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Objective: To express antibacterial peptide thanatin in the prokaryotic expression system and test its antibacterial activity., Methods: The DNA sequence coding for the 21 peptides of thanatin was synthesized using the preferred genetic codes of E. coli, cloned into pTYB11 plasmid, and transformed into E.coli ER2566. The expression of thanatin fused with intein was induced by IPTG in E.coli, and intein-thanatin specifically bound to the column through intein tag was cleaved overnight at 4 degrees celsius; in DTT/cysteine buffer., Results: The cleaved thanatin was eluted with a protein concentration of 245 microg/ml in the first 4 ml. The purified thanatin had showed strong antibacterial activities against G- bacteria such as Shigella flexneri, Klebsiella pneumoniae, Shigella snnei, Escherichia coli O157, toxin producing Escherichia coli, Pseudomonas aeruginosa, and fungi such as Candida albicans, with especial potency in killing drug-resistant Klebsiella pneumoniae, Pseudomonas aeruginosa, and extended-spectrum beta-lactamases (ESBL)-producing E.coli. Eighty strains of drug-resistant (ESBL-producing) and 30 strains of sensitive E. coli were used for anti-bacterial assay, and no significant differences in the antibacterial activity of thanatin were found between the sensitive and drug-resistant E. coli (P>0.05)., Conclusion: The recombinant thanatin obtained shows strong antibacterial activity against drug-resistant and sensitive bacteria, and can be a potential substitute for routine antibiotics in the treatment of G- bacterial infections.

Objective: To verify the miRNA in Aedes albopictus and characterize the expression profile of several miRNAs across all the life stages of Aedes albopictus., Methods: Based on the published miRNA sequences of Anopheles gambiae and Aedes aegypti, 6 DIG-labeled antisense probes were synthesized. The total RNAs from Aedes albopictus in 6 developmental stages (embryo, early larvae, late larvae, pupa, male and female adults) were extracted with a mirVana miRNA isolation kit, loaded onto 15% denaturing polyacrylamide gel and hybridized with the appropriate DIG-labeled probes., Results: Northern blotting detected 5 miRNAs in Aedes albopictus, of which mir-9a was mainly expressed in embryo and larva stages, let-7 in pupa and adult stages, miR-184 in all life stages, mir-M1 only in the embryos and miR-1175 in all the life stages except for embryos. The expression profiles of these miRNAs in Aedes albopictus were similar to those in D. melanogaster and An.stepheni. miR-1174 was not detected in any of the developmental stages of Aedes albopictus., Conclusion: These results present the first direct experimental evidence of miRNA in Aedes albopictus. The expression profiles of the analyzed miRNAs in Aedes albopictus showed stage specificity and conservation with other mosquitoes. Further studies on the functions of these miRNAs may offer new insights in mosquito biology and may lead to novel approaches to the development of insecticides.

Objective: To investigate the effect of cinobufagin on nuclear factor-kappaB (NF-kappaB) pathway of liver cancer cell line HepG2., Methods: Dual-luciferase cis-reporting system was used to detect the relative value of pNF-kappaB-TA-luc upon tumor necrosis factor-alpha (TNF-alpha) stimulation of NF-kappaB pathway. Western blotting was used to detect the protein level of NF-kappaB p65, and RT-PCR was used to detect the gene transcription level of intercellular adhesion molecule-1 (ICAM-1), a target downstream gene of NF-kappaB., Results: At the concentration of 0.25 and 0.5 microg/ml, cinobufagin significantly lowered the relative value of luciferase (P<0.05). The results of Western blotting showed that cinobufagin significantly suppressed the protein expression of NF-kappaB p65. The transcription level of ICAM-1 was reduced by different doses of cinobufagin., Conclusion: The anti-cancer effect of cinobufagin may be related to its activity in inhibiting the activation of NF-kappaB pathway.

Objective: To construct the life cycle of Angiostrongylus cantonensis (A.cantonensis) in laboratory condition., Methods: SD rats were infected orally with the third-stage larvae of A.cantonensis collected from Jiangmen, Guangdong province. Six weeks after infection, the first-stage larvae were isolated from fresh feces of the rats by using Baermann funnel to infect 25 second-generation white jade snails raised in laboratory at the daily dose of 300 000 for 3 consecutive days. Three weeks later, the snails were dissected for counting the third-staged larvae of A.cantonensis, and those positive for A.cantonensis infection were fed directly to 10 fasting rats. The serum samples of the rats were then collected 2 weeks later for examination of specific antibodies using ELISA. The feces of the infected rats were examined microscopically after 6 weeks, and the brain, heart and lungs of the infected rats were dissected to observe the larvae at 3, 5, and 8 weeks, respectively., Results: The 3-stage larvae of A.cantonensis were found in the second-generation snails 3 weeks after infection. The positivity rate of serum specific antibodies was 100% in the 10 rats 2 weeks after feeding of the infected snails. The 1-stage larvae were detected in the feces of the rats 6 weeks after infection, and the fourth-stage larvae were found in the brain of the rats at 3 weeks, while adult worm and eggs were found in the heart and lungs of the infected rats at 5 and 8 weeks., Conclusion: The successful establishment of human colon carcinoma cell line with PRL-3 gene knock-down provide a basis for investigation of the role of PRL-3 gene in the metastasis of human colorectal carcinoma.

Objective: To identify antigens which may help evaluate the therapeutic effect of angiostrongyliasis from adult worm antigen of Angiostrongylus cantonensis., Methods: The adult worm antigens of A. cantonensis were analyzed by Western blotting with the sera of rats infected with A. cantonensis before and after treatment. The sera of rats were tested by enzyme-linked immunosorbent assay (ELISA)., Results: The antigens with relative molecular mass between 38,000 and 78,000 reacted not only with the sera of rats before treatment, but also with that after treatment. The antigens with M(r) between 190,000 and 17,000 reacted with the sera of rats before treatment but not with that after treatment; those with M(r) between 32,000 and 24,000 antigens strongly reacted with the former, but the reaction became much weakened with the latter. The AC32-IgG antibody appeared earlier than the AC-IgG, and disappeared rapidly after treatment. Six of the 10 treated rats became negative for AC-IgG as found by ELISA., Conclusion: The antigens of adult worm antigen of A. cantonensis with M(r) of 190,000, 32,000, 24,000, 17,000 and 16,000 may serve as candidate antigens for therapeutic effect evaluation of angiostrongyliasis.