STK11 Causative Variants and Copy Number Variations Identified in Thai Patients With Peutz-Jeghers Syndrome.

Introduction Peutz-Jeghers syndrome (PJS) is a rare autosomal dominant inherited disorder caused by germline mutations in the serine-threonine kinase 11 (STK11) tumor suppressor gene. This syndrome is characterized by hamartomatous gastrointestinal polyps, mucocutaneous melanin pigmentation, and a higher risk of developing various cancers. Methods We summarized the clinical and molecular characteristics of five unrelated Thai patients with PJS. Denaturing high-performance liquid chromatography (DHPLC) screening, coupled with direct DNA sequencing and multiplex ligation-dependent probe amplification (MLPA), were applied for the molecular analysis of STK11. Results A total of four STK11 pathogenic changeswere identified in the five PJS patients, including two frameshift variants (a novel c.199dup, p.Leu67ProfsTer96 and a known c.834_835del, p.Cys278TrpfsTer6) and two types of copy number variations (CNV), exon 1 deletion and exons 2-3 deletion. Among reported STK11 exonic deletions, exon 1 and exons 2-3 deletions were found to be the two most commonly deleted exons. Conclusion All identified STK11 mutations were null mutations that were associated with more severe PJS phenotypes and cancers. This study broadens the phenotypic and mutational spectrum of STK11 in PJS.

Cepharanthine inhibits dengue virus production and cytokine secretion.

Dengue virus (DENV) infection is a public health problem in tropical and subtropical regions. It can cause a spectrum of clinical manifestations ranging from mild dengue fever (DF) to severe dengue haemorrhagic fever (DHF) and potentially life-threatening disease including dengue shock syndrome (DSS). Severe DENV infection is caused by high viral load and cytokine storm in dengue-infected patients. Currently, there is no specific antiviral drug for DENV infection. An anti-DENV agent that demonstrates inhibitory effects on both DENV replication and cytokine secretion is urgently needed. In this study, cepharanthine (CEP), which is an anti-inflammatory, anti-HIV, and anti-tumor compound isolated from Stephania cepharantha Hayata, was tested for inhibition of DENV infection. We investigated the efficacy of CEP to inhibit DENV infection, replication, and cytokine production. The inhibitory effect of CEP treatment was studied in DENV-infected human chronic myeloid leukemia (K562) cells. The levels of DENV E protein and DENV production were determined by flow cytometry and FFU assay, respectively. CEP treatment significantly reduced viral E protein and viral production in all DENV-1, 2, 3, 4 serotypes. In addition, CEP treatment reduced the IL-6 proinflammatory cytokine production in DENV-infected A549 cells. Taken together, CEP has inhibitory effects on DENV infection specifically at the initial viral replication states and proinflammatory cytokine secretion, and is a promising candidate for further development as an anti-DENV treatment.

Cytoprotective effect of genistein against dexamethasone-induced pancreatic ß-cell apoptosis.

Steroid-induced diabetes is a well-known metabolic side effect of long-term use of glucocorticoid (GC). Our group recently demonstrated dexamethasone-induced pancreatic ß-cell apoptosis via upregulation of TRAIL and TRAIL death receptor (DR5). Genistein protects against pancreatic ß-cell apoptosis induced by toxic agents. This study aimed to investigate the cytoprotective effect of genistein against dexamethasone-induced pancreatic ß-cell apoptosis in cultured rat insulinoma (INS-1) cell line and in isolated mouse islets. In the absence of genistein, dexamethasone-induced pancreatic ß-cell apoptosis was associated with upregulation of TRAIL, DR5, and superoxide production, but downregulation of TRAIL decoy receptor (DcR1). Dexamethasone also activated the expression of extrinsic and intrinsic apoptotic proteins, including Bax, NF-κB, caspase-8, and caspase-3, but suppressed the expression of the anti-apoptotic Bcl-2 protein. Combination treatment with dexamethasone and genistein protected against pancreatic ß-cell apoptosis, and reduced the effects of dexamethasone on the expressions of TRAIL, DR5, DcR1, superoxide production, Bax, Bcl-2, NF-κB, caspase-8, and caspase-3. Moreover, combination treatment with dexamethasone and genistein reduced the expressions of TRAIL and DR5 in isolated mouse islets. The results of this study demonstrate the cytoprotective effect of genistein against dexamethasone-induced pancreatic ß-cell apoptosis in both cell line and islets via reduced TRAIL and DR5 protein expression.

Potential Phosphorylation of Viral Nonstructural Protein 1 in Dengue Virus Infection.

Dengue virus (DENV) infection causes a spectrum of dengue diseases that have unclear underlying mechanisms. Nonstructural protein 1 (NS1) is a multifunctional protein of DENV that is involved in DENV infection and dengue pathogenesis. This study investigated the potential post-translational modification of DENV NS1 by phosphorylation following DENV infection. Using liquid chromatography-tandem mass spectrometry (LC-MS/MS), 24 potential phosphorylation sites were identified in both cell-associated and extracellular NS1 proteins from three different cell lines infected with DENV. Cell-free kinase assays also demonstrated kinase activity in purified preparations of DENV NS1 proteins. Further studies were conducted to determine the roles of specific phosphorylation sites on NS1 proteins by site-directed mutagenesis with alanine substitution. The T27A and Y32A mutations had a deleterious effect on DENV infectivity. The T29A, T230A, and S233A mutations significantly decreased the production of infectious DENV but did not affect relative levels of intracellular DENV NS1 expression or NS1 secretion. Only the T230A mutation led to a significant reduction of detectable DENV NS1 dimers in virus-infected cells; however, none of the mutations interfered with DENV NS1 oligomeric formation. These findings highlight the importance of DENV NS1 phosphorylation that may pave the way for future target-specific antiviral drug design.

Dexamethasone induces pancreatic ß-cell apoptosis through upregulation of TRAIL death receptor.

Long-term medication with dexamethasone - a synthetic glucocorticoid (GC) drug - results in hyperglycemia, or steroid-induced diabetes. Although recent studies revealed that dexamethasone directly induces pancreatic ß-cell apoptosis, its molecular mechanisms remain unclear. In our initial analysis of mRNA transcripts, we discovered the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) pathway may be involved in dexamethasone-induced pancreatic ß-cell apoptosis. In the present study, a mechanism of dexamethasone-induced pancreatic ß-cell apoptosis through the TRAIL pathway was investigated in cultured cells and isolated mouse islets. INS-1 cells were cultured with and without dexamethasone in the presence or absence of a glucocorticoid receptor (GR) inhibitor, RU486. We found that dexamethasone induced pancreatic ß-cell apoptosis in association with the upregulation of TNSF10 (TRAIL) mRNA and protein expression. Moreover, dexamethasone upregulated the TRAIL death receptor (DR5) protein but suppressed the decoy receptor (DcR1) protein. Similar findings were observed in mouse isolated islets: dexamethasone increased TRAIL and DR5 compared to that of control mice. Furthermore, dexamethasone stimulated pro-apoptotic signaling including superoxide production, caspase-8, -9, and -3 activities, NF-κB, and Bax but repressed the anti-apoptotic protein, Bcl-2. All these effects were inhibited by the GR-inhibitor, RU486. Furthermore, knock-down DR5 decreased dexamethasone-induced caspase 3 activity. Caspase-8 and caspase-9 inhibitors protected pancreatic ß-cells from dexamethasone-induced apoptosis. Taken together, dexamethasone induced pancreatic ß-cell apoptosis by binding to the GR and inducing DR5 and TRAIL pathway.

Melatonin Inhibits Dengue Virus Infection via the Sirtuin 1-Mediated Interferon Pathway.

Dengue virus (DENV) is the causative pathogen in the life-threatening dengue hemorrhagic fever and dengue shock syndrome. DENV is transmitted to humans via the bite of an infected Aedes mosquito. Approximately 100 million people are infected annually worldwide, and most of those live in tropical and subtropical areas. There is still no effective drug or vaccine for treatment of DENV infection. In this study, we set forth to investigate the effect of melatonin, which is a natural hormone with multiple pharmacological functions, against DENV infection. Treatment with subtoxic doses of melatonin dose-dependently inhibited DENV production. Cross-protection across serotypes and various cell types was also observed. Time-of-addition assay suggested that melatonin exerts its influence during the post-entry step of viral infection. The antiviral activity of melatonin partly originates from activation of the sirtuin pathway since co-treatment with melatonin and the sirtuin 1 (SIRT1) inhibitor reversed the effect of melatonin treatment alone. Moreover, melatonin could modulate the transcription of antiviral genes that aid in suppression of DENV production. This antiviral mechanism of melatonin suggests a possible new strategy for treating DENV infection.

Protein disulfide isomerase A1 regulates breast cancer cell immunorecognition in a manner dependent on redox state.

Oxidoreductase protein disulphide isomerases (PDI) are involved in the regulation of a variety of biological processes including the modulation of endoplasmic reticulum (ER) stress, unfolded protein response (UPR), ER­mitochondria communication and the balance between pro­survival and pro­death pathways. In the current study the role of the PDIA1 family member in breast carcinogenesis was investigated by measuring ROS generation, mitochondrial membrane disruption, ATP production and HLA­G protein levels on the surface of the cellular membrane in the presence or absence of PDIA1. The results showed that this enzyme exerted pro­apoptotic effects in estrogen receptor (ERα)­positive breast cancer MCF­7 and pro­survival in triple negative breast cancer (TNBC) MDA­MB­231 cells. ATP generation was upregulated in PDIA1­silenced MCF­7 cells and downregulated in PDIA1­silenced MDA­MB­231 cells in a manner dependent on the cellular redox status. Furthermore, MCF­7 and MDA­MB­231 cells in the presence of PDIA1 expressed higher surface levels of the non­classical human leukocyte antigen (HLA­G) under oxidative stress conditions. Evaluation of the METABRIC datasets showed that low PDIA1 and high HLA­G mRNA expression levels correlated with longer survival in both ERα­positive and ERα­negative stage 2 breast cancer patients. In addition, analysis of the PDIA1 vs. the HLA­G mRNA ratio in the subgroup of the living stage 2 breast cancer patients exhibiting low PDIA1 and high HLA­G mRNA levels revealed that the longer the survival time of the ratio was high PDIA1 and low HLA­G mRNA and occurred predominantly in ERα­positive breast cancer patients whereas in the same subgroup of the ERα­negative breast cancer mainly this ratio was low PDIA1 and high HLA­G mRNA. Taken together these results provide evidence supporting the view that PDIA1 is linked to several hallmarks of breast cancer pathways including the process of antigen processing and presentation and tumor immunorecognition.

Peptides targeting dengue viral nonstructural protein 1 inhibit dengue virus production.

Viruses manipulate the life cycle in host cells via the use of viral properties and host machineries. Development of antiviral peptides against dengue virus (DENV) infection has previously been concentrated on blocking the actions of viral structural proteins and enzymes in virus entry and viral RNA processing in host cells. In this study, we proposed DENV NS1, which is a multifunctional non-structural protein indispensable for virus production, as a new target for inhibition of DENV infection by specific peptides. We performed biopanning assays using a phage-displayed peptide library and identified 11 different sequences of 12-mer peptides binding to DENV NS1. In silico analyses of peptide-protein interactions revealed 4 peptides most likely to bind to DENV NS1 at specific positions and their association was analysed by surface plasmon resonance. Treatment of Huh7 cells with these 4 peptides conjugated with N-terminal fluorescent tag and C-terminal cell penetrating tag at varying time-of-addition post-DENV infection could inhibit the production of DENV-2 in a time- and dose-dependent manner. The inhibitory effects of the peptides were also observed in other virus serotypes (DENV-1 and DENV-4), but not in DENV-3. These findings indicate the potential application of peptides targeting DENV NS1 as antiviral agents against DENV infection.

Endoplasmic reticulum stress, unfolded protein response and autophagy contribute to resistance to glucocorticoid treatment in human acute lymphoblastic leukaemia cells.

Acute lymphoblastic leukaemia (ALL) is the most frequent childhood cancer and, although it is highly treatable, resistance to therapy, toxicity and side effects remain challenging. The synthetic glucocorticoid (GC) dexamethasone (Dex) is commonly used to treat ALL, the main drawback of which is the development of resistance to this treatment. The aim of the present study was to investigate potential molecular circuits mediating resistance and sensitivity to GC­induced apoptosis in ALL. The leukaemia cell lines CEM­C7­14, CEM­C1­15 and MOLT4 treated with chloroquine (CLQ), thapsigargin (TG) and rotenone (ROT) were used to explore the roles of autophagy, endoplasmic reticulum (ER) stress/unfolded protein response (UPR) and reactive oxygen species (ROS) generation in the response to GC treatment. ROS levels were associated with increased cell death and mitochondrial membrane potential in rotenone­treated CEM cells. Autophagy inhibition by CLQ exhibited the strongest cytotoxic effect in GC­resistant leukaemia. Autophagy may act as a pro­survival mechanism in GC­resistant leukaemia since increasing trends in beclin­1 and microtubule­associated protein 1 light chain 3α levels were detected in CEM­C1­15 and MOLT4 cells treated with Dex, whereas decreasing trends in these autophagy markers were observed in CEM­C7­14 cells. The intracellular protein levels of the ER stress markers glucose­regulated protein (GRP)78 and GRP94 were stimulated by Dex only in the GC­sensitive cells, suggesting a role of these chaperones in the GC­mediated ALL cell death. Increased cell surface levels of GRP94 were recorded in CEM­C7­14 cells treated with combination of Dex with TG compared with those in cells treated with TG alone, whereas decreasing trends were observed in CEM­C1­15 cells under these conditions. Taken together, the results of the present study demonstrated that autophagy may be a pro­survival mechanism in GC­resistant leukaemia, and by modulating intracellular and surface GRP94 protein levels, Dex is involved in the regulation of ER stress/UPR­dependent cell death and immune surveillance. These observations may be of clinical importance if confirmed in patients.

Expression of androgen receptor and its regulatory molecule Lin28 in non-luminal subtype breast cancer.

Androgen receptor (AR) was associated with favourable outcome in luminal breast cancer. However, the role of AR in non-luminal breast cancer remains inconclusive. The aim of the present study was to evaluate the clinical significance of the AR and its regulatory pathway in non-luminal subtypes of breast cancer. In total, 284 breast cancer patients were recruited from January 2007 to January 2016. Tissue microarrays were constructed from archival paraffin blocks and assessed for AR and its regulatory molecule, Lin28, by immunohistochemistry. The association between AR and Lin28 expression and clinicopathological parameters was analyzed. Results showed that AR and Lin28 were co-expressed. No association between these proteins and clinicopathological parameters, and survival outcome was found. However, a higher proportion of the patients with AR and Lin28 expression were observed in HER2 subtype. In conclusion, Lin28 may be a novel marker for prognosis and targeted for treatment in HER2 subtype breast cancer.

Sirtuin Family Members Selectively Regulate Autophagy in Osteosarcoma and Mesothelioma Cells in Response to Cellular Stress.

The class III NAD+ dependent deacetylases-sirtuins (SIRTs) link transcriptional regulation to DNA damage response and reactive oxygen species generation thereby modulating a wide range of cellular signaling pathways. Here, the contribution of SIRT1, SIRT3, and SIRT5 in the regulation of cellular fate through autophagy was investigated under diverse types of stress. The effects of sirtuins' silencing on cell survival and autophagy was followed in human osteosarcoma and mesothelioma cells exposed to DNA damage and oxidative stress. Our results suggest that the mitochondrial sirtuins SIRT3 and 5 are pro-proliferative under certain cellular stress conditions and this effect correlates with their role as positive regulators of autophagy. SIRT1 has more complex role which is cell type specific and can affect autophagy in both positive and negative ways. The mitochondrial sirtuins (SIRT3 and SIRT5) affect both early and late stages of autophagy, whereas SIRT1 acts mostly at later stages of the autophagic process. Investigation of potential crosstalk between SIRT1, SIRT3, and SIRT5 revealed several feedback loops and a significant role of SIRT5 in regulating SIRT3 and SIRT1. Results presented here support the notion that sirtuin family members play important as well as differential roles in the regulation of autophagy in osteosarcoma vs. mesothelioma cells exposed to DNA damage and oxidative stress, and this can be exploited in increasing the response of cancer cells to chemotherapy.

Serine protease inhibitor AEBSF reduces dengue virus infection via decreased cholesterol synthesis.

Dengue virus (DENV) infection has evolved into a major global health menace and economic burden due to its intensity and geographic distribution. DENV infection in humans can cause a wide range of symptoms including dengue fever (DF), dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). An antiviral agent that is effective against all four serotypes of DENV is urgently needed to prevent and to manage this condition. Reducing the viral load during the early phase of infection may minimize the chance of patients progressing to more severe DHF or DSS. In this study, we set forth to investigate the anti-viral effect of five commercially available protease inhibitors on DENV infection since both viral and host proteases can contribute to effective viral replication. Previously, the serine protease inhibitor AEBSF [4-(2-aminoethyl) benzene sulfonyl fluoride] has been shown to inhibit DENV NS3 protease activity. The results of the present study revealed that DENV genome replication and protein synthesis were significantly inhibited by AEBSF in a dose-dependent manner. AEBSF inhibited the expression of genes such as 3-hydroxy 3-methyl-glutaryl-CoA synthase (HMGCS), 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGCR), and low-density lipoprotein receptor (LDLR). Moreover, AEBSF significantly inhibited HMGCR activity and intracellular cholesterol synthesis after DENV infection. The anti-DENV effect of AEBSF was confirmed in all four DENV serotypes and in three different cell lines. These results indicate that AEBSF reduces DENV infection via both viral and host protease activities.

Drug repurposing of N-acetyl cysteine as antiviral against dengue virus infection.

Liver injury is one of the hallmark features of severe dengue virus (DENV) infection since DENV can replicate in the liver and induce hepatocytes to undergo apoptosis. N-acetyl cysteine (NAC), which is a clinically-used drug for treating acetaminophen toxicity, was found to benefit patients with DENV-induced liver injury; however, its mechanism of action remains unclear. Accordingly, our aim was to repurpose NAC in the preclinical studies to investigate its mechanism of action. Time of addition experiments in HepG2 cells elucidated effectiveness of NAC to reduce infectious virion at pre-, during- and post infection. In DENV-infected mice, NAC improved DENV-associated clinical manifestations, including leucopenia and thrombocytopenia, and reduced liver injury and hepatocyte apoptosis. Interestingly, we discovered that NAC significantly reduced DENV production in HepG2 cells and in liver of DENV-infected mice by induction of antiviral responses via interferon signaling. NAC treatment in DENV-infected mice helped to maintain antioxidant enzymes and redox balance in the liver. Therefore, NAC reduces DENV production and oxidative damage to ameliorate DENV-induced liver injury. Taken together, these findings suggest the novel therapeutic potential of NAC in DENV-induced liver injury and recommend evaluating its efficacy and safety in humans with DENV-induced liver injury.

Vivo-morpholino oligomers strongly inhibit dengue virus replication and production.

Dengue virus (DENV) infection is a worldwide public health problem, which can cause severe dengue hemorrhagic fever (DHF) and life-threatening dengue shock syndrome (DSS). There are currently no anti-DENV drugs available, and there has been an intensive search for effective anti-DENV agents that can inhibit all four DENV serotypes. In this study, we tested whether vivo-morpholino oligomers (vivo-MOs), whose effect on DENV infection has not previously been studied, can inhibit DENV infection. Vivo-MOs were designed to target the top of 3' stem-loop (3' SL) in the 3' UTR of the DENV genome and tested for inhibition of DENV infection in monkey kidney epithelial (Vero) cells and human lung epithelial carcinoma (A549) cells. The results showed that vivo-MOs could bind to a DENV RNA sequence and markedly reduce DENV-RNA, protein, and virus production in infected Vero and A549 cells. Vivo-MOs at a concentration of 4 µM could inhibit DENV production by more than 104-fold when compared to that of an untreated control. In addition, vivo-MOs also inhibited DENV production in U937 cells and primary human monocytes. Therefore, vivo-MOs targeting to the 3' SL in the 3' UTR of DENV genomes are effective and have the potential to be developed as anti-DENV agents.

RNAi screen reveals a role of SPHK2 in dengue virus-mediated apoptosis in hepatic cell lines.

Hepatic dysfunction is a feature of dengue virus (DENV) infection. Hepatic biopsy specimens obtained from fatal cases of DENV infection show apoptosis, which relates to the pathogenesis of DENV infection. However, how DENV induced liver injury is not fully understood. In this study, we aim to identify the factors that influence cell death by employing an apoptosis-related siRNA library screening. Our results show the effect of 558 gene silencing on caspase 3-mediated apoptosis in DENV-infected Huh7 cells. The majority of genes that contributed to apoptosis were the apoptosis-related kinase enzymes. Tumor necrosis factor superfamily member 12 (TNFSF12), and sphingosine kinase 2 (SPHK2), were selected as the candidate genes to further validate their influences on DENV-induced apoptosis. Transfection of siRNA targeting SPHK2 but not TNFSF12 genes reduced apoptosis determined by Annexin V/PI staining. Knockdown of SPHK2 did not reduce caspase 8 activity; however, did significantly reduce caspase 9 activity, suggesting its involvement of SPHK2 in the intrinsic pathway of apoptosis. Treatment of ABC294649, an inhibitor of SPHK2, reduced the caspase 3 activity, suggesting the involvement of its kinase activity in apoptosis. Knockdown of SPHK2 significantly reduced caspase 3 activity not only in DENV-infected Huh7 cells but also in DENV-infected HepG2 cells. Our results were consistent across all of the four serotypes of DENV infection, which supports the pro-apoptotic role of SPHK2 in DENV-infected liver cells.

JNK1/2 inhibitor reduces dengue virus-induced liver injury.

High viral load with liver injury is exhibited in severe dengue virus (DENV) infection. Mitogen activated protein kinases (MAPKs) including ERK1/2 and p38 MAPK were previously found to be involved in the animal models of DENV-induced liver injury. However, the role of JNK1/2 signaling in DENV-induced liver injury has never been investigated. JNK1/2 inhibitor, SP600125, was used to investigate the role of JNK1/2 signaling in the BALB/c mouse model of DENV-induced liver injury. SP600125-treated DENV-infected mice ameliorated leucopenia, thrombocytopenia, hemoconcentration, liver transaminases and liver histopathology. DENV-induced liver injury exhibited induced phosphorylation of JNK1/2, whereas SP600125 reduced this phosphorylation. An apoptotic real-time PCR array profiler was used to screen how SP600125 affects the expression of 84 cell death-associated genes to minimize DENV-induced liver injury. Modulation of caspase-3, caspase-8 and caspase-9 expressions by SP600125 in DENV-infected mice suggests its efficiency in restricting apoptosis via both extrinsic and intrinsic pathways. Reduced expressions of TNF-α and TRAIL are suggestive to modulate the extrinsic apoptotic signals, where reduced p53 phosphorylation and induced anti-apoptotic Bcl-2 expression indicate the involvement of the intrinsic apoptotic pathway. This study thus demonstrates the pivotal role of JNK1/2 signaling in DENV-induced liver injury and how SP600125 modulates this pathogenesis.

Tyrosine kinase/phosphatase inhibitors decrease dengue virus production in HepG2 cells.

Dengue virus is the causative agent of dengue fever, dengue hemorrhagic fever, and dengue shock syndrome. High rates of dengue virus replication and virion production are related to disease severity. To identify anti-DENV compounds, we performed cell-based ELISA testing to detect the level of DENV E protein expression. Among a total of 83 inhibitors, eight were identified as inhibitors with antiviral activity. Epidermal growth factor receptor inhibitor II (EGFR/ErbB-2/ErbB-4 inhibitor II) and protein tyrosine phosphatase inhibitor IV (PTP inhibitor IV) significantly inhibited dengue virus production and demonstrated low toxicity in hepatocyte cell lines. Our results suggest the efficacy of tyrosine kinase/phosphatase inhibitors in decreasing dengue virus production in HepG2 cells.

Drug repurposing of minocycline against dengue virus infection.

Dengue virus infection is one of the most common arthropod-borne viral diseases. A complex interplay between host and viral factors contributes to the severity of infection. The antiviral effects of three antibiotics, lomefloxacin, netilmicin, and minocycline, were examined in this study, and minocycline was found to be a promising drug. This antiviral effect was confirmed in all four serotypes of the virus. The effects of minocycline at various stages of the viral life cycle, such as during viral RNA synthesis, intracellular envelope protein expression, and the production of infectious virions, were examined and found to be significantly reduced by minocycline treatment. Minocycline also modulated host factors, including the phosphorylation of extracellular signal-regulated kinase1/2 (ERK1/2). The transcription of antiviral genes, including 2'-5'-oligoadenylate synthetase 1 (OAS1), 2'-5'-oligoadenylate synthetase 3 (OAS3), and interferon α (IFNA), was upregulated by minocycline treatment. Therefore, the antiviral activity of minocycline may have a potential clinical use against Dengue virus infection.

Mass spectrometric analysis of host cell proteins interacting with dengue virus nonstructural protein 1 in dengue virus-infected HepG2 cells.

Dengue virus (DENV) infection is a leading cause of the mosquito-borne infectious diseases that affect humans worldwide. Virus-host interactions appear to play significant roles in DENV replication and the pathogenesis of DENV infection. Nonstructural protein 1 (NS1) of DENV is likely involved in these processes; however, its associations with host cell proteins in DENV infection remain unclear. In this study, we used a combination of techniques (immunoprecipitation, in-solution trypsin digestion, and LC-MS/MS) to identify the host cell proteins that interact with cell-associated NS1 in an in vitro model of DENV infection in the human hepatocyte HepG2 cell line. Thirty-six novel host cell proteins were identified as potential DENV NS1-interacting partners. A large number of these proteins had characteristic binding or catalytic activities, and were involved in cellular metabolism. Coimmunoprecipitation and colocalization assays confirmed the interactions of DENV NS1 and human NIMA-related kinase 2 (NEK2), thousand and one amino acid protein kinase 1 (TAO1), and component of oligomeric Golgi complex 1 (COG1) proteins in virus-infected cells. This study reports a novel set of DENV NS1-interacting host cell proteins in the HepG2 cell line and proposes possible roles for human NEK2, TAO1, and COG1 in DENV infection.

SB203580 Modulates p38 MAPK Signaling and Dengue Virus-Induced Liver Injury by Reducing MAPKAPK2, HSP27, and ATF2 Phosphorylation.

Dengue virus (DENV) infection causes organ injuries, and the liver is one of the most important sites of DENV infection, where viral replication generates a high viral load. The molecular mechanism of DENV-induced liver injury is still under investigation. The mitogen activated protein kinases (MAPKs), including p38 MAPK, have roles in the hepatic cell apoptosis induced by DENV. However, the in vivo role of p38 MAPK in DENV-induced liver injury is not fully understood. In this study, we investigated the role of SB203580, a p38 MAPK inhibitor, in a mouse model of DENV infection. Both the hematological parameters, leucopenia and thrombocytopenia, were improved by SB203580 treatment and liver transaminases and histopathology were also improved. We used a real-time PCR microarray to profile the expression of apoptosis-related genes. Tumor necrosis factor α, caspase 9, caspase 8, and caspase 3 proteins were significantly lower in the SB203580-treated DENV-infected mice than that in the infected control mice. Increased expressions of cytokines including TNF-α, IL-6 and IL-10, and chemokines including RANTES and IP-10 in DENV infection were reduced by SB203580 treatment. DENV infection induced the phosphorylation of p38MAPK, and its downstream signals including MAPKAPK2, HSP27 and ATF-2. SB203580 treatment did not decrease the phosphorylation of p38 MAPK, but it significantly reduced the phosphorylation of MAPKAPK2, HSP27, and ATF2. Therefore, SB203580 modulates the downstream signals to p38 MAPK and reduces DENV-induced liver injury.