Endre søk
Begrens søket
1234567 101 - 150 of 535
RefereraExporteraLink til resultatlisten
Permanent link
Referera
Referensformat
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Annet format
Fler format
Språk
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Annet språk
Fler språk
Utmatningsformat
  • html
  • text
  • asciidoc
  • rtf
Treff pr side
  • 5
  • 10
  • 20
  • 50
  • 100
  • 250
Sortering
  • Standard (Relevans)
  • Forfatter A-Ø
  • Forfatter Ø-A
  • Tittel A-Ø
  • Tittel Ø-A
  • Type publikasjon A-Ø
  • Type publikasjon Ø-A
  • Eldste først
  • Nyeste først
  • Skapad (Eldste først)
  • Skapad (Nyeste først)
  • Senast uppdaterad (Eldste først)
  • Senast uppdaterad (Nyeste først)
  • Disputationsdatum (tidligste først)
  • Disputationsdatum (siste først)
  • Standard (Relevans)
  • Forfatter A-Ø
  • Forfatter Ø-A
  • Tittel A-Ø
  • Tittel Ø-A
  • Type publikasjon A-Ø
  • Type publikasjon Ø-A
  • Eldste først
  • Nyeste først
  • Skapad (Eldste først)
  • Skapad (Nyeste først)
  • Senast uppdaterad (Eldste først)
  • Senast uppdaterad (Nyeste først)
  • Disputationsdatum (tidligste først)
  • Disputationsdatum (siste først)
Merk
Maxantalet träffar du kan exportera från sökgränssnittet är 250. Vid större uttag använd dig av utsökningar.
  • 101.
    EL Andaloussi-Lilja, Johanna
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Lundqvist, Jessica
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Forsby, Anna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    TRPV1 expression and activity during retinoic acid-induced neuronal differentiation2009Inngår i: Neurochemistry International, ISSN 0197-0186, E-ISSN 1872-9754, Vol. 55, nr 8, s. 768-774Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The transient receptor potential vanilloid subtype 1 (TRPV1) is a Ca2+-permeable channel primarily expressed in dorsal root ganglion neurons. Besides its function in thermogenic nociception and neurogenic inflammation, TRPV1 is involved in cell migration, cytoskeleton reorganisation and in neuronal guidance.  To explore the TRPV1 level and activity during conditions for neuronal maturation, TRPV1-expressing SHSY5Y neuroblastoma cells were differentiated into a neuronal phenotype using all-trans-retinoic acid (RA). We show that RA highly up-regulated the total and cell surface TRPV1 protein expression but the TRPV1 mRNA level was unaffected. The up-regulated receptors were localised to the cell bodies and the developed neurites. Furthermore, RA increased both the basal intracellular free Ca2+ concentration by 30 % as well as the relative capsaicin-induced Ca2+ influx. The results show that TRPV1 protein expression increases during RA-induced differentiation in vitro, which generates an altered intracellular Ca2+ homeostasis.

  • 102.
    El-Andaloussi, S.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Johansson, H.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi och neurotoxikologi.
    Magnusdottir, A.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi och neurotoxikologi.
    Järver, P.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi och neurotoxikologi.
    Lundberg, P.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi och neurotoxikologi.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi och neurotoxikologi.
    TP10, a delivery vector for decoy oligonucleotides targeting the Myc protein2005Inngår i: Journal of Controlled Release, ISSN 0168-3659, E-ISSN 1873-4995, Vol. 110, nr 1, s. 189-201Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    One approach to investigate gene function, by silencing the activity of certain proteins, is the usage of double stranded decoy oligodeoxynucleotides (ds decoy ODNs). Decoy, in this sense, is ds ODNs bearing the consensus binding sequence for a DNA-binding protein. This can be used in clinical settings to attenuate the effect of overexpressed transcription factors in tumor cells. We here choose to target the oncogenic protein Myc. Since oligonucleotides are poorly internalized to cells, a cell-penetrating peptide, TP10, was coupled to the Myc decoy, using two different strategies. Either TP10 was simply mixed with ds decoy ODNs forming complexes through non-covalent electrostatic interactions, or by having a nona-nucleotide overhang in one of the decoy strands, and adding a complementary PNA sequence coupled to an NLS sequence and TP10, which could hybridize to the Myc decoy.

    By using these strategies, uptake was significantly enhanced, especially with the co-incubation approach. Interestingly, various endocytosis inhibitors had no effect on the uptake pattern, suggesting that uptake of these complexes is not mediated via endocytosis. Finally, a decreased proliferative capacity was observed when treating the neuroblastoma cell line N2a with TP10–PNA conjugate hybridized to Myc decoy compared to naked Myc decoy and untreated cells. A dose-dependent decrease in proliferation was also observed in MCF-7 cells, when using both strategies. These results suggest an alternative way to efficiently deliver ds ODNs into cells using the cell-penetrating peptide TP10 and prevent tumor growth by targeting the oncogenic protein Myc.

  • 103. Elmquist, A.
    et al.
    Hansen, Mats
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Langel, Ü
    Structure-activity relationship study of the cell-penetrating peptide pVEC2006Inngår i: Biochim. Biophys. Acta, nr 1758, s. 721-729Artikkel i tidsskrift (Fagfellevurdert)
  • 104.
    Eriksson, Charlotta
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Hallberg, Einar
    Dynamic properties of pore complex proteins in gp210 deficient cells.Manuskript (Annet vitenskapelig)
  • 105.
    Eriksson, Jonas
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Gene therapy tools: oligonucleotides and peptides2016Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Genetic mutations can cause a wide range of diseases, e.g. cancer. Gene therapy has the potential to alleviate or even cure these diseases. One of the many gene therapies developed so far is RNA-cleaving deoxyribozymes, short DNA oligonucleotides that specifically bind to and cleave RNA. Since the development of these synthetic catalytic oligonucleotides, the main way of determining their cleavage kinetics has been through the use of a laborious and error prone gel assay to quantify substrate and product at different time-points. We have developed two new methods for this purpose. The first one includes a fluorescent intercalating dye, PicoGreen, which has an increased fluorescence upon binding double-stranded oligonucleotides; during the course of the reaction the fluorescence intensity will decrease as the RNA is cleaved and dissociates from the deoxyribozyme. A second method was developed based on the common denominator of all nucleases, each cleavage event exposes a single phosphate of the oligonucleotide phosphate backbone; the exposed phosphate can simultaneously be released by a phosphatase and directly quantified by a fluorescent phosphate sensor. This method allows for multiple turnover kinetics of diverse types of nucleases, including deoxyribozymes and protein nucleases.

    The main challenge of gene therapy is often the delivery into the cell. To bypass cellular defenses researchers have used a vast number of methods; one of these are cell-penetrating peptides which can be either covalently coupled to or non-covalently complexed with a cargo to deliver it into a cell. To further evolve cell-penetrating peptides and understand how they work we developed an assay to be able to quickly screen different conditions in a high-throughput manner. A luciferase up- and downregulation experiment was used together with a reduction of the experimental time by 1 day, upscaling from 24- to 96-well plates and the cost was reduced by 95% compared to commercially available assays. In the last paper we evaluated if cell-penetrating peptides could be used to improve the uptake of an LNA oligonucleotide mimic of GRN163L, a telomerase-inhibiting oligonucleotide. The combination of cell-penetrating peptides and our mimic oligonucleotide lead to an IC50 more than 20 times lower than that of GRN163L.

  • 106.
    Eriksson, Jonas
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Kinetic assays for RNA-cleaving deoxyribozymes and other nucleases2016Licentiatavhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    In this thesis two different assays for real-time RNA-cleaving deoxyribozyme and general nuclease kinetics are presented. Previous publications on nuclease kinetic assays have been riddled with drawbacks of labeling, discontinuity, cost etc. To tackle some of the drawbacks two assays were developed; the first specifically for RNA-cleaving deoxyribozymes to allow real-time kinetic measurements independently of whether the deoxyribozyme has low or high levels of secondary structure and when cleaving a full length messenger RNA (mRNA) substrate; the second assay was developed as a means to measure kinetics of virtually any nuclease by utilizing the single ubiquitous phenomenon in nuclease cleavage, the exposure of a phosphate upon hydrolysis of the phosphate backbone.

    In Paper I the assay for RNA-cleaving deoxyribozyme kinetics is presented as a development of a previously published assay. The search for a fluorescent intercalating dye with more preferential properties than ethidium bromide resulted in PicoGreen. This dye allowed the assay to be used for deoxyribozymes with low and high levels of secondary structure as well as using full length mRNA substrates.

    Paper II presents the second assay of this thesis, an assay where phosphates exposed by nuclease cleavage are released from their products by phosphatases; the released inorganic phosphates are quantified in real-time by a biosensor. The assay allows for real-time kinetics without the use of labels (i.e. natural enzymes and substrates). Regardless of whether the nuclease was a protein, nucleic acid-based, an exo- or endonuclease, processive or single-target nuclease the assay suited them equally well.

  • 107.
    Eriksson, Jonas
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Helmfors, Henrik
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    A High-Throughput Kinetic Assay for RNA-Cleaving Deoxyribozymes2015Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 10, nr 8, artikkel-id e0135984Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Determining kinetic constants is important in the field of RNA-cleaving deoxyribozymes (DNAzymes). Using todays conventional gel assays for DNAzyme assays is time-consuming and laborious. There have been previous attempts at producing new and improved assays; however these have drawbacks such as incompatibility with structured DNAzymes, enzyme or substrate modifications and increased cost. Here we present a new method for determining single-turnover kinetics of RNA-cleaving DNAzymes in real-time and in a high-throughput fashion. The assay is based on an intercalating fluorescent dye, PicoGreen, with high specificity for double-stranded DNA and heteroduplex DNA-RNA in this case formed between the DNAzyme and the target RNA. The fluorescence decreases as substrate is converted to product and is released from the enzyme. Using a Flexstation II multi-mode plate reader with built in liquid handling we could automate parts of the assay. This assay gives the possibility to determine single-turnover kinetics for up to 48 DNAzymes simultaneously. As the fluorescent probe is extrinsic there is no need for enzyme or substrate modifications, making this method less costly compared to other methods. The main novelty of this assay is the possibility of using full-length mRNA as the DNAzyme target.

  • 108.
    Eriksson, Jonas
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Quantitative Microplate Assay for Real-Time Nuclease Kinetics2016Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 11, nr 4, artikkel-id e0154099Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Utilizing the phenomenon of nucleases exposing oligonucleotide phosphate backbones to phosphatases we present a novel quantitative method for kinetics of nuclease catalysis. Inorganic phosphate released from nuclease products by phosphatases could be quantified in real-time by a fluorescent sensor of inorganic phosphate. Two different nucleases were employed, showing the versatility of this assay for multiple turnover label-free nuclease studies.

  • 109.
    Eriksson, Olaspers Sara
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Geörg, Miriam
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Sjölinder, Hong
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Sillard, Rannar
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Lindberg, Staffan
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Jonsson, Ann-Beth
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Identification of Cell-Penetrating Peptides That Are Bactericidal to Neisseria meningitidis and Prevent Inflammatory Responses upon Infection2013Inngår i: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 57, nr 8, s. 3704-3712Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Meningococcal disease is characterized by a fast progression and a high mortality rate. Cell-penetrating peptides (CPPs), developed as vectors for cargo delivery into eukaryotic cells, share structural features with antimicrobial peptides. A screen identified two CPPs, transportan-10 (TP10) and model amphipathic peptide (MAP), with bactericidal action against Neisseria meningitidis. Both peptides were active in human whole blood at micromolar concentrations, while hemolysis remained negligible. Additionally, TP10 exhibited significant antibacterial activity in vivo. Uptake of SYTOX green into live meningococci was observed within minutes after TP10 treatment, suggesting that TP10 may act by membrane permeabilization. Apart from its bactericidal activity, TP10 suppressed inflammatory cytokine release from macrophages infected with N. meningitidis as well as from macrophages stimulated with enterobacterial and meningococcal lipopolysaccharide (LPS). Finally, incubation with TP10 reduced the binding of LPS to macrophages. This novel endotoxin-inhibiting property of TP10, together with its antimicrobial activity in vivo, indicates the possibility to design peptide-based therapies for infectious diseases.

  • 110.
    Eriksson Sollenberg, Ulla
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Characterization of galanin receptors using chimeric peptides and site-directed mutagenesis2010Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Galanin, a 29 (30) amino acid neuropeptide, is found throughout both the central and peripheral nervous systems. It signals via three receptors, GalR1-3, all belonging to the rhodopsin-like G-protein coupled receptors. Galanin and its receptors have been implicated in a vast variety of biological processes. To facilitate further characterization of the physiological/pathological roles of galanin, subtype selective ligands targeting the three receptors individually would be of great aid.

    In this thesis the main objective was to provide more information about galanin receptor-ligand interactions, primarily concerning GalR2 and 3.  By using information gained from previously developed chimeric peptides, we designed and synthesized a novel peptide selective towards GalR2 (Paper I). This peptide, M871, binds GalR2 in an inhibitory manner, likely due to its truncated N-terminus and bulky character. In Paper II and IV we performed L-alanine mutagenesis assays of GalR2 and 3 respectively. By point substituting amino acid residues in the receptor sequence, we identified crucial pharmacophores for ligand binding, primarily in transmembrane regions 6 and 7. The targeted residues were selected based on knowledge concerning GalR1 and on conservation between the three receptors. For GalR3 we also conducted a computational docking assay. A homology model was first constructed using three crystallized structures of other receptors also belonging to the Rhodopsin family. Ligands galanin(2-6) and SNAP398299 were then docked to GalR3 in flexible mode. The docking resulted in characterization of GalR3-ligand interactions and conclude that this receptor display a relatively deep and narrow binding pocket. As a result of this, it was hypothesized that the C-terminus of ligands is of importance for GalR3 affinity. An L-alanine scan of ligand was performed (paper III), which confirmed this theory.

    In conclusion, our results give insights into galanin receptor-ligand interactions, information that is relevant for ligand design and drug development.

  • 111. Eriste, Elo
    et al.
    Kurrikoff, Kaido
    Suhorutsenko, Julia
    Osokolkov, Nikita
    Copolovici, Dana Maria
    Jones, Sarah
    Laakkonen, Pirjo
    Howl, John
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. University of Tartu, Estonia.
    Peptide-Based Glioma-Targeted Drug Delivery Vector gHoPe22013Inngår i: Bioconjugate chemistry, ISSN 1043-1802, E-ISSN 1520-4812, Vol. 24, nr 3, s. 305-313Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Gliomas are therapeutically challenging cancers with poor patient prognosis. New drug delivery strategies are needed to achieve a more efficient chemotherapy-based approach against brain tumors. The current paper demonstrates development of a tumor-targeted delivery vector that is based on a cell-penetrating peptide pVEC and a novel glioma-targeting peptide sequence gHo. The unique tumor-homing peptide gHo was identified using in vitro phage display technology. The novel delivery vector, which we designated as gHoPe2, was constructed by a covalent conjugation of pVEC, gHo, and a cargo; the latter could be either a labeling moiety (such as a fluorescent marker) or a cytostatic entity. Using a fluorescent marker, we demonstrate efficient uptake of the vector in glioma cells and selective labeling of glioma xenograft tumors in a mouse model. This is the first time that we know where in vitro phage display has yielded an efficient, in vivo working vector. We also demonstrate antitumor efficacy of the delivery vector gHoPe2 using a well-characterized chemotherapeutic drug doxorubicin. Vectorized doxorubicin proved to be more efficient than the free drug in a mouse glioma xenograft model after systemic administration of the drugs. In conclusion, we have characterized a novel glioma-homing peptide gHo, demonstrated development of a new and potential glioma-targeted drug delivery vector gHoPe2, and demonstrated the general feasibility of the current approach for constructing cell-penetrating peptide-based targeted delivery systems.

  • 112.
    Erlandsson, Mikael
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Methodological studies in solid phase synthesis: Linkers and applications of multi-component condensations2006Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Solid phase synthesis has become an increasingly important tool in the synthesis of oligopolymers and small organic molecules. This thesis covers three important areas in the solid phase synthesis technique: linker strategies, reduction methods and novel routes to complex heterocycles.

    The synthesis of the OMPPA [4-(3-hydroxy-4-metylpentyl)phenyl acetic acid] linker is described. This linker is compatible with the Boc/Bzl protective group strategy, and yields peptide acids upon cleavage from solid support. The OMPPA linker is stable towards “low-acid” treatment and is self-scavenging during final cleavage of the peptide product from solid support. These properties are beneficial for peptide purity and yields.

    The synthesis of the HMPPA [3-(4-hydroxymethylphenylsulfanyl)propanoic acid] linker is described. This safety catch linker is compatible with both Fmoc/tBu- and Boc/Bzl protective group strategies in solid phase peptide synthesis. The HMPPA linker is stable towards super acids yet final cleavage from solid support is performed by a relatively mild reductive acidolysis method, yielding peptide acids. It is suggested that this linker may be useful when synthesizing cyclic peptides on solid support. A new facile method for reducing cystine moieties is described. Adding metallic zinc to cystine containing peptides and proteins dissolved in slightly acidic aqueous and/or non-aqueous solutions, results in rapid disulfide reduction. This method is compatible with functional groups commonly present in peptides and proteins. The solid phase synthesis of oxygen-bridged tetrahydropyridones via a multi-component condensation reaction is described. Expected products were obtained in reasonable yields using both aromatic- and aliphatic ketones. This class of compounds has the general physico-chemical properties that are typical for drugs with high pharmacological activity.

  • 113.
    Erlandsson, Mikael
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Undén, Anders
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    3-(4-hydroxymethylphenylsulfanyl)propanoic acid (HMPPA) as a new safety catch linker in solid phase peptide synthesis2006Inngår i: Tetrahedron Letters, ISSN 0040-4039, E-ISSN 1359-8562, Vol. 32, nr 7, s. 5829-5832Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    A new safety catch linker, 3-(4-hydroxymethylphenylsulfanyl)propanoic acid (HMPPA), is described for use in solid phase peptide synthesis. The linker is readily synthesized from commercially available chemicals in a more cost efficient way compared to similar reported linkers. The HMPPA linker is easily attached to an amino derivatized solid support followed by on-resin oxidation of the thioether to sulfoxide, thereby making the linker very stable towards strong acid treatment. Final resin cleavage is performed by reductive acidolysis.

  • 114. Ertan-Bolelli, Tugba
    et al.
    Bolelli, Kayhan
    Musdal, Yaman
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Yildiz, Ilkay
    Aki-Yalcin, Esin
    Mannervik, Bengt
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Yalcin, Ismail
    Design and synthesis of 2-substituted-5-(4-trifluoromethylphenyl-sulphonamido)benzoxazole derivatives as human GST P1-1 inhibitors2018Inngår i: Artificial Cells, Nanomedicine, and Biotechnology, ISSN 2169-1401, Vol. 46, nr 3, s. 510-517Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The glutathione transferases (GSTs) are a family of widely distributed Phase II detoxification enzymes. GST P1-1 is frequently overexpressed in rat and human tumours. It is suggested that overexpression of hGST P1-1 by human tumor cells may play a role in resistance to cancer chemotherapy. Hence, hGST P1-1 can be a promising target for cancer treatment. In this study, new hGST P1-1 inhibitors, 2-(4-substitutedphenyl/benzyl)-5-(4-trifluoromethylphenylsulphonamido) benzoxazole derivatives (Va-Vk) have been designed and synthesized. Surprisingly, in vitro hGST P1-1 enzyme inhibition studies demonstrated that all of the tested compounds except Vj had better activity than the reference drug EA and it is also correlated with the docking results. Additionally we compared the interactions with hGST P1-1 enzyme of newly synthesized compound Vh (bearing CF3 group) and previously synthesized compound 5f (bearing NO2 group). According to the docking results, compound Vh bound to the hGST P1-1 enzyme with a higher affinity compared to 5f. Therefore, we can consider that these data make a sense and can explain its higher activity. The compounds that obtained from this research could be used as scaffolds in design of new potent hGST P1-1 inhibitors useful in the treatment of the resistance of cancer chemotherapy.

  • 115.
    Ezzat, Kariem
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    CELL PENETRATING PEPTIDES: CHEMICAL MODIFICATION AND FORMULATION DEVELOPMENT2011Licentiatavhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Cell penetrating peptides (CPPs) have been extensively studied and exploited as drug delivery vectors for a wide variety of therapeu-tic cargos. However, several issues remain to be addressed regarding the enhancement of their efficiency and stability. In addition, to be available for patients, CPP-based therapeutics have to be formulated into suitable pharmaceutical forms that can be readily manufactured, transported, stored and conveniently used.In this thesis, three chemically modified CPPs are developed having superior delivery properties for several nucleic acid-based the-rapeutic cargoes including: plasmids, small interfering RNA (siRNA) and splice switching oligonucleutides (SSOs), in different in-vitro and in-vivo models. In Paper I, we show that an N-terminally stearic acid-modified version of transportan-10 (TP10) can form stable nanopar-ticles with plasmids that efficiently transfect different cell types and can mediate efficient gene delivery in-vivo when administrated intra muscularly (i.m.) or intradermaly (i.d.). In paper II, stearyl-TP10 is further modified with pH titratable trifluoromethylquinoline moieties to facilitate endosomal release. The new peptide, denoted PepFect 6 (PF6), elicited robust RNAi responses when complexed with siRNA in several cell models and promoted strong RNAi responses in differ-ent organs following systemic delivery in mice without any associated toxicity. In paper III , a new peptide with ornithine modification, PF14, is shown to efficiently deliver SSOs in different cell models including HeLa pLuc705 and mdx mouse myotubes; a cell culture model of Duchenne‟s muscular dystrophy (DMD). Additionally, we have developed a method for incorporating this delivery system into solid formulation that could be suitable for several therapeutic appli-cations. Solid dispersion technique is utilized and the formed solid formulations are as active as the freshly prepared nanocparticles in solution even when stored at elevated temperatures for several weeks.Taken together, these results demonstrate that certain chemical modifications could drastically enhance the activity and stability of CPPs in-vitro and in-vivo. Moreover, we show that CPP-based thera-peutics could be formulated into convenient and manufacturable do-sage forms.

  • 116.
    Ezzat, Kariem
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Cell-penetrating peptides; chemical modification, mechanism of uptake and formulation development2012Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Gene therapy holds the promise of revolutionizing the way we treat diseases. By using recombinant DNA and oligonucleotides (ONs), gene functions can be restored, altered or silenced according to the therapeutic need. However, gene therapy approaches require the delivery of large and charged nucleic acid-based molecules to their intracellular targets across the plasma membrane, which is inherently impermeable to such molecules. In this thesis, two chemically modified cell-penetrating peptides (CPPs) that have superior delivery properties for several nucleic acid-based therapeutics are developed. These CPPs can spontaneously form nanoparticles upon non-covalent complexation with the nucleic acid cargo, and the formed nanoparticles mediate efficient cellular transfection. In paper I, we show that an N-terminally stearic acid-modified version of transportan-10 (PF3) can efficiently transfect different cell types with plasmid DNA and mediates efficient gene delivery in-vivo when administrated intra muscularly (i.m.) or intradermaly (i.d.). In paper II, a new peptide with ornithine modification, PF14, is shown to efficiently deliver splice-switching oligonucleotides (SSOs) in different cell models including mdx mouse myotubes; a cell culture model of Duchenne’s muscular dystrophy (DMD). Additionally, we describe a method for incorporating the PF14-SSO nanoparticles into a solid formulation that is active and stable even when stored at elevated temperatures for several weeks. In paper III, we demonstrate the involvement of class-A scavenger receptor subtypes (SCARA3 & SCARA5) in the uptake of PF14-SSO nanoparticles, which possess negative surface charge, and suggest for the first time that some CPP-based systems function through scavenger receptors. In paper IV, the ability of PF14 to deliver siRNA to different cell lines is shown and their stability in simulated gastric acidic conditions is highlighted.

    Taken together, these results demonstrate that certain chemical modifications can drastically enhance the activity and stability of CPPs for delivering nucleic acids after spontaneous nanoparticle formation upon non-covalent complexation. Moreover, we show that CPP-based nanoparticles can be formulated into convenient and stable solid formulations that can be suitable for several therapeutic applications. Importantly, the involvement of scavenger receptors in the uptake of such nanoparticles is presented, which could yield novel possibilities to understand and improve the transfection by CPPs and other gene therapy nanoparticles.

  • 117.
    Ezzat, Kariem
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    EL Andaloussi, Samir
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Abdo, Rania
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Peptide-based matrices as drug delivery vehicles2010Inngår i: Current pharmaceutical design, ISSN 1381-6128, E-ISSN 1873-4286, Vol. 16, nr 9, s. 1167-1178Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Peptides, polypeptides and proteins have been extensively studied for their various structural and functional roles in living organisms. However, breakthrough discoveries in the last decades identified some peptide-based matrices that posses the ability to traverse biological membranes, and many peptides, polypeptides and even complete proteins have been shown to have such properties. Hence, these matrices have been successfully used for the intracellular delivery of many therapeutic cargos including small molecules, proteins, peptides, oligonucleutides, plasmids and nanoparticles both in vitro and in vivo. Being neither toxic nor carcinogenic and meanwhile efficient in delivery, they are recognized as very promising vectors to overcome the shortcomings of the available technologies. The characteristics of these peptide-based matrices and their applications in drug delivery are here briefly illustrated together with current challenges and future prospects.

  • 118.
    Ezzat, Kariem
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    EL Andaloussi, Samir
    Zaghloul, Eman M.
    Lehto, Taavi
    Lindberg, Staffan
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Moreno, Pedro M. D.
    Viola, Joana R.
    Magdy, Tarek
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Abdo, Rania
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Guterstam, Peter
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Sillard, Rannar
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Hammond, Suzan M.
    Wood, Matthew J. A.
    Arzumanov, Andrey A.
    Gait, Michael J.
    Smith, C. I. Edvard
    Hällbrink, Mattias
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. University of Tartu, Estonia.
    PepFect 14, a novel cell-penetrating peptide for oligonucleotide delivery in solution and as solid formulation2011Inngår i: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 39, nr 12, s. 5284-5298Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Numerous human genetic diseases are caused by mutations that give rise to aberrant alternative splicing. Recently, several of these debilitating disorders have been shown to be amenable for splice-correcting oligonucleotides (SCOs) that modify splicing patterns and restore the phenotype in experimental models. However, translational approaches are required to transform SCOs into usable drug products. In this study, we present a new cell-penetrating peptide, PepFect14 (PF14), which efficiently delivers SCOs to different cell models including HeLa pLuc705 and mdx mouse myotubes; a cell culture model of Duchenne's muscular dystrophy (DMD). Non-covalent PF14-SCO nanocomplexes induce splice-correction at rates higher than the commercially available lipid-based vector Lipofectamine™ 2000 (LF2000) and remain active in the presence of serum. Furthermore, we demonstrate the feasibility of incorporating this delivery system into solid formulations that could be suitable for several therapeutic applications. Solid dispersion technique is utilized and the formed solid formulations are as active as the freshly prepared nanocomplexes in solution even when stored at an elevated temperatures for several weeks. In contrast, LF2000 drastically loses activity after being subjected to same procedure. This shows that using PF14 is a very promising translational approach for the delivery of SCOs in different pharmaceutical forms.

  • 119.
    Ezzat, Kariem
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Helmfors, Henrik
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Tudoran, Oana
    Juks, Carmen
    Lindberg, Staffan
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Padari, Kärt
    EL Andaloussi, Samir
    Pooga, Margus
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. University of Tartu, Estonia.
    Scavenger receptor-mediated uptake of cell-penetrating peptide nanoparticles with oligonucleotides2012Inngår i: The FASEB Journal, ISSN 0892-6638, E-ISSN 1530-6860, Vol. 26, nr 3, s. 1172-1180Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Cell-penetrating peptides (CPPs) are shortcationic peptides that penetrate cells by interacting withthe negatively charged plasma membrane; however, thedetailed uptake mechanism is not clear. In contrary to theconventional mode of action of CPPs, we show here thata CPP, PepFect14 (PF14), forms negatively charged nanocomplexeswith oligonucleotides and their uptake is mediatedby class-A scavenger receptors (SCARAs). Specificinhibitory ligands of SCARAs, such as fucoidin, polyinosinicacid, and dextran sulfate, totally inhibit the activityof PF14-oligonucleotide nanocomplexes in the HeLapLuc705 splice-correction cell model, while nonspecific,chemically related molecules do not. Furthermore, RNAinterference (RNAi) knockdown of SCARA subtypes(SCARA3 and SCARA5) that are expressed in this cell lineled to a significant reduction of the activity to <50%. Inline with this, immunostaining shows prevalent colocalizationof the nanocomplexes with the receptors, and electronmicroscopy images show no binding or internalizationof the nanocomplexes in the presence of theinhibitory ligands. Interestingly, naked oligonucleotidesalso colocalize with SCARAs when used at high concentrations.These results demonstrate the involvement ofSCARA3 and SCARA5 in the uptake of PF14-oligonucleotidenanocomplexes and suggest for the first time thatsome CPP-based systems function through scavenger receptors,which could yield novel possibilities to understandand improve the transfection by CPPs.

  • 120.
    Ezzat, Kariem
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Zaghloul, Eman M.
    EL Andaloussi, Samir
    Lehto, Taavi
    Hilal, Ramy
    Magdy, Tarek
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Smith, Edvard C. I.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Solid formulation of cell-penetrating peptide nanoparticles with siRNA and their stability in simulated gastric conditions2012Inngår i: Journal of Controlled Release, ISSN 0168-3659, E-ISSN 1873-4995, Vol. 162, nr 1, s. 1-8Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Cell-penetrating peptides (CPPs) are short cationic peptides that have been extensively studied as drug delivery vehicles for proteins, nucleic acids and nanoparticles. However, the formulation of CPP-based therapeutics into different pharmaceutical formulations and their stability in relevant biological environments have not been given the same attention. Here, we show that a newly developed CPP, PepFect 14 (PF14), forms non-covalent nanocomplexes with short interfering RNA (siRNA), which are able to elicit efficient RNA-interference (RNAi) response in different cell-lines. RNAi effect was obtained at low siRNA doses with a unique kinetic profile. Furthermore, we utilized the solid dispersion technique to formulate PF14/siRNA nanocomplexes into solid formulations that were as active as the freshly prepared nanocomplexes in solution. Importantly, the freshly prepared nanocomplexes and solid formulations were stable after incubation with simulated gastric fluid having a pH of 1.2 and containing proteolytic enzymes. These results demonstrate the activity of PF14 in delivering and protecting siRNA in different pharmaceutical forms and biological environments.

  • 121. Fedulova, Natalia
    et al.
    Mannervik, Bengt
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Experimental conditions affecting functional comparison of highly active glutathione transferases2011Inngår i: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 413, nr 1, s. 16-23Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Glutathione transferases (GSTs, EC 2.5.1.18) possess multiple functions and have potential applications in biotechnology. Direct evidence of underestimation of activity of human GST A3-3 and porcine GST A2-2 measured at submicromolar enzyme concentrations is reported here for the first time. The combination of time-dependent and enzyme concentration-dependent loss of activity and the choice of the organic solvent for substrates were found to cause irreproducibility of activity measurements of GSTs. These effects contribute to high variability of activity values of porcine GST A2-2 and human Alpha-class GSTs reported in the literature. Adsorption of GSTs to surfaces was found to be the main explanation of the observed phenomena. Several approaches to improved functional comparison of highly active GSTs are proposed.

  • 122. Fedulova, Natalia
    et al.
    Raffalli-Mathieu, Françoise
    Mannervik, Bengt
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Characterization of porcine Alpha-class glutathione transferase A1-12011Inngår i: Archives of Biochemistry and Biophysics, ISSN 0003-9861, E-ISSN 1096-0384, Vol. 507, nr 2, s. 205-211Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    An Alpha-class glutathione transferase (GST) has been cloned from pig gonads. In addition to two conservative point mutations our nucleotide sequence presents a frame shift resulting from a missing A as compared to a previously published porcine GST A1-1 sequence. The deduced C-terminal amino-acid segment of the protein differs between the two variants. Repeated sequencing of cDNA isolated from different tissues and animals ruled out the possibility of a cloning artifact, and the deduced amino acid sequence of our clone showed higher similarity to related mammalian GST sequences. Hereafter, we refer to our cloned enzyme as GST A1-1 and to the previously published enzyme as GST A1-1(∗). The study of the tissue distribution of the GSTA1 mRNA revealed high expression levels in many organs, in particular adipose tissue, liver, and pituitary gland. Porcine GST A1-1 was expressed in Escherichia coli and its kinetic properties were determined using alternative substrates. The catalytic activity in steroid isomerization reactions was at least 10-fold lower than the corresponding values for porcine GST A2-2, whereas the activity with 1-chloro-2,4-dinitrobenzene was approximately 8-fold higher. Differences in the H-site residues of mammalian Alpha-class GSTs may explain the catalytic divergence.

  • 123. Fernaeus, Sandra
    et al.
    Hälldin, Jonas
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Bedecs, Katarina
    Land, Tiit
    Changed iron regulation in scrapie-infected neuroblastoma cells2005Inngår i: Molecular brain research, ISSN 0169-328X, Vol. 133, nr 2, s. 266-273Artikkel i tidsskrift (Fagfellevurdert)
  • 124.
    Fernaeus, Sandra
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Hälldin, Jonas
    Bedecs, Katarina
    Land, Tiit
    Changed iron regulation in scrapie-infected neuroblastoma cells2005Inngår i: Molecular Brain Research, ISSN 0169-328X, Vol. 133, nr 2, s. 266-273Artikkel i tidsskrift (Fagfellevurdert)
  • 125.
    Fernaeus, Sandra
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Land, Tiit
    Increased iron-induced oxidative stress and toxicity in scrapie-infected neuroblastoma cells2005Inngår i: Neuroscience Letters, ISSN 0304-3940, Vol. 382, nr 3, s. 217-220Artikkel i tidsskrift (Fagfellevurdert)
  • 126.
    Fernaeus, Sandra
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Reis, Katarina
    Bedecs, Katarina
    Land, Tiit
    Increased susceptibility to oxidative stress in scrapie-infected neuroblastoma cells is associated with intracellular iron status2005Inngår i: Neuroscience Letters, ISSN 0304-3940, Vol. 389, nr 3, s. 133-136Artikkel i tidsskrift (Fagfellevurdert)
  • 127.
    Fernaeus, Sandra
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Reis, Katarina
    Hälldin, Jonas
    Bedecs, Katarina
    Land, Tiit
    Differential effects of lipopolysaccharide on the expression of ferritin and transferrin receptor between wild-type and scrapie-infected mouse neuroblastoma cellsManuskript (Annet vitenskapelig)
  • 128.
    Ferreira Vasconcelos, Luis Daniel
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Complexes of cell-penetrating peptides with oligonucleotides: Structure, binding and translocation in lipid membranes2017Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    The fundamental element of life known to man is the gene. The information contained in genes regulates all cellular functions, in health and disease. The ability to selectively alter genes or their transcript intermediates with designed molecular tools, as synthetic oligonucleotides, represents a paradigm shift in human medicine.

    The full potential of oligonucleotide therapeutics is however dependent on the development of efficient delivery vectors, due to their intrinsic characteristics, as size, charge and low bioavailability. Cell-penetrating peptides are short sequences of amino acids that are capable of mediating the transport of most types of oligonucleotide therapeutics to the cell interior. It is the interaction of cell-penetrating peptides with oligonucleotides and the transport of their non-covalently formed complexes across the cellular membrane, that constitutes the main subject of this thesis.

    In Paper I we studied the effects of different types of oligonucleotide cargo in the capacity of cationic and amphipathic peptides to interact with lipid membranes. We found that indeed the cargo sequesters some of the peptide’s capacity to interact with membranes. In Paper II we revealed the simultaneous interaction of different molecular and supramolecular peptide and peptide/oligonucleotide species in equilibrium, with the cellular membrane. In Paper III we developed a series of peptides with improved affinity for oligonucleotide cargo as well as enhanced endosomal release and consequently better delivery capacity. In Paper IV we investigated the effect of saturated fatty acid modifications to a cationic cell-penetrating peptide. The varying amphipathicity of the peptide correlated with the complex physicochemical properties and with its delivery efficiency.

    This thesis contributes to the field with a set of characterized mechanisms and physicochemical properties for the components of the ternary system – cell-penetrating peptide, oligonucleotide and cell membrane – that should be considered for the future development of gene therapy.

  • 129.
    Ferreira Vasconcelos, Luis Daniel
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Oligonucleotide Complexes with Cell-Penetrating Peptides: Structure, Binding, Translocation and Flux in Lipid Membranes2014Licentiatavhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    The ability of cell-penetrating peptides to cross plasma membranes has been explored for various applications, including the delivery of bioactive molecules to inhibit disease-causing cellular processes. The uptake mechanisms by which cell-penetrating peptides enter cells depend on the conditions, such as the cell line the concentration and the temperature. To be used as therapeutics, each novel cell-penetrating peptide needs to be fully characterized, including their physicochemical properties, their biological activity and their uptake mechanism. Our group has developed a series of highly performing, non-toxic cell-penetrating peptides, all derived from the original sequence of transportan 10. These analogs are called PepFects and NickFects and they are now a diverse family of N-terminally stearylated peptides. These peptides are known to form noncovalent, nano-sized complexes with diverse oligonucleotide cargoes. One bottleneck that limits the use of this technology for gene therapy applications is the efficient release of the internalized complexes from endosomal vesicles.

    The general purpose of this thesis is to reveal the mechanisms by which our in house designed peptides enter cells and allow the successful transport of biofunctional oligonucleotide cargo. To reach this goal, we used both biophysical and cell biology methods. We used spectroscopy methods, including fluorescence, circular dichroism and dynamic light scattering to reveal the physicochemical properties. Using confocal and transmission electron microscopy we observed and tracked the internalization and intracellular trafficking. Additionally we tested the biological activity in vitro and the cellular toxicity of the delivery systems.

    We conclude that the transport vectors involved in this study are efficient at perturbing lipid membranes, which correlates with their remarkable capacity to transport oligonucleotides into cells. The improved and distinct capacities to escape from endosomal vesicles can be the result of their different structures and hydrophobicity. These findings extend the knowledge of the variables that condition intracellular Cell-penetrating peptide mediated transport of nucleic acids, which ultimately translates into a small step towards successful non-viral gene therapy.

  • 130.
    Figueroa, Ricardo A.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Gudise, Santhosh
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. Karolinska Institutet, Sweden.
    Hallberg, Einar
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Microtubule-associated nuclear envelope proteins in interphase and mitosis2011Inngår i: Biochemical Society Transactions, ISSN 0300-5127, E-ISSN 1470-8752, Vol. 39, s. 1786-1789Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The LINC (linker of nucleoskeleton and cytoskeleton) complex forms a transcisternal bridge across the NE (nuclear envelope) that connects the cytoskeleton with the nuclear interior. This enables some proteins of the NE to communicate with the centrosome and the microtubule cytoskeleton. The position of the centrosome relative to the NE is of vital importance for many cell functions, such as cell migration and division, and centrosomal dislocation is a frequent phenotype in laminopathic disorders. Also in mitosis, a small group of transmembrane NE proteins associate with microtubules when they concentrate in a specific membrane domain associated with the mitotic spindle. The present review discusses structural and functional aspects of microtubule association with NE proteins and how this association may be maintained over the cell cycle.

  • 131.
    Figueroa, Ricardo A.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för biokemi och biofysik.
    Ramberg, Veronica
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Gatsinzi, Tom
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Samuelsson, Malin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Zhang, Mu
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Iverfeldt, Kerstin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Hallberg, Einar
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Anchored FRET sensors detect local caspase activation prior to neuronal degeneration2011Inngår i: Molecular Neurodegeneration, ISSN 1750-1326, E-ISSN 1750-1326, Vol. 6, s. 35-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    BACKGROUND: Recent studies indicate local caspase activation in dendrites or axons during development and in neurodegenerative disorders such as Alzheimer's disease (AD). Emerging evidences point to soluble oligomeric amyloid-beta peptide as a causative agent in AD.

    RESULTS: Here we describe the design of fluorescence resonance energy transfer (FRET)-based caspase sensors, fused to the microtubule associated protein tau. Specific caspase sensors preferentially cleaved by caspase-3, -6 or -9 were expressed in differentiated human neuroblastoma SH-SY5Y cells. The anchoring of the sensors resulted in high FRET signals both in extended neurites and soma and made analysis of spatiotemporal signal propagation possible. Caspase activation was detected as loss of FRET after exposure to different stimuli. Interestingly, after staurosporine treatment caspase-6 activation was significantly delayed in neurites compared to cell bodies. In addition, we show that exposure to oligomer-enriched amyloid-beta peptide resulted in loss of FRET in cells expressing sensors for caspase-3 and -6, but not -9, in both soma and neurites before neurite degeneration was observed.

    CONCLUSIONS: Taken together, the results show that by using anchored FRET sensors it is possible to detect stimuli-dependent differential activation of caspases and to distinguish local from global caspase activation in live neuronal cells. Furthermore, in these cells oligomer-enriched amyloid-beta peptide induces a global, rather than local activation of caspase-3 and -6, which subsequently leads to neuronal cell death.

  • 132.
    Figueroa, Ricardo
    et al.
    Södertörn University, Sweden; Karolinska Institutet, Sweden.
    Gudise, Santhosh
    Södertörn University, Sweden; Karolinska Institutet, Sweden.
    Larsson, Veronica
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Hallberg, Einar
    Södertörn University, Sweden.
    A transmembrane inner nuclear membrane protein in the mitotic spindle2010Inngår i: Nucleus (Austin), ISSN 1949-1042, Vol. 1, nr 3, s. 249-253Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    We have recently characterized a novel transmembrane protein of the inner nuclear membrane of mammalian cells. The protein has two very interesting features. First, despite being an integral membrane protein it is able to concentrate in the membranes colocalizing with the mitotic spindle in metaphase and anaphase. Hence, the protein was named Samp1, Spindle associated membrane protein 1. Secondly, it displays a functional connection to centrosomes. This article discusses various aspects of Samp1 in relation to possible cellular function(s).

  • 133.
    Fisher, Linda
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Inflammatory cytokines and NFκB in Alzheimer’s disease2006Doktoravhandling, med artikler (Annet vitenskapelig)
    Abstract [en]

    Alzheimer’s disease is the most common form of dementia. It is a neurodegenerative disorder characterized by extracellular senile plaques and intracellular neurofibrillary tangles. The main constituent of the senile plaques is the neurotoxic β-amyloid peptide. Surrounding the senile plaques are activated astrocytes and microglia, believed to contribute to neurotoxicity through secretion of proinflammatory cytokines, like interleukin-1β and interleukin-6. For many inflammatory actions, including the cytokine induction in glial cells, the transcription factor NFκB plays a key role. This suggests that therapeutical strategies aimed to control the development of Alzheimer’s disease could include administration of drugs that hinder NFκB activation.

    The major aim of this thesis was to examine the effects of β-amyloid together with interleukin-1β on cytokine expression as well as NFκB activation in glial cells. The possibility to block NFκB activation, and downstream effects like interleukin-6 expression, by using an NFκB decoy was investigated. The possibility to improve the cellular uptake of the decoy by linking it to a cell-penetrating peptide was also investigated.

    The results obtained provide supportive evidence that inflammatory cytokines are induced by β-amyloid, and that they can indeed potentiate its effects. The results further demonstrate that by blocking NFκB activation, the induction of interleukin-6 expression can be inhibited. By using an improved cellular delivery system, the uptake of the NFκB decoy and hence the downstream cytokine inhibition could be increased. In conclusion, these results demonstrate the possibility to decrease the inflammatory reactions taken place in Alzheimer’s disease brains, which may ultimately lead to a possible way of controlling this disorder.

  • 134.
    Fisher, Linda
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Samuelsson, Malin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Jiang, Yang
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Ramberg, Veronica
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Figueroa, Ricardo
    Södertörn University College, Sweden.
    Hallberg, Einar
    Södertörn University College, Sweden.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Iverfeldt, Kerstin
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Targeting cytokine expression in glial cells by cellular delivery of an NFκB decoy2007Inngår i: Journal of Molecular Neuroscience, ISSN 0895-8696, E-ISSN 1559-1166, Vol. 31, nr 3, s. 209-219Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Inhibition of nuclear factor (NF)-κB has emerged as an important strategy for design of anti-inflammatory therapies. In neurodegenerative disorders like Alzheimer’s disease, inflammatory reactions mediated by glial cells are believed to promote disease progression. Here, we report that uptake of a double-stranded oligonucleotide NF-κB decoy in rat primary glial cells is clearly facilitated by noncovalent binding to a cell-penetrating peptide, transportan 10, via a complementary peptide nucleic acid (PNA) sequence. Fluorescently labeled oligonucleotide decoy was detected in the cells within 1 h only when cells were incubated with the decoy in the presence of cell-penetrating peptide. Cellular delivery of the decoy also inhibited effects induced by a neurotoxic fragment of the Alzheimer β amyloid peptide in the presence of the inflammatory cytokine interleukin (IL) 1β. Pretreatment of the cells with the complex formed by the decoy and the cell-penetrating peptide-PNA resulted in 80% and 50% inhibition of the NF-κB binding activity and IL-6 mRNA expression, respectively.

  • 135. Florén, Anders
    et al.
    Mäger, Imre
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. Tartu University, Estonia.
    Uptake kinetics of cell-penetrating peptides2011Inngår i: Cell-penetrating peptides: Methods and Protocols, Humana Press, 2011, s. 117-128Kapittel i bok, del av antologi (Fagfellevurdert)
    Abstract [en]

    As our knowledge increases about the diversity in uptake mechanisms displayed by cell-penetrating peptides (CPP), the concept of CPP uptake kinetics becomes increasingly complex. Here, we present three different assays that can be used for studying different kinetic aspects of CPP-mediated delivery: intracellular accumulation and membranolytical effects, intracellular CPP-cargo detachment, and finally a functional readout of a biological action from the delivered cargo. Unlike the traditional end-point measurements that give a static postincubation readout, these assays are all dynamic, real-time, in situ measurements obtained during incubation. A combination of some (or all) of these different assays gives us not only interesting kinetic information about the uptake routes but also provides a simple and valuable methodology for the evaluation of potential drug candidates based on the chemical modification of CPPs by cargo attachment.

  • 136.
    Folch, Jaume
    et al.
    Unitat de Bioquimica, Facultat de Medicina i Ciències de la Salut, Universitat Rovira i Virgili, Tarragona, Spain.
    Alvira, Daniel
    Unitat de Farmacologia i Farmacognòsia and Institut de Biomedicina (IBUB), Facultat de Farmàcia, Universitat de Barcelona, Nucli Universitari de Pedralbes, Barcelona, Spain.
    López-Querol, Marta
    Unitat de Farmacologia, Facultat de Medicina i Ciències de la Salut, Universitat Rovira i Virgili, Tarragona, Spain.
    Tajes, Marta
    Unitat de Farmacologia i Farmacognòsia and Institut de Biomedicina (IBUB), Facultat de Farmàcia, Universitat de Barcelona, Nucli Universitari de Pedralbes, Barcelona, Spain.
    Sureda, Francesc X
    Unitat de Farmacologia, Facultat de Medicina i Ciències de la Salut, Universitat Rovira i Virgili, Tarragona, Spain.
    Forsby, Anna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Rimbau, Víctor
    Unitat de Farmacologia i Farmacognòsia, Facultat de Farmàcia, Universitat de Barcelona, Nucli Universitari de Pedralbes, Barcelona, Spain.
    Camins, Antoni
    Unitat de Farmacologia i Farmacognòsia and Institut de Biomedicina (IBUB), Facultat de Farmàcia, Universitat de Barcelona, Nucli Universitari de Pedralbes, Barcelona, Spain.
    Pallàs, Mercè
    Unitat de Farmacologia i Farmacognòsia and Institut de Biomedicina (IBUB), Facultat de Farmàcia, Universitat de Barcelona, Nucli Universitari de Pedralbes, Barcelona, Spain.
    Evaluation of transcriptional activity of caspase-3 gene as a marker of acute neurotoxicity in rat cerebellar granular cells2010Inngår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 24, nr 2, s. 465-471Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Caspase-3 is a key protein involved in the classical apoptosis mechanism in neurons, as in many other cells types. In the present research, we describe the transcriptional activity of caspase-3 gene as a marker of acute toxicity in a primary culture model of rat cerebellar granule neurons (CGNs). CGNs were incubated for 16h in complete medium containing the chemicals at three concentrations (10, 100 μM and 1 mM). A total of 48 different compounds were tested. Gene transcriptional activity was determined by low-density array assays, and by single Taqman caspase-3 assays. Results from the PCR arrays showed that the caspase-3 gene was up-regulated when CGNs were exposed to neurotoxic chemicals. Significative correlations were found between the transcriptional activity of caspase-3 and the activity of some other genes related to apoptosis, cell-cycle and ROS detoxification. In our experiments, acute exposure of CGNs to well-documented pro-apoptotic xenobiotics modulated significantly caspase-3 gene expression, whereas chemicals not related to apoptosis did not modify caspase-3 gene expression. In conclusion, acute exposure of CGNs to neurotoxic compounds modulates the transcriptional activity of genes involved in the classical apoptotic pathway, oxidative stress and cell-cycle control. Transcriptional activity of caspase-3 correlates significantly with these changes and it could be a good indicator of acute neurotoxicity.

  • 137.
    Fors, Lisa
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för ekologi, miljö och botanik.
    Markus, Robert
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Theopold, Ulrich
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Hambäck, Peter A.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för ekologi, miljö och botanik.
    Differences in Cellular Immune Competence Explain Parasitoid Resistance for Two Coleopteran Species2014Inngår i: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 9, nr 9, artikkel-id e108795Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The immune defence of an organism is evolving continuously, causing counteradaptations in interacting species, which in turn affect other ecological and evolutionary processes. Until recently comparative studies of species interactions and immunity, combining information from both ecological and immunological fields, have been rare. The cellular immune defense in insects, mainly mediated by circulating hemocytes, has been studied primarily in Lepidoptera and Diptera, whereas corresponding information about coleopteran species is still scarce. In the study presented here, we used two closely related chrysomelids, Galerucella pusilla and G. calmariensis (Coleoptera), both attacked by the same parasitoid, Asecodes parviclava (Hymenoptera). In order to investigate the structure of the immune system in Galerucella and to detect possible differences between the two species, we combined ecological studies with controlled parasitism experiments, followed by an investigation of the cell composition in the larval hemolymph. We found a striking difference in parasitism rate between the species, as well as in the level of successful immune response (i.e. encapsulation and melanisation of parasitoid eggs), with G. pusilla showing a much more potent immune defense than G. calmariensis. These differences were linked to differences in the larval cell composition, where hemocyte subsets in both naive and parasitised individuals differed significantly between the species. In particular, the hemocytes shown to be active in the encapsulation process; phagocytes, lamellocytes and granulocytes, differ between the species, indicating that the cell composition reflects the ability to defend against the parasitoid.

  • 138.
    Forsby, A
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Bal-Price, A K
    Camins, A
    Coecke, S
    Fabre, N
    Gustafsson, H
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Honegger, P
    Kinsner-Ovaskainen, A
    Pallas, M
    Rimbau, V
    Rodríguez-Farré, E
    Suñol, C
    Vericat, J A
    Zurich, M G
    Neuronal in vitro models for the estimation of acute systemic toxicity.2009Inngår i: Toxicology in vitro : an international journal published in association with BIBRA, ISSN 1879-3177, Vol. 23, nr 8, s. 1564-1569Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The objective of the EU funded integrated project "ACuteTox" is to develop a strategy in which general cytotoxicity, together with organ-specific endpoints and biokinetic features, are taken into consideration in the in vitro prediction of oral acute systemic toxicity. With regard to the nervous system, the effects of 23 reference chemicals were tested with approximately 50 endpoints, using a neuronal cell line, primary neuronal cell cultures, brain slices and aggregated brain cell cultures. Comparison of the in vitro neurotoxicity data with general cytotoxicity data generated in a non-neuronal cell line and with in vivo data such as acute human lethal blood concentration, revealed that GABA(A) receptor function, acetylcholine esterase activity, cell membrane potential, glucose uptake, total RNA expression and altered gene expression of NF-H, GFAP, MBP, HSP32 and caspase-3 were the best endpoints to use for further testing with 36 additional chemicals. The results of the second analysis showed that no single neuronal endpoint could give a perfect improvement in the in vitro-in vivo correlation, indicating that several specific endpoints need to be analysed and combined with biokinetic data to obtain the best correlation with in vivo acute toxicity.

  • 139.
    Forsby, A
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Walum, E
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Polygodial induces inositol phosphate turnover in human neuroblastoma SH-SY5Y cells.1996Inngår i: Neuroscience Letters, ISSN 0304-3940, E-ISSN 1872-7972, Vol. 217, nr 1, s. 50-4Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The pungent sesquiterpenoid unsaturated dialdehydes polygodial and isovelleral, have previously been shown to increase the intracellular free calcium concentration [Ca2+]i in human neuroblastoma SH-SY5Y cells, partly by a release from intracellular Ca2+ stores, whereas the non-pungent compound epipolygodial, had no effect on the [Ca2+]i. In this study, we investigated the effect of isovelleral, polygodial and epipolygodial on inositol phosphate (IP) formation on the assumption that there might be a correlation between the release of intracellular Ca2+ and pungency of the compounds. It was found that polygodial induced IP mobilization in a concentration dependent way, whereas isovelleral had no effect on the IP formation in the SH-SY5Y cells. Phosphoinositide (PPI) turnover was activated by epipolygodial, but only at concentrations 40-fold higher than for polygodial, which emphasizes the importance of the correct stereometry in the dialdehyde configuration for the biological activity of polygodial. The polygodial-induced formation of IP1 was reduced by 71% under extracellular calcium-free conditions, which suggests feedback interactions between the IP formation and the increase in [Ca2+]i to account for a periodic activation of phospholipase C(PLC).

  • 140.
    Forsby, A
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Walum, E
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Sterner, O
    The effect of six sesquiterpenoid unsaturated dialdehydes on cell membrane permeability in human neuroblastoma SH-SY5Y cells.1992Inngår i: Chemico-Biological Interactions, ISSN 0009-2797, E-ISSN 1872-7786, Vol. 84, nr 1, s. 85-95Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The effect of six sesquiterpenes containing an unsaturated dialdehyde functionality, on cell membrane permeability in the human neuroblastoma cell line SH-SY5Y has been studied. The kinetics of the membrane leakage after addition of the sesquiterpenes were determined by measuring the efflux of radioactivity from cells preloaded with tritiated 2-deoxyglucose. The concentrations that gave 5% and 20% efflux of radioactivity as compared with control cells (EC5 and EC20) were determined for each compound. In spite of the structural similarities between the compounds, the effects on cell membrane permeability varied considerably. EC20 for polygodial, which is the most active compound, is 2.5 microM after 20-min incubation, but no leakage could be determined for merulidial even at concentrations as high as 4 mM. Rather, this compound seems to stabilize or fix the cell membrane and a lower efflux of radioactivity was observed as compared to the control cells. A quantitative structure-activity relationship analysis for the five active compounds showed a good correlation between the membrane leakage activity and certain chemical characteristics. Structural features strongly correlated with high activity were found to be: The geometry and the atomic charges of the unsaturated dialdehyde functionality, the dipole moment, the energy difference between the lowest unoccupied molecular orbital and the highest occupied molecular orbital and the lipophilicity.

  • 141.
    Forsby, A
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Witt, R
    Walum, E
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Sesquiterpenoid unsaturated dialdehydes increase the concentration of intracellular free Ca2+ in human neuroblastoma SH-SY5Y cells.1994Inngår i: Natural toxins (Print), ISSN 1056-9014, E-ISSN 1522-7189, Vol. 2, nr 2, s. 89-95Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The effect of the three sesquiterpenoid unsaturated dialdehydes--polygodial, isovelleral, and epipolygodial--on intracellular free Ca2+ concentration, [Ca2+]i, was investigated in the human neuroblastoma cell line SH-SY5Y. [Ca2+]i was measured by the Fura-2 spectrophotofluorometric technique in multi-cell and single-cell experiments. Polygodial and isovelleral induced an increase in [Ca2+]i at low concentrations (0.1 and 0.5 micrograms/ml, respectively) but epipolygodial affected [Ca2+]i only at a high concentration (10 micrograms/ml). The results indicated that there was a relationship between the effect on [Ca2+]i and the pungency of the sesquiterpenes tested. Experiments in a Ca(2+)-free extracellular solution showed that Ca2+ was also released from intracellular calcium stores. Images from single-cell experiments indicated that polygodial induced fluctuations in the [Ca2+]i in some cells. The mechanism behind the sesquiterpene induced increase of the [Ca2+]i was not identified but a possible mobilization of inositol phosphates is considered.

  • 142.
    Forsby, Anna
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Blaauboer, Bas
    Integration of in vitro neurotoxicity data with biokinetic modelling for the estimation of in vivo neurotoxicity.2007Inngår i: Human and Experimental Toxicology, ISSN 0960-3271, E-ISSN 1477-0903, Vol. 26, nr 4, s. 333-338Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Risk assessment of neurotoxicity is mainly based on in vivo exposure, followed by tests on behaviour, physiology and pathology. In this study, an attempt to estimate lowest observed neurotoxic doses after single or repeated dose exposure was performed. Differentiated human neuroblastoma SH-SY5Y cells were exposed to acrylamide, lindane, parathion, paraoxon, phenytoin, diazepam or caffeine for 72 hours. The effects on protein synthesis and intracellular free Ca2+ concentration were studied as physiological endpoints. Voltage operated Ca2+ channel function, acetylcholine receptor function and neurite degenerative effects were investigated as neurospecific endpoints for excitability, cholinergic signal transduction and axonopathy, respectively. The general cytotoxicity, determined as the total cellular protein levels after the 72 hours exposure period, was used for comparison to the specific endpoints and for estimation of acute lethality. The lowest concentration that induced 20% effect (EC20) obtained for each compound, was used as a surrogate for the lowest neurotoxic level (LOEL) at the target site in vivo. The LOELs were integrated with data on adsorption, distribution, metabolism and excretion of the compounds in physiologically-based biokinetic (PBBK) models of the rat and the lowest observed effective doses (LOEDs) were estimated for the test compounds. A good correlation was observed between the estimated LOEDs and experimental LOEDs found in literature for rat for all test compounds, except for diazepam. However, when using in vitro data from the literature on diazepam's effect on gamma-amino butyric acid (GABA)A receptor function for the estimation of LOED, the correlation between the estimated and experimental LOEDs was improved from a 10,000-fold to a 10-fold difference. Our results indicate that it is possible to estimate LOEDs by integrating in vitro toxicity data as surrogates for lowest observed target tissue levels with PBBK models, provided that some knowledge about toxic mechanisms is known.

  • 143.
    Forsby, Anna
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Norman, Kimberly
    EL Andaloussi-Lilja, Johanna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Lundqvist, Jessica
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Wojcik, Beata
    Walczak, Vincent
    Curren, Rodger
    Martin, Katharine
    Tierney, Neena
    Predicting eye stinging potential of baby shampoos by assessing TRPV1 channel activity2012Inngår i: Toxicology Letters, ISSN 0378-4274, E-ISSN 1879-3169, Vol. 211, s. S113-S113Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The Transient Receptor Potential Vanilloid type 1 (TRPV1) receptor is one of the most well characterized pain-inducing receptors. The purpose of this study was to predict human eye stinging of 19 baby bath and shampoo formulations by studying TRPV1 activity. The NociOcular test, a novel recombinant neuronal in vitro model with high expression of functional TRPV1 channels was used to test shampoo formulations containing surfactants, preservatives, and fragrances (sodium laureth sulfate, cocoamidopropylbetaine, cocoglucoside, sodium benzoate, quaternium-15, etc.). The increase in intracellular free Ca2+ was analysed by fluorescence during exposure. TRPV1-specific Ca2+ influx was abolished when the TRPV1 channel antagonist capsazepine was applied to the cells prior to shampoo samples. The positive control, i.e. adult shampoo, was the most active sample tested in the NociOcular test and also induced the worst stinging sensation. The negative control, i.e. marketed baby shampoo, was negative in both tests. Seven of the formulations induced stinging in the human test, and of those six were positive in the NociOcular test. Twelve of the formulations were classified as non-stinging in the human test, and of those 10 were negative in the NociOcular test. None of the established in vitro tests for eye irritation were able to correctly predict the human stinging sensation of the baby products. Our data support that the TRPV1 channel is a principle mediator of eye stinging sensation induced by baby bath and shampoo formulations and that the NociOcular test may be a valuable in vitro tool to predict human eye stinging sensation.

  • 144.
    Forsby, Anna
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Norman, Kimberly G.
    EL Andaloussi-Lilja, Johanna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Lundqvist, Jessica
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Walczak, Vincent
    Curren, Rodger
    Martin, Katharine
    Tierney, Neena K.
    Using Novel In Vitro NociOcular Assay Based on TRPV1 Channel Activation for Prediction of Eye Sting Potential of Baby Shampoos2012Inngår i: Toxicological Sciences, ISSN 1096-6080, E-ISSN 1096-0929, Vol. 129, nr 2, s. 325-331Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The transient receptor potential vanilloid type 1 (TRPV1) channel is one of the most well-characterized pain-inducing receptors. The purpose of this study was to predict human eye stinging of 19 baby bath and shampoo formulations by studying TRPV1 activity, as measured by increase in intracellular free Ca2+. The NociOcular test, a novel recombinant neuronal in vitro model with high expression of functional TRPV1 channels, was used to test formulations containing a variety of surfactants, preservatives, and fragrances. TRPV1-specific Ca2+ influx was abolished when the TRPV1 channel antagonist capsazepine was applied to the cells prior to shampoo samples. The positive control, an adult shampoo that contains cocamide monoethanolamine (CMEA), a known stinging ingredient, was the most active sample tested in the NociOcular test. The negative control, a marketed baby shampoo, was negative in the NociOcular and human tests. Seven of the formulations induced stinging in the human test, and of those six were positive in the NociOcular test. Twelve formulations were classified as nonstinging in the human test, and of those ten were negative in the NociOcular test. There was no correlation between the clinical stinging results for the baby formulations and the data generated from other in vitro eye irritation assays (cytosensor microphysiometer, neutral red uptake, EpiOcular, transepithelial permeability). Our data support that the TRPV1 channel is a principal mediator of eye-stinging sensation induced by baby bath and shampoo formulations and that the NociOcular test may be a valuable in vitro tool to predict human eye stinging sensation.

  • 145. Fossat, P.
    et al.
    Dobremez, E.
    Bouali-Benazzouz, R.
    Favereaux, A.
    Bertrand, S. S.
    Kilk, Kalle
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. University of Tartu, Estonia.
    Leger, C.
    Cazalets, J-R
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. University of Tartu, Estonia.
    Landry, M.
    Nagy, F.
    Knockdown of L calcium channel subtypes: differential effects in neuropathic pain2010Inngår i: Journal of Neuroscience, ISSN 0270-6474, E-ISSN 1529-2401, Vol. 30, nr 3, s. 1073-1085Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    The maintenance of chronic pain states requires the regulation of gene expression, which relies on an influx of calcium. Calcium influx through neuronal L-type voltage-gated calcium channels (LTCs) plays a pivotal role in excitation-transcription coupling, but the involvement of LTCs in chronic pain remains unclear. We used a peptide nucleic acid (transportan 10-PNA conjugates)-based antisense strategy to investigate the role of the LTC subtypes Ca(V)1.2 and Ca(V)1.3 in long-term pain sensitization in a rat model of neuropathy (spinal nerve ligation). Our results demonstrate that specific knockdown of Ca(V)1.2 in the spinal dorsal horn reversed the neuropathy-associated mechanical hypersensitivity and the hyperexcitability and increased responsiveness of dorsal horn neurons. Intrathecal application of anti-Ca(V)1.2 siRNAs confirmed the preceding results. We also demonstrated an upregulation of Ca(V)1.2 mRNA and protein in neuropathic animals concomitant to specific Ca(V)1.2-dependent phosphorylation of the cAMP response element (CRE)-binding protein (CREB) transcription factor. Moreover, spinal nerve ligation animals showed enhanced transcription of the CREB/CRE-dependent gene COX-2 (cyclooxygenase 2), which also depends strictly on Ca(V)1.2 activation. We propose that L-type calcium channels in the spinal dorsal horn play an important role in pain processing, and that the maintenance of chronic neuropathic pain depends specifically on channels comprising Ca(V)1.2.

  • 146. Freimann, Krista
    et al.
    Arukuusk, Piret
    Kurrikoff, Kaido
    Pärnaste, Ly
    Raid, Raivo
    Piirsoo, Andres
    Pooga, Margus
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. University of Tartu, Estonia.
    Formulation of Stable and Homogeneous Cell-Penetrating Peptide NF55 Nanoparticles for Efficient Gene Delivery In Vivo2018Inngår i: Molecular Therapy - Nucleic Acids, ISSN 2162-2531, E-ISSN 2162-2531, Vol. 10, s. 28-35Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Although advances in genomics and experimental gene therapy have opened new possibilities for treating otherwise incurable diseases, the transduction of nucleic acids into the cells and delivery in vivo remain challenging. The high molecular weight and anionic nature of nucleic acids require their packing into nanoparticles for the delivery. The efficacy of nanoparticle drugs necessitates the high bioactivity of constituents, but their distribution in organisms is mostly governed by the physical properties of nanoparticles, and therefore, generation of stable particles with strictly defined characteristics is highly essential. Using previously designed efficient cell-penetrating peptide NF55, we searched for strategies enabling control over the nanoparticle formation and properties to further improve transfection efficacy. The size of the NF55/pDNA nanoparticles correlates with the concentration of its constituents at the beginning of assembly, but characteristics of nanoparticles measured by DLS do not reliably predict the applicability of particles in in vivo studies. We introduce a new formulation approach called cryo-concentration, where we acquired stable and homogeneous nanoparticles for administration in vivo. The cryo-concentrated NF55/pDNA nanoparticles exhibit several advantages over standard formulation: They have long shelf-life and do not aggregate after reconstitution, have excellent stability against enzymatic degradation, and show significantly higher bioactivity in vivo.

  • 147. Freimann, Krista
    et al.
    Arukuusk, Piret
    Kurrikoff, Kaido
    Vasconcelos, Luís Daniel Ferreira
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Veiman, Kadi-Liis
    Uusna, Julia
    Margus, Helerin
    Garcia-Sosa, Alfonso T.
    Pooga, Margus
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. University of Tartu, Estonia.
    Optimization of in vivo DNA delivery with NickFect peptide vectors2016Inngår i: Journal of Controlled Release, ISSN 0168-3659, E-ISSN 1873-4995, Vol. 241, s. 135-143Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    As the field of gene therapy progresses, an increasingly urgent need has arisen for efficient and non-toxic vectors for the in vivo delivery of nucleic acids. Cell-penetrating peptides (CPP) are very efficient transfection reagents in vitro, however, their application in vivo needs improvement. To enhance in vivo transfection we designed various CPPs based on previous knowledge of internalization studies and physiochemical properties of NickFect (NF) nanoparticles. We show that increment of the helicity of these Transportan10 analogues improves the transfection efficiency. We rationally design by modifying the net charge and the helicity of the CPP a novel amphipathic α-helical peptide NF55 for in vivo application. NF55 condenses DNA into stable nanoparticles that are resistant to protease degradation, promotes endosomal escape, and transfects the majority of cells in a large cell population. We demonstrate that NF55 mediates DNA delivery in vivo with gene induction efficiency that is comparable to commercial transfection reagents. In addition to gene induction in healthy mice, NF55/DNA nanoparticles showed promising tumor transfection in various mouse tumor models, including an intracranial glioblastoma model. The efficiency of NF55 to convey DNA specifically into tumor tissue increased even further after coupling a PEG2000 to the peptide via a disulphide-bond. Furthermore, a solid formulation of NF55/DNA displayed an excellent stability profile without additives or special storage conditions. Together, its high transfection efficacy and stability profile make NF55 an excellent vector for the delivery of DNA in vivo.

  • 148. Freimann, Krista
    et al.
    Kurrikoff, Kaido
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. University of Tartu, Estonia.
    Galanin receptors as a potential target for neurological disease2015Inngår i: Expert opinion on therapeutic targets, ISSN 1472-8222, E-ISSN 1744-7631, Vol. 19, nr 12, s. 1665-1676Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    INTRODUCTION: Galanin is a 29/30 amino acid long neuropeptide that is widely expressed in the brains of many mammals. Galanin exerts its biological activities through three different G protein-coupled receptors, GalR1, GalR2 and GalR3. The widespread distribution of galanin and its receptors in the CNS and the various physiological and pharmacological effects of galanin make the galanin receptors attractive drug targets.

    AREAS COVERED: This review provides an overview of the role of galanin and its receptors in the CNS, the involvement of the galaninergic system in various neurological diseases and the development of new galanin receptor-specific ligands.

    EXPERT OPINION: Recent advances and novel approaches in migrating the directions of subtype-selective ligand development and chemical modifications of the peptide backbone highlight the importance of the galanin neurochemical system as a potential target for drug development.

  • 149. Fuxe, K.
    et al.
    Canals, M.
    Torvinen, M.
    Marcellino, D.
    Terasmaa, A.
    Genedani, S.
    Leo, G.
    Guidolin, D.
    Diaz-Cabiale, Z.
    Rivera, A.
    Lundström, L.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Narvaez, J.
    Tanganelli, S.
    Lluis, C.
    Ferré, S.
    Woods, A.
    Franco, R.
    Agnati, L. F.
    Intramembrane receptor-receptor interactions: a novel principle in molecular medicine2007Inngår i: Journal of neural transmission, ISSN 0300-9564, E-ISSN 1435-1463, Vol. 114, nr 1, s. 49-75Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    In 1980/81 Agnati and Fuxe introduced the concept of intramembrane receptor-receptor interactions and presented the first experimental observations for their existence in crude membrane preparations. The second step was their introduction of the receptor mosaic hypothesis of the engram in 1982. The third step was their proposal that the existence of intramembrane receptor-receptor interactions made possible the integration of synaptic (WT) and extrasynaptic (VT) signals. With the discovery of the intramembrane receptor-receptor interactions with the likely formation of receptor aggregates of multiple receptors, so called receptor mosaics, the entire decoding process becomes a branched process already at the receptor level in the surface membrane. Recent developments indicate the relevance of cooperativity in intramembrane receptor-receptor interactions namely the presence of regulated cooperativity via receptor-receptor interactions in receptor mosaics (RM) built up of the same type of receptor (homo-oligomers) or of subtypes of the same receptor (RM type1). The receptor-receptor interactions will to a large extent determine the various conformational states of the receptors and their operation will be dependent on the receptor composition (stoichiometry), the spatial organization (topography) and order of receptor activation in the RM. The biochemical and functional integrative implications of the receptor-receptor interactions are outlined and long-lived heteromeric receptor complexes with frozen RM in various nerve cell systems may play an essential role in learning, memory and retrieval processes. Intramembrane receptor-receptor interactions in the brain have given rise to novel strategies for treatment of Parkinson's disease (A2A and mGluR5 receptor antagonists), schizophrenia (A2A and mGluR5 agonists) and depression (galanin receptor antagonists). The A2A/D2, A2A/D3 and A2A/mGluR5 heteromers and heteromeric complexes with their possible participation in different types of RM are described in detail, especially in the cortico-striatal glutamate synapse and its extrasynaptic components, together with a postulated existence of A2A/D4 heteromers. Finally, the impact of intramembrane receptor-receptor interactions in molecular medicine is discussed outside the brain with focus on the endocrine, the cardiovascular and the immune systems.

  • 150. Fuxe, K.
    et al.
    Marcellino, D.
    Rivera, A.
    Diaz-Cabiale, Z.
    Filip, M.
    Gago, B.
    Roberts, D. C. S.
    Langel, Ülo
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Genedani, S.
    Ferraro, L.
    de la Calle, A.
    Narvaez, J.
    Tanganelli, S.
    Woods, A.
    Agnati, L. F.
    Receptor-receptor interactions within receptor mosaics: Impact on neuropsychopharmacology2008Inngår i: Brain Research Reviews, ISSN 0165-0173, E-ISSN 1872-6321, Vol. 58, nr 2, s. 415-452Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Future therapies for diseases associated with altered dopaminergic signaling, including Parkinson's disease, schizophrenia and drug addiction or drug dependence may substantially build on the existence of intramembrane receptor-receptor interactions within dopamine receptor containing receptor mosaics (RM; dimeric or high-order receptor oligomers) where it is believed that the dopamine D-2 receptor may operate as the 'hub receptor' within these complexes. The constitutive adenosine A(2A)/dopamine D-2 RM, located in the dorsal striatopallidal GABA neurons, are of particular interest in view of the demonstrated antagonistic A(2A)/D-2 interaction within these heteromers; an interaction that led to the suggestion and later demonstration that A(2A) antagonists could be used as novel anti-Parkinsonian drugs. Based on the likely existence of A(2A)/D-2/mGluR5 RM located both extrasynaptically on striato-pallidal GABA neurons and on cortico-striatal glutamate terminals, multiple receptor-receptor interactions within this RM involving synergism between A(2A)/mGluR5 to counteract D-2 signaling, has led to the proposal of using combined mGluR5 and A(2A) antagonists as a future anti-Parkinsonian treatment. Based on the same RM in the ventral striato-pallidal GABA pathways, novel strategies for the treatment of schizophrenia, building on the idea that A(2A) agonists and/or mGluR5 agonists will help reduce the increased dopaminergic signaling associated with this disease, have been suggested. Such treatment may ensure the proper glutamatergic drive from the mediodorsal thalamic nucleus to the prefrontal cortex, one which is believed to be reduced in schizophrenia due to a dominance of D-2-like signaling in the ventral striatum. Recently, A(2A) receptors also have been shown to counteract the locomotor and sensitizing actions of cocaine and increases in A(2A) receptors have also been observed in the nucleus accumbens after extended cocaine self-administration, probably representing a compensatory up-regulation to counteract the cocaine-induced increases in dopamine D-2 and D-3 signaling. Therefore, A(2A) agonists, through antagonizing D-2 and D-3 signaling within A(2A)/D-2 and A(2)/D-3 RM heteromers in the nucleus accumbens, may be found useful as a treatment for cocaine dependence. Furthermore, antagonistic cannabinoid CB1/D-2 interactions requiring A(2A) receptors have also been discovered and possibly operate in CB1/D-2/A(2A) RM located principally on striatal glutamate terminals but also on some ventral striato-pallidal GABA neurons, thereby opening up a new mechanism for the integration of endocannabinoid, DA and adenosine mediated signals. Thus, A(2A), mGluR5 and/or CB1 receptors can form integrative units with D-2 receptors within RM displaying different compositions, topography and localization. Also galaninR/5-HT1A RM probably participates in the transmission of the ascending 5-hydroxytryptamine neurons, where galanin receptors antagonize 5-HT1A recognition and signaling. Subtype specific galanin receptor antagonists may therefore represent novel antidepressant drugs. These results suggest the importance of a complete understanding of the function of these RM with regard to disease. Ultimately receptor-recepor interactions within RM that modify dopaminergic and serotonergic signaling may give new strategies for treatment of a wide range of diseases associated with altered dopaminergic and serotonergic signaling.

1234567 101 - 150 of 535
RefereraExporteraLink til resultatlisten
Permanent link
Referera
Referensformat
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Annet format
Fler format
Språk
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Annet språk
Fler språk
Utmatningsformat
  • html
  • text
  • asciidoc
  • rtf