Search

Microencapsulated phase change materials (MePCMs) based on swelling polymerization in Pickering emulsion were demonstrated. Monodisperse poly(glycidyl methacrylate-co-2-hydroxyethyl methacrylate) P(GMA-co-HEMA) particles were prepared by dispersion polymerization. The introduction of hydrophilic monomer HEMA endowed the obtained particles with suitable surface hydrophilicity for stabilizing Pickering emulsion. MePCMs with PGMA–polystyrene interpenetrating composite shell were formed when noncrosslinked particles were employed as stabilizer, while MePCMs with particlesembedded shell were prepared with crosslinked particles as Pickering stabilizer. Phase change property, thermal stability and thermal reliability of two kinds of MePCMs were investigated by TGA, FT-IR, DSC and cycling test. The temperature of 10% weight loss of MePCMs with interpenetrating shell was 11 °C higher than that of MePCMs with particles-embedded shell. After cycling test, the percentage of leached core materials from MePCMs with interpenetrating shell was approximately one-third of that from MePCMs with particles-embedded shell. The results showed that MePCMs with an integral shell presented better thermal stability, tightness and thermal reliability than MePCMs with particles-embedded shell. The research developed a simple process for MePCMs with excellent performance. [ABSTRACT FROM AUTHOR]

The first guidance on Good Cell Culture Practice (GCCP) dates back to 2005. This document expands this to include aspects of quality assurance for in vitro cell culture focusing on the increasingly diverse cell types and culture formats used in research, product development, testing and manufacture of biotechnology products and cell-based medicines. It provides a set of basic principles of best practice that can be used in training new personnel, reviewing and improving local procedures, and helping to assure standard practices and conditions for the comparison of data between laboratories and experimentation performed at different times. This includes recommendations for the documentation and reporting of culture conditions. It is intended as guidance to facilitate the generation of reliable data from cell culture systems, and is not intended to conflict with local or higher level legislation or regulatory requirements. It may not be possible to meet all recommendations in this guidance for practical, legal or other reasons. However, when it is necessary to divert from the principles of GCCP, the risk of decreasing the quality of work and the safety of laboratory staff should be addressed and any conclusions or alternative approaches justified. This workshop report is considered a first step toward a revised GCCP 2.0. [ABSTRACT FROM AUTHOR]

Hexadecane-loaded microcapsules with SiO2-poly(methyl methacrylate) (PMMA) hybrids as shells were prepared by surface-initiated atom transfer radical polymerization (SI-ATRP) in an oil-in-water Pickering emulsion. ATRP-initiators were introduced onto SiO2 particles by a homemade silane coupling reagent containing a 2-bromoisobutyrate group. SI-ATRP polymerization was conducted on Pickering stabilizers based on activators regenerated by electron transfer (ARGET). TGA results indicated that 80.3% of PMMA chains in the shell were covalently-bonded to SiO2, which was formed by SI-ATRP polymerization. The microcapsules show excellent potential for thermal energy storage, as evidenced by a high melting enthalpy of 161.7 J g-1. [ABSTRACT FROM AUTHOR]

Pickering emulsion polymerization has attracted considerable attention in material fabrication due to its unique surfactant-free character and versatile association of oil, water and particles for a large set of materials. In this study, SiO2 modified with Methacryloxypropyltrimethoxysilane (MPTMS) was employed to prepare Pickering emulsion, and subsequently covalently-bonded polystyrene/SiO2(PS/SiO2) composites were synthesized by Oil-in-water Pickering emulsion polymerization. Optical micrograph, contact angle, thermogravimetric analysis (TGA), Fourier transform infrared spectra (FT-IR), scanning electron microscope (SEM) and dynamic laser scattering (DLS) were employed to characterize the modified SiO2, Pickering emulsion and prepared composites. It was found that prepared composites possess ragged surface morphology and SiO2 concentration has an important effect on the morphology of as-prepared composites. In addition, covalent bond between PS core and SiO2 shell was evidenced by FT-IR. [ABSTRACT FROM AUTHOR]

Background: Wnt3a regulates a canonical signaling pathway in early development that controls the nuclear accumulation of β-catenin and its activation of Lef/Tcf-sensitive transcription of developmentally important genes. Results: Using totipotent mouse F9 teratocarcinoma cells expressing Frizzled-1 and biochemical analyses, we detail the influence of Wnt3a stimulation on the expression, complexation, and subcellular trafficking of key signaling elements of the canonical pathway, i.e., Dishevelled-2, Axin, glycogen synthase kinase-3β, and β-catenin. Cellular content of β-catenin and Axin, and phosphoglycogen synthase kinase-3β, but not Dishevelled-2, increases in response to Wnt3a. Subcellular localization of Axin in the absence of Wnt3a is symmetric, found evenly distributed among plasma membrane-, cytosol-, and nuclear-enriched fractions. Dishevelled-2, in contrast, is found predominately in the cytosol, whereas β-catenin is localized to the plasma membrane-enriched fraction. Wnt3a stimulates trafficking of Dishevelled-2, Axin, and glycogen synthase kinase-3β initially to the plasma membrane, later to the nucleus. Bioluminescence resonance energy transfer measurements reveal that complexes of Axin with Dishevelled-2, with glycogen synthase kinase-3β, and with β-catenin are demonstrable and they remain relatively stable in response to Wnt3a stimulation, although trafficking has occurred. Mammalian Dishevelled-1 and Dishevelled-2 display similar patterns of trafficking in response to Wnt3a, whereas that of Dishevelled-3 differs from the other two. Conclusion: This study provides a detailed biochemical analysis of signaling elements key to Wnt3a regulation of the canonical pathway. We quantify, for the first time, the Wnt-dependent regulation of cellular abundance and intracellular trafficking of these signaling molecules. In contrast, we observe little effect of Wnt3a stimulation on the level of protein-protein interactions among these constituents of Axin-based complexes themselves. [ABSTRACT FROM AUTHOR]

Background: Mammalian receptors that couple to effectors via heterotrimeric G proteins (e.g., beta 2-adrenergic receptors) and receptors with intrinsic tyrosine kinase activity (e.g., insulin and IGF-I receptors) constitute the proximal points of two dominant cell signaling pathways. Receptors coupled to G proteins can be substrates for tyrosine kinases, integrating signals from both pathways. Yeast cells, in contrast, display G protein-coupled receptors (e.g., alpha-factor pheromone receptor Ste2) that have evolved in the absence of receptor tyrosine kinases, such as those found in higher organisms. We sought to understand the motifs in G protein-coupled receptors that act as substrates for receptor tyrosine kinases and the functional consequence of such phosphorylation on receptor biology. We expressed in human HEK 293 cells yeast wild-type Ste2 as well as a Ste2 chimera engineered with cytoplasmic domains of the beta2-adrenergic receptor and tested receptor sequestration in response to activation of the insulin receptor tyrosine kinase. Results: The yeast Ste2 was successfully expressed in HEK 293 cells. In response to alpha-factor, Ste2 signals to the mitogen-activated protein kinase pathway and internalizes. Wash out of agonist and addition of antagonist does not lead to Ste2 recycling to the cell membrane. Internalized Ste2 is not significantly degraded. Beta2-adrenergic receptors display internalization in response to agonist (isoproterenol), but rapidly recycle to the cell membrane following wash out of agonist and addition of antagonist. Beta2-adrenergic receptors display internalization in response to activation of insulin receptors (i.e., cross-regulation), whereas Ste2 does not. Substitution of the cytoplasmic domains of the β2-adrenergic receptor for those of Ste2 creates a Ste2/beta2-adrenergic receptor chimera displaying insulin-stimulated internalization. Conclusion: Chimera composed of yeast Ste2 into which domains of mammalian G proteincoupled receptors have been substituted, when expressed in animal cells, provide a unique tool for study of the regulation of G protein-coupled receptor trafficking by mammalian receptor tyrosine kinases and adaptor proteins. [ABSTRACT FROM AUTHOR]

The objective of this study was to investigate the effect of individual PUFAs on LDL receptor (LDLr) expression in human fibroblasts and HepG2 cells, and to evaluate whether acyl CoA:cholesterol acyltransferase (ACAT) and sterol regulatory element-binding protein 1 (SREBP-1) were involved in the regulation of LDLr expression by fatty acids. When fibroblasts and HepG2 cells were cultured with serum-free defined medium for 48 h, there was a 3- to 5-fold (P < 0.05) increase in LDLr protein and mRNA levels. Incubation of fibroblasts and HepG2 cells in serum-free medium supplemented with 25-hydroxycholesterol (25OH-cholesterol, 5 mg/L) for 24 h decreased LDLr protein and mRNA levels by 50-90% (P < 0.05). Arachidonic acid [AA, 20:4(n-6)], EPA [20:5(n-3)], and DHA [22:6(n-3)] antagonized the depression of LDLr gene expression by 25OH-cholesterol and increased LDLr protein abundance 1-to 3-fold (P < 0.05), but had no significant effects on LDLr mRNA levels. Oleic (18:1), Iinoleic (18:2), and α-linolenic acids [18:3(n-3)] did not significantly affect LDLr expression. ACAT inhibitor (58-035, 1 mg/L) attenuated the regulatory effect of AA on LDLr protein abundance by ∼40% (P < 0.05), but did not modify the regulatory effects of other unsaturated fatty acids in HepG2 cells. The present results suggest that AA, EPA, and DHA increase LDLr protein levels, and that ACAT plays a role in modulating the effects of AA on LDLr protein levels. Furthermore, the effects of the fatty acids appeared to be independent of any change in SREBP-1 protein. [ABSTRACT FROM AUTHOR]

Conjugated linoleic acid (CLA) exerts anticarcinogenic and antiatherosclerotic effects in animals. The present study was conducted to examine the effects of CLA on LDL receptor (LDLr) expression in HepG2 cells, and to evaluate whether the sterol response element binding protein 1 (SREBP-1) and acyl CoA:cholesterol acyltransferase (ACAT) were involved in the regulation of LDLr expression by CLA. When HepG2 cells were cultured with serum-free DMEM for 48 h, there was a three- to five-fold (P < 0.05) increase in LDLr protein and mRNA levels. Incubation of HepG2 cells in serum-free medium supplemented with 25hydroxycholesterol (25OH, 5 mg/L) for 24 h decreased LDLr protein and mRNA by 50-70% (P < 0.05) and mature SREBP-1 by 20-40% (P < 0.05). CLA, but not linoleic acid, antagonized the depressive effects of 25OH and increased both LDLr protein and mRNA abundance twofold (P < 0.05). LDLr protein and mRNA abundance were not different when HepG2 cells were cultured with CLA (0.4 mmol/L) plus 25OH in the presence or absence of an ACAT inhibitor (58-035, 1 mg/L). Furthermore, CLA had no effect on SREBP-1 abundance. These results suggest that CLA upregulates LDLr expression via a mechanism that is independent of ACAT and SREBP-1. KEY WORDS: cholesterol; conjugated linoleic acid; LDL LDL receptor mRNA; LDL receptor protein. [ABSTRACT FROM AUTHOR]