Inhibition of Structural Transformation and Surface Lattice Oxygen Activity for Excellent Stability Li-Rich Mn-Based Layered Oxides.

Li-rich Mn-based layered oxides (LLOs) are one of the most promising cathode materials, which have exceptional anionic redox activity and a capacity that surpasses 250 mA h/g. However, the change from a layered structure to a spinel structure and unstable anionic redox are accompanied by voltage attenuation, poor rate performance, and problematic capacity. The technique of stabilizing the crystal structure and reducing the surface oxygen activity is proposed in this paper. A coating layer and highly concentrated oxygen vacancies are developed on the material's surface, according to scanning electron microscopy, transmission electron microscopy, and X-ray photoelectron spectroscopy. In situ EIS shows that structural transformation and oxygen release are inhibited during the first charge and discharge. Optimized 3@LRMA has an average attenuation voltage of 0.55 mV per cycle (vs 1.7 mV) and a capacity retention rate of 93.4% after 200 cycles (vs 52.8%). Postmortem analysis indicates that the successful doping of Al ions into the crystal structure effectively inhibits the structural alteration of the cycling process. The addition of oxygen vacancies reduces the surface lattice's redox activity. Additionally, surface structure deterioration is successfully halted by N- and Cl-doped carbon coating. This finding highlights the significance of lowering the surface lattice oxygen activity and preventing structural alteration, and it offers a workable solution to increase the LLO stability.

Evaluation of monovalent and bivalent vaccines against lethal Enterovirus 71 and Coxsackievirus A16 infection in newborn mice.

Enterovirus 71 (EV71) and Coxsackievirus A16 (CVA16) have caused severe epidemics of hand, foot and mouth disease (HFMD) in the Asia Pacific in recent years, particularly in infants and young children. This disease has become a serious public health problem, as no vaccines or antiviral drugs have been approved for EV71 and CA16 infections. In this study, we compared four monovalent vaccines, including formalin-inactivated EV71 virus (iEV71), EV71 virus-like particles (VLPs) (vEV71), formalin-inactivated CVA16 virus (iCVA16) and CVA16 VLPs (vCVA16), along with two bivalent vaccines, including equivalent doses of formalin-inactivated EV71+CVA16 virus (iEV71+iCVA16) and EV71+CVA16 VLPs (vEV71+vCVA16). The IgG titers and neutralization antibodies titers demonstrated that there are no immune interference exists between the two immunogens of EV71 and CVA16. IgG subclass isotyping revealed that IgG1 and IgG2b were induced primarily in all vaccine groups. Furthermore, cross-neutralization antibodies were elicited in mouse sera against other sub-genotypes of EV71 and CVA16. In vivo challenge experiments showed that the immune sera from vaccinated animals could confer passive protection to newborn mice against lethal challenge with 14 LD50 of EV71 and 50 LD50 of CVA16. Our results indicated that bivalent vaccination is promising for HFMD vaccine development. With the advantage of having a better safety profile than inactivated virus vaccines, VLPs should be used to combine both EV71 and CVA16 antigens as a candidate vaccine for prevention of HFMD virus transmission.

Identification of luteolin as enterovirus 71 and coxsackievirus A16 inhibitors through reporter viruses and cell viability-based screening.

Hand, foot and mouth disease (HFMD) is a common pediatric illness mainly caused by infection with enterovirus 71 (EV71) and coxsackievirus A16 (CA16). The frequent HFMD outbreaks have become a serious public health problem. Currently, no vaccine or antiviral drug for EV71/CA16 infections has been approved. In this study, a two-step screening platform consisting of reporter virus-based assays and cell viability­based assays was developed to identify potential inhibitors of EV71/CA16 infection. Two types of reporter viruses, a pseudovirus containing luciferase-encoding RNA replicons encapsidated by viral capsid proteins and a full-length reporter virus containing enhanced green fluorescent protein, were used for primary screening of 400 highly purified natural compounds. Thereafter, a cell viability-based secondary screen was performed for the identified hits to confirm their antiviral activities. Three compounds (luteolin, galangin, and quercetin) were identified, among which luteolin exhibited the most potent inhibition of viral infection. In the cell viability assay and plaque reduction assay, luteolin showed similar 50% effective concentration (EC50) values of about 10 µM. Luteolin targeted the post-attachment stage of EV71 and CA16 infection by inhibiting viral RNA replication. This study suggests that luteolin may serve as a lead compound to develop potent anti-EV71 and CA16 drugs.

Analysis of cross-reactive neutralizing antibodies in human HFMD serum with an EV71 pseudovirus-based assay.

Hand, foot and mouth disease, associated with enterovirus 71 (EV71) infections, has recently become an important public health issue throughout the world. Serum neutralizing antibodies are major indicators of EV71 infection and protective immunity. However, the potential for cross-reactivity of neutralizing antibodies for different EV71 genotypes and subgenotypes is unclear. Here we measured the cross-reactive neutralizing antibody titers against EV71 of different genotypes or subgenotypes in sera collected from EV71-infected children and vaccine-inoculated children in a phase III clinical trial (ClinicalTrials.gov Identifier: NCT01636245) using a new pseudovirus-based neutralization assay. Antibodies induced by EV71-C4a were cross-reactive for different EV71 genotypes, demonstrating that C4a is a good candidate strain for an EV71 vaccine. Our study also demonstrated that this new assay is practical for analyses of clinical samples from epidemiological and vaccine studies.

A novel disulfide-stabilized single-chain variable antibody fragment against rabies virus G protein with enhanced in vivo neutralizing potency.

Rabies is a fatal infectious disease requiring efficient protection provided by post-exposure prophylaxis (PEP) with rabies immunoglobulin (RIG). The single-chain Fv fragment (scFv) is a small engineered antigen binding protein derived from antibody variable heavy (V(H)) and light (V(L)) chains. This novel antibody format may potentially replace the current application of RIG to detect and neutralize rabies virus (RV). However, the broad use of scFvs is confined by their generally low stability. In this study, a scFv (FV57) was constructed based on the monoclonal antibody, MAB57, against RV. To enhance its stability and neutralizing potency, a disulfide-stabilized scFv, ds-FV57, was also derived by introduction of cysteines at V(H)44 and V(L)100. Furthermore, the cysteine at V(L)85 of ds-FV57 was mutated to serine to construct ds-FV57(VL85Ser) in order to avoid potential mis-formed disulfide bonds which would alter the affinity of the scFv. The stability and activity of all three proteins expressed in Escherichia coli were evaluated. All of the constructed scFvs could provide efficient protection against RV infection both in vivo and in vitro. However, the stability of ds-FV57(VL85Ser) was notably improved, and its in vitro neutralizing potency against RV infection was enhanced. Our findings from these stabilization modifications support the feasibility of developing scFvs for PEP treatment of rabies.

Thermal stability and structural variations of survivin and its deletants in aqueous solution as revealed by spectroscopy.

Survivin exists as a homodimeric conformation to act as a suppressor of apoptosis in organisms. Previously, we found that the deletants with truncations of N-terminal residues up to Arg18 lost the binding ability to Smac/DIABLO but not the binding force of homodimers. In order to establish the relationship between function and structural stability, thermal unfolding of SurF and its deletants in buffer have been studied in the present paper. The fluorescent results indicated that with the removal of the N-terminus, the thermal stability of the tertiary structure dropped vigorously, especially for SurΔN18. However, using circular dichroism (CD) spectroscopy, we observed that the main unfolding of the secondary structures was not affected very much with N-terminus deletion. Fourier transform infrared (FT-IR) spectroscopy and two-dimensional (2D) correlation analysis were further used to provide structural information that occurred in the main transitions, which were associated with conformational changes of several ß-components and α-helix, followed by the gain of some aggregations and random coils at high temperature. In addition, more aggregates were found to form for the longer N-terminal deletants during the main unfolding.

N-terminal deletion effects of human survivin on dimerization and binding to Smac/DIABLO in vitro.

Survivin, as an apoptosis suppressor, exists as a homodimer interfacing at the N-terminal portion (residues 6-13) of its baculovirus IAP repeat (BIR) domain and a linker segment (residues 89-102). Here we expressed full-length human Survivin (SurF) and a series of its mutants, SurΔN7, SurΔN13, and SurΔN18 with significant truncations of the N-terminus, all of which could still dimerize in solution. Single-molecule force spectroscopy (SMFS) was used to quantitate the unbinding forces of full-length and the mutant homodimers and revealed that the N-terminal residues up to Arg18 were not essential for dimerization. Meanwhile, the binding of SurΔN7 to Smac/DIABLO determined by ELISA was as efficient as the wild-type, but that of SurΔN13 was significantly reduced, and that of SurΔN18 was completely lost. Together, these findings provide direct evidence that the N-terminal sequence of Survivin is not critical for dimer formation but may contribute to correct folding and function of BIR.