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Abstract: The immune system maintains self-tolerance through the interplay of several mechanisms. One of the key mechanisms is the suppression of effector cells through regulatory T cells (Tregs). Recent progress has allowed the identification, analysis, and experimental manipulation of various subpopulations of Tregs. Several major issues regarding Tregs have emerged in relation to organ transplantation, including the questions of whether immunosuppressive drugs influence Tregs and whether Tregs can be used therapeutically to achieve graft acceptance or even tolerance. Although none of the available immunosuppressive drugs have been specifically designed to influence Tregs, they nevertheless do so through various pathways. Notably, current evidence suggests that rapamycin has a favorable effect on Tregs in organ transplant recipients, whereas calcineurin inhibitors (CNI) have markedly negative effects on Tregs. The idea of inducing tolerance through the therapeutic administration of Tregs has received much attention in recent years. To obtain sufficient numbers of recipient-derived Tregs, various strategies are being investigated to expand sorted Treg populations. Rapamycin has been found to significantly promote the in vitro expansion of murine and human Tregs; it may thereby help to generate clinically relevant quantities of Tregs. However, no data have yet been published that would demonstrate that the therapeutic use of Tregs induces transplantation tolerance across full major histocompatibility complex barriers in immunocompetent mice displaying a polyclonal T-cell repertoire. [Copyright &y& Elsevier]

ECP is used since more than 25 years as a rescue therapy in patients with refractory acute cellular rejections (ACR), as an add on treatment in antibody mediated rejection and in patients with CLAD with significant response.Therefore a prospective randomized single centre study was initiated to evaluate the safety and efficacy of adding ECP to standard immunosuppression in preventing ACR episodes in the first 12 months after transplantation. A total of 62 consecutive adult recipients of primary LuTX at the lung transplant center Vienna, Austria (Medical University Vienna) were randomly assigned to receive either standard triple-drug immunosuppressive therapy (Tacrolimus, Mycophenolate Mofetil and prednisone, n=31) or standard triple-drug therapy plus photopheresis (n=31). Primary objective was the incidence of biopsy proven acute cellular rejection episodes in the first 12 month after transplantation. After 12 months of follow-up the mean number acute cellular rejection rates was significantly lower in the ECP group (less high grad rejection episodes ≥A2: mean±SD control group 0.48±0.67 vs ECP group 0.06±0.24, p=0.003) with less need for aggressive rejection therapy (high dose of steroids or ATG). The cumulative A scores after 12 months were 1.0±1.45 in the control group and 0.25±0.48 in the ECP group (p=0.002). The cumulative B scores after 12 months were 1.4±1.06 in the control group and 0.8±0.78 in the ECP group (p= 0.019).Furthermore the number of severe infectious complications was significantly lower in the ECP group (22 vs 5, p=0.002) due to the reduced burden of immunosuppression treatment and the immunomodulatory effects of ECP (increase of Tregs, ⋯.). No difference in survival and CLAD was observed. Addition of ECP to standard triple immunosuppression is associated with a significant lower incidence of acute cellular rejection episodes and a significant reduction of infectious complications after lung transplantation within the first post transplant year. [ABSTRACT FROM AUTHOR]

Alemtuzumab is a monoclonal antibody targeting CD52, one of the most predominant antigens on the surface of lymphocytes. It induces cell lysis and leads to a profound immune cell depletion. However, little is known how T- and B-cell repopulate and if there is predominance towards certain subtypes. B- and T cell subsets were analyzed from whole blood samples of 25 patients, prospectively collected at time of listing (=baseline) and 6 months post-transplant. Cell subsets were measured by FACS. Patients received alemtuzumab induction followed by low-dose tacrolimus and steroids. Median age of the cohort was 59 years (51-64) and 36% of patients were female. Underlying diagnoses were COPD (36%), fibrosis (20%), CF (12%), PPH (4%), EAA (12%) and others (16%). B- and T cell counts significantly decreased compared to baseline (p=0.009, p<0.001). Within B cell population, relative frequency of IgM+CD27−CD38high, IgM+CD27+CD38high and regulatory B cells increased 6 months after Tx (p=0.017, p=0.016, p=0.028, respectively). Within T cell population, CD4+ T cells decreased 6 months after Tx (p<0.001), while relative frequency of CD8+ T cells was significantly higher (p<0.001). A detailed analysis revealed a shift towards CD4+CD57+(p=0,002), PD1+CD4+(p<0.001), CD45RA+CD197-CD4+ (p=0.0013), CD45RA−CD197−CD4+ (p<0.001), CD27−CD28+ (p<0.001), CD27+CD28+ (p<0.001), PD1+CD8+ (p<0.001), CD45RA+CD197−CD8+ (p=0.039) and regulatory FoxP3+-T cells (p=0.005) at 6 months after Tx. After alemtuzumab induction therapy, the composition of B and T cell subsets is altered compared to baseline. Expansion of subsets such as PD1 T cells and regulatory B- and T cells suggests an immunomodulatory effect of alemtuzumab on cell repopulation. [ABSTRACT FROM AUTHOR]

Extracorporeal photopheresis (ECP) is an accepted treatment option for acute and chronic rejection after solid organ transplantation but the mode of action is not fully elucidated. In a randomized controlled trial that investigates the addition of prophylactic use of ECP to a tacrolimus-based immunosuppressive regimen after lung transplantation, a comprehensive phenotypic immune monitoring is performed to assess the immunomodulatory effects of ECP. To date, 40 (from a calculated sample size of 62) end-stage chronic obstructive pulmonary disease (COPD) patients underwent bilateral lung transplantation and were randomized into 2 treatment arms - the control group received standard triple immunosuppression consisting of tacrolimus, mycophenolate mofetil and steroids while the second group additionally received 16 ECP treatments in the first three months after surgery. For monitoring a comprehensive set of leukocyte subsets such as regulatory T cells (CD4+CD25+FoxP3+ Tregs) polychromatic flow cytometry analysis was performed on fresh whole blood samples using validated, standardized, lyophilized monoclonal antibody panels (DuraClone). Samples were acquired before and 3 months after transplantation, when the last ECP treatment was performed, as well as 6 months after transplantation. Statistical analysis was conducted by using a two-tailed paired and unpaired t-test. From 40 double lung transplanted patients, 32 patients have reached their three-month-visit and 28 patients their 6-months-visit. No significant difference in the frequency of Tregs was found between both groups at baseline (p=0.425). At 3 months, Tregs have significantly decreased in the control group (p=0.009) while in the ECP-treated group no significant decline was seen (p=0.141). Six months after transplantation the frequency of Tregs was significantly decreased in both groups when compared to baseline (p=0.043 in control; p=0.030 in ECP-treated). Preliminary data from this interim analysis of double lung transplanted patients receiving ECP as prophylactic treatment provide evidence that during ECP treatments the post-transplant decline in Treg frequency seen with standard immunosuppression is prevented. [ABSTRACT FROM AUTHOR]

Abstract: Background: Cellular immunity plays a major role in rejection of xenografted islets. Depending on the phylogenetical disparity, direct or indirect antigen presentation is predominant. The aim of this study was to analyze in vitro the predominance of direct or indirect presentation, and in vivo the effect of macrophage depletion on concordant and discordant islet xenograft survival. Materials and methods: In vitro, we performed mouse antirat and mouse antihuman mixed lymphocyte reactions (MLR) after depletion of responder or stimulator antigen-presenting cells. In vivo, streptozotocin-induced diabetic C57BL/6 mice were treated by gadolinium chloride to deplete macrophages and rat or human islets were transplanted under the kidney capsule. Islet function was followed by glycemia and xenografts were analyzed at regular intervals for histology. Results: Mouse antirat MLR showed a predominant direct antigen presentation pathway, whereas in mouse antihuman MLR, direct and indirect pathways were similarly involved. Survival of rat islets was not modified by GdCl therapy. In contrast, survival of human islets was significantly prolonged in GdCl-treated mice. Macrophage infiltration was decreased in concordant and discordant GdCl-treated xenografts at day 4, compared to controls. At day 15, macrophage infiltration was similar in all groups. Discussion: Our results indicate that direct antigen presentation is dominant in rejection of concordant islet xenografts and cannot be influenced by host macrophage depletion. Both direct and indirect antigen presentation are involved in rejection of discordant xenogeneic islets. Macrophage depletion or inhibition should be considered as therapeutic tool for discordant islet xenotransplantation. [Copyright &y& Elsevier]

Abstract: Background: Costimulatory blockade has been shown to allow long-term survival of xenogeneic islets. The aim of the present study was to evaluate the role of recipient CD40 and CD154 in the rejection process of concordant and discordant islet xenotransplantation (Tx). Methods: Diabetic C57BL/6 mice, CD40- or CD154 knockout (KO) mice were transplanted with either concordant rat or discordant human islets. Experimental design: group 1, control (ie, C57BL/6 mice received islet Tx without therapy); group 2, C57BL/6 mice received islet Tx with anti-CD154 monoclonal Ab (mAb) therapy; group 3, CD40 KO mice; and group 4, CD154 KO mice were used as recipients without therapy. Mouse anti-rat mixed lymphocyte reactions (MLR) were performed using mouse splenocytes obtained from animals transplanted with rat islets in groups 1 to 4. Results: In group 2, short-term anti-CD154 mAb therapy significantly prolonged rat-to-mouse and human-to-mouse xenograft survival, compared to controls. In CD40-KO and CD154-KO recipients, survival of concordant or discordant islets was not prolonged significantly compared to control groups. Mouse anti-donor rat cellular responses were reduced approximately 50% in group 2 but remained unmodified in groups 3 and 4, when compared to group 1. Conclusions: Improved graft survival and reduced MLR responses against donor cells in vitro among the anti-CD154 mAb-treated mice could be explained by specific targeting of activated T cells with subsequent inactivation by anergy and/or elimination by apoptosis, or complement- or cellular-mediated mechanisms. Rejection of xenografts and strong MLR responses against donor cells in vitro in CD40 or CD154 KO animals is possible through efficient activation of alternate pathways of costimulation. [Copyright &y& Elsevier]