Kinetic of Tissue Metabolism after Oral Administration of Recombinant Attenuated Salmonella Typhimurium Carrying Plasmid pcDNA3s in Mice

摘要

A novel vaccine for hepatitis B virus (HBV) was designed by putting a naked DNA vaccine carrying hepatitis B surface antigen (HBsAg) into live-attenuated salmonella typhimurium. The lung, kidney, spleen, mesenteric lymph node, thymus, gonads, feces, muscle and liver were obtained at 4, 7, 30 and 60 h and 4, 7 week after mucosal immunization by the oral route in mice. Semiquantitative PCR technique was used to detect the distribution and kinetics of plasmid in different tissues after extracting total DNA from tissues. Genomic DNA can be assayed for integrated plasmid by PCR after purification of high molecular weight genomic DNA away from free plasmid using gel electrophoresis. The sensitivity of PCR reaction was determined by PCR amplification of the control tissue genome after administering different copies pcDNA3s plasmid. The plasmid clone in feces bacterial in the intestinal was screened by ampicillin. The results showed that foreign plasmid was detected only in thymus and gonads at fourth week, in kidney at seventh week time. Foreign plasmid mainly survived as fragments in vivo. Each tissue genome was negative after PCR amplification and it was lower than the sensitivity of PCR reaction. No positive plasmid clone was detected in feces bacterium, suggesting that plasmid DNA could not transform into bacterium in the intestine. In conclusion, foreign plasmid can be absorbed by gastrointestinal tract and distributed in different tissues quickly, surviving as the form of fragment in vivo. Foreign plasmid DNA could not integrate into the host genome after oral administration.