ppmFixer: a mass error adjustment for pGlyco3.0 to correct near-isobaric mismatches.

Modern glycoproteomics experiments require the use of search engines due to the generation of countless spectra. While these tools are valuable, manual validation of search engine results is often required for detailed analysis of glycopeptides as false-discovery rates are often not reliable for glycopeptide data. Near-isobaric mismatches are a common source of misidentifications for the popular glycopeptide-focused search engine pGlyco3.0, and in this technical note we share a strategy and script that improves the accuracy of the search utilizing two manually validated datasets of the glycoproteins CD16a and HIV-1 Env as proof-of-principle.

Prognostic Value of an Immune Long Non-Coding RNA Signature in Liver Hepatocellular Carcinoma.

In recent years, there has been a growing recognition of the important role that long non-coding RNAs (lncRNAs) play in the immunological process of hepatocellular carcinoma (LIHC). An increasing number of studies have shown that certain lncRNAs hold great potential as viable options for diagnosis and treatment in clinical practice. The primary objective of our investigation was to devise an immune lncRNA profile to explore the significance of immune-associated lncRNAs in the accurate diagnosis and prognosis of LIHC. Gene expression profiles of LIHC samples obtained from TCGA database were screened for immune-related genes. The optimal immune-related lncRNA signature was built via correlational analysis, univariate and multivariate Cox analysis. Then, the Kaplan-Meier plot, ROC curve, clinical analysis, gene set enrichment analysis, and principal component analysis were performed to evaluate the capability of the immune lncRNA signature as a prognostic indicator. Six long non-coding RNAs were identified via correlation analysis and Cox regression analysis considering their interactions with immune genes. Subsequently, tumor samples were categorized into two distinct risk groups based on different clinical outcomes. Stratification analysis indicated that the prognostic ability of this signature acted as an independent factor. The Kaplan-Meier method was employed to conduct survival analysis, results showed a significant difference between the two risk groups. The predictive performance of this signature was validated by principal component analysis (PCA). Additionally, data obtained from gene set enrichment analysis (GSEA) revealed several potential biological processes in which these biomarkers may be involved. To summarize, this study demonstrated that this six-lncRNA signature could be identified as a potential factor that can independently predict the prognosis of LIHC patients.

Role of Nrf2/HO-1 pathway on inhibiting activation of ChTLR15/ChNLRP3 inflammatory pathway stimulated by E. tenella sporozoites.

The aim of this study is to explore whether Nrf2 antioxidant pathway negatively regulates the ChTLR15/NLRP3 inflammatory pathway stimulated by Eimeria tenella infection. Firstly, levels of molecules in the Nrf2/HO-1 pathway in DF-1 cells pre-treated with an optimized dose of Corilagine or probiotics Levilactobacillus brevis 23017 were quantified using real-time PCR (qRT-PCR) and Western blot. Then, DF-1 cells pre-treated with Corilagine or L. brevis 23017 were stimulated with E. tenella sporozoites, and mRNA levels of molecules in Nrf2/HO-1 and ChTLR15/NLRP3 pathways, protein levels of p-Nrf2, Nrf2, HO-1, ChTLR15 and ChNLRP3, levels of malondialdehyde (MDA) and reactive oxygen species (ROS) were quantified. Further, expression level of Nrf2 and ChTLR15 in DF-1 cells was knocked down by RNA interfering (RNAi) method, and target cells were pre-treated with Corilagine or L. brevis 23017, followed by stimulation with E. tenella sporozoites, and the expression levels of key molecules in Nrf2/HO-1 and ChTLR15/NLRP3 pathways were quantified. The results showed that mRNA and protein levels of key molecules in the Nrf2/HO-1 pathway in DF-1 cells was significantly upregulated after pretreating with 15 µM Corilagine and supernatant of L. brevis 23017. After stimulating with E. tenella sporozoites, levels of molecules in the ChTLR15/NLRP3 pathway, levels of MDA and ROS in DF-1 cells pre-treated with 15 µM Corilagine or bacterial supernatant were all significantly down-regulated. The results from the knock-down experiment also displayed that Corrigine and L. brevis 23017 inhibited the activation of the ChTLR15/ChNLRP3 inflammatory pathway stimulated by E. tenella sporozoites through activating Nrf2/HO-1 antioxidant pathway. This study provides new ideas for the development of novel anticoccidial products.

The transcriptional changes of LrgA discriminates the responsiveness of Staphylococcus aureus towards blue light from that of photodynamic inactivation.

Antimicrobial blue light (aBL) is utilized as a new approach to inhibit the growth of Staphylococcus aureus (S. aureus). Mediated by the endogenous chromophore, aBL possesses the similar photokilling property with aPDI (antimicrobial photodynamic inactivation), however, their mechanistic discrepancies in triggering the death of staphylococcal cells are not yet understood. Here, we describe the use of a 460-nm-LED to curb the viability of S. aureus. According to the results, the bacterial survival was sharply decreased when blue light was applied, reaching a maximum of 4.11 ± 0.04 log10 units. Moreover, the membrane integrity was damaged by aBL, causing the leakage of intracellular DNA. Transcriptomic analysis indicates the divergent gene expression upon either aBL or aPDI, with pathways such as transport, DNA repair, expression regulation and porphyrin massively affected by aBL. Among the commonly regulated genes, LrgA was underpinned on account of its involvement with biofilm formation and protein transport. By comparing the wildtype with the LrgA-overexpressing (LrgA+) strain, the survival rate, membrane penetration, surface structure and biofilm formation were, to a varying degree, improved for LrgA+, which may suggest that LrgA plays essential roles in modulating the responsiveness of S. aureus. Besides, LrgA may function through regulating the expression of autolysis-related systems. Finally, LrgA overexpression did not attenuate but aggravate the impairment induced by aPDI, showcasing a distinct responsive strategy from aBL. Taken together, this study unveils a unique molecular alteration for the aBL-mediated inactivation, providing the basis of utilizing blue light to reduce the harm brought by S. aureus.

Sanguinarine Induces Necroptosis of HCC by Targeting PKM2 Mediated Energy Metabolism.

BACKGROUNDS: Abnormal metabolism is the hallmark of hepatocellular carcinoma. Targeting energy metabolism has become the major focus of cancer therapy. The natural product, sanguinarine, displays remarkable anti-tumor properties by disturbing energy homeostasis; however, the underlying mechanism has not yet been elucidated. METHODS: The anticancer activity of sanguinarine was determined using CCK-8 and colony formation assay. Morphological changes of induced cell death were observed under electron microscopy. Necroptosis and apoptosis related markers were detected using western blotting. PKM2 was identified as the target by transcriptome sequencing. Molecular docking assay was used to evaluate the binding affinity of sanguinarine to the PKM2 molecule. Furthermore, Alb-CreERT2; PKM2loxp/loxp; Rosa26RFP mice was used to construct the model of HCC-through the intervention of sanguinarine in vitro and in vivo-to accurately explore the regulation effect of sanguinarine on cancer energy metabolism. RESULTS: Sanguinarine inhibited tumor proliferation, metastasis and induced two modes of cell death. Molecular docking of sanguinarine with PKM2 showed appreciable binding affinity. PKM2 kinase activity and aerobic glycolysis rate declined, and mitochondrial oxidative phosphorylation was inhibited by sanguinarine application; these changes result in energy deficits and lead to necroptosis. Additionally, sanguinarine treatment prevents the translocation of PKM2 into the nucleus and suppresses the interaction of PKM2 with ß-catenin; the transcriptional activity of PKM2/ß-catenin signaling and its downstream genes were decreased. CONCLUSIONS: Sanguinarine showed remarkable anti-HCC activity via regulating energy metabolism by PKM2/ß-catenin signaling. On the basis of these investigations, we propose that sanguinarine might be considered as a promising compound for discovery of anti-HCC drugs.

Frequency-potency analysis of IgG+ memory B cells delineates neutralizing antibody responses at single-cell resolution.

Identifying individual functional B cell receptors (BCRs) is common, but two-dimensional analysis of B cell frequency versus BCR potency would delineate both quantity and quality of antigen-specific memory B cells. We efficiently determine quantitative BCR neutralizing activities using a single-cell-derived antibody supernatant analysis (SCAN) workflow and develop a frequency-potency algorithm to estimate B cell frequencies at various neutralizing activity or binding affinity cutoffs. In an HIV-1 fusion peptide (FP) immunization study, frequency-potency curves elucidate the quantity and quality of FP-specific immunoglobulin G (IgG)+ memory B cells for different animals, time points, and antibody lineages at single-cell resolution. The BCR neutralizing activities are mainly determined by their affinities to soluble envelope trimer. Frequency analysis definitively demonstrates dominant neutralizing antibody lineages. These findings establish SCAN and frequency-potency analyses as promising approaches for general B cell analysis and monoclonal antibody (mAb) discovery. They also provide specific rationales for HIV-1 FP-directed vaccine optimization.

Antibodies targeting the fusion peptide on the HIV envelope provide protection to rhesus macaques against mucosal SHIV challenge.

The fusion peptide (FP) on the HIV-1 envelope (Env) trimer can be targeted by broadly neutralizing antibodies (bNAbs). Here, we evaluated the ability of a human FP-directed bNAb, VRC34.01, along with two vaccine-elicited anti-FP rhesus macaque mAbs, DFPH-a.15 and DF1W-a.01, to protect against simian-HIV (SHIV)BG505 challenge. VRC34.01 neutralized SHIVBG505 with a 50% inhibitory concentration (IC50) of 0.58 µg/ml, whereas DF1W-a.01 and DFPH-a.15 were 4- or 30-fold less potent, respectively. VRC34.01 was infused into four rhesus macaques at a dose of 10 mg/kg and four rhesus macaques at a dose of 2.5 mg/kg. The animals were intrarectally challenged 5 days later with SHIVBG505. In comparison with all 12 control animals that became infected, all four animals infused with VRC34.01 (10 mg/kg) and three out of four animals infused with VRC34.01 (2.5 mg/kg) remained uninfected. Because of the lower potency of DF1W-a.01 and DFPH-a.15 against SHIVBG505, we infused both Abs at a higher dose of 100 mg/kg into four rhesus macaques each, followed by SHIVBG505 challenge 5 days later. Three of four animals that received DF1W-a.01 were protected against infection, whereas all animals that received DFPH-a.15 were protected. Overall, the protective serum neutralization titers observed in these animals were similar to what has been observed for other bNAbs in similar SHIV infection models and in human clinical trials. In conclusion, FP-directed mAbs can thus provide dose-dependent in vivo protection against mucosal SHIV challenges, supporting the development of prophylactic vaccines targeting the HIV-1 Env FP.

Design, Preparation, and Mechanical Property Study of Polylactic Acid/Ca2BO3Cl: Eu2+, Dy3+ Composite Material for 3D Printing.

To break through the simplification of three-dimensional printing (3DP) materials and realize the application of long-persistent phosphors in more fields, polylactic acid doped with Ca2BO3Cl: Eu2+, Dy3+ was prepared in this study. The structure of the mixtures was analyzed and determined by infrared spectroscopy. The luminescence properties of phosphors and the composites were studied by fluorescence spectra and afterglow decay curve measurements. The yellow light of the mixtures could be attributed to the 5d-4f energy level transition of Eu2+. After the excitation of 254 nm ultraviolet lamp, the luminance and duration of the composites could be clearly observed. The mechanical properties of the composite filaments were tested, including maximal force and elasticity modulus. In particular, the influence of humidity on mechanical properties was analyzed in detail. The prepared composite filaments were printed into hollow dodecahedrons and presented in this study. Therefore, the composites had great properties and might be expected to put into practical applications of 3DP technology.