Analysis of viral proteins of porcine reproductive and respiratory syndrome virus in host protection and early virus-cell interactions

The viral proteins of porcine reproductive and respiratory syndrome virus (PRRSV) were expressed in a baculovirus expression system. They were evaluated for protection of pigs against the respiratory disease and for function in virus-cell interactions. The open reading frames 2 to 6 products were confirmed to be membrane associated proteins as they were detected on the surface of viable insect cells by an immunofluorescence assay. Immunization of pigs with vaccines comprised of a combination of GP2, GP3 and GP4 or a combination of GP5, M and N proteins induced partial protection, but immunization with combination of GP5 and M proteins failed to induce protection. Immunization of pigs with the first two vaccines significantly reduced the severity of clinical respiratory disease (P \u3c 0.001), severity of pneumonia (P \u3c 0.05) and duration of viremia compared with those of control group. These results suggest that in addition to envelope proteins, N protein may also play an important role in inducing host protection. For further characterization of these proteins, monoclonal antibodies (MAbs) were prepared. Most of the MAbs were found to be against conformationally dependent epitopes. Antigenic variation was found in PRRSV field isolates and thirty-three PRRSV isolates were divided into three groups based on their reactivity with the MAbs to GP2, GP3 and M proteins. Early virus-cell interactions in PRRSV infection was evaluated with a virus binding assay on CRL11171 cells. The recombinant ORFs 2 to 6 products failed to block PRRSV binding, implying that there were necessary interactions between viral structural proteins. PRRSV receptor was shown to contain protein moieties as protease treatment of cell monolayers reduced virus binding. The receptor activity was reduced from the cell surface with octylglucoside treatment, but no specific protein band was found by immunoprecipitation of the cell surface extracts, suggesting that the PRRSV receptor is present in low concentration. An 18-kDa viral protein was found to bind to the cells by an adsorption assay. Glycosylation did not affect the binding ability of this protein. The protein was immunoprecipitated with MAb against PRRSV M protein, suggesting its potential involvement in virus binding and/or entry

Inducing Autophagic Cell Death by Nsp5 of Porcine Reproductive and Respiratory Syndrome Virus

Partial funding for Open Access provided by the UMD Libraries' Open Access Publishing Fund.Porcine Reproductive and Respiratory Syndrome (PRRS) leads to severe economic losses to the swine-producing industry. Many unclear questions remain on pathogenesis of PRRS virus (PRRSV), including the mechanism of PRRSV-induced cell death. In this study, we cloned and expressed a PRRSV non-structural protein, nsp5, and discovered that it induced cell death in cultured cells. The nsp5 protein localized in cytoplasm and majority of the protein concentrated in perinuclear region. Along with extension of incubation time, the nsp5 tended to form puncta and polarized besides nucleus. An interesting observation was that the nsp5 expression induced cell death. Cell viability assay showed that the cells with nsp5 expression had over 2-fold more cell death than cells with empty vector. Further study indicated that the nsp5 induced cell death via autophagy. Treatment with 3-MA, an autophagy inhibitor, blocked the nsp5- induced cell death. These results suggest that nsp5 might play an important role in PRRSV-induced cell death. Further examination on the mechanism is warranted

Signal-independent RFF Identification for LTE Mobile Devices via Ensemble Deep Learning

Radio frequency fingerprint (RFF)-based wireless device authentication is an emerging technique to prevent potential spoofing attacks in wireless communications. The random access preamble of the physical random access channel (PRACH) in Long Term Evolution (LTE) systems is the first message sent from a user equipment (UE). However, PRACH preambles change under different evolved Node B (eNB), which will affect the RFF extraction. In this paper, a signal-independent RFF extraction method is first proposed to extract varying LTE PRACH preambles under different LTE eNBs. Residual transient segment (RTS) features from the varying PRACH preambles are extracted for RFF identification. A convolutional neural network (CNN) based ensemble deep learning scheme is proposed to integrate benefits from different RFF features. An experimental system under real operator LTE eNB is designed to capture and identify real UE signals. Experimental results show that the classification accuracy of five UEs can reach more than 95% under the same eNB and 85% under different eNBs. Furthermore, longtime evaluations show that the UE RTS feature is robust over time

Antibacterial performance of a porous Cu-bearing titanium alloy by laser additive manufacturing

Porphyromonas gingivalis (P. gingivalis) is the most common species that causes peri-implantitis. It forms an irreversible dense biofilm and causes inflammation. A novel 3D-printed porous TC4-6Cu alloy was fabricated using selective laser melting (SLM) technology for the dental implant, which is anticipated to inhibit biofilm formation. We attempted to investigate the antibacterial ability and antibacterial mechanism of the 3D-printed porous TC4-6Cu alloy against P. gingivalis. This work used scanning electron microscopy (SEM) and laser confocal microscopy (CLSM) to detect the antimicrobial ability of the alloy against sessile P. gingivalis. The results indicated that the 3D-printed porous TC4-6Cu alloy could cause bacterial fragmentation and deformation. Plate antimicrobial counting experiments showed that the antibacterial rates of the alloy against adherent bacteria and planktonic bacteria after 24 h were 98.05% and 73.92%, respectively. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of Cu2+ were tested to appraise the antibacterial property of the alloy against planktonic P. gingivalis. The relationship between the antibacterial mechanism of the alloy with oxidative stress was evaluated through ROS fluorescence intensity and protein leakage concentration. The results revealed that the alloy significantly eliminated adherent bacteria and inhibited biofilm formation. Moreover, 3D-printed porous TC4-6Cu alloy demonstrated significant bactericidal ability by inducing the production of reactive oxygen species (ROS), which could result in protein leakage from the bacterial cell membrane. This research may open a new perspective on the development and biomedical applications for dental implantation

Cost-Effectiveness Analysis of Capecitabine Plus Oxaliplatin Versus Gemcitabine Plus Oxaliplatin as First-Line Therapy for Advanced Biliary Tract Cancers

Background: In the first-line treatment of biliary tract cancers (BTCs), XELOX (capecitabine plus oxaliplatin) showed comparable clinical efficacy and safety to gemcitabine and oxaliplatin (GEMOX), with fewer visits and better treatment management. Our study aims to investigate the cost-effectiveness of XELOX and GEMOX as the first-line therapy for BTCs from the perspective of the Chinese healthcare systems and to provide valuable suggestions for clinical decision-making.Methods: A Markov model was developed using the phase 3 randomized clinical trial (ClinicalTrials.gov number, NCT01470443) to evaluate the cost-effectiveness of XELOX and GEMOX. Quality-adjusted life-years (QALYs) and incremental cost-effectiveness ratios (ICERs) were used as the primary outcomes of the model. Uncertainty was assessed using univariate and probabilistic sensitivity analysis.Results: The QALYs for the XELOX and GEMOX groups were 0.66 and 0.54, respectively. In China, the total cost of XELOX treatment is US $12,275.51, which is lower than that of the GEMOX regimen. In addition, XELOX is more effective than GEMOX, making it the preferred regimen. A sensitivity analysis determined that XELOX therapy has a stable economic advantage in China.Conclusion: Compared to GEMOX, XELOX is a more cost-effective treatment as a first-line treatment for advanced BTC from the perspective of the Chinese health service system

Viral FLICE Inhibitory Protein of Rhesus Monkey Rhadinovirus Inhibits Apoptosis by Enhancing Autophagosome Formation

Rhesus monkey rhadinovirus (RRV) is a gamma-2 herpesvirus closely related to human herpesvirus 8 (HHV8). RRV encodes viral FLICE inhibitory protein (vFLIP), which has death effector domains. Little is known about RRV vFLIP. This study intended to examine its function in apoptosis. Here we found that RRV vFLIP inhibits apoptosis induced by tumor necrosis factor-α (TNF-α) and cycloheximide. In HeLa cells with vFLIP expression, the cleavage of poly [ADP-ribose] polymerase 1 (PARP-1) and activities of caspase 3, 7, and 9 were much lower than those in controls. Cell viability of HeLa cells with vFLIP expression was significantly higher than control cells after apoptosis induction. However, RRV vFLIP appears unable to induce NF-κB signaling when tested in NF-κB reporter assay. RRV vFLIP was able to enhance cell survival under starved conditions or apoptosis induction. At early time points after apoptosis induction, autophagosome formation was enhanced and LC3-II level was elevated in cells with vFLIP and, when autophagy was blocked with chemical inhibitors, these cells underwent apoptosis. Moreover, RRV latent infection of BJAB B-lymphoblastoid cells protects the cells against apoptosis by enhancing autophagy to maintain cell survival. Knockdown of vFLIP expression in the RRV-infected BJAB cells with siRNA abolished the protection against apoptosis. These results indicate that vFLIP protects cells against apoptosis by enhancing autophagosome formation to extend cell survival. The finding of vFLIP’s inhibition of apoptosis via the autophagy pathway provides insights of vFLIP in RRV pathogenesis

Analysis of viral proteins of porcine reproductive and respiratory syndrome virus in host protection and early virus-cell interactions

The viral proteins of porcine reproductive and respiratory syndrome virus (PRRSV) were expressed in a baculovirus expression system. They were evaluated for protection of pigs against the respiratory disease and for function in virus-cell interactions. The open reading frames 2 to 6 products were confirmed to be membrane associated proteins as they were detected on the surface of viable insect cells by an immunofluorescence assay. Immunization of pigs with vaccines comprised of a combination of GP2, GP3 and GP4 or a combination of GP5, M and N proteins induced partial protection, but immunization with combination of GP5 and M proteins failed to induce protection. Immunization of pigs with the first two vaccines significantly reduced the severity of clinical respiratory disease (P < 0.001), severity of pneumonia (P < 0.05) and duration of viremia compared with those of control group. These results suggest that in addition to envelope proteins, N protein may also play an important role in inducing host protection. For further characterization of these proteins, monoclonal antibodies (MAbs) were prepared. Most of the MAbs were found to be against conformationally dependent epitopes. Antigenic variation was found in PRRSV field isolates and thirty-three PRRSV isolates were divided into three groups based on their reactivity with the MAbs to GP2, GP3 and M proteins. Early virus-cell interactions in PRRSV infection was evaluated with a virus binding assay on CRL11171 cells. The recombinant ORFs 2 to 6 products failed to block PRRSV binding, implying that there were necessary interactions between viral structural proteins. PRRSV receptor was shown to contain protein moieties as protease treatment of cell monolayers reduced virus binding. The receptor activity was reduced from the cell surface with octylglucoside treatment, but no specific protein band was found by immunoprecipitation of the cell surface extracts, suggesting that the PRRSV receptor is present in low concentration. An 18-kDa viral protein was found to bind to the cells by an adsorption assay. Glycosylation did not affect the binding ability of this protein. The protein was immunoprecipitated with MAb against PRRSV M protein, suggesting its potential involvement in virus binding and/or entry.</p