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Given the growing obesity pandemic, the impact of obesity on outcomes of Extracorporeal Membrane Oxygenation (ECMO) would be increasingly relevant to our daily practise. This meta-analysis aims to evaluate the impact of obesity on ECMO outcomes, integrating the latest evidence. Systematic literature search was conducted from inception until December 2024 on MEDLINE, Embase and the Cochrane Library using the terms 'ECMO', 'obesity', and their related terms. Twenty-eight studies were included from 2013 to 2024, including a total of 74,330 ECMO patients (Mean age 52.84 ± 13.55 years). Obese patients had a similar risk of in-hospital or 30-day mortality when compared to non-obese patients (Risk difference -2%, 95%CI -5% to -1%, I 2  = 85%, p  = 0.25). Subgroup analysis of patients on V-V-ECMO showed a trend towards lower mortality in obese patients which did not reach statistical significance (risk difference -6%, 95%CI -13% to 0%, I 2  = 53%, p  = 0.06). Subgroup analysis of patients on V-A-ECMO showed significantly higher mortality in obese patients (risk difference 5%, 95%CI 1% to 9%, I 2  = 54%, p  = 0.007). Regarding secondary outcomes, obesity had no significant association with major bleeding or thrombotic complications (Risk difference 0%, 95%CI -1% to 2%, I 2  = 15%, p  = 0.63). Obesity was associated with significantly shorter hospital length-of-stay (Mean difference -2.92 days, 95% CI -5.03 to -0.80, I 2  = 74%, p  = 0.007), but had no impact on ECMO duration (Mean difference 0.35 days, 95%CI -0.03 to 0.74, I 2  = 41%, p  = 0.07). In summary, our meta-analysis showed that obesity was a favourable prognostic factor in V-V-ECMO. However, obesity increased mortality in V-A-ECMO patients. The modality of ECMO support should be taken into consideration when evaluating ECMO candidacy in individual obese patients., Competing Interests: Declaration of conflicting interestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Lung cancer is the leading cause of cancer-related mortality worldwide, primarily driven by genetic mutations. The most common genetic alterations implicated in lung cancer include mutations in TP53 , KRAS , KEAP1 , NF1 , EGFR , NRF2 , ATM , ALK , Rb1 , BRAF , MET , and ERBB2 . Targeted therapies have been developed to inhibit cancer growth by focusing on these specific genetic mutations. However, either the mutations are undruggable or the efficacy of these therapies is often compromised over time due to the emergence of drug resistance, which can occur through additional mutations in the targeted protein or alternative growth signaling pathways. In recent years, immunotherapy has emerged as a promising approach to enhance the effectiveness of cancer treatment by leveraging the body's immune system. Notable advancements include immune checkpoint inhibitors, monoclonal antibodies targeting cell surface receptors, antibody-drug conjugates, and bispecific antibodies. This review provides an overview of the mechanisms of FDA-approved immunotherapeutic drugs, offering an updated perspective on the current state and future developments in lung cancer therapy. More importantly, the factors that positively and negatively impact the immunotherapy's efficacy will also be discussed.

Background: Chronic kidney disease (CKD) requires accurate prediction of renal replacement therapy (RRT) initiation risk. This study developed deep learning algorithms (DLAs) to predict RRT risk in CKD patients by incorporating medical history and prescriptions in addition to biochemical investigations., Methods: A multi-centre retrospective cohort study was conducted in three major hospitals in Hong Kong. CKD patients with an eGFR < 30ml/min/1.73m 2 were included. DLAs of various structures were created and trained using patient data. Using a test set, the DLAs' predictive performance was compared to Kidney Failure Risk Equation (KFRE)., Results: DLAs outperformed KFRE in predicting RRT initiation risk (CNN + LSTM + ANN layers ROC-AUC = 0.90; CNN ROC-AUC = 0.91; 4-variable KFRE: ROC-AUC = 0.84; 8-variable KFRE: ROC-AUC = 0.84). DLAs accurately predicted uncoded renal transplants and patients requiring dialysis after 5 years, demonstrating their ability to capture non-linear relationships., Conclusions: DLAs provide accurate predictions of RRT risk in CKD patients, surpassing traditional methods like KFRE. Incorporating medical history and prescriptions improves prediction performance. While our findings suggest that DLAs hold promise for improving patient care and resource allocation in CKD management, further prospective observational studies and randomized controlled trials are necessary to fully understand their impact, particularly regarding DLA interpretability, bias minimization, and overfitting reduction. Overall, our research underscores the emerging role of DLAs as potentially valuable tools in advancing the management of CKD and predicting RRT initiation risk., (© 2024. The Author(s).)

A 60-year-old man intubated for airway protection after smoke inhalation was found to have decompensated hypercapnic respiratory failure. Fiberoptic bronchoscopy revealed obstructive airway slough and pseudomembrane, a manifestation of severe inhalation injury. Veno-venous extracorporeal membrane oxygenation was established for stabilization. The airway casts were removed successfully with periprocedural veno-venous extracorporeal membrane oxygenation support., (© The Author(s) 2023. Published by Oxford University Press on behalf of the European Association for Cardio-Thoracic Surgery.)

Pyroglutamic acidosis (PGA) is an underrecognized entity characterised by raised anion gap metabolic acidosis (RAGMA) and urinary hyper-excretion of pyroglutamic acid. It is frequently associated with chronic acetaminophen (APAP) ingestion. We report the case of a 73-year-old man with invasive pulmonary aspergillosis treated with voriconazole and APAP for analgesia with a cumulative dose of 160 g over 40 days. PGA was suspected as he developed severe RAGMA and common causes were excluded. Diagnosis was confirmed via urinary organic acid analysis which showed significant hyper-excretion of pyroglutamic acid. APAP was discontinued, and N-acetylcysteine (NAC) was administered. His RAGMA rapidly resolved following treatment., Competing Interests: Conflict of interest: None declared., (© 2023 Wincy Wing-Sze Ng et al., published by sciendo.)

Introduction: Obesity is associated with an increased risk for non-communicable diseases. Local studies have shown that 33.1% of healthcare providers (HCPs) are overweight, while 21.1% are obese. Interventions that consist of diet, physical exercise and cognitive behavioural training have been shown to be successful in reducing weight., Method: We designed a weight loss programme for our HCPs named the 'Fit and Trimmed Staff programme, which consisted of 3 months of group education on obesity-related health problems led by a doctor, a pharmacist, a nutritionist and an occupational therapist among HCPs. Monthly individual dietary counselling by a nutritionist was also provided for 6 months. We measured the body weight, body mass index, percentage of body fat, visceral fat and percentage of skeletal muscle of the HCPs before and after the intervention., Results: Forty-five (56.25%) HCPs at Simpang Health Clinic were either overweight or obese; the majority of them were drivers and administrative clerks (100%), followed by health attendants (69.2%) and medical assistants (63.6%). At 6 months post-intervention, there was a trend towards a non-significant reduction in the fat percentage (median=-0.8%, P=0.423). Approximately 42% (n=19) of the HCPs lost weight, while 58% gained weight. Weight loss was observed more commonly in the male HCPs (>50%) than in the female HCPs., Conclusion: A weight loss programme solely consisting of health discussion and nutritional advice is inadequate to induce weight reductions. A multimodal approach may be considered in managing weight among HCPs., (© Academy of Family Physicians of Malaysia.)

The neutrophil-to-lymphocyte ratio (NLR), monocyte-to-lymphocyte ratio (MLR), platelet-to-lymphocyte ratio (PLR), and red cell distribution width (RDW) are emerging biomarkers to predict outcomes in general ward patients. However, their role in the prognostication of critically ill patients with pneumonia is unclear. A total of 216 adult patients were enrolled over 2 years. They were classified into viral and bacterial pneumonia groups, as represented by influenza A virus and Streptococcus pneumoniae, respectively. Demographics, outcomes, and laboratory parameters were analysed. The prognostic power of blood parameters was determined by the respective area under the receiver operating characteristic curve (AUROC). Performance was compared using the APACHE IV score. Discriminant ability in differentiating viral and bacterial aetiologies was examined. Viral and bacterial pneumonia were identified in 111 and 105 patients, respectively. In predicting hospital mortality, the APACHE IV score was the best prognostic score compared with all blood parameters studied (AUC 0.769, 95% CI 0.705-0.833). In classification tree analysis, the most significant predictor of hospital mortality was the APACHE IV score (adjusted P = 0.000, χ 2 = 35.591). Mechanical ventilation was associated with higher hospital mortality in patients with low APACHE IV scores ≤ 70 (adjusted P = 0.014, χ 2 = 5.999). In patients with high APACHE IV scores > 90, age > 78 (adjusted P = 0.007, χ 2 = 11.221) and thrombocytopaenia (platelet count ≤ 128, adjusted P = 0.004, χ 2 = 12.316) were predictive of higher hospital mortality. The APACHE IV score is superior to all blood parameters studied in predicting hospital mortality. The single inflammatory marker with comparable prognostic performance to the APACHE IV score is platelet count at 48 h. However, there is no ideal biomarker for differentiating between viral and bacterial pneumonia., (© 2022. The Author(s).)

Antigenic imprinting, which describes the bias of the antibody response due to previous immune history, can influence vaccine effectiveness. While this phenomenon has been reported for viruses such as influenza, there is little understanding of how prior immune history affects the antibody response to SARS-CoV-2. This study provides evidence for antigenic imprinting through immunization with two Sarbecoviruses , the subgenus that includes SARS-CoV-2. Mice were immunized subsequently with two antigenically distinct Sarbecovirus strains, namely SARS-CoV-1 and SARS-CoV-2. We found that sequential heterologous immunization induced cross-reactive binding antibodies for both viruses and delayed the emergence of neutralizing antibody responses against the booster strain. Our results provide fundamental knowledge about the immune response to Sarbecovirus and important insights into the development of pan-sarbecovirus vaccines and guiding therapeutic interventions.

Nafion, a perfluorosulfonic acid proton exchange membrane (PEM), has been widely used in direct methanol fuel cells (DMFCs) to serve as a proton carrier, methanol barrier, and separator for the anode and cathode. A significant drawback of Nafion in DMFC applications is the high anode-to-cathode methanol fuel permeability that results in over 40% fuel waste. Therefore, the development of a new membrane with lower permeability while retaining the high proton conductivity and other inherent properties of Nafion is greatly desired. In light of these considerations, this paper discusses the research findings on developing Nafion-based membranes for DMFC. Several aspects of the DMFC membrane are also presented, including functional requirements, transport mechanisms, and preparation strategies. More importantly, the effect of the various modification approaches on the performance of the Nafion membrane is highlighted. These include the incorporation of inorganic fillers, carbon nanomaterials, ionic liquids, polymers, or other techniques. The feasibility of these membranes for DMFC applications is discussed critically in terms of transport phenomena-related characteristics such as proton conductivity and methanol permeability. Moreover, the current challenges and future prospects of Nafion-based membranes for DMFC are presented. This paper will serve as a resource for the DMFC research community, with the goal of improving the cost-effectiveness and performance of DMFC membranes.

Background: Shewanella species are emerging pathogens that can cause severe hepatobiliary, skin and soft tissue, gastrointestinal, respiratory infections, and bacteremia. Here we reported the largest case series of infections caused by Shewanella species., Aim: To identify the clinical features and risk factors predisposing to Shewanella infections. To evaluate resistance pattern of Shewanella species and appropriateness of antibiotic use in the study cohort., Methods: Patients admitted to a regional hospital in Hong Kong with Shewanella species infection from April 1, 2010 to December 31, 2020 were included. Demographics, antibiotics, microbiology, and outcomes were retrospectively analyzed., Findings: Over the 10 years, we identified 128 patients with Shewanella species infection. 61.7% were male with a median age of 78 (IQR 65-87). Important underlying diseases included hepatobiliary diseases (63.3%), malignancy (26.6%), chronic kidney disease or end-stage renal failure (25.8%), and diabetes mellitus (22.7%). Hepatobiliary infections (60.4%) were the most common clinical manifestation. Majority (92.2%) were infected with Shewanella algae , while 7.8% were infected with Shewanella putrefaciens . The identified organisms were usually susceptible to ceftazidime (98.7%), gentamicin (97.4%), cefoperazone-sulbactam (93.5%) and ciprofloxacin (90.3%). Imipenem-susceptible strains were only present in 76.6% of isolates., Conclusion: This largest case series suggested that Shewanella infections are commonly associated with underlying comorbidities, especially with hepatobiliary diseases and malignancy. Although Shewanella species remained largely susceptible to third and fourth generation cephalosporins and aminoglycosides, carbapenem resistance has been on a significant rise., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Ng, Shum, To and Sridhar.)

Introduction: Despite the ever-growing number of community onset (CO) Pseudomonas aeruginosa (P. aeruginosa) bacteraemia, there is a dearth of district hospital-based research examining this significant infection, which is associated with high mortality. The objectives of this study were as following: (1) to determine the risk factors of CO P. aeruginosa bacteraemia, (2) to compare the 30-day mortality rate between P. aeruginosa and Escherichia coli bacteraemia and (3) to identify the predictors of 30-day mortality for CO gram negative bacteraemia., Methods: This is a retrospective case control study in Hospital Seri Manjung and Hospital Teluk Intan, Perak, Malaysia. P. aeruginosa bacteraemia cases that occurred between 1st January 2015 to 31st December 2019 were included, whilst E. coli bacteraemia cases that occurred within the same period were recruited successively until 1:2 case control ratio was achieved. Subjects below 12-year-old and those with polymicrobial bacteraemia were excluded. Demographic, clinical and treatment data were collected using pre-tested data collection forms by trained investigators., Results: A total of 61 patients with P. aeruginosa bacteraemia and 122 patients with E. coli bacteraemia were included. Recent admission in the earlier three months, regular haemodialysis, immunosuppressive therapy in the past 30 days, chronic wound/pressure sore at presentation and indwelling urinary catheter at presentation were identified as independent predictors of CO pseudomonal bacteraemia. Whilst older age was identified as a negative predictor of CO Pseudomonal bacteraemia (all p<0.05). The 30-day mortality rate was 34.4% in subjects with P. aeruginosa bacteraemia and 27.0% in those with E. coli bacteraemia (p=0.302). Predictors of 30-day mortality for community onset gram negative bacteraemia were as follow: older age, underlying solid tumours, neutropaenia at presentation, in-patient mechanical ventilation, and inpatient nasogastric tube insertion. Unexpectedly, receipt of inappropriate empirical antibiotics which was switched later (delayed and non-delayed switching) was identified as the negative predictors of mortality (all p<0.05)., Conclusion: It is prudent to restrict the usage of empirical anti-pseudomonal antibiotics among individuals at risk as liberal usage of broad-spectrum antibiotics engenders emergence of drug resistant organism, particularly in district setting where community onset pseudomonal bacteraemia remains scarce. Subjects with elevated risk of mortality should receive early escalation of care as per sepsis management guidelines.

The increasing numbers of infected cases of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) poses serious threats to public health and the global economy. Most SARS-CoV-2 neutralizing antibodies target the receptor binding domain (RBD) and some the N-terminal domain (NTD) of the spike protein, which is the major antigen of SARS-CoV-2. While the antibody response to RBD has been extensively characterized, the antigenicity and immunogenicity of the NTD protein are less well studied. Using 227 plasma samples from COVID-19 patients, we showed that SARS-CoV-2 NTD-specific antibodies could be induced during infection. As compared to the results of SARS-CoV-2 RBD, the serological response of SARS-CoV-2 NTD is less cross-reactive with SARS-CoV, a pandemic strain that was identified in 2003. Furthermore, neutralizing antibodies are rarely elicited in a mice model when NTD is used as an immunogen. We subsequently demonstrate that NTD has an altered antigenicity when expressed alone. Overall, our results suggest that while NTD offers a supplementary strategy for serology testing, it may not be suitable as an immunogen for vaccine development., (© 2021 Wiley-VCH GmbH.)

The sequencing of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) in patients with EGFR mutation-positive (EGFRm+) non-small cell lung cancer (NSCLC) remains a matter of controversy. This cohort study analyzed the overall survival (OS) and progression-free survival (PFS) of afatinib compared with erlotinib and gefitinib first-line. EGFRm+, advanced NSCLC patients treated with either afatinib, erlotinib or gefitinib were retrospectively analyzed. A total of 107 patients were included. There was no statistically significant difference in PFS among the 3 groups. In the ≥ 60 years age group, the afatinib group had longer survival compared to the gefitinib group (p = 0.01). Median OS were 19.1, 22.9, and 35.6 months for gefitinib, erlotinib, and afatinib groups, respectively, with statistical significance between the gefitinib and afatinib groups (p = 0.009). Patients on afatinib also had longer median OS than erlotinib and gefitinib pooled together (35.5 versus 21.4 months; hazard ratio = 0.54, p = 0.016), despite similar median PFS. In conclusion, afatinib is a better choice compared to gefitinib or erlotinib for EGFRm+ patients. The OS obtained with afatinib is just 3 months shorter than osimertinib in the FLAURA trial. Direct comparison studies with osimertinib are still needed to determine optimal sequencing., Competing Interests: The authors have declared that no competing interests exist.

Individuals with the 2019 coronavirus disease (COVID-19) show varying severity of the disease, ranging from asymptomatic to requiring intensive care. Although monoclonal antibodies specific to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been identified, we still lack an understanding of the overall landscape of B cell receptor (BCR) repertoires in individuals with COVID-19. We use high-throughput sequencing of bulk and plasma B cells collected at multiple time points during infection to characterize signatures of the B cell response to SARS-CoV-2 in 19 individuals. Using principled statistical approaches, we associate differential features of BCRs with different disease severity. We identify 38 significantly expanded clonal lineages shared among individuals as candidates for responses specific to SARS-CoV-2. Using single-cell sequencing, we verify the reactivity of BCRs shared among individuals to SARS-CoV-2 epitopes. Moreover, we identify the natural emergence of a BCR with cross-reactivity to SARS-CoV-1 and SARS-CoV-2 in some individuals. Our results provide insights important for development of rational therapies and vaccines against COVID-19., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

COVID-19 patients show varying severity of the disease ranging from asymptomatic to requiring intensive care. Although a number of SARS-CoV-2 specific monoclonal antibodies have been identified, we still lack an understanding of the overall landscape of B-cell receptor (BCR) repertoires in COVID-19 patients. Here, we used high-throughput sequencing of bulk and plasma B-cells collected over multiple time points during infection to characterize signatures of B-cell response to SARS-CoV-2 in 19 patients. Using principled statistical approaches, we determined differential features of BCRs associated with different disease severity. We identified 38 significantly expanded clonal lineages shared among patients as candidates for specific responses to SARS-CoV-2. Using single-cell sequencing, we verified reactivity of BCRs shared among individuals to SARS-CoV-2 epitopes. Moreover, we identified natural emergence of a BCR with cross-reactivity to SARS-CoV-1 and SARS-CoV-2 in a number of patients. Our results provide important insights for development of rational therapies and vaccines against COVID-19.

A previously well 21-year-old girl presented to Hospital Teluk Intan, Perak, Malaysia with a short history of fever, vomiting and altered sensorium. She was diagnosed with dengue encephalitis as her dengue NS-1 antigen was positive and her cerebrospinal fluid (CSF) dengue polymerase chain reaction (PCR) was positive with serotype DENV-2. She also had severe hyponatremia due to Syndrome of Inappropriate Antidiuretic Hormone Secretion (SIADH) which caused an episode of seizure. She recovered well with supportive management. SIADH and dengue encephalitis should be considered as one of the differential diagnosis in patients presenting with fever and altered sensorium especially in dengue endemic countries like Malaysia.

Background: Therapeutic drug monitoring of tumor necrosis factor inhibitors, such as adalimumab (ADM), is increasingly being performed for the management of autoimmune diseases. However, there can be significant variation in drug and antibody concentrations obtained by different assay methods. The aim of this study was to compare the performance of 4 enzyme-linked immunosorbent assay (ELISA) kits for measuring ADM and anti-ADM antibodies., Method: Dilutions of ADM or anti-ADM spiked sera were assessed for recovery rate and precision using the following 4 kits: LISA-Tracker (Theradiag, Croissy-Beaubourg, France), Promonitor (Grifols, Barcelona, Spain), Ridascreen (R-Biopharm, Darmstadt, Germany), and Shikari (Matriks Biotek, Gölbaşi/Ankara Turkey). Interference samples were also assessed., Results: At the therapeutic concentration, ADM detection was comparable among the 4 ELISA kits. Lisa-Tracker and Shikari kits produced low-range false positive results in normal sera. Infliximab and etanercept caused false positives in Lisa-Tracker and Shikari kits. Anti-ADM antibody ELISA kits performed differently with spiked samples because of different measuring units and ranges. Ridascreen and Shikari kits were dose responsive across the entire standard curve and correlated well with each other (r = 0.997). Cross reactivity was observed in rheumatoid factor positive sera tested on the Promonitor anti-ADM kit., Conclusions: All ADM kits tested were dose responsive within the therapeutic range and correlated well. The significance of observed low-range false positives and cross reactivity with infliximab in LISA-Tracker and Shikari kits is dependent on the indications received for testing in the laboratory. Anti-ADM ELISA kits produced varied results for spiked sera; however, they showed good precision. Inter-kit variability suggested that anti-ADM levels should be compared only when using the same method.

The World Health Organization has declared the ongoing outbreak of COVID-19, which is caused by a novel coronavirus SARS-CoV-2, a pandemic. There is currently a lack of knowledge about the antibody response elicited from SARS-CoV-2 infection. One major immunological question concerns antigenic differences between SARS-CoV-2 and SARS-CoV. We address this question by analyzing plasma from patients infected by SARS-CoV-2 or SARS-CoV and from infected or immunized mice. Our results show that, although cross-reactivity in antibody binding to the spike protein is common, cross-neutralization of the live viruses may be rare, indicating the presence of a non-neutralizing antibody response to conserved epitopes in the spike. Whether such low or non-neutralizing antibody response leads to antibody-dependent disease enhancement needs to be addressed in the future. Overall, this study not only addresses a fundamental question regarding antigenicity differences between SARS-CoV-2 and SARS-CoV but also has implications for immunogen design and vaccine development., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

The training of a multilayer perceptron neural network (MLPNN) concerns the selection of its architecture and the connection weights via the minimization of both the training error and a penalty term. Different penalty terms have been proposed to control the smoothness of the MLPNN for better generalization capability. However, controlling its smoothness using, for instance, the norm of weights or the Vapnik-Chervonenkis dimension cannot distinguish individual MLPNNs with the same number of free parameters or the same norm. In this paper, to enhance generalization capabilities, we propose a stochastic sensitivity measure (ST-SM) to realize a new penalty term for MLPNN training. The ST-SM determines the expectation of the squared output differences between the training samples and the unseen samples located within their Q -neighborhoods for a given MLPNN. It provides a direct measurement of the MLPNNs output fluctuations, i.e., smoothness. We adopt a two-phase Pareto-based multiobjective training algorithm for minimizing both the training error and the ST-SM as biobjective functions. Experiments on 20 UCI data sets show that the MLPNNs trained by the proposed algorithm yield better accuracies on testing data than several recent and classical MLPNN training methods.

Quality of Life (QOL) is reported to be lower for patients with depression than the general population. This study aims to investigate the mediational effects of protective resource factors (PSFs), such as depressive symptom management ability, self-efficacy, social support and problem-focused coping act, in the relationship between dysfunctional attitudes and QOL. It is hypothesized that these PSFs have different mediating strengths. Self-report questionnaires which aimed to determine the influences of these PSFs through a multiple mediation framework were completed by 80 depressed adult outpatients from the National University Hospital of Singapore. PSFs have different influence on mental and physical QOL. Depressive symptom management ability is the most important PSF mediating both domains and better problem-focused coping abilities demonstrate improvement in the physical domain of QOL. Self-efficacy and social support are shown to be non-significant mediators. The results suggest for future effective interventions to focus primarily on improving depression patients' symptom management ability and problem-focused coping skills to raise their life quality. Furthermore, findings from this study have implications on the future investigation of QOL as a unitary construct., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Undersampling is a widely adopted method to deal with imbalance pattern classification problems. Current methods mainly depend on either random resampling on the majority class or resampling at the decision boundary. Random-based undersampling fails to take into consideration informative samples in the data while resampling at the decision boundary is sensitive to class overlapping. Both techniques ignore the distribution information of the training dataset. In this paper, we propose a diversified sensitivity-based undersampling method. Samples of the majority class are clustered to capture the distribution information and enhance the diversity of the resampling. A stochastic sensitivity measure is applied to select samples from both clusters of the majority class and the minority class. By iteratively clustering and sampling, a balanced set of samples yielding high classifier sensitivity is selected. The proposed method yields a good generalization capability for 14 UCI datasets.

Although illness perception has been shown to be associated with illness outcomes in various chronic physical diseases, the association of illness perception and rumination are not well elucidated in mental disorders. This study aims to investigate the mediational effects of adaptive and maladaptive rumination in the relationship between illness perception and negative emotions (depression, anxiety and stress) in male and female patients (N=110) suffering from depressive disorders. The results showed that maladaptive rumination mediated the relationship between illness perception and negative emotions in both male and female depressive patients. However, no mediating effects of adaptive rumination were found in the relationship between illness perception and negative emotion. Maladaptive rumination mediated the relationship between perceived identity, chronicity of illness, consequences of illness and emotional representation of illness and negative emotions in males. It also mediated the relationship between perceived identity and emotional representation of illness and negative emotions in females. The results, possible clinical implications and limitations of this study are also discussed., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Background: An important step in characterising the function of a gene is identifying the cells in which it is expressed. Traditional methods to determine this include in situ hybridisation, gene promoter-reporter fusions or cell isolation/purification techniques followed by quantitative PCR. These methods, although frequently used, can have limitations including their time-consuming nature, limited specificity, reliance upon well-annotated promoters, high cost, and the need for specialized equipment. In situ PCR is a relatively simple and rapid method that involves the amplification of specific mRNA directly within plant tissue whilst incorporating labelled nucleotides that are subsequently detected by immunohistochemistry. Another notable advantage of this technique is that it can be used on plants that are not easily genetically transformed., Results: An optimised workflow for in-tube and on-slide in situ PCR is presented that has been evaluated using multiple plant species and tissue types. The protocol includes optimised methods for: (i) fixing, embedding, and sectioning of plant tissue; (ii) DNase treatment; (iii) in situ RT-PCR with the incorporation of DIG-labelled nucleotides; (iv) signal detection using colourimetric alkaline phosphatase substrates; and (v) mounting and microscopy. We also provide advice on troubleshooting and the limitations of using fluorescence as an alternative detection method. Using our protocol, reliable results can be obtained within two days from harvesting plant material. This method requires limited specialized equipment and can be adopted by any laboratory with a vibratome (vibrating blade microtome), a standard thermocycler, and a microscope. We show that the technique can be used to localise gene expression with cell-specific resolution., Conclusions: The in situ PCR method presented here is highly sensitive and specific. It reliably identifies the cellular expression pattern of even highly homologous and low abundance transcripts within target tissues, and can be completed within two days of harvesting tissue. As such, it has considerable advantages over other methods, especially in terms of time and cost. We recommend its adoption as the standard laboratory technique of choice for demonstrating the cellular expression pattern of a gene of interest.

Objectives: This study aimed to promote quality end-of-life (EOL) care for nursing home residents, through the establishment of advance care plan (ACP) and introduction of a new care pathway. This pathway bypassed the emergency room (ER) and acute medical wards by facilitating direct clinical admission to an extended-care facility., Design: An audit on a new clinical initiative that entailed the Community Geriatrics Outreach Service, ER, acute medical wards, and an extended-care facility during winter months in Hong Kong., Methods: The participants were older nursing home residents enrolled in an EOL program. We monitored the ratio of clinical to emergency admissions, ACP compliance rate, average length of stay (ALOS) in both acute hospital and an extended-care facility, and mortality rates., Results: A total of 76 patients were hospitalized from January to March 2013. Of them, 30 (39%) were directly admitted to the extended-care facility, either through the liaison of Community Geriatrics Outreach Service (group A, 19/76, 25%) or transferred from the ER (group B, 11/76, 14%). The remaining 46 patients (group C, 61%) were admitted via the ER to acute medical wards following the usual pathway, followed by transfer to an extended-care facility if indicated. The ACP compliance rate was nearly 100%. In the extended-care unit, groups A and C had similar ALOS of 11.8 and 11.1 days, respectively, whereas group B had a shorter stay of 7.6 days. The ALOS of group C in acute medical wards was 3.5 days. The in-hospital mortality rates were comparable in groups A and C of 26% and 28%, respectively, whereas group B had a lower mortality rate of 18%., Conclusions: Nearly 40% of EOL patients could be managed entirely in an extended-care setting without compromising the quality of care and survival. A greater number of patients may benefit from the EOL program by improving the collaboration between community outreach services and ER; and extending hours for direct clinical admission to an extended-care facility., (Copyright © 2014 American Medical Directors Association, Inc. Published by Elsevier Inc. All rights reserved.)

Leishmania parasites alternate between extracellular promastigote stages in the insect vector and an obligate intracellular amastigote stage that proliferates within the phagolysosomal compartment of macrophages in the mammalian host. Most enzymes involved in Leishmania central carbon metabolism are constitutively expressed and stage-specific changes in energy metabolism remain poorly defined. Using (13)C-stable isotope resolved metabolomics and (2)H2O labelling, we show that amastigote differentiation is associated with reduction in growth rate and induction of a distinct stringent metabolic state. This state is characterized by a global decrease in the uptake and utilization of glucose and amino acids, a reduced secretion of organic acids and increased fatty acid β-oxidation. Isotopomer analysis showed that catabolism of hexose and fatty acids provide C4 dicarboxylic acids (succinate/malate) and acetyl-CoA for the synthesis of glutamate via a compartmentalized mitochondrial tricarboxylic acid (TCA) cycle. In vitro cultivated and intracellular amastigotes are acutely sensitive to inhibitors of mitochondrial aconitase and glutamine synthetase, indicating that these anabolic pathways are essential for intracellular growth and virulence. Lesion-derived amastigotes exhibit a similar metabolism to in vitro differentiated amastigotes, indicating that this stringent response is coupled to differentiation signals rather than exogenous nutrient levels. Induction of a stringent metabolic response may facilitate amastigote survival in a nutrient-poor intracellular niche and underlie the increased dependence of this stage on hexose and mitochondrial metabolism.

When reconstructing the three-dimensional (3D) object height profile using the fringe projection profilometry (FPP) technique, the light intensity reflected from the object surface can yield abruptly changing bias in the captured fringe image, which leads to severe reconstruction error. The traditional approach tries to remove the bias by suppressing the zero spectrum of the fringe image. It is based on the assumption that the aliasing between the frequency spectrum of the bias, which is around the zero frequency, and the frequency spectrum of the fringe is negligible. This, however, is not the case in practice. In this paper, we propose a novel (to our knowledge) technique to eliminate the bias in the fringe image using the dual-tree complex wavelet transform (DT-CWT). The new approach successfully identifies the features of bias, fringe, and noise in the DT-CWT domain, which allows the bias to be effectively extracted from a noisy fringe image. Experimental results show that the proposed algorithm is superior to the traditional methods and facilitates accurate reconstruction of objects' 3D models.

The advance of anti-tumour necrosis factor (TNF) therapy had dramatically changed the treatment algorithm of inflammatory bowel disease (IBD). This had significantly improved the quality of life for patients with Crohn's disease (CD) and ulcerative colitis (UC).(1) However, side-effects of anti-TNF treatment were unavoidable with paradoxical inflammation (for example leucocytoclastic vasculitis and psoriasis) being well-known phenomena of anti-TNF therapy.(2) We report a case of infliximab induced cutaneous sarcoidosis in a patient with ulcerative colitis and review the literature., (Crown Copyright © 2012. Published by Elsevier B.V. All rights reserved.)

Leishmania parasites proliferate within nutritionally complex niches in their sandfly vector and mammalian hosts. However, the extent to which these parasites utilize different carbon sources remains poorly defined. In this study, we have followed the incorporation of various (13)C-labeled carbon sources into the intracellular and secreted metabolites of Leishmania mexicana promastigotes using gas chromatography-mass spectrometry and (13)C NMR. [U-(13)C]Glucose was rapidly incorporated into intermediates in glycolysis, the pentose phosphate pathway, and the cytoplasmic carbohydrate reserve material, mannogen. Enzymes involved in the upper glycolytic pathway are sequestered within glycosomes, and the ATP and NAD(+) consumed by these reactions were primarily regenerated by the fermentation of phosphoenolpyruvate to succinate (glycosomal succinate fermentation). The initiating enzyme in this pathway, phosphoenolpyruvate carboxykinase, was exclusively localized to the glycosome. Although some of the glycosomal succinate was secreted, most of the C4 dicarboxylic acids generated during succinate fermentation were further catabolized in the TCA cycle. A high rate of TCA cycle anaplerosis was further suggested by measurement of [U-(13)C]aspartate and [U-(13)C]alanine uptake and catabolism. TCA cycle anaplerosis is apparently needed to sustain glutamate production under standard culture conditions. Specifically, inhibition of mitochondrial aconitase with sodium fluoroacetate resulted in the rapid depletion of intracellular glutamate pools and growth arrest. Addition of high concentrations of exogenous glutamate alleviated this growth arrest. These findings suggest that glycosomal and mitochondrial metabolism in Leishmania promastigotes is tightly coupled and that, in contrast to the situation in some other trypanosomatid parasites, the TCA cycle has crucial anabolic functions.

In optical phase shift profilometry (PSP), parallel fringe patterns are projected onto an object and the deformed fringes are captured using a digital camera. It is of particular interest in real time three-dimensional (3D) modeling applications because it enables 3D reconstruction using just a few image captures. When using this approach in a real life environment, however, the noise in the captured images can greatly affect the quality of the reconstructed 3D model. In this paper, a new image enhancement algorithm based on the oriented two-dimenional dual-tree complex wavelet transform (DT-CWT) is proposed for denoising the captured fringe images. The proposed algorithm makes use of the special analytic property of DT-CWT to obtain a sparse representation of the fringe image. Based on the sparse representation, a new iterative regularization procedure is applied for enhancing the noisy fringe image. The new approach introduces an additional preprocessing step to improve the initial guess of the iterative algorithm. Compared with the traditional image enhancement techniques, the proposed algorithm achieves a further improvement of 7.2 dB on average in the signal-to-noise ratio (SNR). When applying the proposed algorithm to optical PSP, the new approach enables the reconstruction of 3D models with improved accuracy from 6 to 20 dB in the SNR over the traditional approaches if the fringe images are noisy., (© 2011 Optical Society of America)

Introduction: Central serous chorioretinopathy is characterized by serous neurosensory detachment of the macula and it usually resolves spontaneously with good visual prognosis. In some patients, however, the serous retinal detachment might be very extensive and can result in bullous exudative retinal detachment. We evaluated the use of half-dose verteporfin photodynamic therapy for the treatment of bullous retinal detachment in idiopathic central serous chorioretinopathy., Case Presentation: We report the case of a 51-year-old Chinese man who presented with blurred vision in his right eye and superior visual field defect due to bullous variant of central serous chorioretinopathy. No improvement in vision and retinal detachment was noted after three months of observation and a short course of oral acetazolamide. He was then treated with half-dose verteporfin photodynamic therapy and his visual acuity improved from 20/70 to 20/25 within one month of treatment. Three months after photodynamic therapy, there was complete resolution of sub-retinal fluid and bullous retinal detachment. No recurrence of central serous chorioretinopathy was noted in three years of follow-up., Conclusion: We report the beneficial effect of photodynamic therapy with half-dose verteporfin as a treatment option for bullous retinal detachment caused by central serous chorioretinopathy.

Purpose: To characterize various properties of a physiologically-relevant artificial tear solution (ATS) containing a range of tear film components within a complex salt solution, and to measure contact lens parameters and lipid deposition of a variety of contact lens materials after incubation in this ATS., Methods: A complex ATS was developed that contains a range of salts, proteins, lipids, mucin, and other tear film constituents in tear-film relevant concentrations. This ATS was tested to confirm that its pH, osmolality, surface tension, and homogeneity are similar to human tears and remain so throughout the material incubation process, for up to 4 weeks. To confirm that silicone hydrogel and conventional hydrogel contact lens materials do not alter in physical characteristics beyond what is allowed by the International Organization for Standardization (ISO) 18369-2. The diameter, center thickness, and calculated base curve were measured for five different lens materials directly out of the blister pack, after a rinse in saline and then following a two week incubation in the modified ATS. To test the ATS and the effect of its composition on lipid deposition, two lens materials were incubated in the ATS and a modified version for several time points. Both ATS solutions contained trace amounts of carbon-14 cholesterol and phosphatidylcholine, such that deposition of these specific lipids could be quantified using standard methods., Results: This ATS is a complex mixture that remains stable at physiologically relevant pH (7.3-7.6), osmolality (304-306 mmol/kg), surface tension (40-46 dynes/cm) and homogeneity over an incubation period of three weeks or more. The physical parameters of the lenses tested showed no changes beyond that allowed by the ISO guidelines. Incubations with the ATS found that balafilcon A lenses deposit significantly more cholesterol and phosphatidylcholine than omafilcon A lenses (p<0.05) and that removing lactoferrin and immunoglobulin G from the ATS can significantly decrease the mass of lipid deposited., Conclusions: This paper describes a novel complex artificial tear solution specially designed for in-vial incubation of contact lens materials. This solution was stable and did not adversely affect the physical parameters of the soft contact lenses incubated within it and showed that lipid deposition was responsive to changes in ATS composition.

Objective: To compare the survival between patients with synchronous and metachronous colorectal liver metastases after hepatectomy with new generation of peri-operative chemotherapy., Methods: From October 2002 to January 2008, patients receiving hepatectomy for synchronous or metachronous colorectal liver metastasis were studied retrospectively., Results: Fifty-five patients (synchronous group=35, metachronous group=20) underwent hepatectomy for colorectal liver metastases. Besides younger age with male predominance, patients in the synchronous group had more tumour multinodularity and bilobe liver involvement. They had received less hepatic curative hepatectomy (81.1% vs. 100%) with a higher rate of peri-operative chemotherapy (91.4% vs. 50%) and postoperative morbidity (25.7% vs. 0%). However both groups had no statistical significant difference in median overall survival (OS) and disease free survival (DFS). Inferior OS and DFS were observed in the synchronous group for patients who had no peri-operative chemotherapy or those showing poor response to chemotherapy. The most favourable OS is observed in both groups after performing globally curative hepatectomy., Conclusion: Synchronous colorectal liver metastasis is not a poor prognostic factor for survival when compared with the metachronous metastasis. Globally curative hepatectomy in combination of new generation of chemotherapy is recommended for the management of resectable colorectal liver metastasis.

Gastroduodenal artery pseudoaneurysm is a rare but life threatening complication of pancreatitis. Diagnosis and management of it remain challenging. Surgical treatment was associated with a high mortality. Percutaneous transarterial embolization of bleeding artery has recently been advocated as a definitive therapy and can be attempted as the initial measure to control bleeding. We herein report a case of chronic pancreatitis presented with ruptured pseudoaneurysm of gastroduodenal artery which was successfully controlled with transarterial embolisation.

Substance abuse is a major health and social problem among Hong Kong youth and ketamine is the drug most commonly abused. Ketamine abuse is associated with a series of side-effects that include hallucination, nausea, vomiting, elevation of blood pressure, and urinary bladder dysfunction. Here we report three cases of ketamine abuse in which the abusers presented with recurrent epigastric pain and dilated common bile ducts that mimicked choledochal cysts on imaging. The dilated biliary tree may occur more frequently than was once assumed.

Background: Radiofrequency ablation (RFA) has been widely applied for the treatment of hepatocellular carcinoma and liver metastases. The reported mortality and morbidity rates are low. The aim of this study is to evaluate the safety and efficacy of RFA, and compare the results performed percutaneously versus surgically., Patients and Methods: From 2003 to 2006, 79 patients with hepatic malignancies (59 hepatocellular carcinoma, 20 liver metastases) with a total of 110 lesions underwent RFA in our centre. Postablation assessment by CT scan was performed in all patients at 1-, 3- and 6-month intervals. Post-procedural complications, recurrence and survival were analysed., Results: The patients' mean age was 60.0 years. In 46.8% of cases, we used a percutaneous approach; in 53.2% of cases, a surgical approach (8.9% laparoscopic; 44.3% open) was used if percutaneous approach was not feasible. The mean tumour size was 2.4 cm. Within the surgical group, 69% of patients received concomitant operative procedures such as cholecystectomy and hepatectomy. No treatment-related mortality was observed. Immediate complications occurred in five patients (6.3%), including gastric serosal burn (n = 1), ground pad superficial skin burn (n = 1), intra-abdominal bleeding (n = 2) and pleural effusion (n = 1). All patients except one attended subsequent follow-up, with a mean period of 16 months. Ablation was considered complete in 82.3% of patients (percutaneous approach 81.1%, surgical approach 83.3%, p = 0.72). Intrahepatic recurrence was observed in 52.3%, the majority of them located away from the RFA site. Extrahepatic recurrences were observed in 16.9% (percutaneous approach 16.7%, surgical approach 17.1%, p = 0.76). The overall one- and two-year survival rate was 93.7% and 74.4% respectively, and no statistically significant difference was observed between the two approaches., Conclusion: RFA is a safe and effective procedure for treating patients with malignant liver tumours. No difference in short term outcomes was observed between percutaneous and surgical approaches. A more prolonged follow-up study is required to assess longer-term outcomes.

Soy consumption is associated with a lower risk of atherosclerotic disease in the oriental population. Genistein is a soy isoflavone bearing estrogenic properties. Previous experiments in our laboratory demonstrated the potentiation of endothelium-independent relaxation of coronary artery by both estrogen and genistein. The potentiating effects of both estrogen and genistein were mediated through the cAMP-signaling pathway. We hypothesize that genistein could enhance protein kinase A (PKA) activity in porcine coronary artery smooth muscle, thereby offering a mechanism for the potentiation of vascular relaxation by genistein. In our study, a high concentration of genistein (10(-4.5) M) significantly increased PKA activity in porcine coronary artery rings. While genistein at 10(-5.5) M and forskolin at 10(-7) M had no effect on PKA activity, the combination of the two compounds at the prescribed concentrations caused a significant increase in PKA activity. The increase in PKA activity by genistein was abolished by SQ 22536 (adenylate cyclase blocker), but not by NF 449 (Gs protein blocker) or ICI 182780 (estrogen receptor antagonist). Our results suggest that the action of genistein is mediated via adenylate cyclase, but does not appear to involve Gs protein or ICI 182780-sensitive estrogen receptor.

Objective: To evaluate the benefits of laparoscopic versus open resection of liver tumours., Design: Case control study., Setting: Tertiary teaching hospital, Hong Kong., Patients: Data from 25 patients who underwent laparoscopic resections for liver tumours from 2003 to 2006 were compared to a retrospective series of 25 patients who underwent open hepatectomy in a pair-matched design., Main Outcome Measures: Duration of operation, operative morbidity and mortality, blood loss, tumour resection margin, analgesics usage, days to return to an oral diet, duration of postoperative hospital stay, and survival of patients with malignancy., Results: The demographic data and the tumour characteristics were comparable in the two patient groups, as were mortality (0% in both groups) and morbidity rates (4% in both groups). Two (8%) of the patients having laparoscopic resections were converted to open surgery. There was no statistically significant difference between the two groups in terms of operating time or resection margins. However, the laparoscopically treated patients experienced significantly less blood loss (median, 100 vs 250 mL), had shorter hospital stays (median, 4 vs 7 days), were prescribed less analgesia (median morphine dosage, 0.16 vs 0.83 mg per kg body weight), and resumed oral diet earlier (median, 1 vs 2 days). For patients with malignant tumours, there was no significant difference between the two groups in terms of actuarial and disease-free survival., Conclusion: Compared to open hepatectomy, in selected patients laparoscopic liver resection delivers the benefits of decreased blood loss, shorter hospital stay, lesser requirement for analgesics, and an earlier return to an oral diet, without evidence of compromised oncological clearance.

Objectives: To review the reliability of radiological diagnosis and need of regular scans for giant liver haemangioma, in terms of long-term outcome and management options., Design: Retrospective study., Setting: Division of Hepato-biliary and Pancreatic Surgery, Prince of Wales Hospital, Hong Kong., Patients: Patients with giant liver haemangioma noted on initial imaging from February 1996 to July 2006., Main Outcome Measures: Patient demographics, clinical assessments, management, and outcomes., Results: There were 42 female and 22 male patients with a median age of 49 (range, 27-84) years with a suspected haemangioma. The median maximal diameter of the lesions was 5.5 cm (range, 4.0-20.3 cm). They were first detected by ultrasonography (n=45), contrast-enhanced computed tomographic scan (n=18), or magnetic resonance imaging (n=1). Besides regular follow-up scans, 22 patients were investigated further to confirm the diagnosis/exclude malignancy. Finally, 63 patients had a haemangioma and one had a hepatocellular carcinoma. Regarding the patients with haemangiomas, two were operated on for relief of pain and the rest were managed conservatively. The median duration of follow-up was 34 months. Most (54%) of the patients were asymptomatic, but in 17% the haemangioma enlarged to exceed its original size by more than 20%. There were no haemangioma-associated complications., Conclusions: Majority of patients having giant liver haemangioma are asymptomatic and do not suffer complications. If the diagnosis is uncertain, selective further investigations may be necessary. Lesions with a confirmed diagnosis tend to remain static in size; performing regular scans for asymptomatic giant liver haemangiomas may not be necessary.

The generalization error bounds found by current error models using the number of effective parameters of a classifier and the number of training samples are usually very loose. These bounds are intended for the entire input space. However, support vector machine (SVM), radial basis function neural network (RBFNN), and multilayer perceptron neural network (MLPNN) are local learning machines for solving problems and treat unseen samples near the training samples to be more important. In this paper, we propose a localized generalization error model which bounds from above the generalization error within a neighborhood of the training samples using stochastic sensitivity measure. It is then used to develop an architecture selection technique for a classifier with maximal coverage of unseen samples by specifying a generalization error threshold. Experiments using 17 University of California at Irvine (UCI) data sets show that, in comparison with cross validation (CV), sequential learning, and two other ad hoc methods, our technique consistently yields the best testing classification accuracy with fewer hidden neurons and less training time.

As a safer approach to right hepatectomy, Belghiti et al. (J Am Coll Surg 193:109-11, 2001) described a liver-hanging maneuver. However, this procedure is performed blind, with the risks of damaging the small retrohepatic veins and consequential bleeding. To overcome this problem, we modified the procedure so that, instead of performing blind dissection using a long vascular clamp, we use a flexible choledochoscope to dissect the retrohepatic space filled by loose alveolar tissue anterior to the inferior vena cava (IVC). The avascular path is identified by a combination of saline irrigation and gentle movement of the tip of the choledochoscope. Cotton tape can then be passed around the liver parenchyma to elevate the liver away from the anterior surface of the IVC. This modification of Belghiti's liver-hanging maneuver allows direct vision along the plane anterior to the IVC, thus avoiding injury to the retrohepatic veins.

Osteoblastic differentiation is an essential part of bone formation. Dimethyl sulfoxide (DMSO) is a water miscible solvent that is used extensively for receptor ligands in osteoblast studies. However, little is known about its effects on osteoblastogenic precursor cells. In this study, we have used a murine preosteoblast cell line MC3T3-E1 cells to demonstrate that DMSO effectively induces osteoblastic differentiation of MC3T3-E1 cells via the activation of Runx2 and osterix and is dependent upon the protein kinase C (PKC) pathways. We further demonstrated that prolonged activation of PKC pathways is sufficient to induce osteoblastic differentiation, possibly via the activation of PKD/PKCmu.

This study examined 58 sexually abused children referred for clinical psychological services in 1999 in Hong Kong and compared the characteristics and disclosure patterns between those with multiple incidents of abuse and those with single incident. The former group was more likely to have been abused by a family member, took a longer time to disclose, and was more often abused in contexts where the abusers had control and power. Over 40% of subjects also reported presence of other persons during abuse. Myths and misconceptions among professionals are highlighted. Implications on future service development and training needs in child protection are discussed.

A new verotoxin (VT) variant, designated vt2g, was identified from a bovine strain of verocytotoxigenic Escherichia coli (VTEC) serotype O2:H25. When vt2g was aligned with published sequences of vt2 and vt variants, it exhibited the highest DNA sequence homology with vt2 and vt2c. However, vt2g was not detected by vt2-specific primers and probes, although it was partially neutralized by an antiserum to the VT2A subunit. VT2g was cytotoxic for Vero and HeLa cells and was not activated by mouse intestinal mucus. The vt2g gene was detected in 3 of 409 (0.7%) bovine VTEC strains, including serotypes O2:H25, O2:H45 and Ont:H-.

Axin is a multidomain protein that plays a critical role in Wnt signaling, serving as a scaffold for down-regulation of beta-catenin. It also activates the JNK mitogen-activated protein kinase by binding to MEKK1. However, it is intriguing that Axin requires several additional elements for JNK activation, including a requirement for homodimerization, sumoylation at the extreme C-terminal sites, and a region in the protein phosphatase 2A-binding domain. In our present study, we have shown that another MEKK family member, MEKK4, also binds to Axin in vivo and mediates Axin-induced JNK activation. Surprisingly MEKK4 binds to a region distinct from the MEKK1-binding site. Dominant negative mutant of MEKK4 attenuates the JNK activation by Axin. Activation of JNK by Axin in MEKK1-/- mouse embryonic fibroblast cells supports the idea that another MEKK can mediate Axin-induced JNK activation. Expression of specific small interfering RNA against MEKK4 effectively attenuates JNK activation by the MEKK1 binding-defective Axin mutant in 293T cells and inhibits JNK activation by wild-type Axin in MEKK1-/- cells, confirming that MEKK4 is indeed another mitogen-activated protein kinase kinase kinase that is specifically involved in Axin-mediated JNK activation independently of MEKK1. We have also identified an additional domain between MEKK1- and MEKK4-binding sites as being required for JNK activation by Axin. MEKK1 and MEKK4 compete for Axin binding even though they bind to sites far apart, suggesting that Axin may selectively bind to MEKK1 or MEKK4 depending on distinct signals or cellular context. Our findings will provide new insights into how scaffold proteins mediate ultimate activation of different mitogen-activated protein kinase kinase kinases.