Identification of four subsets of human CD3- CD16+ Natural Killer (NK) cells by the expression of clonally distributed functional surface molecules: Correlation between subset assignment of NK clones and ability to mediate specific alloantigen recognition

Alessandro Moretta, Cristina Bottino, Daniela Pende, Gino Tripodi, Giuseppe Tambussi, Oriane Viale, Annamaria Orengo, Maurizio Barbaresi, Annalisa Merli, Ermanno Ciccone, Lorenzo Moretta

Research output: Contribution to journalArticle

Abstract

In previous studies we identified a surface molecule (termed GL183) capable of mediating cell activation and selectively expressed by a subset of human CD3- CD16+ natural killer (NK) cells. In this study we analyzed whether other subset-specific functional molecules were expressed in GL183- NK cells. To this end, mice were immunized with the PE29 (CD3-CD16+GL183-) NK clone. Monoclonal antibodies (mAbs) were selected by screening the hybridoma supernatants for their ability to trigger the cytolytic activity of clone PE29 against the human myelomonocytic leukemia U937. The EB6 mAb (IgG1) triggered the PE29 clone, but not a GL183+ clone used as a control. EB6+ cells ranged between 1 and 13% of peripheral blood lymphocytes and were largely included in the CD3-CD16+CD56+ cell populations (only + cells were CD3+). Analysis of resting or activated CD3-CD16+ populations, or clones for the expression of EB6 or GL183 mAbs, allowed us to identify four distinct, phenotypically stable, NK subsets (EB6+GL183-; EB6+GL183+; EB6-GL183-; EB6-GL183-). Similar to GL183 mAb, the EB6 mAb selectively triggered the NK subset expressing the corresponding surface antigen to lyse human tumor cell lines including U937, IGROV-I, M14, and A549. In addition, EB6 mAb sharply inhibited the cytolytic activity of EB6+ clones against P815, M12, and P3U1 murine target cells. In EB6+GL183+ ("double-positive") clones both EB6 and GL183 mAb inhibited the redirected killing of P815 cells induced by anti-CD16, anti-CD2 mAbs and phytohemagglutinin (PHA). Similar to GL183 molecules, molecules precipitated by EB6 mAb were represented by either single 58-kD chain or double chains of 55 and 58 kD (with no detectable differences in EB6+GL183- or EB6+GL183+ clones). In sequential immunoprecipitation experiments using the double-positive clones CEG52 and CA25.50, preclearing of cell lysates with EB6 or GL183 mAb removed only EB6 or GL183 molecules, respectively, thus indicating that the two antigenic determinants are carried by two distinct molecules. Peptide map analysis indicated that EB6 (or GL183) molecules precipitated from double-positive clones were identical to the corresponding molecules isolated from single-positive ones. On the other hand, comparison of the EB6 and GL183 maps revealed peptides that were unique to each molecule, although most of the major peptides migrated to identical positions. We further investigated whether correlation existed between the phenotypic assignment of NK clones and their ability to mediate specific lysis of normal allogeneic cells. All mixed lymphocyte culture-derived NK clones displaying the previously defined "1 anti-A" specificity expressed the EB6+GL183- surface phenotype. In addition, among peripheral blood-derived CD3-CD16+ clones (with PHA and IL-2), only those expressing the EB6+GL183- phenotype specifically lysed PHA blasts susceptible to lysis by "1 anti-A" clones. Finally, in individuals susceptible to lysis by "1 anti-A" alloreactive clones, EB6+ GL183 NK cells (which would represent autoreactive cells) were virtually absent.

Original languageEnglish
Pages (from-to)1589-1598
Number of pages10
JournalJournal of Experimental Medicine
Volume172
Issue number6
Publication statusPublished - Dec 1 1990

ASJC Scopus subject areas

  • Immunology

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