Identification of an alveolar macrophage-derived activity in bronchial asthma that enhances leukotriene C4 generation by human eosinophils stimulated by ionophore A23187 as a granulocyte-macrophage colony-stimulating factor

The American Review of Respiratory Disease
C J HowellT H Lee

Abstract

Incubation of eosinophils (EOS) with alveolar macrophage (AM) supernatants isolated from asthmatic subjects followed by stimulation with the calcium ionophore A23187 resulted in enhancement of the capacity of EOS to elaborate leukotriene C4 (LTC4) (mean enhancement 169 +/- 37%, n = 31). Pretreatment of EOS with AM supernatants derived from normal individuals did not enhance LTC4 generation as compared with control medium. Enhancement was maximal when EOS were preincubated with a 1:6 dilution of AM supernatants for 5 min at 37 degrees C and were then stimulated with 5 microM A23187 for 15 min. Separation of AM supernatants by size-exclusion HPLC using a TSK G3000 SW column resulted in a peak of enhancing activity with an estimated molecular mass of approximately 30,000 D. Further purification by anion exchange HPLC using a TSK DEAE 5PW column (pH 7.4) resolved the activity into a minor peak at 0.17 M NaCl and a major peak at 0.2 M NaCl. The activities were distinct from interleukin-1 and tumor necrosis factor. Resolution of the major peak of activity by reverse-phase HPLC using a C18 spherisorb ODS column and a slope gradient of 0 to 100% acetonitrile/0.1% trifluoroacetic acid demonstrated a single peak of activity that eluted a...Continue Reading

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