MPO uses endogenous low-molecular–weight/dialyzable substrates in plasma to initiate lipid peroxidation.
(left) LDL (0.2 mg/mL) was incubated with isolated human MPO (30 nM) and an H2O2-generating system composed of G/GO for 12 hours at 37°C in HBSS supplemented with DTPA (100 μM, pH 7.0) as in Figure 3. Where indicated, the filtrate of plasma (Fp), the low-molecular–weight constituents derived from plasma filtered through a 10-kd MWt cutoff filter, was added (50%, vol/vol) to the MPO–LDL reaction mixtures. Isolated MPO, an H2O2-generating system (G/GO) and LDL were added to Fp (Complete System). The content of 9-H(P)ETE formed within endogenous plasma lipids was then determined by LC/ESI/MS/MS as described in “Materials and methods.” (right) Plasma (P) or dialyzed plasma (DP) was incubated with isolated MPO and an H2O2-generating system (G/GO) under conditions similar to those described above. The content of 9-H(P)ETE formed within endogenous plasma lipids was then determined by LC/ESI/MS/MS as described in “Materials and methods.” Where indicated, DP and P were added.
