Effect of the stiffness of one-layer protein-based microcapsules on dendritic cell uptake and endocytic mechanism.

Microcapsules are one of the most promising microscale drug carriers due to their facile fabrication, excellent deformability, and high efficacy in drug storage and delivery. Understanding the effects of their physicochemical properties (size, shape, rigidity, charge, surface chemistry, etc.) on both in vitro and in vivo performance is not only highly significant and interesting but also very challenging. Stiffness, an important design parameter, has been extensively explored in recent years, but how the rigidity of particles influences cellular internalization and uptake mechanisms remains controversial. Here, one-layered lysozyme-based microcapsules with well-controlled stiffness (modulus ranging from 3.49 ± 0.18 MPa to 26.14 ± 1.09 MPa) were prepared and used to investigate the effect of stiffness on the uptake process in dendritic cells and the underlying mechanism. The cellular uptake process and endocytic mechanism were investigated with laser scanning confocal microscopy, mechanism inhibitors, and pathway-specific antibody staining. Our data demonstrated that the stiffness of protein-based microcapsules could be a strong regulator of intracellular uptake and endocytic kinetics but had no obvious effect on the endocytic mechanism. We believe our results will provide a basic understanding of the intracellular uptake process of microcapsules and the endocytic mechanism and inspire strategies for the further design of potential drug delivery microcarriers.

Sheltering proteins from protease-mediated degradation and a de novo strategy for preventing acute liver injury.

Restoring protein functions or supplying proteins is considered one of the most powerful therapeutic strategies for many diseases, but it is mainly limited by the denaturation of proteins during encapsulation and degradation by proteases during in vivo delivery, and limits its delivery. Herein, by encapsulating a protein (catalase, an enzyme) in a hexahistidine-metal assembly (HmA) via a de novo strategy under mild conditions, we demonstrated that HmA could maintain the bioactivity of the enzyme, protect the enzyme from proteinase degradation, and deliver the encapsulated protein for the prevention of disease in an acute liver injury model.

Novel One-Dimensional Covalent Organic Framework as a H+ Fluorescent Sensor in Acidic Aqueous Solution.

Covalent organic frameworks (COFs) represent an emerging class of two- or three-dimensional crystalline porous materials with delicate control over topology, composition, and porosity. Here, we develop a new COF made up of 1,3,6,8-tetrakis(p-formylphenyl)pyrene (TFPPy) and 4,4'-diaminobenzophenone (DABP) that exhibits a rare one-dimensional (1D) structure. The resulting frameworks possess good crystallinity, comparatively high Brunauer-Emmett-Teller (BET) surface area (426 m2/g), and good thermal stability (360 °C). Impressively, this 1D COF shows strong fluorescence and can be used as an excellent H+ sensor in an acidic aqueous solution.

A Facile and Universal Method to Efficiently Fabricate Diverse Protein Capsules for Multiple Potential Applications.

Proteins are considered to be one of the most important highly reproducible and monodisperse building blocks with specific functions in life sciences and material science. Protein capsules and their hybrids composed of protein-polymer conjugates have been intensively explored in drug delivery, catalysis, and cell-mimicking functions. Herein, we present a facile, universal, and efficient method to fabricate the diverse protein capsules, independent of the molecular weight (Mw), isoelectric points (IEP), wettability, amino acid sequence, and functional domains of enumerated proteins. The protein capsules were well characterized by various techniques. Furthermore, their ability to store the original protein functionality was demonstrated, which was mainly embodied in their enzyme responsiveness and good biocompatibility in vitro and in vivo. We believe that these protein capsules have multiple potential applications such as in drug delivery, tissue engineering, catalysis, and other application fields.

The construction and effect of physical properties on intracellular drug delivery of poly(amino acid) capsules.

Constructing intracellular degradable drug delivery vehicles is critical to fully exert the function of loaded drugs. Considering the poly (amino acid) is sensitively degradable to acid and enzyme which indwell in the mature lysosome, we here presented the poly(amino acid) capsules constructed by the synthetic poly(amino acid), (poly-glutamic acid, PGA and poly-ornithine, POR). The fabrication of Dox loaded poly (amino acid) capsules was demonstrated, and was thoroughly characterized by various techniques, including Zetasizer, SEM, TEM, fluorescent microscopy, and confocal laser scan microscopy. By controlling fabrication process, we tuned the carriers with different physical properties (charges and stiffness). Then, we thoroughly investigated the effects of these properties on the intracellular uptake and anti-cancer abilities of various carriers@Dox. In addition, the degradability of poly(amino acid) capsules was studied to reveal the release profiles of the carriers with or without templates from the side aspect. We found the positively charged and stiffer carriers mainly contributed to the cellular uptake process and amount, while both the uptake amount and degradability of the endocytosed carriers@Dox played a critical role on the cytotoxicity. We believe the findings here could pave the way for designing poly(amino acid) capsules or other degradable polymers based on poly(amino acid) as the drug delivery vehicles.

Insight into the mechanism and factors on encapsulating basic model protein, lysozyme, into heparin doped CaCO3.

Porous CaCO3 microparticles are considered as one of the most popular and effective carriers for protein loading. Most of current studies have centered on elucidating particle formation and enhancing the protein loading efficiency, very few reports on the kinetics, driving forces and factors on protein loading process. Here, we took lysozyme as the basic model protein to investigate the kinetics, driving forces on protein loading and factors controlling loading efficiency into porous Hep/CaCO3 microparticles by various techniques (protein quantification assay, QCM-D, SEM, BET, Zeta sizer, TGA, CLSM, CD spectrum, bioactive assay). As revealed, the adsorption process obeyed the pseudo second-order kinetics and Langmuir adsorption model. Doping heparin greatly influenced the detailed texture, pore size, surface area, and maximum loading capacity of lysozyme (LCLys,m). The dependence of LCLys,m on pH reflected the electrostatic interaction mainly contributed to lysozyme adsorption, especially below IEP of lysozyme. But the hydrophobic interaction also played the critical role on lysozyme adsorption at pH around IEP of lysozyme. Accompanying with pH change, the lysozyme orientation shifted from "side on" at lower pH to "end on" at pH around IEP. At proper concentration of NaCl (CNaCl), the loaded lysozyme could be released from Hep/CaCO3 microparticles, making them available for lysozyme reloading. Most importantly, such release-reloading cycle didn't disturb the bioactivity of released lysozyme and following reloading ability. We believe our work will contribute to understand protein adsorption behaviors, improve protein loading efficacy, biomaterials design, tissue engineering and disease treatment.

A facile and efficient strategy to encapsulate the model basic protein lysozyme into porous CaCO3.

Basic proteins play important roles in biological activities and disease treatment, but their high sensitivity to the body environment and short life time have limited their applications. Encapsulating the proteins into carriers has been demonstrated to be an effective way to prolong the protein half life time and to control the release temporally and spatially. However, fabricating protein carriers with high protein loading efficacy under mild conditions is still a big challenge. The capsules generated by the combination of the layer by layer (LBL) technique and sacrificial templates have been extensively investigated for the encapsulation of proteins. Porous CaCO3 is an effective sacrificial template with the ability to load various drugs efficiently under mild conditions, but it shows very poor ability in loading basic proteins. Here, we developed a highly efficient but very simple method to encapsulate lysozyme into porous CaCO3. An efficiency of 99.5% and capacity of 91.6 mg g-1 under very gentle conditions were obtained by doping heparin into porous CaCO3. Most importantly, the activity of the encapsulated lysozyme was almost 100% retained. Furthermore, we evaluated the encapsulation efficiency of the loaded lysozyme during the LBL wrapping process and demonstrated that the loss of the loaded lysozyme was controllable by choosing suitable polyelectrolyte pairs. Considering the multi-interaction mode of heparin with various proteins and the ability to retain the function of the loaded protein demonstrated in our study, we believe that the developed approach has great potential for encapsulating various functional proteins with a wide range of applications in catalysis, disease treatment, and tissue engineering.

A hypothesis on chemical mechanism of the effect of hydrogen.

Many studies have shown that hydrogen can play important roles on the antioxidant, anti-inflammatory and other protective effects. Ohsawa et al have proved that hydrogen can electively and directly scavenge hydroxyl radical. But this mechanism cannot explain more new experimental results. In this article, the hypothesis, which is inspired by H2 could bind to the metal as a ligand, come up to explain its extensive biology effect: Hydrogen could regulate particular metalloproteins by bonding (M-H2 interaction) it. And then it could affect the metabolization of ROS and signal transduction. Metalloproteins may be ones of the target molecules of H2 action. Metal ions may be appropriate role sites for H2 molecules. The hypothesis pointed out a new direction to clarify its mechanisms.