Preparation and characterization of a recombinant poly-epitopic vaccine EgG1Y162-2 (4) against cystic echinococcosis based on the linker GSGGSG / 中国血吸虫病防治杂志

Objective To perform prokaryotic expression and preliminary characterization of the recombinant poly-epitope vaccine EgG1Y162-2 (4) against cystic echinococcosis. Methods The recombinant poly-epitope vaccine EgG1Y162-2 (4) against Echinococcus granulosus based on the linker GSGGSG was subjected to structural three-dimensional (3D) modeling using immunoinformatics to analyze the structural changes and evaluate the antigenicity of the vaccine. The pET30a-EgG1Y162-2 (4) recombinant plasmid was generated using double digestion with EcoR I and Sal I, and then transformed into competent cells. Following protein induction with isopropyl-β-D-thiogalactoside (IPTG), the prokaryotic expression proteins were characterized using Western blotting, and the antigenicity of the recombinant protein was analyzed using sera from cystic echinococcosis patients and health volunteers. Results The four EgG1Y162-2 proteins coupled by the 3D structure of the recombinant vaccine EgG1Y162-2 (4) presented independent and effective expression and good antigenicity. The highest protein expression was detected in the supernatant following induction of the recombinant plasmid pET30a-EgG1Y162-2 (4) by 0.2 mmol/L IPTG at 37 °C for 4 h, and a pure protein component was seen following elution with 60 mmol/L imidazole. Western blotting analysis of the recombinant multiepitope protein HIS-EgG1Y162-2 (4) showed a band at approximately 39 kDa, and this band was recognized by sera from cystic echinococcosis patients. Conclusion A recombinant poly-epitope vaccine EgG1Y162-2 (4) against cystic echinococcosis has been successfully constructed, which provides a preliminary basis for researches on recombinant multi-epitope vaccine against cystic echinococcosis.

Effect of adenosine on autophagy and proliferation of hepatocellular carcinoma cells / 中国药理学通报

Aim To investigate the effect of adenosine on the autophagy and proliferation of hepatocellular carcinoma cells, and improve the curative effect of a-denosine on hepatocellular carcinoma. Methods HepG2 cells were incubated with adenosine, CCK-8 method was used to study the changes of cell prolifera-tion,Western blot was used to study the expression of LC3-Ⅱ and LC3-Ⅰ, and MDC staining was used to observe the number of autophagosomes. Results HepG2 cells were incubated with adenosine(1.0~4.0 mmol·L-1) for 48 h,the proliferation of HepG2 cells were detected at the different time points (12,24,48 h),and the result showed the proliferation was signifi-cantly inhibited by adenosine (P < 0.01). HepG2 cells were incubated with adenosine (0.2,0.5,1.0, 2.0,4.0 mmol·L-1) for 24 h,the ratio of LC3-Ⅱ/LC3-Ⅰ decreased significantly in low concentration of adenosine group (0.2, 0.5 mmol·L-1, P <0.05;1.0 mmol·L-1,P<0.01),and the ratio of LC3-Ⅱ/LC3-Ⅰ increased significantly in higher concentration of adenosine group (4.0 mmol·L-1, P <0.05). HepG2 cells were incubated with adenosine(1.0 mmol·L-1) for 24 h, the ratio of LC3-Ⅱ/LC3-Ⅰ de-creased significantly at 6,12 and 24 h detecting point, the number of autophagosomes were reduced, the low-est ratio of LC3-Ⅱ/LC3-Ⅰ and autophagosomes were observed at 12 h detecting point(P<0.01). Conclu-sions Adenosine inhibits the proliferation of hepato-cellular carcinoma cells,the low concentration of aden-osine inhibits the autophagy,while the high concentra-tion of adenosine increases the autophagy, which is of great significance to reduce multi-drug resistance and improve the therapeutic effect of anti-hepatoma drugs.