Generation of dendritic cells expressing bcr-abl from CD34-positive chronic myeloid leukemia precursor cells.

Patients with a relapse of chronic myeloid leukemia (CML) after allogeneic bone marrow transplantation can be successfully treated with blood mononuclear cells from the original bone marrow donor. However, the antileukemic effect of this treatment is often accompanied by graft-versus-host disease (GVHD). Treatment with cytotoxic T-lymphocyte (CTL) lines or clones that are specifically generated against leukemic antigen-presenting cells from the patient, may separate antileukemic effects from GVHD. In this report we demonstrate that after culturing CD34-positive cells purified from bone marrow of patients with chronic phase CML in medium containing human serum, GM-CSF, TNF alpha, and IL-4 up to 28% of the cultured cells were dendritic cells, characterized by morphology, phenotypic analysis, and their efficient capacity to stimulate allogeneic T lymphocytes. The expression of HLA and costimulatory molecules and the stimulatory capacity of the dendritic cell-enriched cell suspensions were optimal between days 7 and 10 after onset of the cultures. Fluorescence in situ hybridization revealed that all cultured dendritic cells contained the CML specific t(9;22) translocation. PCR analysis showed expression of the translocation specific bcr-abl mRNA. These leukemic dendritic cells may enhance the induction and proliferation of CTL lines and clones with more specificity for the leukemic cells.

Antigen capture and major histocompatibility class II compartments of freshly isolated and cultured human blood dendritic cells.

Dendritic cells (DC) represent potent antigen-presenting cells for the induction of T cell-dependent immune responses. Previous work on antigen uptake and presentation by human DC is based largely on studies of blood DC that have been cultured for various periods of time before analysis. These cultured cells may therefore have undergone a maturation process from precursors that have different capacities for antigen capture and presentation. We have now used immunoelectron microscopy and antigen presentation assays to compare freshly isolated DC (f-DC) and cultured DC (c-DC). f-DC display a round appearance, whereas c-DC display characteristic long processes. c-DC express much more cell surface major histocompatibility complex (MHC) class II than f-DC. The uptake of colloidal gold-labeled bovine serum albumin (BSA), however, is greater in f-DC, as is the presentation of 65-kD heat shock protein to T cell clones. The most striking discovery is that the majority of MHC class II molecules in both f-DC and c-DC occur in intracellular vacuoles with a complex shape (multivesicular and multilaminar). These MHC class II enriched compartments (MIIC) represent the site to which BSA is transported within 30 min. Although MIIC appear as more dense structures with less MHC class II molecules in f-DC than c-DC, the marker characteristics are very similar. The MIIC in both types of DC are acidic, contain invariant chain, and express the recently described HLA-DM molecule that can contribute to antigen presentation. CD19+ peripheral blood B cells have fewer MIIC and surface MHC class II expression than DCs, while monocytes had low levels of MIIC and surface MHC class II. These results demonstrate in dendritic cells the elaborate development of MIIC expressing several of the components that are required for efficient antigen presentation.