Self-assembly of chemically programmed amphiphiles into aqueous nanotubes with a lipophilic lumen

The creation of complex hollow nanostructures with precise control over size and shape represents a great challenge in supramolecular soft materials. Here, we have further developed a bioinspired methodology for the formation of aqueous nanotubes of well-defined dimensions and pore coating through the self-assembly of amphiphiles that are chemically programmed with complementary nucleobases. These nanotubes are endowed with a hydrophobic lumen, whose diameter can be expanded as a function of the monomer length, in which apolar dyes can be efficiently encapsulatedPID2020-116921GB-I00, TED2021-132602B-I00, PID2020-116112RJ-I00, TED2021-131906A-100, PID2021-128313OB-I0

Ultrabright Föster Resonance Energy Transfer Nanovesicles:The Role of Dye Diffusion

The development of contrast agents based on fluorescent nanoparticles with high brightness and stability is a key factor to improve the resolution and signal-to-noise ratio of current fluorescence imaging techniques. However, the design of bright fluorescent nanoparticles remains challenging due to fluorescence self-quenching at high concentrations. Developing bright nanoparticles showing FRET emission adds several advantages to the system, including an amplified Stokes shift, the possibility of ratiometric measurements, and of verifying the nanoparticle stability. Herein, we have developed Förster resonance energy transfer (FRET)-based nanovesicles at different dye loadings and investigated them through complementary experimental techniques, including conventional fluorescence spectroscopy and super-resolution microscopy supported by molecular dynamics calculations. We show that the optical properties can be modulated by dye loading at the nanoscopic level due to the dye's molecular diffusion in fluid-like membranes. This work shows the first proof of a FRET pair dye's dynamism in liquid-like membranes, resulting in optimized nanoprobes that are 120-fold brighter than QDot 605 and exhibit >80% FRET efficiency with vesicle-to-vesicle variations that are mostly below 10%.J.M.-F. gratefully thanks the financial support received by the European Union’s Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement No 712949 (TECNIOspring PLUS) and from the Agency for Business Competitiveness of the Government of Catalonia. We acknowledge the European Commission (EC) FP7-PEOPLE-2013-Initial Training Networks (ITN) “NANO2FUN” project no. 607721 for being the spark that initiates this work and EC project MSCA-RISE-2020 "MICRO4NANO" project no.101007804. This work was also financially supported by Generalitat de Catalunya (grant no. 2017-SGR-918), the Ministry of Economy, Industry, and Competitiveness (Spain), through the “MOTHER” project (MAT2016-80826-R), the Ministry of Science and Innovation of Spain through the grant PID2019-105622RB-I00 (Mol4Bio). ICMAB-CSIC also acknowledges support from the MINECO through the Severo Ochoa Programme FUNFUTURE (SEV-2015-0496 and CEX2019-000917-S). K.D.B. acknowledges the National Science Foundation (CBET-1517273 and CHE-1726345). C.S. and A.P. benefited from the equipment and framework of the COMP-HUB Initiative, funded by the “Departments of Excellence” program of the Italian Ministry for Education, University and Research (MIUR, 2018-2022). We thank the CESGA Supercomputing Center for technical support and the use of computational resources. The contribution of S.I.-T. has been done under the Materials Science PhD program in the Barcelona Autonomous University (UAB). Characterizations of nanovesicles were made at the ICTS “NANBIOSIS”, more specifically by the U6 unit of CIBER-BBN. The authors would like also to thank the collaboration of Hamamatsu Photonics for the quantum yield determinations using the Quantaurus-QY Plus UV–NIR absolute PL quantum yield spectrometer.With funding from the Spanish government through the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2019-000917-S).Peer reviewe

Traction forces at the cytokinetic ring regulate cell division and polyploidy in the migrating zebrafish epicardium

Epithelial repair and regeneration are driven by collective cell migration and division. Both cellular functions involve tightly controlled mechanical events, but how physical forces regulate cell division in migrating epithelia is largely unknown. Here we show that cells dividing in the migrating zebrafish epicardium exert large cell–extracellular matrix (ECM) forces during cytokinesis. These forces point towards the division axis and are exerted through focal adhesions that connect the cytokinetic ring to the underlying ECM. When subjected to high loading rates, these cytokinetic focal adhesions prevent closure of the contractile ring, leading to multi-nucleation through cytokinetic failure. By combining a clutch model with experiments on substrates of different rigidity, ECM composition and ligand density, we show that failed cytokinesis is triggered by adhesion reinforcement downstream of increased myosin density. The mechanical interaction between the cytokinetic ring and the ECM thus provides a mechanism for the regulation of cell division and polyploidy that may have implications in regeneration and cancer

Super-resolution microscopy for nanomedicine research

\u3cp\u3eSuper-resolution microscopy, or nanoscopy, revolutionized the field of cell biology, enabling researchers to visualize cellular structures with nanometric resolution, single-molecule sensitivity, and in multiple colors. However, the impact of these techniques goes beyond biology as the fields of nanotechnology and nanomedicine can greatly benefit from them, as well. Nanoscopy can visualize nanostructures in vitro and in cells and can contribute to the characterization of their structures and nano-bio interactions. In this Perspective, we discuss the potential of super-resolution imaging for nanomedicine research, its technical challenges, and the future developments we envision for this technology.\u3c/p\u3

Electrochemical Investigation of Cellular Uptake of Quantum Dots Decorated with a Proline-Rich Cell Penetrating Peptide

6 páginas, 3 figuras.The use of square wave voltammetry to monitor the cellular uptake, in HeLa cells, of quantum dots (QD) decorated with sweet arrow peptide (SAP) is reported. A SAP derivative containing an additional N-terminal cysteine residue (C-SAP) was synthesized using the solid-phase method and conjugated to QDs. The obtained results show that QDs-SAP either interact with the extracellular cell membrane matrix or translocate the bilayer. The first situation, membrane adsorption, is probably a transient state before cellular uptake. Both confocal microscopy and SWV results support the detection of this cellular internalization process. The developed electrochemical investigation technique can provide valuable insights into the study of peptide-mediated delivery, as well as the design and development of nanoparticle probes for intracellular imaging, diagnostic, and therapeutic applications. In addition, the described electrochemical interrogation is low cost, is easy to use, and offers future interest for diagnostics including cell analysis.This work was supported by MAT2008-03079/NAN (MEC, Madrid); MCI-FEDER (Bio2008-00799, NAN2004-09159-C04- 02 and NANOBIOMED-CONSOLIDER) and the Generalitat de Catalunya (CeRBa and 2005SGR-00663). S. Pujals is supported by a FPU grant from the Ministerio de Ciencia e Innovación of Spain.Peer reviewe

D-SAP: A new, noncytotoxic, and fully protease resistant cell-penetrating peptide

Protease resistant cell-penetrating peptides (CPPs) are promising carriers for drugs unable to cross the cell membrane. As these CPPs are stable in vivo for much longer periods of time compared to other classes of therapeutic peptides, noncytotoxicity is a property sine qua non for their pharmacological development. Described herein is a fully protease resistant CPP that is noncytotoxic at concentrations up to 1 mM. Proteolytic stability was obtained by chiral inversion of the residues of a known self-assembling CPP—from all L-amino acids to all D-amino acids—and then assessed against trypsin and human serum. Circular dichroism studies confirmed the enantiomeric structure of the analogue, and transmission electron microscopy (TEM) studies indicated that the new inverso analogue retains the ability of the original peptide to self-assemble. The results of uptake experiments indicate that the protease-stable (that is, D-amino acid) analogue of the peptide is internalised by cells to the same extent as the protease-susceptible (that is, L-amino acid) parent peptide. Also reported herein are the results of studies on the cellular internalisation mechanism of the all-D analogue, which reveal the steps followed by the peptide upon its entry into the cell.This work was supported by MCYT-FEDER (Bio2005-00295 and NAN2004-09159-C04-02) and the Generalitat de Catalunya (CeRBa and 2005SGR-00663). S.P. is supported by a grant from the Ministerio de Educación y Ciencia of Spain.Peer reviewe

Studying structure and dynamics of self-assembled peptide nanostructures using fluorescence and super resolution microscopy

Understanding the formation and properties of self-Assembled peptide nanostructures is the basis for the design of new architectures for various applications. Here we show the potential of fluorescence and super resolution imaging to unveil the structural and dynamic features of peptide nanofibers with high spatiotemporal resolution

Unveiling protein corona formation around self-propelled enzyme nanomotors by nanoscopy

The interaction of nanoparticles with biological media is a topic of general interest for drug delivery systems and among those for active nanoparticles, also called nanomotors. Herein, we report the use of super resolution microscopy, in particular, stochastic optical reconstruction microscopy (STORM), to characterize the formation of a protein corona around active enzyme-powered nanomotors. First, we characterized the distribution and number of enzymes on nano-sized particles and characterized their motion capabilities. Then, we incubated the nanomotors with fluorescently labelled serum proteins. Interestingly, we observed a significant decrease of protein corona formation (20%) and different composition, which was studied by proteomic analysis. Moreover, motion was not hindered, as nanomotors displayed enhanced diffusion regardless of the protein corona. Elucidating how active particles interact with biological media and maintain their self-propulsion after protein corona formation will pave the way for the use of these systems in complex biological fluids in biomedicine.</p

Studying structure and dynamics of self-assembled peptide nanostructures using fluorescence and super resolution microscopy

\u3cp\u3eUnderstanding the formation and properties of self-Assembled peptide nanostructures is the basis for the design of new architectures for various applications. Here we show the potential of fluorescence and super resolution imaging to unveil the structural and dynamic features of peptide nanofibers with high spatiotemporal resolution.\u3c/p\u3

Nanoscale mapping functional sites on nanoparticles by points accumulation for imaging in nanoscale topography (PAINT)

\u3cp\u3eThe ability of nanoparticles to selectively recognize a molecular target constitutes the key toward nanomedicine applications such as drug delivery and diagnostics. The activity of such devices is mediated by the presence of multiple copies of functional molecules on the nanostructure surface. Therefore, understanding the number and the distribution of nanoparticle functional groups is of utmost importance for the rational design of effective materials. Analytical methods are available, but to obtain quantitative information at the level of single particles and single functional sites, i.e., going beyond the ensemble, remains highly challenging. Here we introduce the use of an optical nanoscopy technique, DNA points accumulation for imaging in nanoscale topography (DNA-PAINT), to address this issue. Combining subdiffraction spatial resolution with molecular selectivity and sensitivity, DNA-PAINT provides both geometrical and functional information at the level of a single nanostructure. We show how DNA-PAINT can be used to image and quantify relevant functional proteins such as antibodies and streptavidin on nanoparticles and microparticles with nanometric accuracy in 3D and multiple colors. The generality and the applicability of our method without the need for fluorescent labeling hold great promise for the robust quantitative nanocharacterization of functional nanomaterials.\u3c/p\u3