Mapping cells and sub-cellular organelles on 2-D gels: ‘new tricks for an old horse’

AbstractNowadays, investigators in all fields are faced with the identification of unknown, up- or down-regulated, modified proteins that they are trying to identify. Two-dimensional (2-D) gel electrophoresis, with its ability to resolve several thousand proteins, is an extremely powerful technique. The current resolution and reproducibility of 2-D gel technology and the establishment of computer assisted 2-D gel protein databases have paved new ways for the identification of proteins

Quality assurance of the solar UV network in the Antarctic

Measuring ultraviolet radiation in the Antarctic region, where weather conditions are extremely challenging, is a demanding task. Proper quality control of the measurements and quality assurance of the data, which are the basis of all scientific use of data, has to be especially well planned and executed. In this paper we show the importance of proper quality assurance and describe the methods used to successfully operate the NILU-UV multichannel radiometers of the Antarctic network stations at Ushuaia, 54S, and Marambio, 64S. According to our experience, even though multichannel instruments are supposed to be rather stable as a function of time, severe drifts can occur in the sensitivity of the channels under these harsh conditions. During 2000–2003 the biggest drifts were 35%, both at Ushuaia and Marambio, with the sensitivity of the channels dropping at different rates. Without proper corrections in the data, this would have seriously affected the calculated UV dose rates. As part of the quality assurance of the network a traveling reference NILU-UV, which was found to be stable, was used to transfer the desired irradiance scale to the site NILU-UV data. Relative lamp tests were used to monitor the stability of the instruments. Each site NILU-UV was scaled channel by channel to the traveling reference by performing solar comparisons. The method of scaling each channel separately was found to be successful, even though the differences between the raw data of the site NILU-UV and the reference instruments were, before the data correction, as much as 40%. After the correction, the mean ratios of erythemally weighted UV dose rates measured during the solar comparisons in 2000–2003 between the reference NILU-UV and the site NILU-UV were 1.007 ± 0.011 and 1.012 ± 0.012 for Ushuaia and Marambio, respectively, when the solar zenith angle varied up to 80. These results make possible the scientific use of NILU-UV data measured simultaneously at quite different locations, e.g., the Antarctic and Arctic, and the method presented is also practicable for other multichannel radiometer networks.S, and Marambio, 64S. According to our experience, even though multichannel instruments are supposed to be rather stable as a function of time, severe drifts can occur in the sensitivity of the channels under these harsh conditions. During 2000–2003 the biggest drifts were 35%, both at Ushuaia and Marambio, with the sensitivity of the channels dropping at different rates. Without proper corrections in the data, this would have seriously affected the calculated UV dose rates. As part of the quality assurance of the network a traveling reference NILU-UV, which was found to be stable, was used to transfer the desired irradiance scale to the site NILU-UV data. Relative lamp tests were used to monitor the stability of the instruments. Each site NILU-UV was scaled channel by channel to the traveling reference by performing solar comparisons. The method of scaling each channel separately was found to be successful, even though the differences between the raw data of the site NILU-UV and the reference instruments were, before the data correction, as much as 40%. After the correction, the mean ratios of erythemally weighted UV dose rates measured during the solar comparisons in 2000–2003 between the reference NILU-UV and the site NILU-UV were 1.007 ± 0.011 and 1.012 ± 0.012 for Ushuaia and Marambio, respectively, when the solar zenith angle varied up to 80. These results make possible the scientific use of NILU-UV data measured simultaneously at quite different locations, e.g., the Antarctic and Arctic, and the method presented is also practicable for other multichannel radiometer networks.S. According to our experience, even though multichannel instruments are supposed to be rather stable as a function of time, severe drifts can occur in the sensitivity of the channels under these harsh conditions. During 2000–2003 the biggest drifts were 35%, both at Ushuaia and Marambio, with the sensitivity of the channels dropping at different rates. Without proper corrections in the data, this would have seriously affected the calculated UV dose rates. As part of the quality assurance of the network a traveling reference NILU-UV, which was found to be stable, was used to transfer the desired irradiance scale to the site NILU-UV data. Relative lamp tests were used to monitor the stability of the instruments. Each site NILU-UV was scaled channel by channel to the traveling reference by performing solar comparisons. The method of scaling each channel separately was found to be successful, even though the differences between the raw data of the site NILU-UV and the reference instruments were, before the data correction, as much as 40%. After the correction, the mean ratios of erythemally weighted UV dose rates measured during the solar comparisons in 2000–2003 between the reference NILU-UV and the site NILU-UV were 1.007 ± 0.011 and 1.012 ± 0.012 for Ushuaia and Marambio, respectively, when the solar zenith angle varied up to 80. These results make possible the scientific use of NILU-UV data measured simultaneously at quite different locations, e.g., the Antarctic and Arctic, and the method presented is also practicable for other multichannel radiometer networks.. These results make possible the scientific use of NILU-UV data measured simultaneously at quite different locations, e.g., the Antarctic and Arctic, and the method presented is also practicable for other multichannel radiometer networks.Fil: Lakkala, K.. Finnish Meteorological Institute; FinlandiaFil: Redondas, A.. Instituto Nacional de Meteorología; EspañaFil: Meinander, O.. Finnish Meteorological Institute; FinlandiaFil: Torres ,Carlos. Instituto Nacional de Meteorología; EspañaFil: Koskela, T.. Finnish Meteorological Institute; FinlandiaFil: Cuevas, Eduardo. Instituto Nacional de Meteorología; EspañaFil: Taalas, P.. Finnish Meteorological Institute; FinlandiaFil: Dahlback, A.. University of Oslo; NoruegaFil: Deferrari, Guillermo Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Austral de Investigaciones Científicas; ArgentinaFil: Edvardsen, K.. Instituto Noruego de Investigación del Aire; NoruegaFil: Ochoa, H.. Ministerio de Relaciones Exteriores, Comercio Interno y Culto. Dirección Nacional del Antártico. Instituto Antártico Argentino; Argentin

Adaptation of an Evaluation System for e-Health Environments

Proceedings of: 14th International Conference, KES 2010, Cardiff, UK, September 8-10, 2010The increase in ageing of European population implies a high cost in economy and society in any European country and it can be reduced if we pay attention and develop home care systems. Evaluation of these systems is a critical and challenging issue but seldom tackled. It is important before evaluating a system to figure out what is the evaluation goal. In our case, such a goal is to evaluate enhanced user experience and beyond the evaluation goal it is also a central concern about what to evaluate. In this paper we propose a multi-agent home care system where we describe how agents coordinate their decisions to provide e-services to patients when at home after hospitalization. Finally we center our proposal on the adaptation of an evaluation system, previously developed, to support the challenges of an e-Health environment and also the multi-user evaluation. These evaluation methods (online/offline) will provide user's (patients, patient's relatives and healthcare professionals) feedback into the system.This work was supported in part by Projects CICYT TIN2008-06742-C02-02/ TSI, CICYT TEC2008-06732-C02-02/TEC, CAM CONTEXTS (S2009/ TIC-1485) and DPS2008-07029-C02-02.Publicad

Cleavage Requirements for Activation of Factor V by Factor Xa

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/65728/1/j.1432-1033.1997.00012.x.pd

Multiple var2csa-Type PfEMP1 Genes Located at Different Chromosomal Loci Occur in Many Plasmodium falciparum Isolates

BACKGROUND:The var2csa gene encodes a Plasmodium falciparum adhesion receptor which binds chondroitin sulfate A (CSA). This var gene is more conserved than other PfEMP1/var genes and is found in all P. falciparum isolates. In isolates 3D7, FCR3/It4 and HB3, var2csa is transcribed from a sub-telomeric position on the left arm of chromosome 12, but it is not known if this location is conserved in all parasites. Genome sequencing indicates that the var2csa gene is duplicated in HB3, but whether this is true in natural populations is uncertain. METHODOLOGY/PRINCIPAL FINDINGS:To assess global variation in the VAR2CSA protein, sequence variation in the DBL2X region of var2csa genes in 54 P.falciparum samples was analyzed. Chromosome mapping of var2csa loci was carried out and a quantitative PCR assay was developed to estimate the number of var2csa genes in P.falciparum isolates from the placenta of pregnant women and from the peripheral circulation of other malaria patients. Sequence analysis, gene mapping and copy number quantitation in P.falciparum isolates indicate that there are at least two loci and that both var2csa-like genes can be transcribed. All VAR2CSA DBL2X domains fall into one of two distinct phylogenetic groups possessing one or the other variant of a large (approximately 26 amino acid) dimorphic motif, but whether either motif variant is linked to a specific locus is not known. CONCLUSIONS/SIGNIFICANCE:Two or more related but distinct var2csa-type PfEMP1/var genes exist in many P. falciparum isolates. One gene is on chromosome 12 but additional var2csa-type genes are on different chromosomes in different isolates. Multiplicity of var2csa genes appears more common in infected placentae than in samples from non-pregnant donors indicating a possible advantage of this genotype in pregnancy associated malaria

Identifying novel genetic determinants of hemostatic balance

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/73734/1/j.1538-7836.2005.01461.x.pd

Human Complement Regulators C4b-Binding Protein and C1 Esterase Inhibitor Interact with a Novel Outer Surface Protein of Borrelia recurrentis

The spirochete Borrelia recurrentis is the causal agent of louse-borne relapsing fever and is transmitted to humans by the infected body louse Pediculus humanus. We have recently demonstrated that the B. recurrentis surface receptor, HcpA, specifically binds factor H, the regulator of the alternative pathway of complement activation, thereby inhibiting complement mediated bacteriolysis. Here, we show that B. recurrentis spirochetes express another potential outer membrane lipoprotein, termed CihC, and acquire C4b-binding protein (C4bp) and human C1 esterase inhibitor (C1-Inh), the major inhibitors of the classical and lectin pathway of complement activation. A highly homologous receptor for C4bp was also found in the African tick-borne relapsing fever spirochete B. duttonii. Upon its binding to B. recurrentis or recombinant CihC, C4bp retains its functional potential, i.e. facilitating the factor I-mediated degradation of C4b. The additional finding that ectopic expression of CihC in serum sensitive B. burgdorferi significantly increased spirochetal resistance against human complement suggests this receptor to substantially contribute, together with other known strategies, to immune evasion of B. recurrentis

Evaluation of Aerosol Delivery of Nanosuspension for Pre-clinical Pulmonary Drug Delivery

Asthma and chronic obstructive pulmonary disease (COPD) are pulmonary diseases that are characterized by inflammatory cell infiltration, cytokine production, and airway hyper-reactivity. Most of the effector cells responsible for these pathologies reside in the lungs. One of the most direct ways to deliver drugs to the target cells is via the trachea. In a pre-clinical setting, this can be achieved via intratracheal (IT), intranasal (IN), or aerosol delivery in the desired animal model. In this study, we pioneered the aerosol delivery of a nanosuspension formulation in a rodent model. The efficiency of different dosing techniques and formulations to target the lungs were compared, and fluticasone was used as the model compound. For the aerosol particle size determination, a ten-stage cascade impactor was used. The mass median aerodynamic diameter (MMAD) was calculated based on the percent cumulative accumulation at each stage. Formulations with different particle size of fluticasone were made for evaluation. The compatibility of regular fluticasone suspension and nanosuspension for aerosol delivery was also investigated. The in vivo studies were conducted on mice with optimized setting. It was found that the aerosol delivery of fluticasone with nanosuspension was as efficient as intranasal (IN) dosing, and was able to achieve dose dependent lung deposition

C4b-Binding Protein Is Present in Affected Areas of Myocardial Infarction during the Acute Inflammatory Phase and Covers a Larger Area than C3

BACKGROUND: During myocardial infarction reduced blood flow in the heart muscle results in cell death. These dying/dead cells have been reported to bind several plasma proteins such as IgM and C-reactive protein (CRP). In the present study we investigated whether fluid-phase complement inhibitor C4b-binding protein (C4BP) would also bind to the infarcted heart tissue. METHODS AND FINDINGS: Initial studies using immunohistochemistry on tissue arrays for several cardiovascular disorders indicated that C4BP can be found in heart tissue in several cardiac diseases but that it is most abundantly found in acute myocardial infarction (AMI). This condition was studied in more detail by analyzing the time window and extent of C4BP positivity. The binding of C4BP correlates to the same locations as C3b, a marker known to correlate to the patterns of IgM and CRP staining. Based on criteria that describe the time after infarction we were able to pinpoint that C4BP binding is a relatively early marker of tissue damage in myocardial infarction with a peak of binding between 12 hours and 5 days subsequent to AMI, the phase in which infiltration of neutrophilic granulocytes in the heart is the most extensive. CONCLUSIONS: C4BP, an important fluid-phase inhibitor of the classical and lectin pathway of complement activation binds to jeopardized cardiomyocytes early after AMI and co-localizes to other well known markers such as C3b

Apolipoprotein M Gene (APOM) Polymorphism Modifies Metabolic and Disease Traits in Type 2 Diabetes

This study aimed at substantiating the associations of the apolipoproein M gene (APOM) with type 2 diabetes (T2D) as well as with metabolic traits in Hong Kong Chinese. In addition, APOM gene function was further characterized to elucidate its activity in cholesterol metabolism. Seventeen APOM SNPs documented in the NCBI database were genotyped. Five SNPs were confirmed in our study cohort of 1234 T2D and 606 control participants. Three of the five SNPs rs707921(C+1871A), rs707922(G+1837T) and rs805264(G+203A) were in linkage disequilibrium (LD). We chose rs707922 to tag this LD region for down stream association analyses and characterized the function of this SNP at molecular level. No association between APOM and T2D susceptibility was detected in our Hong Kong Chinese cohort. Interestingly, the C allele of rs805297 was significantly associated with T2D duration of longer than 10 years (OR = 1.245, p = 0.015). The rs707922 TT genotype was significantly associated with elevated plasma total- and LDL- cholesterol levels (p = 0.006 and p = 0.009, respectively) in T2D patients. Molecular analyses of rs707922 lead to the discoveries of a novel transcript APOM5 as well as the cryptic nature of exon 5 of the gene. Ectopic expression of APOM5 transcript confirmed rs707922 allele-dependent activity of the transcript in modifying cholesterol homeostasis in vitro. In conclusion, the results here did not support APOM as a T2D susceptibility gene in Hong Kong Chinese. However, in T2D patients, a subset of APOM SNPs was associated with disease duration and metabolic traits. Further molecular analysis proved the functional activity of rs707922 in APOM expression and in regulation of cellular cholesterol content