The Mechanism Of Beryllium Sulfate To Human Embryonic Lung Fibroblast Apoptosis

Objective With different concentrations of beryllium sulfate (BeSO4) infected people invitro embryo lung fibroblasts (MRC-5), observe the change of the human embryonic lungfibroblast apoptosis, the Bcl-2, Bax apoptosis related gene, Caspase-3mRNA expression level,inositol1,4,5-triphosphate receptor (IP3R) in cells and inositol1,4,5-triphos-phate (IP3) the change of the content and intracellular calcium ion concentration change,discuss whether BeSO4through IP3R Ca2+channel affect cell apoptosis, and observe IP3Rantagonists and activator intervention change posterity embryo lung cells apoptosis.Methods MRC-5cells cultured in vitro were randomly divided into blank control group,different dose BeSO4infected group, BeSO4+2-APB group and BeSO4+IP3group. Finalconcentration of BeSO4is1、10、100μmol/L，2-APB group and IP3dose of10μmol/L+10μmol/L BeSO4. Cell growth to logarithmic phase, after reaching cell density, according to theexperimental requirements, medicaments respectively develop collecting cells after24h and48h.The following experiments:1. Flow cytometry to detect cell apoptosis rate;2. Real-timePCR detect IP3R Ⅲ, BCL-2, bax, Caspase3mRNA expression；3. ELISA determination ofintracellular IP3R Ⅲ and IP3protein expression;4. Confocal laser scanning microscope todetect intracellular Ca2+.Results1.24h and48h BeSO4low, medium and high dose group compared with the sametime blank control group, MRC-5cell apoptosis rate decline (P<0.01); after BeSO4low,medium and high dose group of infected24h and48h, compared with the blank control group,Bcl-2mRNA expression is rose, Bax and Caspase3mRNA expression is reduced, the Bcl-2/Bax ratio increased (P<0.05, P<0.01); 2. After BeSO4low, medium and high dose group of infected24h and48h, compared with theblank control group, IP3RⅢ mRNA expression is rose(P<0.05，P<0.01)；but the results ofELISA did not find the IP3RⅢ protein increased corresponding(P>0.05); IP3has not beenfound increased, after beryllium sulfate infected MRC-5cells(P>0.05);3.24h BeSO4high dose group compared with blank control group, Ca2+content increased(P<0.05);48h BeSO4middle,high dose group compared with blank control group, Ca2+weredecreased (P<0.05);4. Antagonists and activator intervention effects:24h and48h BeSO4+2-APB group comparedwith the same time BeSO4medium dose group, the MRC-5cell apoptosis rate decline(P<0.05), and lower than the blank control group (P<0.01)；24h and48h BeSO4+IP3groupcompared with the same time BeSO4medium dose group, the MRC-5cell apoptosis rateincreases (P<0.05), but they are still lower than in blank control group (P<0.01);24h and48hBeSO4+IP3group compared with the corresponding phase BeSO4medium dose group ofMRC-5cells IP3concentration increases(P<0.01);24h and48h BeSO4+2-APB groupcompared with the corresponding phase BeSO4medium dose group of Ca2+were decreased(P<0.01), and lower than he corresponding phase blank control group(P<0.05, P<0.01);24hand48h BeSO4+IP3group compared with the corresponding phase BeSO4medium dosegroup of Ca2+were increased (P<0.05, P<0.01).Conclution BeSO4can lead to abnormal human embryonic lung fibroblast apoptosisoccurred, which is possibly related to human embryonic lung fibroblasts of the Bcl-2, Bax,Caspase-3mRNA expression quantity exception and abnormal changes of intracellular Ca2+concentration, by endoplasmic reticulum IP3R Ca2+channel regulation intracellular Ca2+concentration after BeSO4infected MRC-5，the activator of IP3R has a certain improvementeffect for BeSO4apoptosis inhibition.