Abstract. The pathogenesis of plasma leakage during dengue hemorrhagic fever/dengue shock syndrome (DHF/ DSS) is largely unknown. Angiopoietins are key regulators of vascular integrity: Angiopoietin-1 is stored in platelets and maintains vascular integrity, and endothelium-derived angiopoietin-2 promotes vascular leakage. We determined angiopoietin-1 and angiopoietin-2 levels in a cohort of children in Indonesia with DHF/DSS and related them to plasma leakage markers. Patients with DHF/DSS had reduced angiopoietin-1 and increased angiopoietin-2 plasma levels on the day of admission when compared with levels at discharge and in healthy controls. There was an inverse correlation between angiopoietin-1 and markers of plasma leakage and a positive correlation between angiopoietin-2 and markers of plasma leakage. Angiopoietin-1 levels followed the same trend as the soluble platelet activation marker P-selectin and correlated with platelet counts. Dengue-associated thrombocytopenia and endothelial activation are associated with an imbalance in angiopoietin-2: angiopoietin-1 plasma levels. This imbalance may contribute to the transient plasma leakage in DHF/DSS.

BackgroundThrombocytopenia, bleeding and plasma leakage are cardinal features of severe dengue. Endothelial cell activation with exocytosis of Weibel-Palade bodies (WPBs) may play an etiological role in this condition.Methods and Principal FindingsIn a cohort of 73 Indonesian children with dengue hemorrhagic fever (DHF), of which 30 with dengue shock syndrome (DSS), we measured plasma levels of the WPB constituents von Willebrand factor antigen (VWF:Ag), VWF propeptide and osteoprotegerin (OPG), together with activity levels of the VWF-cleaving enzyme ADAMTS-13 and the amount of VWF in a platelet binding conformation (VWF activation factor). Compared with healthy controls (n = 17), children with DHF/DSS had significantly higher levels of VWF:Ag, VWF propeptide and OPG and decreased ADAMTS-13 activity. The VWF activation factor was also significantly higher in DHF/DSS and highest in children who died. There were significant differences in the kinetics of the various WPB constituents: VWF propeptide and OPG levels decreased toward discharge, while VWF:Ag levels were lower than expected at enrollment with plasma levels increasing toward discharge. Moreover, VWF propeptide levels correlated better with markers of disease severity (platelet count, liver enzymes, serum albumin and pleural effusion index) than corresponding VWF levels. Together, these findings suggest that there is consumption of VWF in DHF/DSS. In 4 out of 15 selected children with low ADAMTS-13 levels on admission, we found a remarkable reduction in the large and intermediate VWF multimers in the discharge blood samples, consistent with an acquired von Willebrand disease.ConclusionThese findings suggest that severe dengue is associated with exocytosis of WPBs with increased circulating levels of VWF:Ag, VWF propeptide and OPG. High circulating levels of VWF in its active conformation, together with low ADAMTS-13 activity levels, are likely to contribute to the thrombocytopenia and complications of dengue. During the convalescence phase, qualitative defects in VWF with loss of larger VWF multimers may develop.

Massive pro-inflammatory cytokines production has been correlated with the pathogenesis of severe dengue disease. The active compound of mangosteen fruit pericarps, α-mangostin, has been commonly used as traditional medicine and dietary supplement. We examined the effect of α-mangostin against dengue virus (DENV) infection in human peripheral blood mononuclear cells (PBMC) by the measurement of virus titer and TNF-α and IFN-γ cytokines concentration post infection. Increasing concentration of α-mangostin inhibited virus replication and reduced inflammatory cytokines expression at 24- and 48-h post infection. Our results support the potential use of α-mangostin as anti-antiviral and anti-inflammatory therapies in the treatment of dengue.Graphic Abstract

Background

Mortality in children with severe dengue haemorrhagic fever (DHF) in Indonesia is high. The origin of the elevated plasminogen activator inhibitor-1 (PAI-1) levels in these children is unclear. We measured PAI-1, transforming growth factor-β1 (TGF-β1), platelet counts, plasma leakage and liver function in 71 children with DHF (3-15 years old) and in 30 healthy children. We found that PAI-1 concentrations in children with DHF were significantly higher on admission than on Day 2. Circulating TGF-β1 concentrations on admission were significantly lower in DHF than in controls, but on Day 2 increased towards levels in controls. TGF-β1 and PAI-1 concentrations were not correlated on either day. PAI-1 was correlated with platelet count and serum albumin on admission, and with degree of pleural effusion. Liver function tests were mildly elevated but not correlated with PAI-1. In conclusion, elevated PAI-1 concentrations in DHF were associated with platelet counts and plasma leakage.

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T2DM has increase in global-morbidity and mortality. Oxidative stress and adiponectin-levels are important for insulin-resistance and pancreatic-b-cell-dysfunction in T2DM. Okra fruit is rich of quercetin and phytosterol which have positive-effect for T2DM. Research aimed was to study the effect of okra-flour to adiponectin-levels and total-antioxidant-capacity (TAC) in T2DM. Thirty Wistar-rats were divided randomly in five groups. K1 and (X1, X2 and X3)-treated-groups were in T2DM-condition-induced by high-fat-diet-(HFD)-Streptozotochin-(STZ)-nicotinamid-(NA). Healthy-controls-(K2)-group was also used. Okra-flour was given orally for 28 days at doses of 0.1; 0.2 and 0.3 g/Kg-body-weight/d to X1, X2 and X3-groups, respectively. Statistical program was used to analyse the different between pre-post-intervention, and between groups. Correlations between variables were also analysed. The serum-adiponectin and TAC-levels were measured by ELISA and ABTS-methods, respectively. By comparing pre and post-intervention, adiponectin levels of all-intervention-(X1, X2, X3)-group were  increase (p = 0.027 for X1 and X2; p = 0.028 for X3), while in the same period the decrease were found in group K1 (p = 0.026) and K2 (p = 0.028). Increase-TAC-levels pre-post-intervention was observed in group all-intervention-groups (p = 0.027), while no change in K1 (p = 0.66) and the decrease in group K2 (p = 0.039). Reduce-fasting-blood-glucose-levels pre-post-intervention were shown in the all-intervention-groups (p = 0.028), while for the K1 groups was increase (p = 0.028). There were significant differences between the five-groups on fasting-blood-glucose-levels, adiponectin and TAC-levels, and X3-group showed the highest adiponectin and TAC-levels. Very-strong-correlations were found between glucose-adiponectin-TAC-levels-post-intervention. Okra-flour make better glucose-adiponectin and TAC-levels in T2DM-conditions. Okra dose of 0.30 g/Kg-body-weight/day is the best in increasing adiponectin and TAC-levels.

Abstract. Differential viral recognition by cells bearing Toll-like receptor 4 (TLR4) polymorphisms Asp299Gly and Thr399Ile may influence susceptibility and severity of dengue virus infection. In central Java, Indonesia, we investigated 201 children with dengue hemorrhagic fever (DHF) and 179 healthy controls. Patients and controls were mostly ethnic Javanese. A nearly complete cosegregation of the two mutations was observed. The TLR4 299/399 genotype was found in five patients and four controls. Prevalence of the TLR4 299/399 genotype did not differ significantly between controls and DHF patients or between patients with different severities of DHF. Also, vascular leakage in patients with different TLR4 genotypes did not differ. Thus, the 299/399 TLR4 haplotype has only minor influence on susceptibility and severity of complicated dengue virus infection.