title: Activated platelets rapidly up-regulate CD40L expression and can effectively mature and activate autologous ex vivo differentiated DC.
abstract: Background DC are a promising immunotherapeutic for treatment of infectious/malignant disease. For clinical trials, immature DC generated from precursor cells such as monocytes, using serum-free media containing GM-CSF and IL-4, can be matured with specific cytokines/factors. CD40 ligand (CD40L) plays an important role in DC activation/maturation but is not available for clinical applications. These studies evaluated the feasibility of using activated platelets with elevated CD40L surface expression to stimulate autologous DC maturation. Methods Pilot and clinical scale studies were executed using magnetic/centrifugal separation. Monocyte precursors were differentiated to immature DC with GM-CSF and IL-4 and the ability of activated autologous platelets to mature these cells was evaluated on the basis of phenotype and function. Results In small-scale studies, DC cultures stimulated with activated autologous platelets (CD40L-AP), tumor necrosis factor-alpha (TNF-alpha) or soluble CD40L (sCD40L) up-regulated expression of phenotype markers indicative of activation and maturation. CD86 expression was significantly enhanced (P<0.05) by stimulation with either CD40L-AP (75.5+/-14.5%) or sCD40L (80.5%+/-5.3%) compared with immature DC (55.2+/-14.8%), as were CD80 and CD83. Similarly, in large-scale studies using Isolex 300I to enrich for monocytes and platelets for DC generation/maturation on a clinical scale, stimulation with CD40L-AP increased CD86 and CD80 expression as well as the ability to stimulate allogeneic lymphocytes compared with control cultures. Discussion These results demonstrate that thrombin-activated platelets express CD40L and are effective at inducing matured DC with potent immunogenic activity. Furthermore, these studies demonstrate the feasibility of this approach for clinical immunotherapeutic interventions.
authors.affiliation: Section of Bone Marrow Transplant and Cell Therapy, Rush Presbyterian-St. Luke's Medical Center, Rush Medical School, Chicago, IL, USA.
authors.forename: Ja
authors.initials: J
authors.lastname: Martinson
authors.forename: J
authors.initials: J
authors.lastname: Bae
authors.forename: H-G
authors.initials: HG
authors.lastname: Klingemann
authors.forename: Yk
authors.initials: Y
authors.lastname: Tam
mesh_terms: D015703:Antigens, CD / Q000378:metabolism; D001324:Autoantigens / Q000378:metabolism; D018122:B7-1 Antigen / Q000378:metabolism; D051940:B7-2 Antigen; D001792:Blood Platelets / Q000502:physiology*; D023201:CD40 Ligand / Q000378:metabolism*; D002454:Cell Differentiation; D003713:Dendritic Cells / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D007136:Immunoglobulins / Q000378:metabolism; D018950:Lipopolysaccharide Receptors / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism; D009000:Monocytes / Q000276:immunology / Q000378:metabolism; D015539:Platelet Activation; D013917:Thrombin / Q000494:pharmacology; D015854:Up-Regulation; D000097802:CD83 Antigen
chemical_list: D015703:Antigens, CD; D001324:Autoantigens; D018122:B7-1 Antigen; D051940:B7-2 Antigen; D023201:CD40 Ligand; D007136:Immunoglobulins; D018950:Lipopolysaccharide Receptors; D008562:Membrane Glycoproteins; D013917:Thrombin; D000097802:CD83 Antigen; C497964:CD86 protein, human