[Component analysis of excretory/secretory protein from Trichinella spiralis adult worm].

Objective: To analyze the component of adult worm excretory/secretory protein(AWESP) from Trichinella spiralis using the shotgun method, and find out the active component underlying its regulatory effect on colitis in humans. Methods: The T. spiralis AWESP was prepared, separated by SDS-PAGE, lysed with trypsin, and analyzed by shotgun LC-MS/MS. The protein components were determined with the Masco software and classified using the Gene Ontology(GO) method in cellular components, molecular functions, and biological processes. Results: The AWESPs isolated by SDS-PAGE had a Mr of 15 000-116 000. A total of 280 proteins were revealed by LC-MS/MS, of which 96 were identified by Masco software, 98 were putative, and the remaining 86 were unclear. Preliminary results showed that 4 proteins had regulatory potential for colitis, including cysteine protease inhibitor, serine protease, 53 000 excretory/secretory antigen, and glutathione-S-transferase. GO enrichment analysis showed that the identified proteins had 104 different molecular functions, involved in 363 biological processes. Conclusion: As revealed by the Masco software, T. spiralis AWESP has complex components and 96 have been identified in this study. Four of them are preliminarily shown to be associated with the anti-colitis effect of T. spiralis.

[Construction and analysis of a breast cancer gene-drug network model].

OBJECTIVE: To construct a breast cancer gene-drug network model for extracting and predicting the correlations between breast cancer-related genes and drugs. METHODS: We developed an algorithm based on the ABC principle and the association rules to obtain the correlations between the biological entities. For breast cancer, we constructed 3 different correlations (gene-gene, drug-drug and gene-drug) and used the R language to implement the associated network model. The reliability of the algorithm was verified by ROC curve. RESULTS: We identified 185 breast cancer-associated genes and 98 associations between them, 97 drugs and 170 associations between them. The breast cancer genes-drugs network contained 127 genes and 77 drugs with 384 associations between them. CONCLUSIONS: We identified a large number of different correlations between the breast cancer-related genes and drugs and close correlations between some biological entity pairs that have not yet been reported, which may provide a new strategy for experimental design for testing personalized breast cancer treatment.

[In vitro killing of Trichinella spiralis muscle larvae by exogenous nitric oxide from SNP].

OBJECTIVE: To study the lethal effect of exogenous nitric oxide donor sodium nitroprusside (SNP) on the muscle larvae of Trichinella spiralis in vitro cultivation. METHODS: T. spiralis muscle larvae isolated from the infected BALB/c mice were formulated into a 1,000 larva/ml suspension with RPMI 1640 medium, and 0.1 ml suspension per orifice was cultured with SNP at 37°C in a humidified 5% CO2 atmosphere. The final concentrations of SNP were 0.02, 0.05, 0.10, 0.20, 0.50 and 1.00 mmol/L, respectively, and then the experiments were divided into 5 groups:1.00 mmol/L SNP (control group, Group A), 0.15 mmol/L FeSO4+ 1.00 mmol/L SNP (Group B), 1.00 mmol/L L-cysteine + 1.00 mmol/L SNP (Group C), 0.15 mmol/L FeSO4+ 1.00 mmol/L L-cysteine + 1.00 mmol/L SNP (Group D) and 0.15 mmol/L Hemoglobin + 1.00 mmol/L SNP (Group E). All the groups were incubated with T. spiralis muscle larvae in RPMI 1640 medium. The survivability of the muscle larvae was observed by steromicroscope and the differences of inhibition ratio among these groups were analyzed 4 d after the incubation. Results SNP 0.02 mmol/L was not cytotoxic to the muscle larvae with an inhibition of (5.50 ± 1.80) %. The mortality rates of SNP 0.05, 0.10, 0.20, 0.50, 1.00 mmol/L groups were (20.19±2.71)%, (29.21±2.12)%, (41.81±2.03)%, (47.85±3.79)%, (60.98±5.19)%, respectively, significantly higher than that of the control group[(4.93±0.25) %, all P < 0.051]. There was a positive liner correlation between the mortality of muscle larvae and SNP concentrations in the range of 0.02-1.00 mmol/L. Next, Group A, B, C, D and E led to the mortalities from (60.98±5.19)% to (49.48±1.34)%, (47.29±2.79)%, (26.28±1.37)%, (17.93±3.49)%, respectively, and all the differences between Group A and the other four groups were statistically significant (all P < 0.05). CONCLUSIONS: Exogenous nitric oxide released from SNP can kill the muscle larvae of T. spiralis. However, hemoglobin, L-cysteine, and FeSO4 can reverse the lethal effect on the parasites. The best inhibitor was hemoglobin.

Optimized codon usage enhances the expression and immunogenicity of DNA vaccine encoding Taenia solium oncosphere TSOL18 gene.

Cysticercosis due to larval cysts of Taenia solium, is a serious public health problem affecting humans in numerous regions worldwide. The oncospheral stage-specific TSOL18 antigen is a promising candidate for an anti-cysticercosis vaccine. It has been reported that the immunogenicity of the DNA vaccine may be enhanced through codon optimization of candidate genes. The aim of the present study was to further increase the efficacy of the cysticercosis DNA vaccine; therefore, a codon optimized recombinant expression plasmid pVAX1/TSOL18 was developed in order to enhance expression and immunogenicity of TSOL18. The gene encoding TSOL18 of Taenia solium was optimized, and the resulting opt-TSOL18 gene was amplified and expressed. The results of the present study showed that the codon-optimized TSOL18 gene was successfully expressed in CHO-K1 cells, and immunized mice vaccinated with opt-TSOL18 recombinant expression plasmids demonstrated opt­TSOL18 expression in muscle fibers, as determined by immunohistochemistry. In addition, the codon-optimized TSOL18 gene produced a significantly greater effect compared with that of TSOL18 and active spleen cells were markedly stimulated in vaccinated mice. 3H-thymidine incorporation was significantly greater in the opt-TSOL18 group compared with that of the TSOL18, pVAX and blank control groups (P<0.01). In conclusion, the eukaryotic expression vector containing the codon-optimized TSOL18 gene was successfully constructed and was confirmed to be expressed in vivo and in vitro. The expression and immunogenicity of the codon-optimized TSOL18 gene were markedly greater compared with that of the un-optimized gene. Therefore, these results may provide the basis for an optimized TSOL18 gene vaccine against cysticercosis.

[Prediction and identification of Taenia solium oncosphere TSO45-4B antigen FnIII structure domain linear B cell epitopes].

OBJECTIVE: To predict and identify Taenia solium oncosphere TSO45-4B antigen Fn III structure domain linear B cell epitopes. METHODS: The B cell epitopes of TSO45-4B Fn III structure domain were predicted through the sequence analysis by using bioinformatics online tools and the protein space conformation was predicted by SWISS-MODEL. The peptides were synthesized according to the predicted linear epitopes. The immunoreactivity of sera of cysticercosis patients to the peptides synthesized was tested by using ELISA. RESULTS: Two linear B cell epitopes of TSO45-4B Fn III structure domain were predicted, and one of the predicted epitope peptides synthesized could be recognized by the sera of cysticercosis patients. CONCLUSION: Two linear B cell epitopes of TSO45-4B Fn III structure domain are predicted and one of them has been confirmed successfully.