| 1. |
- Cukalevski, Risto, et al.
(författare)
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Structural Changes in Apolipoproteins Bound to Nanoparticles
- 2011
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Ingår i: Langmuir. - : American Chemical Society (ACS). - 0743-7463 .- 1520-5827. ; 27:23, s. 14360-14369
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Tidskriftsartikel (refereegranskat)abstract
- Nanoparticles are widely used in the pharmaceutical and food industries, but the consequences of exposure to the human body have not been thoroughly investigated. Apolipoprotein A-I (apoAI), the major protein in high-density lipoprotein (HDL), and other lipoproteins are found in the corona around many nanopartides, but data on protein structural and functional effects are lacking. Here we investigate the structural consequences of the adsorption of apoAI, apolipoprotein B100 (apoB100), and HDL on polystyrene nanoparticles with different surface charges. The results of circular dichroism, fluorescence spectroscopy, and limited proteolysis experiments indicate effects on both secondary and tertiary structures. Plain and negatively charged nanoparticles induce helical structure in apoAI (negative net charge) whereas positively charged nanoparticles reduce the amount of helical structure. Plain and negatively charged partides induce a small blue shift in the tryptophan fluorescence spectrum, which is not noticed with the positively charged particles. Similar results are observed with reconstituted HDL. In apoB100, both secondary and tertiary structures are perturbed by all particles. To investigate the generality of the role of surface charge, parallel experiments were performed using human Serum albumin (HSA, negative net charge) and lysozyme (positive net charge). Again, the secondary structure is most affected by nanoparticles carrying an opposite surface charge relative to the protein. Nanoparticles carrying the same net charge as the protein induce only minor structural changes in lysozyme whereas a moderate change is observed for HSA. Thus, surface charge is a critical parameter for predicting structural changes in adsorbed proteins, yet the effect is specific for each protein.
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| 2. |
- Dell'Orco, Daniele, et al.
(författare)
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Modeling the Time Evolution of the Nanoparticle-Protein Corona in a Body Fluid
- 2010
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 5
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Tidskriftsartikel (refereegranskat)abstract
- Background: Nanoparticles in contact with biological fluids interact with proteins and other biomolecules, thus forming a dynamic corona whose composition varies over time due to continuous protein association and dissociation events. Eventually equilibrium is reached, at which point the continued exchange will not affect the composition of the corona. Results: We developed a simple and effective dynamic model of the nanoparticle protein corona in a body fluid, namely human plasma. The model predicts the time evolution and equilibrium composition of the corona based on affinities, stoichiometries and rate constants. An application to the interaction of human serum albumin, high density lipoprotein (HDL) and fibrinogen with 70 nm N-iso-propylacrylamide/N-tert-butylacrylamide copolymer nanoparticles is presented, including novel experimental data for HDL. Conclusions: The simple model presented here can easily be modified to mimic the interaction of the nanoparticle protein corona with a novel biological fluid or compartment once new data will be available, thus opening novel applications in nanotoxicity and nanomedicine.
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| 3. |
- Ferreira, Silvia A., et al.
(författare)
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Biocompatibility of mannan nanogel-safe interaction with plasma proteins
- 2012
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Ingår i: Biochimica et Biophysica Acta. General Subjects. - : Elsevier BV. - 0304-4165. ; 1820:7, s. 1043-1051
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Tidskriftsartikel (refereegranskat)abstract
- Background: Self-assembled mannan nanogels are designed to provide a therapeutic or vaccine delivery platform based on the bioactive properties of mannan to target mannose receptor expressed on the surface of antigen-presenting cells, combined with the performance of nanogels as carriers of biologically active agents. Methods: Proteins in the corona around mannan nanogel formed in human plasma were identified by mass spectrometry after size exclusion chromatography or centrifugation followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Structural changes and time dependent binding of human apolipoprotein A-I (apoA-I) and human serum albumin (HSA) to mannan nanogel were studied using intrinsic tryptophan fluorescence and circular dichroism spectroscopy. The mannan nanogel effect on blood coagulation and fibrillation of Alzheimer's disease-associated amyloid beta peptide and hemodialysis-associated amyloidosis beta 2 microglobulin was evaluated using thrombin generation assay or thioflavin T fluorescence assay, respectively. Results: The protein corona around mannan nanogel is formed through a slow process, is quite specific comprising apolipoproteins B-100, A-I and E and HSA, evolves over time, and the equilibrium is reached after hours to days. Structural changes and time dependent binding of apoA-I and HSA to mannan nanogel are minor. The mannan nanogel does not affect blood coagulation and retards the fibril formation. Conclusions: Mannan nanogel has a high biosafety and biocompatibility, which is mandatory for nanomaterials to be used in biomedical applications. General Significance: Our research provides a molecular approach to evaluate the safety aspects of nanomaterials, which is of general concern in society and science. (C) 2012 Elsevier B.V. All rights reserved.
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| 4. |
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| 5. |
- Hafizi, Sassan, et al.
(författare)
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Tensin2 reduces intracellular phosphatidylinositol 3,4,5-trisphosphate levels at the plasma membrane
- 2010
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Ingår i: Biochemical and Biophysical Research Communications - BBRC. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 399:3, s. 396-401
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Tidskriftsartikel (refereegranskat)abstract
- Tensins are proposed cytoskeleton-regulating proteins. However, Tensin2 additionally inhibits Akt signalling and cell survival. Structural modelling of the Tensin2 phosphatase (PTPase) domain revealed an active site-like pocket receptive towards phosphoinositides. Tensin2-expressing HEK293 cells displayed negligible levels of plasma membrane phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P-3) under confocal microscopy. However, mock-transfected cells, and Tensin2 cells harbouring a putative phosphatase-inactivating mutation, exhibited significant PtdIns(3,4,5)P-3 levels, which decreased upon phosphatidylinositol 3-kinase inhibition with LY294002. In contrast, wtTensin3, mock and mutant cells were identical in membrane PtdIns(3,4,5)P-3 and Akt phosphorylation. In vitro lipid PTPase activity was however undetectable in isolated recombinant PTPase domains of both Tensins, indicating a possible loss of structural stability when expressed in isolation. In summary, we provide evidence that Tensin2, in addition to regulating cytoskeletal dynamics, influences phosphoinositide-Akt signalling through its PTPase domain.
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| 6. |
- Martuszewska, Danuta, et al.
(författare)
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Tensin3 is a negative regulator of cell migration and all four Tensin family members are downregulated in human kidney cancer.
- 2009
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 4:2
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: The Tensin family of intracellular proteins (Tensin1, -2, -3 and -4) are thought to act as links between the extracellular matrix and the cytoskeleton, and thereby mediate signaling for cell shape and motility. Dysregulation of Tensin expression has previously been implicated in human cancer. Here, we have for the first time evaluated the significance of all four Tensins in a study of human renal cell carcinoma (RCC), as well as probed the biological function of Tensin3. PRINCIPAL FINDINGS: Expression of Tensin2 and Tensin3 at mRNA and protein levels was largely absent in a panel of diverse human cancer cell lines. Quantitative RT-PCR analysis revealed mRNA expression of all four Tensin genes to be significantly downregulated in human kidney tumors (50-100% reduction versus normal kidney cortex; P<0.001). Furthermore, the mRNA expressions of Tensins mostly correlated positively with each other and negatively with tumor grade, but not tumor size. Immunohistochemical analysis revealed Tensin3 to be present in the cytoplasm of tubular epithelium in normal human kidney sections, whilst expression was weaker or absent in 41% of kidney tumors. A subset of tumor sections showed a preferential plasma membrane expression of Tensin3, which in clear cell RCC patients was correlated with longer survival. Stable expression of Tensin3 in HEK 293 cells markedly inhibited both cell migration and matrix invasion, a function independent of putative phosphatase activity in Tensin3. Conversely, siRNA knockdown of endogenous Tensin3 in human cancer cells significantly increased their migration. CONCLUSIONS: Our findings indicate that the Tensins may represent a novel group of metastasis suppressors in the kidney, the loss of which leads to greater tumor cell motility and consequent metastasis. Moreover, tumorigenesis in the human kidney may be facilitated by a general downregulation of Tensins. Therefore, anti-metastatic therapies may benefit from restoring or preserving Tensin expression in primary tumors.
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| 7. |
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| 8. |
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| 9. |
- Oslakovic, Cecilia, et al.
(författare)
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Anionic Phospholipids Lose their Procoagulant Properties when Incorporated into High-Density Lipoproteins.
- 2009
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Ingår i: Journal of Biological Chemistry. - 1083-351X. ; 284:9, s. 5896-5904
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Tidskriftsartikel (refereegranskat)abstract
- Blood coagulation involves a series of enzymatic protein complexes that assemble on the surface of anionic phospholipid. To investigate whether apolipoproteins affect coagulation reactions, they where included during the preparation of anionic phospholipid vesicles using a detergent solubilization-dialysis method. Apolipoprotein components of high-density lipoproteins, especially apolipoprotein A-I, had pronounced anticoagulant effect. The anionic phospholipids lost their procoagulant effect when the vesicle preparation method was performed in the presence of apolipoprotein A-I. The anionic phospholipid-apolipoprotein A-I particles were 8-10 nm in diameter and contained around 60-80 phospholipid molecules, depending on the phospholipid composition. The phospholipids of these particles were unable to support the activation of prothrombin by factor Xa in the presence of factor Va, and unable to support binding of factor Va, while binding of prothrombin and factor Xa were efficient. Phospholipid transfer protein was shown to mediate transfer of phospholipids from liposomes to apolipoprotein A-I containing reconstituted high-density lipoprotein. In addition, serum was also shown to neutralize the procoagulant effect of anionic liposomes and to efficiently mediate transfer of phospholipids from liposomes to either apolipoprotein A-I or apolipoprotein B containing particles. In conclusion, apolipoprotein A-I was found to neutralize the procoagulant properties of anionic phospholipids by arranging the phospholipids in surface areas that are too small to accommodate the prothrombinase complex. This anionic phospholipid scavenger function may be an important mechanism to control the exposure of such phospholipids to circulating blood and thereby prevent inappropriate stimulation of blood coagulation.
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| 10. |
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