Blood-brain carrier co-culture models to study nanoparticle penetration : focus on co-culture systems

The blood-brain barrier, as a physical, active transport and metabolic barrier represents the main obstacle in the treatment of central nervous system diseases. The field of nanoparticle delivery systems is rapidly developing and nanocarriers seem to be promising for drug delivery or targeting to the brain. For testing the toxicity, uptake and transcellular transport of nanoparticles culture models of the blood-brain barrier are widely used, including immortalized brain endothelial cell lines, primary brain endothelial cells in static or dynamic culture conditions, and in co-culture systems with glial cells and/or pericytes. This mini-review gives a brief summary of blood-brain barrier co-culture models that were used for testing nanocarriers, the types of different nanoparticle systems that were examined on blood-brain barrier models, and the advantages, limitations and suitability of the blood-brain barrier models for nanoparticle penetration studies

Preparation and investigation of mefenamic acid-polyethylene glycol-sucrose ester solid dispersions

Mefenamic acid (MA) is a widely used non-steroidal anti-inflammatory (NSAID) drug. The adverse effects typical of NSAIDs are also present in the case of MA, partly due to its low water solubility. The aim of this study was to increase the water solubility of MA in order to influence its absorption and bioavailability. Solid dispersions of MA were prepared by the melting method using polyethylene glycol 6000 and different types (laurate, D-1216; palmitate, P-1670; stearate, S-1670) and amounts of sucrose esters as carriers. The X-ray diffraction results show that MA crystals were not present in the products. Dissolution tests carried out in artificial intestinal juice showed that the product containing 10 % D-1216 increased water solubility about 3 times. The apparent permeability coefficient of MA across human Caco-2 intestinal epithelial cell layers was high and, despite the difference in solubility, there was no further increase in drug penetration in the presence of the applied additives

Design and Optimization of Nanostructured Lipid Carrier Containing Dexamethasone for Ophthalmic Use

The aim of this study was to perform a preformulation study of dexamethasone (DXM)-loaded nanostructured lipid carriers (NLCs) for ocular use. Lipid screening was applied to find the most suitable solid and liquid lipids and surfactant for the NLC formulation. The visual observation was proved with XRD measurements for the establishment of the soluble state of DXM. Thermoanalytical measurements indicated that the most relevant depression of the crystallinity index could be ensured when using a 7:3 solid lipid:oil ratio. In order to optimize the NLC composition, a 23 full factorial experimental design was used. It was established that each independent factor (lipid, DXM, and surfactant concentration) had a significant effect on the particle size while in the case of entrapment efficiency, the DXM and surfactant concentrations were significant. Lower surfactant and lipid concentrations could be beneficial because the stability and the entrapment efficacy of NLCs were more favorable. The toxicity tests on human cornea cells indicated good ophthalmic tolerability of NLCs. The in vitro drug release study predicted a higher concentration of the solute DXM on the eye surface while the Raman mapping penetration study on the porcine cornea showed a high concentration of nanocarriers in the hydrophylic stroma layer

NANOPARTICLES: TOXICITY AND PENETRATION ACROSS BIOLOGICAL BARRIERS

Nanoparticles provide new opportunities for drug delivery and human therapy. To fulfill the therapeutical potential of nanoparticles two major aspects, toxicity and penetration across barriers of the body need to be studied. Different ex vivo and in vitro cell culture based models of the skin, nasal, lung, intestinal and blood-brain barriers have been established in our laboratory that can be used for both purposes. Three different types of nanoparticles were tested on the different models. Amorphous nanoparticles from the antiinflammatory drug meloxicam were obtained by by co-grinding with polyvinylpyrrolidone. Nanosized bilayered vesicles of non-ionic surfactants bearing glucose and amino acid ligands were prepared to specifically target solute carriers on the blood-brain barrier [1]. Poly(ferrocenyl silane) redox responsive polymer nanocarriers were also studied [2]. Several methods were applied parallelly to measure the toxicity of nanoparticles. In addition to colorimetric tests like MTT dye reduction assay, release of the cytoplasmic enzyme lactate dehydrogenase cellular events were also monitored in real time. By measuring impedance across microelectrodes covered with cells quantitative information on cell viability and intercellular adherence indicating paracellular permeability could be obtained. Co-culture models of the barriers prepared from primary cultures or human cell lines [3] served for permeability experiments to test the penetration of nanocarriers across cell layers. In the case of the blood-brain barrier a kinetic in vivo study in mice was also performed by near infrared fluorescence time-domain optical imaging. The results indicate that (i) toxicity measurements are very important to obtain the optimal dose of nanoparticles on living cells, (ii) nanonization of drugs can improve drug dissolution, absorption and pharmacokinetics, (iii) targeting of microvesicles increases their penetration across barriers

Identification of Reference Genes for Circadian Studies on Brain Microvessels and Choroid Plexus Samples Isolated from Rats

Delivery of putative compounds of therapeutic value to the brain is limited by brain barriers: the blood–brain barrier located in the endothelium of the brain microvessels (BrMVs) and the blood–cerebrospinal fluid barrier located in the epithelium of the choroid plexus (ChP). Understanding their function and modulation by the circadian clock may enhance the efficacy of brain-targeting therapies. The aim of the present study was to evaluate the stability of 10 reference genes in the BrMV and ChP, isolated from male and female rats at six time points (ZT1, 5, 9, 13, 17, and 21). Gene evaluations were performed by qPCR, analyzed by RefFinder tool, and verified by analyzing the expression of the brain and muscle ARNT-like 1 (Bmal1) using the qPCR and digital PCR methods. We identified as the most stable genes for circadian studies tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein zeta (Ywhaz) and apolipoprotein E (Apoe) for BrMV, and beta actin (Actb) and hypoxanthine-guanine phosphoribosyltransferase (Hprt1) for ChP. After verification, ribosomal protein (Rps18) was also included as a sufficient reference gene. Additionally, the observed gender difference in the Bmal1 oscillations in both BrMV and ChP suggests that separate studies for each gender are recommended

Protection of cultured brain endothelial cells from cytokine-induced damage by α-melanocyte stimulating hormone

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Pitavastatin Ameliorates Lipopolysaccharide-Induced Blood-Brain Barrier Dysfunction

Statins have neuroprotective effects on neurological diseases, including a pleiotropic effect possibly related to blood-brain barrier (BBB) function. In this study, we investigated the effects of pitavastatin (PTV) on lipopolysaccharide (LPS)-induced BBB dysfunction in an in vitro BBB model comprising cocultured primary mouse brain endothelial cells, pericytes, and astrocytes. LPS (1 ng/mL, 24 h) increased the permeability and lowered the transendothelial electrical resistance of the BBB, and the co-administration of PTV prevented these effects. LPS increased the release of interleukin-6, granulocyte colony-stimulating factor, keratinocyte-derived chemokine, monocyte chemotactic protein-1, and regulated on activation, normal T-cell expressed and secreted from the BBB model. PTV inhibited the LPS-induced release of these cytokines. These results suggest that PTV can ameliorate LPS-induced BBB dysfunction, and these effects might be mediated through the inhibition of LPS-induced cytokine production. Clinically, therapeutic approaches using statins combined with novel strategies need to be designed. Our present finding sheds light on the pharmacological significance of statins in the treatment of central nervous system diseases

Application of a 3D-printed, high efficiency sample introduction system for single cell ICP-MS analysis

In this study, the application of an alternative, 3D-printed sample introduction system (SIS) is presented for single cell ICP-MS analysis. The SIS utilized in the experiments was designed by our group with the purpose to enhance single particle detection capabilities. It is able to provide higher transport efficiencies, increased sensitivity, lower background signals and a wider range of detectable particle size in single particle ICP-MS (spICP-MS) analysis compared to the standard SIS. Capitalizing on these features, two single cell ICP-MS (scICP-MS) experiments were done, using single cellular algae (Chlorella) and human endothelial cell suspensions, thereby demonstrating the SIS’s ability to facilitate cell biology related studies such as determining the elemental composition/metal uptake of a single cell or monitoring its cell-tocell variation in cell cultures

Design, Optimization, and Application of a 3D-Printed Polymer Sample Introduction System for the ICP-MS Analysis of Nanoparticles and Cells

Commonly used sample introduction systems for inductively coupled plasma mass spectrometry (ICP-MS) are generally not well-suited for single particle ICP-MS (spICP-MS) applications due to their high sample requirements and low efficiency. In this study, the first completely 3D-printed, polymer SIS was developed to facilitate spICP-MS analysis. The system is based on a microconcentric pneumatic nebulizer and a single-pass spray chamber with an additional sheath gas flow to further facilitate the transport of larger droplets or particles. The geometry of the system was optimized using numerical simulations. Its aerosol characteristics and operational conditions were studied via optical particle counting and a course of spICP-MS measurements, involving nanodispersions and cell suspensions. In a comparison of the performance of the new and the standard (quartz microconcentric nebulizer plus a double-pass spray chamber) systems, it was found that the new sample introduction system has four times higher particle detection efficiency, significantly better signal-to-noise ratio, provides ca. 20% lower size detection limit, and allows an extension of the upper limit of transportable particle diameters to about 25 µm