siRNA 를 이용한 허혈 후 뇌 신경계에서의 유전자 발현 조절

초록

High mobility group box-1 (HMGB-1) is a non-histone DNA binding protein, which is recently identified as cytokine-like mediators in delayed endotoxin lethality and acute lung injury. High serum levels of HMGB-1 in patients with sepsis are associated with increased mortality, and administration of recombinant HMGB-1 produces acute inflammation in animal models of lung injury and endotoxemia. Here we report gene silencing of HMGB1 in the primary cortical culture and in the brain by using small interfering RNA (siRNA). Short hairpin RNA (shRNA) targeting HMGB1 was expressed endogenously under U6 promoter, which was delivered into the postischemic brain via a novel cationic polymer, PAMAM-R. We found that the expression of shRNA was detected beginning 2 days after the administration, which was maintained for more than 5 days. Induction of shRNA for HMGB1 suppressed the endogenous hmgb1 expression between 3 and 7 days after the administration. In addition, we demonstrated that endogenous expression of shRNA can trigger a significant suppression of HMGB1 induction in postischemic brain, which was observed in the ischemic hemisphere after 1 hr of transient middle cerebral artery occlusion (MCAO). A notable reduction in infarct size along with the suppression of the proinflammatory cytokine expression was accompanied by it, indicating that HMGB-1 plays a crucial role in inflammatory process in postischemic brain. The efficacy of plasmid-mediated expression of siRNA shown here extends the application of siRNA to therapies and to in vivo targeting experiments that aim to define the function of specific genes.