Chronic cyclosporine nephropathy is characterized by excessive autophagosome formation and decreased autophagic clearance.
BACKGROUND
The study was performed to investigate the influence of cyclosporine A (CsA)-induced renal injury on autophagy in an experimental model of chronic CsA nephropathy.


METHODS
Three dosages of CsA (7.5, 15, and 30 mg/kg/day) were administered to mice for 4 weeks. The formation of autophagosomes was measured with microtubule-associated protein 1 light chain 3 phospholipid-conjugated form (LC3-II) and beclin-1, and the ability of autophagic clearance was examined with sequestosome-1 (p62). Autophagic vacuoles were visualized and counted using electron microscopy. Double immunolabeling of LC3-II and active caspase-3 was performed to evaluate the association between autophagy and apoptosis. Oxidative stress was evaluated by measuring urinary 8-hydroxy-2'-deoxyguanosine excretion, demonstrating oxidative DNA damage. Antioxidative drugs, pravastatin and N-acetylcysteine, were used to evaluate the role of CsA-induced oxidative stress on autophagy.


RESULTS
CsA treatment increased the expressions of LC3-II and beclin-1 in the kidney in a dose-dependent manner. The number of p62-positive cells was also significantly increased in a CsA dose-dependent manner. Electron microscopy revealed excessive autophagic vacuoles in the CsA group compared with the vehicle group. Expression of active caspase-3 was increased in a CsA dose-dependent manner and was colocalized with LC3-II in the injured area of CsA-treated kidneys. Concurrent pravastatin or N-acetylcysteine treatment reduced urinary excretion of 8-hydroxy-2'-deoxyguanosine and subsequently decreased LC3-II expression and the number of p62-positive cells compared with the CsA group.


CONCLUSIONS
Chronic CsA nephropathy is a state of excessive autophagic vacuoles and decreased autophagic clearance. Oxidative stress may play an importation role in the induction of autophagy.
Convergent Research Consortium for Immunologic Disease, Seoul St. Mary's Hospital, The Catholic University of Korea, Seoul, Korea.
Sun Woo
SW
Lim
Bok Jin
BJ
Hyoung
Shang Guo
SG
Piao
Kyoung Chan
KC
Doh
Byung Ha
BH
Chung
Chul Woo
CW
Yang
D000080242:8-Hydroxy-2'-Deoxyguanosine; D000111:Acetylcysteine / Q000627:therapeutic use; D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D017209:Apoptosis; D001343:Autophagy / Q000187:drug effects*; D053148:Caspase 3 / Q000096:biosynthesis; D016572:Cyclosporine / Q000633:toxicity*; D003849:Deoxyguanosine / Q000031:analogs & derivatives / Q000652:urine; D004305:Dose-Response Relationship, Drug; D051638:Ether-A-Go-Go Potassium Channels / Q000096:biosynthesis; D007674:Kidney Diseases / Q000209:etiology / Q000473:pathology*; D008297:Male; D051379:Mice; D008854:Microscopy, Electron / Q000379:methods; D008869:Microtubule-Associated Proteins / Q000096:biosynthesis; D018384:Oxidative Stress; D010588:Phagosomes / Q000378:metabolism; D017035:Pravastatin / Q000627:therapeutic use; D013997:Time Factors
D000975:Antioxidants; D051638:Ether-A-Go-Go Potassium Channels; C496887:Kcnh3 protein, mouse; C480860:Map1lc3b protein, mouse; D008869:Microtubule-Associated Proteins; D016572:Cyclosporine; D000080242:8-Hydroxy-2'-Deoxyguanosine; D053148:Caspase 3; D003849:Deoxyguanosine; D017035:Pravastatin; D000111:Acetylcysteine