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 West Nile virus (WNV) pseudovirus and evaluate the neutralizing activity of antibody WNV-XH1.Methods:A stable cell line that can package the WNV pseudovirus was established in the early stage to prepare the pseudovirus supernatant. The supernatant was concentrated and infected BHK21 cells to detect the titer of the pseudovirus. After intraperitoneal injection of the pseudovirus into C57BL/J mice, bioluminescence imaging was performed to observe the infection status of the pseudovirus in the mice. After simultaneous infection, blood was collected and ELISA was used to detect NS1 levels in mouse serum. The in vivo functional activity of antibody WNV-XH1 was evaluated using the established mouse infection model.Results:Fluorescence was detected in C57BL/J mice infected with WNV pseudovirus, and the NS1 levels in the peripheral blood serum of mice infected with pseudovirus were significantly higher than those of non infected mice (1.453±0.09vs0.305±0.018). After intravenous administration of WNV-XH1 antibody before the attack, the fluorescence signal in the mice decreased and the serum NS1 level decreased (0.384±0.015).Conclusions:A successful in vivo infection model of WNV pseudovirus was established, and it was confirmed that the antibody WNV-XH1 had a protective effect against WNV pseudovirus infection in vivo.

Patients with severe trauma require an extremely timely treatment and transfusion plays an irreplaceable role in the emergency treatment of such patients. An increasing number of evidence-based medicinal evidences and clinical practices suggest that patients with severe traumatic bleeding benefit from early transfusion of low-titer group O whole blood or hemostatic resuscitation with red blood cells, plasma and platelet of a balanced ratio. However, the current domestic mode of blood supply cannot fully meet the requirements of timely and effective blood transfusion for emergency treatment of patients with severe trauma in clinical practice. In order to solve the key problems in blood supply and blood transfusion strategies for emergency treatment of severe trauma, Branch of Clinical Transfusion Medicine of Chinese Medical Association, Group for Trauma Emergency Care and Multiple Injuries of Trauma Branch of Chinese Medical Association, Young Scholar Group of Disaster Medicine Branch of Chinese Medical Association organized domestic experts of blood transfusion medicine and trauma treatment to jointly formulate Chinese expert consensus on blood support mode and blood transfusion strategies for emergency treatment of severe trauma patients ( version 2024). Based on the evidence-based medical evidence and Delphi method of expert consultation and voting, 10 recommendations were put forward from two aspects of blood support mode and transfusion strategies, aiming to provide a reference for transfusion resuscitation in the emergency treatment of severe trauma and further improve the success rate of treatment of patients with severe trauma.

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.

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 analyze the status and influencing factors of early out of bed activity in patients after coronary artery bypass grafting.Methods:Using the convenient sampling method, patients who underwent coronary artery bypass grafting in Affiliated Hospital of Jining Medical University from January 2018 to December 2020 were selected as the research objects. The general situation questionnaire, the Chinese version of Brief Disease Cognition Questionnaire, Muscle Strength Rating Scale and Visual Analog Scale were used to investigate them. Binomial Logistic regression was used to analyze the influencing factors of early out of bed activity after coronary artery bypass grafting.Results:A total of 100 questionnaires were distributed in this study and 100 were recovered. The effective recovery rate of the questionnaire was 100%. 38% (38/100) of patients with coronary artery bypass grafting performed early out of bed activity within 72 hours after surgery. Binomial Logistic regression analysis showed that vascular bridge selection, postoperative pain, lower limb incision blister, lower limb muscle strength rating and disease awareness were the influencing factors of early out of bed activity after coronary artery bypass grafting ( P<0.05) . Conclusions:Vascular bridge selection, postoperative pain, lower limb incision blister, lower limb muscle strength rating and disease awareness were the influencing factors of early out of bed activity after coronary artery bypass grafting

【Objective】 To investigate the blood compatibility of transfusion sets for single use, the loss, damage and activation of blood components after passage through the transfusion sets. 【Methods】 Transfusion sets (sample A and B) were assessed by comparing samples of blood component taken prior to and after passage through. The following makers of damage/activation were evaluated: Red blood cells (RBCs)-supernatant free hemoglobin (FHb) and potassium(K⁺ ) amount; platelet concentrates (PCs)-pH, hypotonic shock response (HSR), supernatant lactate dehydrogenase(LDH), CD62P; fresh frozen plasma(FFP) – prothrombin fragments 1 and 2 (F1+ 2), fibrinopeptide A (FPA), coagulation factor Ⅻa (FⅫa), Thrombin-antithrombin complex (TAT). 【Results】 After passing through two types of transfusion sets, the loss of RBCs, PCs and FFP were less than 5%. There is no statistic difference for the change of FHb and K⁺ of RBCs(P>0.05). There were no statistics for pH, LDH(U/L) and CD62P(%) after platelet concentrates passing through both transfusion sets. There was no statistics for the HSR after passing through the sample A, while there was a small but statistically significant increase in the HSR of sample B (37.17±6.49 vs 40.75±6.24, P<0.05). After FFP passing through the transfusion giving sets, there were no statistical difference for F1+ 2, FPA, FⅫa and TAT (P>0.05). 【Conclusion】 Two types of transfusion sets caused negligible effects on RBCs, platelet concentrates and FFP.

Objective To evaluate the feasible cervical cancer screening strategies in rural China. Methods The study was based on the health industry scientific research project of National Health Commission in 2015, cervical cancer screening technology and demonstration research suitable for rural areas in China, we collected health economics and epidemiological parameters and established the unscreening model and screening model with Treeage Pro 2011 software. Combining with the data acquired from site investigation, including population screening, treatment-related clinical materials and cost data, we simulated the occurrence and the development of cervical cancer of rural women in China under different screening and intervention programs and predicted the screening effects [cumulative incidence, cumulative risk of disease, life years and quality adjusted life years (QALY), gains] and costs after 20 years, and using health economic evaluation analysis (cost-effectiveness analysis, cost-utility analysis, cost-benefit analysis). Screening programs included five screening strategies [visual inspection with acetic acid/lugol's iodine (VIA/VILI), careHPV, ThinPrep cytology test (TCT), careHPV+TCT, careHPV+VIA/VILI] and three screening intervals (1-year, 3-year, 5-year), a total of fifteen screening programs. Results Compared with no screening, fifteen screening programs reduced the cumulative incidence by 22.65%-51.76%. Compared with TCT or VIA/VILI, for the same screening interval, the reduced cumulative incidence, the amounts of life-year saved and QALY and benefits gained of careHPV were the highest. The cost-effectiveness ratios of these screening programs ranged (0.44-3.24)×104 Yuan per life-year saved, cost-utility ratios ranged (0.15-1.01)×104 Yuan per QALY, benefit-cost ratios ranged 7.73-59.10. The results of incremental cost-effectiveness ratios showed that VIA/VILI every five years, VIA/VILI every three years, careHPV every five years, careHPV every three years and careHPV every year were dominant programs. Conclusions VIA/VILI screening is cost-effective, careHPV is slightly more expensive but more effective. In rural China, careHPV screening every five years could be recommended. This study provides a basis for the determination of cervical cancer screening methods feasible for rural areas in China.

Objective: To evaluate the feasible cervical cancer screening strategies in rural China. Methods: The study was based on the health industry scientific research project of National Health Commission in 2015, cervical cancer screening technology and demonstration research suitable for rural areas in China, we collected health economics and epidemiological parameters and established the unscreening model and screening model with Treeage Pro 2011 software. Combining with the data acquired from site investigation, including population screening, treatment-related clinical materials and cost data, we simulated the occurrence and the development of cervical cancer of rural women in China under different screening and intervention programs and predicted the screening effects [cumulative incidence, cumulative risk of disease, life years and quality adjusted life years (QALY) , gains] and costs after 20 years, and using health economic evaluation analysis (cost-effectiveness analysis, cost-utility analysis, cost-benefit analysis). Screening programs included five screening strategies [visual inspection with acetic acid/lugol's iodine (VIA/VILI), careHPV, ThinPrep cytology test (TCT), careHPV+TCT, careHPV+VIA/VILI] and three screening intervals (1-year, 3-year, 5-year), a total of fifteen screening programs. Results: Compared with no screening, fifteen screening programs reduced the cumulative incidence by 22.65%-51.76%. Compared with TCT or VIA/VILI, for the same screening interval, the reduced cumulative incidence, the amounts of life-year saved and QALY and benefits gained of careHPV were the highest. The cost-effectiveness ratios of these screening programs ranged (0.44-3.24)×10⁴ Yuan per life-year saved, cost-utility ratios ranged (0.15- 1.01)×10⁴ Yuan per QALY, benefit-cost ratios ranged 7.73-59.10. The results of incremental costeffectiveness ratios showed that VIA/VILI every five years, VIA/VILI every three years, careHPV every five years, careHPV every three years and careHPV every year were dominant programs. Conclusions VIA/VILI screening is cost-effective, careHPV is slightly more expensive but more effective. In rural China, careHPV screening every five years could be recommended. This study provides a basis for the determination of cervical cancer screening methods feasible for rural areas in China.