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- Sigurdardottir, Thorgerdur
(författare)
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Actions of Antimicrobial Peptides and Bacterial Components in Inflammation
- 2009
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Antimicrobial peptides are evolutionally ancient parts of the innate immune system and their primary role is to protect us from infections. The human cathelicidin-derived antimicrobial peptide, LL-37, not only possesses broad spectrum antimicrobial activities but is also able to bind and neutralize bacterial lipopolysaccharide (LPS), an important trigger of the widespread inflammatory response contributing to septic shock. LL-37 has been studied as an alternative to conventional antibiotics but clinical trials have been hampered by indications of its toxic effect on mammalian cells and evidence that its antimicrobial effects are inhibited by serum. It has been proposed that the cytotoxicity of LL-37 is due to hydrophobic amino acids. We were able, by removing hydrophobic amino acids from the N-terminal of LL-37, to generate less cytotoxic peptides with retained antimicrobial and LPS–neutralizing actions in serum. By using computer simulation we identified an active domain of LL-37, a 21 amino acid fragment, GKE, displaying similar antimicrobial and LPS-binding activity in vitro as native LL-37 but less toxic and therefore holding promise as a template for the development of peptide antibiotics for treating sepsis. Sepsis due to Gram-positive bacteria is becoming increasingly prevalent. The Gram-positive bacterium Streptococcus pyogenes, carrying a surface protein named M1 being fundamental for its virulence, is the major cause of severe streptococcal infections like streptococcal toxic shock syndrome and necrotizing fasciitis. We found that M1 protein is able to induce vascular nitric oxide (NO) production, which in turn relaxes smooth muscle cells and could thereby contribute to the severe hypotension seen in septic patients. This we confirmed by showing that M1 protein caused hyporesponsiveness to the vasoconstrictor, phenylephrine, in rat aorta. Bacterial compounds are able to activate Toll-like receptors generating an inflammatory response. Experiments using wild type and knockout mice revealed that M1 protein is able to attach to both TLR2 and TLR4 (TLR) in mice, only activating the latter. M1 protein only attached to TLR2 in human blood vessels. LL-37 possesses immunomodulatory effects. In order to explore potential modulatory effects of GKE on vascular nitric oxide production we used the well identified proinflammatory compounds interleukin-1β (IL-1β), M1 protein from Streptococcus pyogenes and lipoteichoic acid (LTA). All three induced vascular NO production in rat aorta. GKE at low concentration inhibited IL-1β-induced NO production, but synergistically increased it at higher concentrations. GKE did not affect the M1 protein induced NO production while GKE inhibited LTA induced NO production. Thus, GKE seems to have complex modulatory effects on vascular nitric oxide production depending on the inflammatory compound used.
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- Sigurdardottir, Thorgerdur, et al.
(författare)
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In Silico Identification and Biological Evaluation of Antimicrobial Peptides Based on Human Cathelicidin LL-37
- 2006
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Ingår i: Antimicrobial Agents and Chemotherapy. - 0066-4804 .- 1098-6596. ; 50:9, s. 2983-2989
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Tidskriftsartikel (refereegranskat)abstract
- Bacterial lipopolysaccharides (LPS) are important triggers of the widespread inflammatory response, which contributes to the development of multiple organ failure during sepsis. The helical 37-amino-acid-long human antimicrobial peptide LL-37 not only possesses a broad-spectrum antimicrobial activity but also binds and neutralizes LPS. However, the use of LL-37 in sepsis treatment is hampered by the fact that it is also cytotoxic. To find a less toxic analog of LL-37, we used in silico analysis to identify amphipathic helical regions of LL-37. A 21-amino-acid fragment (GKE) was synthesized, the biological actions of which were compared to those of two equally long peptides derived from the N and C termini of LL-37 as well as native LL-37. GKE displayed antimicrobial activity against Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Candida albicans, and Candida parapsilosis that was similar to or even stronger than LL-37. GKE, as well as the equally long control peptides, attracted granulocytes in a fashion similar to that of LL-37, while only GKE was as potent as LL-37 in inhibiting LPS-induced vascular nitric oxide production. GKE caused less hemolysis and apoptosis in human cultured smooth muscle cells than LL-37. In summary, we have identified an active domain of LL-37, GKE, which displays antimicrobial activity in vitro and LPS-binding activity similar to those of LL-37 but is less toxic. GKE therefore holds promise as a template for the development of peptide antibiotics for the treatment of sepsis.
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- Sigurdardottir, Thorgerdur, et al.
(författare)
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M1 protein from Streptococcus pyogenes induces nitric oxidemediated vascular hyporesponsiveness to phenylephrine: Involvement of toll-like receptor activation.
- 2010
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Ingår i: Shock. - 1540-0514. ; 34:1, s. 98-104
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Tidskriftsartikel (refereegranskat)abstract
- Streptococcus pyogenes carrying M1 protein causes the severe and increasingly prevalent streptococcal toxic shock syndrome and necrotizing fasciitis. M1 protein is an important virulence factor of Streptococcus pyogenes and induces an inflammatory response in human monocytes. We wanted to investigate if purified M1 protein in solution could induce vascular nitric oxide (NO) production leading to vasopressor hyporesponsiveness. Rat aorta segments were incubated with M1 protein or lipopolysaccharide (LPS) in vitro. M1 protein (10 mug ml) and LPS (1 ng ml) to a similar extent induced NO production and hyporesponsiveness to the vasoconstrictor phenylephrine. Immuno-gold electron microscopy demonstrated that M1 protein binds to toll-like receptor (TLR) 2 as well as TLR4 in mouse aorta but only to TLR2 in human omental artery. Incubation with M1 protein caused a reduction in the contractile response to phenylephrine in aorta segments from wild type and TLR2 knockout but not from TLR4 knockout mice. In conclusion, M1 protein causes vascular NO production leading to hyporesponsiveness to vasopressors via a mechanism involving TLR but the subtypes may be species-dependent. M1 protein could contribute to the circulatory disturbances accompanying severe invasive streptococcal infections.
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