In vitro generation of dendritic cells derived from cryopreserved CD34+ cells mobilized into peripheral blood in lymphoma patients.

Abstract

Dendritic cells (DC) are APC that initiate primary T-cell dependent immune responses. They have been shown to be generated from CD34+ cells in BM, placental/umbilical cord blood (CB), and G-CSF mobilized peripheral blood CD34+ cells (PBSC). In recent clinical studies, DC were used as a vaccine for cancer patients and showed induction of their antitumor effects. Cryopreservation of CD34+ cells is important to extend the availability of cellular therapy with DC. However, little is known about the effect of cryopreservation on the functional maturation of DC. PBSC harvested from lymphoma patients mobilized with G-CSF and undergoing leukapheresis were cryopreserved at -135 degrees C for 3 days. Freshly isolated or cryopreserved PBSC were cultured with GM-CSF/SCF/tumor necrosis factor-alpha (TNF-alpha). After 14 days of culture, DC were harvested, washed, and used for phenotypical and functional analysis. Cryopreserved PBSC, as well as freshly-isolated PBSC cultured for 14 days, gave rise to CD1a+ /CD4+ /CD11c+ /CD14low+ /CD25( -)/CD40+ / CD45RO+/CD80+/CD83+/CD86+/HLA-DR+ cells with dendritic morphology. DC derived from cryopreserved PBSC mobilized with G-CSF showed a similar endocytic capacity and chemotactic migratory capacities when compared with DC derived from freshly-isolated G-CSF mobilized PBSC. These DC also exhibited similar capacities in the primary allogeneic T-cell response. These results indicate that cryopreserved G-CSF mobilized PBSC cultured with GM-CSF/SCF/TNF-alpha gave rise to DC that were morphologically, phenotypically and functionally similar to DC derived from fresh G-CSF mobilized PBSC. The observation indicates the clinical usefulness of cryopreserved CD34+ cells from lymphoma patients.