Effects Of PAS-Na On The Damage Of Primary Cultured Microglia Inflammation Induced By Manganese In Rats

Objective To study manganese on rat primary microglia oxidative damage and inflammatory cytokines interleukin(IL-1 and IL-6),tumor necrosis factor alpha(TNF-?),prostaglandin 2(PGE2)expression,explores the amino sodium salicylate(PAS-Na)intervention of microglia lesions caused by manganese.? methods ?(1)microglia after separation,purification and identification,(1)manganese toxicity test: in 96-well plate cells respectively by five groups with 0,200,300,400 and 500 mol/L concentration of MnCl2 for 24 h.(2)PAS-Na screening test: in 96-well plate cells respectively by four groups with 0,50,150 and 450mol/L concentration of PAS-Na for 24 h.(3)PAS-Na intervention trial:microglia were randomly divided into control,Mn exposed,PAS control,50?150 and 450-PAS group.control,Mn exposed and PAS control group was given normal DMEM culture medium,50?150 and 450-PAS group joined the 400mol/L concentration of Mn Cl2 culture medium for 24 h,after that the control group was given normal DMEM culture medium,the Mn group was given 400mol/L concentration MnCl2 DMEM culture medium,the PAS control group was given 450 mol/L concentration PAS-Na DMEM culture medium.50?150 and450-PAS group was added 50,150 and 450 mol/L concentration PAS-NaDMEM culture medium for 24 h.(2)microglia survival rate is measured by determined by MTT,DCFH-DA labeled microglia oxidative injury,determination of cell supernatant of IL-1?,IL-6,TNF-?,PGE2 levels by enzyme-linked immunosorbent assay,the expression of fluorescence quantitative PCR detection of microglia IL-1?,IL-6 and TNF-?mRNA.Results the cell survival rate: detection of low concentrations of MnCl2 can stimulate primary microglia cells proliferation and increase the survival rate,high concentration of MnCl2 have cytotoxicity on microglia,MnCl2 processing after24 h microglia cell survival rate at 400 mol/L and 500 mol/L were lower than the control group(P<0.05).After PAS-Na treatment,compared with the control group,there was no significant change in cell morphology and survival rate(P>0.05)of each treatment group(24h).After Mn 24 h,50?150 and 450-PAS intervention,the survival rate of Mn group was lower than that of control group(P<0.05)after 24 h treatment.Compared with the Mn group,the survival rate of50?150 and 450-PAS in the intervention group increased significantly(P<0.05).Microglia ros inspection: the number of fluorescent cells in the Mn group was higher than that in the control group,and the number of 50?150 and 450-PAS group was less than that of the Mn group.Inflammatory cytokines in the supernatant: compared with the control group,MnCl2 increased the secretion of IL-1? and TNF-? in the supernatant of microglia(P<0.05).The expression of TNF-? in L,M and H-PAS group was lower than that in Mn group(P<0.05).Microglia IL-1?,TNF-?mRNA expression: Compared with the control group,the expression of IL-1 and TNF-?mRNA in Mn group increased(P<0.05).The expression of IL-1?mRNA in microglia of 50?150 and 450-PAS groups was lower than that in Mn group(P<0.05).The expression of TNF-? mRNA in microglia of 150 and 450-PAS group was lower than that in group Mn(P<0.05).Conclusion(1)excessive manganese exposure can cause damage to microglia,lead to the production of reactive oxygen species in the cell,increase the expression of IL-1?and TNF-?mRNA,and increase the secretion of corresponding inflammatory factors,IL-1 beta and TNF-alpha.(2)PAS-Na can interfere with microglia damage induced by manganese,which may be related to the antioxidant and anti-inflammatory effects of PAS-Na.