Antitumor Effect Of Human PBMC On Bladder Carcinoma Cells Induced By CpG ODN

Background: CpG oligodeoxynucleotides, a fairly new class of reagents that mimic bacterial DNA, have been shown to induce a potent Th1-oriented response that leads to tumor eradication and protective immunity. Moreover, although CpG ODN may initially produce inflammation by way of the innate system, an ensuing adaptive immune response has been demonstrated that leads to immunologic memory and tumor-specific T-cell immunity. In cancer immunotherapy, CpG ODNs have been used as monotherapy, adjuvant for vaccination, and in combination with chemotherapy, radiation treatment, antibody therapy, cellular vaccines. The immune effects of CpG ODNs may be considered in two stages: an early stage of innate immune activation and a later stage of enhancement of adaptive immune responses. Within minutes of exposure of plasmacytoid DCs to CpG, the ODNs appear to enter an endosomal compartment where they interact with TLR9, leading to the activation of cell signaling pathways that culminate in the expression of costimulatory molecules upregulation of the chemokine receptor that causes cell trafficking to the T-cell zone of the lymph nodes, and secretion of Th1-promoting chemokines.Plasmacytoid DCs secrete type I IFN and mature into highly effective antigen-presenting cells. These CpG-induced type I IFNs, cytokines and chemokines trigger a wide range of secondary effects, including NK cell activation and enhanced expression of Fc receptors on effector cells such as polymorphonuclear leukocytes, with a resultant increase in antibodydependent cellular cytotoxicity (ADCC). The study was designed to investigate the function of the effect of CpG ODN on the differentiation, cytokine secreted by PBMC, cytotype of immune cells and anti-bladder-tumor activity of human peripheral blood monocytes. The research aim is to testify the feasibility of CpG ODN as the new immune therapy strategy. Methods: 1. To test lymphocyte proliferation by MTT method after human PBMC was incubated with type A and B CpG ODN. 2.To measure cytokine release by ELISA on human PBMC induced by CpG ODN, including IFN-α, IL-12, IFN-γ and TNF-α. 3. Analysis cell surface marker of human PBMC include NK, B, CD4+ and CD8- T lymphocyte by flow cytometry. Pasmacytoid dendritic cell proportion was measured by flow cytometry yet. 4. To test effect of human PBMC on bladder carcinoma cells induced by CpG ODN by HLA method.Results: 1. In Vitro, CpG ODN could induce human lymphocyte proliferation.OD value of human PBMC, that incubate with high concentration CpG ODN, is twice more than control group.The effect of stimulation is dependent on its dosage. 2. Type A and type B CpG ODN could induced cytokine release of human PBMC include IFN-α, IL-12, IFN-γ and TNF-α. The effect of IFN-α release on type A CpG ODN is high.But there is no difference between control NoCpG ODN group and RPMI1640 group. 3. Expression of CD69 on NK cell, CD4+T and CD8+T cell up-regulated. Expression of CD69 on B cell surface by type A CpG ODN is weaker than by type B CpG ODN. Pasmacytoid dendritic cell proportions were notsignificantly changed among groups. 4. Lysis of human PBMC on bladder carcinoma cells induced by CpG ODN is stronger than NoCpG ODN.Conclusions: CpG ODN could induce Plasmacytoid DCs release cytokines and provide the secong signal which T cell activation needed. Although type A CpG ODN strongly induce pDCs to secrete IFN-α, type B CpG ODNs induce less IFN-α and primarily activate B cells and pDCs, causing them to up-regulate co-stimulatory molecules and secrete Th1 cytokines. CpG ODN could activate cells of the adaptive immune system (lymphocytes) and the innate immune system (NK cells) in a sequence-dependent manner. CpG ODN could induce antitumor effect of human PBMC on bladder carcinoma cells.