Anti-CD47 monoclonal antibody (B6H12) impairs the maturation and function of human dendritic cells.
To investigate the influence and mechanisms of CD47 engagement by its soluble mAb B6H12 on the maturation and function of cultured dendritic cells (DCs), monocyte-derived DCs were propagated in granulocyte-macrophage colony stimulating factor (GM-CSF) combined with lipopolysaccharide (LPS) and interleukin (IL)-4, in the presence or absence of soluble anti-CD47 monoclonal antibodies (anti-CD47 mAbs, B6H12). The characteristic morphology of DCs was identified by using the transmission electron microscopy. Flow cytometry was used to detect the cell surface phenotypes. The concentration of IL-12 P70 in supernatant was measured by ELISA. The antigen-presenting functions of DCs were determined in one-way mixed leukocyte reaction by BrdU-ELISA. Electrophoretic mobility shift assay (EMSA) was applied to examine the activity of NF-kappaB. The results indicated that the anti-CD47 mAbs markedly suppressed the expression of CD80, CD86, CD83, CD1a and HLA-DR on the surface of DCs (P < 0.05). The data of the mixed leukocyte reaction and IL-12 P70 production were consistent with the results by flow cytometry (P < 0.01). Pre-exposure to B6H12 mAb during the development of DCs resulted in a dramatic depletion of the DNA binding activity of NF-kappaB toward nucleus protein. Moreover, such an inhibition effect seemed to be dose dependent. In conclusion, the soluble mAb B6H12 inhibits the expression of the costimulatory molecules and MHCII molecules on the DCs. The antigen-presenting function of DCs was also impaired by B6H12. And these modulations are closely related with the depletive DNA binding activity of NF-kappaB. It is suggested that the soluble B6H12 exerts a negative effect on the maturation and function of in vitro cultured DCs due to inhibition of NF-kappaB binding activity.
Yu
Jing
J
Department of Hematology, Center of Bone Marrow Transplantation, The First Affiliated Hospital, Medical College of Zhejiang University, Hangzhou 310003, China.
Lin
Mao-Fang
MF
D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology*; D015703:Antigens, CD / Q000096:biosynthesis; D018949:Antigens, CD1 / Q000096:biosynthesis; D018122:B7-1 Antigen / Q000096:biosynthesis; D051940:B7-2 Antigen / Q000096:biosynthesis; D051928:CD47 Antigen / Q000276:immunology*; D048429:Cell Size / Q000187:drug effects; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006684:HLA-DR Antigens / Q000096:biosynthesis; D006801:Humans; D007136:Immunoglobulins / Q000096:biosynthesis; D018664:Interleukin-12 / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000096:biosynthesis; D016328:NF-kappa B / Q000378:metabolism; D009841:Oligonucleotides / Q000378:metabolism; D011485:Protein Binding; D000097802:CD83 Antigen
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D018949:Antigens, CD1; D018122:B7-1 Antigen; D051940:B7-2 Antigen; D051928:CD47 Antigen; D006684:HLA-DR Antigens; D007136:Immunoglobulins; D018664:Interleukin-12; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D009841:Oligonucleotides; D000097802:CD83 Antigen; C419393:CD1a antigen