Multicolor cytoenzymatic evaluation of dipeptidyl peptidase IV (CD26) function in normal and neoplastic human T-lymphocyte populations.
Dipeptidyl peptidase IV (DPP IV), also identified as the glycoprotein CD26, is a transmembrane 110- to 120-kDa serine aminopeptidase involved in immune responses by influencing T-cell costimulation and by cleaving cytokines. Additionally, CD26 is a nonintegrin receptor that contains a binding site for extracellular matrix and other molecules. In order to further define the expression and functional activity of this membrane exopeptidase in human T cells, we developed a nondisruptive, four-color cytofluorogenic assay that utilizes three separate antibodies to cell-surface molecules (e.g., CD4/CD8/CD26 and CD19/CD56/CD26) along with a rhodamine 110-conjugated dipeptide substrate that allows the measurement of DPP IV activity in phenotypically defined cells. We found normal human thymi to have notable differences in time-dependent DPP IV activity among the thymocyte subsets defined by their CD4/CD8 phenotype, with CD4-/CD8- thymocytes containing less DPP IV activity than cells expressing CD4 and/or CD8 (i.e., maturing). CD26 positivity was moderately intense in thymocytes and tended to identify cells with higher DPP IV activity. The four-color technique was also used to examine mature peripheral blood lymphocytes, along with an assortment of leukemias and transformed T-cell lines. These experiments revealed that while DPP IV was consistently evident in normal T cells, neoplastic T cells could vary in their expression patterns. Furthermore, the presence (or intensity) of surface CD26 in some abnormal T cells and certain normal peripheral blood mononuclear cells was separable from the level of DPP IV measured intracellularly. Our results established that multicolor cytofluorographic analysis can be a practical means to measure DPP IV activity in various human cell populations. Furthermore, we found that DPP IV activity could vary in T cells according to their differentiation status and that under certain circumstances surface CD26 expression can be disassociated from the level of measured enzyme (i.e., DPP IV) activity.
Ruiz
P
P
Department of Pathology, University of Miami School of Medicine, Florida 33101, USA. pruiz@mednet.med.miami.edu
Zacharievich
N
N
Shenkin
M
M
D002478:Cells, Cultured; D004151:Dipeptides / Q000378:metabolism; D018819:Dipeptidyl Peptidase 4 / Q000097:blood* / Q000378:metabolism*; D005434:Flow Cytometry / Q000379:methods*; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D007945:Leukemia, Lymphoid / Q000201:enzymology / Q000276:immunology*; D012235:Rhodamines / Q000378:metabolism; D013379:Substrate Specificity; D016176:T-Lymphocyte Subsets / Q000201:enzymology / Q000276:immunology*; D013950:Thymus Gland / Q000166:cytology; D014407:Tumor Cells, Cultured
D004151:Dipeptides; D005456:Fluorescent Dyes; D012235:Rhodamines; C049025:rhodamine 110; D018819:Dipeptidyl Peptidase 4