14 research outputs found
Phospholipase A<sub>2</sub>‑Induced Degradation and Release from Lipid-Containing Polymersomes
Hybrid
vesicles, comprising blends of amphiphilic block copolymers
and phospholipids, have attracted significant attention recently because
of their unique combination of chemical and physical properties. We
report a method to make unilamellar hybrid vesicles with diameters
of 100 nm by mixing polybutadiene-<i>block</i>-poly(ethylene
oxide) and phosphocholine lipids using a combination of solvent inversion
and sonication. We show that homogeneous hybrid vesicles are formed
when one component is a minor fraction. At compositions with balanced
mass fractions, separate populations of similarly sized pure liposomes
and hybrid vesicles are indicated. We investigate the release kinetics
of calcein encapsulated in the lumen as hybrid large and giant unilamellar
vesicles (LUVs and GUVs) of different compositions are exposed to
phospholipase A<sub>2</sub> (PLA<sub>2</sub>). PLA<sub>2</sub> hydrolyzes
lipids, which leads to dissolution of lipid domains and provides a
trigger for the release of calcein as pores are formed. We demonstrate
that depending on the polymer mole fraction, block copolymers can
either protect or boost the rate of lipid degradation and thereby
the release rate from nanoscale hybrid vesicles. Strong indications
of lipid phase separation into nanoscale domains in LUVs are observed.
Most importantly, hybrid GUV with lipids in the fluid phase release
calcein slowly as lipids in the liquid-disordered phase do not phase-separate,
but they show the fastest release of all blends as LUVs. This indicates
phase separation on the nanoscale in contrast to on the microscale,
but it also indicates retained high mobility of lipids between the
nanoscale domains, which is absent for lipids in the gel phase. Our
results demonstrate several ways in which nanoscale hybrid vesicles
can and should be optimized for PLA<sub>2</sub>-triggered release
of water-soluble compounds
Stealth Nanoparticles Grafted with Dense Polymer Brushes Display Adsorption of Serum Protein Investigated by Isothermal Titration Calorimetry
Core–shell
nanoparticles receive much attention for their
current and potential applications in life sciences. Commonly, a dense
shell of hydrated polymer, a polymer brush, is grafted to improve
colloidal stability of functional nanoparticles and to prevent protein
adsorption, aggregation, cell recognition, and uptake. Until recently,
it was widely assumed that a polymer brush shell indeed prevents strong
association of proteins and that this leads to their superior “stealth”
properties in vitro and in vivo. We show using <i>T</i>-dependent
isothermal titration calorimetry on well-characterized monodisperse
superparamagnetic iron oxide nanoparticles with controlled dense stealth
polymer brush shells that “stealth” core–shell
nanoparticles display significant attractive exothermic and enthalpic
interactions with serum proteins, despite having excellent colloidal
stability and negligible nonspecific cell uptake. This observation
is at room temperature shown to depend only weakly on variation of
iron oxide core diameter and type of grafted stealth polymer: poly(ethylene
glycol), poly(ethyl oxazoline), poly(isopropyl oxazoline), and poly(<i>N</i>-isopropyl acrylamide). Polymer brush shells with a critical
solution temperature close to body temperature showed a strong temperature
dependence in their interactions with proteins with a significant
increase in protein binding energy with increased temperature. The
stoichiometry of interaction is estimated to be near 1:1 for PEGylated
nanoparticles and up to 10:1 for larger thermoresponsive nanoparticles,
whereas the average free energy of interaction is enthalpically driven
and comparable to a weak hydrogen bond
Complete Exchange of the Hydrophobic Dispersant Shell on Monodisperse Superparamagnetic Iron Oxide Nanoparticles
High-temperature synthesized monodisperse
superparamagnetic iron
oxide nanoparticles are obtained with a strongly bound ligand shell
of oleic acid and its decomposition products. Most applications require
a stable presentation of a defined surface chemistry; therefore, the
native shell has to be completely exchanged for dispersants with irreversible
affinity to the nanoparticle surface. We evaluate by attenuated total
reflectance−Fourier transform infrared spectroscopy (ATR−FTIR)
and thermogravimetric analysis/differential scanning calorimetry (TGA/DSC)
the limitations of commonly used approaches. A mechanism and multiple
exchange scheme that attains the goal of complete and irreversible
ligand replacement on monodisperse nanoparticles of various sizes
is presented. The obtained hydrophobic nanoparticles are ideally suited
for magnetically controlled drug delivery and membrane applications
and for the investigation of fundamental interfacial properties of
ultrasmall core–shell architectures
Selective (Bio)Functionalization of Solid-State Nanopores
We present a method to selectively
(bio)functionalize nanoscale
features with the same materials chemistry. It was successfully combined
with nanosphere lithography to fabricate and functionalize solid-state
nanopores with PEG-brushes, supported lipid membranes, and functional
proteins over large areas. The method is inexpensive, can be performed
without specialized equipment, and can be applied to both topographic
and planar surface modification
Influence of Grafted Block Copolymer Structure on Thermoresponsiveness of Superparamagnetic Core–Shell Nanoparticles
The
morphology and topology of thermoresponsive polymers have a
strong impact on their responsive properties. Grafting onto spherical
particles has been shown to reduce responsiveness and transition temperatures;
grafting of block copolymers has shown that switchable or retained
wettability of a surface or particle during desolvation of one block
can take place. Here, doubly thermoresponsive block copolymers were
grafted onto spherical, monodisperse, and superparamagnetic iron oxide
nanoparticles to investigate the effect of thermal desolvation on
spherical brushes of block copolymers. By inverting the block order,
the influence of core proximity on the responsive properties of the
individual blocks could be studied as well as their relative influence
on the nanoparticle colloidal stability. The inner block was shown
to experience a stronger reduction in transition temperature and transition
enthalpy compared to the outer block. Still, the outer block also
experiences a significant reduction in responsiveness due to the restricted
environment in the nanoparticle shell compared to that of the free
polymer state. The demonstrated pronounced distance dependence importantly
implies the possibility, but also the necessity, to radially tailor
polymer hydration transitions for applications such as drug delivery,
hyperthermia, and biotechnological separation for which thermally
responsive nanoparticles are being developed
Individually Stabilized, Superparamagnetic Nanoparticles with Controlled Shell and Size Leading to Exceptional Stealth Properties and High Relaxivities
Superparamagnetic
iron oxide nanoparticles (SPION) have received immense interest for
biomedical applications, with the first clinical application as negative
contrast agent in magnetic resonance imaging (MRI). However, the first
generation MRI contrast agents with dextran-enwrapped, polydisperse
iron oxide nanoparticle clusters are limited to imaging of the liver
and spleen; this is related to their poor colloidal stability in biological
media and inability to evade clearance by the reticuloendothelial
system. We investigate the qualitatively different performance of
a new generation of individually PEG-grafted core–shell SPION
in terms of relaxivity and cell uptake and compare them to benchmark
iron oxide contrast agents. These PEG-grafted SPION uniquely enable
relaxivity measurements in aqueous suspension without aggregation
even at 9.4 T magnetic fields due to their extraordinary colloidal
stability. This allows for determination of the size-dependent scaling
of relaxivity, which is shown to follow a <i>d</i><sup>2</sup> dependence for identical core–shell structures. The here
introduced core–shell SPION with ∼15 nm core diameter
yield a higher <i>R</i><sub>2</sub> relaxivity than previous
clinically used contrast agents as well as previous generations of
individually stabilized SPION. The colloidal stability extends to
control over evasion of macrophage clearance and stimulated uptake
by SPION functionalized with protein ligands, which is a key requirement
for targeted MRI
Evaluation of High-Yield Purification Methods on Monodisperse PEG-Grafted Iron Oxide Nanoparticles
Fundamental research
on nanoparticle (NP) interactions and development
of next-generation biomedical NP applications relies on synthesis
of monodisperse, functional, core–shell nanoparticles free
of residual dispersants with truly homogeneous and controlled physical
properties. Still, synthesis and purification of e.g. such superparamagnetic
iron oxide NPs remain a challenge. Comparing the success of different
methods is marred by the sensitivity of analysis methods to the purity
of the product. We synthesize monodisperse, oleic acid (OA)-capped,
Fe<sub>3</sub>O<sub>4</sub> NPs in the superparamagnetic size range
(3–10 nm). Ligand exchange of OA for poly(ethylene glycol)
(PEG) was performed with the PEG irreversibly grafted to the NP surface
by a nitrodopamine (NDA) anchor. Four different methods were investigated
to remove excess ligands and residual OA: membrane centrifugation,
dialysis, size exclusion chromatography, and precipitation combined
with magnetic decantation. Infrared spectroscopy and thermogravimetric
analysis were used to determine the purity of samples after each purification
step. Importantly, only magnetic decantation yielded pure NPs at high
yields with sufficient grafting density for biomedical applications
(∼1 NDA-PEG(5 kDa)/nm<sup>2</sup>, irrespective of size). The
purified NPs withstand challenging tests such as temperature cycling
in serum and long-term storage in biological buffers. Dynamic light
scattering, transmission electron microscopy, and small-angle X-ray
scattering show stability over at least 4 months also in serum. The
successful synthesis and purification route is compatible with any
conceivable functionalization for biomedical or biomaterial applications
of PEGylated Fe<sub>3</sub>O<sub>4</sub> NPs
Embedded Plasmonic Nanomenhirs as Location-Specific Biosensors
We introduce a novel optical biosensing
platform that exploits
the asymmetry of nanostructures embedded in nanocavities, termed nanomenhirs.
Upon oblique illumination using plane polarized white light, two plasmonic
resonances attributable to the bases and the axes of the nanomenhirs
emerge; these are used for location-specific sensing of membrane-binding
events. Numerical simulations of the near field distributions confirmed
the experimental results. As a proof-of-concept, we present a model
biosensing experiment that exploits the dual-sensing capability, the
size selectivity offered by the sensor geometry, and the possibility
to separately biochemically modify the nanomenhirs and the nanocavities
for the specific binding of lipid membrane structures to the nanomenhirs
Supported Lipopolysaccharide Bilayers
In this report, the formation of supported lipopolysaccharide
bilayers
(LPS-SLBs) is studied with extracted native and glycoengineered LPS
from Escherichia coli (E. coli) and Salmonella enterica sv
typhimurium (S. typhimurium) to assemble a platform that allows measurement of LPS membrane
structure and the detection of membrane tethered saccharide-protein
interactions. We present quartz crystal microbalance with dissipation
monitoring (QCM-D) and fluorescence recovery after photobleaching
(FRAP) characterization of LPS-SLBs with different LPS species, having,
for example, different molecular weights, that show successful formation
of SLBs through vesicle fusion on SiO<sub>2</sub> surfaces with LPS
fractions up to 50 wt %. The thickness of the LPS bilayers were investigated
with AFM force–distance measurements which showed only a slight
thickness increase compared to pure POPC SLBs. The E. coli LPS were chosen to study the saccharide–protein
interaction between the Htype II glycan epitope and the Ralstonia solanacearum lectin (RSL). RSL specifically
recognizes fucose sugars, which are present in the used Htype II glycan
epitope and absent in the epitopes LPS1 and EY2. We show via fluorescence
microscopy that the specific, but weak and multivalent interaction
can be detected and discriminated on the LPS-SLB platform
Interaction of Size-Tailored PEGylated Iron Oxide Nanoparticles with Lipid Membranes and Cells
Targeted nanomedicine
builds on the concept that nanoparticles
can be directed to specific tissues while remaining inert to others
organs. Many studies have been performed on the synthesis and cellular
interactions of core–shell nanoparticles, in which a functional
inorganic core is coated with a biocompatible polymer layer that should
reduce nonspecific uptake and cytotoxicity. However, work is lacking
that relates structural parameters of the core–shell structure
and colloidal properties directly to interactions with cell membranes
and further correlates these interactions to cell uptake. We have
synthesized monodisperse (SD < 10%), single-crystalline, and superparamagnetic
iron oxide nanoparticles (SPION) of different core size (3–8
nm) that are densely grafted with nitrodopamine-poly(ethylene glycol)
(NDA-PEG(5 kDa)) brushes. We investigated the interactions of the
PEGylated SPION with biomimetic membranes and cancer and kidney cells.
It is shown that a dense homogeneous PEG shell suppresses membrane
interactions and cell uptake but that nanoparticle curvature can influence
membrane interactions for similarly grafted nanoparticles. Weak adsorption
to anionic lipid membranes is shown to correlate with eukaryote cell
uptake and is attributed to double-layer interactions without direct
membrane penetration. This attraction is strongly suppressed during
physiological conditions and leads to unprecedented low cell uptake
and full cell viability when compared to those of traditional dextran-coated
SPION. Less curved (larger core) PEGylated SPION show weaker membrane
adsorption and lower cell uptake due to effectively denser shells.
These results provide a better understanding of design criteria for
core–shell nanoparticles in terms of avoiding nonspecific uptake
by cells, reducing toxicity, and increasing circulation time