Objective To observe the interferon-gamma; (IFN-gamma;), interleukin-2 (IL-2) levels of Th1 cytokine and IL-4、IL-10 levels of Th2 cytokine in serum and culture supernatants of splenic cells of the rats in the prevention of experimental autoimmune uveoretinitis（EAU）by oral tolerance. Methods 72 Lewis rats were randomly divided into EAU group，oral tolerance group (which including 10 mu;g、100 mu;g、1 mg、10 mg of S antigen group respectively) and control group，12 rats in each group. The animal model of EAU was induced by immunization with S antigen（50 mu;g）and Freundrsquo;s complete adjuvant. Oral tolerance 10 mu;g、100 mu;g、1 mg and 10 mg group were fed with 1 ml mixture of 10 mu;g、100 mu;g、1 mg、10 mg S antigen and 1 mg trypsin inhibitor respectively by intubation，once the other day，totally 7 times，and then induced EAU according to above methods；control group was fed with 1 ml mixture of phosphate buffered saline and 1 mg trypsin inhibitor，once the other day，totally 7 times，and then induced EAU. The clinical manifestation of EAU in the eye were recorded,the eyeballs were enucleated at the peak of EAU，followed by pathological grading. Meanwhile the serum was colleced; splentic cells were separated and cultured to collect the supernatant. Cytokine levels of IFN-gamma;, IL-2, IL-4 and IL-10 in serum, cultured supernatant of splenic cells were determined by enzyme-linked immunosorbent assay (ELISA). Results Compared with EAU and control group, the levels of IFN-gamma; and IL-2 (Th1 cytokine) in the serum in 100 mu;g and 1 mg group were decreased while the levels of IL4 and IL10 (Th2 cytokine) were increased，the differences were statistically significant（F=51.9, 68.8, 35.7,7.5,P＜0.01). Compared the levels of Th1 and Th2 cytokines in the serum in 10 mu;g, 10 mg group with EAU and control group, the differences were not statistically significant. In 100 mu;g、1 mg group, the levels of IFN-gamma; and IL-2 (Th1 cytokine) in the culture supernatant of splenic cells were decreased while the levels of IL-4 and IL-10 (Th2 cytokine) were increased, compared with EAU and control group, the differences were statistically significant（F=57.1,15.6,33.1,167.7, P＜0.01）. Compared the levels of Th1 and Th2 cytokine in the culture supernatant of splenic cells in 10 mu;g、10 mg groups with EAU and control group, the difference are not statistically significant. Conclusions In the process to prevent EAU by oral intake, the levels of IFN-gamma; and IL-2 (Th1 cytokine ) were decrease while the levels of IL-4 and IL-10 (Th2 cytokine). Oral administration with too high or low dose of the antigen can not prevent EAU as well as the cytokine levels do not change obviously. Cytokines has played an important role in the prevention of EAU.
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 effect of allogeneic canine cord blood mesenchymal stem cells（cbMSC）transplantation on the distribution of CD4+T and CD8+T in infracted area of hearts. Methods Mononuclear cells of cord blood were isolated by density gradient centrifugation and amplified by adherent culture. 36 adult male dogs were divided into experimental group and control group. Animal models of acute myocardial infarction were established by ligating anterior descending coronary artery. The fourth generations of mesenchymal stem cells (MSC) were transplanted into infarcted area of hearts by left anterior descending coronary artery after 72h induced by 5-aza and transfected by LacZ. The survival of transplanted cells in hearts can be confirmed by βgal expression. CD4+T and CD8+T cells distributed in infarcted area were detected by immunohistochemical staining method. The ImagePlus 5.1 software was used to analyze the images. Results Cells transplanted into infarcted area could survive for a long time. 2, 4, 8 weeks after transplantation, the IOD of CD4+T in experimental group were 44.35±7.03, 19.29±4.11 and 20.27±3.51 respectively, and the CD4+T/CD8+T ratios were 0.63±0.12, 0.51±0.15 and 0.66±0.08. In control group, the IOD of CD4+T at 2, 4, 8 weeks after transplantation were 65.78±10.27, 28.02±2.59, 29.79±6.83, and the CD4+T/CD8+T ratios were 1.28±0.20, 1.34±0.09 and 1.50±0.16. The IOD of CD4+T and CD4+T/CD8+T ratio in experimental group were significantly lower than that in control group. In experimental group the IOD of CD8+T at 2, 4, 8 weeks after transplantation were 69.88±7.84 , 37.80±8.83 and 30.81±7.42, higher than that in control group which were 51.28±10.01, 20.87±4.50 and 19.91±2.87. Conclusion The preliminary results indicated that allogeneic cbMSC transplanted in infarcted area can escape from immune rejection, its mechanism may be associated withdecreasing the amount of CD4+T cells infiltrated in periphery of infarcted area and maintaining CD4+T/CD8+T ratios at a lower level.
ObjectiveTo approach the role of CD4+CD25+ regulatory T cells in the maintenance of immunotolerance in mouse liver allograft. MethodsThe mouse orthotopic liver transplantation was performed. After the liver transplantation immunotolerance induction, antiCD25 monoclonal antibody (PC61) was injected into the recipients with a delayed timing to remove the CD4+CD25+ T cells. The percentage of CD4+CD25+ T cells and the expression of forkhead/winged helix transcription factor (Foxp3) in the recipients were examined. Furthermore, the survival time of the recipient was observed. ResultsC3H/HeJ recipients receiving DBA/2 hepatic allografts survived over 70 d as in the syngeneic liver transplantation (C3H/HeJ recipients receiving C3H/HeJ hepatic grafts). With various protocols of the delayed PC61 treatment, the CD4+CD25+ T cell was completely disappeared as observed. However, the removal of CD4+CD25+ regulatory T cells after the induction of transplantation immunotolerance did not affect the survival of hepatic allografts. ConclusionCD4+CD25+ regulatory T cells are not essential for the maintenance of spontaneous mouse liver transplantation immunotolerance.
Objective To summarize the advancement of immune tolerance in pancreas transplantation.Methods Relevant literatures about immune tolerance in pancreas transplantation, which were published recently domestic and abroad were collected and reviewed. Results The main methods to induce immune tolerance are peripheral tolerance and central tolerance. The induction of chimerism by infusion of donor-specific bone marrow cells is the research hot spot recently. Conclusion The infusion of donor-specific bone marrow cells in combination with one or more peripheral tolerance maybe can induce immune tolerance successfully. However, it should be researched further.
Objective To investigate the expression of transcription factor Foxp3 in the orthotopic liver transplantation by using the inbred rats with spontaneous immune tolerance. Methods The model of orthotopic liver transplantation was established on inbred rats according to double-sleeve technique. The total RNA that was isolated from liver was reversely transcribed into cDNA. The method of real-time fluorescence quantitative PCR (RFQ-PCR) was used to analyze the expression level of Foxp3 mRNA in tolerance group and syngeneic group, respectively. The expression of Scurfin in hepatic tissue was assayed by Western blot and then was analyzed by computer imaging system. Results The expression levels of Foxp3 mRNA and Scurfin in the transplanted liver were significantly lower than those of normal liver within the first week after transplantation. The level of Foxp3 mRNA began to increase on day 7 and reached the peak point on day 14. The expression level of Foxp3 mRNA began to decrease on day 30 but was still higher than the normal value (P＜0.05). The Western blot showed resemble changes on that of Scurfin. Conclusion Transcription factor Foxp3 may play an important role in the spontaneous immune tolerance in the orthotopic liver transplantation of inbred rat.
ObjectiveTo evaluate the effect of pre-infusion of allogeneic lymphoyctes treated with 5-FU on the rat liver graft. MethodsRat liver transplant models from Wistar to SD were established. Four groups were designed as following: control group: only liver transplantation without any other intervention; lymphocytes group: 1 ml of untreated lymphocytes (5×106/ml) from Wistar rats were preinfused into SD rats on day 7 and 4 separately before transplantation; lymphocytes with low concentration of 5-FU group: low concentration 5-FU (7.5 μg) treated lymphocytes were preinfused as above; lymphocytes with high concentration of 5-FU group: high concentration 5-FU (15 μg) treated lymphocytes were preinfused as above. Fas-L and CD8 expression were detected by immunohistochemistry method on day 7 after transplantation. ResultsThe integral opticaldensity (IOD) of Fas-L positive lymphocytes in the lobules of liver and portal areas were higher in lymphocytes with low concentration of 5-FU group than in the other groups （Plt;0.05）. There was no difference between lymphocyte group and lymphocytes with high concentration of 5-FU group （Pgt;0.05）. The IOD of CD8+ expression in lobules of liver was not different among all the three lymphocytes treated groups （Pgt;0.05）. But in portal areas, CD8+ expression was lower in the lymphocytes with low concentration of 5-FU group than in the other groups （Plt;0.05）. ConclusionPreinfusion of lymphocytes treated with low concentration 5-FU can induce graft immune tolerance, the probable mecanism of which is the increasing Fas-L expression in graft.
Objective To summarize the role of costimulatory molecules in inducing immune tolerance of organ transplantation. Methods Domestic and international publications online involving costimulatory molecules and immune tolerance in recent years were collected and reviewed. Results The relationship between costimulatory pathways and transplantation immunity has already been clarified in recent years. The main costimulatory molecules alreadly found mainly include B7-CD28/CTLA4, CD40-CD154, 4-1BB/4-1BBL, and ICOS-B7h, etc. Costimulatory pathways com-inhibition or combining with other immunosuppression methods could obtain stable and long lasting immune tolerance. Conclusions With the development of immunology and molecular biology, costimulatory pathways of T lymphocyte activation will be further interpreted. Other new costimulatory molecules will be discovered in the future, which will afford theory evidence for inducing immune tolerance.