Energetic electron assisted synthesis of highly tunable temperature-responsive collagen/elastin gels for cyclic actuation: macroscopic switching and molecular origins

Thermoresponsive bio-only gels that yield sufficiently large strokes reversibly and without large hysteresis at a well-defined temperature in the physiological range, promise to be of value in biomedical application. Within the present work we demonstrate that electron beam modification of a blend of natural collagen and elastin gels is a route to achieve this goal, viz. to synthesize a bioresorbable gel with largely reversible volume contractions as large as 90% upon traversing a transition temperature that can be preadjusted between 36 °C and 43 °C by the applied electron dose. Employing circular dichroism and temperature depending confocal laser scanning microscopy measurements, we furthermore unravel the mechanisms underlying this macroscopic behavior on a molecular and network level, respectively and suggest a stringent picture to account for the experimental observations. © 2019, The Author(s)

Keratinocytes as Depository of Ammonium-Inducible Glutamine Synthetase: Age- and Anatomy-Dependent Distribution in Human and Rat Skin

In inner organs, glutamine contributes to proliferation, detoxification and establishment of a mechanical barrier, i.e., functions essential for skin, as well. However, the age-dependent and regional peculiarities of distribution of glutamine synthetase (GS), an enzyme responsible for generation of glutamine, and factors regulating its enzymatic activity in mammalian skin remain undisclosed. To explore this, GS localization was investigated using immunohistochemistry and double-labeling of young and adult human and rat skin sections as well as skin cells in culture. In human and rat skin GS was almost completely co-localized with astrocyte-specific proteins (e.g. GFAP). While GS staining was pronounced in all layers of the epidermis of young human skin, staining was reduced and more differentiated among different layers with age. In stratum basale and in stratum spinosum GS was co-localized with the adherens junction component ß-catenin. Inhibition of, glycogen synthase kinase 3β in cultured keratinocytes and HaCaT cells, however, did not support a direct role of ß-catenin in regulation of GS. Enzymatic and reverse transcriptase polymerase chain reaction studies revealed an unusual mode of regulation of this enzyme in keratinocytes, i.e., GS activity, but not expression, was enhanced about 8–10 fold when the cells were exposed to ammonium ions. Prominent posttranscriptional up-regulation of GS activity in keratinocytes by ammonium ions in conjunction with widespread distribution of GS immunoreactivity throughout the epidermis allows considering the skin as a large reservoir of latent GS. Such a depository of glutamine-generating enzyme seems essential for continuous renewal of epidermal permeability barrier and during pathological processes accompanied by hyperammonemia

Illuminating the life of GPCRs

The investigation of biological systems highly depends on the possibilities that allow scientists to visualize and quantify biomolecules and their related activities in real-time and non-invasively. G-protein coupled receptors represent a family of very dynamic and highly regulated transmembrane proteins that are involved in various important physiological processes. Since their localization is not confined to the cell surface they have been a very attractive "moving target" and the understanding of their intracellular pathways as well as the identified protein-protein-interactions has had implications for therapeutic interventions. Recent and ongoing advances in both the establishment of a variety of labeling methods and the improvement of measuring and analyzing instrumentation, have made fluorescence techniques to an indispensable tool for GPCR imaging. The illumination of their complex life cycle, which includes receptor biosynthesis, membrane targeting, ligand binding, signaling, internalization, recycling and degradation, will provide new insights into the relationship between spatial receptor distribution and function. This review covers the existing technologies to track GPCRs in living cells. Fluorescent ligands, antibodies, auto-fluorescent proteins as well as the evolving technologies for chemical labeling with peptide- and protein-tags are described and their major applications concerning the GPCR life cycle are presented

The Concise Guide to PHARMACOLOGY 2023/24: G protein-coupled receptors.

peer reviewedThe Concise Guide to PHARMACOLOGY 2023/24 is the sixth in this series of biennial publications. The Concise Guide provides concise overviews, mostly in tabular format, of the key properties of approximately 1800 drug targets, and about 6000 interactions with about 3900 ligands. There is an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (https://www.guidetopharmacology.org), which provides more detailed views of target and ligand properties. Although the Concise Guide constitutes almost 500 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point-in-time record that will survive database updates. The full contents of this section can be found at http://onlinelibrary.wiley.com/doi/bph.16177. G protein-coupled receptors are one of the six major pharmacological targets into which the Guide is divided, with the others being: ion channels, nuclear hormone receptors, catalytic receptors, enzymes and transporters. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid-2023, and supersedes data presented in the 2021/22, 2019/20, 2017/18, 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the Nomenclature and Standards Committee of the International Union of Basic and Clinical Pharmacology (NC-IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate

Characterization of the Promoter of PRS1 in Saccharomyces cerevisiae Identifies Three Regions Potentially Involved in Control of Expression

The transcription initiation site of the Saccharomyces cerevisiae PRS1 gene was mapped at −179 bp. Measurement of β-galactosidase activity of the successively deleted PRS1 promoter linked to lacZ and integrated at the ura3 locus defined three DNA regions involved in the control of PRS1 expression. Gel shift analysis confirmed the data

Ultralong-Chain-Spaced Crystalline Poly(H-phosphonate)s and Poly(phenylphosphonate)s

Aliphatic poly(H-phosphonate)s were obtained by polyesterification of dimethyl H-phosphonate with bio-based long-chain diols. Nonadecane-1,19-diol, tricosane-1,23-diol, and octatetracontane-1,48-diol with dimethyl H-phosphonate yield the corresponding polyphosphoesters (<b>PPE19H</b>, <b>PPE23H</b>, and <b>PPE48H</b>) with molecular weights (<i>M</i>_n) up to 4.3 × 10⁴ g mol^–1. Postfunctionalization of these polymers via Hirao cross-coupling yields the selectively functionalized poly(H-phosphonate)s <b>PPE19Ph</b>, <b>PPE23Ph</b>, and <b>PPE48Ph</b>. DSC analysis revealed significantly enhanced crystallinities and melting points (up to <i>T</i>_m = 110 °C) with increasing methylene sequence lengths. Hydrolytic degradation of polymer powder of poly-(H-phosphonate) occurred up to 95% in 2 days. The degradation rates decreased with increasing methylene sequence length. After postfunctionalization, degradation occurred only to a minimal extent over 3 months in basic and in acidic media

Direct Observation of Chain Lengths and Conformations in Oligofluorene Distributions from Controlled Polymerization by Double Electron-Electron Resonance

Synthetic polymers are mixtures of different length chains, and their chain length and chain conformation is often experimentally characterized by ensemble averages. We demonstrate that Double-Electron-Electron-Resonance (DEER) spectroscopy can reveal the chain length distribution, and chain conformation and flexibility of the individual n-mers in oligo-(9,9-dioctylfluorene) from controlled Suzuki-Miyaura Coupling Polymerization (cSMCP). The required spin-labeled chain ends were introduced efficiently via a TEMPO-substituted initiator and chain terminating agent, respectively, with an in situ catalyst system. Individual precise chain length oligomers as reference materials were obtained by a stepwise approach. Chain length distribution, chain conformation and flexibility can also be accessed within poly(fluorene) nanoparticles

Multifunctional coatings combining bioactive peptides and affinity-based cytokine delivery for enhanced integration of degradable vascular grafts

Insufficient endothelialization of cardiovascular devices is a high-risk factor for implant failure. Presentation of extracellular matrix (ECM)-derived coatings is a well-known strategy to improve implant integration. However, the complexity of the system is challenging and strategies for applying multifunctionality are required. Here, we engineered mussel-derived surface-binding peptides equipped with integrin (c[RGDfK]) and proteoglycan binding sites (FHRRIKA) for enhanced endothelialization. Surface-binding properties of the platform containing l-3,4-dihydroxyphenylalanine (DOPA) residues were confirmed for hydrophilized polycaprolactone-co-lactide scaffolds as well as for glass and polystyrene. Further, heparin and the heparin-binding angiogenic factors VEGF, FGF-2 and CXCL12 were immobilized onto the peptide in a modular assembly. Presentation of bioactive peptides greatly enhanced human umbilical vein endothelial cell (HUVEC) adhesion and survival under static and fluidic conditions. In subsequent investigations, peptide-heparin-complexes loaded with CXCL12 or VEGF had an additional increasing effect on cell viability, differentiation and migration. Finally, hemocompatibility of the coatings was ensured. This study demonstrates that coatings combining adhesion peptides, glycosaminoglycans and modulators are a versatile tool to convey ECM-inspired multifunctionality to biomaterials and efficiently promote their integration. © 2020 The Royal Society of Chemistry