Abstract:

Ｏｂｊｅｃｔｉｖｅ：To investigate the role of reactive oxygen species on necroptosis in renal tubular epithelial cell.
Ｍｅｔｈｏｄｏｌｏｇｙ：The necroptosis model of HK2 cell was constructed as our previous research.Apocynin,a specific inhibitor of NADPH oxidase was delivered in the necroptosis model.ROS production was detected by Dichlorodihydrofluorescein diacetate.The manner of cell death was identified by flow cytometry.Western Blot was used to determine phosphorylation of receptorinteracting protein kinase 3(RIP3) and mixed lineage kinase domainlike (MLKL)which are essential to necroptosis.
Ｒｅｓｕｌｔｓ：Necroptosis model of HK2 cell was established by TNFα,benzyloxycarbonylValAlaAspfluoromethylketone (zVADfmk) and antimycin A.In this model,PI(+) HK2 cells was increased and phosphorylation of RIP3 and MLKL was augmented.ROS increased in necroptosis group（4329±249 vs 2590±127,P<0001） and could be inhibited by Nec1（3558±108 vs 4329±249,P=0002）.Apocynin not only decreased ROS production(3071±282 vs 4329±249,P<0001）,but also reduced the proportions of necrosis in the necroptosis model（200%±030% vs 699%±279%,P<0001）.Phosphorylated RIP3 and MLKL was also decreased by Apocynin.
Ｃｏｎｃｌｕｓｉｏｎ：
ROS play an important role in necroptosis of HK2 cell.Necroptosis could be ameliorated by inhibiting ROS production.

Key words: reactive oxygen species, necroptosis, renal tubular epithelial cell, acute kidney injury