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Thailand has implemented GNSS Continuously Operating Reference Stations (CORS) and Network Real-Time Kinematic (NRTK) Positioning services collaborated by several government agencies and universities for various applications. This research includes 114 GNSS CORSs; providing 143 triangle loops and covering 63 provinces nationwide, to determine and analyse the obtained horizontal positioning accuracy and their ground station geometries. Triangulation networks are categorised in terms of shapes and baseline distances. Welland ill-conditioned triangles are defined in accordance with different baseline lengths. The horizontal positioning accuracy is computed using static and NRTK positioning determinations whilst static positioning results are defined as the ground truth. Student's T tests are used to ensure the significance of calculated results according to each test case. The results show obtained horizontal positioning accuracies are within the specified accuracy of less than 4 centimetres and do not show any significant differences based on the defined significant level of 0.05. This GNSS CORS triangulation network geometry scheme does not influence the computed horizontal positioning accuracy obtained from NRTK-Virtual Reference Station (VRS) GNSS positioning services; however, the positioning accuracy is still directly due to distances between nearest GNSS CORS or its triangulation network, therefore, further suggestions are provided. [ABSTRACT FROM AUTHOR]

Background and Aims: Current guidelines recommend multiple biopsies from the first (D1) and second (D2) part of duodenum to establish a diagnosis of celiac disease. In this meta-analysis we aimed to find whether D1 biopsy can increase the diagnostic yield of adult celiac disease., Methods: Literature databases were searched until January 2023 for studies reporting diagnosis of celiac disease in the adult population using D1 biopsy. Meta-analysis was done using a random-effects model. Heterogeneity was assessed by I 2 % and 95% prediction interval statistics. Measured outcomes were diagnostic yield with D1 and D2 biopsies and from 4 versus 2 biopsy samples., Results: A total of 16 studies were included in the final analysis. The pooled diagnostic rate of celiac disease from D1 biopsy was 77.4% [95% CI (64.7-86.5, I 2 94%)] and from D2 biopsy was 75.3% [60.8-85.7, I 2 96%]. The pooled rate of increase in diagnostic yield with D1 biopsy was 6.9% I [4.6-10.2, I 2 66%]. The pooled diagnosis rate with 2 biopsy samples were 77.3% [50-92, I 2 93%] and 86.4% I [58.4-96.7, I 2 87%] from D1 and D2 respectively, whereas that with 4 biopsy samples were 83.3% [49.8-96.2, I 2 76%] and 70.5% I [51-84.6, I 2 96%] from D1 and D2, respectively, the difference being non-significant., Conclusion: Our study demonstrates that taking 4 biopsy samples does not incur any additional diagnostic value over taking 2 biopsy samples from each duodenum segment. Although biopsy from the D1 and D2 has similar diagnostic yield in the adult population, there was an overall increase in diagnostic yield with D1 biopsy, especially in those with a patchy disease distribution., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)

Despite the growing disease burden of non-alcoholic fatty liver disease (NAFLD), approved medical treatments to improve or prevent liver fibrosis are effective only in a small number of patients. Recent studies have found the new use of antiplatelet agents for antifibrotic benefits in NAFLD, but human studies are still limited. The goal of this meta-analysis was to combine the findings of existing relevant studies to investigate the effects of antiplatelet therapy in reducing or preventing advanced liver fibrosis in patients with NAFLD. We conducted a systematic literature search in PubMed, EMBASE, and Web of Science databases from inception to January 2021 to identify all original studies that investigated the use of antiplatelet agents in patients with NAFLD. We used the National Institutes of Health's quality assessment tool for observational cohort and cross-sectional studies to assess study quality and risk of bias. The primary outcome was the prevalence of advanced liver fibrosis stage 3-4. Data from each study was combined using the random-effects, generic inverse variance method of DerSimonian and Laird to calculate pooled odds ratio (OR) and 95% confidence intervals (CIs). Of the 2,498 studies identified, 4 studies involving 2,593 patients with NAFLD were included in this study (949 antiplatelet agent users and 1,644 non-antiplatelet agent users). The use of aspirin and/or P2Y12 receptor inhibitors was associated with a lower pooled OR of advanced liver fibrosis in patients with NAFLD (pooled OR = 0.66; 95% CI: 0.53-0.81, I 2  = 0.0%; p < 0.001). This study focuses on the outcome of advanced liver fibrosis in patients with NAFLD. Our study is limited by the small number of studies that were included. Preliminary evidence from this meta-analysis suggests a protective association between antiplatelet therapy and the prevalence of advanced liver fibrosis in patients with NAFLD. Our findings support future research into repositioning an antiplatelet agent as a novel NAFLD treatment., (© 2022. Indian Society of Gastroenterology.)

Competing Interests: Competing interests: None declared.

Parkinson's disease is the most frequent neurodegenerative motor disorder. The clinical syndrome and pathology involve motor disturbance and the degeneration of dopaminergic neurons in the substantia nigra. Root extracts of Withania. somnifera , commonly called Ashwagandha, contain several major chemical constituents known as withanolides. Studies have shown that W. somnifera extracts exhibit numerous therapeutic effects including inflammation and oxidative stress reduction, memory and cognitive function improvement. This study aimed to evaluate the protective effects of KSM-66, W. somnifera root extract, on 6-hydroxydopamine (6-OHDA)-induced toxicity in the human neuroblastoma SH-SY5Y cell line, as well as the associated oxidative response protein expression and redox regulation activity focused on S-glutathionylation. SH-SY5Y cells were treated with 6-OHDA preceded or followed by treatment with the KSM-66 extract. Using KSM-66 concentrations ranging from 0.25 to 1 mg/ml before and after treatment of the cells with 6-OHDA has resulted in an increased viability of SH-SY5Y cells. Interestingly, the extract significantly increased glutathione peroxidase activity and thioltransferase activity upon pre- or post- 6-OHDA treatment. KSM-66 also modulated oxidative response proteins: peroxiredoxin-I, VGF and vimentin proteins upon 6-OHDA pre/post treatments. In addition, the extract controlled redox regulation via S-glutathionylation. Pre-treatment of SH-SY5Y cells with KSM-66 decreased protein-glutathionylation levels in the cells treated with 6-OHDA. The rescue of mitochondria with 0.5 mg/ml KSM-66 extract showed an increase in ATP levels. These findings suggest that W. somnifera root extract acts as a neuroprotectant, thereby introducing a potential agent for the treatment or prevention of neurodegenerative diseases., Competing Interests: The authors declare no conflict of interest., (© 2021 The Author(s).)

Competing Interests: Competing interests: None declared.

Competing Interests: Competing interests: None declared.

Objectives: Globally, the burden of disease caused by alcohol use has been steadily increasing, including in Thailand. In this study, we aim to test the effectiveness of Anderson et al.'s suggested three approaches to change the collective social norms, which comprise of: (1) providing information and an understanding about alcohol use behaviour, its causes and distribution; (2) focusing strategies on groups rather than individuals; and (3) strengthening supportive laws, regulations and approaches., Study Design: We employed a mixed-methods approach. Evidence was gathered from literature review and in-depth interviews with key individuals who are responsible for community-based interventions to alcohol marketing strategies in Thailand., Methods: We chose to focus on two case studies in Nan and Surin provinces, where hospital-based longitudinal data (8 years) were available. Changes in casualties related to the harmful use of alcohol, resulting from interactions between community-based interventions and alcohol marketing during the time of annual festivals were investigated. We employed the theory of change (ToC) defined by Vogel to guide the data collection and analysis. We reviewed literature from online databases and grey literature to generate causal-loop diagrams., Results: We created a causal-loop diagram to describe the complexity of harmful alcohol use, its related factors, context, interventions and outcomes. Over the decade between 2006 and 2015, community-based strategies led to a substantial reduction of casualties (initially a 50% reduction, rising to an 80-90% reduction by the end of the study period) during the time of the festivals., Conclusions: The reduction in injuries and fatalities could be a result of the concerted actions, including legal sanctions of alcohol beverage sales and advertisement, and public education to raise awareness and impart knowledge of the harmful use of alcohol. The actions were organised by a coalition of civil society, health professionals, public authorities and community leaders using hospital-based data on the adverse effects of harmful alcohol use to mobilise political support at the provincial level. The availability of long-term financial support as a catalytic source of funds and the presence of a comprehensive alcohol control act enabled framing and mobilisation of local resources and political support., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2021 The Author(s).)

Background: Over the past 3 decades of tobacco control, Thailand has gained international recognition as a middle-income country with sustained achievement of declining smoking prevalence. However, the number of key Framework Convention on Tobacco Control measures implementation is still far away from the highest-level implementation. As a result, we aim to explore explanatory factors for the paradoxical phenomenon of sustainability in tobacco control in Thailand, to understand what the paradox means, why it happens, and how to take further steps in minimizing the paradox., Methods: We used a mixed-method approach comprising qualitative (review of literature and documents plus Program Sustainability Assessment Tool [PSAT] guided key informant interviews) and semi-quantitative methods (PSAT scoring, Theory of Change [TOC], and causal-loop diagram [CLD]) to synthesize all the findings from the qualitative data., Results: Across all eight domains, sustainability scores at the local level are lower than the national level. The highest total score was in three domains: political support, partnership, and organizational capacity. The lowest total score was for the strategic planning domain. We propose a set of key strategic elements and drivers for future strategic planning., Discussion: Using CLD, we capture a high-level view of tobacco control with dynamic interactions between contexts, mechanisms, interventions, and outcomes. We believe the deep understanding of tobacco control and the proposed strategy to counteract transnational tobacco companies in Thailand will guide future sustainable actions to reduce the prevalence of smoking, especially in the strategic planning domain that has the lowest PSAT score., (© 2020 John Wiley & Sons Ltd.)

Competing Interests: Competing interests: None declared.

The presence of air in the mediastinum is a rare finding called pneumomediastinum. Spontaneous pneumomediastinum is typically benign. Marijuana use has been rarely reported to cause pneumomediastinum. Our case series presents two young men with no significant past medical history who had a history of marijuana use and developed pneumomediastinum after multiple episodes of intractable vomiting. The pathophysiology of spontaneous pneumomediastinum in marijuana users includes cyclic vomiting or microperforation of the esophagus or barotrauma during breathing maneuvers. Most cases can be managed conservatively., (Copyright © 2020 Baylor University Medical Center.)

Dengue virus (DENV) is an arthropod-borne Flavivirus that can cause a range of symptomatic disease in humans. There are four dengue viruses (DENV 1 to 4) and infection with one DENV only provides transient protection against a heterotypic virus. Second infections are often more severe as the disease is potentiated by antibodies from the first infection through a process known as antibody dependent enhancement (ADE) of infection. Phosphorylation is a major post-translational modification that can have marked effects on a number of processes. To date there has been little information on the phosphorylation changes induced by DENV infection. This study aimed to determine global phosphoproteome changes induced by DENV 2 in U937 cells infected under an ADE protocol. A 2-dimensional electrophoretic approach coupled with a phosphoprotein-specific dye and mass spectroscopic analysis identified 15 statistically significant differentially phosphorylated proteins upon DENV 2 infection. One protein identified as significantly differentially phosphorylated, pyruvate kinase M2 (PKM2) was validated. Treatment with a PKM2 inhibitor modestly reduced levels of infection and viral output, but no change was seen in cellular viral protein levels, suggesting that PKM2 acts on exocytic virus release. While the effect of inhibition of PKM2 was relatively modest, the results highlight the need for a greater understanding of the role of phosphoproteins in DENV infection.

Objective: The present study explores factors related to length of stay (LOS) in a rural public hospital in Thailand and assesses the feasibility of using LOS as an ED key performance indicator., Methods: Using a mixed-methods approach, qualitative methods (in-depth interviews, patients' chart review and participatory observations) were used to guide and elaborate findings from quantitative analysis of 555 electronic ED records., Results: Multivariate analysis revealed that age, Emergency Severity Index score and number of laboratory tests were significantly associated with LOS. The qualitative approach provided contradicting evidence on the linkage between LOS and patient outcomes. On the one hand, considering the 4 h rule, a child with asthma was referred to a tertiary care hospital because of deterioration after 4 h of ED care. On the other hand, a woman with sepsis was hospitalised with improved condition despite 7 h of ED care. Interviews revealed the waiting time to see doctors was probably the top priority issue for patients., Conclusions: Factors related to LOS in a rural hospital in Thailand are similar and in contrast to those of a previous study in a medical school setting. Reasons for the discrepancy of findings and implications for improving ED services were discussed. Our data support the notion of controversy in using LOS as a key indicator of ED performance in this rural hospital setting. Thus, it is imperative to not rely on any single throughput or process indicators to monitor ED performance, but to take into account a set of indicators including patient outcomes., (© 2019 Australasian College for Emergency Medicine.)

Background: Evidence has been limited regarding broader emergency systems assessment in low- and middle-income countries. The aim of the present study was to provide the empirical evidence of Thai emergency department (ED) workforce vis-à-vis workload on a national scale, the availability of services for selected high-priority health burdens, and the governance obstacles in addressing the workforce management., Method: One hundred thirty public Thai EDs that provide 24-hour emergency medical care were identified across Thailand as meeting the inclusion criteria. The mailed questionnaires were administered to collect data related to the objectives of the research study., Result: Responses were received from 91 of 130 (70%) hospitals. The median number of patients visiting hospital EDs was 51 221 per year with 32.8% considered nonurgent (ESI levels 4-5). University hospital EDs were staffed with a higher number of ER professionals than EDs of service-based secondary care and tertiary care hospitals under Ministry of Public Health (MOPH). Almost all hospitals expressed concerns about the deficiency of doctors and nurses especially emergency physician (EP) and emergency nurses. The percentage of hospitals reporting the availability of coronary artery catheterization (34%) and thrombolytic infusion for acute thrombotic stroke (24%) was limited. The governance obstacles to manage emergency systems were considered., Conclusion: ED staffing seems to positively correlate with workloads except university hospitals, in our study, which may suggest the influence of teaching status on the allocation of the human resource. Among the governance obstacles in ED management, a better response to nonurgent patients requires flexibility for hospitals to set financial disincentives or mobilization and management of human resources., (© 2019 John Wiley & Sons, Ltd.)

Background: Previous policy implementation in 2012 to incentivize private hospitals in Thailand, a country with universal health coverage, to provide free-of-charge emergency care using DRG-based payment resulted in an equity gap of access and copayment. To bridge the gap, strategic policies involving financial and legal interventions were implemented in 2017. This study aims to assess whether this new approach would be able to fill the gap., Methods: We analyzed an administrative dataset of over 20,206 patients visiting private hospital EDs from April 2017 to October 2017 requested for the preauthorization of access to emergency care in the first 72 hours free of charge. The association between types of insurance and the approval status was explored using logistic regression equation adjusting for age, modes of access, systolic blood pressure, respiratory rate, and Glasgow coma scores., Results and Discussion: The strategic policies implementation resulted in reversing ED payer mix from the most privileged scheme, having the major share of ED visit, to the least privileged scheme. The data showed an increasing trend of ED visits to private hospitals indicates the acceptance of the financial incentive. Obvious differences in degrees of urgency between authorized and unauthorized patients suggested the role of preauthorization as a barrier to the noncritical patient visiting the ED. Furthermore, our study depicted the gender disparity between authorized and unauthorized patients which might indicate a delay in care seeking among critical female patients. Lessons learned for policymakers in low-and-middle income countries attempting to close the equity gap of access to private hospital EDs are discussed.

Introduction: Financing health systems constitutes a key element of well-functioning healthcare system. Prior to 2015, two new financial arrangements (direct-pay and E-claim systems) were introduced on a voluntary basis which aimed to pool more financial resources and improve cash flow of prehospital care systems. The aims of this study were to (1) assess the effects of direct-pay system in terms of (a) timeliness of reimbursement to EMS agencies, (b) changes in clinical care processes, and (c) the outcomes of patient care as compared to previous system; (2) identify the reasons for or against EMS agencies to participate in direct-pay system mechanisms; (3) identify the emerging issues with potential to significantly further the advancement of EMS systems. Using a mixed-methods approach, retrospective datasets of 3,769,399 individual records of call responses from 2015 to 2017 were analyzed which compared EMS units with the direct-pay system against those without in terms of time flow of claim data and patient outcomes. For qualitative data, in-depth interviews were conducted., Results: EMS units participating in both systems had the highest percentages of financial claim being made in time as compared to those not participating in any (p=0.012). However, there were not any practically meaningful differences between EMS units participating and not participating in either of the payment systems in terms of patient care such as appropriateness of response time, airway management, and outcome of treatment. Analysis of data from focus-group and individual interviews ended up with a causal loop diagram demonstrating potential explanatory mechanisms for those findings., Conclusion: It is evident that progress has been made in terms of mobilising more financial inputs and improving financial information flow. However, there is no evidence of any changes in patient outcomes and quality of care. Furthermore, whether the progress is meaningful in filling the gaps of financial demands of the prehospital care systems is still questionable. Room for future improvement of prehospital care systems was discussed with implications for other countries.

The present study aimed to investigate the effect of APAP treatment on the expression of pro-inflammatory cytokines in the astrocytes. The mouse astrocyte cells (C8-D1A) were treated with APAP at the concentration of 100 μM for 24 h, 16 and 28 days. The expressions of pro-inflammatory cytokines and NF-kB were determined using western blot analysis. Furthermore, the expression and localization of phosphorylation of NF-kB were detected by immunohistochemical and immunofluorescent analysis. The ultrastructure of C8-D1A cells was as well monitored. The results revealed that acute APAP treatment (24 h) had no effect on the expression of pro-inflammatory cytokines and pNF-kB. This treatment did not alter the ultrastructure of C8-D1A cells when compared with those in the control cells. However, the results obtained from the study on chronic APAP-treated cells (16 and 28 days) showed the different effect of APAP treatment. The results obtained from western blot analysis showed the increment of pro-inflammatory cytokine (IL-1β and TNF-α) expressions and the activation of NF-kB signaling pathway. Nuclear translocation of pNF-kB and alteration of several cell structures were well observed in the C8-D1A cells with chronic APAP treatment. The results obtained from this study suggest that chronic APAP treatment can induce an upregulation of pro-inflammatory cytokines (IL-1β and TNFα) in astrocytes. This alteration implies the involvement of the activation of NF-kB signaling pathway.

Objective: To identify the candidate protein biomarkers of adult-onset-immunodeficiency (AOID) syndrome using serum proteomics., Methods: Screening and verification phases were performed in the study. A total of 97 serum samples were classified into three groups: AOID patients with opportunistic infections (active AOID), AOID patients without opportunistic infections (inactive AOID), and healthy control. In the screening phase, pooled sera collected from patients and healthy control in each group were separated by 2D-gel electrophoresis, analyzed for differentially expressed proteins and identified for biomarkers using LC/MS. In the verification phase, the protein candidates were selected for confirmation by western blotting., Results: The analysis revealed 35 differentially expressed proteins. Three proteins including haptoglobin, gelsolin, and transthyretin, were selected for verification. The results showed that the levels of haptoglobin in both active and inactive AOID groups were significantly higher than that in the control group, while the levels of gelsolin in the active AOID group were significantly lower than that in the inactive AOID group. The level of transthyretin in the active AOID group was also significantly lower than that in the control group., Conclusions: The comparison of serum proteins between the three groups revealed three candidates which are related to chronic inflammatory diseases. Haptoglobin and transthyretin biomarkers could be applied in clinical assessment for monitor of disease outcome, including for the study of AOID pathogenesis., (Copyright © 2017 Hainan Medical University. Production and hosting by Elsevier B.V. All rights reserved.)

Objective: To generate insights into the mechanism of NVP induced hepatotoxicity., Methods: Liver (HepG2) cells were cultured with various concentrations of NVP. This cell line was chosen because it has low expression of cytochrome P450, allowing evaluation of the effects of NVP rather than specific metabolites. Cytotoxicity was determined using a proliferation assay and cell numbers were monitored using trypan blue exclusion assay for long term culture experiments and apoptosis induction was determined by morphological and biochemical investigation., Results: HepG2 cells treated with the highest concentration of NVP tested (819 μM) initially showed a rounded morphology and all cells had died by week three of exposure. Nuclear condensation and fragmentation, increased Annexin V/propidium iodide staining and caspase 9 activation all supported the induction of apoptosis in HepG2 cells in response to NVP treatment., Conclusions: There is a clear induction of apoptosis in response to NVP which suggests that NVP has significant cytotoxicity, over and above any cytotoxicity of metabolites and may contribute directly to patient hepatotoxicity., (Copyright © 2016 Hainan Medical College. Production and hosting by Elsevier B.V. All rights reserved.)

Previously, our group has demonstrated that chronic paracetamol (APAP) treatment induces alterations to the trigeminovascular nociceptive system in the cortical spreading depression (CSD) migraine animal model. The calcitonin gene related peptide (CGRP) is a key neuropeptide involved in the activation of the trigeminovascular nociceptive system. Therefore, this study examined the expression levels of CGRP in the trigeminal ganglion (TG) after chronic APAP exposure (0, 15, and 30 days) using a CSD model. Rats were divided into control, CSD only, APAP only and APAP treatment with CSD groups. A single injection (i.p.) of APAP (200 mg/kg body weight) was given to the 0-day APAP-treated groups, while the other APAP-treated groups received daily injections for 15 and 30 days. CSD was induced by the topical application of KCl to the parietal cortex. The protein expression of CGRP in the TG was evaluated by immunohistochemistry, and the CGRP mRNA level was investigated by real-time quantitative reverse transcription polymerase chain reaction. The results revealed that the induction of CSD significantly increased the level of CGRP protein but had no effect on CGRP mRNA level. Pretreatment with APAP 1 hour before CSD activation significantly reduced CGRP expression induced by CSD. In contrast, chronic treatment with APAP (15 and 30 days) significantly enhanced CGRP expression in both protein and mRNA levels when compared with the control groups. In combination with CSD, the expression of CGRP further increased in the animal with 30 day treatment. These findings indicate that chronic treatment with APAP induces an increase of CGRP expression in the TG. This alteration may be associated with the increased trigeminovascular nociception observed in our previous studies., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The mechanisms by which MUC1 and p120 catenin contribute to progression of cancers from early transformation to metastasis are poorly understood. Here we show that p120 catenin ARM domains 1, 3-5, and 8 mediate interactions between p120 catenin and MUC1, and that these interactions modulate dynamic properties of cell adhesion, motility, and metastasis of pancreatic cancer cells. We also show that different isoforms of p120 catenin, when coexpressed with MUC1, create cells that exhibit distinct patterns of motility in culture (motility independent of cell adhesion, motility within a monolayer while exchanging contacts with other cells, and unified motility while maintaining static epithelial contacts) and patterns of metastasis. The results provide new insight into the dynamic interplay between cell adhesion and motility and the relationship of these to the metastatic process., (©2013 AACR)

Nevirapine (NVP) is an effective nonnucleoside reverse transcriptase inhibitor (NNRTI) of particular interest as it is often used in resource limited countries. However, one of the main concerns with the use of NVP is hepatotoxicity and elevation of liver enzymes as a consequence of highly active antiretroviral therapy (HAART) containing NVP is more often reported in HIV patients coinfected with hepatitis C virus than in HIV-monoinfected patients. To discover possible markers of NVP induced hepatotoxicity, serum and urine samples from twenty-five HIV or HIV/HCV patients, all of whom had received NVP continuously for at least four months, and healthy controls were subjected to in-solution or in-gel proteomic analysis. A total of 83 differentially regulated proteins consisted of 34 proteins identified in serum by in-solution analysis, 2 proteins identified from serum in a 2D gel electrophoresis analysis, and 47 proteins identified in urine in an in-solution analysis. Three proteins, namely, haptoglobin, Rho-related BTB domain containing protein 3, and death-associated protein kinase 3, were selected for further validation by Western blot analysis and results showed that haptoglobin has potential for further development as an additional marker of NVP induced hepatotoxicity.

Japanese encephalitis virus (JEV) a mosquito-borne flavivirus is a major cause of viral encephalitis in Asia. While the principle target cells for JEV in the central nervous system are believed to be neurons, microglia are activated in response to JEV and have been proposed to act as a long lasting virus reservoir. Viral attachment to a host cell is the first step of the viral entry process and is a critical mediator of tissue tropism. This study sought to identify molecules associated with JEV entry to microglial cells. Virus overlay protein-binding assay (VOPBA) and liquid chromatography-mass spectrometry (LC/MS/MS) identified the 37/67 kDa high-affinity laminin receptor protein and nucleolin as a potential JEV-binding proteins. These proteins were subsequently investigated for a contribution to JEV entry to mouse microglial BV-2 cells together with other possible candidate receptor molecules including Hsp70, Hsp90, GRP78, CD14, and CD4. In antibody mediated inhibition of infection experiments, both anti-laminin receptor and anti-CD4 antibodies significantly reduced virus entry while anti-Hsp70 and 90 antibodies produced a slight reduction. Significant inhibition of virus entry (up to 80%) was observed in the presence of lipopolysaccharide (LPS) which resulted in a complete down-regulation of CD4 and moderate down-regulation of CD14. These results suggest that multiple receptor proteins may mediate the entry of JEV to microglial cells, with CD4 playing a major role., (Copyright © 2012 Wiley Periodicals, Inc.)

Despite the availability of effective vaccines, Japanese encephalitis virus (JEV) infections remain a leading cause of encephalitis in many Asian countries. The virus is transmitted to humans by Culex mosquitoes, and, while the majority of human infections are asymptomatic, up to 30% of JE cases admitted to hospital die and 50% of the survivors suffer from neurological sequelae. Microglia are brain-resident macrophages that play key roles in both the innate and adaptive immune responses in the CNS and are thus of importance in determining the pathology of encephalitis as a result of JEV infection.

Japanese encephalitis virus (JEV), a mosquito-borne Flavivirus, is a major cause of acute encephalitis, and neurons have been proposed to be the principle JEV target cells in the central nervous system. However, clinically, infection with JEV leads to increased levels of cytokines and chemokines in the serum and cerebrospinal fluid (CSF) the levels of which correlate with the mortality rate of patients. This research aimed to study the role of microglial cells in JEV infection. Mouse microglial cells (BV-2) supported the replication of JEV with extracellular production of virus by 10h post-infection, and virus titer reached a maximum (2.55x10(10)pfu/ml) by day 3 post-infection. While apoptosis was induced in response to virus infection, no alteration in nitric oxide production was observed. Microglial cells remained productively infected with JEV for up to 16 weeks without significant morphological alterations, and the released virions were infectious to mouse neuroblastoma (NA) cells. The high virus production and long persistence of JEV in microglial cells suggests that these cells may serve as viral reservoirs for the infection of neurons in the CNS.

While both in vivo and in vitro evidence has suggested that liver cells undergo apoptosis in response to dengue virus infection, little is known about the mechanism of induction. Given that the p53 tumour suppressor gene is a key mediator of apoptosis, we sought to define the role of this gene in response to dengue virus infection. After infection, a p53 wild type liver cell line (HepG2) showed changes consistent with apoptosis including alterations of cell morphology, cellular detachment and DNA laddering. However, p53 was neither up-regulated, nor showed any evidence of complexing with dengue virus proteins as determined by immunoprecipitation. Infection of a p53 null liver cell line (Hep3B) also produced changes consistent with the induction of apoptosis. While the profile of the cells undergoing apoptosis in each cell line was similar as determined by flow cytometry, the absolute levels were markedly different with up to 90% of Hep3B cells undergoing apoptosis compared to only 20% of HepG2 cells at day 5 post infection. By day 7, all Hep3B infected cells were dead. In contrast, it proved possible to culture dengue virus infected HepG2 cells for 3 months. Viral progeny released from the p53 null cell line were nine-fold higher per attached cell than from the p53 wild type cell line. These results suggest that, while induction of apoptosis in liver cells is mediated by a non-p53 regulated pathway, p53 may play a role in restricting the level of viral progeny to below a critical level at which apoptosis is triggered., (Copyright 2004 Wiley-Liss, Inc.)

Three methods were used in succession to screen a whole adult zebrafish cDNA library for expressed p53-like genes. The sequences of the resultant clones describe an open reading frame 1122 nucleotides in length, with another 43 and 940 bases of 5' and 3' untranslated sequence, respectively. The deduced amino acid sequence of the zebrafish p53 protein is 63% identical to that of trout and 48% identical to that of human p53. Two of the three zebrafish clones overlap to span the entire reported cDNA sequence and are identical in their deduced amino acid sequence over their coincident length. The third clone contains a conservative amino acid change, as well as an inserted amino acid subsequently found to be at the junction of exons 2 and 3, suggestive of alternative splicing in the p53 mRNA for this species. Northern analysis demonstrated a zebrafish p53-related transcript to be present and most abundant in zygotes and early-cleavage embryos less than 1 hour after fertilization, thereafter declining to barely detectable levels at 48 hours. A similar temporal expression was detected for the zebrafish L-myc, known to be present in maternally derived RNA, whereas zebrafish N-myc and the zebrafish homologue of the murine T gene were not detectable prior to the onset of zygotic transcription.