Ceramide stimulates the uptake of neutral red in human neutrophils, monocytes, and lymphocytes

Neutral red is a vital stain known to be accumulated in the lysosomes of ne utrophils and monocytes. It is used mainly to identify and detect the activ ated state of these cells. We have found that the extracellular application of physiological ceramide, i.e., a product of sphingomyelin hydrolysis and a newly defined intracellular second-messenger substance, increased the up take of neutral red in a dose-dependent manner in human neutrophils, monocy tes, and lymphocytes, as demonstrated by flow cytometry. Staurosporine was able to totally block this phenomenon, suggesting the involvement of protei n kinase C in the process. These results indicate that the flow-cytometric analysis of ceramide-induced uptake of neutral red can be a new method for the evaluation of lysosome-related activation processes in both phagocytes and lymphocytes.