Modular synthesis of semiconducting graft co-polymers to achieve ‘clickable’ fluorescent nanoparticles with long circulation and specific cancer targeting

Semiconducting polymer nanoparticles (SPNs) are explored for applications in cancer theranostics because of their high absorption coefficients, photostability, and biocompatibility. However, SPNs are susceptible to aggregation and protein fouling in physiological conditions, which can be detrimental for in vivo applications. Here, a method for achieving colloidally stable and low-fouling SPNs is described by grafting poly(ethylene glycol) (PEG) onto the backbone of the fluorescent semiconducting polymer, poly(9,9′-dioctylfluorene-5-fluoro-2,1,3-benzothiadiazole), in a simple one-step substitution reaction, postpolymerization. Further, by utilizing azide-functionalized PEG, anti-human epidermal growth factor receptor 2 (HER2) antibodies, antibody fragments, or affibodies are site-specifically “clicked” onto the SPN surface, which allows the functionalized SPNs to specifically target HER2-positive cancer cells. In vivo, the PEGylated SPNs are found to have excellent circulation efficiencies in zebrafish embryos for up to seven days postinjection. SPNs functionalized with affibodies are then shown to be able to target HER2 expressing cancer cells in a zebrafish xenograft model. The covalent PEGylated SPN system described herein shows great potential for cancer theranostics

Standardization of methods to record Vagus nerve activity in mice

Abstract Background The vagus nerve plays an important role in the regulation of organ function, including reflex pathways that regulate immunity and inflammation. Recent studies using genetically modified mice have improved our understanding of molecular mechanisms in the neural control of immunity. However, mapping neural signals transmitted in the vagus nerve in mice has been limited by technical challenges. Here, we have standardized an experimental protocol to record compound action potentials transmitted in the vagus nerve. Methods The vagus nerve was isolated in Balb/c and B6.129S mice, and placed either on a hook or cuff electrode. The electrical signals from the vagus nerve were digitized using either a Neuralynx or Plexon data acquisition system. Changes in the vagus nerve activity in response to anesthesia, feeding and administration of bacterial endotoxin were analyzed. Results We have developed an electrophysiological recording system to record compound action potentials from the cervical vagus nerve in mice. Cuff electrodes significantly reduce background noise and increase the signal to noise ratio as compared to hook electrodes. Baseline vagus nerve activity varies in response to anesthesia depth and food intake. Analysis of vagus neurograms in different mouse strains (Balb/c and C57BL/6) reveal no significant differences in baseline activity. Importantly, vagus neurogramactivity in wild type and TLR4 receptor knock out mice exhibits receptor dependency of endotoxin mediated signals. Conclusions These methods for recording vagus neurogram in mice provide a useful tool to further delineate the role of vagus neural pathways in a standardized murine disease model

L amino acid transporter structure and molecular bases for the asymmetry of substrate interaction

L-amino acid transporters (LATs) play key roles in human physiology and are implicated in several human pathologies. LATs are asymmetric amino acid exchangers where the low apparent affinity cytoplasmic side controls the exchange of substrates with high apparent affinity on the extracellular side. Here, we report the crystal structures of an LAT, the bacterial alanine-serine-cysteine exchanger (BasC), in a non-occluded inward-facing conformation in both apo and substrate-bound states. We crystallized BasC in complex with a nanobody, which blocks the transporter from the intracellular side, thus unveiling the sidedness of the substrate interaction of BasC. Two conserved residues in human LATs, Tyr 236 and Lys 154, are located in equivalent positions to the Na1 and Na2 sites of sodium-dependent APC superfamily transporters. Functional studies and molecular dynamics (MD) calculations reveal that these residues are key for the asymmetric substrate interaction of BasC and in the homologous human transporter Asc-1.This work was funded by the Spanish Ministry of Science and Innovation (grant SAF2015-64869-R-FEDER), the Fundació la Marató TV3 (20132330), Research Contract with SIDRA Medicine (Qatar), CIBERER ACCI 2017-U731, and the Generalitat de Catalunya (grant SGR2009-1355)