Characterization of the autofluorescence of polymorphonuclear leukocytes, mononuclear leukocytes and cervical epithelial cancer cells for improved spectroscopic discrimination of inflammation from dysplasia

Fluorescence spectroscopy has the potential to improve the in vivo detectio n of intraepithelial neoplasias; however, the presence of inflammation can sometimes result in misclassifications. Inflammation is a common and import ant pathologic condition of epithelial tissues that can exist alone or in c ombination with neoplasia. It has not only been associated with the presenc e of cancer but also with the initiation of cancer by damage induced due to the oxidative activity of inflammatory cells. Microscopic examination of c ervical biopsies has shown increased numbers of polymorphonuclear and monon uclear leukocytes in inflamed tissues mostly confined to the stroma. The pu rpose of this study was to characterize the fluorescence properties of huma n polymorpho- and mononuclear leukocytes and compare their fluorescence to that of cervical cancer cells. Human neutrophils were purified from periphe ral blood and their fluorescence characterized over an excitation range of 250-550 nm, There are four notable excitation emission maxima: the tryptoph an peak at 290 nm excitation, 330 nm emission; the NAD(P)H peak at 350 nm e xcitation, 450 nm emission, the FAD peak at 450 nm excitation, 530 nm emiss ion and an unidentified peak at 500 nm excitation, 530 nm emission. Treatme nt of these peripheral blood neutrophils with 40 nM phorbol myristate aceta te or with the chemotactic peptide formyl-Met-Leu Phe (1 M) demonstrated a significant increase in NAD(P)H fluorescence. Isolated mononuclear cells ha ve similar emission peaks for tryptophan and NAD(P)H and a small broad peak at 450 nm excitation, 530 nm emission suggestive of FAD, Comparison of the fluorescence from leukocytes to epithelial cancer cell fluorescence has de monstrated the presence of these fluorophores in different quantities per c ell. The most notable difference is the high level of tryptophan in cervica l epithelial cancer cells, thus offering the potential for discrimination o f inflammation.