The impact of the pre-treatment interval on antimicrobial efficacy in a biological model

The impact of pre-treatment intervals on the antipseudomonal efficacy of gentamicin, ticarcillin and ceftazidime was studied in an experimental thigh infection model in normal and granulocytopenic mice. Human-equivalent doses were used for simulating human pharmacokinetic profiles of the two study β-lactam drugs. A lethal inoculum of a virulent strain of Pseudomonas aeruginosa was injected into the thigh muscle. Treatment was started at various post-infection intervals. Antimicrobial efficacy was assessed by determinations of surviving organisms at the site of infection, and plasma drug concentrations were determined in the same mice. The age of infection had a substantial impact on antipseudomonal efficacy of the three study drugs even though high, brief supra-MIC concentrations of gentamicin and persistent supra-MIC concentrations of the β-lactam drugs were obtained. A pre-treatment interval of six or more hours abolished the bactericidal effect of all three study drugs despite accumulation of the drugs to multiple-MBC plasma concentrations. We believe that the impact of pre-treatment intervals on antimicrobial efficacy is of paramount importance for the interpretation of antimicrobial activity studies in experimental models of infection, although the mechanisms remain to be elucidate

Vectorisation intra-oculaire [Drug delivery to target the posterior segment of the eye].

Retinal diseases are nowadays the most common causes of vision threatening in developed countries. Therapeutic advances in this field are hindered by the difficulty to deliver drugs to the posterior segment of the eye. Due to anatomical barriers, the ocular biodisponibility of systemically administered drugs remains poor, and topical instillation is not adequate to achieve therapeutic concentrations of drugs in the back of the eye. Ocular drug delivery has thus become one of the main challenges of modern ophthalmology. A multidisciplinary research is being conducted worldwide including pharmacology, biomaterials, ophthalmology, pharmaceutics, and biology. New promising fields have been developed such as implantable or injectable slow release intravitreal devices and degradable polymers, dispersed polymeric systems for intraocular drug delivery, and transscleral delivery devices such as iontophoresis, osmotic pumps or intra-scleraly implantable materials. The first clinical applications emerging from this research are now taking place, opening new avenues for the treatment of retinal diseases

Amikacin, Ceftazidime, and Flucloxacillin against Suspended and Adherent Pseudomonas aeruginosa and Staphylococcus epidermidis in an In Vitro Model of Infection

Bacterial inocula were exposed as suspended cultures or as adherent biofilms on glass beads in a novel in vitro model of infection to oscillating drug concentrations mimicking human serum kinetics during clinical treatment. Amikacin was given once or thrice daily alone or in combination with ceftazidime or flucloxacillin against Pseudomonas aeruginosa or Staphylococcus epidermidis. Killing of adherent bacteria was significantly reduced during single-drug treatment compared with suspended bacteria (P < .001), and β-lactams were more active than amikacin against both suspended and adherent bacteria (P < .01), Amikacin-β-lactam combinations killed the inocula more rapidly and were consistently bactericidal against both suspended and adherent pathogens (P < .05). Once-daily dosing of amikacin produced greater initial killing than thrice daily dosing (P < .05), but both regimens were similarly effective after 48 h. The differences in antibiotic activity against suspended and adherent bacteria may relate to clinical failures in the treatment of foreign-body infections by bacteria sensitive to the administered antibiotics, as determined by standard susceptibility test

Intravitreal Polymeric Nanocarriers with Long Ocular Retention and Targeted Delivery to the Retina and Optic Nerve Head Region

Posterior eye tissues, such as retina, are affected in many serious eye diseases, but drug delivery to these targets is challenging due to various anatomical eye barriers. Intravitreal injections are widely used, but the intervals between invasive injections should be prolonged. We synthesized and characterized (1H NMR, gel permeation chromatography) block copolymers of poly(ethylene glycol), poly(caprolactone), and trimethylene carbonate. These polymers self-assembled to polymersomes and polymeric micelles. The mean diameters of polymersomes and polymeric micelles, about 100 nm and 30–50 nm, respectively, were obtained with dynamic light scattering. Based on single particle tracking and asymmetric flow field-flow fractionation, the polymeric micelles and polymersomes were stable and diffusible in the vitreous. The materials did not show cellular toxicity in cultured human umbilical vein endothelial cells in the Alamar Blue Assay. Pharmacokinetics of the intravitreal nanocarriers in the rabbits were evaluated using in vivo fluorophotometry. The half-lives of the polymersomes (100 nm) and the micelles (30 nm) were 11.4–32.7 days and 4.3–9.5 days. The intravitreal clearance values were 1.7–8.7 µL/h and 3.6–5.4 µL/h for polymersomes and polymeric micelles, respectively. Apparent volumes of distribution of the particles in the rabbit vitreous were 0.6–1.3 mL for polymeric micelles and 1.9–3.4 mL for polymersomes. Polymersomes were found in the vitreous for at least 92 days post-dosing. Furthermore, fundus imaging revealed that the polymersomes accumulated near the optic nerve and retained there even at 111 days post-injection. Polymersomes represent a promising technology for controlled and site-specific drug delivery in the posterior eye segment

Liposomes as delivery systems in the prevention and treatment of infectious diseases

Research on the potential application of liposomes in the prevention and treatment of infectious diseases has focussed on improvement of the therapeutic index of antimicrobial drugs and immunomodulators and on stimulation of the immune response to otherwise weak antigens in vaccines composed of purified micro-organism subunits. In this review current approaches in this field are outlined. The improved therapeutic index of antimicrobial drugs after encapsulation in liposomes is a result of enhanced drug delivery to infected tissue or infected cells and/or a reduction of drug toxicity of potentially toxic antibiotics. Liposomal encapsulation of immunomodulators that activate macrophages aims at reducing the toxicity of these agents and targeting them to the cells of the mononuclear phagocyte system in order to increase the nonspecific resistance of the host against infections. Studies on the immunogenicity of liposomal antigens have demonstrated that liposomes can potentiate the humoral and cell mediated immunity to a variety of antigens

Imaging, quantitation and kinetic modelling of intravitreal nanomaterials

In this study, the intravitreal pharmacokinetics of nanomaterials were investigated in vivo in rats and rabbits. Impact of particle size and shape (spherical, longitudinal) on ocular particle distribution and elimination was investigated with fundus camera, optical coherence tomography and ocular fluorophotometry. Differently sized particles showed prolonged ocular retention and remarkable differences in vitreal elimination, but size dependence was consistent, suggesting that other features have influence on their vitreal kinetics. We also demonstrate that liposomes are eliminated from the rabbit vitreous mainly via the anterior route. Simulation of drug concentrations after injection of intravitreal particles shows the importance of synchronized particle retention and drug release rate for efficient drug delivery. In conclusion, we provide kinetic insights in intravitreally administered nanoparticles to improve retinal drug delivery.Peer reviewe

Diffusion and Protein Corona Formation of Lipid-Based Nanoparticles in the Vitreous Humor : Profiling and Pharmacokinetic Considerations

The vitreous humor is the first barrier encountered by intravitreally injected nanoparticles. Lipid-based nanoparticles in the vitreous are studied by evaluating their diffusion with single-particle tracking technology and by characterizing their protein coronae with surface plasmon resonance and high-resolution proteomics. Single-particle tracking results indicate that the vitreal mobility of the formulations is dependent on their charge. Anionic and neutral formulations are mobile, whereas larger (>200 nm) neutral particles have restricted diffusion, and cationic particles are immobilized in the vitreous. PEGylation increases the mobility of cationic and larger neutral formulations but does not affect anionic and smaller neutral particles. Convection has a significant role in the pharmacokinetics of nanoparticles, whereas diffusion drives the transport of antibodies. Surface plasmon resonance studies determine that the vitreal corona of anionic formulations is sparse. Proteomics data reveals 76 differentially abundant proteins, whose enrichment is specific to either the hard or the soft corona. PEGylation does not affect protein enrichment. This suggests that protein-specific rather than formulation-specific factors are drivers of protein adsorption on nanoparticles in the vitreous. In summary, our findings contribute to understanding the pharmacokinetics of nanoparticles in the vitreous and help advance the development of nanoparticle-based treatments for eye diseases.Peer reviewe

Effect of subconjunctivally injected, liposome-bound, low-molecular-weight heparin on the absorption rate of subconjunctival hemorrhage in rabbits

PURPOSE: To investigate the effect of subconjunctival injection of liposome-bound, low-molecular-weight heparin (LMWH) on the absorption rate of subconjunctival hemorrhages. METHODS: Subconjunctival hemorrhages were induced in both eyes of 30 rabbits by the subconjunctival injection of 0.1 mL of autologous blood from auricular marginal veins. After 8 hours, randomized subconjunctival injections of one of three materials were made: 5 IU/mL liposome-bound LMWH (0.1 mL) in 18 eyes (group A), only liposomes (0.1 mL) in 14 eyes (group B), the free form of LMWH (5 IU/mL, 0.1 mL) in 14 eyes (group C), or no injection in 14 eyes (group D). Subconjunctival hemorrhages were photographed with a digital camera at 8, 24, 48, 72, 96, and 120 hours after induction of subconjunctival hemorrhages, sized with an image analyzer, and compared between groups. RESULTS: Subconjunctival hemorrhages were absorbed faster in group A (liposome-bound LMWH injected) than in with group B (liposome injected). Comparison of groups A and C (free LMWH injected) showed statistical differences in the absorption rates at 96 and 120 hours except at 24, 48, and 72 hours. The mean elapsed time for the complete resorption of subconjunctival hemorrhages was shortest in group A among four groups, whereas group B and the control showed no significant differences. The ocular and systemic absorption of LMWH were significantly lower after injection of the liposome-bound than the free form. CONCLUSIONS: The subconjunctival injection of liposome-bound LMWH appears to enhance subconjunctival hemorrhage absorption in rabbits

Liposomes in tissue engineering and regenerative medicine

Liposomes are vesicular structures made of lipids that are formed in aqueous solutions. Structurally, they resemble the lipid membrane of living cells. Therefore, they have been widely investigated, since the 1960s, as models to study the cell membrane, and as carriers for protection and/or delivery of bioactive agents. They have been used in different areas of research including vaccines, imaging, applications in cosmetics and tissue engineering. Tissue engineering is defined as a strategy for promoting the regeneration of tissues for the human body. This strategy may involve the coordinated application of defined cell types with structured biomaterial scaffolds to produce living structures. To create a new tissue, based on this strategy, a controlled stimulation of cultured cells is needed, through a systematic combination of bioactive agents and mechanical signals. In this review, we highlight the potential role of liposomes as a platform for the sustained and local delivery of bioactive agents for tissue engineering and regenerative medicine approaches. liposomesscaffoldsdelivery systemsbioactive agentsstem cellsThe authors thank the Portuguese Foundation for Science and Technology for the PhD grant to N.S.M. (SFRH/BD/62465/2009), the post-doctoral grants of A.M. (SFRH/BPD/73663/2010). This study was also partly supported by POLARIS (FP7-REGPOT-2012-2013-1), RL3-TECT-NORTE-01-0124-FEDER-000020, co-financed by the North Portugal Regional Operational Programme (ON.2-O Novo Norte), under the National Strategic Reference Framework (NSRF), through the European Regional Development Fund (ERDF), the OsteoGraphy (PTDC/EME-MFE/2008) and MaxBone (PTDC/SAU-ENB/115179/2009) projects

Influence of size and geometry of nanoparticles on cellular internalization pathways

Nanoparticles can be used in biomedical disciplines as carriers for transport of diagnostic as well as therapeutic substances into cells. Variety of different shapes, sizes and different compositions are used experimentally. Despite the discoveries already made in this area, the exact nature of the interaction between a nanoparticle and a cell has not been fully understood yet. The objective of this thesis is to provide the knowledge about possibilities of utilisation and aspects influencing the interaction between the cell membrane and several types of nanoparticles: liposomes, gold nanoparticles and virus-like nanoparticles. The comparison shows that generalisation of the mechanism of nanoparticle entry into the cell is problematic, although it seems that the spherical nanoparticles with the diameter of 50 nm provide the most efficient entry.Nanočástice mohou být v biomedicínských oborech využívány jako nosiče pro dopravu diagnostických a terapeutických látek do buněk. Experimentálně se využívají nanočástice různých tvarů, velikostí a různého složení. Navzdory objevům, kterých bylo již v této oblasti dosaženo, přesná povaha interakcí nanočástic s buňkou zatím není zcela pochopena. Cílem práce je poskytnout poznatky o možnostech využití a aspektech ovlivňujících interakci s buněčnou membránou u několika typů modelových nanočástic: u liposomů, u zlatých nanočástic a u viru podobných nanočástic. Ze srovnání vyplývá, že zobecnění buněčného vstupu pro různé typy nanočástic je problematické, ač se zdá, že obecně efektivnější vstup vykazují kulaté nanočástice s průměrem 50 nm.Department of Genetics and MicrobiologyKatedra genetiky a mikrobiologieFaculty of SciencePřírodovědecká fakult