Objective To study the advances in the relationship between the number of infiltrating dendritic cells and the postoperative prognosis of digestive malignant tumor. MethodsThe literature in recent years on the relationship between the number of infiltrating dendritic cells and the postoperative prognosis of digestive malignant tumor was reviewed.ResultsThe number of infiltrating dendritic cells among esophageal cancer,and gastric carcinoma,colonic cancer and pancreatic cancer was associated with a better prognosis.Conclusion The population density of dendritic cells among the malignant tissue could be regarded as an independent indicator in estimating the postoperative prognosis of malignant tumor.
Objective To study the method of obtaining a large number of dendritic cells （DC）. To study the specific cytotoxicity T lymphocyte (CTL) effect against tumor cells initiated by DC pulsed with peptide of cancer cell. Methods Development of cells with cytologic features of DC in bone marrow cultures supplemented with granulocyte macrophage-colony stimulus factor (GM-CSF) and IL-4. Determining the DC phenotype and the specific structure by electronic microscopy. The CTL effect against pancreatic carcinoma leading by the DC pulsed with tumor cells lysate in vitro was observed. Results A large number of typical DC was proliferated by supplementing with GM-CSF and IL-4 cytokines. DC had specific cell appearance and structure, and highly expressed various cell surface molecules. TNF-α had the ability of stimulating DC mature, the mature DC had the enhancing abilities of antigen presenting and IL-12 self-secreting, as well as, expressed higher levels of CD54， MHC-Ⅱ and CD86 molecules than control group (P＜0.05). T lymphoid cell stimulated by DC without tumor antigen could not recognize and kill the target cells, only if DC pulsed with peptide of cancer cell can lead a b immune response to special tumor cells. The inhibiting ratio of CTL was significantly higher than that in other groups (P＜0.01). Conclusion Bone marrow DC has b ability of inducing special CTL against determined cancer cells after they are pulsed with tumor cell lysate. DC vaccine is probably a new immunotherapeutic method against tumor in the near future.
Objective To investigate the feasibility of dendritic cells ( DCs ) as vector of immunotherapy through intratracheal injection. Methods The DCs obtained from the bone marrow of BALB/ c mice were cultured and isolated with CD11c-positive magnetic beads. Then DCs were overloaded with ovalbumin peptide 323-339 ( OVA 323-339) for 24 hours. The mice in the DC-OVA group were intratrachelly injected DCs overloaded with OVA 323-339 in dose of 2 ×106 cells per mouse. The mice in thenegative control group were intratracheally injected with DCs untreated by OVA 323-339. On the second day,all mice were challenged with 1% OVA in PBS lasting for five days. The asthma animal model established by classic method was used for the positive control. Pathologic changes in lung and cell numbers in bronchoalveolar lavage fluid ( BALF) were assayed 24 hours after challenged. Results Just like the lung tissues from the mice asthma models, the lung tissues from the mice instilled with DCs overloaded with allergen OVA 323-339 showed extensive inflammatory cells infiltration, most of which were eosinophils, neutrophils and lymphocytes. The lung tissues in the DC group showed no obvious inflammation. There were more cells in BALF in the DC-OVA group than that in the DC group. OVA-specific IgE in serum from the DC-OVA group was not significantly different from that in the mice asthma models [ ( 48. 22 ±4. 76) U/mL vs. ( 52. 75 ±4. 03) U/mL, P gt;0. 05] . Conclusion DCs overloaded antigen has the ability of transferring of antigen effectively and may be used as vectors of immunotherapy.
Abstract: Objective To investigate the phonetype and tolerogenic function of semimature dendritic cells (DC) transfected by myeloid differentiation marker 88（MyD88） small interfering ribonucleic acid（siRNA）. Methods Bone marrow of BALB/c mice was inoculated and cultured in vitro，and induced into DC by 10ng/ml recombinated granulocyte macrophage colony stimulating factor (rmGM-CSF) ，then DC was divided into three groups at the 8th day: blank control group: added nothing; lipopolysaccharide （LPS）group: added with 1μg/ml LPS; and experimental group: added with 1μg/ml LPS after transfected by MyD88 siRNA for 4 hours. The phonetype of three groups was analysed by fluorescenceactivated cell sorting (FACS). The concentration of interleukin 10（IL-10）and interleukin 12（IL-12） in culture supernatant was detected by enzyme linked immunosorbent assay（ELISA）.The function of stimulating alloreactive T cell roliferation were evaluated by primary and secondary mixed lymphocyte reaction （MLR）. The cardiac allograft survival time was compared after DC of three groups injected into recipient mice. Results The phonetype of blank control group DC was CD11c＋，CD25-，CD40low，CD80low，CD86low，MHC-Ⅱlow， which could be induced to mature DC by LPS. Experimental group DC was phonetypically semimature DC (CD11c＋，CD25－，CD40mid，CD80low，CD86low，MHC-IIid) and the IL-10/IL-12 ratio of semimature DC increases significantly. yporesponsiveness of alloactive T cells can be induced by experimental group DC and the survive time of heart allograft was prolonged. Conclusion Immature DC could become semimature DC after transfected by MyD88 siRNA and stimulation by LPS. These semimature DC are more tolerogenic than immature DC.
Objective To investigate the effects of FasL gene-modified dendritic cell (DC) on the airway inflammation in mice sensitized/challenged by house dust mite (HDM) allergen.Methods FasL gene-modified DC (FasL-DC) and control DC (nontransfection DC) were administrated into HDM sensitized and challenged mice by intratracheal injection respectively，then HDM sensitized and challenged mice were sacreificed two days later.Total and differentiation cell counts and levels of interleukin-4（IL-4），IL-5 and interferon-γ(IFN-) in bronchoalveolar lavage fluid (BALF) were detected and lung histological features were observed.Results After administration of FasL-DC，lung allergic inflammation was ameliorated while total cell counts，the percentage of eosinophil ,the levels of IL-4 and IL-5 in BALF decreased and the level of IFN- in BALF increased.Conclusion Administrating FasL-DC into HDM sensitized/challenged mice can inhibit Th2 cells activation and ameliorate airway allergic inflammation.
ObjectiveTo explore the antitumor effect of tumor vaccine fused from dendritic cells (DC) and Walker-256 cancer cells on implanted liver cancer in rats and the related mechanism of inhibition for tumor angiogenesis. MethodsWalker-256 cancer cells and mature DC were fused by 50% polyethylene glycol method for preparation of DC-Walker-256 fusion vaccines. Implanted liver cancer models were established through operations on healthy male SD rats at the age of 6－8 weeks. All the rats were divided into four groups, and rats in each group were injected subcutanely with fusion vaccine (group), mixed cultured cells (group), simple DC (group), and PBS (blank control group), respectively. On 28 d after making model, the rats were put to death, the tumor was observed and pathological essays were prepared. All rats’ spleens were collected and prepared into lymphocyte to detect antigenic specificity cytotoxic T lymphocyte (CTL) by enzymelinked immunosorbent spot (ELISPOT) method. The expressions of VEGF, ANG-1, ANG-2, and MVD were detected by immunohistochemistry. ResultsThe numbers of rats survived in the fusion vaccine group, mixed culture cells group, simple DC group, and blank control group was 8, 5, 6, and 3, respectively. The rats in the other three groups except for fusion vaccine group were manifested as inaction, anorexia, and gloomy fur in some degree as well as ascites. The tumorigenesis was found in all survival rats except for two in the fusion vaccine group. The weight of liver tumors of rats in the fusion vaccine group 〔(32.4±9.2) g〕 was significantly lighter than that in the mixed culture cells group 〔(67.3±5.1) g, P=0.031〕, simple DC group 〔(75.0±8.3) g, P=0.019〕, and blank control group 〔(86.6±10.5) g, P=0.008〕, respectively. The number of tumorspecific CTL of rats in the fusion vaccine group was also significantly higher than that in the other three groups (P=0.019, P=0.025, and P=0.001, respectively). The MVD of tumor tissue in the fusion vaccine group was (24.12±2.32) vessels/HP, which was significantly lower than that in the mixed culture cells group 〔(40.34±1.29) vessels/HP, P=0.025〕, simple DC group 〔(42.36±3.16) vessels/HP, P=0.035〕, and blank control group 〔(56.48±5.16) vessels/HP, P=0.006〕, respectively. The MVD of tumor tissue in the mixed cultured cells group and simple DC group was similar (P=0.165), however, which was significantly lower than that in the blank control group (P=0.040 and P=0.043). The positive rate of VEGFA protein expression was 23.2% in the fusion vaccine group, which was significantly lower than that in the mixed culture cells group (42.5%, P=0.031), simple DC group (61.3%, P=0.019), and blank control group (89.6%, P=0.003), respectively. The positive rate of VEGF-A protein expression in the mixed cultured cells and simple DC groups was similar (P=0.089), however, which was significantly lower than that in the blank control group (P=0.027 and P=0.038). The positive rate of ANG-1 protein expression in the fusion vaccine group (43.2%) was not different from that in the mixed culture cells group (46.3%, P=0.292), simple DC group (51.3%, P=0.183), or blank control group (49.6%, P=0.179), respectively, and the difference of pairwise comparison in latter three groups was not significant (P=0.242, P=0.347, and P=0.182). The positive rate of ANG2 protein expression was 19.2% in the fusion vaccine group, which was significantly lower than that in the mixed culture cells group (62.3%, P=0.007), simple DC group (67.3%, P=0.005), and blank control group (71.6%, P=0.004), respectively, however, the difference of pairwise comparison in latter three groups was not significant (P=0.634, P=0.483, and P=0.379). ConclusionFused vaccine can induce CD8+ CTL aiming at tumor cells and establish the effective antitumor immunity in vivo and also downregulate the level of VEGF and ANG-2 to suppress tumor angiogenesis and thereby achieve the purpose of curing tumor.
Objective To investigate the effects of nuclear factor kappa B decoy oligodeoxynucleotides ( NF-κB decoy ODN) transfection on biological characteristics of mature dendritic cells ( mDCs) in mice. Methods Immature DCs were harvested from Balb / c mice bone marrow, followed by the incubation with antigen OVA and LPS, and mature DCs were evaluated by the expressions of CD11c and MHC-Ⅱ detected by FACS. Mature DCs were transfected with NF-κB decoy ODN and the changes of NF-κB activity after the transfection were detected by EMSA. The expressions of the costimulatory molecules( CD40,CD80 and CD86) on DCs were detected by FACS and the proliferation of T cells was tested by mixed lymphocyte reaction( MLR) . Results The mature DCs were cultured successfully. The NF-κB activity of NF-κB decoy ODN transfected DCs was decreased significantly( P lt; 0. 05) . There was no difference in the expressions of CD40 and CD80, but the expression of CD86 was decreased significantly in NF-κB decoy ODN transfection group( P lt; 0. 05) . MLR test showed that the proliferation of T lymphocyte cells was inhibited by NF-κB decoy ODN transfected DCs, but was stimulated bly by the DCs of other groups. Conclusions Mature DCs transfected with NF-κB decoy ODN could inhibit the proliferation and activation of antigenspecical T cells, which was probably related to the down-regulation of CD86 on DCs. This modified DCs might be a promising vaccine for the treatment of asthma in the future.
Objective To observe the effect of transfer of immature mouse myeloid dendritic cells （DC） generated with low-dose granulocyte-macrophage colony-stimulating factor （GM-CSF） on cardiac allograft survival. Methods Mouse DC were generated with standard doses or low doses GM-CSF from bone marrow cells, the phenotype and functional properties of these DC were compared through fluorescence-activated cell sorting（FACS） analysis and mixed lymphocyte reaction（MLR）, 1. 0 × 106 DC generated with low doses GM-CSF were administered to the recipients 7 days before transplantation, and the cardiac allograft survival were observed. Results In contrast to DC generated with standard doses, DC generated with low doses were phenotypically immature DC （CD11c＋, CD80- , CD86- , MHCⅡlow）, and induced allogeneic T cell unresponsiveness, and administration of these DC to recipients prolonged cardiac allograft survival from 6.3±1.2 days to 14.3±1.9 days. Conclusions DC generated from mouse bone marrow progenitors in low doses of GM-CSF are phenotypically and functionally immature, and prolong cardiac allograft survival when they are administered 7 clays before transplantation.
Objective By using small interfering RNAs ( siRNAs) specific for spleen tyrosine kinase ( Syk) , to evaluate the role of Syk in maturation of bone marrow-derived dendritic cells. Methods The fragments of 21-23 bp siRNAs specific for mice Syk were chemo synthesized and transfected into the asthmatic murine bone marrow-derived dendritic cells ( BMDCs) by Lipofectamine 2000 transfection system for 48 hours. Then BMDCs were co-cultured with T cells from the normal mice spleen for 48 hours. The cytokines including IL-4, IL-13, IL-2 and INF-γin supernatant were detect by ELISA. The expression of Syk protein was measured by Western Blot to determine whether the Syk gene was silenced. Results The expression of Syk protein was obviously decreased in the siRNA-interference group. The secretions of IL-4 and IL-13 were significantly inhibited by siRNA interference ( P lt; 0. 05) , but the secretions of IL-2 and INF-γwere not interfered signficantly ( P gt;0. 05) . Conclusion Syk specific siRNA fragments can block the antigen presentation function of dendritic cells and block the activation and differentiation of T cells.
Objective To detect the anti-colon cancer ability of whole cell lysates pulsed dendritic cell (DC) which acts as an adjuvant. Methods Whole cell lysates of LoVo cells were extracted with freeze thawing method, then the monocyte-derived DC were pulsed with this cellular antigen. Subsequently, the capability of antigen pulsed DC to promote T lymphocytes proliferation and the ability of T lymphocytes to kill LoVo cells were detected by 3H-TdR incorporation and lactate dehydrogenase release methods, respectively. Results The whole cell lysates of LoVo cells pulsed DC significantly stimulated T cells proliferation, and the cytotoxicity abilities of primed T cells to kill LoVo cells were also enhanced. Conclusion Tumor cell lysates which act as the cellular antigen to pulse DC can improve the efficacy of anti-cancer immune response， which provides us an experimental evidence for cancer immunotherapy.