Cytokine Expression Profile of Dengue Patients at Different Phases of Illness

Background

Dengue is an important medical problem, with symptoms ranging from mild dengue fever to severe forms of the disease, where vascular leakage leads to hypovolemic shock. Cytokines have been implicated to play a role in the progression of severe dengue disease; however, their profile in dengue patients and the synergy that leads to continued plasma leakage is not clearly understood. Herein, we investigated the cytokine kinetics and profiles of dengue patients at different phases of illness to further understand the role of cytokines in dengue disease.

Methods and Findings

Circulating levels of 29 different types of cytokines were assessed by bead-based ELISA method in dengue patients at the 3 different phases of illness. The association between significant changes in the levels of cytokines and clinical parameters were analyzed. At the febrile phase, IP-10 was significant in dengue patients with and without warning signs. However, MIP-1β was found to be significant in only patients with warning signs at this phase. IP-10 was also significant in both with and without warning signs patients during defervescence. At this phase, MIP-1β and G-CSF were significant in patients without warning signs, whereas MCP-1 was noted to be elevated significantly in patients with warning signs. Significant correlations between the levels of VEGF, RANTES, IL-7, IL-12, PDGF and IL-5 with platelets; VEGF with lymphocytes and neutrophils; G-CSF and IP-10 with atypical lymphocytes and various other cytokines with the liver enzymes were observed in this study.

Conclusions

The cytokine profile patterns discovered between the different phases of illness indicate an essential role in dengue pathogenesis and with further studies may serve as predictive markers for progression to dengue with warning signs.     

Cyclic di‐AMP targets the cystathionine beta‐synthase domain of the osmolyte transporter OpuC

Cellular turgor is of fundamental importance to bacterial growth and survival. Changes in external osmolarity as a consequence of fluctuating environmental conditions and colonization of diverse environments can significantly impact cytoplasmic water content, resulting in cellular lysis or plasmolysis. To ensure maintenance of appropriate cellular turgor, bacteria import ions and small organic osmolytes, deemed compatible solutes, to equilibrate cytoplasmic osmolarity with the extracellular environment. Here, we show that elevated levels of c‐di‐AMP, a ubiquitous second messenger among bacteria, result in significant susceptibility to elevated osmotic stress in the bacterial pathogen Listeria monocytogenes. We found that levels of import of the compatible solute carnitine show an inverse correlation with intracellular c‐di‐AMP content and that c‐di‐AMP directly binds to the CBS domain of the ATPase subunit of the carnitine importer OpuC. Biochemical and structural studies identify conserved residues required for this interaction and transport activity in bacterial cells. Overall, these studies reveal a role for c‐di‐AMP mediated regulation of compatible solute import and provide new insight into the molecular mechanisms by which this essential second messenger impacts bacterial physiology and adaptation to changing environmental conditions.     

Detecting amyloid-beta aggregation with fiber-based fluorescence correlation spectroscopy

Soluble aggregates critically influence the chemical and biological aspects of amyloid protein aggregation, but their population is difficult to measure, especially in vivo. We take an optical fiber-based fluorescence correlation spectroscopy (FCS) approach to characterize a solution of aggregating amyloid-beta molecules. We find that this technique can easily resolve aggregate particles of size 100 nm or greater in vitro, and the size distribution of these particles agrees well with that obtained by conventional FCS techniques. We propose fiber FCS as a tool for studying aggregation in vivo.     

Susceptible and Protective HLA Class 1 Alleles against Dengue Fever and Dengue Hemorrhagic Fever Patients in a Malaysian Population

Background

The human leukocyte antigen alleles have been implicated as probable genetic markers in predicting the susceptibility and/or protection to severe manifestations of dengue virus (DENV) infection. In this present study, we aimed to investigate for the first time, the genotype variants of HLA Class 1(-A and -B) of DENV infected patients against healthy individuals in Malaysia.

Methodology/Principal Findings

This study was carried out with 92 dengue disease patients and 95 healthy controls from three different ethnic groups (Malay, Chinese and Indian) in Malaysia. All patients with clinical and laboratory confirmation of DENV infection were typed for the HLA-A and B loci, using polymerase chain reaction-sequence specific primer techniques. In our total population, a significant increase for HLA-B*53 (P = 0.042, Pc = 1.008) allele and a significant decrease for A*03 (P = 0.015, Pc = 0.18, OR = 5.23, 95% CI = 1.19–23.02) and B*18 (P = 0.017, Pc = 0.408) alleles were noted in DHF patients as compared to healthy donors. We also observed that in the Malay DHF patients, allele B*13 (P = 0.049, Pc = 1.176, OR = 0.18, 95% CI = 0.03–0.90) was present at a significantly higher frequency in this population while allele HLA-B*18 (P = 0.024, Pc = 0.576) was seen to be negatively associated with DHF.

Conclusions/Significance

These are the first findings on genetic polymorphisms in our population and we conclude that: (1) In our total population, HLA-B*53 probably involve in disease susceptibility, while the HLA-A*03 and HLA-B*18 may confer protection from progression to severe disease; (2) In the Malay population, HLA-B*13 and B*18 are probably associated in disease susceptibility and protection, respectively. These results could furnish as a valuable predictive tool to identify ethnically different individuals at risk and/or protection from severe forms of DENV infection and would provide valuable informations for the design of future dengue vaccine.     

Immobilization of acidic lipase derived from Pseudomonas gessardii onto mesoporous activated carbon for the hydrolysis of olive oil

Mesoporous activated carbon (MAC) derived from rice husk is used for the immobilization of acidic lipase (ALIP) produced from Pseudomonas gessardii. The purified acidic lipase had the specific activity and molecular weight of 1473 U/mg and 94 kDa respectively. To determine the optimum conditions for the immobilization of lipase onto MAC, the experiments were carried out by varying the time (10–180 min), pH (2–8), temperature (10–50 °C) and the initial lipase activity (49 × 10³, 98 × 10³, 147 × 10³ and 196 × 10³ U/l in acetate buffer). The optimum conditions for immobilization of acidic lipase were found to be: time—120 min; pH 3.5; temperature—30 °C, which resulted in achieving a maximum immobilization of 1834 U/g. The thermal stability of the immobilized lipase was comparatively higher than that in its free form. The free and immobilized enzyme kinetic parameters (K_m and V_max) were found using Michaelis–Menten enzyme kinetics. The K_m values for free enzyme and immobilized one were 0.655 and 0.243 mM respectively. The immobilization of acidic lipase onto MAC was confirmed using Fourier Transform-Infrared Spectroscopy, X-ray diffraction analysis and scanning electron microscopy.     

Lack of evidence for intermolecular epistatic interactions between adiponectin and resistin gene polymorphisms in Malaysian male subjects

Epistasis (gene-gene interaction) is a ubiquitous component of the genetic architecture of complex traits such as susceptibility to common human diseases. Given the strong negative correlation between circulating adiponectin and resistin levels, the potential intermolecular epistatic interactions between ADIPOQ (SNP+45T > G, SNP+276G > T, SNP+639T > C and SNP+1212A > G) and RETN (SNP-420C > G and SNP+299G > A) gene polymorphisms in the genetic risk underlying type 2 diabetes (T2DM) and metabolic syndrome (MS) were assessed. The potential mutual influence of the ADIPOQ and RETN genes on their adipokine levels was also examined. The rare homozygous genotype (risk alleles) of SNP-420C > G at the RETN locus tended to be co-inherited together with the common homozygous genotypes (protective alleles) of SNP+639T > C and SNP+1212A > G at the ADIPOQ locus. Despite the close structural relationship between the ADIPOQ and RETN genes, there was no evidence of an intermolecular epistatic interaction between these genes. There was also no reciprocal effect of the ADIPOQ and RETN genes on their adipokine levels, i.e., ADIPOQ did not affect resistin levels nor did RETN affect adiponectin levels. The possible influence of the ADIPOQ gene on RETN expression warrants further investigation.     

C-Phycocyanin Confers Protection against Oxalate-Mediated Oxidative Stress and Mitochondrial Dysfunctions in MDCK Cells

Oxalate toxicity is mediated through generation of reactive oxygen species (ROS) via a process that is partly dependent on mitochondrial dysfunction. Here, we investigated whether C-phycocyanin (CP) could protect against oxidative stress-mediated intracellular damage triggered by oxalate in MDCK cells. DCFDA, a fluorescence-based probe and hexanoyl-lysine adduct (HEL), an oxidative stress marker were used to investigate the effect of CP on oxalate-induced ROS production and membrane lipid peroxidation (LPO). The role of CP against oxalate-induced oxidative stress was studied by the evaluation of mitochondrial membrane potential by JC1 fluorescein staining, quantification of ATP synthesis and stress-induced MAP kinases (JNK/SAPK and ERK1/2). Our results revealed that oxalate-induced cells show markedly increased ROS levels and HEL protein expression that were significantly decreased following pre-treatment with CP. Further, JC1 staining showed that CP pre-treatment conferred significant protection from mitochondrial membrane permeability and increased ATP production in CP-treated cells than oxalate-alone-treated cells. In addition, CP treated cells significantly decreased the expression of phosphorylated JNK/SAPK and ERK1/2 as compared to oxalate-alone-treated cells. We concluded that CP could be used as a potential free radical-scavenging therapeutic strategy against oxidative stress-associated diseases including urolithiasis.     

Ameliorative property of Sesbania grandiflora on carbohydrate metabolic enzymes in the liver and kidney of streptozotocin-induced diabetic rats

In diabetic condition, endogenous glucose synthesis will be elevated due to defect in the action of vital enzymes involved in carbohydrate metabolism, which is the main cause for hyperglycemia. The current study was designed to explore the anti-hyperglycemic efficacy of Sesbania grandiflora flower (SGF) extract by evaluating the concentration of C-peptide, insulin, glucose, glycosylated hemoglobin (HbA_1C), hemoglobin (Hb), glycogen and carbohydrate metabolic enzymes activities in diabetic rats. The study found to lower the level of glucose, HbA_1C and simultaneously ameliorated concentrations of C-peptide, insulin, hemoglobin (Hb), glycogen and carbohydrate metabolic enzymes activities in SGF treated (250 mg/kg body weight for 45 days) diabetic rats. Moreover, SGF administered diabetic rats showed diminished consumption of food and water at the same time improved body weight. The results obtained from the present study were compared with glibenclamide treated (600 µg/kg body weight) diabetic rats. SGF were supplemented to normal rats to rule out toxic effect of SGF, to explore any significant alteration in the above parameters. Hence, the results depict that SGF modulated the carbohydrate metabolic enzymes activities through ameliorating the secretion of insulin and diminishing the level of glucose concentration in STZ-induced diabetic rats by its bioactive compounds.     

Phyllanthus spp. Induces Selective Growth Inhibition of PC-3 and MeWo Human Cancer Cells through Modulation of Cell Cycle and Induction of Apoptosis

Background

Phyllanthus is a traditional medicinal plant that has been used in the treatment of many diseases including hepatitis and diabetes. The main aim of the present work was to investigate the potential cytotoxic effects of aqueous and methanolic extracts of four Phyllanthus species (P.amarus, P.niruri, P.urinaria and P.watsonii) against skin melanoma and prostate cancer cells.

Methodology/Principal Findings

Phyllanthus plant appears to possess cytotoxic properties with half-maximal inhibitory concentration (IC₅₀) values of 150–300 µg/ml for aqueous extract and 50–150 µg/ml for methanolic extract that were determined using the MTS reduction assay. In comparison, the plant extracts did not show any significant cytotoxicity on normal human skin (CCD-1127Sk) and prostate (RWPE-1) cells. The extracts appeared to act by causing the formation of a clear “ladder” fragmentation of apoptotic DNA on agarose gel, displayed TUNEL-positive cells with an elevation of caspase-3 and -7 activities. The Lactate Dehydrogenase (LDH) level was lower than 15% in Phyllanthus treated-cancer cells. These indicate that Phyllanthus extracts have the ability to induce apoptosis with minimal necrotic effects. Furthermore, cell cycle analysis revealed that Phyllanthus induced a Go/G1-phase arrest on PC-3 cells and a S-phase arrest on MeWo cells and these were accompanied by accumulation of cells in the Sub-G1 (apoptosis) phase. The cytotoxic properties may be due to the presence of polyphenol compounds such as ellagitannins, gallotannins, flavonoids and phenolic acids found both in the water and methanol extract of the plants.

Conclusions/Significance

Phyllanthus plant exerts its growth inhibition effect in a selective manner towards cancer cells through the modulation of cell cycle and induction of apoptosis via caspases activation in melanoma and prostate cancer cells. Hence, Phyllanthus may be sourced for the development of a potent apoptosis-inducing anticancer agent.     

Lack of Clinical Manifestations in Asymptomatic Dengue Infection Is Attributed to Broad Down-Regulation and Selective Up-Regulation of Host Defence Response Genes

Objectives

Dengue represents one of the most serious life-threatening vector-borne infectious diseases that afflicts approximately 50 million people across the globe annually. Whilst symptomatic infections are frequently reported, asymptomatic dengue remains largely unnoticed. Therefore, we sought to investigate the immune correlates conferring protection to individuals that remain clinically asymptomatic.

Methods

We determined the levels of neutralizing antibodies (nAbs) and gene expression profiles of host immune factors in individuals with asymptomatic infections, and whose cognate household members showed symptoms consistent to clinical dengue infection.

Results

We observed broad down-regulation of host defense response (innate, adaptive and matrix metalloprotease) genes in asymptomatic individuals as against symptomatic patients, with selective up-regulation of distinct genes that have been associated with protection. Selected down-regulated genes include: TNF α (TNF), IL8, C1S, factor B (CFB), IL2, IL3, IL4, IL5, IL8, IL9, IL10 and IL13, CD80, CD28, and IL18, MMP8, MMP10, MMP12, MMP15, MMP16, and MMP24. Selected up-regulated genes include: RANTES (CCL5), MIP-1α (CCL3L1/CCL3L3), MIP-1β (CCL4L1), TGFβ (TGFB), and TIMP1.

Conclusion

Our findings highlight the potential association of certain host genes conferring protection against clinical dengue. These data are valuable to better explore the mysteries behind the hitherto poorly understood immunopathogenesis of subclinical dengue infection.