title: Cryopreservation of adenovirus-transfected dendritic cells (DCs) for clinical use.
abstract: In this study, we examined the effects of cryoprotectant, freezing and thawing, and adenovirus (Adv) transduction on the viability, transgene expression, phenotype, and function of human dendritic cells (DCs). DCs were differentiated from cultured peripheral blood (PB) monocytes following Elutra isolation using granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4) for 6 days and then transduced using an Adv vector with an IL-12 transgene. Fresh, cryopreserved, and thawed transduced immature DCs were examined for their: 1) cellular concentration and viability; 2) antigenicity using an allogeneic mixed lymphocyte reaction (MLR); 3) phenotype (HLA-DR and CD11c) and activation (CD83); and 4) transgene expression based on IL-12 secretion. Stability studies revealed that transduced DCs could be held in cryoprotectant for as long as 75 min at 2-8°C prior to freezing with little effect on their viability and cellularity. Further, cryopreservation, storage, and thawing reduced the viability of the transduced DCs by an average of 7.7%; and had no significant impact on DC phenotype and activation. In summary, cryopreservation, storage, and thawing had no significant effect on DC viability, function, and transgene expression by Adv-transduced DCs.
authors.affiliation: Department of Medical Microbiology School of Health, Namik Kemal University, Takirdağ, Turkey. dumrulgulen@yahoo.com
authors.forename: D
authors.initials: D
authors.lastname: Gülen
authors.forename: S
authors.initials: S
authors.lastname: Maas
authors.forename: H
authors.initials: H
authors.lastname: Julius
authors.forename: P
authors.initials: P
authors.lastname: Warkentin
authors.forename: H
authors.initials: H
authors.lastname: Britton
authors.forename: I
authors.initials: I
authors.lastname: Younos
authors.forename: J
authors.initials: J
authors.lastname: Senesac
authors.forename: Samuel M
authors.initials: SM
authors.lastname: Pirruccello
authors.forename: J E
authors.initials: JE
authors.lastname: Talmadge
mesh_terms: D000256:Adenoviridae / Q000235:genetics*; D001709:Biotechnology; D002470:Cell Survival; D002478:Cells, Cultured; D015925:Cryopreservation; D003451:Cryoprotective Agents / Q000494:pharmacology; D003470:Culture Media; D003713:Dendritic Cells / Q000166:cytology* / Q000276:immunology / Q000378:metabolism; D005434:Flow Cytometry; D005822:Genetic Vectors; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000502:physiology; D006801:Humans; D007167:Immunotherapy; D018664:Interleukin-12 / Q000235:genetics; D015847:Interleukin-4 / Q000502:physiology; D014162:Transfection; D019076:Transgenes
chemical_list: D003451:Cryoprotective Agents; D003470:Culture Media; D018664:Interleukin-12; D015847:Interleukin-4; D016178:Granulocyte-Macrophage Colony-Stimulating Factor