Increased pro-inflammatory cytokines (TNF-alpha and IL-6) and anti-inflammatory compounds (sTNFRp55 and sTNFRp75) in Brazilian patients during exanthematic dengue fever.

Pro-inflammatory cytokines, tumor necrosis factor (TNF-alpha), interleukin-6 (IL-6) and interleukin-1beta (IL-1beta) as well as anti-inflammatory compounds, soluble TNF-Receptor p55 (sTNFRp55), sTNFRp75 and IL-1 receptor antagonist (sIL-1Ra), were investigated in 34 Brazilian cases of dengue fever (DF) originated from a study of exanthematic virosis. The presence of pro-inflammatory cytokines was detected in sera from these patients by ELISA. TNF-alpha and IL-6 levels were significantly higher than control subjects in 32% and 52% patients, respectively. To our knowledge this was the first time a receptor antagonist and soluble receptors for cytokines were detected in sera obtained during exanthematic DF without hemorrhagic manifestations. Both sTNFRp55 and sTNFRp75 were consistently elevated in 42% and 84% patients, respectively. Most patients had IL-1beta levels not different from those of normal subjects, except for one case. Only 16% patients had altered levels of IL-1Ra. Previous studies in dengue hemorrhagic fever patients demonstrated production of these soluble factors; here we observed that they are found in absence of hemorrhagic manifestations. The possible role of these anti-inflammatory compounds in immune cell activation and in regulating cytokine-mediated pathogenesis during dengue infection is discussed.     

Immunopathogenesis of dengue virus infection

Dengue virus infection causes dengue fever (DF), dengue hemorrhagic fever (DHF), and dengue shock syndrome (DSS), whose pathogeneses are not clearly understood. Current hypotheses of antibody-dependent enhancement, virus virulence, and IFN-gamma/TNFalpha-mediated immunopathogenesis are insufficient to explain clinical manifestations of DHF/DSS such as thrombocytopenia and hemoconcentration. Dengue virus infection induces transient immune aberrant activation of CD4/CD8 ratio inversion and cytokine overproduction, and infection of endothelial cells and hepatocytes causes apoptosis and dysfunction of these cells. The coagulation and fibrinolysis systems are also activated after dengue virus infection. We propose a new hypothesis for the immunopathogenesis for dengue virus infection. The aberrant immune responses not only impair the immune response to clear the virus, but also result in overproduction of cytokines that affect monocytes, endothelial cells, and hepatocytes. Platelets are destroyed by crossreactive anti-platelet autoantibodies. Dengue-virus-induced vasculopathy and coagulopathy must be involved in the pathogenesis of hemorrhage, and the unbalance between coagulation and fibrinolysis activation increases the likelihood of severe hemorrhage in DHF/DSS. Hemostasis is maintained unless the dysregulation of coagulation and fibrinolysis persists. The overproduced IL-6 might play a crucial role in the enhanced production of anti-platelet or anti-endothelial cell autoantibodies, elevated levels of tPA, as well as a deficiency in coagulation. Capillary leakage is triggered by the dengue virus itself or by antibodies to its antigens. This immunopathogenesis of DHF/DSS can account for specific characteristics of clinical, pathologic, and epidemiological observations in dengue virus infection.     

Detection of circulant tumor necrosis factor-alpha, soluble tumor necrosis factor p75 and interferon-gamma in Brazilian patients with dengue fever and dengue hemorrhagic fever

Pro-inflammatory cytokines are believed to play an important role in the pathogenesis of dengue infection. This study reports cytokine levels in a total of 54 patients examined in Recife, State of Pernambuco, Brazil. Five out of eight patients who had hemorrhagic manifestations presented tumor necrosis factor-alpha (TNF-alpha) levels in sera which were statistically higher than those recorded for controls. In contrast, only one out of 16 patients with mild manifestations had elevated TNF-alpha levels. The levels of interleukin-6 (IL), IL-1beta tested in 24 samples and IL-12 in 30 samples were not significantly increased. Interferon-g was present in 10 out of 30 patients with dengue. The data support the concept that the increased level of TNF-alpha is related to the severity of the disease. Soluble TNF receptor p75 was found in most patients but it is unlikely to be related to severity since it was found with an equivalent frequency and levels in 15 patients with dengue fever and another 15 with dengue hemorrhagic fever.     

Immune Response to Dengue Virus Infection in Pediatric Patients in New Delhi,India—Association of Viremia,Inflammatory Mediators and Monocytes with Disease Severity

Dengue virus, a mosquito-borne flavivirus, is a causative agent for dengue infection, which manifests with symptoms ranging from mild fever to fatal dengue shock syndrome. The presence of four serotypes, against which immune cross-protection is short-lived and serotype cross-reactive antibodies that might enhance infection, pose a challenge to further investigate the role of virus and immune response in pathogenesis. We evaluated the viral and immunological factors that correlate with severe dengue disease in a cohort of pediatric dengue patients in New Delhi. Severe dengue disease was observed in both primary and secondary infections. Viral load had no association with disease severity but high viral load correlated with prolonged thrombocytopenia and delayed recovery. Severe dengue cases had low Th1 cytokines and a concurrent increase in the inflammatory mediators such as IL-6, IL-8 and IL-10. A transient increase in CD14⁺CD16⁺ intermediate monocytes was observed early in infection. Sorting of monocytes from dengue patient peripheral blood mononuclear cells revealed that it is the CD14+ cells, but not the CD16+ or the T or B cells, that were infected with dengue virus and were major producers of IL-10. Using the Boruta algorithm, reduced interferon-α levels and enhanced aforementioned pro-inflammatory cytokines were identified as some of the distinctive markers of severe dengue. Furthermore, the reduction in the levels of IL-8 and IL-10 were identified as the most significant markers of recovery from severe disease. Our results provide further insights into the immune response of children to primary and secondary dengue infection and help us to understand the complex interplay between the intrinsic factors in dengue pathogenesis.     

Endothelial cells elicit immune-enhancing responses to dengue virus infection

Dengue viruses cause two severe diseases that alter vascular fluid barrier functions, dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). Preexisting antibodies to dengue virus disposes patients to immune-enhanced edema (DSS) or hemorrhagic (DHF) disease following infection by a discrete dengue virus serotype. Although the endothelium is the primary vascular fluid barrier, direct effects of dengue virus on endothelial cells (ECs) have not been considered primary factors in pathogenesis. Here, we show that dengue virus infection of human ECs elicits immune-enhancing EC responses. Our results suggest that rapid early dengue virus proliferation within ECs is permitted by dengue virus regulation of early, but not late, beta interferon (IFN-β) responses. The analysis of EC responses following synchronous dengue virus infection revealed the high-level induction and secretion of immune cells (T cells, B cells, and mast cells) as well as activating and recruiting cytokines BAFF (119-fold), IL-6/8 (4- to 7-fold), CXCL9/10/11 (45- to 338-fold), RANTES (724-fold), and interleukin-7 (IL-7; 128-fold). Moreover, we found that properdin factor B, an alternative pathway complement activator that directs chemotactic anaphylatoxin C3a and C5a production, was induced 34-fold. Thus, dengue virus-infected ECs evoke key inflammatory responses observed in dengue virus patients which are linked to DHF and DSS. Our findings suggest that dengue virus-infected ECs directly contribute to immune enhancement, capillary permeability, viremia, and immune targeting of the endothelium. These data implicate EC responses in dengue virus pathogenesis and further rationalize therapeutic targeting of the endothelium as a means of reducing the severity of dengue virus disease.     

Source and Purity of Dengue-Viral Preparations Impact Requirement for Enhancing Antibody to Induce Elevated IL-1β Secretion: A Primary Human Monocyte Model

Dengue virus is a major global health threat and can lead to life-threatening hemorrhagic complications due to immune activation and cytokine production. Cross-reactive antibodies to an earlier dengue virus infection are a recognized risk factor for severe disease. These antibodies bind heterologous dengue serotypes and enhance infection into Fc-receptor-bearing cells, a process known as antibody-dependent enhancement of infection. One crucial cytokine seen elevated in severe dengue patients is IL-1β, a potent inflammatory cytokine matured by the inflammasome. We used a highly-physiologic system by studying antibody-dependent enhancement of IL-1β in primary human monocytes with anti-dengue human monoclonal antibodies isolated from patients. Antibody-enhancement increased viral replication in primary human monocytes inoculated with supernatant harvested from Vero cells infected with dengue virus serotype 2 (DENV-2) 16681. Surprisingly, IL-1β secretion induced by infectious supernatant harvested from two independent Vero cell lines was not enhanced by antibody. Secretion of multiple other inflammatory cytokines was also independent of antibody signaling. However, IL-1β secretion did require NLRP3 and caspase-1 activity. Immunodepletion of dengue virions from the infectious supernatant confirmed that virus was not the main IL-1β-inducing agent, suggesting that a supernatant component(s) not associated with the virion induced IL-1β production. We excluded RNA, DNA, contaminating LPS, viral NS1 protein, complement, and cytokines. In contrast, purified Vero-derived DENV-2 16681 exhibited antibody-enhancement of both infection and IL-1β induction. Furthermore, C6/36 mosquito cells did not produce such an inflammatory component, as crude supernatant harvested from insect cells infected with DENV-2 16681 induced antibody-dependent IL-1β secretion. This study indicates that Vero cells infected with DENV-2 16681 may produce inflammatory components during dengue virus propagation that mask the virus-specific immune response. Thus, the choice of host cell and viral purity should be carefully considered, while insect-derived virus represents a system that elicits antibody-dependent cytokine responses to dengue virus with fewer confounding issues.     

Serum Proteome and Cytokine Analysis in a Longitudinal Cohort of Adults with Primary Dengue Infection Reveals Predictive Markers of DHF

Background

Infections caused by dengue virus are a major cause of morbidity and mortality in tropical and subtropical regions of the world. Factors that control transition from mild forms of disease such as dengue fever (DF) to more life-threatening forms such as dengue hemorrhagic fever (DHF) are poorly understood. Consequently, there are no reliable methods currently available for early triage of DHF patients resulting in significant over-hospitalization.

Methodology/Principal Findings

We have systematically examined the proteome, cytokines and inflammatory markers in sera from 62 adult dengue patients (44 DF; 18 DHF) with primary DENV infection, at three different times of infection representing the early febrile, defervescence and convalescent stages. Using fluorescent bioplex assays, we measured 27 cytokines in these serum samples. Additionally, we used multiple mass spectrometry methods for iTRAQ-based comparative analysis of serum proteome as well as measurements of protein adducts- 3-nitrotyrosine and 3-chlorotyrosine as surrogate measures of free radical activity. Using multiple methods such as OPLS, MRMR and MSVM-RFE for multivariate feature selection and classification, we report molecular markers that allow prediction of primary DHF with sensitivity and specificity of >80%.

Conclusions/Significance

This report constitutes a comprehensive analysis of molecular signatures of dengue disease progression and will help unravel mechanisms of dengue disease progression. Our analysis resulted in the identification of markers that may be useful for early prediction of DHF during the febrile phase. The combination of highly sensitive analytical methods and novel statistical approaches described here forms a robust platform for biomarker discovery.     

IL-33/ST2 Correlates with Severity of Haemorrhagic Fever with Renal Syndrome and Regulates the Inflammatory Response in Hantaan Virus-Infected Endothelial Cells

Background

Hantaan virus (HTNV) causes a severe lethal haemorrhagic fever with renal syndrome (HFRS) in humans. Despite a limited understanding of the pathogenesis of HFRS, the importance of the abundant production of pro-inflammatory cytokines has been widely recognized. Interleukin 33 (IL-33) has been demonstrated to play an important role in physiological and pathological immune responses. After binding to its receptor ST2L, IL-33 stimulates the Th2-type immune response and promotes cytokine production. Depending on the disease model, IL-33 either protects against infection or exacerbates inflammatory disease, but it is unknown how the IL-33/ST2 axis regulates the immune response during HTNV infection.

Methodology/Principal Findings

Blood samples were collected from 23 hospitalized patients and 28 healthy controls. The levels of IL-33 and soluble ST2 (sST2) in plasma were quantified by ELISA, and the relationship between IL-33, sST2 and the disease severity was analyzed. The role of IL-33/sST2 axis in the production of pro-inflammatory cytokines was studied on HTNV-infected endothelial cells. The results showed that the plasma IL-33 and sST2 were significantly higher in patients than in healthy controls. Spearman analysis showed that elevated IL-33 and sST2 levels were positively correlated with white blood cell count and viral load, while negatively correlated with platelet count. Furthermore, we found that IL-33 enhanced the production of pro-inflammatory cytokines in HTNV-infected endothelial cells through NF-κB pathway and that this process was inhibited by the recombinant sST2.

Conclusion/Significance

Our results indicate that the IL-33 acts as an initiator of the “cytokine storm” during HTNV infection, while sST2 can inhibit this process. Our findings could provide a promising immunotherapeutic target for the disease control.     

Differential Expression of Toll-like Receptors in Dendritic Cells of Patients with Dengue during Early and Late Acute Phases of the Disease

Background

Dengue hemorrhagic fever (DHF) is observed in individuals that have pre-existing heterotypic dengue antibodies and is associated with increased viral load and high levels of pro-inflammatory cytokines early in infection. Interestingly, a recent study showed that dengue virus infection in the presence of antibodies resulted in poor stimulation of Toll-like receptors (TLRs), thereby facilitating virus particle production, and also suggesting that TLRs may contribute to disease pathogenesis.

Methodology/Principal Findings

We evaluated the expression levels of TLR2, 3, 4 and 9 and the co-stimulatory molecules CD80 and CD86 by flow cytometry. This was evaluated in monocytes, in myeloid and plasmacytoid dendritic cells (mDCs and pDCs) from 30 dengue patients with different clinical outcomes and in 20 healthy controls. Increased expression of TLR3 and TLR9 in DCs of patients with dengue fever (DF) early in infection was detected. In DCs from patients with severe manifestations, poor stimulation of TLR3 and TLR9 was observed. In addition, we found a lower expression of TLR2 in patients with DF compared to DHF. Expression levels of TLR4 were not affected. Furthermore, the expression of CD80 and CD86 was altered in mDCs and CD86 in pDCs of severe dengue cases. We show that interferon alpha production decreased in the presence of dengue virus after stimulation of PBMCs with the TLR9 agonist (CpG A). This suggests that the virus can affect the interferon response through this signaling pathway.

Conclusions/Significance

These results show that during dengue disease progression, the expression profile of TLRs changes depending on the severity of the disease. Changes in TLRs expression could play a central role in DC activation, thereby influencing the innate immune response.     

Role of mitogen-activated protein kinase signaling in the pathogenesis of dengue virus infection

Dengue virus (DENV) infection is a disease that is endemic to many parts of the world, and its increasing prevalence ranks it among the diseases considered to be a significant threat to public health. The clinical manifestations of DENV infection range from mild dengue fever (DF) to more severe dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). Increased proinflammatory cytokines and vascular permeability, both of which cause organ injury, are the hallmarks of severe dengue disease. Signs of liver injury were observed in studies using hepatic cell lines, mouse models, and autopsy specimens from DENV-infected patients, and these signs substantiated the effects of inflammatory responses and hepatic cell apoptosis. Mitogen-activated protein kinases (MAPK) are involved in inflammatory responses and cellular stress during viral infections. The roles of MAPK signaling in DENV infection were reviewed, and published data indicate MAPK signaling to be involved in inflammatory responses and hepatic cell apoptosis in both in vitro cultures and in vivo models. Modulation of MAPK signaling ameliorates the inflammatory responses and hepatic cell apoptosis in DENV infection. This accumulation of published data relative to the role of MAPK signaling in inflammatory responses and cell apoptosis in DENV infection is elucidatory, and may help to accelerate the development of novel or repositioned therapies to treat this unpredictable and often debilitating disease.     

Elevated levels of interleukin-13 and IL-18 in patients with dengue hemorrhagic fever

Interleukin (IL)-13 is produced by T helper 2 (Th2)-type cells and inhibits the production of proinflammatory cytokines by activated monocytes, while IL-18 is a pleiotropic cytokine that induces interferon-gamma and plays an important role in the development of Th1-type cells. Role of the shift from a Th1-type response to Th2-type has been suggested in the pathogenesis of dengue hemorrhagic fever (DHF). This study was undertaken to investigate the possible protective/pathogenic role of IL-13 and IL-18 in patients with DHF. Sera were collected from a total of 84 patients with various grades of dengue illness and 21 normal healthy controls and tested for IL-13 and IL-18 levels using commercial enzyme-linked immunosorbent assay kits. The results showed that very low levels of IL-13 (4+/-3 pg ml(-1)) and IL-18 (15+/-4 pg ml(-1)) were detected in the sera of healthy controls. In dengue patients, the levels of IL-13 and IL-18 were the highest in the patients with DHF grade IV (205+/-103 pg ml(-1) and 366+/-155 pg ml(-1), respectively) and the lowest in patients with dengue fever (22+/-12 pg ml(-1) and 76+/-50 pg ml(-1), respectively). Both the cytokines appeared (IL-13=20+/-11 pg ml(-1) and IL-18=70+/-45 pg ml(-1)) during the first 4 days of illness and reached peak levels (IL-13=204+/-96 pg ml(-1) and IL-18=360+/-148 pg ml(-1)) by day 9 onwards. The presence of high levels of IL-13 and IL-18 during severe illness and late phases of the disease suggests that both of these cytokines may contribute to the shift from a Th1- to Th2-type response and thus to the pathogenesis of DHF.     

Dengue Virus Infection Mediates HMGB1 Release from Monocytes Involving PCAF Acetylase Complex and Induces Vascular Leakage in Endothelial Cells

High mobility group box 1 (HMGB1) protein is released from cells as a pro-inflammatory cytokine in response to an injury or infection. During dengue hemorrhagic fever (DHF)/dengue shock syndrome (DSS), a number of pro-inflammatory cytokines are released, contributing to disease pathogenesis. In this study, the release of HMGB1 from human myelogenous leukemia cell line K562 and primary peripheral blood monocytes (PBM) cells was examined during dengue virus (DV)-infection. HMGB1 was shown to translocate from cell nuclei to the cytoplasm in both K562- and PBM-infected cells. The translocation of HMGB1 from the nucleus to the cytoplasm was shown to be mediated by the host cell p300/CBP-associated factor (PCAF) acetylase complex in K562 cells. In addition, DV capsid protein was observed to be the putative viral protein in actuating HMGB1 migration from the nucleus to cytoplasm through the involvement of PCAF acetylase. HMGB1 was released from DV-infected K562 cells into the extracellular milieu in a multiplicity of infection (M.O.I.)-independent manner and its release can be inhibited by the addition of 1-5 mM of ethyl pyruvate (EP) in a dose-dependent manner. Application of DV-infected K562 cell culture supernatants to primary endothelial cells induced vascular permeability. In contrast, supernatants from DV-infected K562 cells treated with EP or HMGB1 neutralizing antibody were observed to maintain the structural integrity of the vascular barrier.     

Secondary heterologous dengue infection risk: Disequilibrium between immune regulation and inflammation?

Increased serum levels of cytokines released by cells of the immune response have been detected in patients suffering from dengue disease. Likewise, secondary infections by a different dengue virus serotype result in a highest risk of development of the severe dengue disease. Both findings suggest that the memory immune response is one of the key players in the pathogenesis of this disease. Here we take advantage of the particular Cuban epidemiological situation in dengue to analyze a broad spectrum of cell-mediated immune response mediators at mRNA and protein level. Evidences for a regulatory immune pattern in homologous (TGF-β, IL-10) vs. pro-inflammatory pattern (IFN-γ, TNF-α) in heterologous dengue virus re-challenge were found, suggesting a possible association with the higher incidence of severe dengue cases in the latter case.     

High plasma levels of soluble ST2 but not its ligand IL-33 is associated with severe forms of pediatric dengue

Identification of early determinants of dengue disease progression, which could potentially enable individualized patient care are needed at present times. Soluble ST2 (sST2) has been recently reported to be elevated in the serum of children older than 2 years old and adults with dengue infection and it was correlated with secondary infections as well as with severe presentations of the disease. The mechanism by which secreted ST2 is linked to severe dengue and plasma leakage remains unclear. One possibility is that IL-33 ligand may be elevated, contributing to membrane bound ST2 as part of the immune activation in dengue infection. We determined plasma levels of sST2 and the ligand IL-33 in 66 children with acute secondary dengue infections clinically classified using the guidelines of the World Health Organization, 2009. Dengue infection showed significant increases in cytokines IL-12p70, IL-10, IL-8, IL-6, IL-1β and TNFα measured by flow cytometry based assay compared to uninfected individuals. In contrast, IL-33 levels remained unchanged between infected and uninfected individuals. The levels of sST2 positively correlated with values of IL-6 and IL-8 and inversely correlated with number of median value of platelet levels. In addition to circulating cytokine positive correlations we found that sST2 and isoenzyme creatine kinase-MB (CK-MB), a marker of myocardial muscle damage present in severe dengue cases were associated. Our pediatric study concluded that in dengue infections sST2 elevation does not involve concomitant changes of IL-33 ligand. We propose a study to assess its value as a predictor factor of disease severity.     

老年大鼠神经组织星形胶质细胞的分选培养与炎性特征分析*

目的: 建立优化的年轻与老年大鼠神经组织星形胶质细胞的分离纯化方法,比较年轻大鼠与老年大鼠星形胶质细胞的形态、功能差异,探讨老化后星形胶质细胞的功能改变及其在衰老过程中发挥的可能机制。方法: 采用50%-35%的percoll密度梯度离心法分选年轻(2月龄)和老年(20月龄)SD大鼠的大脑与脊髓星形胶质细胞;每组细胞设置3个复孔,培养72 h后,采用免疫荧光检测星形胶质细胞特异性标志物胶质纤维酸性蛋白(GFAP),观察不同年龄阶段星形胶质细胞的形态特征;qPCR检测衰老标志(p16、p21)的表达,β-半乳糖苷酶染色检测星形胶质细胞的衰老情况;qPCR检测促炎因子(IL-1β、TNF-α)与抗炎因子(IL-10)的表达水平。结果: 采用50%-35%的percoll梯度分选得到的星形胶质细胞的数量多、活性好、纯度高达95%以上,可用于后续实验。与年轻大鼠神经组织的星形胶质细胞相比,分选自老年大鼠神经组织的星形胶质细胞在细胞形态上偏向激活态,突起较少;星形胶质细胞β-半乳糖苷酶染色阳性率升高,p16、p21表达也明显增多(P<0.01);老年大鼠神经组织的星形胶质细胞的促炎因子(IL-1β、TNF-α)表达升高(P<0.05),抗炎因子(IL-10)表达有所降低(P<0.05)。结论: 50%-35%的percoll梯度可以作为大鼠神经组织星形胶质细胞的分选纯化、原代培养的方法;随着年龄的增加,星形胶质细胞发生细胞老化,表现出促炎症表型,促进神经系统的炎性衰老,可能是神经系统老化及神经退行性疾病的机制之一。     

Activated peripheral lymphocytes with increased expression of cell adhesion molecules and cytotoxic markers are associated with dengue fever disease

The immune mechanisms involved in dengue fever and dengue hemorrhagic/dengue shock syndrome are not well understood. The ex vivo activation status of immune cells during the dengue disease in patients was examined. CD4 and CD8 T cells were reduced during the acute phase. Interestingly, CD8 T cells co-expressing activation marker HLA-DR, Q, P, and cytolytic granule protein-Tia-1 were significantly higher in dengue patients than in controls. Detection of adhesion molecules indicated that in dengue patients the majority of T cells (CD4 and CD8) express the activation/memory phenotype, characterized as CD44HIGH and lack the expression of the na?ve cell marker, CD62L LOW. Also, the levels of T cells co-expressing ICAM-1 (CD54), VLA-4, and LFA-1 (CD11a) were significantly increased. CD8 T lymphocytes expressed predominantly low levels of anti-apoptotic molecule Bcl-2 in the acute phase, possibly leading to the exhibition of a phenotype of activated/effector cells. Circulating levels of IL-18, TGF-b1 and sICAM-1 were significantly elevated in dengue patients. Early activation events occur during acute dengue infection which might contribute to viral clearance. Differences in expression of adhesion molecules among CD4 and CD8 T cells might underlie the selective extravasation of these subsets from blood circulation into lymphoid organs and/or tissues. In addition, activated CD8 T cells would be more susceptible to apoptosis as shown by the alteration in Bcl-2 expression. Cytokines such as IL-18, TGF-b1, and sICAM-1 may be contributing by either stimulating or suppressing the adaptative immune response, during dengue infection, thereby perhaps establishing a relationship with disease severity.     

D-dimer and CoV-2 spike-immune complexes contribute to the production of PGE2 and proinflammatory cytokines in monocytes

An overreactive inflammatory response and coagulopathy are observed in patients with severe form of COVID-19. Since increased levels of D-dimer (DD) are associated with coagulopathy in COVID-19, we explored whether DD contributes to the aberrant cytokine responses. Here we show that treatment of healthy human monocytes with DD induced a dose dependent increase in production of pyrogenic mediator, Prostaglandin E2 (PGE2) and inflammatory cytokines, IL-6 and IL-8. The DD-induced PGE2 and inflammatory cytokines were enhanced significantly by co-treatment with immune complexes (IC) of SARS CoV-2 recombinant S protein or of pseudovirus containing SARS CoV-2 S protein (PVCoV-2) coated with spike-specific chimeric monoclonal antibody (MAb) containing mouse variable and human Fc regions. The production of PGE2 and cytokines in monocytes activated with DD and ICs was sensitive to the inhibitors of β2 integrin and FcγRIIa, and to the inhibitors of calcium signaling, Mitogen-Activated Protein Kinase (MAPK) pathway, and tyrosine-protein kinase. Importantly, strong increase in PGE2 and in IL-6/IL-8/IL-1β cytokines was observed in monocytes activated with DD in the presence of IC of PVCoV-2 coated with plasma from hospitalized COVID-19 patients but not from healthy donors. The IC of PVCoV-2 with convalescent plasma induced much lower levels of PGE2 and cytokines compared with plasma from hospitalized COVID-19 patients. PGE2 and IL-6/IL-8 cytokines produced in monocytes activated with plasma-containing IC, correlated well with the levels of spike binding antibodies and not with neutralizing antibody titers. Our study suggests that a combination of high levels of DD and high titers of spike-binding antibodies that can form IC with SARS CoV-2 viral particles might accelerate the inflammatory status of lung infiltrating monocytes leading to increased lung pathology in patients with severe form of COVID-19.     

An emerging role for the anti-inflammatory cytokine interleukin-10 in dengue virus infection

Infection with dengue virus (DENV) causes both mild dengue fever and severe dengue diseases, such as dengue hemorrhagic fever and dengue shock syndrome. The pathogenic mechanisms for DENV are complicated, involving viral cytotoxicity, immunopathogenesis, autoimmunity, and underlying host diseases. Viral load correlates with disease severity, while the antibody-dependent enhancement of infection largely determines the secondary effects of DENV infection. Epidemiological and experimental studies have revealed an association between the plasma levels of interleukin (IL)-10, which is the master anti-inflammatory cytokine, and disease severity in patients with DENV infection. Based on current knowledge of IL-10-mediated immune regulation during infection, researchers speculate an emerging role for IL-10 in clinical disease prognosis and dengue pathogenesis. However, the regulation of dengue pathogenesis has not been fully elucidated. This review article discusses the regulation and implications of IL-10 in DENV infection. For future strategies against DENV infection, manipulating IL-10 may be an effective antiviral treatment in addition to the development of a safe dengue vaccine.     

Early prolonged neutrophil activation in critically ill patients with sepsis

We hypothesised that plasma concentrations of biomarkers of neutrophil activation and pro-inflammatory cytokines differ according to the phase of rapidly evolving sepsis. In an observational study, we measured heparin-binding protein (HBP), myeloperoxidase (MPO), IL-6 and IL-8 in 167 sepsis patients on intensive care unit admission. We prospectively used the emergence of the first sepsis-associated organ dysfunction (OD) as a surrogate for the sepsis phase. Fifty-five patients (of 167, 33%) developed the first OD > 1 h before, 74 (44%) within ± 1 h, and 38 (23%) > 1 h after intensive care unit admission. HBP and MPO were elevated at a median of 12 h before the first OD, remained high up to 24 h, and were not associated with sepsis phase. IL-6 and IL-8 rose and declined rapidly close to OD emergence. Elevation of neutrophil activation markers HBP and MPO was an early event in the evolution of sepsis, lasting beyond the subsidence of the pro-inflammatory cytokine reaction. Thus, as sepsis biomarkers, HBP and MPO were not as prone as IL-6 and IL-8 to the effect of sample timing.