Single-Cell Sequencing of Peripheral Mononuclear Cells Reveals Distinct Immune Response Landscapes of COVID-19 and Influenza Patients.

The coronavirus disease 2019 (COVID-19) pandemic poses a current world-wide public health threat. However, little is known about its hallmarks compared to other infectious diseases. Here, we report the single-cell transcriptional landscape of longitudinally collected peripheral blood mononuclear cells (PBMCs) in both COVID-19- and influenza A virus (IAV)-infected patients. We observed increase of plasma cells in both COVID-19 and IAV patients and XIAP associated factor 1 (XAF1)-, tumor necrosis factor (TNF)-, and FAS-induced T cell apoptosis in COVID-19 patients. Further analyses revealed distinct signaling pathways activated in COVID-19 (STAT1 and IRF3) versus IAV (STAT3 and NFκB) patients and substantial differences in the expression of key factors. These factors include relatively increase of interleukin (IL)6R and IL6ST expression in COVID-19 patients but similarly increased IL-6 concentrations compared to IAV patients, supporting the clinical observations of increased proinflammatory cytokines in COVID-19 patients. Thus, we provide the landscape of PBMCs and unveil distinct immune response pathways in COVID-19 and IAV patients.

High throughput screening of pure silica zeolites for CF4 capture from electronics industry gas.

The capture and separation of CF4 from CF4/N2 mixture gas is a crucial issue in the electronics industry, as CF4 is a commonly used etching gas and the ratio of CF4 to N2 directly affects process efficiency. Utilizing high-throughput computational screening techniques and grand canonical Monte Carlo (GCMC) simulations, we comprehensively screened and assessed 247 types of pure silicon zeolite materials to determine their adsorption and separation performance for CF4/N2 mixtures. Based on screening, the relationships between the structural parameters and adsorption and separation properties were meticulously investigated. Four indicators including adsorption selectivity, working capacity, adsorbent performance score (APS), and regenerability (R%) were used to evaluate the performance of adsorbents. Based on the evaluation, we selected the top three best-performing zeolite structures for vacuum swing adsorption (LEV, AWW and ESV) and pressure swing adsorption (AVL, ZON, and ERI) processes respectively. Also, we studied the preferable adsorption sites of CF4 and N2 in the selected zeolite structures through centroid density distributions at the molecule level. We expect the study may provide some valuable guidance for subsequent experimental investigations on adsorption and separation of CF4/N2.

Comprehensive Analysis of Fibroblast Activation Protein Expression in Interstitial Lung Diseases.

Rationale: Sustained activation of lung fibroblasts and the resulting oversynthesis of the extracellular matrix are detrimental events for patients with interstitial lung diseases (ILDs). Lung biopsy is a primary evaluation technique for the fibrotic status of ILDs, and is also a major risk factor for triggering acute deterioration. Fibroblast activation protein (FAP) is a long-known surface biomarker of activated fibroblasts, but its expression pattern and diagnostic implications in ILDs are poorly defined. Objectives: The present study aims to comprehensively investigate whether the expression intensity of FAP could be used as a potential readout to estimate or measure the amounts of activated fibroblasts in ILD lungs quantitatively. Methods: FAP expression in human primary lung fibroblasts as well as in clinical lung specimens was first tested using multiple experimental methods, including real-time quantitative PCR (qPCR), Western blot, immunofluorescence staining, deep learning measurement of whole slide immunohistochemistry, as well as single-cell sequencing. In addition, FAP-targeted positron emission tomography/computed tomography imaging PET/CT was applied to various types of patients with ILD, and the correlation between the uptake of FAP tracer and pulmonary function parameters was analyzed. Measurements and Main Results: Here, it was revealed, for the first time, FAP expression was upregulated significantly in the early phase of lung fibroblast activation event in response to a low dose of profibrotic cytokine. Single-cell sequencing data further indicate that nearly all FAP-positive cells in ILD lungs were collagen-producing fibroblasts. Immunohistochemical analysis validated that FAP expression level was closely correlated with the abundance of fibroblastic foci on human lung biopsy sections from patients with ILDs. We found that the total standard uptake value (SUV) of FAP inhibitor (FAPI) PET (SUVtotal) was significantly related to lung function decline in patients with ILD. Conclusions: Our results strongly support that in vitro and in vivo detection of FAP can assess the profibrotic activity of ILDs, which may aid in early diagnosis and the selection of an appropriate therapeutic window.

Single-cell RNA sequencing reveals enhanced antitumor immunity after combined application of PD-1 inhibitor and Shenmai injection in non-small cell lung cancer.

BACKGROUND: Immune checkpoint inhibitors (ICIs) have altered the clinical management of non-small cell lung cancer (NSCLC). However, the low response rate, severe immune-related adverse events (irAEs), and hyperprogressive disease following ICIs monotherapy require attention. Combination therapy may overcome these limitations and traditional Chinese medicine with immunomodulatory effects provides a promising approach. Shenmai injection (SMI) is a clinically effective adjuvant treatment for cancer with chemotherapy and radiotherapy. Therefore, the combined effects and mechanisms of SMI and programmed death-1 (PD-1) inhibitor against NSCLC was focused on this study. METHODS: A Lewis lung carcinoma mouse model and a lung squamous cell carcinoma humanized mouse model were used to investigate the combined efficacy and safety of SMI and PD-1 inhibitor. The synergistic mechanisms of the combination therapy against NSCLC were explored using single-cell RNA sequencing. Validation experiments were performed using immunofluorescence analysis, in vitro experiment, and bulk transcriptomic datasets. RESULTS: In both models, combination therapy alleviated tumor growth and prolonged survival without increasing irAEs. The GZMAhigh and XCL1high natural killer (NK) cell subclusters with cytotoxic and chemokine signatures increased in the combination therapy, while malignant cells from combination therapy were mainly in the apoptotic state, suggesting that mediating tumor cell apoptosis through NK cells is the main synergistic mechanisms of combination therapy. In vitro experiment confirmed that combination therapy increased secretion of Granzyme A by NK cells. Moreover, we discovered that PD-1 inhibitor and SMI combination blocked inhibitory receptors on NK and T cells and restores their antitumoral activity in NSCLC better than PD-1 inhibitor monotherapy, and immune and stromal cells exhibited a decrease of angiogenic features and attenuated cancer metabolism reprogramming in microenvironment of combination therapy. CONCLUSIONS: This study demonstrated that SMI reprograms tumor immune microenvironment mainly by inducing NK cells infiltration and synergizes with PD-1 inhibitor against NSCLC, suggested that targeting NK cells may be an important strategy for combining with ICIs. Video Abstract.

Aortic Regurgitation Requiring Unplanned Surgery following Transcatheter Closure of Ventricular Septal Defect in Children: Incidence and Risk Factors.

INTRODUCTION: Our aim was to investigate the incidence and risk factors for aortic regurgitation (AR) requiring unplanned surgery after transcatheter closure of ventricular septal defect (VSD) in children. METHODS: Medical records of 876 children with VSD who underwent transcatheter closure from July 2009 to September 2018 in our hospital were retrospectively reviewed. Groups with and without new-onset or increasing AR requiring unplanned surgery were compared. Univariate and multivariate analyses were used to identify the possible risk factors. Smoothing plot and threshold effect analysis were carried out to find the relationship between possible factors and risk of new-onset or increasing AR. RESULTS: A total of 29 children (3.3%) underwent unplanned surgery after transcatheter closure owing to new-onset or increasing AR, including 6 children with new-onset AR and 23 children with increasing AR. Multivariate regression analysis revealed that preoperative mild AR (OR: 60.39, 95% CI: 11.53-316.30, p < 0.001), larger ratio between diameter to body surface area (OR: 1.25, 95% CI: 1.01-1.55, p = 0.039), intracristal VSD (OR: 34.09, 95% CI: 4.07-285.65, p < 0.001), and shorter distance from the upper edge of defect to the aortic valve (or the sub-aortic rim) (OR: 0.12, 95% CI: 0.05-0.27, p < 0.001) were risk factors for new-onset or increasing AR requiring unplanned surgery. And, low risk of AR after muscular VSD transcatheter closure was found. An L-shaped nonlinear relationship between the sub-aortic rim and the risk of new-onset or increasing AR was observed, and the risk of new-onset or increasing AR with the sub-aortic rim up to the turning point (2 mm) (adjusted OR: 0.00, 95% CI: 0.00-0.08; p =0.001). With a median time of 7.3 years' follow-up, no new-onset or increasing AR has been found for children who initially did not have unplanned surgery. CONCLUSION: Preoperative mild AR, larger ratio between diameter to body surface area, intracristal VSD, and shorter distance of the sub-aortic rim (especially <2 mm) could increase the risk of new-onset or increasing AR requiring unplanned surgery after transcatheter closure of VSD.

scDeepSort: a pre-trained cell-type annotation method for single-cell transcriptomics using deep learning with a weighted graph neural network.

Advances in single-cell RNA sequencing (scRNA-seq) have furthered the simultaneous classification of thousands of cells in a single assay based on transcriptome profiling. In most analysis protocols, single-cell type annotation relies on marker genes or RNA-seq profiles, resulting in poor extrapolation. Still, the accurate cell-type annotation for single-cell transcriptomic data remains a great challenge. Here, we introduce scDeepSort (https://github.com/ZJUFanLab/scDeepSort), a pre-trained cell-type annotation tool for single-cell transcriptomics that uses a deep learning model with a weighted graph neural network (GNN). Using human and mouse scRNA-seq data resources, we demonstrate the high performance and robustness of scDeepSort in labeling 764 741 cells involving 56 human and 32 mouse tissues. Significantly, scDeepSort outperformed other known methods in annotating 76 external test datasets, reaching an 83.79% accuracy across 265 489 cells in humans and mice. Moreover, we demonstrate the universality of scDeepSort using more challenging datasets and using references from different scRNA-seq technology. Above all, scDeepSort is the first attempt to annotate cell types of scRNA-seq data with a pre-trained GNN model, which can realize the accurate cell-type annotation without additional references, i.e. markers or RNA-seq profiles.

CircRNAs: A Promising Star for Treatment and Prognosis in Oral Squamous Cell Carcinoma.

CircRNAs are a class of endogenous long non-coding RNAs with a single-stranded circular structure. Most circRNAs are relatively stable, highly conserved, and specifically expressed in tissue during the cell and developmental stages. Many circRNAs have been discovered in OSCC. OSCC is one of the most severe and frequent forms of head and neck cancer today, with a poor prognosis and low overall survival rate. Due to its prevalence, OSCC is a global health concern, characterized by genetic and epigenomic changes. However, the mechanism remains vague. With the advancement of biotechnology, a large number of circRNAs have been discovered in mammalian cells. CircRNAs are dysregulated in OSCC tissues and thus associated with the clinicopathological characteristics and prognosis of OSCC patients. Research studies have demonstrated that circRNAs can serve as biomarkers for OSCC diagnosis and treatment. Here, we summarized the properties, functions, and biogenesis of circRNAs, focusing on the progress of current research on circRNAs in OSCC.

Managing Secondary Phase Lead Iodide in Hybrid Perovskites via Surface Reconstruction for High-Performance Perovskite Solar Cells with Robust Environmental Stability.

Rationally managing the secondary-phase excess lead iodide (PbI2 ) in hybrid perovskite is of significance for pursuing high performance perovskite solar cells (PSCs), while the challenge remains on its conversion to a homogeneous layer that is robust stable against environmental stimuli. We herein demonstrate an effective strategy of surface reconstruction that converts the excess PbI2 into a gradient lead sulfate-silica bi-layer, which substantially stabilizes the perovskite film and reduces interfacial charge transfer barrier in the PSCs device. The perovskite films with such bi-layer could bear harsh conditions such as soaking in water, light illumination at 70 % relative humidity, and the damp-thermal (85 °C and 30 % humidity) environment. The resulted PSCs deliver a champion efficiency up to 24.09 %, as well as remarkable environmental stability, e.g., retaining 78 % of their initial efficiency after 5500 h of shelf storage, and 82 % after 1000 h of operational stability testing.

Intracellular hydroxyproline imprinting following resolution of bleomycin-induced pulmonary fibrosis.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a fatal lung disease with few treatment options. The poor success in developing anti-IPF strategies has impelled researchers to reconsider the importance of the choice of animal model and assessment methodologies. Currently, it is still not settled whether the bleomycin-induced lung fibrosis mouse model finally returns to resolution. METHODS: This study aimed to follow the dynamic fibrotic features of bleomycin-treated mouse lungs over extended durations through a combination of the latest technologies (micro-computed tomography imaging and histological detection of degraded collagens) and traditional methods. In addition, we also applied immunohistochemistry to explore the distribution of all hydroxyproline-containing molecules. RESULTS: As determined by classical biochemical methods, total lung hydroxyproline contents reached a peak at 4 weeks after bleomycin injury and maintained a steady high level thereafter until the end of the experiments (16 weeks). This result seemed to partially contradict with the changes of other fibrosis evaluation parameters, which indicated a gradual degradation of collagens and a recovery of lung aeration after the fibrosis peak. This inconsistency was well reconciled by our data from immunostaining against hydroxyproline and fluorescent peptide staining against degraded collagen, together showing large amounts of hydroxyproline-rich degraded collagen fragments detained and enriched within the intracellular regions at 10 or 16 weeks rather than at 4 weeks after bleomycin treatment. CONCLUSIONS: Our present data not only offer respiratory researchers a new perspective towards the resolution nature of mouse lung fibrosis, but also remind them to be cautious when using the hydroxyproline content assay to evaluate the severity of fibrosis.

[Modern research on Chinese medicine based on single-cell omics: technologies and strategies].

As one of the most advanced technologies, single-cell omics technology develops rapidly in recent years. Based on different technical strategies, it enables unbiased and high-throughput access to multiple omics information at single-cell resolution. So far, single-cell omics technology, by virtue of its great powder in resolving tissue heterogeneity, has become a revolutionary tool to deeply understand the functional structure of tissues, reveal complex disease processes, and elucidate drug mechanisms of action. In view of the technical challenges in deconstructing the complexity of Chinese medicine and clarifying the modern scientific connotation of traditional Chinese medicine(TCM) theory, single-cell omics technology has huge application potential in the discovery of pharmacodynamic substances, construction of action networks, and elucidation of integrated regulatory mechanisms, which brings new opportunities for modern research in TCM. The present study briefly introduced three representative single-cell omics technologies, i.e., single-cell transcriptome sequencing, spatial transcriptomics, and single-cell multimodal omics, and their main application patterns. On this basis, an outlook was proposed on the strategies and applications for modern research in TCM using single-cell omics technology.

The breadth of concomitant virological features in a family cluster outbreak of COVID-19 pneumonia.

An outbreak of a novel coronavirus disease (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) had emerged in 2019 and rapidly posed a global epidemic. Here, we report the breadth of concomitant virological features of a family cluster with COVID-19. The period of virus shedding is significantly different between upper respiratory and feces samples. Even the SARS-CoV-2 virus titers were undetectable in feces, it could be positive again soon and likely related to fluctuated inflammation levels (interleukin-6, etc.) and lowered immune responses (CD4 + T lymphocyte, etc.). Our findings expand the novel understanding of the breadth of concomitant virological features during a non-severe family cluster of COVID-19.

HBx Protein Contributes to Liver Carcinogenesis by H3K4me3 Modification Through Stabilizing WD Repeat Domain 5 Protein.

BACKGROUND AND AIMS: Cancer is typically considered as a genetic and epigenetic disease. Although numerous studies have indicated that an aberrant structure, function, or expression level of epigenetic enzymes contribute to many tumor types, precisely how the epigenetic mechanisms are involved in the hepatitis B virus (HBV)-induced hepatocellular carcinoma (HCC) remains unknown. APPROACH AND RESULTS: In this study, we found that the WD repeat domain 5 protein (WDR5)-a core subunit of histone H3 lysine 4 methyltransferase complexes, which catalyze the generation of histone H3 lysine 4 trimethylation (H3K4me3) modification-is highly expressed in HBV-related HCC and promotes HCC development. WDR5 plays a critical role in HBV-driven cell proliferation and tumor growth in mice, and the WDR5-0103 small-molecule inhibitor of WDR5 activity compromises HBV- and hepatitis B x protein (HBx)-driven tumor proliferation. The aberrantly high WDR5 protein level was found to involve HBx through its stabilization of the WDR5 protein by inhibiting the interaction between the damage-specific DNA-binding protein 1/cullin-4 and WDR5, causing decreased ubiquitination of the WDR5 protein. HBx was found to colocalize with WDR5 on chromatin genome wide and promotes genome-wide H3K4me3 modification by means of WDR5. Furthermore, the recruitment of HBx to promoters of target genes relied on its interaction with WDR5 through its α-helix domain. WDR5 was also found to promote HBV transcription through H3K4 modification of covalently closed circular DNA minichromosome, and WDR5-0103 was able to inhibit HBV transcription. Finally, the in vitro and in vivo data further proved that HBx exerted its tumor-promoting function in a WDR5-dependent manner. CONCLUSIONS: Our data reveals that WDR5 is a key epigenetic determinant of HBV-induced tumorigenesis and that the HBx-WDR5-H3K4me3 axis may be a potential therapeutic target in HBV-induced liver pathogenesis.

A recombinant influenza virus with a CTLA4-specific scFv inhibits tumor growth in a mouse model.

Oncolytic viruses (OV) have shown excellent safety and efficacy in preclinical and clinical studies. Influenza A virus (IAV) is considered a promising oncolytic virus. In this report, we generated a recombinant influenza virus expressing an immune checkpoint blockade agent targeting CTLA4. Using reverse genetics, a recombinant influenza virus, termed rFlu-CTLA4, encoding the heavy chain of a CTLA4 antibody on the PB1 segment and the light chain of the CTLA4 antibody on the PA segment was produced. RFlu-CTLA4 could replicate to high titers, and antibodies were produced in the allantoic fluid of infected eggs. Furthermore, the selective cytotoxicity of the virus was higher in various hepatocellular carcinoma cancer cell lines than in the normal cell line L02 in vitro, as indicated by MTS assays. More importantly, in a subcutaneous H22 mouse hepatocarcinoma model, intratumoral injections of rFlu-CTLA4 inhibited the growth of treated tumors and increased the overall survival of mice compared with injections of the PR8 virus. Taken together, these results warrant further exploration of this novel recombinant influenza virus for its potential use as a single or combination agent for cancer immunotherapy.

Long noncoding RNA FAM66C promotes tumor progression and glycolysis in intrahepatic cholangiocarcinoma by regulating hsa-miR-23b-3p/KCND2 axis.

Long noncoding RNAs (lncRNAs) are known to be the important regulators in cancer progression. However, the role of lncRNA FAM66C (FAM66C) is yet to be investigated in intrahepatic cholangiocarcinoma (ICC). This study aimed to investigate the effects and related mechanisms of FAM66C in ICC. Human ICC tissues and cell lines were collected. The expression levels of FAM66C, hsa-miR-23b-3p (miR-23b-3p), and KCND2 were detected by qRT-RCR. The transfection experiments were employed to measure the effect of FAM66C on cell viabilities, migration, and invasion in ICC cells by CCK-8, transwell assays. Glycolysis was investigated by glucose consumption, lactate production and ATP levels. The dual-luciferase reporter and RNA pull down assays were conducted as a means of confirming the interactions between FAM66C, miR-23b-3p, and KCND2. Furthermore, the levels of the EMT-associated proteins (KCND2, GLUT1, PKM2, and LDHA) in ICC cells were detected by western blot. FAM66C was increased in ICC tissues and cells, increased cell viability, glycolysis, migration and invasion, and decreased apoptosis were shown in FAM66C overexpressing cells. Mechanistic analyses revealed that FAM66C regulated the downstream target gene KCND2 by sponging miR-23b-3p. FAM66C effect on ICC was further validated in murine xenograft assays. FAM66C knockdown cells gave rise to tumors that were smaller in size, consistent with the role of FAM66C as a promoter of in vivo tumor growth. These data revealed that FAM66C was able to drive ICC tumor progression and glycolytic activity via the miR-23b-3p/KCND2 axis, indicating FAM66C may be a viable target for treating ICC.

SET domain containing 1B gene is mutated in primary hepatic neuroendocrine tumors.

Primary hepatic neuroendocrine tumors (PHNETs) are extremely rare NETs originating from the liver. These tumors are associated with heterogeneous prognosis, and few treatment targets for PHNETs have been identified. Because the major genetic alterations in PHNET are still largely unknown, we performed whole-exome sequencing of 22 paired tissues from PHNET patients and identified 22 recurring mutations of somatic genes involved in the following activities: epigenetic modification (BPTF, MECP2 and WDR5), cell cycle (TP53, ATM, MED12, DIDO1 and ATAD5) and neural development (UBR4, MEN1, GLUL and GIGYF2). Here, we show that TP53 and the SET domain containing the 1B gene (SETD1B) are the most frequently mutated genes in this set of samples (3/22 subjects, 13.6%). A biological analysis suggests that one of the three SETD1B mutants, A1054del, promotes cell proliferation, migration and invasion compared to wild-type SETD1B. Our work unveils that SETD1B A1054del mutant is functional in PHNET and implicates genes including TP53 in the disease. Our findings thus characterize the mutational landscapes of PHNET and implicate novel gene mutations linked to PHNET pathogenesis and potential therapeutic targets.

Virus-like particle vaccine by intranasal vaccination elicits protective immunity against respiratory syncytial viral infection in mice.

Respiratory syncytial virus (RSV) is a leading cause of lower respiratory infection in infants and children, but there is still no licensed vaccine available. In this report, we developed virus-like particle (VLP) vaccines based on the Bac-to-Bac baculovirus expression system, consisting of an influenza virus matrix (M1) protein and the RSV fusion protein (F) or glycoprotein (G). These RSV VLPs were identified by western blot analysis and electron microscopy. Female BALB/c mice immunized intranasally (i.n.) with RSV-F VLPs, RSV-G VLPs, or both showed viral-specific antibody responses against RSV. Total IgG, IgG1, IgG2a, and mucosal IgA were detected in mice with RSV-F plus RSV-G VLPs, revealing potent cellular and mucosal immune responses. Moreover, we found that these mixed RSV VLPs conferred enhanced protection against live RSV challenges, showing significant decreases in lung viral replication and obvious attenuation of histopathological changes associated with viral infections. These results demonstrate that RSV-F plus RSV-G VLPs by intranasal vaccination is a promising vaccine candidate that warrants further evaluation using cotton rat and primate models.

Human Basic Fibroblast Growth Factor Inhibits Tau Phosphorylation via the PI3K/Akt-GSK3ß Signaling Pathway in a 6-Hydroxydopamine-Induced Model of Parkinson's Disease.

BACKGROUND: Basic fibroblast growth factor (bFGF) has been increasingly investigated due to its neuroprotection in neurodegenerative disorders. Because there are still no cures for any of these disorders, it is crucial to identify new therapeutic targets and screen potential drugs. The increased phosphorylation of tau at Ser396 leads to intracellular tau accumulation, which forms neurofibrillary tangles in Parkinson's disease (PD). In this study, neuroprotection by bFGF was observed, and the mechanisms related to its regulation of phosphorylated tau were investigated. METHODS: bFGF-loaded liposome carriers were intranasally administered to rats. The neuroprotective effects of bFGF were assessed in a PD model induced by 6-hydroxydopamine (6-OHDA) in vivo and in vitro. The phosphorylation of tau was measured, and the PI3K/Akt-GSK3ß signaling pathway was investigated. RESULTS: Our study demonstrated that liposomes markedly assisted in the delivery of bFGF to the striatum and substantia nigra of rats and enhanced the neuroprotective effects of bFGF on dopaminergic neurons. bFGF treatment significantly ameliorated the behavioral deficits induced by 6-OHDA, rescued the loss of tyrosine hydroxylase-positive neurons and increased the number of Nissl bodies. bFGF reduced the phosphorylation of tau and GSK3ß and increased the phosphorylation of PI3K/Akt. CONCLUSION: Liposomes markedly assisted in the delivery of bFGF to the brain and enhanced the neuroprotective effects of bFGF by inhibiting the phosphorylation of tau. bFGF down-regulated the phosphorylation of tau by increasing the phosphorylation of GSK3ß via the PI3K/Akt signaling pathway. These findings provide a new vision of bFGF as a potential therapy for PD.

Characterization of recombinant influenza A virus as a vector expressing respiratory syncytial virus fusion protein epitopes.

Respiratory syncytial virus (RSV) is the most common cause of respiratory infection in infants and the elderly, and no vaccine against this virus has yet been licensed. Here, we report a recombinant PR8 influenza virus with the RSV fusion (F) protein epitopes of the subgroup A gene inserted into the influenza virus non-structural (NS) gene (rFlu/RSV/F) that was generated as an RSV vaccine candidate. The rescued viruses were assessed by microscopy and Western blotting. The proper expression of NS1, the NS gene product, and the nuclear export protein (NEP) of rFlu/RSV/F was also investigated using an immunofluorescent assay. The rescued virus replicated well in the MDCK kidney cell line, A549 lung adenocarcinoma cell line and CNE-2Z nasopharyngeal carcinoma cell line. BALB/c mice immunized intranasally with rFlu/RSV/F had specific haemagglutination inhibition antibody responses against the PR8 influenza virus and RSV neutralization test proteins. Furthermore, intranasal immunization with rFlu/RSV/F elicited T helper type 1-dominant cytokine profiles against the RSV strain A2 virus. Taken together, our findings suggested that rFlu/RSV/F was immunogenic in vivo and warrants further development as a promising candidate vaccine.

Evaluation of the safety and brain-related tissues distribution characteristics of TAT-HaFGF via intranasal administration.

Disabilities triggered by neurodegeneration mainly result in mortality in the elderly, and patients with neurodegenerative disease also display deficits in olfactory function. Therefore drug distribution to the brain through intranasal administration has become one of the most difficult challenges in the treatment of central nervous system (CNS) diseases. TAT-human acidic fibroblast growth factor (HaFGF) is a new fused protein retaining the neuroprotective activities of HaFGF, and is a promising prospect in the treatment of neurodegenerative diseases. TAT (a cell-penetrating peptide) contains a high relative abundance of positively charged amino acids such as lysine and arginine, which have a powerful attraction to the negatively charge on the nasal epithelial membrane. The present study focused on the evaluation of the safety and absorption characteristics of TAT-HaFGF following intranasal administration. After TAT-HaFGF intranasal administration (100, 300, 600 µg/kg) for 5 weeks, hematoxylin-eosin (HE) staining showed no pathology in any of the investigated tissues and organs. The expression of olfactory marker protein (OMP) was observed with immunohistochemical staining, which showed no altered expression in the sensory neurons of the nasal epithelium. Nasal ciliotoxicity studies carried out using an in situ palate model and optical microscope showed that TAT-HaFGF had no nasal ciliotoxicity. The distribution of the TAT-HaFGF following intranasal administration was assessed using a radioisotopic tracing method. Radioactivity was observed in the brain after 15 min. This became stronger at 30 min and weaker at 1 h. All of the results confirmed the in vivo safety of TAT-HaFGF via intranasal administration.

Development of an immunochromatographic kit for rapid detection of human influenza B virus infection.

BACKGROUND: Type B influenza virus is a major epidemic strain responsible for considerable mortality and morbidity. METHODS: A colloidal gold immunochromatographic strip for the rapid detection of human influenza B virus was developed. This test is based on membrane chromatography and uses colloidal gold conjugated with influenza B virus anti-NP monoclonal antibody as the tracer. The assembled test strip was housed in a plastic case. RESULTS: The colloid gold strip (CGS) specifically detected all influenza B viruses tested and did not react with other respiratory viruses. Compared with SYBR Green real-time PCR, the sensitivity and specificity of the CGS test was 89.76% and 99.56%, respectively, and the consistency ratio between CGS and real-time PCR was 96.06% in detecting influenza B virus in 710 nasopharyngeal swabs from patients with influenza-like illness in the hospital. CONCLUSIONS: The CGS array developed in this study enabled typing of influenza B viruses in human clinical specimens. Thus, together with the advantages of rapid detection and easy operation without requiring specialized personnel and equipment, this technique is a convenient and relatively inexpensive diagnostic tool for large-scale screening of clinical samples.