Coexistence of adeno-associated virus 2 with adenovirus 18 or herpesvirus may be associated with severe lingual papillomatosis in an immunocompromised individual.

Next-generation sequencing (NGS) has significantly improved the accuracy and efficiency of pathogen diagnosis for a wide range of diseases. In this study, viral metagenomics analysis was conducted on fecal and tissue samples from a 13-year-old recipient of hematopoietic stem cell transplantation (HSCT) afflicted with severe lingual papillomatosis. The analysis revealed a high abundance of adeno-associated virus 2 (AAV2), alongside potential helper viruses, herpesvirus type 1 (HSV-1), and the uncommon adenovirus serotype 18 (AdV18). Although a direct causal relationship was not definitively established, the concurrence of these viruses indicated a plausible link to the development of severe lingual papillomatosis in immunocompromised individuals. Notably, the study generated a complete genome sequence of AdV18, offering insights into adenovirus genetic variability, origin, and pathogenicity. Noteworthy findings include three amino acid substitutions in the polymerase and one in the hexon, distinguishing them from previously published strains of AdV18. Phylogenetic analysis unveiled a close relationship between both the polymerase and hexon regions of AdV18 in our study and previously reported AdV18 sequences. This study underscores the pivotal role of comprehensive viral scrutiny in elucidating infections among HSCT patients with lingual papillomatosis.

Suggestion of GLYAT gene underlying variation of bone size and body lean mass as revealed by a bivariate genome-wide association study.

Bone and muscle, two major tissue types of musculoskeletal system, have strong genetic determination. Abnormality in bone and/or muscle may cause musculoskeletal diseases such as osteoporosis and sarcopenia. Bone size phenotypes (BSPs), such as hip bone size (HBS), appendicular bone size (ABS), are genetically correlated with body lean mass (mainly muscle mass). However, the specific genes shared by these phenotypes are largely unknown. In this study, we aimed to identify the specific genes with pleiotropic effects on BSPs and appendicular lean mass (ALM). We performed a bivariate genome-wide association study (GWAS) by analyzing ~690,000 SNPs in 1,627 unrelated Han Chinese adults (802 males and 825 females) followed by a replication study in 2,286 unrelated US Caucasians (558 males and 1,728 females). We identified 14 interesting single nucleotide polymorphisms (SNPs) that may contribute to variation of both BSPs and ALM, with p values <10(-6) in discovery stage. Among them, the association of three SNPs (rs2507838, rs7116722, and rs11826261) in/near GLYAT (glycine-N-acyltransferase) gene was replicated in US Caucasians, with p values ranging from 1.89 × 10(-3) to 3.71 × 10(-4) for ALM-ABS, from 5.14 × 10(-3) to 1.11 × 10(-2) for ALM-HBS, respectively. Meta-analyses yielded stronger association signals for rs2507838, rs7116722, and rs11826261, with pooled p values of 1.68 × 10(-8), 7.94 × 10(-8), 6.80 × 10(-8) for ALB-ABS and 1.22 × 10(-4), 9.85 × 10(-5), 3.96 × 10(-4) for ALM-HBS, respectively. Haplotype allele ATA based on these three SNPs was also associated with ALM-HBS and ALM-ABS in both discovery and replication samples. Interestingly, GLYAT was previously found to be essential to glucose metabolism and energy metabolism, suggesting the gene's dual role in both bone development and muscle growth. Our findings, together with the prior biological evidence, suggest the importance of GLYAT gene in co-regulation of bone phenotypes and body lean mass.

Intranasal immunization with a helper-dependent adenoviral vector expressing the codon-optimized fusion glycoprotein of human respiratory syncytial virus elicits protective immunity in BALB/c mice.

BACKGROUND: Human respiratory syncytial virus (RSV) is a serious pediatric pathogen of the lower respiratory tract. Currently, there is no clinically approved vaccine against RSV infection. Recent studies have shown that helper-dependent adenoviral (HDAd) vectors may represent effective and safe vaccine vectors. However, viral challenge has not been investigated following mucosal vaccination with HDAd vector vaccines. METHODS: To explore the role played by HDAd as an intranasally administered RSV vaccine vector, we constructed a HDAd vector encoding the codon optimized fusion glycoprotein (Fsyn) of RSV, designated HDAd-Fsyn, and delivered intranasally HDAd-Fsyn to mice. RESULTS: RSV-specific humoral and cellular immune responses were generated in BALB/c mice, and serum IgG with neutralizing activity was significantly elevated after a homologous boost with intranasal (i.n.) application of HDAd-Fsyn. Humoral immune responses could be measured even 14 weeks after a single immunization. Immunization with i.n. HDAd-Fsyn led to effective protection against RSV infection on challenge. CONCLUSION: The results indicate that HDAd-Fsyn can induce powerful systemic immunity against subsequent i.n. RSV challenge in a mouse model and is a promising candidate vaccine against RSV infection.

Tracking of Mutational Signature of SARS-CoV-2 Omicron on Distinct Continents and Little Difference was Found.

The Omicron variant is currently ravaging the world, raising serious concern globally. Monitoring genomic variations and determining their influence on biological features are critical for tracing its ongoing transmission and facilitating effective measures. Based on large-scale sequences from different continents, this study found that: (i) The genetic diversity of Omicron is much lower than that of the Delta variant. Still, eight deletions (Del 1-8) and 1 insertion, as well as 130 SNPs, were detected on the Omicron genomes, with two deletions (Del 3 and 4) and 38 SNPs commonly detected on all continents and exhibiting high-occurring frequencies. (ii) Four groups of tightly linked SNPs (linkage I-IV) were detected, among which linkage I, containing 38 SNPs, with 6 located in the RBD, increased its occurring frequency remarkably over time. (iii) The third codons of the Omicron shouldered the most mutation pressures, while the second codons presented the least flexibility. (iv) Four major mutants with amino acid substitutions in the RBD were detected, and further structural analysis suggested that the substitutions did not alter the viral receptor binding ability greatly. It was inferred that though the Omicron genome harbored great changes in antigenicity and remarkable ability to evade immunity, it was immune-pressure selected. This study tracked mutational signatures of Omicron variant and the potential biological significance of the SNPs, and the linkages await further functional verification.

Comparative analysis of human respiratory syncytial virus evolutionary patterns during the COVID-19 pandemic and pre-pandemic periods.

The COVID-19 pandemic has resulted in the implementation of strict mitigation measures that have impacted the transmission dynamics of human respiratory syncytial virus (HRSV). The measures also have the potential to influence the evolutionary patterns of the virus. In this study, we conducted a comprehensive analysis comparing genomic variations and evolving characteristics of its neutralizing antigens, specifically F and G proteins, before and during the COVID-19 pandemic. Our findings showed that both HRSV A and B exhibited an overall chronological evolutionary pattern. For the sequences obtained during the pandemic period (2019-2022), we observed that the HRSV A distributed in A23 genotype, but formed into three subclusters; whereas the HRSV B sequences were relatively concentrated within genotype B6. Additionally, multiple positively selected sites were detected on F and G proteins but none were located at neutralizing antigenic sites of the F protein. Notably, amino acids within antigenic site III, IV, and V of F protein remained strictly conserved, while some substitutions occurred over time on antigenic site Ø, I, II and VIII; substitution S389P on antigenic site I of HRSV B occurred during the pandemic period with nearly 50% frequency. However, further analysis revealed no substitutions have altered the structural conformations of the antigenic sites, the vial antigenicity has not been changed. We inferred that the intensive public health interventions during the COVID-19 pandemic did not affect the evolutionary mode of HRSV.

The protective immunity induced by intranasally inoculated serotype 63 chimpanzee adenovirus vector expressing human respiratory syncytial virus prefusion fusion glycoprotein in BALB/c mice.

Human respiratory syncytial virus (RSV) is a ubiquitous pediatric pathogen causing serious lower respiratory tract disease worldwide. No licensed vaccine is currently available. In this work, the coding gene for mDS-Dav1, the full-length and prefusion conformation RSV fusion glycoprotein (F), was designed by introducing the stabilized prefusion F (preF) mutations from DS-Cav1 into the encoding gene of wild-type RSV (wtRSV) F protein. The recombinant adenovirus encoding mDS-Cav1, rChAd63-mDS-Cav1, was constructed based on serotype 63 chimpanzee adenovirus vector and characterized in vitro. After immunizing mice via intranasal route, the rChAd63-mDS-Cav1 induced enhanced neutralizing antibody and F-specific CD8+ T cell responses as well as good immune protection against RSV challenge with the absence of enhanced RSV disease (ERD) in BALB/c mice. The results indicate that rChAd63-mDS-Cav1 is a promising mucosal vaccine candidate against RSV infection and warrants further development.

Genetic diversity and molecular evolution of human respiratory syncytial virus A and B.

Human respiratory syncytial viruses (RSVs) are classified into two major groups (A and B) based on antigenic differences in the G glycoprotein. To investigate circulating characteristics and phylodynamic history of RSV, we analyzed the genetic variability and evolutionary pattern of RSVs from 1977 to 2019 in this study. The results revealed that there was no recombination event of intergroup. Single nucleotide polymorphisms (SNPs) were observed through the genome with the highest occurrence rate in the G gene. Five and six sites in G protein of RSV-A and RSV-B, respectively, were further identified with a strong positive selection. The mean evolutionary rates for RSV-A and -B were estimated to be 1.48 × 10-3 and 1.92 × 10-3 nucleotide substitutions/site/year, respectively. The Bayesian skyline plot showed a constant population size of RSV-A and a sharp expansion of population size of RSV-B since 2005, and an obvious decrease 5 years later, then became stable again. The total population size of RSVs showed a similar tendency to that of RSV-B. Time-scaled phylogeny suggested a temporal specificity of the RSV-genotypes. Monitoring nucleotide changes and analyzing evolution pattern for RSVs could give valuable insights for vaccine and therapy strategies against RSV infection.

Biological Significance of the Genomic Variation and Structural Dynamics of SARS-CoV-2 B.1.617.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants have been emerging and circulating globally since the start of the COVID-19 pandemic, of which B.1.617 lineage that was first reported in India at the end of 2020, soon became predominant. Tracing genomic variations and understanding their impact on the viral properties are the foundations for the vaccine and drug development and for the mitigation measures to be taken or lifted. In this study, 1,051 near-complete genomes and 1,559 spike (S) sequences belonging to the B.1.617 were analyzed. A genome-wide spread of single nucleotide polymorphisms (SNPs) was identified. Of the high frequency mutations identified, 61% (11/18) involved structural proteins, despite two third of the viral genome encoding nonstructural proteins. There were 22 positive selection sites, mostly distributed across the S protein, of which 16 were led by non-C to U transition and should be of a special attention. Haplotype network revealed that a large number of daughter haplotypes were continually derived throughout the pandemic, of which H177, H181 H219 and H286 from the ancestor haplotype H176 of B.1.617.2 were widely prevalent. Besides the well known substitutions of L452R, P681R and deletions of E156 and F157, as well as the potential biological significance, structural analysis in this study still indicated that new amino acid changes in B.1.617, such as E484Q and N501Y, had reshaped the viral bonding network, and increasingly sequenced N501Y mutant with a potential enhanced binding ability was detected in many other countries in the follow-up monitoring. Although we can't conclude the properties of all the mutants including N501Y thoroughly, it merits focusing on their spread epidemically and biologically.

Evaluation of the Safety and Immune Efficacy of Recombinant Human Respiratory Syncytial Virus Strain Long Live Attenuated Vaccine Candidates.

Human respiratory syncytial virus (RSV) infection is the leading cause of lower respiratory tract illness (LRTI), and no vaccine against LRTI has proven to be safe and effective in infants. Our study assessed attenuated recombinant RSVs as vaccine candidates to prevent RSV infection in mice. The constructed recombinant plasmids harbored (5' to 3') a T7 promoter, hammerhead ribozyme, RSV Long strain antigenomic cDNA with cold-passaged (cp) mutations or cp combined with temperature-sensitive attenuated mutations from the A2 strain (A2cpts) or further combined with SH gene deletion (A2cptsΔSH), HDV ribozyme (δ), and a T7 terminator. These vectors were subsequently co-transfected with four helper plasmids encoding N, P, L, and M2-1 viral proteins into BHK/T7-9 cells, and the recovered viruses were then passaged in Vero cells. The rescued recombinant RSVs (rRSVs) were named rRSV-Long/A2cp, rRSV-Long/A2cpts, and rRSV-Long/A2cptsΔSH, respectively, and stably passaged in vitro, without reversion to wild type (wt) at sites containing introduced mutations or deletion. Although rRSV-Long/A2cpts and rRSV-Long/A2cptsΔSH displayed  temperature-sensitive (ts) phenotype in vitro and in vivo, all rRSVs were significantly attenuated in vivo. Furthermore, BALB/c mice immunized with rRSVs produced Th1-biased immune response, resisted wtRSV infection, and were free from enhanced respiratory disease. We showed that the combination of ΔSH with attenuation (att) mutations of cpts contributed to improving att phenotype, efficacy, and gene stability of rRSV. By successfully introducing att mutations and SH gene deletion into the RSV Long parent and producing three rRSV strains, we have laid an important foundation for the development of RSV live attenuated vaccines.

Analysis of Continuous Mutation and Evolution on Circulating SARS-CoV-2.

Monitoring the mutation and evolution of the virus is important for tracing its ongoing transmission and facilitating effective vaccine development. A total of 342 complete genomic sequences of SARS-CoV-2 were analyzed in this study. Compared to the reference genome reported in December 2019, 465 mutations were found, among which, 347 occurred in only 1 sequence, while 26 occurred in more than 5 sequences. For these 26 further identified as SNPs, 14 were closely linked and were grouped into 5 profiles. Phylogenetic analysis revealed the sequences formed 2 major groups. Most of the sequences in late period (March and April) constituted the Cluster II, while the sequences before March in this study and the reported S/L and A/B/C types in previous studies were all in Cluster I. The distributions of some mutations were specific geographically or temporally, the potential effect of which on the transmission and pathogenicity of SARS-CoV-2 deserves further evaluation and monitoring. Two mutations were found in the receptor-binding domain (RBD) but outside the receptor-binding motif (RBM), indicating that mutations may only have marginal biological effects but merit further attention. The observed novel sequence divergence is of great significance to the study of the transmission, pathogenicity, and development of an effective vaccine for SARS-CoV-2.

High-throughput screening of active compounds against human respiratory syncytial virus.

Human respiratory syncytial virus (RSV) is one of the predominant pathogens causing lower respiratory tract infection in infants and young children worldwide, whereas there is so far no vaccine or drug against RSV infection for clinical use. In this work, we developed and validated a fluorescence-based high-throughput screening (HTS) assay to identify compounds active against RSV, using RSV-mGFP, a recombinant RSV encoding enhanced green fluorescent protein (EGFP). Thereafter, among 54,800 compounds used for our screen, we obtained 62 compounds active against RSV. Among these hits, azathioprine (AZA) and 6-mercaptopurine (6-MP) were identified as RSV inhibitors with half maximal inhibitory concentration (IC50) values of 6.69 ±â€¯1.41 and 3.13 ±â€¯0.98 µM, respectively. Further experiments revealed that they functioned by targeting virus transcription or/and genome replication. In conclusion, the established HTS assay is suitable to screen anti-RSV compounds, and the screened two hits of AZA and 6-MP, as potential anti-RSV agents targeting RSV genome replication/transcription, are worthy of further investigation on their anti-RSV activity in vivo.

A Built-In CpG Adjuvant in RSV F Protein DNA Vaccine Drives a Th1 Polarized and Enhanced Protective Immune Response.

Human respiratory syncytial virus (RSV) is the most significant cause of acute lower respiratory infection in children. However, there is no licensed vaccine available. Here, we investigated the effect of five or 20 copies of C-Class of CpG ODN (CpG-C) motif incorporated into a plasmid DNA vaccine encoding RSV fusion (F) glycoprotein on the vaccine-induced immune response. The addition of CpG-C motif enhanced serum binding and virus-neutralizing antibody responses in BALB/c mice immunized with the DNA vaccines. Moreover, mice vaccinated with CpG-modified vaccines, especially with the higher 20 copies, resulted in an enhanced shift toward a Th1-biased antibody and T-cell response, a decrease in pulmonary pathology and virus replication, and a decrease in weight loss after RSV challenge. This study suggests that CpG-C motif, cloned into the backbone of DNA vaccine encoding RSV F glycoprotein, functions as a built-in adjuvant capable of improving the efficacy of DNA vaccine against RSV infection.

Construction and Characterization of a Recombinant Human Respiratory Syncytial Virus Encoding Enhanced Green Fluorescence Protein for Antiviral Drug Screening Assay.

Human respiratory syncytial virus (RSV) is the single most important cause of lower respiratory tract disease in infants and young children and a major viral agent responsible for respiratory tract disease in immunosuppressed individuals and the elderly, but no vaccines and antiviral drugs are available. Herein the recombinant RSV (rRSV) encoding enhanced green fluorescence protein (EGFP, rRSV-EGFP) was constructed and the potential for screening anti-RSV drugs was investigated. The recombinant plasmid of pBRATm-rRSV-EGFP, containing T7 transcription cassette composed of T7 promoter, RSV antigenomic cDNA with EGFP gene, HDV ribozyme (δ), and T7 terminator in the order of 5' to 3', was constructed and cotransfected into BHK/T7-9 cells together with helper plasmids encoding N, P, L, and M2-1 gene, respectively. The rescued rRSV-EGFP was confirmed by increasing expression of EGFP over blind passages and by RT-PCR. rRSV-EGFP was comparable to the other two recombinant RSVs encoding red fluorescent protein (RFP, rRSV-RFP) or luciferase (Luc, rRSV-Luc) in the growth kinetic, and there was a difference in sensitivity between them for screening anti-RSV agents based on infection of HEp-2 cells. The EGFP-encoding rRSV has been constructed and rescued successfully and has the potential for high-throughput anti-RSV drug screening in vitro.

Amyloid ß-Derived Diffusible Ligands (ADDLs) Induce Abnormal Autophagy Associated with Aß Aggregation Degree.

Autophagy is disturbed in Alzheimer's disease (AD) and maintaining normal autophagy homeostasis is a new therapeutic strategy for AD treatment. Amyloid ß-derived diffusible ligands (ADDLs), the most toxic species of which are oligomeric forms of amyloid ß peptide (Aß) that originate from amyloid ß precursor protein (APP) via autophagy; however, whether ADDLs are involved in autophagy-related AD pathogenesis remains unclear. In this study, we primarily defined the specific subsets of ADDLs, A-0, A-12, A-24, and A-48, which were generated from ADDL aggregation mixtures at different time courses of assembly. The secondary structures of ADDL subsets were detected by circular dichroism (CD). Neuronal or non-neuronal cells were exposed to the subsets of ADDLs in vitro, and then, autophagic markers were detected. Our results first showed that exogenous or endogenous LC3 puncta (autophagosomes) were induced in the cytoplasm of cells exposed to ADDLs and that the LC3 puncta were the strongest with A-24 exposure. Then, the CD spectroscopy data also indicated that the proportion of α-helices decreased, whereas the proportion of ß-strands and ß-turns increased during ADDL assembly from 0 to 24 h. In addition, the quantitative Western blot data demonstrated that the ratio of LC3B-II/I was significantly increased, and SQSTM1/p62 decreased over time. Finally, our results indicated that the level of phosphorylated p70 S6 kinase (p-p70 S6 kinase), which is a substrate protein in the MTOR pathway, and the ratio of p-p70 S6 kinase/p70 S6 kinase significantly decreased following A-24 exposure. Taken together, our data suggest that ADDL-induced abnormal autophagy is correlated with Aß aggregation degree and the MTOR pathway, which might contribute to ADDL-induced AD pathogenesis.

Enhanced humoral and CD8+ T cell immunity in mice vaccinated by DNA vaccine against human respiratory syncytial virus through targeting the encoded F protein to dendritic cells.

Human respiratory syncytial virus (RSV) is the most important cause of serious lower respiratory tract infection in infants, the elderly, and the immunocompromised population. There is no licensed vaccine against RSV until now. It has been reported that targeting antigen to DEC205, a phagocytosis receptor on dendritic cells (DCs), could induce enhanced CD4+ and CD8+ T cell responses in mice. To develop RSV DNA vaccine and target the encoded antigen protein to DCs, the ectodomain of fusion glycoprotein (sF, amino acids: 23-524) of RSV was fused with anti-DEC205 single-chain Fv fragment (scDEC) and designated scDECF. Following successful expression from the recombinant plasmid of pVAX1/scDECF, the recombinant protein of scDECF was found capable of specifically binding to DEC205 receptor on CHOmDEC205 cells, and facilitating uptake of RSV F by DC2.4 cells in vitro. Furthermore, the higher levels of RSV-specific IgG antibody responses and neutralization antibody titers, as well as RSV F-specific CD8+ T cell responses were induced in mice immunized intramuscularly by pVAX1/scDECF than by the control plasmid of pVAX1/scISOF encoding sF protein fused with isotype matched control single-chain Fv fragment (scISO). Compared with pVAX1/scISOF, both the ratio of IgG2a/IgG1, >1, and the enhanced IFN-γ cytokine were induced in mice following pVAX1/scDECF immunization, which exhibited a Th1 dominant response in pVAX1/scDECF vaccinated mice. Notably, the elevated efficiency of RSV F protein bound by DCs in vivo could also be observed in mice inoculated by pVAX1/scDECF. Collectively, these results demonstrate the enhanced IgG and CD8+ T cell immune responses have been induced successfully by DNA vaccine against RSV by targeting F antigen to DCs via the DEC205 receptor, and this DC-targeting vaccine strategy merits further investigation.

A single intranasal administration of virus-like particle vaccine induces an efficient protection for mice against human respiratory syncytial virus.

Human respiratory syncytial virus (RSV) is an important pediatric pathogen causing acute viral respiratory disease in infants and young children. However, no licensed vaccines are currently available. Virus-like particles (VLPs) may bring new hope to producing RSV VLP vaccine with high immunogenicity and safety. Here, we constructed the recombinants of matrix protein (M) and fusion glycoprotein (F) of RSV, respectively into a replication-deficient first-generation adenoviral vector (FGAd), which were used to co-infect Vero cells to assemble RSV VLPs successfully. The resulting VLPs showed similar immunoreactivity and function to RSV virion in vitro. Moreover, Th1 polarized response, and effective mucosal virus-neutralizing antibody and CD8+ T-cell responses were induced by a single intranasal (i.n.) administration of RSV VLPs rather than intramuscular (i.m.) inoculation, although the comparable RSV F-specific serum IgG and long-lasting RSV-specific neutralizing antibody were detected in the mice immunized by both routes. Upon RSV challenge, VLP-immunized mice showed increased viral clearance but decreased signs of enhanced lung pathology and fewer eosinophils compared to mice immunized with formalin-inactivated RSV (FI-RSV). In addition, a single i.n. RSV VLP vaccine has the capability to induce RSV-specific long-lasting neutralizing antibody responses observable up to 15 months. Our results demonstrate that the long-term and memory immune responses in mice against RSV were induced by a single i.n. administration of RSV VLP vaccine, suggesting a successful approach of RSV VLPs as an effective and safe mucosal vaccine against RSV infection, and an applicable and qualified platform of FGAd-infected Vero cells for VLP production.

DNA vaccine encoding central conserved region of G protein induces Th1 predominant immune response and protection from RSV infection in mice.

Human respiratory syncytial virus (RSV) can cause serious infection in the lower respiratory tract, especially in infants, young children, the elderly and the immunocompromised population worldwide. Previous study demonstrated the polypeptide (amino acids 148-198) of RSV attachment (G) glycoprotein, corresponding to the central conserved region and encompassing CX3C chemokine motif, could induce antibodies and protection from RSV challenge in mice [1,2]. In this study, we evaluated the immune efficacy of the recombinant DNA vaccine of pVAX1/3G148-198 encoding RSV G protein polypeptide. RSV specific serum IgG antibodies with neutralizing activity were stimulated following prime-boost immunization of pVAX1/3G148-198 intramuscularly, and the ratio of IgG2a/IgG1 was 4.93, indicating a Th1 biased immune response. After challenged intranasally with RSV Long, the vaccinated mice showed both decreased lung RSV titers, pulmonary inflammation and body weight loss. The results suggest that pVAX1/3G148-198 DNA vaccine may be an effective RSV vaccine candidate, and deserves further exploration.

Single chain variable fragment against aß expressed in baculovirus inhibits abeta fibril elongation and promotes its disaggregation.

Alzheimer's disease (AD) is the most common form of age-related dementia, and the most urgent problem is that it is currently incurable. Amyloid-ß (Aß) peptide is believed to play a major role in the pathogenesis of AD. We previously reported that an Aß N-terminal amino acid targeting monoclonal antibody (MAb), A8, inhibits Aß fibril formation and has potential as an immunotherapy for AD based on a mouse model. To further study the underlying mechanisms, we tested our hypothesis that the single chain fragment variable (scFv) without the Fc fragment is capable of regulating either Aß aggregation or disaggregation in vitro. Here, a model of cell-free Aß "on-pathway" aggregation was established and identified using PCR, Western blot, ELISA, transmission electron microscopy (TEM) and thioflavin T (ThT) binding analyses. His-tagged A8 scFvs was cloned and solubly expressed in baculovirus. Our data demonstrated that the Ni-NTA agarose affinity-purified A8 scFv inhibited the forward reaction of "on-pathway" aggregation and Aß fibril maturation. The effect of A8 scFv on Aß fibrillogenesis was markedly more significant when administered at the start of the Aß folding reaction. Furthermore, the results also showed that pre-formed Aß fibrils could be disaggregated via incubation with purified A8 scFv, which suggested that A8 scFv is involved in the reverse reaction of Aß aggregation. Therefore, A8 scFv was capable of both inhibiting fibrillogenesis and disaggregating matured fibrils. Our present study provides valuable insight into the regulators of ultrastructural dynamics of cell-free "on-pathway" Aß aggregation and will assist in the development of therapeutic strategies for AD.

Sublingual administration of a helper-dependent adenoviral vector expressing the codon-optimized soluble fusion glycoprotein of human respiratory syncytial virus elicits protective immunity in mice.

Sublingual (s.l.) immunization has been described as a convenient and safe way to induce mucosal immune responses in the respiratory and genital tracts. We constructed a helper-dependent adenoviral (HDAd) vector expressing a condon-optimized soluble fusion glycoprotein (sFsyn) of respiratory syncytial virus (HDAd-sFsyn) and explored the potential of s.l. immunization with HDAd-sFsyn to stimulate immune responses in the respiratory mucosa. The RSV specific systemic and mucosal immune responses were generated in BALB/c mice, and the serum IgG with neutralizing activity was significantly elevated after homologous boost with s.l. application of HDAd-sFsyn. Humoral immune responses could be measured even 14weeks after a single immunization. Upon challenge, s.l. immunization with HDAd-sFsyn displayed an effective protection against RSV infection. These findings suggest that s.l. administration of HDAd-sFsyn acts as an effective and safe mucosal vaccine against RSV infection, and may be a useful tool in the prevention of RSV infection.

Novel APP K724M mutation causes Chinese early-onset familial Alzheimer's disease and increases amyloid-ß42 to amyloid-ß40 ratio.

Alzheimer's disease (AD) is the most common neurodegenerative disorder among the elderly individuals. Although there are several million cases of AD estimated in China with the most population in the world, no Chinese early-onset familial AD caused by new APP gene mutation has ever been reported. Here, we first described a Chinese family with early-onset AD that was inherited in autosomal dominant manner, and the age of onset was 46.6 ± 7.7 years (n = 5; range, 40-58 years). By using genetic analysis of 3 collected patients' DNA samples, we identified a heterozygous APP gene mutation (g.275363A>T, K724M according to APP770). Finally, when APP695 with K724M mutation was ectopically expressed in HEK293 cell, the ratio of amyloid-ß42 to amyloid-ß40 was 2.23-fold higher than that of wild-type control. Together, our data suggest that APP K724M gene mutation may contribute to the cause of this Chinese early-onset familial AD.