Costimulatory function of umbilical cord blood CD14+ and CD34+ derived dendritic cells.
Dendritic cells (DCs) consist of a heterogeneous population of hematopoietic cells characterized by their unique dendritic morphology, their efficient antigen-presenting capability to activate naive CD4(+) and CD8(+) T cells, and their lack of lineage specific markers. Functional properties comparing umbilical cord blood monocyte-derived and umbilical cord blood stem cell-derived DCs have not yet been investigated. CD14(+) monocytes and CD34(+) stem cells were isolated from human umbilical cord blood and were induced to differentiate into dendritic cells using 100 ng/mL granulocyte-macrophage colony stimulating factor (GM-CSF), 25 ng/mL IL-4, 2.5 ng/mL tumor necrosis factor-alpha (TNF-alpha), 100 ng/mL GM-CSF, 25 ng/mL stem cell factor, and 2.5 ng/mL TNF-alpha, respectively. Flow cytometric analysis revealed that the 14-day-old dendritic cells were CD80(+), CD86(+), CD83(+), CD54(+), CD1a(+), CD11b(+), CD11c(+), HLA-DR(+), CD34(-), CD3(-), CD19(-), CD14(-), and CD16(-). Reverse transcription polymerase chain reaction was employed to detect expression of mRNA for CD80 and CD86. Differentiating monocytes initially expressed CD86 while CD80 appeared on day 2. Differentiating stem cells expressed CD80 and CD86 on day 2 of culture. The surface expression of CD80 and CD86 was studied over the course of differentiation. Mixed lymphocyte reaction was employed to evaluate the two types of lineage-derived DCs. Prior to the functional assay, CD14(+) and CD34(+) derived DCs were stimulated for 18 h with 0.1 mg/mL and 1.0 mg/mL E. coli lipopolyssacharide, respectively. Monoclonal antibodies (mabs) to CD80 and CD86 were employed to assess their costimulatory roles. A decrease of stimulation as depicted by decreased T cell activation was significant with mabs to both CD80 and CD86 on monocyte-derived DCs while only mabs to CD86 induced decreased T cell activation by stem cell-derived DCs. The varied functional role of CD80 and CD86 costimulatory molecules is associated with DC differentiation from distinct cord blood isolated hematopoietic lineages. These studies demonstrate that DC association with distinct hematopoietic lineages is of relevance in transplantation and vaccine therapies.
Dilioglou
Smaroula
S
Department of Pathology, University of Mississippi Medical Center, Jackson, MS 39216-4505, USA.
Cruse
Julius M
JM
Lewis
Robert E
RE
D000911:Antibodies, Monoclonal; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D018952:Antigens, CD34 / Q000378:metabolism; D018122:B7-1 Antigen / Q000378:metabolism; D051940:B7-2 Antigen; D002454:Cell Differentiation; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000166:cytology* / Q000276:immunology / Q000378:metabolism*; D005260:Female; D005312:Fetal Blood / Q000166:cytology* / Q000276:immunology / Q000378:metabolism*; D005434:Flow Cytometry; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000378:metabolism; D006801:Humans; D018950:Lipopolysaccharide Receptors / Q000378:metabolism; D007959:Lymphocyte Culture Test, Mixed; D008562:Membrane Glycoproteins / Q000378:metabolism; D009000:Monocytes / Q000166:cytology / Q000378:metabolism; D011247:Pregnancy; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D018952:Antigens, CD34; D018122:B7-1 Antigen; D051940:B7-2 Antigen; C497964:CD86 protein, human; D018950:Lipopolysaccharide Receptors; D008562:Membrane Glycoproteins; D012333:RNA, Messenger