Myeloperoxidase-Based Chemiluminescence of Polymorphonuclear Leukocytes and Monocytes

Abstract

Luminol and lucigenin chemiluminescence (CL) responses produced by separated human blood polymorphonuclear leukocytes (pmn) and monocytes (mono) have been studied following stimulation with the surface-receptor agonist fMLP (a synthetic chemotactic peptide) and the protein kinase C activator phorbol myristate acetate (PMA). Pmn produced two- to threefold the luminol CL and superoxide anion (O2-) levels of mono; lucigenin CL was similar for both cell-types. The myeloperoxidase (MPO) inhibitor salicylhydroxamic acid (SHA) abrogated luminol but not lucigenin CL in both cell types, but did not further inhibit the already grossly subnormal luminol CL responses seen with MPO-deficient cells which produced normal lucigenin CL. SHA also profoundly inhibited the luminol CL response in a cell-free MPO-H2O2 system. Mono lucigenin CL does not appear to specifically measure O2- production. These data show that luminol CL provides a useful measure of pmn and also mono MPO activity. However, analysis of the effects of various reactive oxygen species (ROS) scavengers, assessed on phagocyte and cell-free CL systems (both MPO-H2O2 and superoxide generating) suggest that the luminol CL signal is not entirely dependent on MPO activity.

Induction of reactive oxygen intermediates in human monocytes by tumour cells and their role in spontaneous monocyte cytotoxicity

“…Hydrogen peroxide, O 2 -. , and myeloperoxidase are implicated in CL generated by monocytes stimulated with PMA and fMLP (McNally and Bell, 1996). However, in our hands, CL induced by tumour cells was not dependent on hydrogen peroxide generation because catalase had no effect.…”

Enterobacter cloacae leukotoxin: modulation of reactive oxidant species generated by neutrophils

“…Moreover, the rate of ROS generation in the presence of PMA reached a negative slope of CL (À4.0 Â 10 3 ) with 100 mg toxin/mL (Table 3). PMA is considered an important activator of protein kinase C (18,19). However, in the present paper an inhibitory effect of high leukotoxin concentrations was observed on the ROS stimulation produced by PMA.…”

Bioactive flavonoids and saponins from Climacoptera obtusifolia

“…These results are in agreement with the observation of Chen et al (2002), who showed that compound 3 at a high concentration can suppress superoxide generation, induced by phorbol 12-myristate 13-acetate (PMA) and formyl-methionyl-leucyl-phenylalanine (fMLP), in a dose dependant manner. However, the system we employed (serum opsonized luminol dependant-chemiluminescence) can detect varieties of free radicals, such as superoxide, H 2 O 2 , OH, and HOCl (McNally and Bell, 1996).…”