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[Effect of benzo [a] pyrene on HSP70 expression in rat cortical neurons in vitro]
1School of Public Health, Shanxi Medical University, Taiyuan 030001, China. q_xuguoyao@163.com
Objective:
To explore the effect of benzo [a] pyrene (B [a] P) on heat shock protein 70 (HSP70) expression in rat cortical neurons in vitro.
Methods:
SD rat cortical neurons were isolated and cultured, then they were exposed 24h with the doses of B [a] P (0, 0.1, 0.5, 1,5 and 10 micromol/L)and exposed 24h, 48h, 72h with doses of B [a] P (0, 0.5 and 5 micromol/L), observing dose-response relationship and time-dependent relationship, respectively. Including the 3% volume of S9 mixture was added in the medium. The neurons viability was detected by MTT assay. Western blot was used to measure the expression of HSP70 and immunofluorescence histochemistry was used to observe the expression and location of HSP70.
Results:
When neurons were exposed 24h with B [a] P, compared with the control group, the expression of HSP70 was no significant difference in the group of 0.1 micromol/L dose of B [a] P (P > 0.05). The expression of HSP70 in groups from 0.5 to 10 micromol/L doses of B [a] P gradually decreased and was significantly reduced compared with 0.1 micromol/L group (P < 0.05). The difference between 10 micromol/L group and control group was significant (P < 0.05). After neurons were exposed 24, 48 and 72h with B [a] P in groups of 0, 0.5 and 5 micromol/L, HSP70 expression at different time-piont in 0.5, 5 micromol/L groups were significantly lower than the control group (P < 0.05). HSP70 expression decreased with time extend in 0.5, 5 micromol/L groups and the difference between time points of 24h and 72h were significantly different (P < 0.05). Immunohistochemistry showed that HSP70 expression was only in the cytoplasm in control group, but fluorescence expressed strongly and expressed in both cytoplasm and nuclear in 0.1 micromol/L group. As the dose increased, the expression of HSP70 in nucleus gradually increased and the fluorescence intensity gradually decreased.
Conclusion:
High doses of metabolites of B [ a] P (0.5 - 10 micromol/L) inhibit the expression of HSP70 in cultured cortical neurons in rat, and there were relationships of dose and time-dependent between the dose and the inhibition. With the B [a] P dose increased HSP70 gradually moved to the nucleus.