Adenoviral vector mediated-expression of caspase-3 siRNA on apoptosis induced by lipopolysaccharide

Objective: To construct the recombinant adenovirus expressing small RNA of rats caspase-3 and observe the down-regulation effect of caspase-3 in neurons induced by lipopolysaccharide(LPS) in vitro. Methods: pShuttleH1-siCas3 containing Oligo DNA of the targeting sequences and pEGFPC1-Cas3 containing caspase-3 and EGFP sequences were constructed respectively.pShuttleH1-siCas3 and pEGFPC1-Cas3 were co-transfected to the 293 cells by liposomes to determine interfering efficacy by flow cytometry. pShuttleH1-siCas3 was linearized and transformed into E. coli BJ5183 cells containing backbone plasmid pAdEasy-1. The recombinant plasmid was transfected into 293 cells to package the adenovirus Ad-siCas3. The titers of adenovirus were determined by the specific 50% tissue culture infection dosage method. After virus infected the cultured hippocampus neurons, LPS-induced apoptosis and caspase-3 mRNA expression were observed. Results: It was identified that the sequence of target gene was correctly inserted into the genome of virus.The expression of green fluorescence protein was reduced by pShuttleH1-siCas3 in 293 cells. The titer of recombinant adenovirus was 1.06×1010 pfu/ml. After virus infection, caspase-3 mRNA was greatly reduced and neurons apoptosis was suppressed. Conclusion: The recombinant adenovirus expressing rats caspase-3 siRNA were successfully constructed, which may probably be further used in pain therapy by its anti-apoptosis effect.

Taxonomy of Ochroconis, genus including opportunistic pathogens on humans and animals

The genus Ochroconis(Sympoventuriaceae,Venturiales)is revised and currently contains 13 species for which the phylogenetic position has been determined using multilocus sequencing.The older generic name Scolecobasidium is considered to be of doubtful identity because the type specimen is ambiguous.Within the Ochroconis lineage,phylogenetic distances of all markers analyzed are exceptionally large,both between and within species.A new genus Verruconis is proposed for the neurotropic opportunist Ochroconis gallopava.Species accepted within the lineages are keyed out on the basis of phenotypic characters.Main ecological traits within each species are discussed.Verruconis species are thermophilic and one of them is an important agent of infection in the brain,while Ochroconis is mesophilic,several species causing infections in cold-blooded animals.