A new microfluidic methodology to assess the haemocompatibility of magnetic nanoparticles designed for theranostic applications

The remarkable physicochemical properties of magnetic nanoparticles (MNPs) at the nanoscale have boosted the development of new and promising strategies for the simultaneous diagnosis and treatment of diseases, particularly in cancer therapy Ð the so-called theranostic applications (1). In these strategies, the intrinsic superparamagnetic properties of MNPs have been exploited to gain access into multifunctional systems able to simultaneously perform as enhanced magnetic resonance imaging (MRI) contrast agents, efficient nanocarriers for drug delivery and nanoheaters in magnetic hyperthermia based therapy (2), among others.

Haemocompatibility of iron oxide nanoparticles synthesized for theranostic applications: a high-sensitivity microfluidic tool

The poor heating efficiency of the most reported magnetic nanoparticles (MNPs), allied to the lack of comprehensive biocompatibility and haemodynamic studies, hampers the spread of multifunctional nanoparticles as the next generation of therapeutic bio-agents in medicine. The present work reports the synthesis and characterization, with special focus on biological/toxicological compatibility, of superparamagnetic nanoparticles with diameter around 18 nm, suitable for theranostic applications (i.e. simultaneous diagnosis and therapy of cancer). Envisioning more insights into the complex nanoparticle-red blood cells (RBCs) membrane interaction, the deformability of the human RBCs in contact with magnetic nanoparticles (MNPs) was assessed for the first time with a microfluidic extensional approach, and used as an indicator of haematological disorders in comparison with a conventional haematological test, i.e. the haemolysis analysis. Microfluidic results highlight the potential of this microfluidic tool over traditional haemolysis analysis, by detecting small increments in the rigidity of the blood cells, when traditional haemotoxicology analysis showed no significant alteration (haemolysis rates lower than 2 %). The detected rigidity has been predicted to be due to the wrapping of small MNPs by the bilayer membrane of the RBCs, which is directly related to MNPs size, shape and composition. The proposed microfluidic tool adds a new dimension into the field of nanomedicine, allowing to be applied as a highsensitivity technique capable of bringing a better understanding of the biological impact of nanoparticles developed for clinical applications.

Effect of solvent; enhancing the wettability and engineering the porous structure of a calcium phosphate/agarose composite for drug delivery

Tissue engineering deals with the regeneration of tissues for bone repair, wound healing, drug delivery, etc., and a highly porous 3D artificial scaffold is required to accommodate the cells and direct their growth. We prepared 3D porous calcium phosphate ((hydroxyapatite/beta-tricalcium phosphate)/agarose, (HAp/beta-TCP)/agarose) composite scaffolds by sol-gel technique with water (WBS) and ethanol (EBS) as solvents. The crystalline phases of HAp and beta-TCP in the scaffolds were confirmed by X-ray diffraction (XRD) analysis. The EBS had reduced crystallinity and crystallite size compared to WBS. WBS and EBS revealed interconnected pores of 1 mu m and 100 nm, respectively. The swelling ratio was higher for EBS in water and phosphate buffered saline (PBS). An in vitro drug loading/release experiment was carried out on the scaffolds using gentamicin sulphate (GS) and amoxicillin (AMX). We observed initial burst release followed by sustained release from WBS and EBS. In addition, GS showed more extended release than AMX from both the scaffolds. GS and AMX loaded scaffolds showed greater efficacy against Pseudomonas than Bacillus species. WBS exhibited enhanced mechanical properties, wettability, drug loading and haemocompatibility compared to EBS. In vitro cell studies showed that over the scaffolds, MC3T3 cells attached and proliferated and there was a significant increase in live MC3T3 cells. Both scaffolds supported MC3T3 proliferation and mineralization in the absence of osteogenic differentiation supplements in media which proves the scaffolds are osteoconducive. Microporous scaffolds (WBS) could assist the bone in-growth, whereas the presence of nanopores (EBS) could enhance the degradation process. Hence, WBS and EBS could be used as scaffolds for tissue engineering and drug delivery. This is a cost effective technique to produce scaffolds of degradable 3D ceramic-polymer composites.

Effect of acyl chain length on transfection efficiency and toxicity of polyethylenimine

Polyethylenimine (PEI) is an efficient nonviral gene delivery vector because of its high buffering capacity and DNA condensation ability. In our study, the amino groups on the polymeric backbone were acylated using acetic or propionic anhydride to alter the protonation behaviour and the hydrophilic/hydrophobic balance of the polymer. The concentration of acylated primary amines was determined using trinitrobenzene sulphonic acid assay. Results showed that our modified polymers had lower buffering capacities in solutions compared to PEI. The polymers were complexed with plasmid encoding enhanced green fluorescent protein at three different ratios (1:1, 1:2 and 1:10 w/w DNA to polymer) to form polyplexes and their toxicities and transfection efficiencies were evaluated in HEK 293 cells. Acylation reduced the number of primary amines on the polymer and the surface charge, improving haemocompatibility and reducing cytotoxicity. The reduction in the concentration of amino groups helped to optimise DNA compaction and facilitated polyplex dissociation in the cell, which increased transfection efficiency of the modified polymers compared to the parent polymer. Polymers with buffering capacities greater than 50% and less than 80% relative to PEI, showed higher transfection efficiencies than PEI. The propionic anhydride modified polymers had appropriate interactions with DNA which provided both DNA compaction and polyplex dissociation. These systems interacted better with the cell membrane because of their slightly higher lipophilicity and formed polyplexes which were less cytotoxic than polyplexes of acetic anhydride modified polymers. Among the vectors tested, 1:0.3 mol/mol PEI:propionic anhydride in a 1:2 w/w DNA:polymer composition provided the best transfection system with improved transfection efficiency and reduced cytotoxicity.

Biocompatible, biomimetic ampholyte materials

New ampholyte biomaterial compounds containing ampholyte moieties are synthesized and integrated into polymeric assemblies to provide hydrophilic polymers exhibiting improved biocompatibility, haemocompatibility, hydrophilicity non-thrombogenicity, anti-bacterial ability, and mechanical strength, as well as suitability as a drug delivery platform.

Counter-ion effect on surfactant-DNA gel particles as controlled DNA delivery systems

The ability to entrap drugs within vehicles and subsequently release them has led to new treatments for a number of diseases. Based on an associative phase separation and interfacial diffusion approach, we developed a way to prepare DNA gel particles without adding any kind of cross-linker or organic solvent. Among the various agents studied, cationic surfactants offered particularly efficient control for encapsulation and DNA release from these DNA gel particles. The driving force for this strong association is the electrostatic interaction between the two components, as induced by the entropic increase due to the release of the respective counter-ions. However, little is known about the influence of the respective counter-ions on this surfactant-DNA interaction. Here we examined the effect of different counter-ions on the formation and properties of the DNA gel particles by mixing DNA (either single-(ssDNA) or double-stranded (dsDNA)) with the single chain surfactant dodecyltrimethylammonium (DTA). In particular, we used as counter-ions of this surfactant the hydrogen sulfate and trifluoromethane sulfonate anions and the two halides, chloride and bromide. Effects on the morphology of the particles obtained, the encapsulation of DNA and its release, as well as the haemocompatibility of these particles are presented, using counter-ion structure and DNA conformation as controlling parameters. Analysis of the data indicates that the degree of counter-ion dissociation from the surfactant micelles and the polar/hydrophobic character of the counter-ion are important parameters in the final properties of the particles. The stronger interaction with amphiphiles for ssDNA than for dsDNA suggests the important role of hydrophobic interactions in DNA.