Several biological barriers must be overcome to achieve efficient nonviral gene delivery. These barriers include target cell uptake, lysosomal degradation, and dissociation from the carrier. In this study, we compared the differences in the uptake mechanism of cationic, star-shaped polymer/MMP-9siRNA complexes (β-CD-(D3)7/MMP-9siRNA complexes: polyplexes) and commercial liposome/MMP-9siRNA complexes (Lipofectamine® 2000/MMP-9siRNA complexes: liposomes). The uptake pathway and transfection efficiency of the polyplexes and liposomes were determined by fluorescence microscopy, flow cytometry, and reverse transcriptase-polymerase chain reaction. The occurrence of intracellular processing was assessed by confocal laser scanning microscopy. Endosomal acidification inhibitors were used to explore the endosomal escape mechanisms of the polyplexes and lysosomes. We concluded that the polyplexes were internalized by non-caveolae- and non-clathrin-mediated pathways, with no lysosomal trafficking, thereby inducing successful transfection, while the majority of liposomes were internalized by clathrin-dependent endocytosis (CDE), caveolae-mediated endocytosis, and macropinocytosis, and only CDE induced successful transfection. Liposomes might escape more quickly than polyplexes, and the digestion effect of acidic organelles on liposomes was faint compared to the polyplexes, although both complexes escaped from endolysosomes via the proton sponge mechanism. This may be the key aspect that leads to the lower transfection efficiency of the β-CD-(D3)7/MMP-9siRNA complexes. The present study may offer some insights for the rational design of novel delivery systems with increased transfection efficiency but decreased toxicity.

The next-generation closure device for interventional treatment of congenital heart disease is regarded to be biodegradable, yet the corresponding biomaterial technique is still challenging. Herein, we report the first fully biodegradable atrial septal defect (ASD) occluder finally coming into clinical use, which is made of biodegradable poly(l-lactic acid) (PLLA). We characterized the physico-chemical properties of PLLA fibers as well as the raw polymer and the operability of the as-fabricated occluders. Cell behaviors on material were observed, and in vivo fiber degradation and inflammatory responses were examined. ASD models in piglets were created, and 44 PLLA ASD occluders were implanted via catheter successfully. After 36 months, the PLLA ASD occluders almost degraded without any complications. The mechanical properties and thickness between newborn and normal atrial septum showed no significant difference. We further accomplished the first clinical implantation of the PLLA ASD occluder in a four-year boy, and the two-year follow-up up to date preliminarily indicated safety and feasibility of such new-generation fully biodegradable occluder made of synthetic polymers.

The adoptive transfer of the natural regulatory B cells and macrophages should be a useful treatment for inflammation and autoimmune disease. However, it is usually difficult to isolate these cells from the tissues and expand them. Here, we investigated the feasibility of adoptively transferring peritoneal cells (PCs) as a treatment for DSS-induced colitis. We found that peritoneal cavity can provide an easily accessible site for harvesting enough number of PCs, namely, two-dose PCs for the treatment from a mouse in one operation. Adoptive therapy of these cells from healthy mice or those with disease is effectively in reducing the disease activity score. The natural B cells and macrophages of the infused PCs can selectively migrate to lesion sites and regulate the expression of Stat3, NF-κB, Smad3 and Smad7. Additionally, PCs exert dual activity of IL-10 and TGF-β secreted spontaneously by both peritoneal B cells and macrophages, which in turn enhance the induction of regulatory B cells and Macrophages in microenvironment of inflammation. Moreover, PCs can re-establish immunological tolerance in the OVA-immunized mice. Thus, our findings provide a new strategy for colitis therapy and could be of importance in additional exploration of other inflammation and autoimmune diseases therapy.

In this study, one of the commonly used MXene (Ti3C2Tx) and β nucleated isotactic polypropylene (β-iPP)/MXene composites of different compositions were fabricated. The effects of MXene on non-isothermal crystallization and polymorphic behavior of β-iPP/MXene composites were comparatively studied. The non-isothermal crystallization kinetics indicates that for all samples, the lower cooling rates promote composites to crystallize at higher temperatures. When MXene and β-Nucleating agent (β-NA) are added separately, the crystallization temperature of composites shifts towards higher temperatures at all cooling rates. When MXene and β-NA are added simultaneously, the composite shows different cooling rate dependence, and the effects of improving crystallization temperatures is more obvious under rapid cooling. The activation energy of four samples iPP, iPP/MXene, iPP/β-NA, and iPP/MXene/β-NA were -167.5, -185.5, -233.8, and -218.1 kJ/mol respectively, which agree with the variation tendency of crystallization temperatures. The polymorphic behavior analysis obtained from Differential Scanning calorimetry (DSC) is affected by two factors: the ability to form β-crystals and the thermal stability of β-crystals. Because β-crystals tend to recrystallize to α-crystals below a critical temperature, to eliminate the effect of β-α recrystallization, the melting curves at end temperatures Tend = 50 °C and Tend = 100 °C are comparatively studied. The results show that more thermally unstable β-crystals would participate in β-α recrystallization with higher cooling rates. Moreover, thermal stability of β-crystals is improved by adding MXene. To further verify these findings, samples of three different thermal conditions were synthesized and analyzed by DSC, X-Ray Diffraction (XRD), and Polarized Light Optical Microscopy (PLOM), and the results were consistent with the above findings. New understandings of synthesizing β-iPP/MXene composites with adjustable morphologies and polymorphic behavior were proposed.

Medical apparatus and instruments, such as vascular grafts, are first exposed to blood when they are implanted. Therefore, blood compatibility is considered to be the critical issue when constructing a vascular graft. In this regard, the coating method is verified to be an effective and simple approach to improve the blood compatibility as well as prevent the grafts from blood leakage. In this study, polyester fabric is chosen as the substrate to provide excellent mechanical properties while a coating layer of polyurethane is introduced to prevent the blood leakage. Furthermore, gelatin is coated on the substrate to mimic the native extracellular matrix together with the improvement of biocompatibility. XPS and FTIR analysis are performed for elemental and group analysis to determine the successful coating of polyurethane and gelatin on the polyester fabrics. In terms of blood compatibility, hemolysis and platelet adhesion are measured to investigate the anticoagulation performance. In vitro cell experiments also indicate that endothelial cells show good proliferation and morphology on the polyester fabric modified with such coating layers. Taken together, such polyester fabric coated with polyurethane and gelatin layers would have a promising potential in constructing vascular grafts with expected blood compatibility and biocompatibility without destroying the basic mechanical requirements for vascular applications.

Environmentally sound composites reinforced with natural fibers or particles interest many researchers and engineers due to their great potential to substitute the traditional composites reinforced with glass fibers. However, the sensitivity of natural fiber-reinforced composites to water has limited their applications. In this paper, wood powder-reinforced polypropylene composites (WPCs) with various wood content were prepared and subjected to water absorption tests to study the water absorption procedure and the effect of water absorbed in the specimens on the mechanical properties. Water soaking tests were carried out by immersion of composite specimens in a container of distilled water maintained at three different temperatures, 23, 60 and 80 °C. The results showed that the moisture absorption content was related to wood powder percentage and they had a positive relationship. The transfer process of water molecules in the sample was found to follow the Fickian model and the diffusion constant increased with elevated water temperature. In addition, tensile and bending tests of both dry and wet composite samples were conducted and the results indicated that water absorbed in composite specimens degraded their mechanical properties. The tensile strength and modulus of the composites reinforced with 15, 30, 45 wt % wood powder decreased by 5.79%, 17.2%, 32.06% and 25.31%, 33.6%, 47.3% respectively, compared with their corresponding dry specimens. The flexural strength and modulus of the composite samples exhibited a similar result. Furthermore, dynamic mechanical analysis (DMA) also confirmed that the detrimental effect of water molecules on the composite specimens.

An anomalous plasticizing effect was observed in polymer/ionic liquid (IL) solutions by applying broad range of rheological techniques. Poly(ethylene oxide)(PEO)/IL solutions exhibit stronger dynamic temperature dependence than pure PEO, which is in conflict with the knowledge that lower-Tg solvent increases the fractional free volume. For poly(methy methacrylate)(PMMA)/IL solutions, the subtle anomaly was detected from the fact that the effective glass transition temperature Tg,eff of PMMA in IL is higher than the prediction of the self-concentration model, while in conventional polymer solutions, Tg,eff follows the original Fox equation. Observations in both solutions reveal retarded segmental dynamics, consistent with a recent simulation result (Macromolecules, 2018, 51, 5336) that polymer chains wrap the IL cations by hydrogen bonding interactions and the segmental unwrapping delays their relaxation. Start-up shear and nonlinear stress relaxation tests of polymer/IL solutions follow a universal nonlinear rheological behavior as polymer melts and solutions, indicating that the segment-cation interaction is not strong enough to influence the nonlinear chain orientation and stretch. The present work may arouse the further theoretical, experimental, and simulation interests in interpreting the effect of complex polymer-IL interaction on the dynamics of polymer/IL solutions.

Despite the initial evidence on the role of zinc and zinc transporters in cancer prevention, little attention has been paid to the zinc-derived compounds. In the present work, we reported a strategy to prepare a kind of zinc-releasing container with enhanced biocompatibility and release dynamics using ZIF-8 nanocrystals as the sacrificial templates. Transmission electron microscopy (TEM) analysis demonstrated that the ZIF-8 nanocrystals were gradually etched out in the aqueous media within 48 h, resulting in hollow nanocapsules. Notably, we found the self-polymerization of dopamine can form nanoshells around the ZIF-8 nanocrystals, which served as a type of functional membranes during the release of zinc. More interestingly, PDA@ZIF-8⁻based nanohybrids expressed stronger inhibition to the cancer cell growth, which implied that the nanohybrids could be a drug carrier for chemotherapy. This study broadens the biomedical application of ZIF-8 and also provides a versatile strategy toward the development of multifunctional delivery system.

Hydrogels can be utilized to extract energy from salinity gradients when river water mixes with seawater. Saline-sensitive hydrogels exhibit a reversible swelling/shrinking process when they are, alternately, exposed to fresh and saline water. We present a comparison of several poly(acrylic acid)-based hydrogels, including poly(acrylic acid) (PAA), poly(acrylic acid-co-vinylsulfonic acid) (PAA/PVSA), and poly(4-styrenessulfonic acid-co-maleic acid) interpenetrated in a poly(acrylic acid) network (PAA/PSSA-MA). The hydrogels were synthesized by free radical polymerization, copolymerization, and by semi-IPN (interpenetrating polymer network). The hydrogels were placed in a piston-like system to measure the recovered energy. Semi-IPN hydrogels exhibit a much higher recovered energy compared to the copolymer and PAA hydrogel. The recovered energy of 60 g swollen gel was up to 4 J for the PAA/PSSA-MA hydrogel. The obtained energy per gram dried gel was up to 13.3 J/g. The swelling volume of the hydrogels was maintained for 30 cycles without decline in recovered energy.

Allosteric communication between distant parts of proteins controls many cellular functions, in which metal ions are widely utilized as effectors to trigger the allosteric cascade. Due to the involvement of strong coordination interactions, the energy landscape dictating the metal ion binding is intrinsically rugged. How metal ions achieve fast binding by overcoming the landscape ruggedness and thereby efficiently mediate protein allostery is elusive. By performing molecular dynamics simulations for the Ca2+ binding mediated allostery of the calmodulin (CaM) domains, each containing two Ca2+ binding helix-loop-helix motifs (EF-hands), we revealed the key role of water-bridged interactions in Ca2+ binding and protein allostery. The bridging water molecules between Ca2+ and binding residue reduces the ruggedness of ligand exchange landscape by acting as a lubricant, facilitating the Ca2+ coupled protein allostery. Calcium-induced rotation of the helices in the EF-hands, with the hydrophobic core serving as the pivot, leads to exposure of hydrophobic sites for target binding. Intriguingly, despite being structurally similar, the response of the two symmetrically arranged EF-hands upon Ca2+ binding is asymmetric. Breakage of symmetry is needed for efficient allosteric communication between the EF-hands. The key roles that water molecules play in driving allosteric transitions are likely to be general in other metal ion mediated protein allostery.

Coronavirus disease 2019 (COVID-19) is caused by a novel coronavirus (SARS-CoV-2) and represents the causative agent of a potentially fatal disease that is a public health emergency of international concern. Coronaviruses, including SARS-CoV-2, encode an envelope (E) protein, which is a small, hydrophobic membrane protein; the E protein of SARS-CoV-2 shares a high level of homology with severe acute respiratory syndrome coronavirus (SARS-CoV). In this study, we provide insights into the function of the SARS-CoV-2 E protein channel and the ion and water permeation mechanisms using a combination of in silico methods. Based on our results, the pentameric E protein promotes the penetration of cation ions through the channel. An analysis of the potential mean force (PMF), pore radius and diffusion coefficient reveals that Leu10 and Phe19 are the hydrophobic gates of the channel. In addition, the pore exhibits a clear wetting/dewetting transition with cation selectivity under transmembrane voltage, indicating that it is a hydrophobic voltage-dependent channel. Overall, these results provide structure-based insights and molecular dynamic information that are needed to understand the regulatory mechanisms of ion permeability in the pentameric SARS-CoV-2 E protein channel.

The interactions between chiral molecules and cell membranes have attracted more and more attention in recent decades, due to their importance in molecular science and medical applications. It is observed that some peptides composed of different chiral amino acids may have distinct interactions with a membrane. How does the membrane exhibit a selective behavior related to the chirality of the peptides? Microscopically, the interactions between the peptides and the membrane are poorly understood. In this work, we study the interactions between an amphipathic peptide (C6) and POPC membrane with simulations. The kinetics and thermodynamics of peptide enantiomers during the adsorption to the membrane are characterized with direct simulations and umbrella sampling. It is observed that there are slow kinetics for the peptide composed of D-type amino acids. Along the observed pathways, the free energy landscapes are determined with umbrella sampling techniques. A free-energy barrier for the peptide composed of D-amino acids is observed, which is consistent with the kinetic observations. The results indicate the concurrent adsorption and rotation of the peptide helix. The local interactions between the peptides and the membrane are examined in detail, including the contact interactions between the peptides and the membrane, and the distributions of the lipids around the peptide. There are observable differences of the local interactions for the cases related to different peptide enantiomers. These results further demonstrate the importance of the rotation of peptide helix during the adsorption. More interestingly, all these kinetic differences between peptide enantiomers can be explained based on the conformations of the residue Trp and interactions between Trp and lipid molecules. These results give us a molecular understanding of the mechanism of the chirality-dependent peptide-membrane interactions, and may provide clues to designing systems which are sensitive to the chirality of membranes.

How RNA sequences fold to specific tertiary structures is one of the key problems for understanding their dynamics and functions. Here, we study the folding process of an H-type RNA pseudoknot by performing a large-scale all-atom MD simulation and bias-exchange metadynamics. The folding free energy landscapes are obtained and several folding intermediates are identified. It is suggested that the folding occurs via multiple mechanisms, including a step-wise mechanism starting either from the first helix or the second, and a cooperative mechanism with both helices forming simultaneously. Despite of the multiple mechanism nature, the ensemble folding kinetics estimated from a Markov state model is single-exponential. It is also found that the correlation between folding and binding of metal ions is significant, and the bound ions mediate long-range interactions in the intermediate structures. Non-native interactions are found to be dominant in the unfolded state and also present in some intermediates, possibly hinder the folding process of the RNA.

The catalytic cycle of the enzyme adenylate kinase involves large conformational motions between open and closed states. A previous single-molecule experiment showed that substrate binding tends to accelerate both the opening and the closing rates and that a single turnover event often involves multiple rounds of conformational switching. In this work, we showed that the repeated conformational transitions of adenylate kinase are essential for the relaxation of incorrectly bound substrates into the catalytically competent conformation by combining all-atom and coarse-grained molecular simulations. In addition, free energy calculations based on all-atom and coarse-grained models demonstrated that the enzyme with incorrectly bound substrates has much a lower free energy barrier for domain opening compared to that with the correct substrate conformation, which may explain the the acceleration of the domain opening rate by substrate binding. The results of this work provide mechanistic understanding to previous experimental observations and shed light onto the interplay between conformational dynamics and enzyme catalysis.

Variations in the sugar component (ribose or deoxyribose) and the nature of the phosphodiester linkage (3'-5' or 2'-5' orientation) have been a challenge for genetic information transfer from the very beginning of evolution. RNA polymerase II (pol II) governs the transcription of DNA into precursor mRNA in all eukaryotic cells. How pol II recognizes DNA template backbone (phosphodiester linkage and sugar) and whether it tolerates the backbone heterogeneity remain elusive. Such knowledge is not only important for elucidating the chemical basis of transcriptional fidelity but also provides new insights into molecular evolution. In this study, we systematically and quantitatively investigated pol II transcriptional behaviors through different template backbone variants. We revealed that pol II can well tolerate and bypass sugar heterogeneity sites at the template but stalls at phosphodiester linkage heterogeneity sites. The distinct impacts of these two backbone components on pol II transcription reveal the molecular basis of template recognition during pol II transcription and provide the evolutionary insight from the RNA world to the contemporary 'imperfect' DNA world. In addition, our results also reveal the transcriptional consequences from ribose-containing genomic DNA.

Mussels can strongly adhere to hydrophilic minerals in sea habitats by secreting adhesive proteins. The adhesion ability of these proteins is often attributed to the presence of Dopa derived from posttranslational modification of Tyr, whereas the contribution of structural feature is overlooked. It remains largely unknown how adhesive proteins overcome the surface-bound water layer to establish underwater adhesion. Here, we use molecular dynamics simulations to probe the conformations of adhesive protein Pvfp-5β and its salt-tolerant underwater adhesion on superhydrophilic mica. Dopa and positively charged basic residues form pairs, in this intrinsically disordered protein, and these residue pairs can lead to firm surface binding. Our simulations further suggest that the unmodified Tyr shows similar functions on surface adhesion by forming pairing structure with a positively charged residue. We confirm the presence of these residue pairs and verify the strong binding ability of unmodified proteins using nuclear magnetic resonance spectroscopy and lap shear tests.

Nonviral gene therapy holds great promise but has not delivered treatments for clinical application to date. Lack of safe and efficient gene delivery vectors is the major hurdle. Among nonviral gene delivery vectors, poly(β-amino ester)s are one of the most versatile candidates because of their wide monomer availability, high polymer flexibility, and superior gene transfection performance both in vitro and in vivo. However, to date, all research has been focused on vectors with a linear structure. A well-accepted view is that dendritic or branched polymers have greater potential as gene delivery vectors because of their three-dimensional structure and multiple terminal groups. Nevertheless, to date, the synthesis of dendritic or branched polymers has been proven to be a well-known challenge. We report the design and synthesis of highly branched poly(β-amino ester)s (HPAEs) via a one-pot "A2 + B3 + C2"-type Michael addition approach and evaluate their potential as gene delivery vectors. We find that the branched structure can significantly enhance the transfection efficiency of poly(β-amino ester)s: Up to an 8521-fold enhancement in transfection efficiency was observed across 12 cell types ranging from cell lines, primary cells, to stem cells, over their corresponding linear poly(β-amino ester)s (LPAEs) and the commercial transfection reagents polyethyleneimine, SuperFect, and Lipofectamine 2000. Moreover, we further demonstrate that HPAEs can correct genetic defects in vivo using a recessive dystrophic epidermolysis bullosa graft mouse model. Our findings prove that the A2 + B3 + C2 approach is highly generalizable and flexible for the design and synthesis of HPAEs, which cannot be achieved by the conventional polymerization approach; HPAEs are more efficient vectors in gene transfection than the corresponding LPAEs. This provides valuable insight into the development and applications of nonviral gene delivery and demonstrates great prospect for their translation to a clinical environment.

RNA polymerase II (Pol II) surveils the genome, pausing as it encounters DNA lesions and base modifications and initiating signals for DNA repair among other important regulatory events. Recent work suggests that Pol II pauses at 5-carboxycytosine (5caC), an epigenetic modification of cytosine, because of a specific hydrogen bond between the carboxyl group of 5caC and a specific residue in fork loop 3 of Pol II. This hydrogen bond compromises productive NTP binding and slows down elongation. Apart from this specific interaction, the carboxyl group of 5caC can potentially interact with numerous charged residues in the cleft of Pol II. However, it is not clear how other interactions between Pol II and 5caC contribute to pausing. In this study, we use Markov state models (a type of kinetic network models) built from extensive molecular dynamics simulations to comprehensively study the impact of 5caC on Pol II translocation. We describe two translocation intermediates with specific interactions that prevent the template base from loading into the Pol II active site. In addition to the previously observed state with 5caC constrained by fork loop 3, we discovered a new intermediate state with a hydrogen bond between 5caC and fork loop 2. Surprisingly, we find that 5caC may curb translocation by suppressing kinking of the helix bordering the active site (the bridge helix) because its high flexibility is critical to translocation. Our work provides new insights into how epigenetic modifications of genomic DNA can modulate Pol II translocation, inducing pauses in transcription.

As a zero-dimensional (0D) nanomaterial, graphene quantum dot (GQD) has a unique physical structure and electrochemical properties, which has been widely used in biomedical fields, such as bioimaging, biosensor, drug delivery, etc. Its biological safety and potential cytotoxicity to human and animal cells have become a growing concern in recent years. In particular, the potential DNA structure damage caused by GQD is of great importance but still obscure. In this study, molecular dynamics (MD) simulation was used to investigate the adsorption behavior and the structural changes of single-stranded (ssDNA) and double-stranded DNA (dsDNA) on the surfaces of GQDs with different sizes and oxidation. Our results showed that ssDNA can strongly adsorb and lay flat on the surface of GQDs and graphene oxide quantum dots (GOQDs), whereas dsDNA was preferentially oriented vertically on both surfaces. With the increase of GQDs size, more structural change of adsorbed ssDNA and dsDNA could be found, while the size effect of GOQD on the structure of ssDNA and dsDNA is not significant. These findings may help to improve the understanding of GQD biocompatibility and potential applications of GQD in the biomedical field.

Gel syneresis is a common problem in gel treatment for oil recovery applications. In this study, a stable gel was prepared in a soft brine by using a water-soluble phenolic resin as a crosslinker, nanoparticles as a stabilizer, and partially hydrolyzed polyacrylamide (HPAM) or copolymers with different contents of 2-acrylamido-2-methylpropane sulfonic acid (AMPS) groups as polymers. The syneresis behavior of the gels formed in a soft brine was evaluated upon aging in hard brines. The results show that when the salinity of the hard brine is lower than 30,000 mg/L, the gel expands, and its strength decreases; when the salinity of the hard brine is higher than 50,000 mg/L, the gel exhibits syneresis, and its strength increases. The effects of various influencing factors on the gel syneresis behavior were also evaluated. It was found that optimizing the polymer structure and adding nanoparticles can effectively overcome gel syneresis and enhance gel stability. Based on the research described in this paper, some proposals for designing salt-resistant polymer gels are presented.