Pathogenesis Of Dengue Virus Infection Research Articles

Current understanding of the pathogenesis of Dengue virus infection and therapeutic diet for controlling associated symptoms

Current understanding of the pathogenesis of Dengue virus infection and therapeutic diet for controlling associated symptoms

The relationship between changes in IL-2 / IL-18 and liver enzyme with dengue severity.

A synergistic impact of virulence capacity of dengue viruses and host immune response contributes to the pathogenesis of dengue virus infection (DENV). Elevation of hepatic enzyme and cytokine storm is the major contributory factor producing severity. IL-2 and IL-18 both play a critical role in generating immunogenicity during viral infection. The goal of the study was to evaluate the correlation between alterations in IL-2 and IL-18 levels with alterations in hepatic enzymes in dengue-infected patients. Accordingly, a total of 91 NS1/IgM confirmed DENV infected patients were selected for the IL-2 and IL-18 evaluation using a standard ELISA assay. Biochemical, haematological parameters were recorded at the time of admission. Interestingly, raised levels of IL-2 (p<0.0001) and IL-18 (p<0.0001) was obtained in severe dengue as compared to non-severe dengue patients. A significant positive correlation was observed between IL and 2 levels and SGOT (r=0.3168 with p=0.002), SGPT (r=0.569 with p<0.0001). Similarly IL-18 was also highly associated with SGOT (r=0.387 with p=0.0002) and SGPT (r=0.407 with p<0.0001) in dengue infected patient.Our data reveal that a rising amount of IL-2 and IL-18 in initial stage of the disease could be predictors for developing severe form of dengue infection.

Association of single nucleotide polymorphisms in TNF-α (−308G/A and −238G/A) to dengue: Case-control and meta-analysis study

Dengue is an acute viral disease whose clinical condition is related to the interaction of factors related to the Dengue virus (DENV), environment and the host, with the immunity of the human host contributing a substantial role in the pathogenesis of DENV infection. Studies have demonstrated that single nucleotide polymorphisms (SNPs) in the promoter regions of cytokine genes such as tumor necrosis factor (TNF-α) affect transcription and/or expression; and therefore, may influence the pathogenesis of infectious diseases, such as dengue. Consequently, the objective of this study was to assess through a case-control study whether there was an association between the presence of SNPs −308G/A and −238G/A in the TNF-α gene and 158 patients with dengue and 123 controls. No association was found between the SNPs and the dengue cases in the study population. We then performed a meta-analysis, retrieving data from case-control studies in the literature for the same polymorphisms. For SNP-308G/A, the GG genotype was associated with dengue fever (DF) risk (OR = 1.24, 1.00–1.53; p = 0.05; I2 = 0%), while the GA genotype (OR = 0.75, 0.60–0.93; p = 0.01; I2 = 0%) and allele A (OR = 0.75, 0.60–0.93; p = 0.01; I2 = 0%) were associated with protection. The genotype GG population in the Asian continent (OR = 1.81 [1.06, 3.09], p = 0.03, I2 = 0%) and American (OR = 1.29 [1.00, 1.65], p = 0.05, I2 = 0%) was also associated with protection in the comparison between the cases versus the control group. In each comparison, the dominant model AA + GA (p < 0.00001) conferred protection. For SNP-238G/A the GA genotype was associated with risk for dengue hemorrhagic fever (DHF; OR = 2.17, 1.28–3.67; p = 0.004; I2 = 0%)), and the dominant AA + GA model (p < 0.00001) was associated with protection in each comparison. In summary, our results did not associate SNPs in the TNF-α gene to dengue in the Brazilian northeast population. However, combined literature data suggested the effect of the GG and GA genotypes of the SNP-308G/A on risk and protection, respectively, in Asian and American populations.

Association of FCN2 polymorphisms and Ficolin-2 levels with dengue fever in Vietnamese patients

Background and objectiveThe human ficolin-2, encoded by FCN2, recognizes pathogen-associated acetylated residues on their cell surfaces and activates the lectin complement cascade. This study aimed to investigate the contribution of human ficolin-2 and the functional FCN2 genetic variants in dengue virus (DENV) infection and in clinical progression. MethodsFCN2 genetic polymorphisms in the promoter, intron 7 and exon 8 were genotyped in 279 patients with dengue fever and in 200 healthy controls by direct Sanger sequencing. The ficolin-2 levels were measured in serum samples by ELISA and correlated with clinical data. ResultsThe frequencies of +6031GG, +6220GG and +6424TT genotypes were significantly higher in dengue patients compared to healthy controls indicating an increased risk of dengue fever. The SNPs rs11103563 (+6031A/G), rs7872508 (+6220 T/G), and rs7851696 (+6424G/T) significantly regulated ficolin-2 levels in dengue patients (P < 0.0001). Ficolin-2 levels were increased in patients with dengue and Dengue with Warning Signs (DWS) compared to healthy controls (P < 0.0001 and P = 0.038, respectively). Ficolin-2 levels were significantly increased after 10–14 days of admission in both dengue and DWS patients and then slightly decreased after three weeks of discharge, indicating that ficolin-2 levels were modulated during the progression of dengue fever. In addition, ficolin-2 levels were negatively correlated with AST levels and positively correlated with platelet counts. ConclusionsFCN2 polymorphisms are associated with dengue fever in the Vietnamese population. Ficolin-2 levels are modulated during the progression of dengue fever and correlated with clinical parameters and thus may play a possible role in the pathogenesis of DENV infection.

Role of NS1 antibodies in the pathogenesis of acute secondary dengue infection

The role of NS1-specific antibodies in the pathogenesis of dengue virus infection is poorly understood. Here we investigate the immunoglobulin responses of patients with dengue fever (DF) and dengue hemorrhagic fever (DHF) to NS1. Antibody responses to recombinant-NS1 are assessed in serum samples throughout illness of patients with acute secondary DENV1 and DENV2 infection by ELISA. NS1 antibody titres are significantly higher in patients with DHF compared to those with DF for both serotypes, during the critical phase of illness. Furthermore, during both acute secondary DENV1 and DENV2 infection, the antibody repertoire of DF and DHF patients is directed towards distinct regions of the NS1 protein. In addition, healthy individuals, with past non-severe dengue infection have a similar antibody repertoire as those with mild acute infection (DF). Therefore, antibodies that target specific NS1 epitopes could predict disease severity and be of potential benefit in aiding vaccine and treatment design.

Substitution of the precursor peptide prevents anti-prM antibody-mediated antibody-dependent enhancement of dengue virus infection

Antibody-dependent enhancement (ADE) is currently considered as the mechanism underlying the pathogenesis of severe dengue disease. Many studies have shown that precursor (pr) peptide-specific antibodies do not efficiently neutralize infection but potently promote ADE of dengue virus (DENV) infection. To explore the effect of pr peptide substitution on neutralization and ADE of DENV infection, the rabbit anti-prM polyclonal antibodies (pAbs) and anti-JEVpr/DENV-M pAbs were prepared, and the neutralization and ADE of these two pAbs were further compared. Here, we report that both anti-JEVpr/DENV-M and anti-prM pAbs exhibited broad cross-reactivity and only partial neutralization with four DENV serotypes and immature DENV. Rabbit anti-prM pAbs showed a significant enhancement in a broad range of serum dilutions. However, there was no statistically significant difference in the enhancing activity of rabbit anti-JEVpr/DENV-M pAbs at various levels of dilution. These results demonstrate that anti-prM antibody-mediated ADE can be prevented by JEV pr peptide replacement. The present study contribute further to research on the pathogenesis of DENV infection.

Levels of Circulating Tumor Necrosis Factor-α in Children with Symptomatic Dengue Evaluated by ELISA and Bead-Based Assays.

Tumor necrosis factor (TNF)-α is a key cytokine in the pathogenesis of dengue virus infection, and its accurate detection in several types of human samples is critical. The enzyme-linked immunosorbent assay (ELISA) is the gold standard for the detection of TNF-α, but multiplexed bead-based assays such as cytometric bead array (CBA) are now frequently used. Here, using ELISA and two CBAs commercially available, we measured TNF-α concentrations in plasma and serum from children with acute dengue virus infection and healthy controls. To evaluate the detection efficiency and factors affecting it, spiked recovery and immune complex dissociation assays were also performed. The levels of TNF-α evaluated by ELISA in paired serum and plasma samples from children with dengue positively correlated (rho = 0.99, p < 0.0001). Children with dengue had higher levels of plasma TNF-α than those of healthy children (p = 0.004). The ELISA detected TNF-α in a higher number of plasma samples than the CBA (p < 0.0001), and both methods only correlated when TNF-α was evaluated in buffer-based solutions but not in plasma, indicating the presence of a factor interfering with the detection of TNF-α in plasma. The recovery of several types of human recombinant TNF-α was dramatically decreased in plasma but not in tissue culture media (p ≤ 0.01), and this effect was similar in the plasma obtained from the children with dengue or the healthy controls. The dissociation of immune complexes did not improve TNF-α recovery. Dilution of the plasma samples increased the recovery of TNF-α, but at high concentrations of the cytokine. In short, plasma affects the efficiency of TNF-α detection, and this effect should be considered in the measurement of this cytokine.

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.

Comprehensive mapping infection-enhancing epitopes of dengue pr protein using polyclonal antibody against prM.

Dengue vaccine development is considered a global public health priority, but the antibody-dependent enhancement (ADE) issues have critically restricted vaccine development. Recent findings have demonstrated that pre-membrane (prM) protein was involved in dengue virus (DENV) infection enhancement. Although the importance of prM antibodies have been well characterized, only a few epitopes in DENV prM protein have ever been identified. In this study, we screened five potential linear epitopes located at positions pr1 (1-16aa), pr3 (13-28aa), pr4 (19-34aa), pr9 (49-64aa), and pr10 (55-70aa) in pr protein using peptide scanning and comprehensive bioinformatics analysis. Then, we found that only pr4 (19-34aa) could elicit high-titer antibodies in Balb/c mice, and this epitope could react with sera from DENV2-infected patients, suggesting that specific antibodies against epitope peptide pr4 were elicited in both DENV-infected mice and human. In addition, our data demonstrated that anti-pr4 sera showed limited neutralizing activity but significant ADE activity toward standard DENV serotypes and imDENV. Hence, it seems responsible to hypothesize that anti-pr4 serum was infection-enhancing antibody and pr4 was infection-enhancing epitope. In conclusion, we characterized a novel infection-enhancing epitope on dengue pr protein, a finding that may provide new insight into the pathogenesis of DENV infection and contribute to dengue vaccine design.

Pathogenic roles of macrophage migration inhibitory factor during dengue virus infection.

Dengue virus (DENV) infection is the most common cause of viral hemorrhagic fever, which can lead to life-threatening dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS). Hemorrhage and plasma leakage are two major hallmarks of DHF/DSS. Because the mechanisms causing these pathogenic changes are unclear, there is no effective therapy against DHF/DSS. In this review, we focus on the possible pathogenic effects of a pleiotropic cytokine, macrophage migration inhibitory factor (MIF), on the pathogenesis of DENV infection. MIF is a critical mediator of the host immune response and inflammation, and there is a correlation between the serum levels of MIF and disease severity in dengue patients. Furthermore, MIF knock-out mice exhibit less severe clinical disease and lethality. However, the role of MIF in the pathogenesis of DHF/DSS is not limited to immune cell recruitment. Recent evidence indicates that DENV infection induced MIF production and may contribute to vascular hyperpermeability and viral replication during DENV infection. The expression of both adhesion and coagulation molecules on MIF-stimulated monocytes and endothelial cells is also increased, which may contribute to inflammatory and anticoagulatory states during DHF/DSS. Therefore, blocking MIF production or its function may provide a solution for the treatment and prevention of DHF/DSS.

The Potent and Broadly Neutralizing Human Dengue Virus-Specific Monoclonal Antibody 1C19 Reveals a Unique Cross-Reactive Epitope on the bc Loop of Domain II of the Envelope Protein

ABSTRACTFollowing natural dengue virus (DENV) infection, humans produce some antibodies that recognize only the serotype of infection (type specific) and others that cross-react with all four serotypes (cross-reactive). Recent studies with human antibodies indicate that type-specific antibodies at high concentrations are often strongly neutralizing in vitro and protective in animal models. In general, cross-reactive antibodies are poorly neutralizing and can enhance the ability of DENV to infect Fc receptor-bearing cells under some conditions. Type-specific antibodies at low concentrations also may enhance infection. There is an urgent need to determine whether there are conserved antigenic sites that can be recognized by cross-reactive potently neutralizing antibodies. Here, we describe the isolation of a large panel of naturally occurring human monoclonal antibodies (MAbs) directed to the DENV domain II fusion loop (FL) envelope protein region from subjects following vaccination or natural infection. Most of the FL-specific antibodies exhibited a conventional phenotype, characterized by low-potency neutralizing function and antibody-dependent enhancing activity. One clone, however, recognized the bc loop of domain II adjacent to the FL and exhibited a unique phenotype of ultrahigh potency, neutralizing all four serotypes better than any other previously described MAb recognizing this region. This antibody not only neutralized DENV effectively but also competed for binding against the more prevalent poor-quality antibodies whose binding was focused on the FL. The 1C19 human antibody could be a promising component of a preventative or therapeutic intervention. Furthermore, the unique epitope revealed by 1C19 suggests a focus for rational vaccine design based on novel immunogens presenting cross-reactive neutralizing determinants.

Challenges for the formulation of a universal vaccine against dengue

Dengue is rapidly becoming a disease of an escalating global public health concern. The disease is a vector-borne disease, transmitted by the bite of an Aedes spp. mosquito. Dynamic clinical manifestations, ranging from asymptomatic, flu-like febrile illness, dengue fever (DF) to dengue hemorrhagic fever (DHF) with or without dengue shock syndrome (DSS), make the disease one of the most challenging to diagnose and treat. DF is a self-limited illness, while DHF/DSS, characterized by plasma leakage resulting from an increased vascular permeability, can have severe consequences, including death. The pathogenesis of dengue virus infection remains poorly understood, mainly due to the lack of a suitable animal model that can recapitulate the cardinal features of human dengue diseases. Currently, there is no specific treatment or antiviral therapy available for dengue virus infection and supportive care with vigilant monitoring is the principle course of treatment. Since vector control programs have been largely unsuccessful in preventing outbreaks, vaccination seems to be the most viable option for prevention. There are four dengue viral serotypes and each one of them is capable of causing severe dengue. Although immunity induced by infection by one serotype is effective in protection against the homologous viral serotype, it only has a transient protective effect against infection with the other three serotypes. The meager cross protective immunity generated wanes over time and may even induce a harmful effect at the time of subsequent secondary infection. Thus, it is imperative to have a vaccine that can elicit equal and long-lasting immunity to all four serotypes simultaneously. Numerous tetravalent vaccines are currently either in the pipeline for clinical trials or under development. For those frontrunner tetravalent vaccines in clinical trials, despite good safety and immunogenicity profiles registered, issues such as imbalanced immune responses between serotypes and questions with regard to whether the optimum formulation have been identified remain unresolved. This review centers on these issues and offers strategies that may improve the tetravalent vaccine formulation.

Mapping the Interactions of Dengue Virus NS1 Protein with Human Liver Proteins Using a Yeast Two-Hybrid System: Identification of C1q as an Interacting Partner

Dengue constitutes a global health concern. The clinical manifestation of this disease varies from mild febrile illness to severe hemorrhage and/or fatal hypovolemic shock. Flavivirus nonstructural protein 1 (NS1) is a secreted glycoprotein that is displayed on the surface of infected cells but is absent in viral particles. NS1 accumulates at high levels in the plasma of dengue virus (DENV)-infected patients, and previous reports highlight its involvement in immune evasion, dengue severity, liver dysfunction and pathogenesis. In the present study, we performed a yeast two-hybrid screen to search for DENV2 NS1-interacting partners using a human liver cDNA library. We identified fifty genes, including human complement component 1 (C1q), which was confirmed by coimmunoprecipitation, ELISA and immunofluorescence assays, revealing for the first time the direct binding of this protein to NS1. Furthermore, the majority of the identified genes encode proteins that are secreted into the plasma of patients, and most of these proteins are classified as acute-phase proteins (APPs), such as plasminogen, haptoglobin, hemopexin, α-2-HS-glycoprotein, retinol binding protein 4, transferrin, and C4. The results presented here confirm the direct interaction of DENV NS1 with a key protein of the complement system and suggest a role for this complement protein in the pathogenesis of DENV infection.

CLEC5A is critical for dengue virus–induced inflammasome activation in human macrophages

AbstractPersistent high fever is one of the most typical clinical symptoms in dengue virus (DV)–infected patients. However, the source of endogenous pyrogen (eg, IL-1β) and the signaling cascade leading to the activation of inflammasome and caspase-1, which are essential for IL-1β and IL-18 secretion, during dengue infection have not been elucidated yet. Macrophages can be polarized into distinct phenotypes under the influence of GM-CSF or M-CSF, denoted as GM-Mφ and M-Mφ, respectively. We found that DV induced high levels of IL-1β and IL-18 from GM-Mφ (inflammatory macrophage) and caused cell death (pyroptosis), whereas M-Mφ (resting macrophage) did not produce IL-1β and IL-18 on DV infection even with lipopolysaccharide priming. This observation demonstrates the distinct responses of GM-Mφ and M-Mφ to DV infection. Moreover, up-regulation of pro-IL-1β, pro-IL-18, and NLRP3 associated with caspase-1 activation was observed in DV-infected GM-Mφ, whereas blockade of CLEC5A/MDL-1, a C-type lectin critical for dengue hemorrhagic fever and Japanese encephalitis virus infection, inhibits NLRP3 inflammasome activation and pyrotopsis in GM-Mφ. Thus, DV can activate NLRP3 inflammasome via CLEC5A, and GM-Mφ plays a more important role than M-Mφ in the pathogenesis of DV infection.

P034 Inhibition of macrophage migration inhibitory factor reduces dengue virus replication

Introduction Dengue virus (DENV) infection can cause life-threatening dengue hemorrhage fever and shock syndrome (DHF/DSS). Currently there is no effective treatment to prevent the development of DHF/DSS, except supporting care. Macrophage migration inhibitory factor (MIF) is a pro-inflammatory cytokine that plays important roles in the pathogenesis of DHF/DSS (1). Because autophagy, which can be induced by reactive oxygen species (ROS), is involved in DENV replication (2), the effect of MIF and ROS on DENV replication in human hepatoma cell line Huh-7 cells was studied. Methods MIF inhibitor, ISO-1 as well as ROS scavenger N-acetyl-L-cysteine (NAC) were applied in DENV-infected Huh-7 cells. In addition, MIF knockdown Huh-7 cells were generated by shRNA transfection to further confirm MIF is involved in DENV infection-induced autophagy. DENV replication was determined by secreted alkaline phosphatase (SEAP) assay as well as flow cytometry. Results We demonstrated that, in the presence of ISO-1 as well as NAC, DENV replication in Huh-7 cells was reduced. Diminished MIF expression by shRNA transfection further confirmed DENV replication and autophagy-induced by DENV were reduced in MIF knockdown cells. Conclusion Taken together, these data suggest that therapeutically blocking MIF by its inhibitor or antibody may not only reduce proinflammatory response but also virus replication during DENV infection. References (1) Assuncao-Miranda I, Amaral FA, et al. Contribution of macrophage migration inhibitory factor to the pathogenesis of dengue virus infection. FASEB J 2010;24(1):218–28. (2) Lee YR, Lei HY, et al. Autophagic machinery activated by dengue virus enhances virus replication. Virology 2008;374(2):240–8.

The Relation Between Levels of TNF Alpha, IL-1B, PGE2 and PLA with the Severity Degree of Dengue Hemorrhagic

The pathogenesis of dengue virus infection is still being debated. Based on the existing data, there is a strong evidence that the immunopathological mechanism plays a role in dengue virus infection with various complications. Some unknown immune responses play a role in the pathogenesis of dengue virus infection. Researchers are trying to establish the role of several inflammatory mediators such as PLA2, IL-1, TNF-, PGE2, PGI2, Thromboxane A2, Leucotrien and MPTP, in relation to the severity degree of the dengue virus infection. The aim of this study is to recognize the relation between the severity degrees of DHF (Dengue Hemorrhagic Fever) patients and the immunological profile in the sub-cellular level, such as PLA2, IL-1, TNF-, PGE2 and PLA 2. The collected data was processed and presented analytically. The relation between each parameter (TNFα, SPLA2, PGE2,IL-1β) and the degree of DHF was analyzes, using Spearman's correlation analysis, ordinal regression, and using ANOVA. It was shown that there was no relation between the levels of TNF-ά, PGE2, IL-1β, and SPLA2 in patients with various degrees of DHF, but there were significant differences between DHF grade 1 and 3, and also 2 and 3, on IL-1β. There were increased levels of the four parameters in dengue grade 1 to 2, but decreased levels in grade 3. This can be caused by inflammatory processes, but the severity degree of DHF can also be influenced by complement, thromboxane, and leukotrien.

The Role Activity of Complement, TNFα &amp; IL12 in Pathogenesis Dengue Virus

Dengue Virus infection is always found in some part of the world especially South East Asia including Indonesia. The pathogenesis of Dengue Virus infection is still controversial. The aim of this study is to analyze the role complement activity, TNFα &amp; IL12 in Dengue Virus infection especially in pathogenesis of Dengue Virus infection. Cross sectional study had been done since February 2009 in Dr. Soetomo Hospital Surabaya. Blood Sera of Dengue Virus infection were collected from Dengue Fever, and Dengue Hemorrhagic Fever patient who had been care in Paediatric. Dengue patients and time schedule for taking blood sample for examination CH50, TNFα &amp; IL12 as follow: on the first day on admission, the second day, the third day. Study groups of patients as follow: Dengue Fever, 36; Dengue Hemorrhagic Fever grade I, 37; Dengue Hemorrhagic Fever grade II, 10; Dengue Hemorrhagic Fever grade III, 18; Dengue Hemorrhagic Fever grade IV, 6. In this study found that the higher activity complement which lower level CH50 was more identified on Dengue Shock Syndrome and Dengue Hemorrhagic Fever grade III than Dengue Fever cases. A concept of our study was focusing on manifestation of vascular leakage, measurement of complement activity CH50, TNFα &amp; IL12 and clinical manifestation Dengue Hemorrhagic Fever. The examination of TNFα &amp; IL12 in our study supported the role the activity complement. The conclusion are measurement CH50, TNFα &amp; IL12 can be used as a predictive factor of the degree of Dengue Virus infection

Development of an antibody-dependent enhancement assay for dengue virus using stable BHK-21 cell lines expressing FcγRIIA

Dengue virus (DENV) causes a wide range of symptoms, from mild febrile illness, dengue fever (DF), to severe life threatening illness, dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). Subneutralizing concentrations of antibody to DENV enhance DENV infection in FcγR positive cells. This phenomenon is known as antibody-dependent enhancement (ADE). ADE is considered to be a risk factor for DHF and DSS. To develop an ADE assay for DENV, two stable BHK-21 cell lines were established that express FcγRIIA (BHK-FcγRIIA). The BHK-FcγRIIA cell lines were used in an ADE assay with monoclonal antibody (4G2) to DENV, and DENV antibody-positive human sera. Virus growth was quantified directly in BHK-FcγRIIA cells with a standard plaque assay procedure. ADE was detected with monoclonal antibody (4G2) to DENV. ADE was also detected with DENV antibody-positive human sera, but not with DENV antibody-negative human sera. The new ADE assay using BHK-FcγRIIA cells is simple and practical, and is useful for defining the role of ADE in the pathogenesis of DENV infection.

Contribution of macrophage migration inhibitory factor to the pathogenesis of dengue virus infection

Dengue fever is an emerging viral disease transmitted by arthropods to humans in tropical countries. Dengue hemorrhagic fever (DHF) is escalating in frequency and mortality rates. Here we studied the involvement of macrophage migration inhibitory factor (MIF) in dengue virus (DENV) infection and its pathogenesis. Patients with DHF had elevated plasma concentrations of MIF. Both leukocytes from these patients and macrophages from healthy donors infected in vitro with DENV showed a substantial amount of MIF within lipid droplets. The secretion of MIF by macrophages and hepatocytes required a productive infection and occurred without an increase in gene transcription or cell death, thus indicating active secretion from preformed stocks. In vivo infection of wild-type and mif-deficient (Mif(-/-)) mice demonstrated a role of MIF in dengue pathogenesis. Clinical disease was less severe in Mif(-/-) mice, and they exhibited a significant delay in lethality, lower viremia, and lower viral load in the spleen than wild-type mice. This reduction in all parameters of severity on DENV infection in Mif(-/-) mice correlated with reduced proinflammatory cytokine concentrations. These results demonstrated the contribution of MIF to the pathogenesis of dengue and pointed to a possible beneficial role of neutralizing MIF as an adjunctive therapeutic approach to treat the severe forms of the disease.

Essential role of platelet-activating factor receptor in the pathogenesis of Dengue virus infection

Severe dengue infection in humans causes a disease characterized by thrombocytopenia, increased levels of cytokines, increased vascular permeability, hemorrhage, and shock. Treatment is supportive. Activation of platelet-activating factor (PAF) receptor (PAFR) on endothelial cells and leukocytes induces increase in vascular permeability, hypotension, and production of cytokines. We hypothesized that activation of PAFR could account for the major systemic manifestations of dengue infection. Inoculation of adult mice with an adapted strain of Dengue virus caused a systemic disease, with several features of the infection in humans. In PAFR(-/-) mice, there was decreased thrombocytopenia, hemoconcentration, decreased systemic levels of cytokines, and delay of lethality, when compared with WT infected mice. Treatment with UK-74,505, an orally active PAFR antagonist, prevented the above-mentioned manifestations, as well as hypotension and increased vascular permeability, and decreased lethality, even when started 5 days after virus inoculation. Similar results were obtained with a distinct PAFR antagonist, PCA-4246. Despite decreased disease manifestation, viral loads were similar (PAFR(-/-)) or lower (PAFR antagonist) than in WT mice. Thus, activation of PAFR plays a major role in the pathogenesis of experimental dengue infection, and its blockade prevents more severe disease manifestation after infection with no increase in systemic viral titers, suggesting that there is no interference in the ability of the murine host to deal with the infection. PAFR antagonists are disease-modifying agents in experimental dengue infection.