Plague was classified as a natural focus disease,occurred in remote areas of tratfic block in the past.But with the development of society and economy,plague of long distance transmission risk is growing in the source regions and densely populated areas.In order to effectively prevent and control the spread of plague outbreaks are occurring,and long distance transmission spread in Qinghai Province,and adhere to the implementation of the long-distance transmission of plague prevention and control strategies,avoid threats and reduced long distance spread of plague.

Objective To observe the effect of p53 protein on smooth muscle cell(VSMC)in rabbit artery balloon injury.Methods Restenosis model of carotid artery after balloon injury was established in rabbits.30 rabbits were divided into 2 groups,the sham group(n = 6)and the vascular injury group(n = 24).With H.E.staining and automatic image analysis system,we investigated artery morphology alteration and measured the area of arterial intima and media.The expressions of p53 protein were detected by immunohistochemical analysis.Results With H.E.staining and automatic image analysis and immunohistochemistry,the results showed that the expression of p53 was significantly reduced and the intima area was increased in model group compared with the sham group(P ＜0.01).But the expression of p53 in media was remarkably reduced compared with intima(P ＜ 0.01).Conclusions The possible mechanism of preventing arterial restenosis in the balloon injury might be related with p53,which may be through inhibiting neointimal proliferation in arterial restenosis .

To obtain active protein of pIL-18 expression in Lactococcus lactis, and to observe its biological activity, the total RNA was extracted as template from peripheral blood mononuclear cells. Porcine interleukin 18 (pIL-18) was amplified by RT-PCR. The resulting fragment was cloned into pAMJ399 L. lactis vector, and then transformed to L. lactis MG1363 cells by electroporation. Expression of pIL-18 protein was detected by SDS-PAGE and Western-blotting. Bioactivity of the product was tested by pig spleen lymphocyte proliferation test and cytopathogenic effect inhibition assay. The result of Western blotting and bioactivity test shows that the molecular weight of pIL-18 protein was 19 kDa. The react line was observed in both supernatant and precipitated of the recombinant bacteria pAMJ399-pIL18/MG1363. The expressed pIL-18 can promote the proliferation of pig spleen lymphocyte, and significantly inhibit virus multiplication. As conclusion, porcine interleukin-18 was successfully expressed in L. lactis, and the product was biologically active.
Animals ; Blotting, Western ; Electrophoresis, Polyacrylamide Gel ; Electroporation ; Genetic Vectors ; Interleukin-18 ; biosynthesis ; Lactococcus lactis ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Swine

The coding sequence for the mature peptide of porcine lactoferrin (Plf) was synthesized according to the codon usage of lactobacillus, to establish optimized porcine lactoferrin Lactobacillus expression system. The gene was ligated into the Xho I/BamH I site of Lactobacillus expression vector pPG612.1 and the recombinant plasmid pPG612.1-plf was transformed individually into Lactobacillus casei ATCC393, Lactobacillus pentosus KLDS1.0413, Lactobacillus plantarum KLDS1.0344 or Lactobacillus paracasei KLDS1.0652 by electroporation. After induction with xylose, expression of the recombinant proteins was detected by Western blotting and confocal laser scanning microscopy. Secretion of recombinant Plf proteins from four recombinant species was determined quantitatively by ELISA. The antibacterial activities of recombinant proteins were measured by agar diffusion method. The result shows that Plf was correctly expressed in four species of recombinant lactobacillus, with molecular weight of about 73 kDa. The expression levels in recombinant Lactobacillus casei, Lactobacillus pentosus, Lactobacillus plantarum, Lactobacillus paracasei were 9.6 μg/mL, 10.8 μg/mL, 12.5 μg/mL and 9.9 μg/mL, respectively. Antimicrobial activity experiment shows that the recombinant proteins were active against E. coli, Staphylococcus aureus, Salmonella typhimurium, Listeria, Pasteurella. The recombinant Plf expressed by recombinant Lactobacillus plantarum showed the best antibacterial activity among all recombinant lactobacillus species. These data represent a basis for the development and application of porcine lactoferrin from recombinant lactobacillus.
Animals ; Anti-Bacterial Agents ; biosynthesis ; Lactobacillus ; metabolism ; Lactoferrin ; biosynthesis ; Recombinant Proteins ; biosynthesis ; Swine

0.05). Conclusion pcDNA3.1/SjTs-1 induced the mucosal and systemic immune response and partial protection against the challenge of S.japonicum by the intranasal vaccinations of mice.

To evaluate the immune responses of recombinant Lactobacillus casei 393 expressing Porcine Epidemic Diarrhea Viral (PEDV) N protein as oral vaccine, n gene of PEDV was subcloned into the expression vector pPG-1, and then transformed into L. casei 393 by electroporation, resulting in recombinant strain pPG-1-n/L, casei 393. The recombinant strains were induced to express interest protein, which was detected by Western blotting, immunofluorescence microscopy and the whole bacteria ELISA. And then BALB/C mice were used as an animal model immunized with recombinant strains by oral administration, and the immune efficacy was analyzed. The recombinant PEDV N protein showed the antigenic specificity, and was located on the bacterial cell walls of pPG-1-n transformed L. casei. The results of ELISA showed that the mice immunized with recombinant strains could produce remarkable special sIgA level in the feces, and high level of anti-PEDV N protein IgG in the serum (P < 0.01), but the induced antibodies in serum did not demonstrated neutralizing effect. Statistical significant difference was observed among the spleen lymphocyte proliferation index (LPI) among the immunization groups of mice and control groups. And there was significant increase. of IFN-gamma and IL-4 contents in the supernatant of spleen cell culture in immunized group. In conclusion, the oral immunizations with recombinant L. casei 393 can induce significant specific mucosal PEDV N-specific IgA response as well as serum IgG responses, and can evoke both mucosal immune and system immune responses.
Administration, Oral ; Animals ; Antibody Formation ; Coronavirus Infections ; prevention & control ; Female ; Immunity, Mucosal ; immunology ; Lactobacillus casei ; genetics ; metabolism ; Mice ; Mice, Inbred BALB C ; Nucleocapsid Proteins ; biosynthesis ; genetics ; immunology ; Porcine epidemic diarrhea virus ; immunology ; Recombinant Proteins ; biosynthesis ; genetics ; immunology ; Swine ; Viral Vaccines ; administration & dosage ; immunology

Objective To investigate the role of cell adhesion molecule L1 like (CALL) in the genesis and development of esophageal squamous cell carcinoma (ESCC).Methods From July 2007 to December 2010,a total of 100 patients with ESCC who received radical resection of esophageal cancer were enrolled.The ESCC tissues and corresponding tumor-adjacent normal tissues were obtained.The expression of CALl was determined by tissue microarray technology and immunohistochemical staining.The CALL over-expressed esophageal cancer cell line was established.The effects of CALL on cell migration and invasion were detected by wound-healing assay and Transwell assay,respectively.The effects of CALL on actin microfilament was analyzed by filamentous actin (F-actin) staining.Chi square test,Fisher's exact test,multivariate analysis and t test were performed for statistical analysis.Results The positive expression rate of CALL in ESCC tissues was 56 ％ (56/100),which was lower than that of tumor-adjacent normal tissues (95％,95/100),and the difference was statistically significant (x2=41.114,P＜0.01).There were statistically significant differences in CALL expression at protein level among patients with ESCC of different differentiation degree,different pathological T stage,lymph node metastasis and different TNM stage (x2=13.702,5.317,21.453,Fisher's exact test;all P＜ 0.05).The five year disease related survival rate of ESCC patients with down-regulated expression of CALL was 0(0/49),which was lower than those with normal CALL expression (25.5％,13/51),and the difference was statistically significant (x2 =43.338,P＜0.01).The median survival time of CALL expression down-regulated group was 17 months,and that of normal expressed group was 38 months.CALL expression was an independent risk factor of disease special survival rate (hazard ratio (HR) 0.353,95％ confidence interval (CI) 0.188 to 0.666,P=0.001).The results of wound-healing assay showed that the migration ability of CALL overexpressed CALL-k30 cells was lower than that of Vec-k30 cells in control group on 24 hours after wound.The results of Transwell invasion test showed the number of migrating cells penetrating CALL k30 cells attached to the inferior surface of the membrane was 44.000±13.748,which was less than that of the Vec k30 cells (154.333±25.007),and the difference was statistically significant (t=5.136,P=0.036).The results of F-actin staining demonstrated that actin filaments of CALL-k30 cells was 234.667 ± 65.118,which was lower than that of Vec-k30 cells (597.000± 119.929),and the difference was statistically significant (t=4.707,P=0.042).Conclusions CALL lowers the migration and invasion abilities of esophageal cancer cells by inhibiting F-actin microfilaments.Its abnormal expression may play an important role in the genesis,development and prognosis of ESCC.

Objective:To explore the role and molecular mechanism of hepatocyte nuclear factor 4γ (HNF4γ) in proliferation and stemness of gastric cancer.Methods:A total of 102 cases of paraffin-embedded gastric cancer tissues and matched adjacent gastric tissues and 42 cases of fresh-frozen tissues derived from gastric patients who received radical gastrectomy were collected from the First Affiliated Hospital of Zhengzhou University between 2012 to 2015. The expression of HNF4γ was tested by immunohistochemical staining, quantitative real-time polymerase chain reaction (qRT-PCR). HNF4γ overexpressed (AGS-HNF4γ) and shRNA silenced (HGC27-shHNF4γ) gastric cell lines were established. The effects of HNF4γ on cell proliferation and stemness were verified by XTT, clone formation and sphere formation assay. The expression of CD44 was detected by western blot.Results:The mRNA expression level of HNF4γ in fresh-frozen gastric cancer tissue was (12.43±2.702), which was significantly higher than (3.639±1.109) in normal tissue ( P<0.001). The high protein expression rate of HNF4γ in paraffin-embedded gastric cancer tissues was 41.2% (42/102), which was significantly higher than 8.8% (9/102) in normal gastric mucosa tissue ( P< 0.001). The protein expression of HNF4γ was closely related to the tumor differentiation, infiltration depth, lymph node metastasis and tumor stage ( P<0.05). The median survival interval of patients with HNF4γ high expression was 25 months, the 3-year survival rate was 4.8% (2/42), significantly lower than 38 months and 51.7% (31/60) of patients with normal HNF4γ expression ( P<0.001). The proliferation and CD44 protein expression of AGS-HNF4γ cells were significantly higher than those of the AGS-Vector cells. The number of clone formation, sphere formation rate of AGS-HNF4γ cells were 243.5±24.5 and (83.5±3.9)%, significantly higher than 81.0±16.0 and (21.8±5.6)% of AGS-Vector cells ( P=0.030 and P=0.010, respectively). The proliferation and CD44 protein expression of HGC27-shHNF4 cells were significantly lower than those of the HGC27-vector cells. The number of clone formation, sphere formation rate of HGC27-shHNF4 cells were 26.0±1.0 and (20.8±8.4)%, significantly higher than 83.5±4.5 and (72.5±4.8)% of HGC27-vector cells ( P=0.006 and P=0.030, respectively). Conclusions:HNF4γ is upregulated in the gastric cancer tissues and related with the poor prognosis of patients with gastric cancer. Overexpression of HNF4γ promotes the proliferation and remains the stemness of gastric cancer cells by upregulating the expression of CD44.

Objective:To explore the role and molecular mechanism of hepatocyte nuclear factor 4γ (HNF4γ) in proliferation and stemness of gastric cancer.Methods:A total of 102 cases of paraffin-embedded gastric cancer tissues and matched adjacent gastric tissues and 42 cases of fresh-frozen tissues derived from gastric patients who received radical gastrectomy were collected from the First Affiliated Hospital of Zhengzhou University between 2012 to 2015. The expression of HNF4γ was tested by immunohistochemical staining, quantitative real-time polymerase chain reaction (qRT-PCR). HNF4γ overexpressed (AGS-HNF4γ) and shRNA silenced (HGC27-shHNF4γ) gastric cell lines were established. The effects of HNF4γ on cell proliferation and stemness were verified by XTT, clone formation and sphere formation assay. The expression of CD44 was detected by western blot.Results:The mRNA expression level of HNF4γ in fresh-frozen gastric cancer tissue was (12.43±2.702), which was significantly higher than (3.639±1.109) in normal tissue ( P<0.001). The high protein expression rate of HNF4γ in paraffin-embedded gastric cancer tissues was 41.2% (42/102), which was significantly higher than 8.8% (9/102) in normal gastric mucosa tissue ( P< 0.001). The protein expression of HNF4γ was closely related to the tumor differentiation, infiltration depth, lymph node metastasis and tumor stage ( P<0.05). The median survival interval of patients with HNF4γ high expression was 25 months, the 3-year survival rate was 4.8% (2/42), significantly lower than 38 months and 51.7% (31/60) of patients with normal HNF4γ expression ( P<0.001). The proliferation and CD44 protein expression of AGS-HNF4γ cells were significantly higher than those of the AGS-Vector cells. The number of clone formation, sphere formation rate of AGS-HNF4γ cells were 243.5±24.5 and (83.5±3.9)%, significantly higher than 81.0±16.0 and (21.8±5.6)% of AGS-Vector cells ( P=0.030 and P=0.010, respectively). The proliferation and CD44 protein expression of HGC27-shHNF4 cells were significantly lower than those of the HGC27-vector cells. The number of clone formation, sphere formation rate of HGC27-shHNF4 cells were 26.0±1.0 and (20.8±8.4)%, significantly higher than 83.5±4.5 and (72.5±4.8)% of HGC27-vector cells ( P=0.006 and P=0.030, respectively). Conclusions:HNF4γ is upregulated in the gastric cancer tissues and related with the poor prognosis of patients with gastric cancer. Overexpression of HNF4γ promotes the proliferation and remains the stemness of gastric cancer cells by upregulating the expression of CD44.

Application of the PCR-derived technology in gene identification and genotypes of different ecotype Yersinia pestis to make the high-throughput experimental results can reflect the epidemic history and compare the diversity in genome, pathogenicity, so that results from these experiments provide an important basis for clinical diagnosis, treatment and origin. But the experiment should be considered typing ability, practicality, budget and other experimental factors or conditions, because each PCR-derivative technology has advantages and disadvantages.
Genotype ; Humans ; Polymerase Chain Reaction ; Virulence ; Yersinia pestis