IL-4 deficiency prevents eosinophilic rejection and uncovers a role for neutrophils in the rejection of MHC class II disparate skin grafts

BACKGROUND: Acute rejection of MHC class II-disparate bm12 skin grafts by C57BL/6 recipient mice is characterized by massive graft infiltration by eosinophils, together with increased intragraft amounts of IL-4 and IL-5 mRNA. IL-5 blockade prevents the intragraft eosinophil infiltration and prolongs the survival of skin allografts. As the differentiation of T cell precursors into Th2 cells is largely driven by IL-4, we investigated the role of IL-4 in MHC class II-disparate allograft rejection. METHODS: We performed skin grafts from MHC class II incompatible bm12 mice into wild-type C57BL/6 mice (IL-4) or C57BL/6 IL-4 deficient mice (IL-4). Graft survival, in vitro T cell reactivity, and histology were compared. RESULTS: We observed that 50% of IL-4 mice rapidly rejected their bm12 allograft, whereas the other 50% retained their graft 60 days after transplantation. Histological examination of bm12 allografts retained by IL-4 mice showed a normal appearance with no inflammatory infiltrate and no eosinophils. Among IL-4 mice that acutely rejected their bm12 skin graft, we observed a dense polymorphonuclear infiltrate. The depletion of neutrophils significantly prolonged bm12 graft survival. CONCLUSIONS: Eosinophil infiltrates, typical of MHC class II disparate acute skin graft rejection, are critically dependent on the availability of IL-4. IL-4 mice reject MHC class II disparate skin grafts by a pathway of rejection where neutrophils play a direct causal role.     

Interferon-gamma is necessary for initiating the acute rejection of major histocompatibility complex class II-disparate skin allografts.

BACKGROUND: Although interferon (IFN)gamma has immunostimulatory functions, it is not essential for the acute rejection of fully allogeneic grafts in mice. It is not known whether IFNgamma plays a critical role in the acute rejection of MHC class I- or MHC class II-disparate allografts. METHODS: We studied the survival of skin allografts transplanted from fully allogeneic (BALB/c), MHC class I-disparate (bml), or MHC class II-disparate (bm12) donors to C57BL/6 wild-type (IFNgamma+/+) and IFNgamma gene-knockout (IFNgamma-/-) recipients. We also investigated the in vitro responses of IFNgamma+/+ and IFNgamma-/- T cells to MHC class II-disparate splenocytes. RESULTS: We found that IFNgamma-/- recipients reject BALB/c and bml skin grafts at the same rate as IFNgamma+/+ mice but are not capable of rejecting bm12 skin. Despite the inability of IFNgamma-/- mice to reject bm12 skin grafts, IFNgamma-/- T cells displayed vigorous proliferation and cytotoxic responses when stimulated with bm12 splenocytes in vitro. Furthermore, priming IFNgamma-/- recipients with bm12 splenocytes enabled these mice to reject bm12 skin grafts at a normal rate and to mount a cutaneous delayed-type hypersensitivity response to the bm12 antigen. CONCLUSION: The data demonstrate that IFNgamma is not necessary for generating effector mechanisms associated with acute transplant rejection but that it is required for initiating alloimmune responses to MHC class II-disparate skin grafts.     

Unexpected effects of viral interleukin-10-secreting dendritic cells in vivo: preferential inhibition of TH2 responses

Viral interleukin (IL)-10 (vIL-10) has been widely described as an immunoregulatory cytokine that does not possess the T-cell costimulatory activities of cellular IL-10; it was therefore believed to be a more potent tolerogenic mediator. The immunosuppressive properties of this cytokine are partly attributed to its capacity to render dendritic cells (DCs) unable to undergo full maturation and to activate T cells. We reported here that myeloid DCs retrovirally transduced with vIL-10 had an impaired production of IL-12 and a decreased expression of MHC class II molecules but had minor defects in costimulatory molecule expression and no alteration on CCR5 and CCR7 expression. In mixed leukocyte reaction, vIL-10-transduced C57BL/6 bm12 (MHC class II mismatch) DCs had a reduced capacity to stimulate C57BL/6 wild-type CD4+ T-cell proliferation. We show that bm12 vIL-10-transduced DC administration in CD8-/- C57BL/6 mice promoted IFN-gamma production, down-regulated TH2-type cytokine production, and did not induce skin graft tolerance. These findings suggest that vIL-10-transduced DC may surprisingly facilitate Th1-type inflammatory responses in vivo.     

Differential effects of myeloid dendritic cells retrovirally transduced to express mammalian or viral interleukin-10 on cytotoxic T lymphocyte and natural killer cell functions and resistance to tumor growth

BACKGROUND: Genetic engineering of dendritic cells (DC) to express immunosuppressive molecule(s) offers potential for therapy of allograft rejection and autoimmune disease. Viral (v) interleukin (IL)-10, encoded by the Epstein-Barr virus, is highly homologous to mammalian (m) IL-10, but lacks certain of its T-cell stimulatory activities. Our aim was to evaluate and compare the influence of vIL-10 and mIL-10 gene transfer on the T-cell and natural killer (NK) cell stimulatory activity of DC, and their impact on the growth of transplantable tumors. METHODS: Myeloid DC progenitors, propagated from the bone marrow of C57BL/6J (H2b) mice in granulocyte-macrophage colony-stimulating factor + IL-4, were transduced using retroviral supematant from the BOSC ecotropic packaging cell line. The function of the IL-b gene-modified DC was assessed by examining their ability to induce naive allogeneic T-cell proliferation and cytotoxic T lymphocyte (CTL) generation. MCA205 (H2b) sarcoma cells mixed with either vIL-10-, mIL-10-, or Zeo (control gene)-transduced DC were inoculated intradermally into C57BL/6J (syngeneic) or BALB/cJ (H2d) (allogeneic) recipients, which were monitored for tumor growth. The role of specific host effector cell populations in tumor resistance was determined by antibody depletion. RESULTS: Compared with control gene-modified DC, both vIL-10- and mIL-10-transduced DC, which secreted the transgene product, showed reduced surface expression of MHC class II and costimulatory molecules, and impaired ability to induce T-cell proliferation. vIL-10-transduced DC were also inhibited with respect to CTL induction but did not affect the generation of NK cells. By contrast, mIL-10-transduced DC augmented CTL generation and NK cell activity. In the tumor transplant model, vIL-10-transduced DC enhanced tumor growth both in syngeneic and allogeneic hosts, whereas mIL-10-transduced cells inhibited tumor development. Depletion of CD4+ or CD8+ T cells or NK cells in mice given mIL-10-transduced DC reversed this therapeutic effect. CONCLUSION: mIL-10 gene-modified myeloid DC promote CTL and NK cell-mediated responses and inhibit tumor growth. By contrast, vIL-10-engineered DC, which elicit diminished CTL responses and do not promote NK cell activity, seem to have therapeutic potential for inhibition of T cell-mediated immunity.     

Impact of donor immune cells in pancreatic islet transplantation

Autoimmune-mediated cytotoxicity may cause pancreatic islet transplant failure, leading to recurrent diabetes. Protection of islet grafts depends on immunosuppressive control, which may also prevent autoimmune recurrence of diabetes. In this study, we compared the survival of syngeneic islet transplants using different strains of donor mice. We observed extended functional survival in the islet grafts from donors lacking the genetic background and potential of autoimmunity. Without immunosuppression, the islet grafts of NOR and immune-deficient NOD. Scid donors functioned up to 3 weeks in syngeneic islet transplants compared to 3-day survivals with the grafts from NOD donors. T-cell proliferation and activation markers, CD44 and CD69, were upregulated in NOD donors, suggesting that T-cell activation had occurred prior to pancreas procurement. Systemic delivery of a recombinant adenoassociated viral vector (AAV) encoding the viral (vIL-10) IL-10 gene (AAV vIL-10) in NOD recipients protected syngeneic islets from autoimmune destruction. Alternatively, pretreatment of NOD donor mice with AAV vIL-10 prolonged islet graft survival in untreated NOD recipients. Both studies indicate the effectiveness of vIL-10 gene therapy in autoimmune regulation. These results suggest that a donor factor may exist in autoimmune-prone donors. Therefore, autoimmune recurrence of diabetes may result from donor immune cells transferred during islet transplantation. The AAV vIL-10 gene therapy suppressed previously activated donor T cells and protected the grafted islets from autoimmune-mediated destruction.     

Delayed rejection of MHC class II-disparate skin allografts in mice treated with farnesyltransferase inhibitors

Farnesyltransferase inhibitors (FTIs), developed as anti-cancer drugs, have the potential to modulate immune responses without causing nonspecific immune suppression. We have investigated the possibility that FTIs, by affecting T cell cytokine secretion, can attenuate alloreactive immune responses. The effects of FTIs on murine alloreactive T cells were determined both in vitro, by measuring cytokine secretion or cell proliferation in mixed lymphocyte cultures, and in vivo, by performing skin allografts from H-2(bm12) mice to MHC class II-disparate B6 mice. We found that two different FTIs, ABT-100 and L-744,832, blocked secretion of IFN-gamma, IL-2, IL-4, and TNF-alpha from na?ve T cells in vitro. ABT-100 and L-744,832 blocked cytokine production from both CD4(+) and CD8(+) na?ve T cells stimulated with CD3 and CD28 antibodies, but only if the cells were pretreated with the FTIs for 48 h. Proliferation of alloreactive T cells in mixed lymphocyte cultures was blocked by either FTI. We also found that the proliferation of enriched T cells stimulated with IL-2 was blocked by ABT-100 treatment. In mice with an MHC class II-disparate skin graft, rejection of primary allografts was significantly delayed by treatment with either ABT-100 or L-744,832. Secondary rejection in mice previously primed to the alloantigen was found to be unaffected by L-744,832 treatment. We have shown that FTIs can block T cell cytokine secretion and attenuate alloreactive immune responses. The ability of FTIs to block secretion of cytokines, including IFN-gamma and IL-4, from na?ve T cells provides a likely biological mechanism for the specific suppression of class II MHC-mediated allorejection. These results demonstrate that FTIs may have useful immunomodulatory activity due to their ability to delay priming to alloantigens.     

Syngeneic bone marrow transduced with a recombinant retroviral vector to express endoplasmic reticulum signal-sequence-deleted major histocompatibility complex class-I alloantigen can induce specific immunologic unresponsiveness in vivo

BACKGROUND: Long-term survival of fully allogeneic cardiac grafts can be induced in mice through transduction of recipient bone marrow cells (BMCs) with a recombinant retroviral vector encoding a single full-length major histocompatibility complex (MHC) class I alloantigen. This study investigated whether cell surface expression of the transduced MHC antigen was necessary for the induction of specific unresponsiveness.METHOD The signal sequence for translocation into the endoplasmic reticulum was deleted from H-2K (SDELKb). Syngeneic BMCs from CBA.Ca (H2k) recipients were transduced with an MFG retroviral vector encoding either wild-type Kb or the mutant SDELKb and reinfused in conjunction with an anti-CD4 therapy. Four weeks later, the recipients underwent transplantation with a fully allogeneic C57BL/10 cardiac graft. Graft survival and the development of transplant arteriosclerosis were assessed. RESULTS: Expression of both the wild-type Kb or SDELK in recipient CBA mice before transplantation resulted in prolonged survival of C57BL/10 grafts. Grafts from recipients pretreated with SDELKb developed 48%+/-22% intimal proliferation compared with 61%+/-21% in grafts from recipients pretreated with wild-type Kb. However, this difference did not reach statistical significance.CONCLUSION Cell surface expression, and therefore direct recognition, of an MHC class I alloantigen is not required to induce long-term survival of fully allogeneic cardiac grafts after retroviral transduction of recipient BMCs.     

转染病毒白细胞介素10基因对同种小鼠心脏移植生存的影响

目的了解病毒白细胞介素10(vIL10)基因对同种小鼠移植心脏的影响。方法用小鼠干细胞病毒(MSCVneo)逆转录病毒表达系统和vIL10cDNA构建MSCVneovIL10重组体,在体外转导CBA(H2K)小鼠的造血干细胞,输入经致死量照射(900rad)的同基因CBA(H2K)小鼠体内,8周后以vIL10表达阳性的CBA(H2K)小鼠(12只,实验组)作为受体,C57BL/6(H2b)小鼠为供体,进行同种腹部异位心脏移植,以未处理、移植无病毒转染的造血干细胞(HSCs)、移植空MSCVneo转染的HSCs鼠(均为10只)为对照组。在移植后第8天,每组受体动物各杀死5只,取出移植心脏行组织病理学、CD+4与CD+8T淋巴细胞浸润(免疫荧光抗体法)检查,IL2、IL4、IL6、小鼠IL10(mIL10)、γ干扰素(IFNγ)、诱导型一氧化氮合酶(iNOS)、B71、B72mRNA表达的测定[逆转录聚合酶链反应(RTPCR)法],余下的动物观察移植心脏存活时间。结果(1)实验组移植心脏存活时间(800±333)d,其他对照组分别为(104±10),(116±11)与(112±17)d(P<001);(2)实验组心肌炎性细胞浸润稀少、心肌结构完整、无明显血管炎,急性排斥反应Ⅰ级,而其他对照组则出现严重的炎性细胞浸润、心肌灶性坏死、心内膜炎、血管炎和心肌出血,急性排斥反应均为Ⅲ级;(3)实验组移植物内IL2、IFNγ、B71、和B72和iNOS的mRNA表达明显下调,而其     

Monoclonal Antibody Specific for TIRC7 Induces Donor-specific Anergy and Prevents Rejection of Cardiac Allografts in Mice

T cell immune response c-DNA (TIRC7) is up-regulated during the early stages of T-cell activation in response to alloantigens. In this study, we analyzed the effects of newly developed monoclonal antibodies (mAb) against TIRC7 in acute cardiac allograft rejection. Fully vascularized heterotopic allogeneic heart transplantation was performed in mice across a full-mismatch barrier (C57Bl/10 into CBA). Recipients received seven injections (day 0-7) of a novel anti-TIRC7 mAb or remained untreated. Graft survival, histology and ex vivo lymphocyte functions were tested. Targeting of TIRC7 with an anti-TIRC7 mAb diminishes lymphocyte infiltration into grafts resulting in delay of morphological graft damage and prolongation of allograft survival. The lymphocytes from anti-TIRC7 mAb-treated animals exhibit hypo-responsiveness without evidence of lymphocyte depletion against the donor allo-antigens. Proliferation and expression of interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha) were down-regulated while interleukin-4 (IL-4) and IL-10 expression were spared. Moreover, anti-TIRC7 mAb enhanced up-regulation of CTLA-4 expression but suppressed up-regulation of CD25 on stimulated lymphocytes in vitro and in vivo. Ligation of TIRC7 has important effects on the regulation of co-stimulatory signaling pathways associated with suppressing of T-cell activation. Targeting of TIRC7 may therefore provide a novel therapeutic approach for modulating T cell immune responses during organ transplantation.     

Interleukin-10 fails to modulate low shear stress-induced neointimal hyperplasia.

INTRODUCTION: Overexpression of the anti-inflammatory cytokine interleukin-10 (IL-10) blocks atherosclerotic events in vivo, and IL-10 has been recently hailed as an "immunologic scalpel" for atherosclerosis. Alternatively, mice lacking IL-10 receiving atherogenic diets have increased occlusive lesions. It remains unclear whether such IL-10 modulation broadly applies to other forms of occlusive arterial remodeling. We hypothesized that lack of IL-10 would exacerbate, and exogenous or overexpression of IL-10 would abrogate low shear stress-induced neointimal hyperplasia (NIH). METHODS: Wild-type (WT) and IL-10-deficient (IL-10-/-) mice underwent unilateral common carotid artery (CCA) ligation. Low shear stress in the patent ligated artery results in remodeling and formation of neointima containing BrdU and SMC alpha-actin-positive cells. Additional groups of WT mice underwent CCA ligation and were treated daily with intraperitoneal saline or 1 microg of human IL-10. Chronic delivery gene therapy approaches were also utilized to define the role of IL-10 signaling. WT mice were treated adventitially with 1 x 10(10) adenovirus/green fluorescent protein (Ad/gfp) and an Ad/empty control to confirm the veracity of adventitial delivery. Then, Ad viral IL-10 (vIL-10), Ad/empty, and virus buffer alone were applied directly to the adventitia of the CAA immediately following ligation. In separate experiments, 1 x 10(10) Ad/empty or Ad/vIL-10 was injected intramuscularly. CCAs were perfusion fixed 28 days postligation, the time at which NIH is near maximum. RESULTS: IL-10-/- mice developed identical NIH areas compared to WT controls. Mice receiving IL-10 demonstrated NIH equivalent to saline controls. Mice receiving intramuscular or adventitial Ad/IL-10 developed high serum levels of IL-10 yet formed NIH areas similar to those of controls. Serum IL-10 levels were significantly higher (P = 0.04) with adventitial delivery. Mice treated adventitially with Ad/gfp showed reliable transfection of cells within the adventitia of CAA. No antibody to human IL-10 was found in the sera of intraperitoneal IL-10-treated mice, which failed to attenuate NIH. CONCLUSION: Under the conditions of this experiment, lack of IL-10 does not exacerbate low shear stress-induced NIH, nor does exogenous administration or overexpression of IL-10 attenuate it. Despite high serum levels of vIL-10 in mice treated with ad/vIL-10 adventitially, there appears to be no therapeutic effect despite the confirmed transfection of adventitial cells. Discrepancies between these findings and previous research may be related to IL-10 dosing, inflammation induced by the adenoviral vector, or disparities between the NIH models.     

Viral interleukin-10 gene therapy to induce tolerance to solid organ transplants in mice

In this study, a novel gene therapy approach to prolong allograft survival was designed. Autologous (syngeneic) hematopoietic stem cell-enriched bone marrow cells (HSC; lin(-)) engineered with the vIL-10 gene (vIL-10-HSC) were injected (4 to 6 x 10(6) cells, i.v.) into lethally (9.5 Gy) or sublethally (4 Gy) irradiated CBA/J mice 6 weeks prior to allogeneic heart (C57BL/6) transplantation (Tx). Cardiac allograft survival was significantly (P <.004) prolonged in lethally (71 +/- 40 days) and sublethally (114 +/- 15 days) irradiated mice that received vIL-10-HSC compared to controls that received no HSC (11 +/- 1 days), unengineered HSC, or vector-DNA-engineered HSC (12 to 16 days). Tolerant graft histopathology demonstrated mild arteritis/venulitis (grade 0.7) and rejection (grade 1.0). Intragraft expression of costimulatory molecules (B7.1, B7.2), cytokines (IL-2, IL-4, mIL-10, IFN-gamma), and iNOS molecules were markedly lower in tolerant grafts that survived for >100 days; recipient T lymphocytes demonstrated hyporeactivity to donor and third-party antigens in mixed lymphocyte cultures. These findings have important implications and potential therapeutic applications in transplantation and autoimmune diseases.     

Survival of fetal skin grafts is prolonged on the human peripheral blood lymphocyte reconstituted-severe combined immunodeficient mouse/skin allograft model

BACKGROUND: Fetal tissue is considered to be immune privileged and is under extensive investigation as a source of tissue for transplantation. In this paper, we analyzed the immune properties of human fetal and neonatal skin before and after transplantation to severe combined immunodeficient (SCID) mice. Using a human peripheral blood mononuclear cell reconstituted SCID (huPBMC-SCID) mouse model of allograft rejection, we compared the immune response to transplanted fetal and neonatal skin. METHODS: We analyzed human fetal (55-122 days of gestation) and neonatal skin samples by routine histology and immunohistochemistry for the expression of (MHC class I and II antigens before and after transplantation to SCID mice. After transplantation, we injected the mice with huPBMCs and analyzed the survival of neonatal and fetal skin grafts both visually and microscopically. RESULTS: We detected no class II expression in fetal skin of all gestational ages and only weak class I expression after 89 days compared with abundant class I and II expression in neonatal skin before transplantation. When transplanted to SCID mice, fetal skin grafts differentiated and expressed class I and II, but the levels were lower than neonatal grafts. In mice injected with huPBMCs, rejection of neonatal grafts started on day 5, and by day 9 all grafts were rejected. In contrast, rejection of fetal skin grafts was significantly delayed. Rejection started on day 13 and was complete by day 23 (P<0.00005). Histology sections from the rejected grafts showed marked CD3+ T cell infiltration in the human skin with a sharp demarcation between the human and mouse skin, with no T-cell infiltration in the mouse skin. CD4+ and CD8+ T cells were present in the rejected sites in similar densities. CONCLUSIONS: Our results show that fetal skin differentiates and expresses increased amounts of MHC class I and class II antigens when transplanted to SCID mice. However, these levels are much lower than the levels found in neonatal skin. We demonstrate that the survival of human fetal skin allografts is markedly prolonged compared with that of neonatal skin grafts in the huPBMC-SCID mouse model. Our results support the hypothesis that low levels of MHC antigen expression lead to a delay in the rejection of fetal skin and further demonstrate the utility of the huPBMC-SCID mouse model to investigate the molecular and cellular mechanisms of the immune response to human fetal tissues.     

病毒白细胞介素-10转基因在造血干细胞中的表达

目的　观察逆转录病毒转病毒白细胞介素 10 (vIL 10 )基因在体内的表达。方法　用MSCVneo vIL 10重组体在体外转导CBA (H 2 K)小鼠的造血干细胞 (HSCs) ,给经致死照射(90 0rads)的 2 0只同基因CBA(H 2 K)小鼠注入经MSCVneo vIL 10转染的HSCs ,2× 10 6HSCs/只。酶联免疫吸附测定 (ELISA)、逆转录 聚合酶链反应 (RT PCR )、Westernblot分析vIL 10的表达。结果　移植MSCVneo vIL 10转染HSCs的 2 0只小鼠 ,移植后 8周用ELISA检测 ,其中 15只小鼠血清的vIL 10浓度为 :2 70～ 13 40ng/L ,5只小鼠血清的vIL 10为阴性。 12周后有 2只小鼠vIL 10测不出 ,13只小鼠长时间表达vIL 10达 6个月。对照组小鼠血清vIL 10均为阴性。RT PCR和Westernblot证实小鼠的器官均有vIL 10的mRNA和蛋白的表达。结论　逆转录病毒能有效地将vIL 10基因导入造血干细胞并在体内长时间表达。     

Viral IL-10 Gene Transfer Inhibits the Expression of Multiple Chemokine and Chemokine Receptor Genes Induced by Inflammatory or Adaptive Immune Stimuli

Previously we have shown that adenovirus-mediated gene transfer and expression of vIL-10 are able to prolong cardiac allograft survival, through the inhibition of the immune response to both alloantigen and adenoviral antigens. In the current study, we have defined further mechanisms of Ad.vIL-10-mediated prolongation of cardiac allograft survival. E1- and E3-deleted adenoviral vectors encoding beta-galactosidase or vIL-10 were transferred into grafts at the time of transplantation, chemokine and chemokine receptor expression were evaluated by a pathway-specific cDNA array, and the results were confirmed with real time RT-PCR on selected genes. Ischemic injury, alloantigen and adenovirus vector induced the expression of multiple pro-inflammatory chemokines in the grafts, which likely amplify allograft rejection. Most of these Th1-related chemokine genes were inhibited or down-regulated by Ad.vIL-10 administration, which may help to decrease leukocytic infiltration and improve graft survival. Among the potent Th1 type chemokines inhibited were the CXCR3 ligands CXCL9 and CXCL10, which could directly inhibit vector-mediated gene expression in myoblasts, although targeting CXCR3 or its ligands did not prolong allograft survival with vIL-10 gene transfer. Ad.vIL-10 administration also induced the expression of the Th2-associated chemokines eotaxin-2 and MIP-1 gamma, suggesting Th1 to Th2 immune deviation. These results demonstrated that the vIL-10 gene transfer inhibits chemokine expression, preventing stimulation of innate and adaptive immunity.     

Comparison of Abeta(b-/-), H2-DM(-), and CIITA(-/-) in second-set skin allograft rejection.

BACKGROUND: Responses against donor MHC antigens are the major contributor to allograft rejection. Currently, it is unclear whether both direct and indirect recognition pathways are necessary and/or sufficient for allograft rejection. Previously, we found donor MHC class II and H2-DM to have dramatic effects on cardiac allograft survival. METHODS: Here, we used H2-DM(-) mice, which express CLIP-MHC class II complexes, and CIITA(-/-) mice, which lack all class II proteins, to examine the role of direct and indirect recognition on skin allograft rejection. Recipients were primed with donor cultured keratinocytes and later tested for accelerated memory response by challenge with full-thickness tail skin grafts. RESULTS: As previously reported, Abeta(b-/-) grafts survived longer than wild-type grafts, while H2-DM(-) grafts were rejected as rapidly as wild-type grafts. Skin grafts deficient for both beta2m and H2-DM survived longer than grafts lacking only H2-DM, but not as long as Abeta(b-/-) grafts. Additionally, CIITA(-/-) grafts survived as long as Abeta(b-/-) grafts. CONCLUSIONS: The delayed rejection of Abeta(b-/-) compared to H2-DM(-) suggests that indirect recognition of surface-expressed donor MHC class II is sufficient to mediate rapid skin allograft rejection. The equivalent survival of CIITA(-/-) and Abeta(b-/-) grafts suggests that indirect presentation of donor class II molecules (Aalpha or Ebeta) present in Abeta(b-/-) but not CIITA(-/-) mice does not contribute to graft rejection. These results reveal a modest role for surface-expressed donor class II in primed keratinocyte rejection, but also reveal a dramatic contrast to the cardiac allograft system and indicate tissue/organ-specific mechanisms of rejection.