Background-Donor brain death upregulates expression of inflammatory mediators in the heart. It is hypothesized that these nonspecific changes trigger and amplify acute rejection in unmodified recipients compared with hearts from normal living donors. We examined the inflammatory and immunological consequences of gradual-onset donor brain death on cardiac allografts after transplantation. Methods and Results-Functioning hearts were engrafted from normotensive donors after 6 hours of ventilatory support.Hearts from brain-dead rats (Fisher, F344) were rejected significantly earlier (meanϮSD, 9.3Ϯ0.6 days) by their (Lewis) recipients than hearts from living donor controls (11.6Ϯ0.7 days, Pϭ0.03). The inflammatory response of such organs was accelerated, with rapid expression of cytokines, chemokines, and adhesion molecules and brisk infiltration of associated leukocyte populations. Upregulation of major histocompatibility class II antigens increased organ immunogenicity. Acute rejection evolved in hearts from brain-dead donors more intensely and at a significantly faster rate than in controls. Conclusions-Donor brain death is deleterious to transplanted hearts. The resultant upregulation of inflammatory factors provokes host immune mechanisms and accelerates the acute rejection process in unmodified hosts. (Circulation.

ObjectiveTo define the potential influences of donor brain death on organs used for transplantation. Summary Background DataDonor brain death causes prompt upregulation of inflammatory mediators on peripheral organs. It is hypothesized that this antigen-independent insult may influence the rate and intensity of host alloresponsiveness after engraftment. MethodsThe rates of survival of unmodified Lew recipients sustained by kidney allografts from brain-dead, normal anesthetized, and anesthetized ventilated F344 donors were compared. Brain death was induced by gradually increasing intracranial pressure under electroencephalographic control. Tracheotomized brain-dead animals and anesthetized controls were mechanically ventilated for 6 hours before transplant nephrectomy. The rate and intensity of the acute rejection event were examined by histology, immunohistology, and reverse transcriptase-polymerase chain reaction. ResultsAnimals bearing kidneys from brain-dead donors died of renal failure secondary to acute rejection at a significantly faster rate than those from anesthetized living controls or anesthetized animals ventilated for 6 hours. Within 3 hours after placement and reperfusion of brain-dead donor grafts, significant neutrophil infiltration was observed, followed by increasing numbers of macrophages and T cells. mRNA of proinflammatory mediators detected in kidneys within 6 hours of brain death and upregulated even before transplantation increased thereafter and appeared to accelerate and amplify host alloresponsiveness, as manifested by the rapid expression of chemokines, cytokines, adhesion molecules, and major histocompatibility complex class II antigens in the engrafted organ. The process evolved in the controls less intensely and at a slower rate. ConclusionsDonor brain death is a significant risk factor for peripheral organs used for transplantation. The activated state of such organs appears to trigger host immune mechanisms that accelerate the process of acute rejection. The effects of this central injury may explain in part the less satisfactory performance of cadaver organs in human transplantation compared with those from living sources.Transplantation has evolved as the treatment of choice for many patients with end-stage organ disease. Success has improved progressively, particularly in the short term. However, despite a functional survival rate of most grafts of more than 80% at 1 year, the ultimate goal of providing long-term treatment for an irreversible process has not been achieved; the rate of attrition over time has remained relatively stable throughout the entire experience.1 Whereas early immune-mediated injury is considered to be primarily responsible for graft dysfunction and failure, the influence of nonimmunologic, antigen-independent events may have been underestimated; there appears to be a continuum between inflammatory changes secondary to an initial nonspecific insult and the onset of alloresponsiveness. This concept has been emphasized by recent pooled data from the United...

IntroductionRegulatory T cells (Tregs) play a dominant role in transplantation tolerance, as shown by adoptive cell transfer for the prevention of organ graft rejection and GVHD in experimental models. [1][2][3][4][5][6] Translating Treg therapies to humans requires cell isolation and purification, because transfer of the accompanying effector T cells would exacerbate rather than ameliorate human disease. Naturally occurring Treg populations comprise 1%-3% of the human T-cell repertoire. 7 Tregs are characterized by the constitutive expression of CD4, the IL-2 receptor ␣ chain (CD25), and high levels of a nuclear transcription factor, forkhead box p3 (Foxp3), which is critical for their development and suppressive function. [8][9][10] Tregs express little or no IL-7 receptor ␣ chain (CD127). 11 Therefore, the constitutive expression of CD25 and low expression of CD127 have been used for identifying and purifying Tregs. 12,13 Immunomagnetic cell-separation technology has been adapted for this application. Adoptive transfer of freshly isolated human Tregs has prevented GVHD in patients treated with allogeneic hematopoietic stem cell transplantations partially depleted of conventional T cells. 14 However, T cell-replete grafts are more commonly used, and prevention of GVHD after T cell-replete transplantations require the transfer of a higher number of Tregs that can only be obtained through ex vivo expansion.Tregs can be expanded by more than 1000-fold by anti-CD3 and anti-CD28 Ab-coated beads, but loss of regulatory function and acquisition of effector functions have been a concern with this technology. The addition of rapamycin to artificial APCs has apparently solved this problems, because Tregs maintained Foxp3 expression and in vivo suppressive function in immunodeficient mice. 15 Because in vivo Treg protection from GVHD requires alloantigen-specific (allo-specific) engagement of the TCR, 16 adoptive immunotherapy with allo-specific Tregs offers more advantages than polyspecific Tregs: selective rather than broad immunosuppression, ability to control the conditions for antigen presentation, and economy of scale because they are a fraction of the entire Treg population. Antigen-specific Tregs expanded in response to dendritic cells (DCs) presenting a single self-peptide 17 and suppressed experimental autoimmune diabetes more efficiently than polyclonal Tregs. 18 Human allospecific Tregs could also expand ex vivo, 19 but only by 10-to 20-fold. 18 Because Tregs constitute 1%-3% of human blood T cells, and effective suppression requires a Treg:effector T cell ratio of 1:1, we surmise that therapeutic applications will require expanding allo-specific Tregs by over 100-fold. 20 There is a need to improve on expansion protocols before allo-specific Tregs can be brought to the clinic. Sagoo et al 21 have recently adopted a 2-step process for the large-scale expansion of allo-specific Tregs: primary Treg activation by DCs presenting the specific alloantigen directly, followed by sorting Tregs for expression of the a...

It is highly desirable that immature dendritic cells (DC) used for tolerance induction maintain steady immature state with predominant interleukin (IL)-10 production. In this study, we attempted to develop DC with durable immaturity and other tolerogenic features by using dexamethasone (Dex). We found DC derived from human monocytes in the presence of 10 À7 M Dex were negative for CD1a. Compared with control transduced DC (Ctrl-DC), Dex-DC expressed lower CD40, CD80 and CD86 but equivalent human leucocyte antigen-DR. Both immature Dex-and Ctrl-DC did not express CD83. Nevertheless, upon stimulation of lipopolysaccharide (LPS) or CD40 ligand, the expression of CD40, CD80, CD83 and CD86 was upregulated on Ctrl-DC but not on Dex-DC. The immaturity of Dex-DC was durable following Dex removal. Interestingly, Dex-DC maintained production of large amount of IL-10 and little IL-12 five days after Dex removed. Further study indicated that high-level IL-10 production by Dex-DC was associated with high-level phosphorylation of extracellular signal-regulated kinase (ERK) as blockade of this enzyme markedly attenuated IL-10 production. Furthermore, Dex-DC sustained the capability of high phosphorylation of ERK and IL-10 production 5 days after Dex removal. In addition, Dex-DC had significantly lower activity in stimulating T-cell proliferation. Neutralization of IL-10, to some extent, promoted DC maturation activated by LPS, as well as T-cell stimulatory activity of Dex-DC. The above findings suggest that IL-10-producing Dex-DC with durable immaturity are potentially useful for induction of immune tolerance.