterning by regulating runx2b in zebrafish and provide potential targets for the treatment of diseases or syndromes associated with decreased skeletal development.

The cardioprotective effects of high-density lipoprotein cholesterol (HDL-C) and apolipoprotein A1 (apoA-I) are well documented, but their effects in the direction of the cardiac differentiation of embryonic stem cells are unknown. We evaluated the effects of exogenous apoA-I expression on cardiac differentiation of ESCs and maturation of ESC-derived cardiomyocytes. We stably over-expressed full-length human apoA-I cDNA with lentivirus (LV)-mediated gene transfer in undifferentiated mouse ESCs and human induced pluripotent stem cells. Upon cardiac differentiation, we observed a significantly higher percentage of beating embryoid bodies, an increased number of cardiomyocytes as determined by flow cytometry, and expression of cardiac markers including α-myosin heavy chain, β-myosin heavy chain and myosin light chain 2 ventricular transcripts in LV-apoA-I transduced ESCs compared with control (LV-GFP). In the presence of noggin, a BMP4 antagonist, activation of BMP4-SMAD signaling cascade in apoA-I transduced ESCs completely abolished the apoA-I stimulated cardiac differentiation. Furthermore, co-application of recombinant apoA-I and BMP4 synergistically increased the percentage of beating EBs derived from untransduced D3 ESCs. These together suggests that that pro-cardiogenic apoA-I is mediated via the BMP4-SMAD signaling pathway. Functionally, cardiomyocytes derived from the apoA-I-transduced cells exhibited improved calcium handling properties in both non-caffeine and caffeine-induced calcium transient, suggesting that apoA-I plays a role in enhancing cardiac maturation. This increased cardiac differentiation and maturation has also been observed in human iPSCs, providing further evidence of the beneficial effects of apoA-I in promoting cardiac differentiation. In Conclusion, we present novel experimental evidence that apoA-I enhances cardiac differentiation of ESCs and iPSCs and promotes maturation of the calcium handling property of ESC-derived cardiomyocytes via the BMP4/SMAD signaling pathway.

Bone loss induced by hypoxia is associated with various pathophysiological conditions, however, little is known about the effects of hypoxia and related signaling pathways on osteoblast differentiation and bone formation. Because bone marrow-derived mesenchymal stem cells (MSCs) survive under hypoxic conditions and readily differentiate into osteoblasts by standard induction protocols, they are a good in vitro model to study the effects of hypoxia on osteoblast differentiation.Using human MSCs, we discovered TWIST, a downstream target of HIF-1α, was induced under hypoxia and acted as a transcription repressor of RUNX2 through binding to the E-box located on the promoter of type 1 RUNX2. Suppression of type 1 RUNX2 by TWIST under hypoxia further inhibited the expression of BMP2, type 2 RUNX2 and downstream targets of RUNX2 in MSCs.Our findings point to the important role of hypoxia-mediated signalling in osteogenic differentiation in MSCs through direct regulation of RUNX2 by TWIST, and provide a method for modifying MSC osteogenesis upon application of these cells in fracture healing and bone reconstruction.

BACKGROUND: Obesity is a multifactorial disease that arises from complex interactions between genetic predisposition and environmental factors. Leptin is central to the regulation of energy metabolism and control of body weight in mammals. METHODOLOGY/PRINCIPAL FINDINGS: To better recapitulate the complexity of human obesity syndrome, we applied N-ethyl-N-nitrosourea (ENU) mutagenesis in combination with a set of metabolic assays in screening mice for obesity. Mapping revealed linkage to the chromosome 6 within a region containing mouse Leptin gene. Sequencing on the candidate genes identified a novel T-to-A mutation in the third exon of Leptin gene, which translates to a V145E amino acid exchange in the leptin propeptide. Homozygous Leptin(145E/145E) mutant mice exhibited morbid obesity, accompanied by adipose hypertrophy, energy imbalance, and liver steatosis. This was further associated with severe insulin resistance, hyperinsulinemia, dyslipidemia, and hyperleptinemia, characteristics of human obesity syndrome. Hypothalamic leptin actions in inhibition of orexigenic peptides NPY and AgRP and induction of SOCS1 and SOCS3 were attenuated in Leptin(145E/145E) mice. Administration of exogenous wild-type leptin attenuated hyperphagia and body weight increase in Leptin(145E/145E) mice. However, mutant V145E leptin coimmunoprecipitated with leptin receptor, suggesting that the V145E mutation does not affect the binding of leptin to its receptor. Molecular modeling predicted that the mutated residue would form hydrogen bond with the adjacent residues, potentially affecting the structure and formation of an active complex with leptin receptor within that region. CONCLUSIONS/SIGNIFICANCE: Thus, our evolutionary, structural, and in vivo metabolic information suggests the residue 145 as of special function significance. The mouse model harboring leptin V145E mutation will provide new information on the current understanding of leptin biology and novel mouse model for the study of human obesity syndrome.

BACKGROUND: Satellite RNAs (satRNAs), virus parasites, are exclusively associated with plant virus infection and have attracted much interest over the last 3 decades. Upon virus infection, virus-specific small interfering RNAs (vsiRNAs) are produced by dicer-like (DCL) endoribonucleases for anti-viral defense. The composition of vsiRNAs has been studied extensively; however, studies of satRNA-derived siRNAs (satsiRNAs) or siRNA profiles after satRNA co-infection are limited. Here, we report on the small RNA profiles associated with infection with Bamboo mosaic virus (BaMV) and its two satellite RNAs (satBaMVs) in Nicotiana benthamiana and Arabidopsis thaliana. METHODOLOGY/PRINCIPAL FINDINGS: Leaves of N. benthamiana or A. thaliana inoculated with water, BaMV alone or co-inoculated with interfering or noninterfering satBaMV were collected for RNA extraction, then large-scale Solexa sequencing. Up to about 20% of total siRNAs as BaMV-specific siRNAs were accumulated in highly susceptible N. benthamiana leaves inoculated with BaMV alone or co-inoculated with noninterfering satBaMV; however, only about 0.1% of vsiRNAs were produced in plants co-infected with interfering satBaMV. The abundant region of siRNA distribution along BaMV and satBaMV genomes differed by host but not by co-infection with satBaMV. Most of the BaMV and satBaMV siRNAs were 21 or 22 nt, of both (+) and (-) polarities; however, a higher proportion of 22-nt BaMV and satBaMV siRNAs were generated in N. benthamiana than in A. thaliana. Furthermore, the proportion of non-viral 24-nt siRNAs was greatly increased in N. benthamiana after virus infection. CONCLUSIONS/SIGNIFICANCE: The overall composition of vsiRNAs and satsiRNAs in the infected plants reflect the combined action of virus, satRNA and different DCLs in host plants. Our findings suggest that the structure and/or sequence demands of various DCLs in different hosts may result in differential susceptibility to the same virus. DCL2 producing 24-nt siRNAs under biotic stresses may play a vital role in the antiviral mechanism in N. benthamiana.

BACKGROUND: Numerous studies were performed to illuminate mechanisms of tumorigenesis and metastases from gene expression profiles of Head and Neck Squamous Cell Carcinoma (HNSCC). The objective of this review is to conduct a network-based meta-analysis to identify the underlying biological signatures of the HNSCC transcriptome. METHODS AND FINDINGS: We included 63 HNSCC transcriptomic studies into three specific categories of comparisons: Pre, premalignant lesions v.s. normal; TvN, primary tumors v.s. normal; and Meta, metastatic or invasive v.s. primary tumors. Reported genes extracted from the literature were systematically analyzed. Participation of differential gene activities across three progressive stages deciphered the evolving nature of HNSCC. In total, 1442 genes were verified, i.e. reported at least twice, with ECM1, EMP1, CXCL10 and POSTN shown to be highly reported across all three stages. Knowledge-based networks of the HNSCC transcriptome were constructed, demonstrating integrin signaling and antigen presentation pathways as highly enriched. Notably, functional estimates derived from topological characteristics of integrin signaling networks identified such important genes as ITGA3 and ITGA5, which were supported by findings of invasiveness in vitro. Moreover, we computed genome-wide probabilities of reporting differential gene activities for the Pre, TvN, and Meta stages, respectively. Results highlighted chromosomal regions of 6p21, 19p13 and 19q13, where genomic alterations were shown to be correlated with the nodal status of HNSCC. CONCLUSIONS: By means of a systems-biology approach via network-based meta-analyses, we provided a deeper insight into the evolving nature of the HNSCC transcriptome. Enriched canonical signaling pathways, hot-spots of transcriptional profiles across the genome, as well as topologically significant genes derived from network analyses were highlighted for each of the three progressive stages, Pre, TvN, and Meta, respectively.

Objective: This scoping review aimed to map existing research on adverse events encountered during telerehabilitation delivery, across rehabilitation populations. This includes identifying characteristics of adverse events (frequency/physical/non-physical, relatedness, severity) and examining adverse events by different modes of telerehabilitation delivery and disease states. Introduction: Telerehabilitation, a subset of telemedicine, has gained traction during the COVID-19 pandemic for remote service delivery. However, no prior scoping review, systematic review, or meta-analysis has identified and summarized the current primary research on adverse events in telerehabilitation. Understanding adverse events, such as falls during physiotherapy or aspiration pneumonia during speech therapy, is crucial for identifying limitations and optimizing delivery through risk mitigation and quality indicators. This understanding could also help to improve the uptake of telerehabilitation among clinicians and patients. This review addresses this gap by summarizing published literature on adverse events during telerehabilitation. Methods: The review followed the Joanna Briggs Institute framework and adhered to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses Extension for Scoping Reviews guidelines. The review protocol was registered and published on Open Science Framework. A comprehensive search across multiple databases (MEDLINE ALL/EMBASE/APA PsycINFO/CENTRAL/CINAHL) was conducted. Screening, extraction, and synthesis were performed in duplicate and independently. Data extraction followed the Template for Intervention Description and Replication framework and also involved extraction on authors, publication year (pre- or post-COVID), population, sample size, and modes of telerehabilitation delivery (asynchronous, synchronous, hybrid). For synthesis, data were summarized quantitatively using numerical counts and qualitatively via content analysis. The data were grouped by intervention type and by type of adverse event. Inclusion criteria: This scoping review included qualitative and quantitative studies published between 2013–2023, written in English, and conducted in any geographic area. All modes of telerehabilitation delivery were included. Systematic reviews, meta-analyses, commentaries, protocols, opinion pieces, conference abstracts, and case series with fewer than five participants were excluded. Results: The search identified 11,863 references, and 81 studies were included in this review with a total of 3,057 participants (mean age:59.3 years; females:44.6%). Modes of telerehabilitation delivery (whether asynchronous, synchronous or hybrid) used in the studies included videoconferencing (52), phone calls (25), text messaging (4), email (6), mobile apps (10), and internet-based virtual reality systems (3). A total of 295 adverse events occurred during 84,534 sessions (0.3%), with the majority being physical (e.g., falls or musculoskeletal pain), non-serious/non-severe/mild, and unrelated to (i.e., not caused by) to the telerehabilitation provided. Conclusions: From the 81 included studies, telerehabilitation was delivered with related adverse events being rare, and mostly characterized as mild/non-severe. A comparable occurrence of adverse events (~30%) was found between asynchronous and synchronous telerehabilitation studies. When categorized by disease type, cardiac telerehabilitation studies had the most frequent adverse events. Detailed reporting of telerehabilitation interventions and adverse event characteristics is recommended for future studies (i.e., use of TIDieR reporting guidelines). Telerehabilitation has the potential to make rehabilitation services more accessible to patients; however, more evidence on the safety of telerehabilitation is needed. [ABSTRACT FROM AUTHOR]

Background: Food-drug interaction (FDI) is prevalent in our day-to-day life. Widely recognised as drug expert, pharmacists are responsible to provide patient education, including on FDI, to ensure optimum safety and efficacy of treatment. Most pharmacists have knowledge and experience regarding FDI to certain extent. However, the level of knowledge, attitude and practice (KAP) towards FDI among pharmacists were yet to elucidated for many countries, including for Malaysia. Objective: This study aims to assess pharmacists' KAP on FDI, their inter-association, as well as association with sociodemographic characteristics in Sabah, Malaysia. Methods & materials: A cross-sectional study was conducted from 1 to 31 May 2023, using online, self-administered Google Form questionnaire, involving 24 government hospitals and 113 government health clinics in Sabah. Results: A total of 273 (or 35.5%) out of 768 pharmacists responded. Over two-third were female and serving government hospitals (79.6%). Mean score of KAP were 72.5 ± 14.3%, 93.2 ± 8.3% and 56.0 ± 16.7%, respectively, reflected good attitude but moderate knowledge and practice. Knowledge gaps identified include common medications such as paracetamol and metformin.Additionally, 28.2% respondents reported lack of FDI coverage during undergraduate, while merely 17.2% have received formal training on FDI after started working. Although 89.0% respondents agree that informing patients about possible FDIs is their responsibility, only 35.9% of the respondents usually or always practiced this. Such discrepancy maybe attributed to insufficient pharmacists' knowledge on FDI, where pharmacists with good level of knowledge have almost 2 times higher odd for good level of practice, compared to those with poor/moderate knowledge (odds ratio, OR: 1.92; 95% CI 1.02–3.61; p = 0.040) in this study. Conclusion: There are significant training gaps to be filled in. Pharmacist possessing good knowledge is a prerequisite for better pharmacy practice. Adequate education strategies covering FDI should be emphasised for all pharmacy undergraduates and practising pharmacists. [ABSTRACT FROM AUTHOR]

BackgroundIndividuals with anorexia nervosa (AN) are often cognitively rigid and behaviorally over-controlled. We previously showed that adult females recovered from AN relative to healthy comparison females had less prefrontal activation during an inhibition task, which suggested a functional brain correlate of altered inhibitory processing in individuals recovered from AN. However, the degree to which these functional brain alterations are related to disease state and whether error processing is altered in AN individuals is unknown.Methodology/principal findingsIn the current study, ill adolescent AN females (n = 11) and matched healthy comparison adolescents (CA) with no history of an eating disorder (n = 12) performed a validated stop signal task (SST) during functional magnetic resonance imaging (fMRI) to explore differences in error and inhibitory processing. The groups did not differ on sociodemographic variables or on SST performance. During inhibitory processing, a significant group x difficulty (hard, easy) interaction was detected in the right dorsal anterior cingulate cortex (ACC), right middle frontal gyrus (MFG), and left posterior cingulate cortex (PCC), which was characterized by less activation in AN compared to CA participants during hard trials. During error processing, a significant group x accuracy (successful inhibit, failed inhibit) interaction in bilateral MFG and right PCC was observed, which was characterized by less activation in AN compared to CA participants during error (i.e., failed inhibit) trials.Conclusion/significanceConsistent with our prior findings in recovered AN, ill AN adolescents, relative to CA, showed less inhibition-related activation within the dorsal ACC, MFG and PCC as inhibitory demand increased. In addition, ill AN adolescents, relative to CA, also showed reduced activation to errors in the bilateral MFG and left PCC. These findings suggest that altered prefrontal and cingulate activation during inhibitory and error processing may represent a behavioral characteristic in AN that is independent of the state of recovery.

Tumor associated macrophages (TAMs) are recruited from the circulation to the tumor site, and can undergo a spectrum of phenotypic changes, with two contrasting activation states described in the literature: the M1 and M2 phenotypes. We previously identified a population of TAMs that express proliferating cell nuclear antigen (PCNA) and are associated with high grade, hormone receptor negative breast cancers and poor outcomes. In the present exploratory study we again found that high PCNA(+) TAM counts in pre-treatment tumor biopsies (102 invasive breast cancer cases from the I-SPY 1 Trial, a prospective neoadjuvant trial with serial core biopsies and gene array data) were associated with high grade, hormone receptor negativity, and decreased recurrence free survival. We explored the association of these PCNA(+) TAMs with the expression of M1 and M2 related genes and, contrary to expectation, observed that high PCNA(+) TAM levels were associated with more M1- than M2-related genes. An immune gene signature, derived from cytotoxic T cell and MHC Class II genes (Tc/ClassII), was developed and we found that high PCNA(+) TAM counts, in the context of a low Tc/ClassII signature score, were associated with significantly worse recurrence free survival in all cases and in hormone receptor negative only cases. We observed similar results using a gene signature-proxy for PCNA(+) TAMs in a larger independent set of 425 neoadjuvant-treated breast cancer cases. The results of this exploratory study indicate that high numbers of PCNA(+) TAMs, in the absence of an anti-tumor immune microenvironment (as indicated by a low Tc/ClassII signature score), are associated with poor outcomes in breast cancer patients treated with neoadjuvant chemotherapy. This, along with the observation that PCNA(+) TAMs were associated predominantly with M1-related genes, may provide new insights into the role of the immune microenvironment in breast cancer.