Effects of penehyclidine hydrochloride on Nrf2/ARE signaling pathway during endotoxin-induced acute lung injury in neonate rats

Abstract

Objective: To evaluate the effects of penehyclidine hydrochloride on the nuclear factor erythroid 2-related factor 2/antioxidant responsive element (Nrf2/ARE) signaling pathway during endotoxin-induced acute lung injury(ALI) in neonate rats. Methods: Forty 7-day-old Wistar rats weighing 12-18 g were randomly divided into 4 groups (n=10) using a random number table: normal saline group(NS group), acute lung injury(ALI group), penehyclidine hydrochloride group(PHC group) and penehyclidine hydrochloride+ Nrf2 siRNA plasmid group(PNS group). The ALI model was induced with intraperitoneal endotoxin (5.0 mg/kg) in groups ALI, PHC and PNS. In groups PHC and PNS, penehyclidine hydrochloride (2.0 mg/kg) was injected intraperitoneally at 1 h before ALI respectively, while the equal volume of normal saline was administered in groups NS and ALI. The animal of PNS group were inhaled adenovirus packaging of Nrf2-siRNA three times (one time a day) before modeling. At 4 h after endotoxin injection, the rats were sacrificed. The lungs were collected to determine the wet/dry(W/D) lung weight ratio. The expression of Nrf2 and heme oxygen and enzyme 1(HO-1) were determined by Western blotting, contents of tumor necrosis factor-alpha(TNF-α),interleukin10 (IL-10)were determined by enzyme-linked immunosorbent assay(ELISA). The cell apoptosis were determined by transferase-mediated deoxyuridine triphosphate-biotin nick end labeling(TUNEL),and the apoptotic index was calculated. Results: The W/D ratio in NS, ALI, PHC and PNS groups were (4.2±0.1), (9.6±0.7), (6.5±0.6), (8.3±1.3) respectively. The apoptotic index were (3.7±0.5)%, (31.5±3.2)%, (17.6±4.2)%, (28.1±3.5)%respectively.The contents of TNF-α were (10.3±1.6), (98.5±8.5), (68.5±6.7), (89.9±8.5) pg/ml respectively. The contents of IL-10 were (7.9±0.6), (102.8±9.3), (72.5±5.8), (97.7±9.1) pg/ml respectively.The expression of Nrf2 were (23.2±7.6), (79.8±13.0), (155.5±16.7), (12.0±3.3) respectively. The expression of HO-1 were (31.7±8.6), (90.8±10.3), (147.6±22.5), (61.4±9.7) respectively. There were statistically significant differences among different groups (F=86.013, 154.897, 328.810, 374.198, 333.965, 125.274, all P<0.05). Compared with group NS, the W/D ratio, apoptotic index and the contents of TNF-α, IL-10 increased, the expression of Nrf2 and HO-1 up-regulated in group ALI and group PHC (all P<0.05). Compared with group ALI, the W/D ratio, apoptotic index and the contents of TNF-α, IL-10 decreased, the expression of Nrf2 and HO-1 up-regulated in group PHC (all P<0.05). Compared with group ALI, no significant differences were found in the W/D ratio, apoptotic index and the contents of TNF-α, IL-10 in group PNS(all P>0.05), while the expression of Nrf2 and HO-1 down-regulated in group PNS (all P<0.05). Compared with group PHC, the W/D ratio,apoptotic index and the contents of TNF-α,IL-10 increased, the expression of Nrf2 and HO-1 down-regulated in group PNS (all P<0.05). Conclusion: Nrf2/ARE signaling pathway is involved in the reduction of ALI by penehyclidine hydrochloride in neonate rats.