Drug delivery of 6-bromoindirubin-3'-glycerol-oxime ether employing poly(D,L-lactide-co-glycolide)-based nanoencapsulation techniques with sustainable solvents.

BACKGROUND: Insufficient solubility and stability of bioactive small molecules as well as poor biocompatibility may cause low bioavailability and are common obstacles in drug development. One example of such problematic molecules is 6-bromoindirubin-3'-glycerol-oxime ether (6BIGOE), a hydrophobic indirubin derivative. 6BIGOE potently modulates the release of inflammatory cytokines and lipid mediators from isolated human monocytes through inhibition of glycogen synthase kinase-3 in a favorable fashion. However, 6BIGOE suffers from poor solubility and short half-lives in biological aqueous environment and exerts cytotoxic effects in various mammalian cells. In order to overcome the poor water solubility, instability and cytotoxicity of 6BIGOE, we applied encapsulation into poly(D,L-lactide-co-glycolide) (PLGA)-based nanoparticles by employing formulation methods using the sustainable solvents Cyrene™ or 400 g/mol poly(ethylene glycol) as suitable technology for efficient drug delivery of 6BIGOE. RESULTS: For all preparation techniques the physicochemical characterization of 6BIGOE-loaded nanoparticles revealed comparable crystallinity, sizes of about 230 nm with low polydispersity, negative zeta potentials around - 15 to - 25 mV, and biphasic release profiles over up to 24 h. Nanoparticles with improved cellular uptake and the ability to mask cytotoxic effects of 6BIGOE were obtained as shown in human monocytes over 48 h as well as in a shell-less hen's egg model. Intriguingly, encapsulation into these nanoparticles fully retains the anti-inflammatory properties of 6BIGOE, that is, favorable modulation of the release of inflammation-relevant cytokines and lipid mediators from human monocytes. CONCLUSIONS: Our formulation method of PLGA-based nanoparticles by applying sustainable, non-toxic solvents is a feasible nanotechnology that circumvents the poor bioavailability and biocompatibility of the cargo 6BIGOE. This technology yields favorable drug delivery systems for efficient interference with inflammatory processes, with improved pharmacotherapeutic potential.

The Effect of Stereocomplexation and Crystallinity on the Degradation of Polylactide Nanoparticles.

Polymeric nanoparticles (PNPs) are frequently researched and used in drug delivery. The degradation of PNPs is highly dependent on various properties, such as polymer chemical structure, size, crystallinity, and melting temperature. Hence, a precise understanding of PNP degradation behavior is essential for optimizing the system. This study focused on enzymatic hydrolysis as a degradation mechanism by investigation of the degradation of PNP with various crystallinities. The aliphatic polyester polylactide ([C3H4O2]n, PLA) was used as two chiral forms, poly l-lactide (PlLA) and poly d-lactide (PdLA), and formed a unique crystalline stereocomplex (SC). PNPs were prepared via a nanoprecipitation method. In order to further control the crystallinity and melting temperatures of the SC, the polymer poly(3-ethylglycolide) [C6H8O4]n (PEtGly) was synthesized. Our investigation shows that the PNP degradation can be controlled by various chemical structures, crystallinity and stereocomplexation. The influence of proteinase K on PNP degradation was also discussed in this research. AFM did not reveal any changes within the first 24 h but indicated accelerated degradation after 7 days when higher EtGly content was present, implying that lower crystallinity renders the particles more susceptible to hydrolysis. QCM-D exhibited reduced enzyme adsorption and a slower degradation rate in SC-PNPs with lower EtGly contents and higher crystallinities. A more in-depth analysis of the degradation process unveiled that QCM-D detected rapid degradation from the outset, whereas AFM exhibited delayed changes of degradation. The knowledge gained in this work is useful for the design and creation of advanced PNPs with enhanced structures and properties.

Rutile facet-dependent fibrinogen conformation: Why crystallographic orientation matters.

Previous studies implied that single crystalline rutile surfaces have the ability to guide the functionality of adsorbed blood plasma proteins. However, a clear relation between the rutile crystallographic orientation and conformation of adsorbed proteins is still missing. Here, we examine the adsorption characteristics of human plasma fibrinogen (HPF) on atomically flat single rutile crystals with (110), (100), (101) and (001) facets. By direct visualization of individual protein molecules through atomic force microscopy (AFM) imaging, the distinct conformations of HPF were determined depending on rutile surface crystallographic orientation. In particular, dominant trinodular and globular conformation was found on (110) and (001) facets, respectively. The observed variations of HPF conformation were reasoned from the surface water contact angle and surface energy point of view. By analyzing AFM-based force measurements, statistically significant changes in surface energies of rutile surfaces covered with HPF were determined and linked to HPF conformation. Furthermore, the facet-dependent structural rearrangement of HPF was indirectly confirmed through deconvolution of high-resolution X-ray photoelectron spectroscopy (XPS) carbon and nitrogen spectra. The globular, and thus native-like HPF conformation observed on (001) facet, was reflected in the lowest level of amino group formation. We propose that the mechanism behind the crystallographic orientation-induced HPF conformation is driven by the facet-specific surface hydrophilicity and energy. From the biomedical material perspective, our results demonstrate that the conformation of HPF can be guided by controlling the crystallographic orientation of the underlying material surface. This might be beneficial to the field of titanium-based biomaterials design and development.

Self-assembled fibrinogen-fibronectin hybrid protein nanofibers with medium-sensitive stability.

Hybrid protein nanofibers (hPNFs) have been identified as promising nano building blocks for numerous applications in nanomedicine and tissue engineering. We have recently reported a nature-inspired, self-assembly route to create hPNFs from human plasma proteins, i.e., albumin and hemoglobin. However, it is still unclear whether the same route can be applied to other plasma proteins and whether it is possible to control the composition of the resulting fibers. In this context, to further understand the hPNFs self-assembly mechanism and to optimize their properties, we report herein on ethanol-induced self-assembly of two different plasma proteins, i.e., fibrinogen (FG) and fibronectin (FN). We show that by varying initial protein ratios, the composition and thus the properties of the resulting hPNFs can be fine-tuned. Specifically, atomic force microscopy, hydrodynamic diameter, and zeta potential data together revealed a strong correlation of the hPNFs dimensions and surface charge to their initial protein mixing ratio. The composition-independent prompt dissolution of hPNFs in ultrapure water, in contrast to their stability in PBS, indicates that the molecular arrangement of FN and FG in hPNFs is mainly based on electrostatic interactions. Supported by experimental data we introduce a feasible mechanism that explains the interactions between FN and FG and their self-assembly to hPNFs. These findings contribute to the understanding of dual protein interactions, which can be beneficial in designing innovative biomaterials with multifaceted biological and physical characteristics.

Distinct endocytosis and immune activation of poly(lactic-co-glycolic) acid nanoparticles prepared by single- and double-emulsion evaporation.

Background: Poly(lactic-co-glycolic) acid (PLGA) nanoparticles can be prepared by emulsion-solvent-evaporation from o/w and w1/o/w2 emulsions. Aims: To elaborate similarities and differences regarding mechanical, morphological and physicochemical properties, as well as endocytosis and dose-dependent immune responses by primary human leukocytes between nanoparticles prepared by these two methods. Methods: Fluorescently labeled as well as TLR agonist (R848)-loaded PLGA nanoparticles were prepared via both single- and double-emulsion solvent evaporation. Results: Particles prepared by both methods were similar in chemical composition and surface charge but exhibited slight differences in size and morphology. Pronounced differences were found for loading, dissolution and mechanical properties. The particles were differently endocytosed by monocytes and induced qualitatively and quantitatively different immune responses. Conclusions: Variations in nanoparticle preparation can affect particle-derived immunological characteristics.

Sustainable preparation of anti-inflammatory atorvastatin PLGA nanoparticles.

In the present study, the anti-inflammatory lipophilic drug atorvastatin was encapsulated in poly(D,L-lactide-co-glycolide) (PLGA) using a sustainable method in comparison to the standard emulsion-diffusion-evaporation technique. For the sustainable method the organic solvent ethyl acetate was fully replaced by 400 g/mol poly(ethylene glycol) (PEG 400). Both techniques led to the formation of nanoparticles with comparable sizes of about 170 to 247 nm depending on the polymer type, with monomodal size distribution and negative zeta potential. All nanoparticles demonstrated a high biocompatibility in a shell-less hen's egg model and displayed an anti-inflammatory effect in human monocytes. The use of PEG 400 resulted in plasticizing effects and a lower crystallinity of the PLGA nanoparticles as determined by differential scanning calorimetry and Raman spectroscopy, which correlated with a faster drug release. Interestingly, the particles prepared by the sustainable method showed a crystallinity and drug release kinetics similar to nanoparticles made of PEG-PLGA using the standard method. Conclusively, the sustainable method is a fast and easy to perform technique suitable to prepare atorvastatin-loaded PLGA nanoparticles avoiding toxic and environmentally damaging drawbacks frequently associated with classical organic solvents.