ObjectiveTo investigate the effects of form deprivation on the morphology of different types of RGC in mice.MethodsSixty B6.Cg-Tg (Thy1-YFP) HJrs/J transgenic mice were randomly assigned to form-deprived group (n=28) and control group (n=32). The right eyes of mice in the form-deprived group were covered by an occluder for 2 weeks as experimental eyes. The right eyes of mice in the control group were taken as control eyes. Before and 2 weeks after form deprivation, the refraction and ocular biometrics were measured; RGC were stained with Bra3a antibody and counted; the morphology of RGC was reconstructed with Neuroexplore software after immunohistochemical staining. The data was compared among experimental eyes, fellow eyes and control eyes by one-way analysis of variance.ResultsTwo weeks after form deprivation, the axial myopia was observed in the experimental eyes (refraction: F=15.009, P＜0.001; vitreous chamber depth: F=3.360, P=0047; ocluar axial length: F=5.011, P=0013). The number of RGC in central retina of the experimental eyes was decreased compared with the fellow eyes and the control eyes (F=4.769, P=0.035). The reconstructed RGC were classified into 4 types according to their dendritic morphology. Form deprivation affected all 4 types of RGC but in a different way. Among them, 3 types of RGC were likely contribute to form vision perception. Form deprivation increased the dendrite branches in these types of ganglion cells. However, form deprivation decreasd dendrite segment numbers in both eyes and the intersection and length insholl analyse type 4 ganglion cells which were morphologically identified as ipRGC.ConclusionForm deprivation distinguishingly affects the morphology of different types of RGC, indicating that form vision and non-form vision play different role in myopia development.
Objective To investigate the expression of Bloom syndrome (BLM) helicase and tumor infiltrating dendritic cells (TIDC) in colon cancer tissues and their relationship with the prognosis of patients after surgery. Methods Onehundred and sixty-eight patients with colon cancer who underwent surgical resection in our hospital from June 2014 to August 2016 were selected as the research objects. The specimens of surgically resected colon cancer tissues and adjacent tissues archived by the pathology department were obtained, and the expression of BLM helicase and TIDC density were detected by immunohistochemistry. Pearson was used to analyze the correlation between BLM helicase expression and TIDC density, and the relationship between the expression of BLM helicase and TIDC density and the clinicopathological features of colon cancer was analyzed by using χ2 test or Wilcoxon rank test. The influencing factors of postoperative survival of patients with colon cancer were analyzed by Cox proportional hazards regression model. Results The relative expression of BLM helicase in colon cancer tissues was higher than that in adjacent tissues (1.49±0.33 vs. 1.02±0.17), while the TIDC density was lower than that in adjacent tissues [(9.53±2.36)% vs. (12.36±2.37)%], the differences were statistically significant (P<0.05). Pearson correlation analysis showed that there was a negative correlation between the expression of BLM helicase and TIDC density (r=–0.588, P<0.05). The expression of BLM helicase and TIDC density were correlated with tumor differentiation, clinical stage and lymph node metastasis (P<0.05). That is, those with high expression of BLM helicase and low density of TIDC had low degree of tumor differentiation, late clinical grade, and higher ratio of lymph node metastasis. Sixty-three cases died (37.5%) during the follow-up period (16–60 months, medium follow up 45 months). Log-rank analysis showed that the 5-year cumulative survival rate of the BLM helicase-low expression group was higher than that of the high expression group, and that of the TIDC-low density group was lower than that of the high density group (P<0.05). Cox regression analysis showed that the high expression of BLM helicase, low density of TIDC, low degree of tumor differentiation, late stage and lymph node metastasis were risk factors affecting the postoperative survival of patients with colon cancer (P<0.05). Conclusion The abnormal expression of BLM helicase and TIDC density in colon cancer tissues are related to the degree of differentiation and lymph node metastasis, which are risk factors affecting the long-term survival of patients with colon cancer.
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.
Objective To explore the effect of dendritic cells (DCs) allergized by K-ras mutant peptide on expressions of chemokines CCL19, CCL22, and cytoskeletal protein fascin-1. Methods DCs were derived from peripheral blood in the presence of granuloceyte/macrophage-colony stimulating factor, interleukin (IL) -4 in vitro. The DCs were collected on day 7 after culture, and were divided into non-K-ras mutant peptide group (addition of RPMI 1604 culture solution 50 μg/ml) and K-ras mutant peptide group (addition of K-ras mutant peptide 50 μg/ml). Phenotype was identified by flow cytometry. The morphological structure was observed by scanning and transmission electron microscopies, respectively. The expressions of IL-12, CCL19, and CCL22 were tested continuously by enzyme-linked immunosorbent assay (ELISA). The expression of cytoskeletal protein fascin-1 was determined by Western blot. Results ①The expressions of CD1a, CD80, and CD86 after loading K-ras mutant peptide were higher than that before loading K-ras mutant peptide (Plt;0.01). ②The DCs with petal-like and branch-like profections after loading were observed under scanning electron microscopy; The DCs with irregular shapes, branch-like or burr-like were showed under transmission electron microscopy. ③The expressions of IL-12, CCL19, and CCL22 in the Kras mutant peptide group were higher than those in the non-K-ras mutant peptide group at different times (6, 12, 24, and 48 h) after loading Kras mutant peptide (Plt;0.01). ④The expression of fascin-1 in the K-ras mutant peptide group was also higher than that in the non-K-ras mutant peptide group (Plt;0.01). Conclusion K-ras mutant peptide can promote DC to mature and improve the expression of chemokines and cytoskeletal protein which will strengthen DC migration.
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.
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 To study the mechanism of immune hyporesponsiveness of allograft rejection induced by transfusion nonpufsed allopeptide syngeneic immature dendritic cell (imDC) generated from recipient bone marrow progenitors and to explore a possible strategy for liver allograft protection in clinic. Methods Forty experimental rats were randomly divided into 4 group: control group, cyclosporine A (CsA) group, mature DC (mDC) group and imDC group. In control group, Wistar rats only received liver transplantation. In CsA group, Wistar rats underwent liver transplantation plus CsA treatment 〔10 mg/（kg·d)〕. In mDC group, recipient-derived mDC 1×106 were infused intravenously through the penile vein to Wistar rats. In imDC group, ImDC with the dose of 1×106 were injected into Wistar rats via the dorsum vein of penile. In each group, five recipients were killed on the 10th day after transplantation, the other five recipients were left to observe survival time. The levels of ALT, AST, TBIL, IL-2, IFN-γ, IL-4 and IL-10 were detected. The acute rejection and the expression of FasL/Fas in the grafts were detected by HE and immunohistochemical staining. Western blot was used to detect Scurfin protein expression of CD4+ CD25+ T cells. Results The median survival time of the liver allografts in CsA group and imDC group were significantly longer than that in control group and mDC group ( P ＜ 0.05). The levels of ALT and TBIL in control group and mDC group were significantly higher than those in CsA group and imDC group ( P ＜ 0.05). Compared with CsA group and imDC group, the levels of IL-2 and IFN-γ were higher but the levels of IL-4 and IL-10 were lower in control group and mDC group ( P ＜ 0.01). Slightly or no rejection reaction was found in CsA group and imDC group ( P ＜ 0.05). The Scurfin protein expressions of CD4+ CD25+ T cells of imDC group were significantly higher than those of other three groups. Conclusion Application of nonpufsed allopeptide syngeneic recipient-derived imDC is an effective way to induce immune hyporesponsiveness by blocking indirect recognition in rat liver transplantation model. Survival span is significantly prolonged by its protective effect. The mechanism of immune hyporesponsiveness induced by imDC transfusion might be involved in some aspects: T cell apoptosis, immune deviation of Thl/Th2 cytokine net and inhibition of T lymphocytes responsiveness by regulatory T cells.
ObjectiveTo review the role of dendritic cells (DC) in immune metabolism of rheumatoid arthritis (RA). MethodsLiterature on the role of DC in the immune metabolism of RA was extensively reviewed in recent years, and the metabolic characteristics of RA, the role of DC in RA, the correlation between the immune metabolism of DC and pathogenesis of RA, and the treatment were summarized and analyzed. Results DC promotes the progression of RA under hypoxia, increased glycolysis, inhibition of oxidative phosphorylation, and decreased lipid metabolism. Moreover, many DCs (especially conventional DC and monocyte-derived DC) have different functions and phenotypic characteristics in RA, which are closely related to the occurrence and development of RA. Conclusion DC plays an important role in the immune metabolism of RA, and immunometabolism therapy based on DC can provide targeted therapy for the treatment of RA.
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.