Background— In allograft rejection, recipient leukocytes and alloantibodies first target donor endothelial cells. Although the leukocyte integrin Mac-1 (α Mβ2 , CD11b/CD18) facilitates cell–cell interactions among leukocytes and interactions between leukocytes and endothelial cells or platelets, its role in allograft survival and vasculopathy is incompletely defined. Methods and Results— This study examined parenchymal rejection and graft arterial disease after total allomismatched cardiac transplantation (BALB/c donor heart and B6 recipients) in wild-type (WT) and Mac-1-deficient (Mac-1 −/− ) recipients. Recipient Mac-1 deficiency attenuated parenchymal rejection and significantly prolonged cardiac allograft survival from 8.3±1.3 days in WT recipient allografts (n=18) to 13.8±2.3 days in Mac-1 −/− recipient allografts (n=6; P <0.0001). Accumulation of neutrophils and macrophages significantly decreased in Mac-1 −/− compared with WT recipients. Adoptive transfer of WT but not Mac-1 −/− macrophages to Mac-1 −/− recipients exacerbated parenchymal rejection and reduced allograft survival; in contrast, adoptive transfer of WT neutrophils did not affect graft survival. Mac-1 −/− macrophages expressed significantly lower levels of costimulatory molecules both in vivo and in vitro, and mixed lymphocyte reaction using alloantigen-primed Mac-1 −/− macrophages resulted in significantly lower antigen-presenting function than for WT macrophages. Tumor necrosis factor-α production also fell in cultures with Mac-1 −/− macrophages. Despite attenuation of acute rejection, recipient Mac-1-deficiency did not prevent late graft arterial disease. Conclusions— These studies demonstrate critical participation of Mac-1 in alloresponses during cellular allograft rejection. These observations establish a molecular target for modulating recipient responses to prolong graft survival.

Background The calcium-binding proteins myeloid-related protein (MRP)-8 (S100A8) and MRP-14 (S100A9) form MRP-8/14 heterodimers (S100A8/A9, calprotectin) that regulate myeloid cell function and inflammatory responses, and serve as early serum markers for monitoring acute allograft rejection. Despite functioning as a pro-inflammatory mediator, the pathophysiological role of MRP-8/14 complexes in cardiovascular disease is incompletely defined. This study investigated the role of MRP-8/14 in cardiac allograft rejection using MRP-14-deficient mice (MRP14-/-) that lack MRP-8/14 complexes. Methods and Results We examined parenchymal rejection (PR) after major histocompatibility complex (MHC) class II allomismatched cardiac transplantation (bm12 donor heart and B6 recipients) in wild-type (WT) and MRP14-/- recipients. Allograft survival averaged 5.9 ± 2.9 weeks (n=10) in MRP14-/- recipients, compared to > 12 weeks (n = 15, p < 0.0001) in WT recipients. Two weeks after transplantation, allografts in MRP14-/- recipients had significantly higher PR scores (2.8 ± 0.8, n=8) than did WT recipients (0.8 ± 0.8, n=12, p<0.0001). Compared to WT recipients, allografts in MRP14-/- recipients had significantly increased T-cell and macrophage infiltration, as well as increased mRNA levels of IFN-γ and IFN-γ–associated chemokines (CXCL9, CXCL10, and CXCL11), IL-6, and IL-17, with significantly higher levels of Th17 cells. MRP14-/- recipients also had significantly more lymphocytes in the adjacent paraaortic lymph nodes than did WT recipients (cell number per lymph node: 23.7 ± 0.7 × 105 for MRP14-/- vs. 6.0 ± 0.2 × 105 for WT, p < 0.0001). The dendritic cells (DCs) of the MRP14-/- recipients of bm12 hearts expressed significantly higher levels of the co-stimulatory molecules CD80 and CD86 than did those of WT recipients 2 weeks after transplantation. Mixed leukocyte reactions using allo-EC-primed MRP14-/- DCs resulted in significantly higher antigen-presenting function than reactions using WT DCs. Ovalbumin-primed MRP14-/- DCs augmented proliferation of OT-II CD4+ T cells with increased IL-2 and IFN-γ production. Cardiac allografts of B6 MHC class II-/- hosts and of B6 WT hosts receiving MRP14-/- DCs had significantly augmented inflammatory cell infiltration and accelerated allograft rejection, compared to WT DCs from transferred recipient allografts. Bone marrow–derived MRP14-/- DCs infected with MRP-8 and MRP-14 retroviral vectors showed significantly decreased CD80 and CD86 expression compared to controls, indicating that MRP-8/14 regulates B7-costimulatory molecule expression. Conclusion Our results indicate that MRP-14 regulates B7 molecule expression and reduces antigen presentation by DCs, and subsequent T-cell priming. The absence of MRP-14 markedly increased T-cell activation and exacerbated allograft rejection, indicating a previously unrecognized role for MRP-14 in immune cell biology.

Background-Graft arterial disease (GAD) limits long-term solid-organ allograft survival. The thickened intima in GAD contains smooth muscle-like cells (SMLCs), leukocytes, and extracellular matrix. The intimal SMLCs in mouse GAD lesions differ from medial smooth muscle cells in their function and phenotype. Although intimal SMLCs may originate by migration and modulation of donor medial cells or by recruitment of host-derived precursors, the mechanisms that underlie their localization within grafts and the factors that drive these processes remain unclear. Methods and Results-This study of aortic transplantation in mice demonstrated an important function for chemokines beyond their traditional role in leukocyte recruitment and activation. Intimal SMLCs, but not medial smooth muscle cells, express functional CC chemokine receptor-1 (CCR1) and respond to RANTES by increased migration and proliferation. Although RANTES infusion in vivo promoted inflammatory cell accumulation in the adventitia of aortic allografts of wild-type and CCR1-deficient recipients, it increased GAD intimal thickening with SMLC proliferation in only the wild-type hosts. Aortic allografts transplanted into CCR1-deficient mice after wild-type bone marrow transplantation did not develop intimal lesions, which indicates that CCR1-bearing inflammatory cells do not contribute to intimal lesion formation. Moreover, RANTES induced SMLC proliferation in vitro but did not promote medial smooth muscle cell growth. Blockade of CCR5 attenuated RANTES-induced T-cell and monocyte/macrophage proliferation but did not affect RANTES-induced SMLC proliferation, consistent with a larger role of CCR1-binding chemokines in SMLC migration and proliferation and GAD development. Conclusions-These studies provide a novel mechanistic insight into the formation of vascular intimal hyperplasia and suggest a novel therapeutic strategy for preventing allograft arteriopathy. (Circulation. 2009;120:1800-1813.)Key Words: chemokines Ⅲ chemokine receptors Ⅲ smooth muscle cells Ⅲ cytokines Ⅲ arteriosclerosis Ⅲ pathogenesis Ⅲ transplantation D espite contemporary therapies that effectively limit acute rejection, solid-organ allografts can still develop severe and diffuse arterial intimal hyperplastic lesions (graft arterial disease [GAD]) that constitute a major obstacle to long-term success. GAD lesions consist primarily of smooth muscle-like cells (SMLCs) and extracellular matrix, admixed with infiltrating mononuclear leukocytes, 1,2 and can lead to progressive organ ischemia. Clinical Perspective on p 1813GAD occurs predominantly as a consequence of immunemediated allospecific vascular injury in grafts. 1 Host inflammatory cells initially interact with allografts at the vascular wall interface. Alloresponses induce the expression of cytokines, chemokines, adhesion molecules, and growth factors from activated donor endothelial cells and medial smooth muscle cells (SMCs), as well as from host inflammatory cells. The inflammatory events subsequently induce the accumulation and p...