Adaptation of the disector method to rare small organelles in TEM sections exemplified by counting synaptic bodies in the rat pineal gland

The disector is the only objective method for quantifying particles of variable size in a given volume. With this method, cell organelles are identified on adjacent sections, but only those present in one section are counted. When counting extremely rare structures in transmission electron microscope sections (physical disector), the usual procedure of counting on electron micrographs is limited for economic reasons (e.g. micrographs highly outnumbering the investigated structures). Hence, to apply this unbiased stereological method, a modification of the physical disector concerning 3 aspects has been developed. (1) The prerequisite of screening large corresponding tissue areas (here ∼65000 μm²) was fulfilled by examining tissue areas along the edges of ultrathin sections. (2) The size of the counting frame was determined by measuring the lengths of the section margins (minus a guard area) by means of a Morphomat. This value was multiplied by the width of the investigated tissue zone, corresponding to the diameter of the electron microscope viewing screen. (3) Disector counting was carried out simultaneously on both sections (bidirectional disector) to improve efficiency. In the present study tiny synaptic bodies (SBs) were quantitated by disector in a rat pineal gland, yielding ∼30 SBs/1000 μm³. By contrast, single section profile counts of SBs amounted to 90 SBs/20000 μm². Since the presently described adaptation of the disector is time-consuming, it is proposed to determine a proportion factor allowing to estimate number of structures per volume based on single section profile counts. This would decrease the evaluation time by more than 50%.     

Repeated immunization with plasmid DNA formulated in poly(lactide-co-glycolide) microparticles is well tolerated and stimulates durable T cell responses to the tumor-associated antigen cytochrome P450 1B1

Injection of microparticle-encapsulated DNA elicits immune responses to plasmid-encoded antigens in mice and humans. Cytochrome P450 CYP1B1 (CYP1B1) is a member of the CYP1 P450 enzyme family that is overexpressed in a variety of solid tumors. The work described herein was performed to study the kinetics of stimulating T cell responsiveness with an encapsulated DNA encoding CYP1B1 and provides support for the clinical development of this formulation. Immunization of HLA-A2/Kb transgenic mice with human CYP1B1 encoding plasmid DNA formulated in poly(lactide-co-glycolide) (PLG) microparticles elicits CD8+ T cells that respond to human CYP1B1-positive target cells. The duration of the immune response, the effect on the immune response of multiple injections, and the safety of repeated injections were studied. These results show that the PLG-encapsulated DNA therapeutic elicits durable immune responses to CYP1B1, the responses are dependent on repeat immunization, and that the formulation is well tolerated.     

Size-Controllable Synthesis of Monodisperse Magnetite Microparticles Leading to Magnetically Tunable Colloidal Crystals

Colloidal crystals (CCs) are periodic arrays of monodisperse microparticles. Such CCs are very attractive as they can be potentially applicable as versatile photonic devices such as reflective displays, sensors, lasers, and so forth. In this article, we describe a promising methodology for synthesizing monodisperse magnetite microparticles whose diameters are controllable in the range of 100–200 nm only by adjusting the base concentration of the reaction solution. Moreover, monodisperse magnetite microparticles in aqueous suspensions spontaneously form the CC structures under an external magnetic field, leading to the appearance of Bragg reflection colors. The reflection peak can be blue-shifted from 730 nm to 570 nm by the increase in the external magnetic field from 28 mT to 220 mT. Moreover, the reflection properties of CCs in suspension depend on the microparticle concentration in suspension and the diameter of the magnetite microparticles. Both fine-control of microparticle diameter and investigation of magneto-optical properties of CCs would contribute to the technological developments in full-color reflective displays and sensors by utilizing these monodisperse magnetite microparticles.     

Pancreas fat content,insulin homeostasis and circulating endothelial microparticles in male essential hypertensive patients

The pancreas fat content has been poorly investigated in essential hypertension. The authors aim to relate pancreas and liver fat content with parameters measuring insulin resistance, beta‐cell function and also with markers of endothelial dysfunction and platelet or endothelial cell destruction. The authors studied a group of 40 male hypertensive patients with well‐controlled blood pressure, maintaining a stable weight, and having not changed their medication during the last year. Pancreas fat content was correlated with HOMA‐IR (r = .616, p < .001), HOMA‐S (r = −.439, p < .005), beta cell function parameter (r = .457, p < .005), and QUICKI (r = .412, p < .01), whereas liver fat was not patients in the highest quartile of pancreas fat content had more circulating endothelial microparticles than patients in the other quartiles (median 129 [94.3–200] vs. 60.9 [49.4–88.8], p = .002). However, patients in the highest quartile of the pancreas fat content distribution did not differ from the lowest in hyperemic response after ischemia nor circulating platelet microparticles count. Liver fat content was not related to any of the parameters studied. In a multivariate stepwise binary logistic regression analysis (Wald Method) circulating endothelial microparticles remain significantly associated with pancreas fat content after adjusting for confounding factors, such as tobacco, diabetes mellitus, hypercholesterolemia, or metabolic syndrome. Our results reflect that in essential hypertension, pancreas fat content is superior to liver fat to study beta‐cell functionality and insulin resistance. Moreover, the authors described for the first time that pancreas fat content is related to endothelial cell destruction. Further studies are needed to confirm this point.     

Corrosion Protection of Q235 Steel Using Epoxy Coatings Loaded with Calcium Carbonate Microparticles Modified by Sodium Lignosulfonate in Simulated Concrete Pore Solutions

In this study, calcium carbonate (CaCO₃) microparticles having pH-sensitive properties were loaded with sodium lignosulfonate (SLS), a corrosion inhibitor. Scanning electron microscope (SEM), UV–VIS spectrophotometer (UV-vis), X-ray diffraction (XRD), and attenuated total reflection-Fourier-transform infrared spectroscopy (ATR-FTIR) were applied to evaluate the properties of the synthetic microparticles. This material could lead to the release of corrosion inhibitor under different pH conditions of the aqueous media. However, the extent of release of the corrosion inhibitor in the acidic media was higher, leading to enhanced shielding effect of the Q235 steel. These microparticles can serve as anti-corrosion additive for epoxy resin-coated Q235 steel. Electrochemical experiments were used to assess the anti-corrosive ability of the epoxy coatings in simulated concrete pore (SCP) solution, confirming the superior corrosion inhibition of the epoxy coating via incorporation of 5 wt % calcium carbonate microparticles loaded with SLS (SLS/CaCO₃). The physical properties of coating specimens were characterized by water absorption, contact angle, adhesion, and pencil hardness mechanical tests.     

Imbalance between the circulating endothelium-derived apoptotic microparticles and the endothelial colony-forming units of progenitor cells in patients undergoing diagnostic coronary angiography

PurposeThe involvement of the circulating endothelium-derived microparticles (EMPs) and the endothelial progenitor cells (EPCs) has been shown in the pathogenesis of coronary artery disease (CAD). The current study aimed to explore whether the Friesinger index is associated with the levels of the apoptotic CD144+/CD31+/annexin V+ ​EMPs and the number of endothelial colony-forming units of progenitor cells in patients undergoing coronary angiography.Patients and methodsFifty-seven patients with a median age of 62 years (range: 48–84 years) were enrolled. Quantification of the apoptotic CD144⁺/CD31⁺/annexin V⁺ EMPs was performed by flow cytometry. The number of endothelial colony-forming units defined by CFU-Hill was assessed by cell culture.ResultsThere was a positive correlation between the Friesinger index and the circulating levels of the apoptotic CD144⁺/CD31⁺/annexin V⁺ EMPs (rho=0.817, p<0.001), whereas a negative correlation was found with the number of CFU-Hill (rho ​= ​− 0.649, p<0.001). Multivariable logistic analysis showed that the risk of having moderate/severe CAD was five times greater among male patients (OR:5.32; 95% CI: 1.19 - 16.33; p=0.038) and almost one and a half times higher among those with a higher level of apoptotic CD144⁺/CD31⁺/annexin V⁺ EMPs (OR:1.74; 95% CI: 1.23 - 2.28; p=0.001). Finally, the circulating levels of apoptotic EMPs labelled for CD144⁺/CD31⁺/annexin V⁺ presented a good discrimination of moderate/severe CAD, with an AUC of 0.85 (95% CI ​= ​0.74 - 0.96; p< 0.001).ConclusionsModerate or severe CAD is associated with increased levels of apoptotic EMPs and reduced EPC colony-forming capacity, increasing the occurrence of endothelial injuries.     

Solid lipid microparticles containing loratadine prepared using a Micromixer

Solid lipid microparticles were investigated as a taste-masking approach for a lipophilic weak base in a suspension. The idea was that the drug concentration in the aqueous phase of a suspension might be reduced by its partitioning into the solid lipid particles. Loratadine, as a model drug, was used to prepare Precirol® ATO 5 microparticles by a Micromixer. The effects of three process variables: drug loading, PVA concentration and water/lipid ratio on the microparticle size, encapsulation efficiency, surface appearance, in-vitro release and drug partitioning in a suspension were studied. Loratadine release was slow in simulated saliva and very fast at the pH of stomach. In suspension of loratadine lipid microparticles, drug was released into the aqueous phase to the same concentration as in a drug suspension. Therefore, the usefulness of these microparticles for taste-masking in liquids is limited. However, they might be useful for taste-masking in solid dosage forms.     

Non-invasive endotracheal delivery of paclitaxel-loaded alginate microparticles

Aerosolized chemotherapeutics leads to higher, localized and continuous concentrations of active agents in lung tissue with lower side effects for other organs. The present study was performed on jugular vein cannulated rats which endothracheally received 4 mg/kg of free paclitaxel powder (Free-PTX), paclitaxel-loaded alginate microparticles (PTX-ALG-MPs) and i.v. paclitaxel (Anzatax^®). Pharmacokinetic parameters for Free-PTX and PTX-ALG-MPs contain higher AUC, mean residence time (MRT),half-life and bioavailability, with lower elimination constant (k_e). Statistical analysis showed that the amount of paclitaxel per gram of lung tissue after 0.5, 6 and 24 h after administration of Free-PTX was lower than PTX-ALG-MPs. Lung tissue AUC for Free-PTX was lower than PTX-ALG-MPs. According to the obvious advantages obtained, such as dose lowering and increasing paclitaxel residence time and half-life. It should be noted that cell cytotoxicity test on normal airway cell lines was not examined in this study but due to previous reports on safety of inhaled paclitaxel, it can be suggested that pulmonary delivery of paclitaxel can be a useful non-invasive route of administration compared with i.v administration.     

Formulation and evaluation of novel tableted chitosan microparticles for the controlled release of clozapine

Controlled release formulations of clozapine microparticulated tablets were prepared by using chitosan. Microparticles were characterized for particle size and size distribution. Microparticles were compressed into tablets using the directly compressible excipients. SEM photographs of the fractured part of the tablet revealed the presence of discrete particles in the tablets, suggesting that the system chosen is ideal for tableting. Drug release from the tableted microparticles exhibited an initial burst effect, but the release decreased with increasing extent of cross-linking. Tablets were coated with chitosan or cellulose acetate, which significantly lowered the initial burst effect when compared to uncoated tablets. Drug release from chitosan-coated tablets was slightly higher than the tablets coated with cellulose acetate. Tablets prepared were effective in delivering clozapine over a period of 12?h.     

The role of particle physico-chemical properties in pulmonary drug delivery for tuberculosis therapy

Abstract

There is increasing interest in the use of inhaled aerosol drug therapy for the treatment of tuberculosis (TB). A number of methods of preparation of particles have been employed including spray drying, solvent evaporation, emulsion and phospholipid methods to create microparticles, macroaggregated nanoparticles, solid lipid nanoparticles and liposomes. Each of these methods involves the use of different proportions of additives to aid in the particle formation or to achieve important physico-chemical properties such as ease of dispersion. While these approaches all have merit their practical value is limited by constraints on dose and means of delivery as an aerosol in order to achieve a therapeutic effect. A review of a number of approaches is presented and placed in the context of the need for effective aerosol delivery systems for the treatment of TB as a guide to selection of appropriate excipients, processes and delivery strategies to support product development activities.