Roles Of ATM/Chk2-p53 Signaling Pathway In B[a]p-induced Neuronal Apoptosis

Objective:To study the mechanism of ATM/Chk2-p53 signal pathway in B[a]p-induced brain cortical neuronal apoptosis sfter exposure to B[a]p in SD rats. Methods:One hundred and fifty male SD rats were randomly sorted based on weights and assigned to five groups. Five dogage groups(0,1.0,2.5,6.25mg/kg.bw) rats were received intraperitoneal injection every other day for 1,2 and 3 months respectively. Three rats were fixed under heart perfusion every group and brains were cut for histologic examination. The rest of rats were cut under anaesthetization and cortex was taken for next experiment. The expression levels of ATM,ATMser794,Chk2, Chk2thr68, P53, P53ser20 and Cleaved Caspase3 in cortex were test by West-blotting.Cortical neuronal apoptosis were detected by TUNEL; BPDE-DNA adducts in rats cortex were detected by Immunohistochemistry. Results:1After B[a]P exposure,cortical neuronal apoptosis rates were significantly increased.Apoptosis rates depends on infected time and infected dose. When rats were exposed to 2.5mg/kg B[a]P for 1,2 and 3 months, cortical neuronal apoptosis rates were 1.59、1.93 and 2.58 folds of control groups(P<0.05). As exposed B[a]P dose at 6.25mg/kg for 1,2 and 3months cortical neuronal apoptosis rates were 1.85、2.69 and 4.75 folds of control groups(P<0.05). The apoptosis rate is 23.49% after B[a]P exposed 3mouths(P<0.05). The interactions of dose and time on the indexes of cortical neuronal apoptosis rates were statistically significant(F=13.923,P<0.001) with the increase of exposure time and dose.2 With the increase of exposure time and dose, BPDE-DNA adducts in cortex were raised. When rats were exposed to 6.25mg/kg B[a]P for 3month,BPDE-DNA adducts increased 161.5% compared with the control(P<0.05).The interactions of dose and time on the indexes of BPDE-DNA adducts were statistically significant(F=2.731,P=0.022) with the increase of exposure time and dose.3 As exposed B[a]P dose at 1.0, 2.5 and 6.25mg/kg for 3months ATM protein expression were increased 20.7%, 17.2% and 28.7%(P<0.05)of control groups(P<0.05). As exposed B[a]P dose at 1.0mg/kg B[a]P for 1,2 and 3months ATMser794 protein expression wereincreased11.5% 、34.7% and 35.0%(P<0.05) of control groups.When rats were exposed to 2.5mg/kg B[a]P for 1,2 and 3 months, ATMser794 protein expression wereincreased 31.9%、17.9% and 45.5%(P<0.05) of control groups. As exposed B[a]P dose at 6.25mg/kg B[a]P for 1,2 and 3months ATMser794 protein expression wereincreased 46.4% 、 34.6% and 62.3%(P<0.05) of control groups.As exposed B[a]P dose at 1.0mg/kg B[a]P for 2 and 3months Chk2 protein expression wereincreased 63.5% and 61.6%(P<0.05) of control groups.When rats were exposed to 2.5mg/kg B[a]P for 2 and 3 months, Chk2 protein expression wereincreased60.0% and 87.7%( P<0.05) of control groups. As exposed B[a]P dose at 6.25mg/kg B[a]P for 1,2 and 3months Chk2 protein expression wereincreased77.3%、85.8% and 165.8%(P<0.05) of control groups.When rats were exposed to 6.25mg/kg B[a]P for 2 and 3 months, Chk2Thr68 protein expression wereincreased 92.2%, 112.3%(P<0.05) of control groups.As exposed B[a]P dose at 1.0mg/kg B[a]P for 2 and 3months P53 protein expression wereincreased 35.9% and 117.8%(P<0.05) of control groups.When rats were exposed to 2.5mg/kg B[a]P for1, 2 and 3 months, P53 protein expression wereincreased 83.3%、92.1% and 158.9%(P<0.05) of control groups. As exposed B[a]P dose at 6.25mg/kg B[a]P for 1,2 and 3months P53 protein expression wereincreased108.3% 、150.0% and 255.3%(P<0.05) of control groups.As exposed B[a]P dose at 6.25mg/kg B[a]P for 1,2 and 3months P53ser20 protein expression wereincreased 31.7%、92.4% and 173.4%(P<0.05) of control groups.4 When rats were exposed to 2.5mg/kg B[a]P for1, 2 and 3 months, Cleaved Caspase3 protein expression wereincreased 49.4%、61.3% and 77.7%(P<0.05) of control groups. As exposed B[a]P dose at 6.25mg/kg B[a]P for 1,2 and 3months Cleaved Caspase3 protein expression wereincreased 55.9%、94.3% and 111.1%(P<0.05) of control groups.As exposed B[a]P dose at 1.0mg/kg B[a]P for 1,2 and 3months P53 protein expression wereincreased 43.5%、30.3% and 69.5%(P<0.05) of control groups.When rats were exposed to 2.5mg/kg B[a]P for1, 2 and 3 months, Pu ma protein expression wereincreased 77.4%、87.7% and 144.1%(P<0.05) of control groups. As exposed B[a]P dose at 6.25mg/kg B[a]P for 1,2 and 3months P53 protein expression wereincreased 93.5%、137.8% and 208.5%(P<0.05) of control groups. Conclusions:B[a]P can induce cortical neuronal apoptosis and DNA damage in SD rats. The damage is more serious with infected time high and infected dose increase. The ATM/Chk2-p53 signle pathway protein and Cleaved Caspase3、puma protein expression wereincreased after B[a]P exposure.ATM/Chk2-p53 and P53 pathway may be the key factor by DNA damage in B[a]P-induced cortical neuronal apoptosis.