Mihaela Delcea

Stimuli-responsive LbL capsules and nanoshells for drug delivery ☆

Review of basic principles and recent developments in the area of stimuli responsive polymeric capsules and nanoshells formed via layer-by-layer (LbL) is presented. The most essential attributes of the LbL approach are multifunctionality and responsiveness to a multitude of stimuli. The stimuli can be logically divided into three categories: physical (light, electric, magnetic, ultrasound, mechanical, and temperature), chemical (pH, ionic strength, solvent, and electrochemical) and biological (enzymes and receptors). Using these stimuli, numerous functionalities of nanoshells have been demonstrated: encapsulation, release including that inside living cells or in tissue, sensors, enzymatic reactions, enhancement of mechanical properties, and fusion. This review describes mechanisms and basic principles of stimuli effects, describes progress in the area, and gives an outlook on emerging trends such as theranostics and nanomedicine.

Multicompartmental Micro- and Nanocapsules: Hierarchy and Applications in Biosciences

Multicompartmentalized micro- and nanocapsules allow simultaneous delivery of several vectors or biomolecules; they are the next generation of carriers with increased complexity. Here we overview multicompartment micro- and nanocapsules and present a road-map for future developments in the field. Four basic building block structures are demonstrated, three isotropic: concentric, pericentric, and innercentric, and one anisotropic: acentric. As an elaborate implementation of multicompartmentalization, an enzyme-catalyzed reaction inside the same capsule carrying both an enzyme and a substrate is shown. Applications of multicompartmentalized microcapsules for simultaneous multiple drug delivery in bio-medicine are discussed.

Nanoplasmonics for dual-molecule release through nanopores in the membrane of red blood cells

A nanoplasmonics-based opto-nanoporation method of creating nanopores upon laser illumination is applied for inducing diffusion and triggered release of small and large molecules from red blood cells (RBCs). The method is implemented using absorbing gold nanoparticle (Au-NP) aggregates on the membrane of loaded RBCs, which, upon near-IR laser light absorption, induce release of encapsulated molecules from selected cells. The binding of Au-NPs to RBCs is characterized by Raman spectroscopy. The process of release is driven by heating localized at nanoparticles, which impacts the permeability of the membrane by affecting the lipid bilayer and/or trans-membrane proteins. Localized heating and temperature rise around Au-NP aggregates is simulated and discussed. Research reported in this work is relevant for generating nanopores for biomolecule trafficking through polymeric and lipid membranes as well as cell membranes, while dual- and multi-molecule release is relevant for theragnostics and a wide range of therapies.

Mechanobiology: Correlation Between Mechanical Stability of Microcapsules Studied by AFM and Impact of Cell-Induced Stresses

Intracellular delivery of proteins, peptides, and other biomolecules [ 1,2 ] by microcapsules is of growing importance not only in applied biomedical research, but also in fundamental cell biology. [ 3 ] Although microcapsules [ 2 , 4–8 ] have the potential to reveal information about mechanobiology and cell mechanics, previous investigations of their mechanical properties have only been used for designing delivery vehicles with improved mechanical strength. [ 9–11 ] Accordingly, the force-deformation behavior of polyelectrolyte capsules has been intensively studied. [ 12–14 ] Several parameters such as temperature, number of layers, and encapsulated contents infl uence the mechanical properties of the capsules. It has been shown that incubation at high temperatures results in drastic changes of the morphology of polymeric capsules accompanied by increasing stiffness. [ 7 , 15–17 ] Furthermore, delivery and release, including pulsed release, [ 18 ] remotely controllable release, [ 1 ]

Anti–protamine-heparin antibodies: incidence, clinical relevance, and pathogenesis

Protamine, which is routinely used after cardiac surgery to reverse the anticoagulant effects of heparin, is known to be immunogenic. Observing patients with an otherwise unexplained rapid decrease in platelet count directly after protamine administration, we determined the incidence and clinical relevance of protamine-reactive antibodies in patients undergoing cardiac-surgery. In vitro, these antibodies activated washed platelets in a FcγRIIa-dependent fashion. Using a nonobese diabetic/severe combined immunodeficiency mouse model, those antibodies induced thrombocytopenia only when protamine and heparin were present but not with protamine alone. Of 591 patients undergoing cardiopulmonary bypass surgery, 57 (9.6%) tested positive for anti-protamine-heparin antibodies at baseline and 154 (26.6%) tested positive at day 10. Diabetes was identified as a risk factor for the development of anti-protamine-heparin antibodies. In the majority of the patients, these antibodies were transient and titers decreased substantially after 4 months (P < .001). Seven patients had platelet-activating, anti-protamine-heparin antibodies at baseline and showed a greater and more prolonged decline in platelet counts compared with antibody-negative patients (P = .003). In addition, 2 of those patients experienced early arterial thromboembolic complications vs 9 of 584 control patients (multivariate analysis: odds ratio, 21.58; 95% confidence interval, 2.90-160.89; P = .003). Platelet-activating anti-protamine-heparin antibodies show several similarities with anti-platelet factor 4-heparin antibodies and are a potential risk factor for early postoperative thrombosis.

Patchiness of Embedded Particles and Film Stiffness Control Through Concentration of Gold Nanoparticles

Patchy particles are fabricated using a method of embedding-into and extracting-from thick, biocompatible, gel-like HA/PLL films. Control over the patchiness is achieved by adjusting the stiffness of films, which affects embedding and masking of particles. The stiffness is adjusted by the concentration of gold nanoparticles adsorbed onto the surface of the films.

Anisotropic multicompartment micro- and nano-capsules produced via embedding into biocompatible PLL/HA films

We present a novel strategy to fabricate anisotropic multicompartment Janus capsules by embedding larger containers into a soft poly-L-lysine/hyaluronic acid (PLL/HA) polymeric film, followed by adsorption of smaller containers on top of their unmasked surface. This research is also attractive for developing substrates for cell cultures.

Thermal stability, mechanical properties and water content of bacterial protein layers recrystallized on polyelectrolyte multilayers

The influence of the temperature on the surface topology, layer thickness and density of recrystallized bacterial S-layers from Bacillus sphaericus CCM2177 on polyelectrolyte multilayers in contact with liquid water was investigated. A quartz crystal microbalance with dissipation (QCM-D) was used to monitor the build-up of the polyelectrolyte multilayer and the adsorption of S-layer protein (1600 ng cm-2). The critical temperature (55 °C) at which the S-layer loses its 2-D structure was obtained from atomic force microscopy (AFM) and confirmed by neutron reflectometry (NR) experiments. The process of S-layer denaturation was found to be irreversible. Aggregates of denatured S-proteins resist lower loads than the crystalline nanostructure formed from folded S-proteins. The combination of the QCM-D results with the scattering length density and film thickness (14 nm) obtained from neutron reflectometry studies permitted the estimation of the density of adsorbed S-protein together with the bound water (M = 1.16 g cm-3), the dry protein scattering length density (2.02 × 10-6 Å-2) and the S-protein mass density (1.48 g cm-3). The results confirmed that S-proteins form very loosely packed layers on polyelectrolyte multilayers incorporating a water volume fraction of around 68%.

Binding of anti-platelet factor 4/heparin antibodies depends on the thermodynamics of conformational changes in platelet factor 4.

The chemokine platelet factor 4 (PF4) undergoes conformational changes when complexing with polyanions. This can induce the antibody-mediated adverse drug effect of heparin-induced thrombocytopenia (HIT). Understanding why the endogenous protein PF4 becomes immunogenic when complexing with heparin is important for the development of other negatively charged drugs and may also hint toward more general mechanisms underlying the induction of autoantibodies to other proteins. By circular dichroism spectroscopy, atomic force microscopy, and isothermal titration calorimetry we characterized the interaction of PF4 with unfractionated heparin (UFH), its 16-, 8-, and 6-mer subfractions, low-molecular-weight heparin (LMWH), and the pentasaccharide fondaparinux. To bind anti-PF4/heparin antibodies, PF4/heparin complexes require (1) an increase in PF4 antiparallel β-sheets exceeding ∼30% (achieved by UFH, LMWH, 16-, 8-, 6-mer), (2) formation of multimolecular complexes (UFH, 16-, 8-mer), and (3) energy (needed for a conformational change), which is released by binding of ≥11-mer heparins to PF4, but not by smaller heparins. These findings may help to synthesize safer heparins. Beyond PF4 and HIT, the methods applied in the current study may be relevant to unravel mechanisms making other endogenous proteins more vulnerable to undergo conformational changes with little energy requirement (eg, point mutations and post-translational modifications) and thereby predisposing them to become immunogenic.

Fabrication of quantum dot microarrays using electron beam lithography for applications in analyte sensing and cellular dynamics.

Quantum dot (QD) based micro-/nanopatterned arrays are of broad interest in applications ranging from electronics, photonics, to sensor devices for biomedical purposes. Here, we report on a rapid, physico-chemically mild approach to generate high fidelity micropattern arrays of prefunctionalized water-soluble quantum dots using electron beam lithography. We show that such patterns retain their fluorescence and bioaffinity upon electron beam lithography and, based on the streptavidin-biotin interaction, allow for detection of proteins, colloidal gold nanoparticles and magnetic microparticles. Furthermore, we demonstrate the applicability of QD based microarray patterns differing in their shape (circles, squares, grid-like), size (from 1 to 10 μm) and pitch distance to study the adhesion, spreading and migration of human blood derived neutrophils. Using live cell confocal fluorescence microscopy, we show that pattern geometry and pitch distance influence the adhesion, spreading and migratory behavior of neutrophils. Research reported in this work paves the way for producing QD microarrays with multiplexed functionalities relevant for applications in analyte sensing and cellular dynamics.