Nerve growth factor is preformed in and activates human peripheral blood eosinophils

Background: Recent studies have shown that nerve growth factor (NGF) is produced by and can act on several immune-inflammatory cells. Objectives: The objective of this study was to study the effects of NGF on human peripheral blood eosinophils and assess whether these cells produce and store NGF. Methods: Eosinophils were purified by negative immunoselection (magnetic cell sorting systems, purity 98% to 100%) from 13 subjects (9 to 26 years old) with mild blood eosinophilia, mainly of allergic origin. Eosinophils were incubated with NGF (50 to 1000 ng/mL), and supernatants were collected for measurement of eosinophil peroxidase (EPO, 20 minutes, colorimetric enzymatic assay) and IL-6 (12 hours, ELISA), Eosinophil viability was evaluated by Trypan blue test (days 2, 3, and 4), NGF content in freshly isolated eosinophils, after ultrasound disruption, was determined with a 2-site immunoenzymatic assay. The presence of mRNA for NGF was evaluated by reverse transcription PCR. Results: NGF caused EPO release (highly significant at 1000 ng/mL NGF), IL-6 release from eosinophils was not higher than IL-6 spontaneously released into culture medium alone, NGF did not significantly affect the number of viable eosinophils. NGF was found in the eosinophil sonicates (1.5 to 17.8 pg/mL per 10(6) cells). Similarly, mRNA for NGF,vas detected by reverse transcription PCR in the freshly isolated eosinophils. Conclusions: NGF activates human peripheral blood eosinophils from subjects with mild eosinophilia to selectively release inflammatory mediators. Eosinophils store and produce NGF. Therefore the capability of NGF to induce a secretory response and its production and storage by circulating human eosinophils suggest a possible role for NGF in conditions associated with eosinophilia, including allergic disease.