Ikappab-alpha Transfer The Gene Therapy Of Acute Lung Injury

Acute lung injury occurs as a result of lung inflammation initiated by either infectious or noninfectious stimuli, accompanied with lung neutrophil accumulation . The excessive SIRS (systemic inflammatory response syndyome), induced by an elevated level of proinflammatory mediators combined with a decreased expression of anti-inflammatory molecules is a critical component of acute lung injury . Expression of proinflammatory genes is regulated by transcript ional mechanisms. Nuclear factor-κB (NF-κ B) is one critical transcription factor required for maximal expression of many cytokines involved in the pathogenesis of acute lung injury. The activation of NF- K B is associated with the generation of acute lung injury. Blocking the activation of NF-κ B and down-regulating production of inflammatory mediators could prove to be an important methods to the therapy of acute lung injury .1κB is the inhibitor of NF-κ B. In our research, we clone the Chinese IKB-α gene, construct the eukaryotic express vectors Ad-AN I KB-α and investigate its inhibitory effect to NF-κB in vitro and in vivo, aim to provide clinical information for gene therapy of acute lung injury .Contents:1. Prepare the model of acute lung injury by IPS ip, investigate the expression of NF-κB.2. Clone the Chinese I κB-α gene by RT-PCR from placenta, modify its gene by delete the NH2-terminal 32 and 36 amino acids(AN IKB-OC) .Constructe its eukaryotic express vectors pShuttle/N I κB-α.3. Constructe adenovirus vectors: Ad-N I κ B-α and Ad-LacZ.4. Investigate the inhibitory effect of N I κB-α to NF-κB in vitro and in vivo by western blotting , ELISA and immunohistochemistry.Results:1. The espression of NF-κB in lung tissue of mouse with ALIchanges according to the time after IPS ip.It becomes strongest 3 hours after IPS stimulation, and then decreases.2. The activation of NF-κB induced by LPS is limitedsignificantly after transferring N IκB-α into A549 cells.3. Adenovirus vectors can infect alveolar epithelial cells effectively and express the target gene highly.4. Both NF-κB activity and neutrophil differential count , TNF-α, IL-6 content in BALF of mouse with ALI induced by LPS after AN IκB-α transgene for lung are decreased significantly.Conclusion:N IκB-α transgene for lung can inhibit NF-κ B activity and downregulate the expression of TNF-α IL-6 in BALF of mouse with ALI, suggesting that AN I κB-α might play an anti-inflammatory role in ALI induced by LPS .