The Influnce Of Nano-Silicon Dioxide And Benzopyrene Separated Or Combined In TNF-α-TNFR Signal Pathway

Objective:To explore the expression level of TNF-α—TNFR in BEAS-2B, A549 and YTMLC cell lines,which are stimulated by nano-silicon dioxide and benzopyrene alone or combined.Methods:(1)Measure the lethal concentration of 50%(LC50) of BEAS-2B,A549 and YTMLC when stimulated by nano-silicon dioxide and benzopyrene alone or combined; (2) Using nano-silicon dioxide and benzopyrene alone or combined to stimulate the BEAS-2B,A549 and YTMLC cell lines,RIPA method to extract the protein of three cell lines when stimulating happens in12h,24h and 48h.measure the concentration of proteins by Ultra-micro spectrophotometer,draw a standard curve.(3)Using western blot to detect the expression level of Caspase3、Caspase8、 NF-κB、P100/P52、P105/P50、TNF-a of three cell lines in three timings.(4)Using SPSS 20.0 to make statistical analysis.Results:(1)IC50 of nano-SiO2 in BEAS-2B cell line equals to71.400±0.785 mg/L,IC50 of nano-SiO2 in A549 cell line equals to 72.452±0.538 mg/L,IC50 of nano-SiO2 in YTMLC cell line equal to 70.110±0.329 mg/L;IC50 of Benzopyrene in BEAS-2B cell line equals to 7.623±0.443μmol/L,IC50 of Benzopyrene in A549 cell line equals to 7.623±0.726 μmol/L,IC50 of Benzopyrene in YTMLC cell line equal to 8.521±0.811μmol/L;(2)BEAS-2B cell lines:Compared with control group,the expression level of TNF-α increases in Separate groups and combined group when stimulated with 12h,there is no difference between the combined and separate group,at the same time,the expression level of Caspase3, Caspase8, P100 of three cell lines have little difference;when stimulated with 24h,the expression level of TNF-α、 caspase3 and caspase8 in separated groups and combined group are higher than control group.compared with separated group,the expression level of TNF-a,caspase3,P100 and caspase8 increases;when stimulated with 48h,the expression level of TNF-a in combined group are higher than separated group and control group,compared with separated groups and control group.the expression level of caspase3,P100 and caspase8 are much lower in combined group(barely no expression) (3) A549 cell lines:Compared with control group and combined group, when stimulated with 12h,the expression level of TNF-a,P100,caspase8 increases while caspase3 reduced in combined group;when stimulated with 24h,the expression level of TNF-α、caspase3 in combined group are higher than control group and separated group.the expression level of P100 and caspase8 in each group have little difference. when stimulated with 48h,the expression level of P100 and caspase8 in combined group are lower than separated group and control group,the expression level of caspase3 and TNF-a in each group have little difference. (4)YTMLC cell lines:when stimulated with 12h,,the expression level of TNF-a,caspase8 reduces while caspase3 increases in combined group and separated group,the expression level of P100 in each group have little difference;when stimulated with 24h,the expression level of TNF-α、caspase3,caspase8, P100 in combined group are higher than control group and separated group.when stimulated with 48h,the expression level of TNF-a and caspase8 in combined group induced while caspase3 increases,the expression level of P100 increases in separated group and combined group.Conclusion(s):1.Using benzopyrene and nano-dioxide silicon combined is more poisonous than using them separately, it might be the reason that cancerization chance of bronchial epithelial cells increase; 2.Using benzopyrene and nano-dioxide silicon combined would interference the normal expression of the key protein of TNF-a-TNFR in cell,TNF-α-TNFR might be the possible pathway that cancerization happened in bronchial epithelial cells.