Effects of extracellular pH on neutrophil superoxide anion production, and chemiluminescence augmented with luminol, lucigenin or DMNH

Abstract

When neutrophils were stimulated with f-met-leu-phe (fMLP) or phorbol myristate acetate (PMA) chemiluminescence (CL) augmented with luminol, lucigenin or 7-dimethylamino-naphthalene-1,2-dicarbonic acid hydrazide (DMNH) showed a strong dependence on the pH of the medium. CL increased in an exponential fashion when pH was increased from 6.5-9.2. All three luminescent intermediaries showed this pH-dependency but luminol was affected most. Unstimulated CL showed a similar increase with pH. Also neutrophil superoxide anion production (in the cytochrome C reduction assay) increased with pH, but not to the same extent as CL. CL induced by xanthine-oxidase plus hypoxanthine increased likewise with pH, and that increase was more pronounced than what was observed from the same reaction in the cytochrome C assay. The lucigenin-amplified neutrophil CL showed the best correlation to the superoxide production from neutrophils. Thus, several effects underlie the variation of CL with pH but the pH dependency of the luminescent intermediaries contribute substantially, which must be considered.