Objective:To establish an antibody expression system to reduce the antibody-dependent enhancement (ADE) effect of target antibody.Methods:Site-directed mutagenesis was used to mutate the 234 and 235 sites of the Fc region of the mammalian cell antibody expression vector-L234A and L235A to establish the antibody expression vector pFRT-IgG1κ-FcM. An antibody Wt-WNV with significant ADE effect obtained in previous work was selected and expressed by the pFRT-IgG1κ-FcM system to obtain mutant antibody FcM-WNV. The binding ability of FcM-WNV to target antigen West Nile virus envelope protein-DⅢ (WNV E-DⅢ) was detected by ELISA, and the its binding ability to human high-affinity IgG Fc receptor hFcγRⅠ (hCD64 ) was analyzed by flow cytometry. The neutralizing activity of FcM-WNV in vitro was detected by pseudovirus infection of host cells (BHK21 and K562). Results:The expression levels of FcM-WNV and Wt-WNV were comparable, and FcM-WNV could recognize and bind to WNVE-DIII in a concentration-dependent manner. Compared with Wt-WNV, the binding ability of FcM-WNV to hCD64 was significantly weakened, showing a significant decrease in fluorescence intensity. Consistent with the previous experimental results, Wt-WNV at a concentration of 5 μg/ml significantly enhanced the infection of K562 by WNV pseudovirus, while FcM-WNV at a concentration of 5 μg/ml could effectively block pseudovirus infection in both K562 and BHK21 cells.Conclusions:The established antibody expression system can effectively reduce the ADE effect of the target antibody.

Objective:To establish an in vivo infection model of H5N1 pseudovirus and to detect the neutralizing activity of FHA3 antibody using this model. Methods:Based on the sequence information of hemagglutinin (HA) and neuraminidase (NA) of A/Anhui/1/2005/H5N1 strain, two recombinant plasmids of pcDNA3.1-HA5 and pcDNA3.1-NA1 were constructed. The two plasmids and plasmid pNL4-3.Luc.R-E- were co-transfected into 293T cells to prepare H5N1 pseudovirus supernatant. The morphology of pseudovirus particles in the supernatant was observed by electron microscopy. MDCK cells were infected with the pseudovirus supernatant and the virus titer was detected. BALB/c mice were injected with the pseudovirus supernatant by intraperitoneal injection and subjected to bioluminescence imaging at 2, 5, 8, and 12 d after infection to detect the pseudovirus infection in vivo. The functional activity of FHA3 antibody in vivo was evaluated using the established mouse infection model. Results:The recombinant plasmids pcDNA3.1-HA5 and pcDNA3.1-NA1 were correctly constructed and could be used to prepare pseudovirus supernatants of high titer by co-transfecting 293T cells with the plasmid pNL4-3.Luc.R-E-. The virus particles were round under electron microscope. H5N1 pseudovirus-infected mice exhibits strong fluorescence signals, which were attenuated by FHA3 treatment before challenge.Conclusions:The in vivo infection model of H5N1 pseudovirus was successfully constructed and FHA3 antibody was proved to be protective against the pseudovirus infection.

Flaviviruses are a class of positive-strand RNA viruses mainly transmitted by arthropods, which can cause high mortality and morbidity worldwide. At present, there is no specific therapy. Therapeutic antibodies bring hope for the treatment of flavivirus infection, but the antibody-dependent enhancement (ADE) effect induced by flavivirus infection can lead to disease progression. The ideal therapeutic antibodies against flaviviruses should not only treat flavivirus infection, but also avoid the harm caused by ADE. Therefore, researchers have optimized some of the antibodies to seek the best therapeutic antibodies. This review briefly described the research progress and mechanism of therapeutic antibodies against flaviviruses as well as some strategies to reduce the ADE effect induced by the therapeutic antibodies.

Neutralizing monoclonal antibodies against dengue virus cross-react with other flaviviruses due to the sequence homology between them, such as the antibodies targeting envelope protein and non-structural protein 1. Apart from exerting protective effects in infected animals, cross-neutralizing antibodies could also cause antibody-dependent enhancement (ADE) in vivo. This review summarized the progress in cross-reactivity of neutralizing antibodies against dengue virus.

Objective:To prepare and identify a broad-spectrum antibody FHA3 targeting influenza A virus hemagglutinin (HA).Methods:According to the single-chain antibody fragment (scFv) sequence, the heavy chain (VH) and light chain (VL) variable regions of FHA3 were amplified by PCR and a recombinant plasmid pFRT-IgG1κ-FHA3 was constructed by linking the expression vector pFRT-IgG1κ. FHA3 was expressed in the ExpiCHO system and purified by affinity purification. The binding activity of FHA3 to influenza A virus HA was detected by ELISA. The neutralizing activity of FHA3 was detected in vitro by infecting host cells with pseudovirus. Results:SDS-PAGE showed that high-purity FHA3 was obtained. FHA3 could bind to H1N1 HA, H2N2 HA, H3N2 HA, H5N1 HA, H7N9 HA and H9N2 HA in a concentration-dependent manner. FHA3 had good neutralizing activity in vitro that was it could effectively block the invasion of H5N1 and H7N9 pseudoviruses into target cells at a low concentration of 5 μg/ml and H1N1 pseudovirus at 0.012 5 μg/ml. Conclusions:A broad spectrum antibody targeting HA protein of influenza A virus with neutralizing activity in vitro was obtained.

Objective:To prepare and identify a functional antibody FNA1 targeting the neuraminidase (NA) of influenza A virus N1 subtype.Methods:According to single-chain antibody fragment (scFv) sequence, the heavy chain and light chain variable region sequences of FNA1 were synthesized, and the recombinant expression plasmid pFRT-IgG1κ-FNA1 was constructed by linking the expression vector pFRT-IgG1κ. The FNA1 antibody was expressed in ExpiCHO cells and purified using affinity purification technique. The binding ability of FNA1 to the target proteins, influenza A virus N1 subtype NA antigens, was detected by ELISA. Flow cytometry was performed to analyze the binding ability of FNA1 to the NA antigens expressed on the surface of cell membrane. The in vitro activity of FNA1 against NA was evaluated by infecting 293T cells with pseudovirus. Results:Protein electrophoresis showed that FNA1 with high purity was obtained. FNA1 specifically recognized and bound to N1 subtype NA antigens in a concentration-dependent manner. FNA1 could effectively block NA activity by binding to N1 subtype NA protein expressed on the surface of cell membrane, thus inhibiting the release of packaged pseudovirus from cell surface and further inhibiting target cell infection.Conclusions:An antibody FNA1 targeting influenza A virus N1 subtype NA with in vitro functional activity was obtained.

Objective? To explore the effects of home visit directed by Peplau interpersonal relationship model on negative emotions and medication adherence in elderly bronchial asthma patients. Methods? From May 2015 to May 2017, we selected 90 elderly bronchial asthma patients in the First Affiliated Hospital of Zhengzhou University as subjects by convenience sampling. All of the patients were divided into observation group and control group with the method of random number table, 45 cases in each group. Observation group carried out home visit nursing based on the Peplau interpersonal relationship model for elderly bronchial asthma patients. Control group adopted routine follow-up care. The medication adherence and disease cognition of patients both groups were recorded with the Medication Adherence Report Scale for Asthma (MARS-A) as well as Insight and Treatment Attitude Questionnaire (ITAQ), and the anxiety and depression of patients both groups before and after intervention were assessed with the Hamilton Anxiety Scale (HAMA) and Hamilton Depression Scale (HAMD). We also observed the incidence of bronchial asthma acute attack in patients one year after discharge. Results? When home visit was completed and one year after discharge, the medication adherence of observation group was higher than that of control group, and the score of ITAQ was higher than that of control group and that at discharge with statistical differences (P＜ 0.05). One year after discharge, the incidence of bronchial asthma acute attack of observation group was lower than that of control group with a statistical difference (P＜ 0.05). When home visit was completed and one year after discharge, the scores of HAMA and HAMD of observation group were lower than those of control group with statistical differences (P＜0.05). Conclusions? Home visit directed by Peplau interpersonal relationship model help to reduce the incidence of acute attack of elderly bronchial asthma patients external hospital, relieve the negative emotions and improve disease cognition as well as medication adherence.

Objective? To explore the application of three-dimensional quality assessment model of structure-process-outcome in continued nursing care for elderly patients with lung cancer after operation. Methods? By purposive sampling, 48 elderly patients discharged after radical resection of lung cancer from January to December 2016 were taken as control group and received routine continuous nursing care. Another 51 elderly patients discharged after radical resection of lung cancer from June 2017 to June 2018 were taken as the observation group. Continuous nursing program based on structure-process-outcome quality assessment model was applied for the observation group. The effect of intervention was assessed by using Strategies Used by People to Promote Health (SUPPH) and Quality of Life Questionnaire-30(QLQ-30). Results? At 6 months after discharge, the scores of each dimension of SUPPH scale in the observation group were statistically higher than those in the control group (P＜ 0.05). In the observation group the scores of "physical function", "role function", "social function" and "overall health" of QLQ-30 scale were higher than the control group, the scores of "fatigue", "pain", "insomnia", "lack of appetite", "constipation", and "diarrhea" were all lower than the control group with statistical significance (P＜0.05). Conclusions? The continuous nursing care based on the three-dimensional quality assessment model of structure-process-outcome for elderly patients with lung cancer can help to improve their sense of self-efficacy and quality of life.

Objective To explore the application effects of nursing team in patients with acute multiple trauma in multiple disciplinary team (MDT) model. Methods A convenient sampling method was used to select 82 patients with multiple injuries who were treated since the establishment of the trauma center in the First Affiliated Hospital of Zhengzhou University from July 2016 to June 2018. From July 2014 to June 2016, a total of 55 patients with multiple trauma before the establishment of the trauma center were seclected as the control group. The experimental group was used MDT model for rescue, and the control group was used traditional model for rescue. The doctors' arrival time, emergency examination time, emergency stay time, rescue success rate and postoperative complication rate of the specialists in the two groups were compared. Results The doctors' arrival time in the experimental group was (16.80±3.57) min, the time of emergency examination was (22.36±3.49) min, and the time of emergency stay was (38.19±8.18) min, which was shorter than those of the control group (23.27±5.76), (45.69±7.75), (55.49±13.67) min, the differences between the two groups were statistically significant (P＜0.01). The successful rescue rate of the treatment group was 96.3% (79/82), which was higher than 87.3% (48/55) of the control group. The difference between the two groups was statistically significant (P＜ 0.05). The postoperative complication rate was 3.7% (3/82) in the experimental group and 12.7% (7/55) in the control group, the difference between the two groups was statistically significant (P＜ 0.05). Conclusions Patients with multiple trauma received treatment with MDT model can effectively shorten the time of doctors' arrival, emergency examination time, emergency stay time, improve the success rate of treatment, and reduce the incidence of postoperative complications.

Objective To construct a shRNA lentiviral vector targeting the gene encoding tumor necrosis factor alpha-induced protein 8 (TNFAIP8) in RAW264. 7 cells, a mouse macrophage cell line, and to investigate the effects of TNFAIP8 gene silencing on the functions of mouse macrophages. Methods The shRNA sequence targeting TNFAIP8 gene was designed and DNA oligos containing small hairpin frame was synthesized. The double-stranded DNA was cloned into pLKO. 1-TRC vector after annealing. The recombi-nant vector was verified by using double enzyme digestion and gene sequencing. Lentiviruses were prepared by transfecting the constructed vector into 293T cells. Fluorescent quantitative RT-PCR and Western blot as-say were performed to detect the expression of TNFAIP8 at mRNA and protein levels after infecting the RAW264. 7 cells with lentiviruses. Flat dish adhesion experiment and wound-healing assay were used to evaluate the effects of TNFAIP8 gene silencing on the adhesion and migration of RAW264. 7 cells. Results The recombinant lentiviral vector was successfully constructed as indicated by double enzyme di-gestion and gene sequencing analysis. The expression of TNFAIP8 in RAW264. 7 cells at both mRNA and protein levels were significantly down-regulated after lentivirus infection (P<0. 05). Moreover, TNFAIP8 gene silencing significantly impaired the cell adhesion ability of RAW264. 7 cells after 15 min, 30 min or 2 hours of culture. Compared with the cells in control group, the RAW264. 7 cells harboring silenced TN-FAIP8 gene looked round with a smaller number of cellular extensions. The wound-healing assay showed that less TNFAIP8 gene-silenced RAW264. 7 cells migrated into the wounded area as compared with the cells in control group after 24 hours of culture (P<0. 05). The wound-healing rates of the experimental and control groups were 25% and 50%, respectively. Conclusion The recombinant lentiviral vector containing shRNA targeting the TNFAIP8 gene was successfully constructed. Transfecting the RAW264. 7 cells with the con-structed vector significantly silenced the expression of TNFAIP8 gene and inhibited the adhesion and migra-tion of these cells.