Nanopatterned Protein Films Directed by Ionic Complexation with Water-Soluble Diblock Copolymers

The use of ionic interactions to direct both protein templating and block copolymer self-assembly into nanopatterned films with only aqueous processing conditions is demonstrated using block copolymers containing both thermally responsive and pH responsive blocks. Controlled reversible addition–fragmentation chain-transfer (RAFT) polymerization is employed to synthesize poly­(<i>N</i>-isopropylacrylamide-<i>b</i>-2-(dimethylamino)­ethyl acrylate) (PNIPAM-<i>b</i>-PDMAEA) diblock copolymers. The pH-dependent ionic complexation between the fluorescent protein, mCherry, and the ionic PDMAEA block is established using dynamic light scattering (DLS) and UV–vis spectroscopy. DLS shows that the size of the resulting coacervate micelles depends strongly on pH, while UV–vis spectroscopy shows a correlation between the protein’s absorption maximum and the ionic microenvironment. Zeta potential measurements clearly indicate the ionic nature of the complex-forming interactions. Spin-casting was used to prepare nanostructured films from the protein–block copolymer coacervates. After film formation, the lower critical solution temperature (LCST) of the PNIPAM blocks allows the nanomaterial to be effectively immobilized in aqueous environments at physiological temperatures, enabling potential use as a controlled protein release material or polymer matrix for protein immobilization. At pH 9.2 and 7.8, the release rates are at least 10 times faster than that at pH 6.4 due to weaker interaction between protein and PNIPAM-<i>b</i>-PDMAEA (PND) diblock copolymer. Because of the ionic environment in which protein is confined, the majority of the protein (80%) remains active, independent of pH, even after having been dehydrated in vacuum and confined in the films

Salami-like Electrospun Si Nanoparticle-ITO Composite Nanofibers with Internal Conductive Pathways for use as Anodes for Li-Ion Batteries

We report novel salami-like core–sheath composites consisting of Si nanoparticle assemblies coated with indium tin oxide (ITO) sheath layers that are synthesized via coelectrospinning. Core–sheath structured Si nanoparticles (NPs) in static ITO allow robust microstructures to accommodate for mechanical stress induced by the repeated cyclical volume changes of Si NPs. Conductive ITO sheaths can provide bulk conduction paths for electrons. Distinct Si NP-based core structures, in which the ITO phase coexists uniformly with electrochemically active Si NPs, are capable of facilitating rapid charge transfer as well. These engineered composites enabled the production of high-performance anodes with an excellent capacity retention of 95.5% (677 and 1523 mAh g^–1, which are based on the total weight of Si-ITO fibers and Si NPs only, respectively), and an outstanding rate capability with a retention of 75.3% from 1 to 12 C. The cycling performance and rate capability of core–sheath-structured Si NP-ITO are characterized in terms of charge-transfer kinetics

Effect of N-acetyl cysteine (Nac), Mito-TEMPO, and Mdivi-1 treatment on mitochondrial morphology in 3T3-L1 cells after differentiation.

<p>(A) DsRed2-Mito-expressing 3T3-L1 cells treated with 1 μg/mL insulin for 6 days with or without 10 mM Nac, 200 μM Mito-TEMPO, or 50 μM Mdivi-1 after MDI treatment for 48 h. Mitochondrial morphology was determined by confocal microscopy. (B) Graph showing average mitochondrial length. (C) Graph indicating mitochondrial morphology. Data in the bar graphs represent the means ± SEM of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001 compared with untreated or insulin-treated cells. ^##p < 0.01, ^###p < 0.001 compared with untreated cells.</p

Effect of N-acetyl cysteine (Nac), Mito-TEMPO, and Mdivi-1 on lipid accumulation in 3T3-L1 cells during adipogenesis.

<p>3T3-L1 adipocytes were pretreated with 10 mM Nac, 200 μM Mito-TEMPO, or 50 μM Mdivi-1 for 30 min and then with insulin during adipogenesis. (A) Accumulated lipid was stained with Oil Red O reagent and quantified by absorbance at 490 nm (B). Data in the bar graph represent the means ± SEM of three independent experiments. *p < 0.05, ***p < 0.001, compared with untreated or insulin-treated cells. ^###p<0.001, compared with insulin-treated cells.</p

Effect of N-acetyl cysteine (Nac), Mito-TEMPO, and Mdivi-1 treatment on antioxidant related genes in 3T3-L1 cells after differentiation.

<p>(A) 3T3-L1 adipocytes were pretreated with 10 mM Nac, 200 μM Mito-TEMPO, or 50 μM Mdivi-1 for 30 min and then with insulin during adipogenesis. Cells were lysed and proteins were subjected to Western blot analysis analysis with the indicated antibodies; antioxidant genes (Prx1, Prx2, Prx3, Prx5, SOD1, and SOD2). Data in the bar graph represent the means ± SEM of three independent experiments. * p< 0.05, ** p< 0.01, *** p< 0.001, compared with not treated cell. # p< 0.05, ## p< 0.01, ### p< 0.001, compared with insulin-treated cell.</p

Effect of N-acetyl cysteine (Nac), Mito-TEMPO, and Mdivi-1 treatment on intracellular and mitochondrial ROS production in 3T3-L1 cells after differentiation.

<p>(A) 3T3-L1 adipocytes were pretreated with 10 mM Nac, 200 μM Mito-TEMPO, or 50 μM Mdivi-1 for 30 min and then with insulin during adipogenesis. After 8 days of the initiation of differentiation, cells were used in CM-H₂DCF-DA (A, B) and Mito-SOX (A, C) stained assays. Data in the bar graphs represent the means ± SEM of three independent experiments. ***p < 0.001 compared with untreated or insulin-treated cells.</p

Effect of N-acetyl cysteine (Nac), Mito-TEMPO, and Mdivi-1 on mitochondrial dynamics-related genes in 3T3-L1 cells after differentiation.

<p>3T3-L1 adipocytes were pretreated with 10 mM Nac, 200 μM Mito-TEMPO, or 50 μM Mdivi-1 for 30 min and then with insulin during adipogenesis. Cells were lysed and proteins were subjected to western blot analysis with the indicated antibodies: mitochondrial dynamics-related genes (OPA1, Mfn1, Mfn2, p-Drp1 (Ser616), and Drp1). Data in the bar graphs represent the means ± SEM of three independent experiments. **p < 0.01, ***p < 0.001, compared with untreated cells. ^#p < 0.05, ^##p < 0.01, ^###p < 0.001 compared with insulin-treated cells.</p

Effect of N-acetyl cysteine (Nac), Mito-TEMPO, and Mdivi-1 on the expression of adipogenic related genes in 3T3-L1 cells after differentiation.

<p>3T3-L1 adipocytes were pretreated with 10 mM Nac, 200 μM Mito-TEMPO, or 50 μM Mdivi-1 for 30 min and then with insulin during adipogenesis. Cells were lysed and proteins were subjected to western blot analysis with the indicated antibodies: adipogenic genes (PPARγ, C/EBPα, p-AKT, AKT, GLUT4, and aP2). Data in the bar graphs represent the means ± SEM of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001, compared with untreated cells. ^#p < 0.05, ^##p < 0.01, ^###p < 0.001 compared with insulin-treated cells.</p

Tuning the “Roadblock” Effect in Kinesin-Based Transport

Major efforts are underway to harness motor proteins for technical applications. Yet how to best attach cargo to microtubules that serve as kinesin-driven “molecular shuttles” without compromising transport performance remains challenging. Furthermore, microtubule-associated proteins (MAPs) can block motor protein-powered transport in neurons, which can lead to neurodegenerative diseases. Again it is unclear how different physical roadblock parameters interfere with the stepping motion of kinesins. Here, we employ a series of MAPs, tailored (strept)­avidins, and DNA as model roadblocks and determine how their geometrical, nanomechanical, and electrochemical properties can reduce kinesin-mediated transport. Our results provide insights into kinesin transport regulation and might facilitate the choice of appropriate cargo linkers for motor protein-driven transport devices

Tuning the “Roadblock” Effect in Kinesin-Based Transport

Major efforts are underway to harness motor proteins for technical applications. Yet how to best attach cargo to microtubules that serve as kinesin-driven “molecular shuttles” without compromising transport performance remains challenging. Furthermore, microtubule-associated proteins (MAPs) can block motor protein-powered transport in neurons, which can lead to neurodegenerative diseases. Again it is unclear how different physical roadblock parameters interfere with the stepping motion of kinesins. Here, we employ a series of MAPs, tailored (strept)­avidins, and DNA as model roadblocks and determine how their geometrical, nanomechanical, and electrochemical properties can reduce kinesin-mediated transport. Our results provide insights into kinesin transport regulation and might facilitate the choice of appropriate cargo linkers for motor protein-driven transport devices