Differential impacts of clinical, anatomical, and procedural factors on early and late mortality following open thoracoabdominal aortic repair: a retrospective observational study.

BACKGROUND: The operative outcomes of thoracoabdominal aortic aneurysms (TAAAs) are challenged by high operative mortality and disabling complications. This study aimed to explore the baseline clinical, anatomical, and procedural risk factors that impact early and late outcomes following open repair of TAAAs. METHODS: We reviewed the medical records of 290 patients who underwent open repair of TAAAs between 1992 and 2020 at a tertiary referral center. Determinants of early mortality (within 30 days or in hospital) were analyzed using multivariable logistic regression models, while those of overall follow-up mortality were explored using multivariable Cox proportional hazards models and landmark analyses. RESULTS: The rates of early mortality and spinal cord deficits were 13.1% and 11.0%, respectively, with Crawford extent II showing the highest rates. In the logistic regression models, older age (P < 0.001), high cardiopulmonary bypass (CPB) time (P < 0.001), and low surgical volume of the surgeon (P < 0.001) emerged as independent factors significantly associated with early mortality. During follow-up (median, 5.0 years; interquartile range, 1.1-7.6 years), 82 late deaths occurred (5.7%/patient-year). Cox proportional hazards models demonstrated that older age (P < 0.001) and low hemoglobin level (P = 0.032) were significant risk factors of overall mortality, while the landmark analyses revealed that the significant impacts of low surgical volume (P = 0.017), high CPB time (P = 0.002), and Crawford extent II (P = 0.017) on mortality only remained in the early postoperative period, without significant late impacts (all P > 0.05). CONCLUSION: There were differential temporal impacts of perioperative risk variables on mortality in open repair of TAAAs, with older age and low hemoglobin level having significant impacts throughout the postoperative period, and low surgical volume, high CPB time, and Crawford extent II having impacts in the early postoperative phase.

Predictors of Late Adverse Events in Patients with Surgically Treated Type I Aortic Dissection.

OBJECTIVE: Residual aortic dissection (AD) following DeBakey type I AD repair is associated with a high rate of adverse events that need additional intervention or surgery. This study aimed to identify clinical and early post-operative computed tomography (CT) imaging factors associated with adverse events in patients with type I AD after ascending aorta replacement. METHODS: This single centre, retrospective cohort study included consecutive patients with type I AD who underwent ascending aorta replacement from January 2011 to December 2017 and post-operative CT within three months. The primary outcome was AD related adverse events, defined as AD related death and re-operation due to aortic aneurysm or impending rupture. The location and size of the primary intimal tears, aortic diameter, and false lumen status were evaluated. Regression analyses were performed to identify factors associated with AD related adverse events. A decision tree model was used to classify patients as high or low risk. RESULTS: Of 103 participants (55.43 ± 13.94 years; 49.5% male), 24 (23.3%) experienced AD related adverse events. In multivariable Cox regression analysis, connective tissue disease (hazard ratio [HR] 15.33; p < .001), maximum aortic diameter ≥ 40 mm (HR 4.90; p < .001), and multiple (three or more) intimal tears (HR 7.12; p < .001) were associated with AD related adverse events. The three year cumulative survival free from AD related events was lower in the high risk group with aortic diameter ≥ 40 mm and multiple intimal tears (41.7% vs. 90.9%; p < .001). CONCLUSION: Early post-operative CT findings indicating a maximum aortic diameter ≥ 40 mm and multiple intimal tears may predict a higher risk of adverse events. These findings suggest the need for careful monitoring and more vigilant management approaches in these cases.

Prognostic impact of the E/e' ratio in patients with chronic severe aortic regurgitation undergoing aortic valve replacement.

OBJECTIVES: The study objective was to evaluate the clinical implication of left ventricular diastolic dysfunction in patients with chronic severe aortic regurgitation undergoing aortic valve replacement. METHODS: We reviewed the medical records of 323 patients (age, 56.3 ± 14.1 years; 111 female) who underwent aortic valve replacement for chronic severe aortic regurgitation between 2005 and 2019. Left ventricular diastolic dysfunction was assessed by the ratio of peak left ventricular inflow velocity over mitral annular velocity (E/e'). The study end point was the composite of death and heart failure requiring hospital admission. RESULTS: The E/e' ratio was significantly correlated with age, left atrial dimension, left ventricular end-diastolic volume, mitral regurgitation grade, and tricuspid regurgitation grade (all P < .001). During follow-up (1748.3 patient-years), death and heart failure occurred in 36 patients (2.06/patient-year) and 9 patients (0.53/patient-year), respectively. In multivariable analysis, E/e' ratio (per 5 increment, hazard ratio, 1.32; 95% confidence interval, 1.02-1.71; P = .03), age (hazard ratio, 1.06; 95% confidence interval, 1.03-1.10; P < .001), and left ventricular ejection fraction (hazard ratio, 0.94; 95% confidence interval, 0.90-0.98; P = .002) were independent predictors of death and heart failure. The 5-year heart failure-free survival was 94.9% ± 1.7% in patients with E/e' less than 15% and 84.2% ± 4.2% in patients with E/e' 15 or greater (P < .001). CONCLUSIONS: The E/e' ratio was significantly associated with adverse outcomes in patients with chronic severe aortic regurgitation undergoing aortic valve replacement and may be useful as a prognostic marker in such patients.

Effect of preoperative patient education and simulated mouth breathing training on opioid requirements in the post-anesthesia care unit after nasal surgery: a randomized controlled study.

BACKGROUND: A simulated education, prior to surgery about postoperative nasal stuffiness and ease of breathing through the mouth may help patients tolerate discomfort after nasal surgery. This study aimed to investigate the effect of preoperative simulated education on immediate postoperative opioid requirements in patients undergoing elective nasal surgery. METHODS: This randomized controlled trial of 110 patients undergoing nasal surgery randomly allocated patients into either a control (group C) or an education group (group E). One day before surgery, patients in group E were intensively trained to breathe through the mouth by using a nasal clip, with informative explanations about inevitable nasal obstruction and discomfort following surgery. Patients in group C were provided with routine preoperative information. Total intravenous anesthesia (TIVA) with propofol and remifentanil was used for anesthesia. No further opioid was used for analgesia intraoperatively. The primary outcome was index opioid (fentanyl) requirements at the post-anesthesia recovery unit (PACU). Secondary outcomes were emergence agitation, pain scores at the PACU, and postoperative recovery using the Quality of Recovery-15 (QoR15-K). RESULTS: The rate of opioid use in the PACU was 51.0% in the group E and 39.6% in the group C (p = 0.242). Additional request for analgesics other than index opioid was not different between the groups. Emergence agitation, postoperative pain severity, and QoR15-K scores were comparable between the groups. CONCLUSION: Preoperative education with simulated mouth breathing in patients undergoing nasal surgery did not reduce opioid requirements. TRIAL REGISTRATION: KCT0006264; 16/09/2021; Clinical Research Information Services ( https://cris.nih.go.kr ).

Effect of Tulobuterol Patch Versus Placebo on the Occurrence of Respiratory Adverse Events in Children Undergoing Tonsillectomies: A Randomized Controlled Trial.

BACKGROUND: Perioperative respiratory adverse events are common in children. We aimed to evaluate the effect of the transdermal ß-2 agonist, tulobuterol, compared with that of placebo on the incidence of perioperative respiratory adverse events in pediatric patients undergoing tonsillectomy. METHODS: In this triple-blinded (patient, anesthesia provider, and outcome assessor) randomized controlled trial, 188 patients were randomly allocated to receive tulobuterol or a placebo. The tulobuterol groups received a tulobuterol patch (1 mg) masked with a bandage, whereas the placebo only received the bandage. The assigned bandage was applied to the patients 8 to 10 hours before the surgery. The primary outcome was the occurrence of any perioperative respiratory adverse events: oxygen desaturation <95%, airway obstruction, laryngospasm, bronchospasm, severe coughing, or stridor. The outcomes were evaluated using the average relative effect test, which estimates the effect of individual components of a composite outcome and then averages effects across components. RESULTS: A total of 88 and 94 patients who received tulobuterol and placebo, respectively, were analyzed. The incidence of any perioperative respiratory adverse event was lower with tulobuterol (n = 13/88; 14.7%) than that with the placebo (n = 40/94; 42.5%), with an estimated average relative risk (95% confidence interval) across components of 0.35 (0.20-0.60; P < .001). The symptoms of airway obstruction were lower with tulobuterol (n = 8/88; 9.0%) than that with the placebo (n = 32/94; 34.0%), with relative risk (95% CI) of 0.31 (0.17-0.56; P < .001). The occurrence of severe coughing was lower with tulobuterol (n = 1/88; 1.1%) than that with the placebo (n = 8/94; 8.5%), with relative risk (95% CI) of 0.15 (0.03-0.68; P = .014). CONCLUSIONS: In preschool children undergoing tonsillectomy, the preoperative application of a tulobuterol patch could decrease the occurrence of perioperative respiratory adverse events. Further studies are needed to elucidate the effect of the tulobuterol patch in a broad spectrum of pediatric anesthesia.

Left atrial appendage preservation versus closure during surgical ablation of atrial fibrillation.

OBJECTIVE: There is limited evidence regarding the effectiveness of left atrial appendage (LAA) closure during surgical ablation of atrial fibrillation (AF) in yielding superior clinical outcomes. This study aimed to evaluate the association of LAA closure versus preservation with the risk of adverse clinical outcomes among patients undergoing surgical ablation during cardiac surgery. METHODS: We evaluated 1640 patients (aged 58.8±11.5 years, 898 women) undergoing surgical ablation during cardiac surgery (including mitral valve (MV), n=1378; non-MV, n=262) between 2001 and 2018. Of these, 804 had LAA preserved, and the remaining 836 underwent LAA closure. Comparative risks of stroke and mortality between the two groups were evaluated after adjustments with inverse-probability-of-treatment weighting (IPTW). Longitudinal echocardiographic data (n=9674, 5.9/patient) on transmitral A-wave and E/A-wave ratio were analysed by random coefficient models. RESULTS: Adjustment with IPTW yielded patient cohorts well-balanced for baseline profiles. During a median follow-up of 43.5 months (IQR 19.0-87.3 months), stroke and death occurred in 87 and 249 patients, respectively. The adjusted risk of stroke (HR 0.85; 95% CI 0.52-1.39) and mortality (HR 0.80; 95% CI 0.61 to 1.05) did not differ significantly between the two groups. Echocardiographic data demonstrated higher transmitral A-wave velocity (group-year interaction, p=0.066) and lower E/A-wave ratio (group-year interaction, p=0.045) in the preservation group than in the closure group. CONCLUSIONS: LAA preservation during surgical AF ablation was not associated with an increased risk of stroke or mortality. Postoperative LA transport functions were more favourable with LAA preservation than with LAA closure.

Long-Term Outcomes of Surgical Ablation for Atrial Fibrillation: Impact of Ablation Lesion Sets.

Background: The lesion sets for surgical ablation of atrial fibrillation (AF) that provide optimal outcomes have remained controversial. Objectives: We evaluated the effects of left-atrial (LA) ablation of AF compared with bi-atrial (BA) ablation on the clinical and rhythm outcomes, and examined the predictors for AF recurrence and permanent pacing in consideration of ablation lesion sets. Methods: Between 2001 and 2018, 1,965 patients underwent surgical ablation during cardiac surgery at our institution. Among these, 796 and 1,169 patients underwent LA and BA ablation, respectively. The clinical outcomes were evaluated after propensity score adjustment, with death accounting for a competing event. The probability of AF recurrence was estimated with the generalized estimating equations model. Results: The patients with BA ablation had morbidities greater than those with LA ablation. The probability of AF recurrence at 1 and 5 years was 13.9% and 37.1% in patients with LA ablation, and 11.2% and 30.1% in those with BA ablation (hazard ratio [HR]: 1.24; 95% confidence interval [CI]: 0.96-1.61; P = 0.100). After adjustment, LA ablation was associated with a decreased risk of early death (<30 days) (odds ratio [OR]: 0.56; 95% CI: 0.31-0.96; P = 0.041) and new-onset dialysis (OR 0.47; 95% CI: 0.27-0.78; P = 0.003). However, the risk of overall mortality (HR: 1.03; 95% CI: 0.75-1.41; P = 0.878) and permanent pacing (HR: 0.68; 95% CI: 0.43-1.06; P = 0.091) was comparable between the 2 groups. Conclusions: The risk of AF recurrence and adverse events was comparable between the 2 ablation lesion sets. BA ablation was not related to an increased risk of permanent pacing.

The prognostic nutritional index and postoperative complications after curative lung cancer resection: A retrospective cohort study.

OBJECTIVE: The prognostic nutritional index is a score that represents a patient's immune-nutritional status based on the lymphocyte count and serum albumin concentration. We hypothesized that preoperative prognostic nutritional index is associated with postoperative complications and long-term outcomes after curative resection of lung cancer. METHODS: We retrospectively analyzed 1011 patients with pathologic stage I-III adenocarcinoma and squamous cell carcinoma who underwent open thoracotomy for curative resection of lung cancer. The preoperative prognostic nutritional index was calculated as follows based on preoperative laboratory data: 10 × serum albumin (g/dL) + 5 × total lymphocyte count (/nL). The cutoff value of prognostic nutritional index (cutoff value: 50) was obtained by receiver operating characteristics curve and patients were classified as high and low groups. Outcomes were compared with the use of propensity scores and inverse probability weighting adjustment to reduce treatment selection bias. RESULTS: The low group exhibited more postoperative complications (34% [96/285] vs 24% [174/726]; P = .002) especially pneumonia (13% [36/285] vs 6% [41/756]; P < .001) and delirium (10% [29/285] vs 5% [36/726]; P = .002), and greater in-hospital mortality (4% [11/285] vs 1% [9/726]; P = .007) than the high group. A low prognostic nutritional index was associated with greater postoperative pulmonary complications [odds ratio, 1.7; 95% confidence interval, 1.3-2.3], lower recurrence-free survival (hazard ratio, 1.3; 95% confidence interval, 1.1-1.5), and overall survival (hazard ratio, 1.5; 95% confidence interval, 1.2-1.8) after balancing the covariables. CONCLUSIONS: The preoperative prognostic nutritional index was associated with postoperative pulmonary complications and long-term outcomes after curative resection of non-small cell lung cancer.

The effect of preventive use of corticosteroids on postoperative complications after esophagectomy: A retrospective cohort study.

Corticosteroids have been empirically administered to reduce the rate of acute respiratory distress syndrome (ARDS) after esophagectomy. However, their efficacy remains controversial, and corticosteroids may increase the risk of graft dehiscence and infection, which are major concerns after esophagectomy. Therefore, we compared the incidence of composite complications (ARDS, graft dehiscence and infection) after esophagectomy between patients who received a preventive administration of corticosteroids and those who did not. All patients who underwent esophagectomy from 2010 to 2015 at a tertiary care university hospital were reviewed retrospectively (n = 980). Patients were divided into Steroid (n = 120) and Control (n = 860) groups based on the preventive administration of 100 mg hydrocortisone during surgery. The primary endpoint was the incidence of composite complications. The incidence of composite complications was not different between the Control and Steroid groups (17.4% vs. 21.7% respectively; P = 0.26). The incidence rates of complications in each category were not different between the Control and Steroid groups: ARDS (3.8% vs. 5.0%; P = 0.46), graft dehiscence (4.8% vs. 6.7%; P = 0.37), and infection (12.8% vs. 15.8%; P = 0.36). Propensity score matching revealed that composite complications (20.0% vs. 21.7%; P = 0.75), ARDS (4.3% vs. 5.2%; P = 0.76) and infection (16.5% vs. 15.7%; P = 0.86) were not different between the Control and Steroid group, but the incidence of graft dehiscence was higher in the Steroid group than in the Control group (0.9% vs. 7.0%; P = 0.0175). In conclusions, the preventive use of corticosteroids did not reduce the incidence of ARDS, but may be related to an increased incidence of graft dehiscence. Therefore, routine administration of corticosteroids to prevent ARDS is not recommended in esophagectomy.

A retrospective analysis on the relationship between intraoperative hypothermia and postoperative ileus after laparoscopic colorectal surgery.

Postoperative ileus (POI) is an important factor prolonging the length of hospital stay following colorectal surgery. We retrospectively explored whether there is a clinically relevant association between intraoperative hypothermia and POI in patients who underwent laparoscopic colorectal surgery for malignancy within the setting of an enhanced recovery after surgery (ERAS) program between April 2016 and January 2017 at our institution. In total, 637 patients were analyzed, of whom 122 (19.2%) developed clinically and radiologically diagnosed POI. Overall, 530 (83.2%) patients experienced intraoperative hypothermia. Although the mean lowest core temperature was lower in patients with POI than those without POI (35.3 ± 0.5°C vs. 35.5 ± 0.5°C, P = 0.004), the independence of intraoperative hypothermia was not confirmed based on multivariate logistic regression analysis. In addition to three variables (high age-adjusted Charlson comorbidity index score, long duration of surgery, high maximum pain score during the first 3 days postoperatively), cumulative dose of rescue opioids used during the first 3 days postoperatively was identified as an independent risk factor of POI (odds ratio = 1.027 for each 1-morphine equivalent [mg] increase, 95% confidence interval = 1.014-1.040, P <0.001). Patients with hypothermia showed significant delays in both progression to a soft diet and discharge from hospital. In conclusion, intraoperative hypothermia was not independently associated with POI within an ERAS pathway, in which items other than thermal measures might offset its negative impact on POI. However, as it was associated with delayed discharge from the hospital, intraoperative maintenance of normothermia is still needed.

Morphine Suppresses Lung Cancer Cell Proliferation Through the Interaction with Opioid Growth Factor Receptor: An In Vitro and Human Lung Tissue Study.

BACKGROUND: There have been inconsistent reports on whether opioids promote or inhibit lung cancer growth. In this study, we suggest that opioid growth factor receptor (OGFR), a negative regulator of cell proliferation, is a binding site of morphine and is involved in subsequent morphine-induced lung cancer growth suppression. METHODS: The expression and distribution of OGFR in human lung cancer tissues and cell lines were assessed with immunohistochemistry and real-time reverse transcription polymerase chain reaction. The human lung cancer cell line, H1975 (adenocarcinoma), which overexpressed OGFR but not µ-opioid receptors, was selected for further analysis to verify the interaction between morphine and OGFR and the impact of morphine on cancer cell growth. RESULTS: OGFR was expressed in lung cancer tissues and all cancer cell lines tested. Adenocarcinoma showed a higher OGFR expression than squamous cell carcinoma (reverse transcription polymerase chain reaction relative quantitation value: median [interquartile range], 13.1 [9.3-20.0] vs 4.3 [2.2-6.6]; P = 0.003). OGFR expression showed an inverse correlation with cell proliferation (r = -0.92, P = 0.0001). Morphine treatment reduced the median H1975 cell number by approximately 23% (P = 0.03). Growth suppression by morphine was attenuated when OGFR was knocked down. A confocal experiment demonstrated binding of morphine to OGFR. Growth suppression by morphine occurred in the S phase of the cell cycle. CONCLUSIONS: Lung cancer tissues and cell lines express OGFR. Morphine interacts with OGFR and may suppress lung cancer progression.

Functional Roles of Aromatic Residues and Helices of Papiliocin in its Antimicrobial and Anti-inflammatory Activities.

A cecropin-like peptide, papiliocin, isolated from the swallowtail butterfly Papilio xuthus, possesses high selectivity against gram-negative bacteria. Since Trp(2) and Phe(5) are highly conserved residues in cecropin-like peptides, we investigated the role of Trp(2) and Phe(5) in antibacterial activity. Substitution of Trp(2) and Phe(5) in papiliocin with Ala (papiliocin-2A and papiliocin-5A) revealed that Trp(2) is a key residue in its antibacterial activities. In order to understand the structural requirements for papiliocin function and to design shorter, but more potent, peptide antibiotics, we designed papiliocin constructs, PapN (residues Arg(1)-Ala(22) from the N-terminal amphipathic helix). PapN exhibited significant broad-spectrum antibacterial activities without cytotoxicity. Bactericidal kinetics of peptides against E.coli showed that papiliocin completely and rapidly killed E.coli in less than 10 minutes at 2× MIC concentration, while papiliocin-2A and papiliocin-5A killed four times more slowly than papiliocin. The PapN series peptides permeabilized bacterial membranes less effectively than papiliocin, showing no antibacterial activities in an hour. The results imply that the Trp(2) and Phe(5) in the amphipathic N-terminal helix are important in the rapid permeabilization of the gram-negative bacterial membrane. The hydrophobic C-terminal residues permeabilize the hydrophobic bacterial cell membrane synergistically with these aromatic residues, providing selectivity against gram-negative bacteria.

Profiling of viral proteins expressed from the genomic RNA of Japanese encephalitis virus using a panel of 15 region-specific polyclonal rabbit antisera: implications for viral gene expression.

Japanese encephalitis virus (JEV), a mosquito-borne flavivirus, is closely related to West Nile (WN), yellow fever (YF), and dengue (DEN) viruses. Its plus-strand genomic RNA carries a single open reading frame encoding a polyprotein that is cleaved into three structural (C, prM/M, and E) and at least seven nonstructural (NS1/NS1', NS2A, NS2B, NS3, NS4A, NS4B, and NS5) proteins, based on previous work with WNV, YFV, and DENV. Here, we aimed to profile experimentally all the viral proteins found in JEV-infected cells. We generated a collection of 15 JEV-specific polyclonal antisera covering all parts of the viral protein-coding regions, by immunizing rabbits with 14 bacterially expressed glutathione-S-transferase fusion proteins (for all nine viral proteins except NS2B) or with a chemically synthesized oligopeptide (for NS2B). In total lysates of JEV-infected BHK-21 cells, immunoblotting with these antisera revealed: (i) three mature structural proteins (~12-kDa C, ~8-kDa M, and ~53-kDa E), a precursor of M (~24-kDa prM) and three other M-related proteins (~10-14 kDa); (ii) the predicted ~45-kDa NS1 and its frameshift product, ~58-kDa NS1', with no evidence of the predicted ~25-kDa NS2A; (iii) the predicted but hardly detectable ~14-kDa NS2B and an unexpected but predominant ~12-kDa NS2B-related protein; (iv) the predicted ~69-kDa NS3 plus two major cleavage products (~34-kDa NS3N-term and ~35-kDa NS3C-term), together with at least nine minor proteins of ~16-52 kDa; (v) the predicted ~14-kDa NS4A; (vi) two NS4B-related proteins (~27-kDa NS4B and ~25-kDa NS4B'); and (vii) the predicted ~103-kDa NS5 plus at least three other NS5-related proteins (~15 kDa, ~27 kDa, and ~90 kDa). Combining these data with confocal microscopic imaging of the proteins' intracellular localization, our study is the first to provide a solid foundation for the study of JEV gene expression, which is crucial for elucidating the regulatory mechanisms of JEV genome replication and pathobiology.

Risk factors of emergence agitation in adults undergoing general anesthesia for nasal surgery.

OBJECTIVES: To identify the incidence and the risk factors of emergence agitation in adults undergoing general anesthesia for nasal surgery. METHODS: We retrospectively examined 792 patients aged ≥18 years who underwent general anesthesia for elective nasal surgery between July 2012 and August 2013. Patients in the postanesthesia care unit with a Richmond Agitation Sedation Scale≥+1 at any time were considered to have emergence agitation. RESULTS: The overall incidence of emergence agitation is 22.2%. From multivariate regression analysis, the following six variables were found to be significantly associated with emergence agitation (P<0.05): younger age, recent smoking, sevoflurane anesthesia, postoperative pain on the numerical rating scale (NRS)≥5, presence of a tracheal tube, and presence of a urinary catheter. Presence of a tracheal tube was the greatest risk factor, increasing the risk of developing emergence agitation by approximately fivefold (odds ratio, 5.448; 95% confidence interval, 2.973 to 9.982). Younger age was also a strong risk factor (odds ratio, 0.975 for each 1-year increase; 95% confidence interval, 0.964 to 0.987). Current smoking, sevoflurane anesthesia, postoperative pain of NRS≥5, and the presence of a urinary catheter nearly doubled the risk of emergence agitation. CONCLUSION: Emergence agitation following general anesthesia is a common complication in adult nasal surgery patients. To reduce the occurrence and consequences of agitation episodes, elimination of the associated risk factors is necessary, especially in at-risk patients.

A molecularly cloned, live-attenuated japanese encephalitis vaccine SA14-14-2 virus: a conserved single amino acid in the ij Hairpin of the Viral E glycoprotein determines neurovirulence in mice.

Japanese encephalitis virus (JEV), a mosquito-borne flavivirus that causes fatal neurological disease in humans, is one of the most important emerging pathogens of public health significance. JEV represents the JE serogroup, which also includes West Nile, Murray Valley encephalitis, and St. Louis encephalitis viruses. Within this serogroup, JEV is a vaccine-preventable pathogen, but the molecular basis of its neurovirulence remains unknown. Here, we constructed an infectious cDNA of the most widely used live-attenuated JE vaccine, SA14-14-2, and rescued from the cDNA a molecularly cloned virus, SA14-14-2MCV, which displayed in vitro growth properties and in vivo attenuation phenotypes identical to those of its parent, SA14-14-2. To elucidate the molecular mechanism of neurovirulence, we selected three independent, highly neurovirulent variants (LD50, <1.5 PFU) from SA14-14-2MCV (LD50, >1.5×105 PFU) by serial intracerebral passage in mice. Complete genome sequence comparison revealed a total of eight point mutations, with a common single G1708→A substitution replacing a Gly with Glu at position 244 of the viral E glycoprotein. Using our infectious SA14-14-2 cDNA technology, we showed that this single Gly-to-Glu change at E-244 is sufficient to confer lethal neurovirulence in mice, including rapid development of viral spread and tissue inflammation in the central nervous system. Comprehensive site-directed mutagenesis of E-244, coupled with homology-based structure modeling, demonstrated a novel essential regulatory role in JEV neurovirulence for E-244, within the ij hairpin of the E dimerization domain. In both mouse and human neuronal cells, we further showed that the E-244 mutation altered JEV infectivity in vitro, in direct correlation with the level of neurovirulence in vivo, but had no significant impact on viral RNA replication. Our results provide a crucial step toward developing novel therapeutic and preventive strategies against JEV and possibly other encephalitic flaviviruses.

Metformin inhibits tumor cell migration via down-regulation of MMP9 in tamoxifen-resistant breast cancer cells.

Metformin is the most widely used anti-diabetic drug in the world. Recent evidence indicates that metformin could potentially inhibit tumorigenesis. In the present study, we found that metformin inhibited cell migration and invasion of phorbol 12-myristate 13-acetate-induced MCF-7 and tamoxifen-resistant MCF-7 breast cancer cells. This inhibition was correlated with the modulation of matrix metalloproteinase-9 (MMP9) via the suppression of its expression and proteolytic activity. These results indicate that metformin leads to the suppression of migration and invasion through regulation of MMP9 and it may have potential as an anticancer drug for therapy in human breast cancer, especially of chemoresistant cancer cells.

Nrf2-mediated HO-1 induction coupled with the ERK signaling pathway contributes to indirect antioxidant capacity of caffeic acid phenethyl ester in HepG2 cells.

The objective of this study is to investigate the contributing effect of the nuclear transcription factor-erythroid 2-related factor 2 (Nrf2)-mediated signaling pathway on the indirect antioxidant capacity of caffeic acid phenethyl ester (CAPE) against oxidative stress in HepG2 cells. The result of an antioxidant response element (ARE)-luciferase assay showed that CAPE stimulated ARE promoter activity resulting in increased transcriptional and translational activities of heme oxygenase-1 (HO-1). In addition, CAPE treatment enhanced Nrf2 accumulation in the nucleus and the post-translational phosphorylation level of extracellular signal-regulated kinase (ERK) among several protein kinases tested. Treatment with ERK inhibitor U126 completely suppressed CAPE-induced ERK phosphorylation and HO-1 expression, but it only partly inhibited CAPE-induced Nrf2 accumulation and ARE promoter. Using the 2',7'-dichlorofluorescein-diacetate (DCFH-DA) method, the cellular antioxidant capacity of CAPE against 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH)- or H2O2-induced oxidative stress also was shown to be partially suppressed by the ERK inhibitor. From the overall results it is proposed that the indirect antioxidant activity of CAPE against oxidative stress in HepG2 cells is partially attributed to induction of HO-1, which is regulated by Kelch-like erythroid-cell-derived protein with CNC homology (ECH)-associated protein 1 (Keap1)-independent Nrf2 activation relying on post-translational phosphorylation of ERK.

Genistein inhibits osteoclastic differentiation of RAW 264.7 cells via regulation of ROS production and scavenging.

Genistein, a phytoestrogen, has been demonstrated to have a bone-sparing and antiresorptive effect. Genistein can inhibit the osteoclast formation of receptor activator of nuclear factor-κB ligand (RANKL)-induced RAW 264.7 cells by preventing the translocation of nuclear factor-κB (NF-κB), a redox-sensitive factor, to the nucleus. Therefore, the suppressive effect of genistein on the reactive oxygen species (ROS) level during osteoclast differentiation and the mechanism associated with the control of ROS levels by genistein were investigated. The cellular antioxidant capacity and inhibitory effect of genistein were confirmed. The translation and activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 1 (Nox1), as well as the disruption of the mitochondrial electron transport chain system were obviously suppressed by genistein in a dose-dependent manner. The induction of phase II antioxidant enzymes, such as superoxide dismutase 1 (SOD1) and heme oxygenase-1 (HO-1), was enhanced by genistein. In addition, the translational induction of nuclear factor erythroid 2-related factor 2 (Nrf2) was notably increased by genistein. These results provide that the inhibitory effects of genistein on RANKL-stimulated osteoclast differentiation is likely to be attributed to the control of ROS generation through suppressing the translation and activation of Nox1 and the disruption of the mitochondrial electron transport chain system, as well as ROS scavenging through the Nrf2-mediated induction of phase II antioxidant enzymes, such as SOD1 and HO-1.

Ligation of CD47 induces G1 arrest in EBV-transformed B cells through ROS generation, p38 MAPK/JNK activation, and Tap73 upregulation.

CD47 is expressed in normal activated cells as well as in several tumors. It also has been implicated as having antiangiogenic and antimetastatic properties, but its roles in Epstein-Barr virus (EBV)-transformed B cells are still not fully understood. Herein, we report that EBV infection induced CD47 surface expression on B cells, and CD47 ligation with anti-CD47 mAb (B6H12) reduced cell proliferation and induced G1 arrest. CD47-induced G1 arrest was mediated through increased cyclin-dependent kinase inhibitors (CDKi) and a simultaneously decreased CDK/cyclins, and p38 MAPK/JNK activation preceded binding of CDKi-CDK. Moreover, reactive oxygen species (ROS) generation and upregulation of both TAp73 and ER stress sensor proteins were detected after CD47 ligation, and p38 inhibitor SB203580 and JNK inhibitor SP600125 blocked upregulation of TAp73 and cell cycle arrest. We investigated whether ROS generation is the initial event of CD47-mediated G1 arrest because ROS scavenger NAC effectively abrogated the majority of CD47-mediated responses but SB203580 and SP600125 did not block ROS production. Taken together, we concluded that CD47 ligation on EBV-transformed B cells led to G1 arrest by ROS generation and, subsequently, there was p38 MAPK/JNK pathway activation, ER stress triggering, and TAp73 upregulation. Our findings provide data supporting CD47 as a feasible target for EBV-associated tumor therapy.

Core needle biopsy can minimise the non-diagnostic results and need for diagnostic surgery in patients with calcified thyroid nodules.

PURPOSE: To evaluate the role of core needle biopsy (CNB) for calcified thyroid nodules. METHODS: Between October 2008 and July 2011, 264 patients underwent ultrasound-guided CNB for 272 calcified thyroid nodules at our institution. We retrospectively evaluated the incidence of technical failure, non-diagnostic readings, and the diagnostic performance of CNB, and analysed the relationship between the types of calcification and the CNB results. Finally, the incidence of diagnostic surgery was calculated. RESULTS: The incidence of technical failure was 1.1 % (3/275) and that of non-diagnostic results was 0.7 % (2/272). The diagnostic accuracy, sensitivity, specificity, positive predictive value, and negative predictive value of CNB were 94.7 %, 89.5 %, 100 %, 100 %, and 90.2 %, respectively. There were no significant differences according to the calcification subtype for either the non-diagnostic results or the incidence of technical failure (P > 0.99 and P > 0.99). CNB could prevent diagnostic surgery for 92.9 % (13/14) of the patients who showed more than two non-diagnostic results in previous FNA. CONCLUSIONS: CNB can minimise the non-diagnostic results as well as diagnostic surgery in patients with calcified thyroid nodules. Therefore, CNB may be used as a first-line diagnostic tool for calcified thyroid nodules rather than FNA. KEY POINTS: CNB results show the low incidence of technical failure (1.1 %, 3/275). • CNB results show the low non-diagnostic rate (0.7 %, 2/272). There were no significant differences according to the calcification subtype. CNB can prevent unnecessary diagnostic surgery in 92.9 % (13/14).