Procyanidin B2 Inhibits NLRP3 Inflammasome Activation And Protects Against Oxidative Stress Injury In H9C2 Cells

ObjectiveTo investigate the protective effects of proanthocyanidin B2（PCB2）on H9C2 cardiomyocyte injury induced by hydrogen peroxide（H₂O₂）or high glucose（HG）,and to explore its possible mechanism.MethodsThe H₂O₂ injury protection experiments were grouped into normal control group,H₂O₂ injury group,H₂O₂+PCB2（5μmol/L）,H₂O₂+PCB2（15μmol/L）,H₂O₂+PCB2（45μmol/L）.The high glucose damage protection experiments were divided into normal control group,HG injury group,HG+PCB2（5μmol/L）,HG+PCB2（15μmol/L）,HG+PCB2（45μmol/L）.MTT assay was used to detect the viability of H9C2 cardiomyocytes.Western blotting was used to detect NOD-like receptor protein-3（NLRP3）,cysteine-containing aspartate proteolytic enzyme（caspase-1）,and leukocyte-mediated Protein expression levels of prime（IL）-1βand IL-18.The amount of reactive oxygen species（ROS）in the cells was detected by fluorescent dye dihydroethidine（DHE）staining.The activity of lactate dehydrogenase（LDH）,malondialdehyde（MDA）content,the activities of total antioxidant enzymes（T-AOC）,superoxide dismutase（SOD）,catalase（CAT）and glutathione peroxidase（GSH-Px）were detected.Results（1）Compared with the control group,when H₂O₂ was applied to H9C2cardiomyocytes,the cell viability level decreased significantly,and the release of lactate dehydrogenase（LDH）increased,indicating that H₂O₂ caused cell damage.Compared with the H₂O₂ group,PCB2 increased cell viability,decreased the release of LDH,and the effect was dose-dependent,indicating that PCB2 can alleviate oxidative damage of H9C2 cardiomyocytes induced by H2O2.（2）Compared with the control group,the expression levels of NLRP3,caspase-1,IL-1βand IL-18 protein were increased in the H₂O₂ group.Compared with the H₂O₂ group,the expression levels of NLRP3,caspase-1,IL-1βand IL-18 protein in the PCB2 group were reduced in a dose-dependent manner.（3）Compared with the control group,the production of MDA and ROS in the H₂O₂ group increased.Compared to H₂O₂ PCB2 treatment reduced MDA and ROS production in a dose-dependent manner.（4）The activities of T-AOC,SOD,CAT and GSH-Px in the H₂O₂ group were lower than those in the control group.However,the treatment of PCB2increased the activity of T-AOC,SOD,CAT and GSH-Px in a dose-dependent manner compared to the H₂O₂ group.（5）Compared with the control group,the activity level of cardiomyocytes in the HG group decreased,and the release of lactate dehydrogenase（LDH）increased,causing cell damage.Compared with the HG group,the activity of the cells in the PCB2 group increased,the release of LDH decreased,and the effect was concentration-dependent,indicating that PCB2 can alleviate the damage of H9C2 cardiomyocytes induced by HG.（6）Compared with the control group,HG increased the levels of MDA and ROS in cardiomyocytes.Compared to HG,the production of MDA and ROS in the PCB2 group was reduced.（7）Compared with the control group,the activities of T-AOC,SOD,CAT,and GSH-Px in the HG group were decreased.The activity of T-AOC,SOD,CAT and GSH-Px in the PCB2 group was increased compared with the HG group.ConclusionProanthocyanidin B2 has protective effects against H₂O₂ or HG-induced oxidative stress damage in H9C2 cardiomyocytes.Its mechanism is related to the inhibition of the activation of NLRP3 inflammasome.