Shikonin induces glioma cell necroptosis in vitro by ROS overproduction and promoting RIP1/RIP3 necrosome formation  

作　　者：Bin LU Xu GONG Zong-qi WANG Ye DING Chen WANG Tian-fei LUO Mei-hua PIAO Fan-kai MENG Guang-fan CHI Yi-nan LUO Peng-fei GE 

机构地区：[1]Department of Neurosurgery, First Hospital of Jilin University, Changchun 130021, China [2]Research Center of Neuroscience, First Hospital of Jilin University, Changchun 130021, China [3]Department of Hand Surgery, First Hospital of Jilin University, Changchun 130021, China [4]Department of Neurology, First Hospital of Jilin University, Changchun 130021, China [5]Department of Anesthesiology, First Hospital of Jilin University, Changchun 130021, China [6]Department of Neurosurgery, People's Hospital of Jilin Province, Changchun 130021, China [7]Key Laboratory of Pathobiology, Ministry of Education, Jilin University, Changchun 130021, China

出　　处：《Acta Pharmacologica Sinica》2017年第11期1543-1553,共11页中国药理学报（英文版）

基　　金：This work was supported by the National Natural Science Foundation of China （81372697）, the Program for New Century Excellent Talents in University （NCET-12-0233）, the Changbaishan Scholar Project of Jilin Province （2013026）, the Scientific Research Foundation of Jilin province （20150414013GH and 20160101127JC）, and the Bethune Project B of Jilin University （No 2012203）.

摘　　要：Necroptosis is a type of programmed necrosis regulated by receptor interacting protein kinase 1 （RIP1） and RIP3. Necroptosis is found to be accompanied by an overproduction of reactive oxygen species （ROS）, but the role of ROS in regulation of necroptosis remains elusive. In this study, we investigated how shikonin, a necroptosis inducer for cancer cells, regulated the signaling leading to necroptosis in glinoma cells in vitro. Treatment with shikonin （2-10 pmol/L） dose-dependently triggered necrosis and induced overproduction of intracellular ROS in rat C6 and human SHG-44, U87 and U251 glioma cell lines. Moreover, shikonin treatment dose- dependently upregulated the levels of RIP1 and RIP3 and reinforced their interaction in the glioma cells. Pretreatment with the specific RIP1 inhibitor Nec-1 （100 pmol/L） or the specific RIP3 inhibitor GSK-872 （5 pmol/L） not only prevented shikonin-induced glioma cell necrosis but also significantly mitigated the levels of intraceliular ROS and mitochondrial superoxide. Mitigation of ROS with MnTBAP （40 pmol/L）, which was a cleaner of mitochondrial superoxide, attenuated shikonin-induced glioma cell necrosis, whereas increasing ROS levels with rotenone, which improved the mitochondrial generation of superoxide, significantly augmented shikonin-caused glioma cell necrosis. Furthermore, pretreatment with MnTBAP prevented the shikonin-induced upregulation of RIP1 and RIP3 expression and their interaction while pretreatment with rotenone reinforced these effects. These findings suggest that ROS is not only an executioner of shikonin-induced glioma cell necrosis but also a regulator of RIP1 and RIP3 expression and necrosome assembly.Necroptosis is a type of programmed necrosis regulated by receptor interacting protein kinase 1 （RIP1） and RIP3. Necroptosis is found to be accompanied by an overproduction of reactive oxygen species （ROS）, but the role of ROS in regulation of necroptosis remains elusive. In this study, we investigated how shikonin, a necroptosis inducer for cancer cells, regulated the signaling leading to necroptosis in glinoma cells in vitro. Treatment with shikonin （2-10 pmol/L） dose-dependently triggered necrosis and induced overproduction of intracellular ROS in rat C6 and human SHG-44, U87 and U251 glioma cell lines. Moreover, shikonin treatment dose- dependently upregulated the levels of RIP1 and RIP3 and reinforced their interaction in the glioma cells. Pretreatment with the specific RIP1 inhibitor Nec-1 （100 pmol/L） or the specific RIP3 inhibitor GSK-872 （5 pmol/L） not only prevented shikonin-induced glioma cell necrosis but also significantly mitigated the levels of intraceliular ROS and mitochondrial superoxide. Mitigation of ROS with MnTBAP （40 pmol/L）, which was a cleaner of mitochondrial superoxide, attenuated shikonin-induced glioma cell necrosis, whereas increasing ROS levels with rotenone, which improved the mitochondrial generation of superoxide, significantly augmented shikonin-caused glioma cell necrosis. Furthermore, pretreatment with MnTBAP prevented the shikonin-induced upregulation of RIP1 and RIP3 expression and their interaction while pretreatment with rotenone reinforced these effects. These findings suggest that ROS is not only an executioner of shikonin-induced glioma cell necrosis but also a regulator of RIP1 and RIP3 expression and necrosome assembly.

关 键 词：SHIKONIN glioma necrosome ROS mitochondrial superoxide RIP1 RIP3 Nec-1 GSK-872 MnTBAP ROTENONE 

分 类 号：R[医药卫生]