| Objective: To investigate the protective effect of proteasome inhibitor MG132 on intestine and lung injury caused by intestinal ischemia-reperfusion (I/R) of rats through Nrf2/Keap1-ARE pathway.Methods: Thirty-two male SD rats were randomly divided into four groups, control group(A), I/R group(B), control+MG132 pretreatment group(C) and I/R+MG132 pretreatment group(D) (n=8). The I/R model was established by blocking the superior mesenteric artery (SMA) with clap for 1 hour and reperfusion for 2 hours. Pretreatment group was administrated with intraperitonal injection of 0.5 mg/kg MG132 at 30 minutes before ischemia. The control group carried out only laparotomy and isolation of the SMA without clamping. Blood, lung and intestine tissue samples were quickly collected after two hours reperfusion, rinsed in cold saline and stored correspondingly at appropriate conditions. Serum TNF-α, IL-6 and 26S proteasome activities were measured.The immunohistochemical expression and western blot of intestine and lung Nrf2 and HO-1 were preformed.Rusults: Intestinal ischemia-reperfusion induced lung and intestine injury, as was seen as from significant increase of MPO, TNF-αand IL-6. The SOD activities decreased, which implied that intestinal ischemia-reperfusion injury (IIR) reduced the antioxidant capacity of lung and intestinal tissue. However, MG132 pretreatment provided protection against intestinal ischemia-reperfusion induced lung and intestine injury, which could be observed from an improved lung and intestine histological structure, significant decrease of 26S proteasome, MPO, TNF-αand IL-6 activities. Mg132 also enhanced SOD activities and attenuated toxicant caused by oxidative damage. Compared with the normal group, elevation of SOD activities and reduction of 26S proteasome activities were observed. Western blot analysis revealed that IIR increased the expression level of nuclear Nrf2 due to the ROS generated by oxidative damage, which suggested the celluar defensive mechanism of mammalians. MG132 also facilitated the translocation of Nrf2 into the nucleus and this induction of the IIR with the MG132 pretreatment was more powerful than of IIR. The I/R+MG132 group had the highest expression level of Nrf2, possibly due to the dual induction effect afforded by IIR and MG132. The expression level of cytoplasm HO-1 corresponded to the nuclear level of Nrf2, as this enzyme was mainly regulated by the transcriptional factor Nrf2. Immunohistochemisty also provided the same results of Nrf2 and HO-1 expression.Conclusion: The study reveals that intestinal ischemia-reperfusion injury could result in apparent intestine and lung damage. MG132, as a proteasome inhibitor, can ablate intestine and lung injury caused by intestinal I/R, increase the antioxidant and phaseⅡenzymes. The mechanism is complicated, one possible mechanism is attributed to the inhibition of enhanced oxidative stress through an activation of the Nrf2/Keap1-ARE pathway. |
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