Calcification of polyurethane-based biomaterials implanted subcutaneously in rats: role of porosity and fluid absorption in the mechanism of mineralization

The object of this study was to investigate the mineralization of polyurethane in the rat subcutaneous environment, in comparison to collageneous biomaterials, which would facilitate investigation of the calcification mechanism in implantable biomaterials. This model was compared to anin vitro model of non-spontaneous formation of calcium phosphate precipitate. To study the role of porosity/fluid absorption capacity, various biomaterials such as polyurethane, hydrophilic polyurethane foam, Type I collagen sponges, and bioprosthetic heart valve tissue were examined inin vitro andin vivo models. In view of the exceedingly low levels of polyurethane calcification, it seems that the rat subcutaneous model is not suitable for routine studies of the mechanism of polyurethane calcification. The calcification extent of both collageneous and hydrophilic polyurethane, was found to be in good correlation with the water absorption capacity of these biomaterials. It is suggested that the water capacity of the biomaterial determines the bulk level of calcification, which in turn is generated and propagates via the inherent affinity sites of the biomaterial to Ca²⁺.     

Morphology and mechanical behavior of TTCP-derived calcium phosphate cement subcutaneously implanted in rats

A pre-hardened, TTCP-derived CPC was immersed in Hanks’ solution as well as subcutaneously implanted into abdomen of rats. The implant-soft tissue interfacial morphology was examined and properties of the CPC were evaluated and compared under in vitro and in vivo conditions. The results indicate that the surface of immersed samples appeared rougher and more porous than that of implanted samples and was covered with a layer of fine apatite crystals. The CPC samples implanted for 4 weeks or longer were surrounded by a layer of fibrous tissue, which was further surrounded by a soft tissue capsule comprising numerous fat cells. The soft tissue capsule had a non-uniform distribution in thickness, which increased most significantly between 4 weeks and 12 weeks after implantation. None of polymorphic cells, osteoblast cells or bone cells adjacent to the implant were observed. The majority of original TTCP powder was transformed into apatite after 1 day of either immersion in Hanks’ solution or implantation. The average porosity values of samples immersed in Hanks’ solution for 4 weeks or longer were significantly larger than those immersed for 1 day or 1 week. The porosity values of samples implanted for different times were not significantly different. The DTS values of Hanks’ solution-immersed samples largely decreased after a few weeks of immersion. The implanted samples maintained their strengths throughout the study.     

Biocompatible transferrin-conjugated sodium hexametaphosphate-stabilized gold nanoparticles: synthesis, characterization, cytotoxicity and cellular uptake

The feasibility of using gold nanoparticles (AuNPs) for biomedical applications has led to considerable interest in the development of novel synthetic protocols and surface modification strategies for AuNPs to produce biocompatible molecular probes. This investigation is, to our knowledge, the first to elucidate the synthesis and characterization of sodium hexametaphosphate (HMP)-stabilized gold nanoparticles (Au-HMP) in an aqueous medium. The role of HMP, a food additive, as a polymeric stabilizing and protecting agent for AuNPs is elucidated. The surface modification of Au-HMP nanoparticles was carried out using polyethylene glycol and transferrin to produce molecular probes for possible clinical applications. In vitro cell viability studies performed using as-synthesized Au-HMP nanoparticles and their surface-modified counterparts reveal the biocompatibility of the nanoparticles. The transferrin-conjugated nanoparticles have significantly higher cellular uptake in J5 cells (liver cancer cells) than control cells (oral mucosa fibroblast cells), as determined by inductively coupled plasma mass spectrometry. This study demonstrates the possibility of using an inexpensive and non-toxic food additive, HMP, as a stabilizer in the large-scale generation of biocompatible and monodispersed AuNPs, which may have future diagnostic and therapeutic applications.     

Size,shape and surface chemistry of nano-gold dictate its cellular interactions,uptake and toxicity

Colloidal gold is undoubtedly one of the most extensively studied nanomaterials, with 1000s of different protocols currently available to synthesise gold nanoparticles (AuNPs). While developments in the synthesis of AuNPs have progressed rapidly in recent years, our understanding of their biological impact, with particular respect to the effect of shape, size, surface characteristics and aggregation states, has struggled to keep pace. It is generally agreed that when AuNPs are exposed to biological systems, these parameters directly influence their pharmacokinetic and pharmacodynamic properties by influencing AuNPs distribution, circulation time, metabolism and excretion in biological systems. However, the rules governing these properties, and the science behind them, are poorly understood. Therefore, a systematic understanding of the implications of these variables at the nano–bio interface has recently become a topic of major interest. This Review Article attempts to ignite a discussion around the influence of different physico-chemical parameters on biological activity of AuNPs, while focussing on critical aspects of cellular interactions, uptake and cytotoxicity. The review also discusses emerging trends in AuNP uptake and toxicity that are leading to technological advances through AuNP-based therapy, diagnostics and imaging.     

Quantitative measurements of protein interactions in a crowded cellular environment

Quantitative measurements of protein interaction strengths, crucial for describing signaling networks and predicting cellular responses to environmental stimuli, are typically performed in dilute buffer solutions. However, protein-protein interactions in cells occur within the context of a crowded system, which is characterized by a high macromolecular concentration. In this paper, we explore the utility of cell-derived vesicles as a model crowded environment for quantitative FRET measurements of protein-protein interactions. We show that the FRET efficiency, and the donor and acceptor concentrations, can be calculated in each vesicle. We also introduce the "quantitative imaging Foster resonance energy transfer" method as a tool that can yield protein interaction strengths within these vesicles.     

Intracellular delivery of etoposide loaded biodegradable nanoparticles: cytotoxicity and cellular uptake studies

The preferred delivery systems for anticancer drugs would be the one which would have selective and effective destruction of cancer cells. In the present study etoposide (ETO) loaded nanoparticles (NP) were prepared using PLGA (ETO-PLGA NP), PLGA-MPEG block copolymer (ETO-PLGA-MPEG NP) and PLGA-Pluronic copolymer (ETO-PLGA-PLU NP) and they were evaluated for cytotoxicity and cellular uptake studies using two cancer cell lines, L1210 and DU145. The IC50 values for L1210 cells were 18.0, 6.2, 4.8 and 5.4 microM and for DU145 cells the IC50 values were 98.4, 75.1, 60.1 and 71.3 microM for ETO, ETO-PLGA NP, ETO-PLGA-MPEG NP and ETO-PLGA-PLU NP respectively. The increased cytotoxicities were attributed to increased uptake of the NPs by the cells. Moreover the ETO loaded PLGA-MPEG NP and PLGA-Pluronic NP showed a sustained cytotoxic effect till 5 days on both the cell lines. Results of the long term cytotoxicity study concluded that the drug loaded PLGA nanoparticulate formulations were efficient in decreasing the viability of the L1210 cells over a period of three days, whereas the pure drug exerted its maximum efficiency on the day one itself. Z-stack confocal images of NPs showed fluorescence activity in each section of DU 145 and L1210 cells indicating that the nanoparticles were internalized by the cells. The study concluded that ETO loaded PLGA NPs had higher cytotoxicity compared with that of the free drug and ETO-PLGA-MPEG NP and ETO-PLGA-PLU NP had higher cell uptake efficiency compared with that of ETO-PLGA NP. The developed PLGA based NPs shows promise to be used for cancer therapy.     

Development of docetaxel nanocapsules for improving in vitro cytotoxicity and cellular uptake in MCF-7 cells

The aim of this study was to fabricate docetaxel loaded nanocapsules (DTX-NCs) with a high payload using Layer-by-Layer (LbL) technique by successive coating with alternate layers of oppositely charged polyelectrolytes. Developed nanocapsules (NCs) were characterized in terms of morphology, particle size distribution, zeta potential (ζ-potential), entrapment efficiency and in vitro release. The morphological characteristics of the NCs were assessed using transmission electron microscopy (TEM) that revealed coating of polyelectrolytes around the surface of particles. The developed NCs successfully attained a submicron particle size while the ζ-potential of optimized NCs alternated between (+) 34.64?±?1.5 mV to (?) 33.25?±?2.1 mV with each coating step. The non-hemolytic potential of the NCs indicated the suitability of the developed formulation for intravenous administration. A comparative study indicated that the cytotoxicity of positively charged NCs (F4) was significant higher (p?in vitro on MCF-7 cells. Furthermore, cell uptake studies evidenced a higher uptake of positive NCs (≥1.2 fold) in comparison to negative NCs. In conclusion, formulated NCs are an ideal vehicle for passive targeting of drugs to tumor cells that may result in improved efficacy and reduced toxicity of encapsulated drug moiety.     

Micropatterned co-culture of hepatocyte spheroids layered on non-parenchymal cells to understand heterotypic cellular interactions

Abstract

Microfabrication and micropatterning techniques in tissue engineering offer great potential for creating and controlling cellular microenvironments including cell–matrix interactions, soluble stimuli and cell–cell interactions. Here, we present a novel approach to generate layered patterning of hepatocyte spheroids on micropatterned non-parenchymal feeder cells using microfabricated poly(ethylene glycol) (PEG) hydrogels. Micropatterned PEG-hydrogel-treated substrates with two-dimensional arrays of gelatin circular domains (φ = 100 μm) were prepared by photolithographic method. Only on the critical structure of PEG hydrogel with perfect protein rejection, hepatocytes were co-cultured with non-parenchymal cells to be led to enhanced hepatocyte functions. Then, we investigated the mechanism of the functional enhancement in co-culture with respect to the contributions of soluble factors and direct cell–cell interactions. In particular, to elucidate the influence of soluble factors on hepatocyte function, hepatocyte spheroids underlaid with fibroblasts (NIH/3T3 mouse fibroblasts) or endothelial cells (BAECs: bovine aortic endothelial cells) were compared with physically separated co-culture of hepatocyte monospheroids with NIH3T3 or BAEC using trans-well culture systems. Our results suggested that direct heterotypic cell-to-cell contact and soluble factors, both of these between hepatocytes and fibroblasts, significantly enhanced hepatocyte functions. In contrast, direct heterotypic cell-to-cell contact between hepatocytes and endothelial cells only contributed to enhance hepatocyte functions. This patterning technique can be a useful experimental tool for applications in basic science, drug screening and tissue engineering, as well as in the design of artificial liver devices.     

Evaluation of systemic exposure of nanoparticle suspensions subcutaneously administered to mice regarding stabilization,volume, location,concentration and size

Different routes of administration are likely to result in very different outcomes due to different availability or plasma profile. The objective of the present study was to evaluate the pharmacokinetic profile after different subcutaneous (s.c.) administration of nanoparticle suspensions of a lipophilic compound to mice. Pharmacokinetics of the selected test compound and the effect of drug concentration, particle size, location of administration, volume given and particle stabilizers were studied. Adding PEGylated lipids or pluronic F-127 to the negatively charged surface of the nanoparticles increased the stability of the particles and the bioavailability. The in vivo studies demonstrated linear absorption kinetics for the selected model compound up to at least 500?µmol/kg. Absorption from upper-neck resulted in different systemic exposure compared to administration in the hip. The former was preferred if a prolonged C_max was desired while the latter ensured a flat profile for approximately 24 hours. Administering the double volume (but the same dose) had no effect on the pharmacokinetics, whereas smaller particle size significantly increased the exposure.     

Mesenchymal stem cellular adhesion and cytotoxicity study of random biopolyester scaffolds for tissue engineering

Mesenchymal stem cells (MSCs) were isolated from the bone marrow of rabbits and inoculated respectively on 3D scaffolds of poly(3-Hydroxybutyrate-co-3-Hydroxyvalerate) (PHBV), poly(butylenes succinate) (PBS) and different blends (100/0, 80/20, 50/50, 20/80, 0/100) (Wt%) in vitro. It was found that the (50/50) blends possessed the best performance on adhesion and cytotoxicity of MSCs. The scanning electronic microscopy (SEM) results showed that the (50/50) blends had the appropriate roughness for MSCs to attach and grow, which may be used as a suitable biomaterial to create small caliber vascular grafts.     

Nanostructured lipid carriers loaded with simvastatin: effect of PEG/glycerides on characterization,stability, cellular uptake efficiency and in vitro cytotoxicity

Objective: The aim of this study is to evaluate the use of PEG/glycerides of different HLB; oleoyl macrogol-6-glycerides (Labrafil^® M 1944 CS) and caprylocaproylmacrogol-8-glycerides (Labrasol^®), compared to Labrafac lipophile^® as PEG-free glyceride in the preparation of nanostructured lipid carriers (NLCs). PEG/glycerides are suggested to perform a dual function; as the oily component, and as the PEG-containing substrate required for producing the PEGylated carriers without physical or chemical synthesis.

Methods: Lipid nanocarriers were loaded with simvastatin (SV) as a promising anticancer drug. An optimization study of NLC fabrication variables was first conducted. The effect of lyophilization was investigated using cryoprotectants of various types and concentrations. The prepared NLCs were characterized in terms of particle size (PS), size distribution (PDI), zeta potential (ZP), drug entrapment, in vitro drug release, morphology and drug–excipient interactions. The influence of glycerides?±?PEG on the cytotoxicity of SV was evaluated on MCF-7 breast cancer cells, in addition to the cellular uptake of fluorescent blank NLCs.

Results: The alteration between different oil types had a significant impact on PS, ZP and drug release. Both sucrose and trehalose showed the lowest increase in PS and PDI of the reconstituted lyophilized NLCs. The in vitro cytotoxicity and cellular uptake studies indicated that SV showed the highest antitumor effect on MCF-7 cancer cells when loaded into Labrasol^® NLCs demonstrating a high cellular uptake as well.

Conclusion: The study confirms the applicability of PEG/glycerides in the development of NLCs. Encapsulating SV in Labrasol^®-containing NLC could enhance the antitumor effect of the drug.     

Effect of surface charge on the cellular uptake and cytotoxicity of fluorescent labeled cellulose nanocrystals

Probing of cellular uptake and cytotoxicity was conducted for two fluorescent cellulose nanocrystals (CNCs): CNC-fluorescein isothiocyanate (FITC) and newly synthesized CNC-rhodamine B isothiocyanate (RBITC). The positively charged CNC-RBITC was uptaken by human embryonic kidney 293 (HEK 293) and Spodoptera frugiperda (Sf9) cells without affecting the cell membrane integrity. The cell viability assay and cell-based impedance spectroscopy revealed no noticeably cytotoxic effect of the CNC-RBITC conjugate. However, no significant internalization of negatively charged CNC-FITC was observed at physiological pH. Indeed, the effector cells were surrounded by CNC-FITC, leading to eventual cell rupture. As the surface charge of CNC played an important role in cellular uptake and cytotoxicity, facile surface functionalization together with observed noncytotoxicity rendered modified CNC as a promising candidate for bioimaging and drug delivery systems.     

Sigma-2 receptor ligand anchored telmisartan loaded nanostructured lipid particles augmented drug delivery,cytotoxicity, apoptosis and cellular uptake in prostate cancer cells

Recently, the anticancer activity of telmisartan (TEL) has been discovered against prostate cancer. Nevertheless, despite favorable therapeutic profile, poor aqueous solubility and suboptimal oral bioavailability hamper the anticancer efficacy of TEL. Therefore, in this investigation, sigma-2 receptor ligand, 3-(4-cyclohexylpiperazine-1-yl) propyl amine (CPPA) anchored nanostructured lipid particles of telmisartan (CPPA-TEL-NLPs) were engineered using stearic acid for targeting prostate cancer, PC-3 cells. The mean particle size of TEL-NLPs was measured to be 25.4?±?3.2?nm, significantly (p?p?p?In vitro drug release study was conducted to determine the drug delivery potential of tailored nanoparticles. TEL-NLPs released 93.36% of drug significantly (p?50 of CPPA-TEL-NLPs was measured to be 20.3?µM significantly (p?50 of 41.3?µM, significantly (p?>?0.05) not different from 43.4?µM, exhibited by TEL-NLPs in PNT-2 cells. We elucidated that CPPA-TEL-NLPs entered the PC-3 cells via receptor mediated endocytosis pathway and thus exhibited superior cytotoxicity, apoptosis and greater extent of cellular uptake in PC-3 cells. In conclusion, CPPA-TEL-NLPs may be a promising nanomedicine and warrant further in vivo investigations for gaining clinical success.     

Nano- and microcrystals of griseofulvin subcutaneously administered to rats resulted in improved bioavailability and sustained release

Griseofulvin is a commonly used antifungal agent which is administered per oral (p.o.). The oral administration route, however, shows rather low bioavailability. The aim of this study was to improve the bioavailability and to evaluate and interpret the pharmacokinetic profiles after subcutaneous (s.c.) administration of crystalline griseofulvin nano- and microsuspensions. Both formulations were injected at 5 and 500 µmol/kg to rats. For the lower concentration, the profiles were similar after s.c. injection but extended as compared to p.o. administration. For the higher concentration, injection of microsuspension resulted in a maintained plateau whereas the nanosuspension resulted in an obvious peak exposure followed by extended elimination. Both suspensions showed improved exposure with dose. The differences in peak exposures between nano- and microparticles, at the high dose, were mainly ascribed to differences in dissolution rate, experimentally determined in vitro, using spectroscopic methods. The extended appearance in the circulation may depend on the physicochemical properties of the compound and the physiological conditions at the injection site. The bioavailability was improved for both formulations compared with an orally administered nanosuspension, suggesting the s.c. route to be a preferred administration option for compounds with low oral bioavailability regarding both overall exposure and extended efficacy.     

Preparation and evaluation of charged solid lipid nanoparticles of tetrandrine for ocular drug delivery system: pharmacokinetics,cytotoxicity and cellular uptake studies

In this study, tetrandrine-loaded cationic solid lipid nanoparticles (TET-CNP) and solid lipid nanoparticles (TET-NP) were prepared by the emulsion evaporation-solidification at low temperature method. The particle size, zeta potential, and entrapment efficiency of TET-CNP and TET-NP were characterized. The results showed that the TET-CNP and TET-NP had average diameters of (15.29?±?1.34) nm and (18.77?±?1.23) nm with zeta potentials of (5.11?±?1.03) mV and (?8.71?±??1.23) mV and entrapment efficiencies of (94.1?±?2.37)% and (95.6?±?2.43)%, respectively. In vitro release studies indicated that the TET-CNP and TET-NP retained the drug entity better than tetrandrine ophthalmic solutions (TET-SOL). In the pharmacokinetics studies, the AUC values of TET-CNP and TET-NP were 1.96-fold and 2.00-fold higher than that of TET-SOL (?p?C_max values of TET-CNP and TET-NP were 2.45-fold and 2.53-fold higher than that of the TET-SOL (p?相似文献   

Preparation,characterization and testing of implanted 37Ar-layers

³⁷Ar atoms were produced via the ³⁷Cl(p, n)³⁷Ar reaction using the 30 MeV proton beam of the Cyclone 30 cyclotron of the UCL and a NaCl target. After ionization, ³⁷Ar²⁺ ions were implanted at 20 keV into 20 μm thick Al foils. The number of ³⁷Ar atoms in the layers was determined by measurement of the 2.62 keV KX-rays (emitted during the EC-decay of ³⁷Ar) with a gas-flow proportional counter. Under vacuum and under intense thermal neutron irradiation, the implantation was found to be stable. Furthermore, the layers were found to be suited for (n, p) and (n, α) measurements.     

Response to polyetherimide based composite materials implanted in muscle and in bone

The in-vivo response to a composite material obtained with polyetherimide (PEI) reinforced with carbon/glass fibers was investigated by histological methods by implanting cylinders in muscle and in bone of the New Zealand White rabbit. A common metallic alloy, widely used in orthopaedic surgery, was used as control (Stellite). The aim of the study was to analyze the biological response towards the surface of the material. Composite implants and metallic implants did not induce adverse or inflammatory reactions. The morphological picture produced was similar, in muscle and in bone, for both materials. In muscle, cylinders were confined by an extremely thin fibrous layer and the overall appearance of the muscular tissue was normal. In bone, cylinders were confined by a nearly annular rim of newly formed bone. From these data it is possible to derive that the response to PEI-based composite material is comparable with the response to metallic substrate and, then, the material can be suitable for clinical application. ©1999 Kluwer Academic Publishers     

Magnetism in GaN layers implanted by La, Gd, Dy and Lu

We present a complex study of rare earth elements implanted GaN layers grown by low pressure metal-organic vapor phase epitaxy on c-plane sapphire substrates. Gd, Dy, La and Lu ions were implanted with energies of 200 keV and doses ranging from 5 × 10¹³ to 4 × 10¹⁷ atoms.cm^− 2. The chemical composition and concentration profiles of ion-implanted layers were studied by secondary ion mass spectrometry and Rutherford back scattering. The structural properties of the layers were characterized by Rutherford back scattering/channeling and X-ray diffraction reciprocal space mapping. Gd implanted layers exhibit ferromagnetic behavior persisting up to ~ 720 K. Since the ferromagnetic behavior was not observed in the case of La and Lu implanted layers, it cannot be attributed to the structural damage of the layer. Based on the fact that the samples are electrically conducting we conclude that the ferromagnetism can be associated with doped electrons mediating the ferromagnetic interaction between local moments on Gd and Dy.