Evaluation and optimization of anti-breast cancer effect of dendritic cell vaccine derived from cord blood CD34+hematopoietic stem cells in vitro

WANG Miao; XIE Hui-ping; YAN Ke-qing; CUI Hong-fei; LI Wei; ZHANG Zhi-fei; JIN Yuan-yuan

doi:10.20251/j.cnki.1003-3734.2025.04.008

Abstract：Objective:To optimize the induction and culture method of umbilical cord blood CD34+hematopoietic stem cell derived dendritic cell(DC)in vitro,prepare DC vaccine loaded with tumor cell lysate,and explore the anti-breast cancer activity of DC vaccine.Methods:CD34+hematopoietic stem cells were obtained after umbilical blood mononuclear cells were sorted by immunomagnetic beads.The purity of CD34+hematopoietic stem cells was determined by flow cytometry,and DC was obtained by amplification culture and induced differentiation culture.The influence of different factors added in different combinations and different times on the phenotype of the obtained DC was compared.Flow cytometry was used to detect the antiphagocytotic activity of DC on FITC-OVA,CCK-8 method was used to detect the ability of DC vaccine to stimulate the proliferation of heterologous umbilical blood mononuclear cells,and LDH method was used to detect the in vitro killing activity of T cells activated by DC vaccine loaded with tumor cell lysate antigen on breast cancer cells.Results:During amplification and culture,IMDM medium+GM-CSF(100 ng·mL-1)+SCF(50 ng·mL-1)+Flt-3L(100 ng·mL-1)+TPO(100 ng·mL-1)+10% FBS+1% P/S condition culture group and IMDM medium+GM-CSF(100 ng·mL-1)+SCF(50 ng·mL-1)+10% FBS+1% P/S,there was no significant difference in the phenotype of harvested DCs,but the former group could harvest more DCs.In the induction differentiation stage,the expression of CD80 and CD83 of the harvested DC was higher when TNF-α was added 48 hours after the addition of GM-CSF and IL-4 than at other time.Flow cytometry showed that immature DC had strong antigen-phagocytosis activity.DC vaccine showed a strong ability to stimulate the proliferation of heterologous cord blood mononuclear cells.T cells activated by DC vaccine loaded with tumor cell lysate antigen showed significant killing activity against different breast cancer cell lines in vitro.Conclusion:Cord blood hematopoietic stem cells can differentiate into a considerable number of functional DC,and DC vaccine loaded with tumor cell lysate antigen can activate T cells and have significant killing activity against breast cancer cell line.