EMBRYONIC STEM-CELLS AND EMBRYOID BODIES EXPRESS LYMPHOCYTE COSTIMULATORY MOLECULES

Despite the importance of the costimulatory proteins B7-1 (CD80), B7-2 (CD86), and their counterreceptors CD28 and CTLA-4 (CD154) in the reg ulation of T cell proliferation in the adult immunological system, the initial appearance of these proteins during embryonic development has not been investigated. Using in vitro cultures of undifferentiated mo use embryonic stem (ES) cells and differentiating embryoid bodies as a model of very early embryonic development, we examined these cells fo r the presence of mRNA and protein corresponding to the B7 and CD28 fa milies of costimulatory molecules. By flow cytometry, a stochastically regulated subpopulation of B7-1(+) cells comprising 33% of total cell s was detected in ES cell cultures, while negligible staining was foun d for B7-2, CTLA-4, and CD28. When ES cells were differentiated into e mbryoid bodies for 12 days, a CD45(+) subpopulation of embryoid body c ells were found to stain positively for B7-1, B7-2, and CD28. RT-PCR c onfirmed cell staining data by revealing amplification products corres ponding to B7-1, B7-2, and CD28 in corresponding samples. Very low lev els of CTLA-4 amplification products were found in all samples; howeve r, surface staining of CTLA-4 was never detected. The functional capac ity of ES cell B7-1 to bind its ligand was verified by the ability of the soluble fusion protein CTLA-4 -Ig to bind ES cells and the ability of this reagent to block anti-B7-1 antibody binding in cell based com petition assays. These results demonstrate that expression of costimul atory molecules arises very early during in vitro development and sugg ests that the early embryonic environment may utilize cellular signali ng systems analogous to those seen in the immune system. (C) 1998 Acad emic Press.