AIM: To probe the effect of azelastine hydrochloride on experimental asthma and its mechanism. METHODS: Experimental asthma models of guinea pigs induced by histamine and acetylcholine in vivo as well as guinea pig tracheal spirals in vitro were used in this experiment. RESULTS: Azelastine hydrochloride inhibited asthma induced by histamine and acetylcholine in guinea pigs in a dose dependent manner, prolonged the incubation period of histamine and acetylcholine induced asthma (P

Objective To observe the effects of bolus volume on pharyngeal and upper esophageal sphincter pressures and durations in healthy volunteers by using high-resolution manometry (HRM).Methods Twentyfour health subjects were recruited and asked to swallow three volumes of bolus (3 ml,5 ml and 10 ml) in the neutral head position.Pressure and duration measurements were acquired by utilizing a high-resolution solid-state manometer,with an emphasis on the hypopharynx and upper esophageal sphincter (UES).Variables including UES residual pressure,UES relaxation duration,maximum hypopharygeal pressure and hypopharyngeal pressure duration were analyzed across bolus volumes and consistencies by using three-way repeated measures analysis of variance (ANOVA) to investigate influence of bolus volume.Results UES residual pressure [-1.71 mmHg(3 ml thick liquid)vs.-4.68 mmHg(10 ml thick liquid)],UES relaxation duration[590.45 ms(3 ml thick liquid) vs.702.49 ms (10 ml thick liquid)],maximum hypopharygeal pressure [169.91 mmHg (3 ml thick liquid) vs.204.42 mmHg (10 ml thick liquid)] and hypopharyngeal pressure duration(P ＜0.05) varied significantly across bolus volumes when swallowing water or thick liquid.The UES relaxation duration,UES residual pressure and maximum hypopharyngeal pressure had a direct positive relationship with bolus volume.There was significant differences with regard to UES relaxation duration [685.75 ms(3 ml paste)vs.772.27 ms (10 ml paste)] but not to UES residual pressure (P ＞ 0.05) and maximum hypopharyngeal pressure (P ＞ 0.05) across bolus volume when swallowing paste.Conclusions Difference in hypopharyngeal pressure and duration,UES residual pressure and duration were detected across varying bolus volumes.Consideration of these variables is paramount in understanding normal and pathological swallowing.

M PU employed as control center,the nerve stimulator can produce different stimul ation modes easily.The high-quality constant-current pulse and the safety of th e patient are ensured through two-steps current stabilizer,pulse width monitor and the float to ground.Both needle electrode and surface electrode can be used to achieve supramaximal stimulation.

Objective To explore the diagnostic value of serum procalcitonin ,C‐reactive protein and white blood cell count in children with different diseases .Methods Retrospective analysis 94 cases of pathogenic infectious children from June 2013 to May 2014 in our hospital ,according to the results of pathogen detection was divided into bacterial infection 36 cases ,mycoplasma infec‐tion group 28 cases ,30 cases of viral infection ,detection and analysis serum PCT ,CRP and WBC levels .Results Bacterial infection group serum PCT ,CRP and WBC were (2 .41 ± 0 .94)ng/mL ,(47 .91 ± 18 .26)mg/L and (13 .18 ± 6 .03) × 109/L ,significantly higher than the mycoplasma infection and viral infection group (F=133 .4 ,F=60 .1 ,F=8 .5 ,P<0 .05);diagnosis of bacterial in‐fections ,PCT sensitivity and specificity were 92 .11% and 91 .05% ,positive and negative predictive value of 89 .84 % and 94 .01%were significantly higher than CPR and WBC ,Mycoplasma infection as the control group ,PCT ,CRP and WBC in the diagnosis of bacterial infections ,the area of under the ROC curves were 0 .816 ,0 .728 and 0 .614 ,respectively .Conclusion Serum PCT for the i‐dentification of bacterial infections has a high diagnostic value ,worth generalizing and applying .

Objective To explore the clinical feature of severe enterovirus 71 (EV71) associated hand foot and mouth disease (HFMD),and genotype of EV71.Methods Fluorescent quantitation PCR was done for detecting EV71.RT-PCR was performed to amplify VP1 for sequencing and identifying genotype.A retrospective analysis was performed based on the clinical data of 15 cases with severe EV71 infection.Results EV71 nucleotide was positive in all 15 cases.The genotype of EV71 was C4.All cases had abnormal temperature and followed with nervous symptoms in the early stage.Average time was 1.26 days from onset to severe complications appearance.Eleven cases progressed to neurogenic pulmonary edema.Four cases accepted nasal continuous positive airway pressure.Eleven cases accepted oral trachea cannula mechanical ventilation.Except for 3 cases died,one case abandoned,others 11 cases were cured.Conclusion The isolated strains of EV71 in this study are all C4 genotype.All cases with severe EV71 infection were followed with nervous symptoms in the early stage,most of whom would progress to neurogenic pulmonary edema.The mortality would be cut down by using mechanical ventilation in early stage.

In order to construct recombinant baculovirus carrying Schistosoma japonicum 26 ku glutathione S-transferase gene (Sj26), and observe the expression of Sj26 in mammalian cells, the Sj26 gene was amplified with plasmid pGEX-3X as template by PCR, and then recombined into T vector for sequencing. Sj26 gene was inserted into the downstream of CMV promoter of donor plasmid pFBDGC, and the recombinant donor plasmid pFBDGC-Sj26 transformed into DH10Bac, then the recombinant bacmid AcCMVSj26 was isolated and transfected into Sf9 cells. The recombinant baculovirus was harvested and final titer of vAcCMVSj26 was measured. BHK cells were transducted with recombinant baculovirus in vitro. By using Western blot, the expression of 26 ku glutathione S-transferase (GST) was detected. The results showed that after enzyme digestion and sequencing, the donor plasmid was successfully constructed. PCR confirmed that pFBDGC-Sj26 and Bacmid homologous recombination occurred in E. coli. After transfection of Sf9 cells with recombinant Bacmid, recombinant baculovirus was replicated in Sf9 cells and expressed green fluorescent protein. PCR further revealed recombinant baculovirus contained Sj26. The titer of the harvested baculovirus was 1.24 x 10(8). Western blot demonstrated that recombinant baculovirus could express 26 ku GST in BHK cells. It was concluded that Sj26 recombinant baculovirus was successfully constructed, and the 26 ku GST was expressed in mammalian cells.

AIM: To detect the effect of conjunction matrigel with mammary fad pat(MFP)implantation on the tumorigenesis, proliferation, apoptosis and metastasis of Her2 positive and negative breast cancer model. METHODS: The Her2 positive BT 474 and Her2 negative MDA-MB 231 breast cancer cells were injected into MFP of nude mice with or without matrigel to establish breast cancer model. The tumor volume was measured every 3 d. Followed up for 30 d after implantation, the nude mice were killed and the tumors and associated organs were dissected for pathological sectioning and staining with hematoxylin and eosin. The time of tumor formation and the tumorigenesis were determined after implantation. The tumor volume and metastasis rate were calculated and compared with each other. The proliferation and apoptosis of Her2 positive and negative tumors were also determined. RESULTS: Matrigel and MFP implantation technology shortened the time of tumorigenesis significantly(P<0.01). The tumorigenesis rate of BT 474 and MDA-MB 231 breast cancer cells did not show any different(P>0.05). The metastasis rate of MDA-MB 231 breast cancer cells were improved from 25.0% to 37.5%(P<0.05). CONCLUSION: Matrigel and MFP implantation can be combined to shorten the time of tumor formation by two kinds of breast cancer cells, and improve the metastasis of Her2 negative MDA-MB 231 cells. Using matrigel does not show any effect of proliferation and apoptosis on Her2 positive and negative breast cancer cells.

OBJECTIVETo investigate the expression of Toll-like receptor (TLR) mRNA in enterovirus 71(EV-A71) infected human Jurkat T cells and clarify the role of TLRs in the pathogenesis of EV-A71 infection-induced inflammation.

METHODSEV-A71 strains were isolated from feces of children patients with hand, foot and mouth disease in 2014 by Shenzhen Center for Disease Control and Prevention. Human Jurkat T cells were infected with 200 μl EV-A71 at 10(3) cell culture infective dose 50%(CCID50)/ml. The expression of TLR1-TLR10 mRNA in human Jurkat T cells was assessed at different exposure time by RT-PCR. Levels of TLR7 mRNA expression were detected by real-time PCR, and levels of myeloid differentiation factor 88 (MyD88) by western blot. The cytokine secretion of interleukin (IL)-6, IL-8 and Tumor Necrosis Factor α (TNF-α) was analyzed by ELISA assay.

RESULTSThe relative expression level of TLR7 mRNA in human Jurkat T cells were 1.26 ± 0.15, 1.75 ± 0.20, 2.26 ± 0.23 and 3.74 ± 0.62 in 6, 12, 24 and 48 h after EV-A71 infection, which the differences were significant with mock-infected group(t values were -2.96, -6.38, -9.57, -7.71; P<0.05). Western blot showed that the protein expression levels of MyD88 had increased 1.34 times and 2.17 times in 24 h and 48 h after EV-A71 infection compared with mock-infected group. After infected for 24 h and 48 h, the levels of IL-6 were (302.86 ± 38.11), (179.70 ± 14.50) pg/ml, which were significantly higher than mock-infected group (176.42 ± 9.60), (179.70 ± 14.50) pg/ml (t values were -5.57, -18.54, P<0.05). The levels of TNF-α in EV-A71 infected group (100.81 ± 9.81) pg/ml was higher than that in mock-infected group (56.19 ± 6.94) pg/ml, and the difference was significant (t=-6.43, P=0.003).

CONCLUSIONTLR7 is the main pattern recognition receptor responsible for EV-A71 recognition in immune cells, which then leads to the activation of TLR7 downstream signaling and the production of proinflammatory cytokines.

Blotting, Western ; Cell Line ; Enterovirus A, Human ; pathogenicity ; Enzyme-Linked Immunosorbent Assay ; Humans ; Interleukin-6 ; metabolism ; Interleukin-8 ; metabolism ; Myeloid Differentiation Factor 88 ; metabolism ; Real-Time Polymerase Chain Reaction ; T-Lymphocytes ; immunology ; virology ; Toll-Like Receptor 7 ; immunology ; Tumor Necrosis Factor-alpha ; metabolism

FMDV 2A peptide was introduced as a linker between GP5 and M protein of porcine reproduction and respiratory syndrome virus (PRRSV) to allow automatic self-cleavage the polyproteins. This strategy simultaneously displayed the neutralizing action of GP5 protein and cell-mediated immunity of M protein. We put them into the expression cassette of adenovirus vector. The results of RT-PCR, IFA and Western blotting showed that GP5 and M protein were not only expressed correctly, but also self-cleavaged and assemble heterodimers formation. To detect the advantages of rAd-GP5-2A-M, we also constructed some other recombinant adenoviruses (rAd-GP5, rAd-M and rAd-GP5-M) as control. After inoculated subcutaneously into BALB/c mice, the four recombinant adenoviruses can induce PRRSV-specific antibodies and cell-mediated immune response, but the level of humoral and cell-mediated immune response against PRRSV induced by rAd-GP5-2A-M is the strongest among the four recombinant adenoviruses. All of these suggested that it is possible to develop one multi-gene engineering vaccine utilizing FMDV 2A peptide, and also provided a novel strategy for developing other viral disease vaccine.
Adenoviridae ; genetics ; metabolism ; Animals ; Female ; Immunization ; Mice ; Mice, Inbred BALB C ; Recombinant Fusion Proteins ; genetics ; immunology ; metabolism ; Swine ; Vaccines, Synthetic ; immunology ; Viral Envelope Proteins ; genetics ; immunology ; metabolism ; Viral Matrix Proteins ; genetics ; immunology ; metabolism ; Viral Vaccines ; immunology