Objective:To explore the immunogenecity of four Dengue virus-specific peptides in mice.Methods:Each peptide of four Dengue virus-specific peptides (C45-57KLVMAFIAFLRFL,E396-408SSIGKMFEATARG,NS323-35YRILQRGLLGRSQ,and NS3141-155NREGKIVGLYGNGVV) was used to immunize BALB/c mice and C57BL/6 mice,respectively.Three weeks later,mice were killed and splelocytes were prepared.Splelocytes were stimulated by the same peptide and intracellular cytokine staining (ICS) assay was used to detect the percentages of peptide-specific IFN-? or IL-4 producing CD4+ T cells in total CD4+ T cells.Results:Peptide C45-57 induced peptide-specific IFN-?+ CD4+ T cells(0.72%?0.04% vs 0.04%?0.02%,P

Aim To provide practical method for screening human carbonic anhydrase Ⅱ(hCA Ⅱ) inhibitors in drug discovery.Methods hCA Ⅱ protein was obtained from induced BL21(DE3) E.coli containing plasmid pET-28b-hCA Ⅱ.The hCA Ⅱ activity was detected under pH 7.6 and 25℃ by its esterase activity which could decompose PNPA to increase the photoabsorption at 348 nm. After the assay conditions were finally selected, 35 new compounds were tested.Results A practical method for screening hCA Ⅱ inhibitors was successfully constituted by using recombinant hCA Ⅱ protein expressed in E.coli as the source of hCA Ⅱ enzyme.10 new compounds had better inhibitory effect and 9 new compounds had the same inhibitory effect on hCA Ⅱ compared with acetazolamide.Conclusions The hCA II inhibitor screening technique constituted in this work possesses advantages of being reliable, rapid, and practical. 19 new compounds are worth further research for developing high efficiency and low side effect drugs used for high-altitude illness.

Objective To study protein variation between PtenL/LMEFs and Pten△/△MEFs cells.Methods Two-dimensional electrophoresis(2-DE) was employed to compare the differential expression proteins between PtenL/LMEFs and Pten△/△MEFs cells.Six differential expression proteins were digested in gel by enzyme and the mass of generated peptides was measured by matrix assisted laser desorption ionization time of flight mass spectrometry(MALDI-TOF-MS).The data obtained from peptide mass fingerprinting(PMF) were searched using the Internet available database and five proteins were identified.Results Compared with that of PtenL/LMEFs,expression level of proteins including phosphoglycerate mutase 1(PGAM1) and peptidyl-prolyl cis-trans isomerase C(PPIC) was up-regulated,whereas expression level of Transgelin 2 was down-regulated in Pten△/△MEFs cells.B and F proteins were both identified to be peroxiredoxin-6.They had similar molecular weight but different PI which might be caused by post-translation modification.B protein was only expressed in Pten△/△MEFs cells.Conclusion The protein profile of Pten△/△MEFs cells displayed obvious difference compared to that of PtenL/LMEFs cells.The results implied that various distinct different proteins might lead to cancer.

Objective To identify hepatitis C virus(HCV)-specific cytotoxicity T lymphocytes (CTL)epitopes by the combination of T epitopes prediction software and in vitro assays.Methods HCVspecific CTL epitopes were predicted by T epitope prediction software Rankpep and then candidate HCV-specific CTL epitopes were selected.Candidate HCV-specific CTL epitopes were used to stimulate PBMC of HCV-infected patients and healthy volunteers.and then enzyme-linked immunospot(ELISPOT)and intracellular cytokine staining(ICS)were used to measure the frequencies of IFN-γ-producing cells in total PBMC and the percentages of IFN-γ+CD8+T cells in total CD8+T cells,respectively.Results Five candidate CTL epitopes[NS3 450(TVPQDAVSR),NS3 594(GPTLLYRL),Ns4b 78(sMMAFSAAL),NS5a 416(SEENVSVVF)and NS5a 367(TVSSALAEL)]were used to stimulate PBMC of ten HCV-infected patients and two healthy volunteers.PBMC of seven HCV-infected patients secreted IFN-γ while PBMC of healthy volunteers did not produce IFN-γ.The frequencies of peptide-specific IFN-γ-producing cells ranged from 5 to 36 SFC/105 PBMC and the percentages of peptide-specific IFN-γ+CD8+T cells ranged from 0.02%-0.25%.Conclusion Resuhs of ELISPOT assay and ICS assay confirm that these five peptides NS3 450,NS3 594,NS4b 78,NS5a 416 and NS5a 367 are identified as Hovel HCV-specific CTL epitopes.

Objective To develop a superparamagnetic iron oxide nanoparticles ( SPIO ) based on MRI probe specifically targeting human epidermal growth factor receptor 2 (HER2) and explore its value as MRI positive contrast agents in vitro.Methods (1) The superparamagnetic iron oxide ( PS) was obtained by means of classical coprecipitation in polylactic acid solution , then coupled with fluorescein isothiocyanate (FITC) labeled LTVSPYW to develop the targeted probe ( FITC-LTVSPWY-PS).The particle size was measured under transmission electron microscope.Relaxation rate was detected by 3.0 T MR scanner.(2) Climbing films of human breast cancer cell MCF-7 were prepared and incubated with FITC-LTVSPWY-SPIO, then fluorescence distribution was observed under inverted microscope.And distribution of iron particles was confirmed by prussian blue staining.(3) MCF-7 and MDA-MB-231 cells were incubated with FITC-LTVSPWY-SPIO and PS, respectively.MCF-7 incubated with FITC-LTVSPWY-PS were used as experimental group, MCF-7 treated with PS as control group , and cells added with nothing as blank group.There were 3 samples in each group.The MR imaging was performed only once and T 2 WI signal intensity of cells was recorded.The comparison of T 2 signal intensity among groups was conducted by using one-way ANOVA.Results The core and surface size of nanoparticles were (13.9 ±1.6) nm and (122.0 ±5.5) nm respectively.Zeta potential and relaxation rate of the FITC-LTVSPWY-PS were ( -30.7 ±2.2 ) mV and 70.7 m· M-1 · s-1 respectively, and the PS were (28.1 ±2.8) mV and 72.1 m· M-1 · s-1 respectively.The fluorescence could be seen on the surface of MCF-7 cells, and the prussian blue staining showed that FITC-LTVSPWY-PS could specifically target HER 2-positive cells.The low signal on T 2 WI was observed in MCF-7 cells incubated with FITC-LTVSPWY-PS, whereas cells treated with PS and blank group showed equal signals , the T2 values were ( 61.8 ±5.7 ) , ( 101.6 ±2.5 ) and ( 103.5 ±1.9 ) ms respectively.Significant difference existed among these groups ( F =355.698, P <0.05 ).Conclusions The MR targeting probe FITC-LTVSPWY-PS was prepared successfully , its physical characterization and magnetic properties could target the HER 2 highly expressing on the surface of breast cancer cells and meet the need of targeted imaging.It provides an important tool for MR molecular imaging.

Objective:To investigate the short-term effects of articular injection of hyaluronic acid combined with glucocorticoid in patients with knee osteoarthritis.Methods:From October 2017 to June 2018, a total of 188 patients diagnosed with knee osteoarthritis received parallel articular injection. There were 60 cases with mild knee osteoarthritis, 72 with moderate and 56 with severe according to the WOMAC knee functional score. There patients were divided into group rank Ⅰ48 cases, Ⅱ 49 cases, Ⅲ 45 cases, Ⅳ 46 cases according to the knee joint X-ray Kellgren-Lawrence classification. The unified treatment regimen was 2.5 ml Sodium Hyaluronate (SHA) injection for the first time, SHA 2.5 ml and compound betamethasone injection (CBI) 1 ml for the second week, and 2.5 ml of SHA for the third week. WOMAC score and Lequesne index were used to evaluate joint function before the first injection and after SHA and SHA+CBI injection. The improvement rate of Lequesne index ≥30% or improvement rate of WOMAC score ≥25% was regarded as effective treatment.Results:Lequesne index and WOMAC score decreased gradually in the mild, moderate and severe groups after 3 weeks of injection. Among these patients, the improvement rates of Lequesne index after SHA injection and SHA+CBI injection were 36.44%±8.46% and 49.26%±13.75% in the mild group, 23.09%±12.61% and 30.66%±14.95% in the moderate group, and 10.50%±8.78% and 11.07%±6.52% in the severe group. The improvement rate of WOMAC score in the mild group after SHA injection and after SHA+CBI injection was greater than 25%. After SHA injection, the improvement rate of WOMAC score was 13.06%±10.21% in the moderate group, and 27.49%±13.61% after SHA+CBI injection. Those in severe group were all less than 25%. Kendall's staub correlation analysis results showed that there was a strong positive correlation between WOMAC function score and X-ray Kellgren-Lawrence classification ( r=0.744, P<0.001). The Lequesne index and WOMAC scores of the Kellgren-Lawrence X-ray classification decreased gradually after 3 weeks of injection. The improvement rate of Lequesne index period in group rank Ⅰ after SHA and SHA+CBI injection was 36.64%±10.05% and 52.00%±8.19%, respectively. That for group rank Ⅱ was 32.05%±8.09% and 41.95%±10.53%, group rank Ⅲ 16.93%±10.34% and 27.77%±10.25%, group rank Ⅳ 7.52%±5.53% and 7.60%±6.66%. The improvement rate of WOMAC score period in group rank Ⅰ after SHA and SHA+CBI injection was 29.48%±11.77% and 42.59%±13.55%, respectively. That for group rank Ⅱ was 26.72%±10.21% and 30.49%±16.90%, group rank Ⅲ 13.78%±5.96% and 23.05%±9.52%, group rank Ⅳ 4.77%±3.80% and 4.27%±4.23%. Conclusion:For mild or X-ray classification Ⅰ, Ⅱ knee osteoarthritis patients, articular injection SHA or SHA+CBI are effective. Further, SHA+CBI is better than single injection of SHA. SHA+CBI injection was effective for moderate knee osteoarthritis patients. For severe or X-ray classification Ⅲ, Ⅳ patients, SHA or SHA+CBI injection at interval are invalid.

Objective:To observe the results of electroacupuncture (EA) on the resuscitation of a rat model of asphyxia cardiac arrest (CA). And to explore its effect on the neurologic deficits and hemodynamic instability of post-cardiac arrest syndrome (PCAS).Methods:A total of 107 male SD rats were randomly divided into sham, CA, and EA groups. Each group received arterial catheterization and tracheal intubation. The sham group was not induced asphyxia. Asphyxial cardiac arrest was established by endotracheal tube clamping. Rats in the CA group received basic respiratory support and fluid resuscitation in return of spontaneous circulation (ROSC) and rats in the EA group received EA at Baihui based on the treatment of CA group after ROSC, with a dense-dispersed wave at frequencies of 4-20 Hz, while the current intensity was adjusted minimum to induce a twitch of the scalp, the course of treatment was 30 minutes. The baseline data, hemodynamics after ROSC, neurological deficit score (NDS), pathological changes of brain tissue, and levels of serum biomarker were recorded and compared among the three groups. The 72-hour survival of rats was analyzed by Kaplan-Meier survival curve. Hematoxylin-eosin (HE) staining was used to observe the pathological changes of necrotic neurons in the hippocampal CA1 region of rat brain. Meanwhile, Nissl staining and TdT-mediated dUTP nick-end labeling (TUNEL) were used to detect cell apoptosis and injury.Results:Compared with the CA group, the mean arterial pressure (MAP) in the EA group increased significantly at 15 minutes after ROSC [mmHg (1 mmHg≈0.133 kPa): 125.00 (94.00, 136.25) vs. 92.00 (72.00, 122.50), P < 0.05]. There was no significant difference in the NDS score between the EA group and the sham group. Still, the NDS score of the rats in the CA group at 6 hours after ROSC were significantly lower than that in the sham group (46.00±10.61 vs. 80.00±0.00, P < 0.05). Kaplan-Meier survival curve analysis showed that EA did not improve the 72-hour survival rate of rats (100% in the sham group, 25% in the CA group, and 30% in the EA group, P > 0.05). The analysis by TUNEL showed that the apoptosis rate of neurons in CA1 region of the hippocampus in EA group at 6 hours after ROSC was significantly lower than that in CA group [(62.84±2.67)% vs. (71.29±3.70)%, P < 0.05]. Compared with the CA group, the level of serum S100 calcium binding protein B (S100B) in the EA group at 6 hours after ROSC was significantly lower (ng/L: 19.30±13.87 vs. 132.28±31.67, P < 0.05), but there were no significant differences in the levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) between these two groups. Conclusions:In the present study, EA at Baihui can stabilize the hemodynamic, moreover, it has a particular neuroprotective effect on PCAS rats. Still, EA at Baihui does not reduce the systemic inflammatory response and improve the survival rate of rats, and its mechanism remains to be verified in further research.

OBJECTIVE: To compare the effects of ~(56)Fe~(17+),~(12)C~(6+)ion beams and~(60)Co γ rays on chromosome aberration in human lymphoblastoid cells. METHODS: The human lymphoblastoid cells were divided into 0. 1,0. 3,0. 5,0. 7,1. 0,2. 0 Gy irradiated groups and 0. 0 Gy control group. They were separately exposed to ~(56)Fe~(17+)ion beams( linear energy transfer was 400. 0 ke V/μm),~(12)C~(6+)ion beams( linear energy transfer was 26. 0 ke V/μm) and~(60)C γ rays. Chromosome specimens were harvested 48 hours after irradiation. The effects of different radiation on dicentric plus centric ring( “d + r”) aberration rate and chromosome aberration in human lymphoblastoid cells were detected by light microscope with artificial counting. RESULTS: The “d + r”aberration rates induced by 0. 3-2. 0 Gy ~(12)C~(6+)ion beams were significantly higher than those of ~(56)Fe~(17+)ion beams and~(60)Co γ rays at the same dose( P < 0. 017). Chromosome aberration cell rates of 0. 1-2. 0 Gy ~(12)C~(6+)ion beams were significantly higher than those of ~(56)Fe~(17+)ion beams and~(60)C γ rays at the same dose( P < 0. 017). At the dose range of 0. 0-2. 0 Gy,chromosome aberration effects of three kinds of radiations were gradually increased( P < 0. 01). The relative biological effectiveness of ~(56)Fe~(17+)ion beams was lower than that of ~(12)C~(6+)ion beams.CONCLUSION: The chromosome aberration induced by ~(12)C~(6+)ion beams was more serious than that of~(60)Co γ rays and ~(56)Fe~(17+)ion beams.