FIGURE 1.

Effect of up- and down-regulation of CYP2B1 expression on anti-Fx1A-induced H₂O₂ generation and cytotoxicity in GECs. A and B, GECs were infected with the CYP2B1-expressing adenovirus (adv) or an empty adenovirus (−adv), followed by treatment with 4 mg/ml anti-Fx1A, and then incubated with FHS or HIS (5%) for 1 h. Generation of H₂O₂ was measured between 0.5 to 2 h after treatment by the oxidant-sensitive fluorescent dye dichlorodihydrofluorescein diacetate (DCF) (A). Data at each time point in both the adenovirus and empty adenovirus groups represent net complement-mediated generation of H₂O₂ by FHS minus the endogenous generation of H₂O₂ by HIS. Cytotoxicity was determined by LDH release (B). Data in both the adenovirus and empty adenovirus groups represent net complement-mediated percentage of LDH release by FHS minus the endogenous LDH release by HIS. C and D, the CYP2B1 was silenced in GECs infected with CYP2B1 adenovirus for 48 h. GECs were then treated with anti-Fx1A followed by FHS or HIS. A negative siRNA (−ve siRNA) group was included to reflect the specificity of our silencing effect. H₂O₂ generation (C) and cytotoxicity (D) were measured as described. Values are means ± S.E. (n = 3). *, p < 0.05 compared with the empty adenovirus (C) or negative siRNA (D).