| 11. |
- Brolin, M, et al.
(författare)
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Simultaneous transcription of duplicated var2csa gene copies in individual Plasmodium falciparum parasites
- 2009
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Ingår i: Genome Biology. - : Springer Science and Business Media LLC. - 1465-6906 .- 1474-760X. ; 10:10, s. R117-
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Tidskriftsartikel (refereegranskat)abstract
- Background: Single nucleotide polymorphisms are common in duplicated genes, causing functional preservation, alteration or silencing. The Plasmodium falciparum genes var2csa and Pf332 are duplicated in the haploid genome of the HB3 parasite line. Whereas the molecular function of Pf332 remains to be elucidated, VAR2CSA is known to be the main adhesin in placental parasite sequestration. Sequence variations introduced upon duplication of these genes provide discriminative possibilities to analyze allele-specific transcription with a bearing towards understanding gene dosage impact on parasite biology. Results: We demonstrate an approach combining real-time PCR allelic discrimination and discriminative RNA-FISH to distinguish between highly similar gene copies in P. falciparum parasites. The duplicated var2csa variants are simultaneously transcribed, both on a population level and intriguingly also in individual cells, with nuclear co-localization of the active genes and corresponding transcripts. This indicates transcriptional functionality of duplicated genes, challenges the dogma of mutually exclusive var gene transcription and suggests mechanisms behind antigenic variation, at least in respect to the duplicated and highly similar var2csa genes. Conclusions: Allelic discrimination assays have traditionally been applied to study zygosity in diploid genomes. The assays presented here are instead successfully applied to the identification and evaluation of transcriptional activity of duplicated genes in the haploid genome of the P. falciparum parasite. Allelic discrimination and gene or transcript localization by FISH not only provide insights into transcriptional regulation of genes such as the virulence associated var genes, but also suggest that this sensitive and precise approach could be used for further investigation of genome dynamics and gene regulation.
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| 12. |
- Chan, Sherwin, et al.
(författare)
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Regulation of PfEMP1-VAR2CSA translation by a Plasmodium translation-enhancing factor
- 2017
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Ingår i: Nature Microbiology. - : Springer Science and Business Media LLC. - 2058-5276. ; 2:7
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Tidskriftsartikel (refereegranskat)abstract
- Pregnancy-associated malaria commonly involves the binding of Plasmodium falciparum-infected erythrocytes to placental chondroitin sulfate A (CSA) through the PfEMP1-VAR2CSA protein. VAR2CSA is translationally repressed by an upstream open reading frame. In this study, we report that the P. falciparum translation enhancing factor (PTEF) relieves upstream open reading frame repression and thereby facilitates VAR2CSA translation. VAR2CSA protein levels in var2csa-transcribing parasites are dependent on the expression level of PTEF, and the alleviation of upstream open reading frame repression requires the proteolytic processing of PTEF by PfCalpain. Cleavage generates a C-terminal domain that contains a sterile-alpha-motif-like domain. The C-terminal domain is permissive to cytoplasmic shuttling and interacts with ribosomes to facilitate translational derepression of the var2csa coding sequence. It also enhances translation in a heterologous translation system and thus represents the first non-canonical translation enhancing factor to be found in a protozoan. Our results implicate PTEF in regulating placental CSA binding of infected erythrocytes.
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| 13. |
- Ch'ng, Jun-Hong, et al.
(författare)
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Epitopes of anti-RIFIN antibodies and characterization of rif-expressing Plasmodium falciparum parasites by RNA sequencing
- 2017
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 7
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Tidskriftsartikel (refereegranskat)abstract
- Variable surface antigens of Plasmodium falciparum have been a major research focus since they facilitate parasite sequestration and give rise to deadly malaria complications. Coupled with its potential use as a vaccine candidate, the recent suggestion that the repetitive interspersed families of polypeptides (RIFINs) mediate blood group A rosetting and influence blood group distribution has raised the research profile of these adhesins. Nevertheless, detailed investigations into the functions of this highly diverse multigene family remain hampered by the limited number of validated reagents. In this study, we assess the specificities of three promising polyclonal anti-RIFIN antibodies that were IgG-purified from sera of immunized animals. Their epitope regions were mapped using a 175,000-peptide microarray holding overlapping peptides of the P. falciparum variable surface antigens. Through immunoblotting and immunofluorescence imaging, we show that different antibodies give varying results in different applications/assays. Finally, we authenticate the antibody-based detection of RIFINs in two previously uncharacterized non-rosetting parasite lines by identifying the dominant rif transcripts using RNA sequencing.
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| 14. |
- Elfstrand, Lidia, et al.
(författare)
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From starch to starch microspheres: Factors controlling the microspheres quality
- 2006
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Ingår i: Stärke. - : Wiley. - 0038-9056. ; 58:8, s. 381-390
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Tidskriftsartikel (refereegranskat)abstract
- Starch microparticle is a dosage form suitable for encapsulation of protein drugs. The microparticle characteristics can be influential factors for consecutive sustained drug release from the starch matrix and impact its ability to be coated by poly(D,L-lactide-co-glycolide). In this study, six types of starch microparticles were investigated and characterized. Desired qualities of the microspheres such as spherical form of the particles, well-defined and sharp particle contours were coincident with coarse surface morphology, sharp and distinct peaks in DSC and X-ray graphs. It was concluded that the quality of the starch microparticles was influenced by molecular properties of the starch material as well as type of buffer, and these factors were related to recrystallization kinetics, amount and quality of ordered/crystalline structure of final starch microspheres and their microscopic appearance.
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| 15. |
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| 16. |
- Elfstrand, Lidia, et al.
(författare)
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Recrystallization of waxy maize starch during manufacturing of starch microspheres for drug delivery: Optimization by experimental design
- 2007
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Ingår i: Carbohydrate Polymers. - : Elsevier BV. - 0144-8617. ; 68:3, s. 568-576
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Tidskriftsartikel (refereegranskat)abstract
- Gelatinized starch/water dispersions with a water content of 70% w/w were studied in order to evaluate the influence of the incubation time at two fixed temperatures (6 and 37 degrees C) on the recrystallization behavior. The starch material was produced from waxy maize starch by acid hydrolysis followed by a mechanical treatment, i.e. a high pressure homogenization. Empirical models for the recrystallization of the model starch dispersions were investigated by using a central composite circumscribed (CCC) design in two factors involving I I trials. The rebuilt crystalline structure was estimated by measuring the melting transition parameters with differential scanning calorimetry (DSC). The designed models for the melting enthalpy, the melting transition temperatures, and the melting interval as the responses were reproducible and predictable and the results were analyzed by using a response surface modeling. Operative conditions for the formation of the most thermally stable crystalline structure with the highest possible crystallinity were determined. It was also concluded that the decisive factor for the crystallinity of the given starch material was a prolonged storage at 6 'C, while thermal characteristics of the crystallites could be controlled by storage at 37 degrees C. An interaction effect was observed between the incubation time at these two experimental temperatures regarding enthalpy values and the melting interval of the samples. (c) 2006 Elsevier Ltd. All rights reserved.
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| 17. |
- Fredenberg, Susanne, et al.
(författare)
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Development of mass transport resistance in poly(lactide-co-glycolide) films and particles - A mechanistic study.
- 2011
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Ingår i: International Journal of Pharmaceutics. - : Elsevier BV. - 1873-3476 .- 0378-5173. ; 409:1-2, s. 194-202
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Tidskriftsartikel (refereegranskat)abstract
- Poly(d,l-lactide-co-glycolide) (PLG) is the most frequently used biodegradable polymer in the controlled release of an encapsulated drug. The purpose of this work was to explain the surprisingly slow diffusion through this polymer, and locate the major source of mass transport resistance. Diffusion of human growth hormone (hGH) and glucose through PLG films was undetectable (using a diffusion cell), although the degraded polymer contained several times more water than polymer mass. In vitro release of hGH from PLG-coated particles also showed a surprisingly slow rate of release. Non-porous regions inside the PLG films were detected after three weeks of degradation using dextran-coupled fluorescent probes and confocal microscopy. The findings were supported by scanning electron microscopy. Diffusion through PLG films degraded for five weeks was significantly increased when the porosity of both surfaces was increased due to the presence of ZnCl(2) in the buffer the last 3 days of the degradation period. The results indicated high mass transport resistance inside the films after three weeks of degradation, and at the surfaces after five weeks of degradation. These results should also be applicable to microparticles of different sizes. Knowledge of the reason for transport resistance is important in the development of pharmaceuticals and when modifying the rate of drug release.
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| 18. |
- Fredenberg, Susanne, et al.
(författare)
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Pore formation and pore closure in Poly(D,L-lactide-co-glycolide) films.
- 2011
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Ingår i: Journal of Controlled Release. - : Elsevier BV. - 1873-4995 .- 0168-3659. ; 150, s. 142-149
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Tidskriftsartikel (refereegranskat)abstract
- Pore formation and pore closure in poly(D,L-lactide-co-glycolide)-based drug delivery systems are two important processes as they control the release of the encapsulated drug. The phenomenon pore closure was investigated by studying the effects of the pH and the temperature of the release medium, and the properties of the polymer. Poly(D,L-lactide-co-glycolide) (PLG) films were subjected to a pore forming pre-treatment, and then pore closure was observed simultaneously with changes in glass transition temperature, wettability (contact angle), water absorption and mass remaining. To further understand the effect of pH, combined pore formation and pore closure were studied at different pH values. Pore closure was increased in a release medium with low pH, with a low-molecular-weight PLG of relatively low degree of hydrophobicity, or at high temperature. Pore closure occurred by two different mechanisms, one based on polymer-polymer interactions and one on polymer-water interactions. The mobility of the PLG chains also played an important role. The surface of the PLG films were more porous at pH 5-6 than at lower or higher pH, as pore formation was relatively fast and pore closure were less pronounced in this pH range. The pH had a significant impact on the porous structure, which should be kept in mind when evaluating experimental results, as the pH may be significantly decreased in vitro, in vivo and in situ. The results also show that the initial porosity is very important when using a high-molecular-weight PLG.
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| 19. |
- Fredenberg, Susanne, et al.
(författare)
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The mechanisms of drug release in poly(lactic-co-glycolic acid)-based drug delivery systems-A review.
- 2011
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Ingår i: International Journal of Pharmaceutics. - : Elsevier BV. - 1873-3476 .- 0378-5173. ; 415:1-2, s. 34-52
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Tidskriftsartikel (refereegranskat)abstract
- Poly(d,l-lactic-co-glycolic acid) (PLGA) is the most frequently used biodegradable polymer in the controlled release of encapsulated drugs. Understanding the release mechanisms, as well as which factors that affect drug release, is important in order to be able to modify drug release. Drug release from PLGA-based drug delivery systems is however complex. This review focuses on release mechanisms, and provides a survey and analysis of the processes determining the release rate, which may be helpful in elucidating this complex picture. The term release mechanism and the various techniques that have been used to study release mechanisms are discussed. The physico-chemical processes that influence the rate of drug release and the various mechanisms of drug release that have been reported in the literature are analyzed in this review, and practical examples are given. The complexity of drug release from PLGA-based drug delivery systems can make the generalization of results and predictions of drug release difficult. However, this complexity also provides many possible ways of solving problems and modifying drug release. Basic, generally applicable and mechanistic research provides pieces of the puzzle, which is useful in the development of controlled-release pharmaceuticals.
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| 20. |
- Goel, Suchi, et al.
(författare)
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RIFINs are adhesins implicated in severe Plasmodium falciparum malaria
- 2015
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Ingår i: Nature Medicine. - : Springer Science and Business Media LLC. - 1078-8956 .- 1546-170X. ; 21:4, s. 314-317
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Tidskriftsartikel (refereegranskat)abstract
- Rosetting is a virulent Plasmodium falciparum phenomenon associated with severe malaria. Here we demonstrate that P. falciparum-encoded repetitive interspersed families of polypeptides (RIFINs) are expressed on the surface of infected red blood cells (iRBCs), bind to RBCs-preferentially of blood group A-to form large rosettes and mediate microvascular binding of iRBCs. We suggest that RIFINs have a fundamental role in the development of severe malaria and thereby contribute to the varying global distribution of ABO blood groups in the human population.
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