| IntroductionOrgan transplantation has become a well-established modality for treatment of end stage disease.Allograft rejection will still be the main cause of death in transplant recipients who survived an indefinite time. Allograft rejection is a specific cellular immuneresponse mainly due to T cell activation. While the activation of the help T cells(Th cell) plays a critical role in the response stage. The activated Th cells can be transformed into effective Th cells and secret lots of cytokine which act as an important medium in the following immunoresponse stage. They induce the activation,proliferation and differentiation of effective lymphocytes to proceed allograft rejection. These cytokines can be divided into two groups, Thl and Th2, according to its effect on the allograft immunoresponse.Interleukin 10(IL-10) is a kind of Th2 cytokine. Previous researchs have demonstrated that IL-10 can reduce the expression of major histocompatibility complex II (MHCII) molecule on the surface of antigen-presenting cell(APC), decrease the secretion of Thl cytokin, inhibit the activation of cytotoxic T cells(CTL), macrophages and NK cells. Furthermore, in the organ transplantation field, recent reports have confirmed that the localized immunosuppression in the allograft is parellel to expression level of IL-10. The high expression of IL-10 can act an obvious immunosuppression of allograft rejection.In the present research we transfer IL-10 gene into cardiomyocytes in transplanted heart in mice and investigated gene transfer efficiency and expression in allograft as well as the intrinsic IL-10 expression and its implication in allo-immune response by means of pathology, RT-PCR, immunohistochemical staining. We also explored whether extrinsic IL-10 is able to inhibit the allograft rejection and their possible mechanism.Materials and MethodsA total of 200 mice (donors and recipients) were used in our experiments. Inbred female Balb/c (H-2d) mice were used as syngeneic donors and C57BL/6(H-2b) were used as allogeneic donors. All recipients were Balb/c (H-2d) mice who were randomly divided as follows: Group A: Syngeneic control (Balb/c-Balb/c) ;Group B: C57BL/6-Balb/c+NS; Group C: C57BL/6-Balb/c+pORF-ITRs; Group D: C57BL/6-Balb/c+pORF-mIL-10; Group E: C57BL/6-Balb/c+pORF-ITRs-mIL-10. All donors of group D, E, F were treated with 30ul plasmid/lipofectin complex by coronary perfusion through the ascending aorta before tranlplantation and donars of group B treated with 30ul NS. All groups were subdivided into day 1, 3, 5, 7 posttransplantation respectively for sample harvesting, and additional subgroups for observation of general situation and survival time. Graft specimens were harvested to determine morphological changes by pathological examination and gene expression by reverse transcription-polymerase chain reaction (RT-PCR), as well as protein expression by immunohistochemical staining. The graft infiltrated T cell subsets were also analyzed by immunohistochemical staining.All data were expressed as mean SD. Statistical analysis was performed by SPSS 12.0 software. Kaplan-Meier was used to analyse the survival rate between each groups Mann Whitney U test was used to determine the difference of graft survival time between each groups. One-way ANOVA and LSD Test were also used for parametric data analysis between groups, respectively. Probability P values <0.05 were considered different and P values <0.01 significantly different.ResultsAll transplanted hearts of group A survived more than 100 days. The mean survival time of transplanted hearts in group B and C was 8. 125 + 0. 991 days and 7. 715+0. 756 days respectively. Prolonged survival time of transplanted hearts were found within group D and group E as compared to that in group B and group C (15. 714+2.498 days and 17.857 + 1.864 days, respectively). The similar results were also found within group E as compared to that in group D. No signs of rejection were found at any time point in group A with minimal myocardial inflammatory infiltratio... |
PDF Full Text Request