Construction Onephropathy Model In The Minipigs And Intervention Study

Objective: Acute kidney injury (AKI) is a common and serious clinical problemand is associated with high mortality, and few preventive and therapeutic options. Itis also an important risk factor for the eventual progression to end-stage renaldisease and the development of chronic kidney disease, which further imposesenormous medical and economic burdens on society. Aminoglycoside antibiotics areextensively used in the treatment of Gram-negative bacterial infection, such asgentamicin and kanamycin. However, the major clinical problem in the use ofgentamicin is gentamicin nephrotoxicity. Cytosolic gentamicin acts on mitochondria,and thus activates the intrinsic pathway of apoptosis, produces oxidative stress byincreasing superoxide anions and hydroxyl radicals, which further contributes to celldeath. Because autophagy is a mechanism of removing damaged biologicalmaterials and dysfunctional organelles, autophagy is very important to maintain thecellular homeostasis. Reactive oxygen species (ROS) caused by mitochondrialdysfunction could have a major role in gentamicin nephrotoxicity. Importantly,autophagy can degrade the damaged mitochondria and alleviate the oxidativedamage in cells. However, the role of autophagy in gentamycin nephrotoxicity andthe effect of the antioxidant N-acetyl-L-cysteine and rapamycin, a specific inhibitorof mTOR on the autophagy in the gentamicin nephrotoxicity is unclear.In this study, we investigated the changes of autophagy in gentamicin-inducednephrotoxicity in the minipigs kidneys.Methods:In the dose-response study of gentamicin, the minipigs received a dailyi.m. injection for10days of gentamicin sulfate (5,6.85,10,15,20,40,60,80and100mg/kg, each group=6). For the gentamicin time course study, minipigs receivedan i.m. injection of80mg/kg gentamicin daily for0,1,3,5,7and10days. We examined the levels of Scr, BUN, autophagy, oxidative damage and apoptosis.Finally, we compared the effects of gentamicin alone, andN-acetyl-L-cysteine(NAC)/rapamycin combination with gentamicin in the minipigskidneys.Results: First, in the dose-response study of gentamicin, the levels of BUN andcreatinine were increased significantly in80mg/kg and100mg/kg gentamicincompared to the control group. Second, in the time course study of gentamicin, thelevels of BUN and creatinine and ATN score were increased significantly in80mg/kg gentamicin daily for10days compared to the0day group.The levels ofp62/SQSTM1, polyubiqintin aggregates, p-Parkin, AMBRA1and PINK1wereincreased significantly in the10days group. Conversely, the levels of Bnip3andHIF-1α were decreased significantly in the10days group compared to the0daysgroup. In addition, the makers of oxidative damage, such as the4HNE, γ-H2AX andprotein carbonyl were increased obviously in the10days group.Analysis bytransmission electron microscope demonstrated swelling and disintegration ofcristae in the mitochondria of the10days group.Third, compared with thegentamicin nephrotoxicity group, NAC and rapamycin enhanced the level ofautophagy, decreased the oxidative damage，but cannot ameliorate the renal functionand the tubular necrosis in the kidneys.Conclusions: Firstly, nephrotoxicity is a dose-limiting side and time-dependenteffect of gentamicin in the minipigs. With gentamicin, it was demonstrated that aprogressive decrease of renal function, the level of autophagy, especially mitophagyand the increase of apoptosis and oxidative damage in minipigs kidneys.