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.     

Differential patterns of synaptotagmin7 mRNA expression in rats with kainate- and pilocarpine-induced seizures

Previous studies in rat models of neurodegenerative disorders have shown disregulation of striatal synaptotagmin7 mRNA. Here we explored the expression of synaptotagmin7 mRNA in the brains of rats with seizures triggered by the glutamatergic agonist kainate (10 mg/kg) or by the muscarinic agonist pilocarpine (30 mg/kg) in LiCl (3 mEq/kg) pre-treated (24 h) rats, in a time-course experiment (30 min-1 day). After kainate-induced seizures, synaptotagmin7 mRNA levels were transiently and uniformly increased throughout the dorsal and ventral striatum (accumbens) at 8 and 12 h, but not at 24 h, followed at 24 h by somewhat variable upregulation within different parts of the cerebral cortex, amigdala and thalamic nuclei, the hippocampus and the lateral septum. By contrast, after LiCl/pilocarpine-induced seizures, there was a more prolonged increase of striatal Synaptotagmin7 mRNA levels (at 8, 12 and 24 h), but only in the ventromedial striatum, while in some other of the aforementioned brain regions there was a decline to below the basal levels. After systemic post-treatment with muscarinic antagonist scopolamine in a dose of 2 mg/kg the seizures were either extinguished or attenuated. In scopolamine post-treated animals with extinguished seizures the striatal synaptotagmin7 mRNA levels (at 12 h after the onset of seizures) were not different from the levels in control animals without seizures, while in rats with attenuated seizures, the upregulation closely resembled kainate seizures-like pattern of striatal upregulation. In the dose of 1 mg/kg, scopolamine did not significantly affect the progression of pilocarpine-induced seizures or pilocarpine seizures-like pattern of striatal upregulation of synaptotagmin7 mRNA. In control experiments, equivalent doses of scopolamine per se did not affect the expression of synaptotagmin7 mRNA. We conclude that here described differential time course and pattern of synaptotagmin7 mRNA expression imply regional differences of pathophysiological brain activation and plasticity in these two models of seizures.     

Balance of human choline kinase isoforms is critical for cell cycle regulation: implications for the development of choline kinase-targeted cancer therapy

The enzyme choline kinase (CK), which catalyzes the phosphorylation of choline to phosphorylcholine in the presence of ATP, has an essential role in the biosynthesis of phosphatidylcholine, the major constituent of all mammalian cell membranes. CK is encoded by two separate genes expressing the three isoforms CKα1, CKα2 and CKβ that are active as homodimeric or heterodimeric species. Metabolic changes observed in various cancer cell lines and tumors have been associated with differential and marked up-regulation of the CKα genes, and specific inhibition of CKα activity has been proposed as a potential anti-cancer strategy. As a result, less attention has been given to CKβ and its interaction with CKα. With the aim of profiling the intracellular roles of CKα and CKβ, we used RNA interference (RNAi) as a molecular approach to down-regulate the expression of CK in HeLa cells. Individual and simultaneous RNAi-based silencing of the CK α and β isoforms was achieved using different combinations of knockdown strategies. Efficient knockdown was confirmed by immunodetection using our isoform-specific antibodies and by quantitative real-time PCR. Our analyses of the phenotypic consequences of CK depletion showed the expected lethal effect of CKα knockdown. However, CKβ- and CKα +?CKβ-silenced cells had no aberrant phenotype. Therefore, our results support the hypothesis that the balance of the α and β isoforms is critical for cancer cell survival. The suppression of the cancer cell killing effect of CKα silencing by simultaneous knockdown of both isoforms implies that a more effective CK-based anti-cancer strategy can be achieved by reducing cross-reactivity with CKβ.     

Imaging of the cell surface interface using objective coupled widefield surface plasmon microscopy

We report on the development and on the first use of the widefield surface plasmon (WSPR) microscope in the examination of the cell surface interface at submicron lateral resolutions. The microscope is Kohler illuminated and uses either a 1.45 numerical aperture (NA) oil immersion lens, or a 1.65 NA oil immersion lens to excite surface plasmons at the interface between a thin gold layer and a glass or sapphire cover slip. Like all surface plasmon microscope systems the WSPR has been proven in previous studies to also be capable of nanometric z-scale resolutions. In this study we used the system to image the interface between HaCaT cells and the gold layer. Imaging was performed in air using fixed samples and the 1.45 NA objective based system and also using live cells in culture media using the 1.65 NA based system. Imaging in air enabled the visualisation of high resolution and high-contrast submicron features identified by vinculin immunostaining as component of focal contacts and focal adhesions. In comparison, imaging in fluid enabled cell surface interfacial interactions to be tracked by time-lapse video WSPR microscopy. Our results indicate that the cell surface interface and thus cell signalling mechanisms may be readily interrogated in live cells without the use of labelling techniques.     

Sulfidation of NiMn‐Layered Double Hydroxides/Graphene Oxide Composites toward Supercapacitor Electrodes with Enhanced Performance

Supercapacitors can deliver high‐power density and long cycle stability, but the limited energy density due to poor electronic and ionic conductivity of the supercapacitor electrode has been a bottleneck in many applications. A strategy to prepare microflower‐like NiMn‐layered double hydroxides (LDH) with sulfidation is delineated to reduce the charge transfer resistance of supercapacitor electrode and realize faster reversible redox reactions with notably enhanced specific capacitance. The incorporation of graphite oxide (GO) in NiMn LDH during sulfidation leads to simultaneous reduction of GO with enhanced conductivity, lessened defects, and doping of S into the graphitic structure. Cycling stability of the sulfidized composite electrode is enhanced due to the alleviation of phase transformation during electrochemical cycling test. As a result, this sulfidation product of LDH/GO (or LDHGOS) can reach a high‐specific capacitance of 2246.63 F g^?1 at a current density of 1 A g^?1, and a capacitance of 1670.83 F g^?1 is retained at a high‐current density of 10 A g^?1, exhibiting an outstanding capacitance and rate performance. The cycling retention of the LDHGOS electrode is also extended to ≈ 67% after 1500 cycles compared to only ≈44% of the pristine NiMn LDH.     

Mapping global cropland and field size

A new 1 km global IIASA‐IFPRI cropland percentage map for the baseline year 2005 has been developed which integrates a number of individual cropland maps at global to regional to national scales. The individual map products include existing global land cover maps such as GlobCover 2005 and MODIS v.5, regional maps such as AFRICOVER and national maps from mapping agencies and other organizations. The different products are ranked at the national level using crowdsourced data from Geo‐Wiki to create a map that reflects the likelihood of cropland. Calibration with national and subnational crop statistics was then undertaken to distribute the cropland within each country and subnational unit. The new IIASA‐IFPRI cropland product has been validated using very high‐resolution satellite imagery via Geo‐Wiki and has an overall accuracy of 82.4%. It has also been compared with the EarthStat cropland product and shows a lower root mean square error on an independent data set collected from Geo‐Wiki. The first ever global field size map was produced at the same resolution as the IIASA‐IFPRI cropland map based on interpolation of field size data collected via a Geo‐Wiki crowdsourcing campaign. A validation exercise of the global field size map revealed satisfactory agreement with control data, particularly given the relatively modest size of the field size data set used to create the map. Both are critical inputs to global agricultural monitoring in the frame of GEOGLAM and will serve the global land modelling and integrated assessment community, in particular for improving land use models that require baseline cropland information. These products are freely available for downloading from the http://cropland.geo-wiki.org website.     

Site localization of sialyl Lewis(x) antigen on alpha1-acid glycoprotein by high performance liquid chromatography-electrospray mass spectrometry

A simple, fast and sensitive method was developed to verify the presence of the sialyl Lewis(x) antigen on an N-linked glycoprotein. High performance liquid chromatography-electrospray mass spectrometry (HPLC-ESI/MS) was used to identify which of the five N-linked glycosylation sites of human plasma alpha1-acid-glycoprotein (orosomucoid, OMD) contain the sialyl Lewis(x) antigen. OMD was digested with proteolytic enzymes and analyzed by reversed phase chromatography coupled with on-line ESI/MS. A tandem mass spectrometry experiment was designed to detect the presence of the sialyl Lewis(x) antigen based on the observation of an 803 mass to charge ratio ( m/z ) ion produced in the intermediate pressure region of the ESI interface. The ESI/MS signal at m/z 803 is consistent with an oxonium ion for a glycan structure containing NeuAc, Gal, GlcNAc, and Fuc. The identity of the m/z 803 ion was confirmed by ESI/MS/MS analysis of the m/z 803 fragment ion and comparison with a sialyl Lewis(x) standard. The stereochemistry and linkage positions were assigned using previous NMR analysis but could be determined with permethylation analysis if necessary. The analysis of OMD gave a pattern showing signal for the sialyl Lewis(x) antigen coeluting with each of the five N-linked glycopeptides. The ability to monitor sialyl Lewis(x) expression at each of the five sites is of interest in the study of OMD's role in inflammatory diseases.