In the present study,
we investigated lipid membrane interactions
of silica nanoparticles as carriers for the antimicrobial peptide
LL-37 (LLGDFFRKSKEKIGKEFKRIVQRIKDFLRNLVPRTES).
In doing so, smooth mesoporous nanoparticles were compared to virus-like
mesoporous nanoparticles, characterized by a “spiky”
external surface, as well as to nonporous silica nanoparticles. For
this, we employed a combination of neutron reflectometry, ellipsometry,
dynamic light scattering, and ζ-potential measurements for studies
of bacteria-mimicking bilayers formed by palmitoyloleoylphosphatidylcholine/palmitoyloleoylphosphatidylglycerol.
The results show that nanoparticle topography strongly influences
membrane binding and destabilization. We found that virus-like particles
are able to destabilize such lipid membranes, whereas the corresponding
smooth silica nanoparticles are not. This effect of particle spikes
becomes further accentuated after loading of such particles with LL-37.
Thus, peptide-loaded virus-like nanoparticles displayed more pronounced
membrane disruption than either peptide-loaded smooth nanoparticles
or free LL-37. The structural basis of this was clarified by neutron
reflectometry, demonstrating that the virus-like nanoparticles induce
trans-membrane defects and promote incorporation of LL-37 throughout
both bilayer leaflets. The relevance of such effects of particle spikes
for bacterial membrane rupture was further demonstrated by confocal
microscopy and live/dead assays on Escherichia coli bacteria. Taken together, these findings demonstrate that topography
influences the interaction of nanoparticles with bacteria-mimicking
lipid bilayers, both in the absence and presence of antimicrobial
peptides, as well as with bacteria. The results also identify virus-like
mesoporous nanoparticles as being of interest in the design of nanoparticles
as delivery systems for antimicrobial peptides.
High and low density lipoproteins (HDL and LDL) are thought to play vital roles in the onset and development of atherosclerosis; the biggest killer in the western world. Key issues of initial lipoprotein (LP) interactions at cellular membranes need to be addressed including LP deposition and lipid exchange. Here we present a protocol for monitoring the in situ kinetics of lipoprotein deposition and lipid exchange/removal at model cellular membranes using the non-invasive, surface sensitive methods of neutron reflection and quartz crystal microbalance with dissipation. For neutron reflection, lipid exchange and lipid removal can be distinguished thanks to the combined use of hydrogenated and tail-deuterated lipids. Both HDL and LDL remove lipids from the bilayer and deposit hydrogenated material into the lipid bilayer, however, the extent of removal and exchange depends on LP type. These results support the notion of HDL acting as the ‘good’ cholesterol, removing lipid material from lipid-loaded cells, whereas LDL acts as the ‘bad’ cholesterol, depositing lipid material into the vascular wall.
Herein,
we report on the formation of cross-linked antimicrobial
peptide-loaded microgel multilayers. Poly(ethyl acrylate-co-methacrylic acid) microgels were synthesized and functionalized
with biotin to enable the formation of microgel multilayers cross-linked
with avidin. Microgel functionalization and avidin cross-linking were
verified with infrared spectroscopy, dynamic light scattering, and
z-potential measurements, while multilayer formation (up to four layers)
was studied with null ellipsometry and quartz crystal microbalance
with dissipation (QCM-D). Incorporation of the antimicrobial peptide
KYE28 (KYEITTIHNLFRKLTHRLFRRNFGYTLR) into the microgel
multilayers was achieved either in one shot after multilayer formation
or through addition after each microgel layer deposition. The latter
was found to strongly promote peptide incorporation. Further, antimicrobial
properties of the peptide-loaded microgel multilayers against Escherichia coli were investigated and compared to those
of a peptide-loaded microgel monolayer. Results showed a more pronounced
suppression in bacterial viability in suspension for the microgel
multilayers. Correspondingly, LIVE/DEAD staining showed promoted disruption
of adhered bacteria for the KYE28-loaded multilayers. Taken together,
cross-linked microgel multilayers thus show promise as high load surface
coatings for antimicrobial peptides.
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