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Characterization of Differential Toll-like Receptor Responses below the Optical Diffraction Limit

Aaron, Jesse S. ; Carson, Bryan D. ; Timlin, Jerilyn A.

Small (Weinheim an der Bergstrasse, Germany), 2012-10, Vol.8 (19), p.3041-3049 [Periódico revisado por pares]

Weinheim: WILEY-VCH Verlag

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  • Título:
    Characterization of Differential Toll-like Receptor Responses below the Optical Diffraction Limit
  • Autor: Aaron, Jesse S. ; Carson, Bryan D. ; Timlin, Jerilyn A.
  • Assuntos: Animals ; Cell Line ; Cell Membrane - metabolism ; clusters ; Escherichia coli - metabolism ; imaging ; lipopolysaccharides ; Lipopolysaccharides - chemistry ; Lipopolysaccharides - metabolism ; Macrophages - metabolism ; Mice ; Microscopy - methods ; Optical Imaging ; receptors ; stochastic optical reconstruction microscopy ; Toll-Like Receptor 4 - chemistry ; Toll-Like Receptor 4 - metabolism ; Yersinia pestis - metabolism
  • É parte de: Small (Weinheim an der Bergstrasse, Germany), 2012-10, Vol.8 (19), p.3041-3049
  • Notas: istex:57D1E3EEC1DD12064DA1C75A3B9236C48B16B2B9
    ark:/67375/WNG-59QVS0K1-X
    ArticleID:SMLL201200106
    ObjectType-Article-1
    SourceType-Scholarly Journals-1
    ObjectType-Feature-2
    content type line 23
    AC04-94AL85000
    SAND2012-0259J
    USDOE National Nuclear Security Administration (NNSA)
  • Descrição: Many membrane receptors are recruited to specific cell surface domains to form nanoscale clusters upon ligand activation. This step appears to be necessary to initiate cell signaling, including pathways in innate immune system activation. However, virulent pathogens such as Yersinia pestis (the causative agent of plague) are known to evade innate immune detection, in contrast to similar microbes (such as Escherichia coli) that elicit a robust response. This disparity has been partly attributed to the structure of lipopolysaccharides (LPS) on the bacterial cell wall, which are recognized by the innate immune receptor TLR4. It is hypothesized that nanoscale differences exist between the spatial clustering of TLR4 upon binding of LPS derived from Y. pestis and E. coli. Although optical imaging can provide exquisite details of the spatial organization of biomolecules, there is a mismatch between the scale at which receptor clustering occurs (<300 nm) and the optical diffraction limit (>400 nm). The last decade has seen the emergence of super‐resolution imaging methods that effectively break the optical diffraction barrier to yield truly nanoscale information in intact biological samples. This study reports the first visualizations of TLR4 distributions on intact cells at image resolutions of <30 nm using a novel, dual‐color stochastic optical reconstruction microscopy (STORM) technique. This methodology permits distinction between receptors containing bound LPS from those without at the nanoscale. Importantly, it is also shown that LPS derived from immunostimulatory bacteria result in significantly higher LPS–TLR4 cluster sizes and a nearly twofold greater ligand/receptor colocalization as compared to immunoevading LPS. A dual‐color stochastic optical reconstruction microscopy technique is employed to gain insight into the nanoscale organization of the innate immune system receptor TLR4. Data indicate significant changes in TLR4 clustering behavior within the cell membrane in response to immunostimulatory and immunoevading bacterial antigens, thereby shedding light on virulence mechanisms of highly pathogenic microbes such as Yersinia pestis.
  • Editor: Weinheim: WILEY-VCH Verlag
  • Idioma: Inglês
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  • Realização: ABCD-USP USP   Logos de Redes Sociais: Freepik

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