Objective To examine the expression of ? defensin genes and inflammatory cytokines genes in rat airway epithelium after intraperitoneal injection of emulsified isoflurane.Methods Thirty-six SD rats were assigned into six groups(A-F) randomly.The tissue from rat airway epithelium were harvested at 1 h after intraperitoneal injection of normal saline,30% fat milk and 8% emulsified isofluranes of 4 different dosages(0.25,0.5,1.0 and 1.5 times of ED50).Then expressions of ? defensin(BD-1,BD-2),tumor necrosis factor-alpha(TNF-?),interleukin beta(IL-1?),IL-8 mRNA were measured by semiquantitative polymerase chain reaction(PCR) using GAPDH as an internal standard.Results Except BD-1 and IL-8 mRNA,marked increases in expressions of TNF-?,IL-1? and BD-2 genes were observed with a dosage-dependent manner.Conclusion The expressions of BD-2 genes and inflammatory cytokines genes(IL-1?,TNF-?)in rat airway epithelium increase significantly in a dosage-dependent manner after intraperitoneal injection of emulsified isoflurane.

Objective To construct the bait plasmid of HNP-3 mature peptide in yeast two-hybrid system and examine whether the recombinant bait plasmid has self-activating and toxicity effect.Methods Using RT-PCR technique,the cDNA fragments of HNP-3 mature peptide gene were amplified from the extracted RNA in cultured HL-60 cells.The fragment was firstly cloned into pBluescript-SK-II vector,confirmed by sequencing,then sub-cloned into the bait plasmid pGBKT7 and identified with PCR and sequence analysis techniques.The recombinant plasmid was introduced into the yeast cell AH109,and its self-activating and toxicity effect was tested by auxotrophic selective culture.Results DNA sequencing indicated that the inserted fragment in pBluescript-SK-II vector was HNP-3 mature peptide gene sequence,and the sub-cloned recombinant pGBKT7-HNP-3 was no mismatch.The recombinant bait plasmid didn't have self-activating effect and did not show toxicity to yeast AH109 cell.Conclusion The bait plasmid of HNP-3 mature peptide was constructed successfully.This was helpful for investigating the proteins interacting with HNP-3 mature peptide by yeast two-hybrid technique.