ALTERATIONS IN HUMAN B-CELL CALCIUM HOMEOSTASIS BY POLYCYCLIC AROMATIC-HYDROCARBONS - POSSIBLE ASSOCIATIONS WITH CYTOCHROME-P450 METABOLISMAND INCREASED PROTEIN-TYROSINE PHOSPHORYLATION

Previous studies performed in this laboratory have shown that certain benzo(a)pyrene (BaP) metabolites, such as benzo(a)pyrene-7,8-dihydro d iol (BaP-7,8-diol) and benzo(a)pyrene-7,8-dihydro-diol-9, 10-epoxide ( BPDE), were more effective in elevating intracellular Ca2+ in normal h uman peripheral blood mononuclear cell (HPBMC) T and B cells than was BaP. Additionally, it has been shown that the suppression of human T c ell mitogenesis produced by polycyclic aromatic hydrocarbons (PAHs) an d certain BaP metabolites is reversed by treatment with alpha-naphthof lavone (ANF), a cytochrome P450 1A and 1B inhibitor. ANF also diminish es the elevation in intracellular calcium (Ca2+) produced by BaP in HP BMC. In the present studies, we further defined the relationships betw een intracellular Ca2+ elevation produced by BaP and two immunotoxic P 450-derived metabolites, BaP-7,8-diol and BPDE in the Daudi human B ce ll line. At 1, 4, and 18 h, both BaP-7,8-diol and BPDE produced a sign ificant rise in intracellular Ca2+. This effect, however, was not obse rved with BaP or benzo(e)pyrene (BeP), a nonimmunotoxic PAH. To evalua te the potential role of cytochrome P450 metabolism in PAH-induced Ca2 + elevation, Daudi cells were pretreated with ANF for 4 h, followed by treatment with BaP metabolites for 18 h. ANF completely reversed the rise in Ca2+ produced by BaP-7,8-diol, but had no effect on the Ca2+ e levation produced by BPDE. These results suggest that BPDE may be the ultimate P450 metabolite responsible for Ca2+ elevation in human B cel ls. BaP-7,8-diol and BPDE were found to increase tyrosine phosphorylat ion in Daudi whole cell lysates and to increase tyrosine phosphorylati on of two important Src-related protein tyrosine kinases (PTKs), Lyn a nd Syk. Inhibition of tyrosine phosphorylation by herbimycin A was fou nd to largely prevent the increase in intracellular Ca2+ produced by B aP-7,8-diol and BPDE, suggesting that Ca2+ elevation is coupled to inc reased tyrosine phosphorylation in Daudi. BPDE was found to produce a statistically significant increase in tyrosine phosphorylation of Lyn and Syk within 10 min of exposure. Collectively, these data demonstrat e that certain P350-derived metabolites of BaP may be responsible for PTK activation and an increase intracellular Ca2+, which may alter ant igen receptor signaling in human B cells. (C) 1998 Academic Press.