The mechanism of interactions between natural killer (NK) cells and dendritic cells (DCs) in regulating the early stage of innate immune response and the subsequent adaptive immune response has been elucidated in recent years. NK-DC interactions result in the activation of NK cells and the maturation and/or apoptosis of DCs through direct cell-to-cell contact and releasing cytokines. NK cells can control and enhance DC-mediated antiviral immune response by inducing DCs to mature in favor of Th1 immune response and providing antigens to DCs and killing immature DCs (iDCs). DCs stimulate NK cells through soluble and cell-contact activators, thereby increasing their proliferation, survival and cytotoxicity to shape the innate immune response. Recent studies have shown that interactions between NK cells and DCs also play a critical role in several antiviral responses. Therefore, this paper reviewed the NK-DC interactions and their relationship with antiviral responses, thus providing a theoretical basis for studying the molecular mechanism of NK-DC interactions in viral infectious diseases.

Objective To analyze how enterovirus D68 (EV-D68) protease 2A affects the anti-vi-ral interferon typeⅠ(IFN-Ⅰ) pathway in 293T cells following infection. Methods Western blot was used to detect the expression of recombinant protease 2A, IFN-α and signal transducers and activators of tran-scription 1 (STAT1) at protein level. Expression of EV-D68 viral protein (VP1) and protease 2A was ana-lyzed by immunofluorescence at different time points. Cytopathic effects were recorded to calculate 50％ cell culture infective dose ( CCID50 ) . Expression of the genes involved in the anti-viral IFN-Ⅰ pathway was measured by real-time PCR (RT-PCR). Results The recombinant plasmid pCLIPf-2A was successfully constructed and the expression of recombinant protease 2A could be detected by Western blot 24 h after transfection. The recombinant protease 2A promoted the proliferation of EV-D68 at the late stage of infection and induced the production of IFN-α. Expression of the genes involved in the anti-viral IFN-Ⅰ pathway at mRNA level was up- or down-regulated to different degrees with various trends in different groups following infection. Expression of STAT1 was enhanced in all groups. Conclusions EV-D68 protease 2A promoted the activation of anti-viral IFN-Ⅰpathway in response to viral infection and enhanced the proliferation of virus at the late stage of infection.

Objective To investigate the role of CXC chemokine ligand 5 (CXCL5) in the patho-genesis of dextran sulfate sodium (DSS)-induced inflammatory bowel disease (IBD). Methods A mouse model of IBD was established by giving 3% DSS in drinking water. Influences of CXCL5 knockout on mouse body weight, clinical symptoms, survival rate, pathological injury and the secretion of inflammatory cyto-kines were analyzed. Results CXCL5 levels in serum of mice with DSS-induced IBD were significantly higher than those of the normal control group. DSS-induced weight gain, death, pathological damages and inflammatory cytokine secretion were alleviated in mice after knocking out CXCL5. Conclusion CXCL5 might promote the secretion of inflammatory cytokines in mice with DSS-induced acute colitis and aggravate pathological damages,suggesting that CXCL5 might be a potentially important candidate target for the treat-ment of IBD.

Objective To investigate the characteristics of VP1 gene hypervariable region in hu-man enterovirus type 68(HEV68)strains isolated in China. Methods Nucleotide sequences of the VP1 gene in the Chinese strains and strains isolated in other countries were aligned by using Clustal W in the MEGA6 program. The phylogenetic trees were constructed by using Neighbor-Joining(NJ)method in the MEGA6 program. Sequence of the amino acids encoded by that region was analyzed by compared with that of the standard strain Fermon. Results A total of 80 strains of EV68 had been isolated in China by the end of 2015. Most of the mutations occurred in BC and DE loops. The mutation sites lied in the VP1 gene of Chi-nese isolates were at 83,89,91,94,96,97,98,102,109,139,141,142,143,144 and 147. Glycine was missing from most of the amino acid sequences encoded by the VP1 gene of Chinese strains. The phylo-genetic analysis indicated that 53 and 21 EV68 strains isolated in China belonged to B and C clades,respec-tively. Conclusion Compared with the standard strain Fermon,the Chinese strains changed a lot in BC-loop and DE-loop,which were associated with the antigenicity and virulence of EV68. The EV68 strains iso-lated in China belonged to B and C clades.

Objective To express and purify the recombinant UL7 protein of herpes simplex virus 1 (HSV-1), to prepare the corresponding UL7-specific polyclonal antibody and to preliminarily analyze the expression of UL7 protein during the proliferation of HSV-1. Methods The UL7 gene was amplified by PCR and then cloned into the pGEX-5X-1 vector for expression of UL7 protein in the prokaryotic expression system. The constructed expression plasmid, pGEX-5X-1-UL7, was transformed into E. coli BL21 (DE3) to induce the expression of UL7 protein by IPTG. The purified GST-UL7 fusion protein was used as antigen to inject the ICR mouse for the preparation of polyclonal antibody specific for UL7 protein. The titer and speci-ficity of the polyclonal antibody were analyzed by using indirect ELISA and Western blot assay, respectively. The UL7 protein-specific polyclonal antibody was used to detect the expression of UL7 protein at different time points after infecting Vero cells with HSV-1. Results The GST-UL7 fusion protein was efficiently ex-pressed in E. coli BL21 (DE3). The UL7 protein-specific polyclonal antibody was prepared with high titer (1 ∶ 105) and high specificity as indicated by the indirect ELISA and Western blot assay. The expression of UL7 protein was detected at different time points after infecting Vero cells with HSV-1. Conclusion The GST-UL7 fusion protein was obtained successfully and the UL7 protein-specific polyclonal antibody was pre-pared. Accompany with the proliferation of HSV-1, the expression of UL7 protein was detected at different time points by using the polyclonal antibody.

Objective To establish a non-traumatic mouse model of acid aspiration-induced lung injury which al-lows longitudinal studies.Method C57BL/6 mice were anesthetized and orotracheally intubated with a 20 gauge angio-catheter guided by optical fiber.The mice were subsequently placed in the right lateral decubitus position and external com-pression to the left lung was manually applied.A polyethylene catheter was advanced into the right lung and used to instill either hydrochloric acid (2.5μL/g, 0.1 mol/L, pH 1.5) or saline as control.Then the mice were recovered with supple-mental oxygen for 4 hours.The pulmonary physiological function and survival of mice within 2 weeks after surgery were as-sessed.Results Methylene blue instillation showed that the staining fluid went into the right lung of the non-traumatically intubated mice.The survival rate of the mice with non-traumatic instillation was 80%, statistically significantly higher than those with tracheostomy instillation.Histological examination and lung function ( wet/dry ratio, elastance and arterial oxy-gen saturation) assay demonstrated that acid instillation caused a profound pathological changes and functional impairment of the lung.Besides, acid aspiration into the mouse lung caused a significant increase in neutrophil infiltration in mouse pulmonary alveoli and high concentrations of inflammatory factors (TNF-α, IL-6, CXCL1 and CXCL2) in the bronchoalve-olar lavage fluid.Conclusions We successfully established a mouse model of acid aspiration-induced lung injury, which may serve as a reliable model for longitudinally studying pulmonary immune-inflammatory mechanism in humans.

Objective To investigate the epidemic patterns and the characteristics of influenza in Chi-na through a Meta-analysis based on the studies published in domestic literatures.Methods Related articles published during 2005 to 2012 were screened out from domestic databases and analyzed through a Meta-analysis with Review Manager 5.0 software.Results Twenty-two articles covering 957 901 patients with influenza-like-illness (ILI) and 148 233 pathogen samples were screened out according to the inclusion criteria.No significant difference with the ILI diagnosis rate was found between subjects at age 0-4 years and those at age 15-59 years. Higher ILI diagnosis rates were observed in those two groups as compared with subjects elder than 60 years old. Most of the pathogen samples were carried by subjects aged 25-59 years.More influenza virus strains were isola-ted in 2009 as compared with those of the seven other years (OR=2.25, 95%CI=1.27-3.70).There was sta-tistical difference between the numbers of influenza A H1N1 and seasonal influenza A strains (OR=2.25, 95%CI=1.30-3.91) .Significant difference was also observed between the numbers of influenza A and influenza B strains (OR=4.05, 95%CI=2.53-6.47).Conclusion There was significant difference with the diagnosis rate between subjects aged 0-4 years and those aged≥60 years.More attention should be paid to people at high risk of infection (0-4 years old and≥60 years old) and those at 25-29 years with high mobility and social inter-course for the timely prevention and control of pandemic influenza.The detection rate of influenza virus strains was increased during the outbreak of novel influenza A H1N1 infection in 2009.After that outbreak, the detec-tion rate of novel influenza A H1N1 strains was 2.25 times the rate of seasonal influenza strains.The detection rate of influenza A was 4.05 times the rate of influenza B virus strains.Therefore, it is necessary to strengthen the surveillance for influenza A virus and other epidemic influenza virus strains.

Objective To establish adapted strain of seasonal influenza virus influenza A virus [A/ Hong Kong/1968 (H3N2)] in mice and explore the molecular mechanism of adaption preliminary.Methods The mice model was induced by nose dropping with wild type H3N2 influenza virus,by continuous passage in the lung of mice and detection of the clinical manifestation,viral load,virus titer,pathology,cytokines,to get the adapted strain of seasonal influenza A virus [A/Hong Kong/1968 (H3N2)].Results After 7 times continuous passage in the lung of mice,virulence was increased,viral load and virus titer were increased either.Genome sequencing and alignment indicated that the HA and NA gene was mutated.Conclusion The influenza virus H3N2 of mice lung adaption can be acquired from wild type seasonal H3N2 by continuous passage in the lung tissues of mice.Mutation of Ile to Val at residue 248 of HA and Val to Ile at residue 444of NA protein may be the key virulence determinant.

OBJECTIVETo investigate the effect of temperature on the stability of intermediate and final products of inactivated enterovirus 71 vaccine, which was prepared in human diploid cells.

METHODSThe different batches of harvest viral cultures, the vaccine stock solutions and the final productions of inactivated enterovirus 71 vaccine were stored at different temperatures. The samples of viral culture stored at -20°C or 4°C were harvested at 0, 6, 12 and 24 months later. The samples of vaccine stock solutions stored at -20°C were harvested at 0, 6, 12 and 24 months later, and that stored at 4°C were harvested at 0, 1, 3, 6 and 12 months later. The samples of finial products were harvested at different time points (0, 6, 12 and 24 months for storing at 4°C; 0, 7, 14, 28, 42 and 60 d for storing at 25°C; 0, 3, 7, 14 and 21 d for storing at 37°C). The viral titer, antigen content, antigen purity, endotoxin content, effectiveness, pH and appearance of samples were determined, respectively. A total of 1 800 BLAB/c mice were immunized by vaccine and 150 control mice were injected by diluents without antigen via intraperitoneal. The tail vein blood (500 µl per mouse) from 1 950 mice were harvested after 4 weeks post injected. The neutralization antibody titers of the serum were tested to calculate the half effective dose (ED50) of final products. All results were analyzed using analysis of variance to compare the differences of the above indexes.

RESULTSThe viral titers of harvest viral culture of inactivated EV71 vaccine were (6.67 ± 0.13), (6.56 ± 0.09), (6.52 ± 0.04), (6.39 ± 0.16) lgCCID50/ml (CCID50, the half cell culture infective dose) after 0, 6, 12 and 24 months storage at -20°C; and (6.67 ± 0.13), (6.41 ± 0.13), (6.19 ± 0.18), (5.97 ± 0.09) lgCCID50/ml at 4°C. The viral titers reduced with time (F = 9.81 or 44.16, P < 0.05). The antigen contents of the vaccine stock solution were maintained at (3 626.67 ± 1 382.56) EU/ml within 3 months at 4°C, but were (2 080.00 ± 876.36), (951.17 ± 346.35) EU/ml at 6 and 12 months, respectively. The ED50 of the final production were (31.00 ± 2.71), (32.93 ± 3.22), (39.37 ± 3.44) and (46.04 ± 3.25) EU/ml after 0, 6, 12 and 24 months storage at 4 °C, but were (31.00 ± 2.71), (32.23 ± 2.66), (34.70 ± 1.77), (40.04 ± 2.10), (47.78 ± 1.93) and (56.97 ± 0.50) EU/ml at 0, 7, 14, 28, 42 and 60 days at 25°C, and were (31.00 ± 0.00), (36.20 ± 0.00), (41.87 ± 0.50), (53.25 ± 0.50) and (64.84 ± 0.58) EU/ml at 0, 3, 7, 14 and 21 days at 37°C, respectively. The ED50 had increased with the time by and had significantly differences compared with the beginning level (F = 28.49, 215.15 or 156.12, P < 0.05).

CONCLUSIONThere is a good stability of the intermediate and final productions of inactivated enterovirus 71 (EV71) vaccines, within 24 months at -20°C or 6 months at 4°C storage for viral culture, 24 months at -20°C or 3 months at 4°C storage for stock solution and 24 months at 4°C or 28 d at 25°C or 7 d at 37°C storage for finial vaccine.

Animals ; Drug Storage ; methods ; Enterovirus A, Human ; Humans ; Immunization ; Mice ; Vaccination ; Vaccine Potency ; Vaccines, Inactivated

In light of the scarcity of reports on the interaction between HSV-1 nucleocapsid protein UL25 and its host cell proteins,the purpose of this study is to use yeast two-hybrid screening to search for cellular proteins that can interact with the UL25 protein.C9orf69,a protein of unknown function was identified.The interaction between the two proteins under physiological conditions was also confirmed by biological experiments including co-localization by fluorescence and immunoprecipitation.A preliminary study of the function of C9orf69 showed that it promotes viral proliferation.Further studies showed that C9orf69 did not influence viral multiplication efficiency by transcriptional regulation of viral genes,but indirectly promoted proliferation via interaction with UL25.