| Background: Radiotherapy is one of the most effective treatments for primary or metastatic brain tumors.However,while killing malignancies,radiotherapy inevitably damages surrounding normal brain tissue,which may induce neurological and cerebrovascular pathologies,causing progressive and irreversible cognitive impairment in patients.Although the exact pathological mechanism of radiation-induced brain injury remains unclear,radiation-induced persistent neuroinflammatory response in the brain,including microglial overactivation and production of a series of pro-inflammatory factors,may play an important role in the pathogenic process.Therefore,inhibiting the activation of microglial pro-inflammatory phenotype is an important strategy to limit neuroinflammatory progression after radiotherapy.Lycium barbarum polysaccharide(LBP)is a major active ingredient in the traditional Chinese medicine lycium barbarum.Several studies have shown its anti-inflammatory,antioxidant,and neuroprotective properties,but the effects and specific mechanism of inhibiting microglial overactivation and oxidative stress induced by ionizing radiation remain to be investigated.Objective:To investigate the effects of LBP on radiation-induced overactivation and oxidative stress in BV2 microglial cells,and to further elucidate the underlying molecular mechanism of the action.Methods: Mouse BV2 cell lines were cultured in DMEM supplement with 10% FBS and were divided into control group(no radiation and LBP treatment),radiation group(radiation only),100 μg/ml LBP group(100 μg/ml LBP and radiation),200 μg/ml LBP group(200 μg/ml LBP and radiation),400 μg/ml LBP group(400 μg/ml LBP and radiation).Cells were pretreated with different concentrations of LBP for 12 h before Xray irradiation,and were collected 12 h after irradiation.(1)Cell viability was detected by CCK-8 assay.(2)Griess kit was used to detect the concentration of NO in the culture medium.(3)The protein expressions of TNF-α,IL-1β,p65,p-p65,IκBα,p-IκBα,IKKβ,p-IKKβ,HO-1 and NQO-1 were detected by western blot.(3)Immunocytofluorescence assay was used to detect the expression of BV2 polarization markers i NOS,Arg-1,and YM-1.(4)The level of intracellular ROS was detected by DCFH-DA fluorescent probe.The enzymatic activities of SOD and GPx were evaluated by SOD and GPx detection kits.Result:(1)Pretreatment with LBP did not alter the survival rate of BV2 cells after irradiation.(2)LBP reduced the radiation-induced production of inflammatory factors(TNF-α、IL-1β、NO)in BV2 cells.(3)Compared with the radiation group,the i NOS protein expression of LBP group was decreased,and the Arg-1 and Ym-1 protein expressions were increased.(4)LBP inhibited the nuclear translocation of p65 protein and reduced the expressions of p-p65,p-IκBα and p-IKKβ protein in the NF-κB pathway after irradiation.(5)X-ray significantly induced the production of ROS in BV2 cells,but LBP did not change the level of intracellular ROS.(6)Compared with the control group,X-ray and LBP treatment did not change the activities of SOD and GPx,as well as the expressions of HO-1 and NQO-1 protein.Conclusions:(1)LBP suppressed the production of inflammatory factors and promoted the transformation of BV2 cells from M1 pro-inflammatory phenotype to M2anti-inflammatory phenotype after radiation by negatively regulating IKKβ/IκBα/NF-κB pathway.(2)LBP could not eliminate the radiation-induced oxidative stress,or improve the antioxidant capacity of BV2 cells. |
