Priming Effect On Murine Dendritic Cells Of Complex Of Antigen Of Elemene-combo Tumor Cell Vaccine And Heat Shock Protein 70 Of BCG (H_TA-HSP70_BCG)

Heat shock protein (HSPs) are the most important of molecular chaperones. HSPs are engaged in processing and presenting of antigens. Exogeneous HSPs bound with CD91 of dendritic cells (DCs) and induced maturation of DCs. In tumor cells HSPs bound with mutant or injuried proteins /peptides to form HSP-peptide complexes. HSP-peptide complexes extracted from tumor cells could induce immunity to the tumor and might be used as tumor vaccines. Their application in treatment of human cancer was restricted by insufficient amount. Dendritic cell was an antigen presenting cell(APC) with potent function. DCs loaded with tumor antigens might be used as tumor vaccine to induce immune responses to tumor. Researche on antigen carriers which conveyed tumor antigen to and primed DCs was an impotent DC field.Objectives: Our previous reports showed that immunization with complex of rumor antigens of Hca-F elemene combo -tumor cell vaccine and HSP70 of BCG (HTA-HSP70BCG ) could induce potent immune protective effects against Hca-F in mice. In this experiment normal murine DCs were primed with HTA-HSP70BCG.Their proliferation and antigen presenting function were evaluated.Methods: The dendritic cells were isolated and grown from spleen or bone marrow of 615 inbred mice using complete RPMI 1640 media supplemented with GM-CSF(100ng/ml) and IL-4(100ng/ml). 18-20 hours later, DCs were pulsed with HTA-HSP70BCG,HTA or HSP70BcG for 3 days. The cells were harvested. Their proliferation and stimulating effects on spleen nonadherent cells were evaluated with MTT assay. Their capability of endocytosing FITC labled dextran was assayed with FACscan, Morphological changes of DC were observed in electron microscope.Results: 1. Priming effect of HtA-HSPTOgcc on normal murine DCs. DCs pulsed with HTA-HSP70BCG, HTA or HSP70BcG proliferated in the presense of GM-CSF and IL-4, proliferation index of spleen DCs was 2.119±0.121, 1.135±0.130 and 1.269±0.029, respectively(P < 0.05); The index of bone marrow DCs was 2.107±0.013, 1.126±0.127 and 1.271±0.014,respectively(P < 0.05). 2. Morphological changes of DC pulsed with HTA-HSP70BCG: There were much dendric protrusionsand richer mitochondrions in DC pulsed with HTA-HSP70BCC. 3. The activating effect of DCs pulsed with HTA-HSP70BCG on normal nonadherent spleen cells. DCs pulsed with HTA-HSP70BCG,HTA or HSP70BCG were mixed with normal nonadherent spleen cells of 615 mice and cultured in presence of GM-CSF and EL-4.Cells stimulated with pulsed DCs showed strong proliferation than cells stimulated not pulsed DCs. The proliferation index of spleen DCs plus nonadherent spleen cells was 2.004±0.018, 1.137±0.069 and 1.285±0.014, respectively. The index of bone marrow DCs plus nonadherent spleen cells was 1.927±0.073, 1.311±0.013 and 1.162±0.036, respectively(P < 0.05). 4. Capability of endocytosis of FITC tabled dextran. The results showed that DCs pulsed with HTA-HSP70BCG had more potent capability to uptake antigens. DCs pulsed with HTA-HSP70BCG had more potent capacity to endocytose FITC labled dextran than DCs pulsed with HTA and HSP70BCG.The percent of DCs endocytosed FITC labled dextran of spleen DCs was 55.2%, 0.54% and 0.25%, the percent of bone marrow DCs was 58.61 %, 0.25%, and 0.25%.Conclusion: The results showed that HTA-HSP70BCG could priming DCs from spleen and bone marrow, DCs pulsed with HTA-HSP70BcG had more potent capability to stimulate nonadherent spleen cells and uptake antigens than DCs pulsed with HTA and HSP70BCG. HTA-HSP70BCG might be used to induce immune responses to tumor cells as a tumor vaccine.