Mechanisms of small ubiquitin-like modifier-2/3-ylation of GluK2 receptor in brain injury after deep hypothermic circulatory arrest

Li Bin; Liu Aijun; Liu Jian; Su Junwu; Liu Yinglong

doi:10.3760/j.issn.1673-4777.2019.03.011

Abstract：Objective To explore the role of small ubiquitin-like modifier-2/3 (SUMO2/3)-ylation of GluK2 receptor in brain injury after deep hypothermic circulatory arrest (DHCA) in piglet model.Methods Eighteen piglets were randomly divided into cardiopulmonary bypass (CPB) group,DHCA group and selective antegrade cerebral perfusion(SACP) group,with 6 piglets in each group.Two hours after making DHCA and SACP models,the animals were executed and hippocampal brain tissues were removed.Western blotting was used to test SUMO2/3-ylation,expressions of GluK2,p-MLK3,MLK3,p-JNK3 and JNK3.Co-immunoprecipitation was used to detect the SUMO2/3-ylation of GluK2 and the interaction between GluK2 and MLK3.TUNEL was used to observe cell apoptosis in hippocampus and immunohistochemistry was used to detect expressions of Bax,Bcl-2 and Caspase-3.Results In hippocampus,DHCA was sufficient to significantly increase SUMO2/3-ylation and activation of GluK2 [(1.438 ±0.011),(1.965 ±0.107)] compared with SACP group[(1.220 ±0.039),(1.372 ±0.196),P ＜ 0.05] and CPB group[(0.443 ± 0.025),(0.986 ± 0.181),P ＜ 0.01].Expressions of GluK2-SUMO2/3 and GluK2-MLK3 were significantly higher in DHCA group than those in other two groups (P ＜ 0.01),which could activate phosphorylations of MLK3 and JNK3 (P ＜ 0.05).TUNEL assay demonstrated more neuronal apoptosis (64.8％) of hippocampus in DHCA group than in CPB group(15.8％) and SACP group(47.0％) (P ＜0.001).Immunohistochemistry indicated high levels of Bax and Caspase-3 and low level of Bcl-2 in hippocampus in DHCA group than in other two groups.Conclusion SUMO2/3-ylation of GluK2 in hippocampus may promote its binding with MLK3 to activate the phosphorylation of MLK3 and MLK3-JNK3 pathway,which may be responsible for brain injury after DHCA.