ABSTRACT
BACKGROUND: Immature dendritic cells have strong antigen uptake and processing ability, but the lack of a variety of costimulatory molecules cannot activate and proliferate initial T cells to produce immune response, which can lead to T cell anergy, thus inducing low immune response or antigen immune specific tolerance. Simultaneously, immature dendritic cells can induce hyporeactivity of allogeneic antigen-specific T cells, thus prolonging the survival time of transplanted organs. OBJECTIVE: To investigate the effect of immature dendritic cells on liver rejection after orthotopic liver transplantation in rats and its mechanism. METHODS: According to the weight of DA and Lewis rats, the rats were randomly divided into three groups, and the liver transplantation model of DA-Lewis rats was established by “two-cuff” method. The rats of control group received no measures. The rats of cyclosporine group were treated with 10 mg/kg cyclosporine from the second day after operation, once a day, for 7 days. The rats of the immature dendritic cell group were injected with 1×106 immature dendritic cells from bone marrow of DA rats one day before operation through dorsal penile vein; the injection was repeated twice with an interval of 10 minutes. The livers of all these rats were removed 7 days after operation. Hematoxylin-eosin staining was used to observe the pathological changes. The mRNA and protein expressions of SHIP, AKT, IKK and IKβ in these three groups were detected by qRT-PCR and western blot assay. RESULTS AND CONCLUSION: (1) Compared with the control group, the survival time of cyclosporine group and immature dendritic cell group was significantly longer (P < 0.05). (2) In cyclosporine group and immature dendritic cell group, the number of infiltrating mononuclear cells and lymphocytes in the portal area of liver tissue was less, the structure of hepatic lobule was not significantly damaged, and the inflammatory cells in hepatic artery, portal vein and bile duct were significantly less than those in control group, which did not reach the level of severe acute rejection. (3) Compared with the control group, the mRNA expression of IKβ in the cyclosporine group and immature dendritic cell group was increased, while the mRNA expression of SHIP, AKT and IKK significantly decreased (P < 0.05). (4) Compared with the control group, the expression of SHIP and IKβ protein significantly increased, IKK and AKT protein significantly decreased in the immature dendritic cell group (P < 0.05). Compared with the control group, the expression of SHIP and IKK protein significantly decreased, AKT and IKβ protein expression significantly increased in the cyclosporine group (P < 0.05). (5) Results confirm that immature dendritic cells can slow down the severe acute rejection, delay the survival time of liver and reduce the T cell immune response ability of allogeneic liver transplantation.
ABSTRACT
PURPOSE: The purpose of this study was to investigate the efficacy of stereotactic body radiation therapy (SBRT) as a tumor-associated antigen (TAA) presentation method for dendritic cell (DC) sensitization and evaluate its effect in combination with immunotherapy using an intratumoral injection of immature DCs (iDCs). MATERIALS AND METHODS: CT-26 colon carcinoma cell was used as a cancer cell line. Annexin V staining and phagocytosis assays were performed to determine the appropriate radiation dose and incubation time to generate TAAs. BALB/c mice were used for in vivo experiments. Cancer cells were injected into the right legs and left flanks to generate primary and metastatic tumors, respectively. The mice were subjected to radiation therapy (RT) alone, intradermal injection of electroporated DCs alone, or RT in combination with iDC intratumoral injection (RT/iDC). Tumor growth measurement and survival rate analysis were performed. Enzyme-linked immunospot and cytotoxicity assays were performed to observe the effect of different treatments on the immune system. RESULTS: Annexin V staining and phagocytosis assays showed that 15 Gy radiation dose and 48 hours of incubation was appropriate for subsequent experiments. Maximum DC sensitization and T-cell stimulation was observed with RT as compared to other TAA preparation methods. In vivo assays revealed statistically significant delay in the growth of both primary and metastatic tumors in the RT/iDC group. The overall survival rate was the highest in the RT/iDC group. CONCLUSION: The combination of SBRT and iDC vaccination may enhance treatment effects. Clinical trials and further studies are warranted in the future.
Subject(s)
Animals , Mice , Annexin A5 , Cell Line , Colon , Dendritic Cells , Immune System , Immunotherapy , Injections, Intradermal , Leg , Methods , Phagocytosis , Radiation Dosage , Radiosurgery , Survival Rate , T-Lymphocytes , VaccinationABSTRACT
Objective:To explore the role of γδ T cells in the transdifferentiation of immature dendritic cells(imDC) into osteoclasts(OC). Methods:(1) Peripheral blood mononuclear cells(PBMNC) were cultured with zoledronate(Zol) and recombinant human interleukin-2(IL-2),and PBMNC from healthy volunteers were cultured with granulocyte macrophage colony-stimulating factor (GM-CSF) and recombinant human interleukin-4(IL-4) to differentiate into imDC,which were then cultured with receptor activator nuclear factor к B ligand(RANKL) and macrophage colony-stimulating factor(M-CSF) to differentiate into OC. The purity of γδ T cells,and phenotype changing of OC transdifferentiated from imDC were investigated by flow cytometry. (2) Co-culture system was es-tablished using millicell inserts.γδT cells isolated with immune magnetic bead were placed in the upper compartment and imDC in the lower compartment in the ratio of 10∶1. To explore the role of γδ T cells during differentiation of imDC into OC,tartrate resistant acid phosphatase( TRAP) staining and bone resorption observation staining were used. Tumor necrosis factor-alpha( TNF-α) of supernatant liquid from different cultures was measured using ELISA(Enzyme linked immunosorbent assay) kit. Results:(1) γδT cells can be ex-panded from PBMNC of MM patients, and the production capacity was similar to that of healthy volunteers ( 68. 87%± 20. 94% vs 69. 33%±16. 84%,P>0. 05 ) . ( 2 ) OC could be transdifferentiated from imDC when cultured with RANKL and M-CSF. ( 3 ) The number of TRAP+ multinuclear cell and the absorption area of dentine were significantly lower in the group of imDC indirectly co-cultured with γδ T cells than in the group of control imDC(5.67±0.58 vs 28.33±2.08,4.97%±4.3% vs 28.47%±12.8%, respectively). (4) Under the circumstance of γδ T cell-imDC indirect coculture,TNF-α got significantly higher. Conclusion: γδ T cells might inhibit the transdifferentiation of imDC into OC.γδ T cells-based immunotherapy is expected to be a new treatment for myeloma bone disease.
ABSTRACT
BACKGROUND: Dendritic cells induce tolerance in the immature state besides the function of antigen presenting cells in the mature state. Using this effect, it will be overcome the rejection in organ transplantation. In this research we are to confirm the tolerance effect and induction mechanism in immature dendritic cell cultured with low-dose GM-CSF. METHODS: Dendritic cells were cultured from BALB/c (H-2d) bone marrow cells in the low concentration of GM-CSF (5U/mL, GM(lo)DC) for 10 days. The phenotype and functional properties of these GM(lo)DC were compared to those of standard BM-DC cultures generated in the high concentrations of GM-CSF (200U/mL) with lipopolysaccharide (GM(hi)DC). RESULTS: Compare to mature DC, GM(lo)DC expressed the low level of CD80 and CD86 and these cells were weak stimulators of allogeneic T cell responses by mixed lymphocyte reaction. The administration of GM(lo)DC prolonged allogeneic skin graft survival (median survival time 11.3+/-1.2 days, compared with 6.6+/-0.8 days in nontreated controls). The effects of these cells were donor specific but couldn't be sustained for a long period. CONCLUSIONS: GM(lo)DC were phenotypically immature and they didn't induce allogeneic T cell responses compared with GM(hi)DC in vitro. It is suggested that the less expression of costimulatory molecule in GM(lo)DC induce the tolerance effect.