These results emphasize the importance of specifically screening heart transplantation candidates for the presence of IgG antibodies directed against MHC class II molecules and suggest that strategies aimed at their reduction may have an impact on the onset and frequency of high-grade cellular rejections after transplantation.
In depth phenotyping and functional analysis of skin dendritic cell subsets suggests that the function of Langerhans cells may not be conserved between mouse and human and supports the idea that human Langernhans cells may be a relevant therapeutic target.
Murine heterotopic tracheal allografts develop obliterative airway disease (OAD), a suitable model of chronic lung allograft rejection. This model, however, fails to account for the behavior of the allograft when adjacent to recipient airway tissues, particularly the epithelium. This study was performed to determine the immunologic role of the epithelium in development of OAD. BALB/c (H2 d ) tracheal allografts were transplanted orthotopically into C57BL/6 (H2 b ) mice and harvested 14-150 days post-transplantation. The phenotype of the allograft epithelium after orthotopic transplantation was determined with immunofluorescent staining. Orthotopic BALB/c tracheal allografts harvested at 28 days were re-transplanted heterotopically into BALB/c or C57BL/6 mice, harvested after 28 days, and assessed for OAD. Orthotopic allografts displayed mild cellular infiltration, no fibrosis and preserved epithelium at 28 days post-transplant. The presence of recipient-derived epithelium within the allograft was demonstrated with immunofluorescent staining at day 14. Significantly, BALB/c orthotopic allografts re-transplanted heterotopically into BALB/c mice developed OAD by day 28, whereas BALB/c orthotopic allografts re-transplanted heterotopically into C57BL/6 mice did not. Repopulation of orthotopic tracheal allografts with recipient-derived epithelium confers a protective effect against OAD after heterotopic re-transplantation. This indicates that the airway epithelium plays a crucial role in OAD development.
We hypothesized that localized IL-10 gene transfer can induce alloreactive T cell apoptosis and tested this hypothesis with liposome-mediated ex vivo intracoronary IL-10 gene transfer using a functional heterotopic allograft heart transplant model in rabbits. Localized IL-10 overexpression prolonged cardiac allograft survival over three folds. In parallel with the time-course of IL-10 overexpression, the percentage of apoptotic CD3+ cells among total CD3+ cells was significantly increased in the gene therapy group (36.5+/-3.9%) compared with that in the control group (6.2+/-2.6%, P<0.01) on postoperative day (POD) 3-6, and it was further increased (45.8+/-5.7%) on POD7-10. Apoptotic CD4+ and CD8+ cells were also significantly increased in the gene group (P<0.01). In contrast, the percentage of apoptotic myocytes significantly decreased from 10.1+/-0.8% in the control group to 3.5+/-0.4% in the gene group on POD7-10 (P<0.01). This reduction was inversely correlated with the increase in the percentages of apoptotic CD4+ and CD8+ cells (P<0.01). The percentage of caspase-3 positive myocytes was significantly reduced, although percentages of caspase-3 positive CD4+ and CD8+ cells were markedly increased in the gene group (P<0.01). Moreover, about 60-80% of apoptotic T lymphocytes expressed Fas in the gene group compared with less than 10% in the control group (P<0.01). These results suggest that localized IL-10 gene transfer induces alloreactive T cell apoptosis via the Fas/FasL pathway that may contribute to the alleviated acute rejection, improved cardiac function, and prolonged survival in the IL-10 gene-treated cardiac allografts.
Activation-induced cell death and cytokine deprivation are demonstrated by peripheral T cell populations at the conclusion of natural immune responses, and each of these processes is modulated by the immunosuppressive cytokine interleukin (IL)-10 in vitro. This study employs a clinically relevant in vivo model of IL-10 gene transfer with heterotopically transplanted cardiac allografts to determine the mechanisms of the effects of IL-10 on T cell survival. IL-10 protein overexpression within allografts 4-5 days after gene transfer augments apoptosis of CD4+ and CD8+ graft-infiltrating lymphocytes by 7.1-fold (P < 0.001) and 6.0-fold (P < 0.001), respectively. Graft-infiltrating T cells express 10-fold more proapoptotic Fas (P < 0.01) and 30-fold more Bax (P < 0.01) than controls. The fractions of activated caspase-8 (FADD-like IL-1beta-converting enzyme) and activated caspase-9 were increased 7- and 2.3-fold, respectively, in IL-10 gene-treated allografts at postoperative day 4-5. These changes in the Fas-Fas ligand pathway and Bcl-2 mitochondrial apoptosis regulation are enhanced by complete suppression of antiapoptotic FADD-like IL-1beta-converting enzyme inhibitory protein (FLIP) (from 30.5 to 0.0%, P < 0.01) and Bcl-xL (from 22.5 to 0.1%, P = 0.03) expression among these cells from the earliest days after gene transfer. Although changes in proteins of Fas- and Bcl-2-mediated apoptosis signaling occur, only the levels of Fas and FLIP correlate to the rate of apoptosis of graft-infiltrating CD3 lymphocytes and histological rejection scores. These results indicate that dichotomous apoptosis-regulatory pathways are affected by IL-10 gene therapy, but Fas-mediated mechanisms of activation-induced cell death more substantially contribute to the greater cell death of graft-infiltrating T cells after ex vivo IL-10 gene transfer.
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