Minor Mismatch Research Articles

Plasmons in graphene moiré superlattices.

Moiré patterns are periodic superlattice structures that appear when two crystals with a minor lattice mismatch are superimposed. A prominent recent example is that of monolayer graphene placed on a crystal of hexagonal boron nitride. As a result of the moiré pattern superlattice created by this stacking, the electronic band structure of graphene is radically altered, acquiring satellite sub-Dirac cones at the superlattice zone boundaries. To probe the dynamical response of the moiré graphene, we use infrared (IR) nano-imaging to explore propagation of surface plasmons, collective oscillations of electrons coupled to IR light. We show that interband transitions associated with the superlattice mini-bands in concert with free electrons in the Dirac bands produce two additive contributions to composite IR plasmons in graphene moiré superstructures. This novel form of collective modes is likely to be generic to other forms of moiré-forming superlattices, including van der Waals heterostructures.

UGT2B17 minor histocompatibility mismatch and clinical outcome after HLA-identical sibling donor stem cell transplantation

Minor histocompatibility Ags (mHags) have been implicated in the pathogenesis of GVHD after allogeneic hematopoietic stem cell transplantation (HSCT). Uridine diphospho-glucuronosyltransferase 2B17 (UGT2B17) gene deletion may act as a mHag and its association with acute GVHD (aGVHD) has been described. We retrospectively studied the clinical impact of a UGT2B17 mismatch in a cohort of 1127 patients receiving a HSCT from an HLA-identical sibling donor. UGT2B17 mismatch was present in 69 cases (6.1%). Incidence of severe aGVHD was higher in the UGT2B17 mismatched pairs (22.7% vs 14.6%), but this difference was not statistically significant (P: 0.098). We did not detect differences in chronic GVHD, overall survival, relapse-free survival, transplant-related mortality or relapse. Nevertheless, when we analyzed only those patients receiving grafts from a male donor (616 cases), aGVHD was significantly higher in the UGT2B17 mismatched group (25.1% vs 12.8%; P: 0.005) and this association was confirmed by the multivariate analysis (P: 0.043; hazard ratio: 2.16, 95% confidence interval: 1.03-4.57). Overall survival was worse for patients mismatched for UGT2B17 (P: 0.005). We conclude that UGT2B17 mismatch has a negative clinical impact in allogeneic HSCT from HLA-identical sibling donors only when a male donor is used. These results should be confirmed by other studies.

Detuning Effect Between the Cavity Resonance and Atomic Transition Frequencies on Noise Flux Profiles of Class-C Lasers

The class-C lasers are well recognized to have very narrow atomic transition bandwidths. Therefore, a minor mismatch between the cavity resonance and atomic transition frequencies (CAF) is normally led to the large modifications in the gain and noise profiles. The motion of class-C lasers is described by the three variables of cavity electric field, atomic population inversion, and dipole moment of atoms that are coupled to each other by the optical Maxwell-Bloch equations. The fluctuating features of these three variables are completely elaborated by solving their equations of motion in the presence of the cavity Langevin force. The results peculiarly indicate that the noise profile of the cavity electric field exactly mimics the amplification profile of an input signal by the laser in both below and above threshold states. The both gain and noise profiles simultaneously tend to infinity at the normalized pumping rates and CAF mismatches determined by the laser stability theory. The noise flux profiles are finally confirmed by illustrating the flux conservation.

Advanced control with parameter estimation of batch transesterification reactor

The objective of this work is to enhance the economic performance of a batch transesterification reactor producing biodiesel by implementing advanced, model based control strategies. To achieve this goal, a dynamic model of the batch reactor system is first developed by considering reaction kinetics, mass balances and heat balances. The possible plant-model mismatch due to inaccurate or uncertain model parameter values can adversely affect model based control strategies. Therefore, an evolutionary algorithm to estimate the uncertain parameters is proposed. It is shown that the system is not observable with the available measurements, and hence a closed loop model predictive control cannot be implemented on a real system. Therefore, the productivity of the reactor is increased by first solving an open-loop optimal control problem. The objective function for this purpose optimizes the concentration of biodiesel, the batch time and the heating and cooling rates to the reactor. Subsequently, a closed-loop nonlinear model predictive control strategy is presented in order to take disturbances and model uncertainties into account. The controller, designed with a reduced model, tracks an offline determined set-point reactor temperature trajectory by manipulating the heating and cooling mass flows to the reactor. Several operational scenarios are simulated and the results are discussed in view of a real application. With the proposed optimization and control strategy and no parameter mismatch, a revenue of 2.76$min−1 can be achieved from the batch reactor. Even with a minor parameter mismatch, the revenue is still 2.01$min−1. While these values are comparable to those reported in the literature, this work also accounts for the cost of energy. Moreover, this approach results in a control strategy that can be implemented on a real system with limited online measurements.

Anti-A and anti-B: what are they and where do they come from?

Intravenous immunoglobulin (IVIG) is made from thousands of donors having a variety of blood groups. All of the donors being used for IVIG production, with the exception of group AB donors, have in their plasma antibodies of variable titer commonly known as isohemagglutinins or ABO antibodies. As blood groups O and A are the most commonly found in the world population, most of the plasma used in IVIG production is from donors having these blood groups, with group B and group AB donors being fewer in number. Consequently, all batches of IVIG contain antibodies that are reactive with individuals of group A, group B, and group AB. These antibodies were originally discovered by Dr Karl Landsteiner in the early 1900s and are now known to consist of immunoglobulin (Ig)M, IgG, and IgA classes. As the process for producing IVIG results in almost exclusively IgG, isohemagglutinins contained in IVIG are of this immunoglobulin class. ABO antibodies are highly clinically significant and, because of this, blood bank cross-matching is done to ensure that blood of the correct type is transfused into recipients to avoid a so-called major mismatch or major incompatibility that can cause significant morbidity and often death. Administration of IVIG, which contains ABO antibodies, is often infused into individuals who have the corresponding ABO antigens, commonly called a minor mismatch, and although not as significant as a major mismatch, the isohemagglutinins contained in the IVIG have some risk for a significant transfusion reaction due to the ABO incompatibility. Indeed, currently there is no way to match IVIG to recipients according to blood type, so when IVIG is administered to group A, B, or AB recipients, there is potential for transfusion reactions analogous to a blood transfusion mismatch. For this reason, strict guidelines have been put into place to restrict the titers of the ABO antibodies contained in IVIG. This review will provide background information about the discovery and biochemistry of the ABO antigens and discuss the various isohemagglutinins that are found in plasma of the different ABO blood types and their potential clinical significance. In addition, a brief discussion of the controversial topic of the origins of these antibodies will conclude this review.

B7-H1 regulates autoimmunity in αCTLA-4, bm12 transfer and pristane-induced models through distinct sexually dimorphic and non-dimorphic effects (BA4P.128)

Abstract Females have more autoimmunity through poorly understood mechanisms. Regulatory T cells (Tregs) defend against autoimmunity. We show that B7-H1 signals dampen estrogen (E2)-driven Treg mTOR activation through estrogen receptor-β, maintaining Treg function in E2. B7-H1-/- female mice (but not males or WT of either sex) developed αCTLA-4-induced autoimmunity, rescued by E2 removal or B7-H1+/+ dendritic cell (DC) transfer. Monocytes or B or T cells also gave protective B7-H1 signals to restore Treg function in E2. B7-H1-mediated E2 protection was during Treg differentiation and was not via CD80, PD-1 or Treg B7-H1. E2 also reduced male B7-H1-/- Treg function in vitro and in vivo. Sex-matched B7-H1-/-bm12 splenocyte transfer (minor MHC mismatch) caused lupus symptoms (albuminuria, alopecia) earlier in B7-H1-/- females than B7-H1-/- males or WT. B7-H1-/- males also sickened faster than WT males, showing sex-independent B7-H1 effects. Transfer of B7-H1+/+bm12 splenocytes into B7-H1-/- mice did not induce rapid symptoms, suggesting rescue by B7-H1 signals in bm12 cells. In pristane-induced lupus, B7-H1-/- females had earlier proteinuria than WT females. Plasmacytoid DC or granulocytes from female B7-H1-/- produced more IFN-α in vitro with pristane+TLR7 agonist than WT females. Thus, B7-H1 regulates autoimmunity through diverse sexually-dimorphic and non-dimorphic mechanisms. Defining B7-H1 effects in autoimmunity will help understand female predilection and immunopathogenesis.

Impact of ABO blood group mismatch in alemtuzumab-based reduced-intensity conditioned haematopoietic SCT.

The impact of ABO incompatibility on clinical outcomes following haematopoietic SCT (HSCT) remains controversial. This retrospective study assessed the effect of ABO mismatch on transplant outcomes and transfusion requirements in 594 patients undergoing reduced-intensity conditioned (RIC) HSCT with alemtuzumab in three UK transplant centres. We found no significant effects of minor, major or bidirectional ABO mismatch on overall survival, relapse-free survival, nonrelapse mortality or relapse incidence. Although the rate of acute GVHD was unaffected by ABO mismatch, the incidence of extensive chronic GVHD was higher in patients with minor and major mismatch compared with those who were ABO matched (hazard ratio (HR) 1.74, P=0.032 for minor, HR 1.69 P=0.0036 for major mismatch). Red cell and platelet transfusion requirements in the first 100 days post transplant did not differ by ABO mismatch. In this large UK series, ABO mismatch in RIC HSCT has no clinically significant effect on survival outcomes but appears to modify susceptibility to extensive chronic GVHD.

CD19 CAR T Cells Maintain Efficacy in the Allogeneic Environment but Mediate Acute Graft-Versus-Host-Disease Only in the Presence CD19+ Acute Lymphoblastic Leukemia

A significant portion of patients with relapsed acute lymphoblastic leukemia (ALL) who are eligible for treatment with chimeric-antigen-receptor (CAR) T cells have undergone a previous hematopoietic stem cell transplantation. We and others have previously demonstrated that the allogeneic environment, even in the presence mild graft-versus-host disease (GVHD), severely impairs tumor-directed T-cell immunity. To study the behavior of CAR cells in the allogeneic setting we chose a murine model of minor mismatched allogeneic T-cell depleted bone marrow transplantation (BMT) followed by CAR T (CD19-28-z) cell infusion in recipients bearing a murine B-precursor ALL model (positive for CD19). Importantly, the leukemia persists following myeloablative radiation thus mimicking post-transplant minimal residual disease. Donor-derived, post-transplant injection of CD19 CAR T cells eliminated residual leukemia in the syngeneic as well as the allogeneic settings. CD19 CAR T cells harvested from allogeneic recipients maintained in-vitro ability to produce IFN-gamma and degranulate in the presence of ALL ex vivo, at levels comparable to syngeneic CD19 CAR T cells. However, CAR T cells administered in the allogeneic environment had the potential to mediate severe acute GVHD with early mortality of recipients not typically seen in this minor mismatch model. This occurred across multiple T cell doses capable of clearing leukemia (0.3e6-5e6), in transduced CD19 CAR T cells generated from donors tolerized to allogeneic antigens, and when CD19 CAR T cell infusion was delayed following BMT. In-vivo tracking of transferred cells showed comparable expansion and persistence of the CD19 CAR T cells in the allogeneic and syngeneic environments. However, syngeneic CAR T cells tended to develop later into central memory T cells (CD62L+CD44+), whereas the profile of allogeneic cells was significantly skewed toward effector T cells (CD62L-CD44+). Remarkably, the process of CAR-driven acute GVHD in this minor mismatch model was only seen in the presence of leukemia, indicating a CAR-target interaction influences the induction of GVHD. Indeed, pro-inflammatory cytokines IFN-gamma and IL-6 were elevated only in the presence of both ALL and CAR T cells, whereas TNF alpha levels were undetectable in all instances. We are currently testing whether neutralization of cytokines can prevent GVHD in these models. Altogether, these data demonstrate efficacy of CD19 CAR to clear residual leukemia in an immunocompetent mouse model, and maintain initial cytotoxicity despite the potentially suppressive allogeneic environment. However, we demonstrated potential risks of allogeneic CAR T cells aggravating GVHD in the presence of residual leukemia, likely via cytokine-mediated manner. Clinically, as IL-6 was shown to be significant in cytokine release syndrome, this may represent a murine model to study potential interventions. DisclosuresNo relevant conflicts of interest to declare.

MiR-146a Regulates the TRAF6/TNF-Axis in Donor T Cells during Graft-Versus-Host Disease

▪Introduction: Acute graft-versus-host disease (GvHD) arises from the attack of recipient tissues by donor allogeneic T cells and represents one of the major limitations of allogeneic hematopoietic cell transplantation (allo-HCT). In spite of many clinical trials, the standard immunosuppressive regimens for prevention of acute GvHD have improved little in the last two decades. Hence, a better understanding of the biology of acute GvHD may improve therapeutic options. MicroRNA-146a (miR-146a) was found to be increased in the sera of patients with GvHD. Therefore, we aimed to decipher the role of miR-146a in allogeneic donor T cells during GvHD by functional studies and in patients undergoing allo-HCT by single nucleotide polymorphism (SNP) analysis.Methods: We used two different murine major MHC mismatch models for acute GvHD. Recipient mice were conditioned with irradiation before transplantation of bone marrow and either wildtype or miR-146a deficient T cells from allogeneic donor mice. Furthermore, genomic DNA from 289 patients that underwent allo-HCT and their respective hematopoietic stem cell donors was isolated in order to determine their miR-146a rs2910164genotype.Results: We observed miR-146a upregulation in T cells of mice developing acute GvHD compared to untreated mice in a major MHC and a minor histocompatibility antigen mismatch model. Transfer of miR-146a deficient T cells caused increased GvHD severity, elevated TNF serum levels and reduced survival. Conversely, the phytochemical induction of miR-146a or its overexpression in donor T cells using a specific miR-146a mimic reduced GvHD severity. TNF receptor-associated factor 6 (TRAF6), a verified target of miR-146a, was upregulated in miR-146a-/- T cells following alloantigen stimulation. Higher TRAF6 levels translated into increased NF-κB activity and TNF production in miR-146a-/- T cells, while other pro-inflammatory cytokine levels were unaffected. The detrimental effect of miR-146a deficiency in T cells could be antagonized by TNF blockade in vivo. Moreover, in contrast to WT T cells, over expression of miR-146a in Tnf deficient T cells had no effect on their alloreactivity. In the human system, the minor genotype of the SNP rs2910164, which causes reduced miR-146a expression, was more frequent in patients developing acute GvHD grade III/IV compared to all other allo-HCT recipients (n=289).Conclusions: Taken together we show that miR-146a functions as a negative regulator of the TRAF6/TNF-axis in allogeneic donor T cells during GvHD, leading to reduced TNF transcription. Given our observation on the predictive role of the SNP leading to decreased miR-146a expression in acute GvHD in patients and the possibility to exogenously enhance miR-146a expression, we provide a novel and targeted molecular approach to mitigate GvHD. DisclosuresNo relevant conflicts of interest to declare.

MyD88 in Donor Bone Marrow Cells Is Critical for Protection from Acute Intestinal Graft-Versus-Host Disease

Allogeneic hematopoietic stem cell transplantation is an important therapeutic modality used to treat malignancies of hematopoietic origin, such as leukemia and lymphoma. However, development of graft-versus-host disease (GVHD) causes non-relapse mortality and substantial morbidity of recipients. Myeloid differentiation factor 88 (MyD88), a major adaptor mediating TLR signaling, is also known to deliver pro-inflammatory signals. Activation of inflammatory signaling through MyD88 plays a key role in the expansion of myeloid-derived suppressor cells (MDSC) which are a heterogeneous population of immature myeloid ells with anti-inflammatory activity. To explore the contribution of MyD88 expressed by donor bone marrow (BM) cells to development of GVHD, we induced GVHD using T-cell-depleted BM (TCD-BM) isolated from MyD88-deficient (MyD88KO) mice and T cells isolated from wild-type (WT) mice. We employed C57BL/6 (H-2b) → B6D2F1 (H-2b/d) mouse model of GVHD, which differ at major and minor histocompatibility loci. Lethally irradiated B6D2F1 recipient mice were transplanted with either T cell-depleted bone marrow (TCD BM, 5 x 106) from either WT or MyD88KO mice together with WT spleen T cells (1 x 106). Transplantation with MyD88KO TCD BM aggravated GVHD; serious gut damage was evident, with infiltration of T cells specifically into the intestines of recipients. GVHD hosts transplanted with MyD88KO TCD BM exhibited markedly reduced expansion of MDSC. GVHD aggravation after transplantation with MyD88KO TCD-BM, associated with high-level T cell infiltration into the intestine and insignificant expansion of MDSC, was reproduced in another minor histocompatibility mismatch model (C57BL/6 → BALB.B). We next examined allogeneic T cells in the spleens of GVHD hosts in terms of the expression levels of CCR9, which are known to be associated with T cell migration to the intestinal mucosa and the proportion of CCR9 positive cells in CFSE low CD8+ T cells was higher in recipients of MyD88KO TCD BM than WT controls. In parallel, the levels of CCL25 were more highly expressed in the gut of MyD88KO recipients than WT controls. Mixed leukocyte cultures of CFSE- labeled C57BL/6 T cells and irradiated B6DF1 feeder cells were prepared in the presence of MDSC isolated from MyD88KO or WT mice. T cells, co-incubated with MDSC isolated from MyD88KO BM, exhibited a greater extent of CFSE dilution and less Annexin V staining, compared to T cells co-incubated with cells from WT BM. Moreover, MDSC from recipients of MyD88KO TCD BM exhibited a reduced suppressive function, compared to their WT counterparts. Next, we determined whether insufficient expansion of and ineffective suppression by MDSC caused severe GVHD in recipients of MyD88KO TCD BM. Supplementation of transplanted mice with MDSC from WT mice, not from MyD88KO mice, attenuated the severity of GVHD and reduced intestinal T cell infiltration in recipients of MyD88KO TCD BM. To verify the importance of MyD88-mediated signaling by MDSC in protection against severe GVHD, we determined if transplantation with TCD-BM cells containing high levels of MDSC attenuated the severity of GVHD. Pre-treatment of BM donors with lipopolysaccharide increased the frequencies of MDSC and the amounts of MyD88 transcripts in TCD-BM transplant, and alleviated the severity of GVHD and intestinal T-cell infiltration. To explore whether MDSC expansion levels could be used to predict the severity of intestinal GVHD, the T/MDSC ratios were calculated in blood of patients at the time of engraftment and were significantly higher in patients with intestinal GVHD ≥ grade 2. In conclusion, we have shown that MyD88-dependent MDSC expansion from donor BM is critical for protection against fatal acute intestinal GVHD. DisclosuresNo relevant conflicts of interest to declare.

Therapeutic Effects of MLN4924 on Sclerodermatous Chronic Graft Versus Host Disease

▪Allogeneic hematopoietic stem cell transplantation (HSCT) remains the only curative option for the treatment of a variety of hematological diseases. However, high incidences of graft versus host disease (GVHD) and tumor relapse remain as major hurdles to this approach. While recent advances in transplant preparation and prophylactic regimens have lowered the incidence of acute GVHD, chronic GVHD (cGVHD) still remains a major cause of long-term morbidity and mortality. Recent studies have suggested that post-translation modifications (PTMs) may serve as therapeutic targets during GVHD. PTMs of proteins are important pathways that confer biochemical alterations, such as acetylation, phosphorylation, and ubiquitination. Targeting PTMs by using small molecular inhibitors has demonstrated potent anti-tumor effects, as well as immune modulatory properties. We have previously shown that bortezomib, an inhibitor of PTMs pathways, can successfully ameliorate sclerodermatous cGVHD responses in both preclinical and clinical studies. Here we reported a new small molecular targeting agent, MLN4924, a selective inhibitor of NEDD8-activating enzyme, on the treatment of cGVHD.MLN4924 is a selective inhibitor of NEDD8-activating enzyme and has been shown to be well tolerated at higher doses and exhibit potent anti-tumor activity in murine models and in early clinical trials. Furthermore, MLN4924 can suppress dendritic cell functions, as well as their production of proinflammatory cytokines during allogeneic responses. Due to these findings, we sought to test the treatment effects of MLN4924 on cGVHD responses. To investigate this, we utilized a minor MHC mismatch murine model by transferring donor B10.D2 (H-2d) cells into BALB/c (H-2d) recipients. MLN4924 or vehicle control (10% 2-hydroxy-propyl-beta cyclodextrin) was injected intraperitoneally on day 20, the point at which skin sclerodermatous cGVHD is initiated. Compared with the vehicle groups, animals that received either bortezomib (0.1 mg/kg) or MLN4924 (20 mg/kg) developed significantly less cGVHD, as determined by clinical scores (2.34±0.34 versus 1.03±0.37 versus 0.8±0.14, respectively; P <0.01). Surprisingly, a preventive approach started from either day 0 or day 10 post-HSCT resulted in higher cGVHD scores compared to early therapeutic interventions, started from Day 20, (3.3 and 2.37± 0.18 versus 0.8±0.14, respectively). The temporally dependent efficacy of MLN4924 suggests that PTMs may play a suppressive role in the early development of cGVHD pathogenesis while worsening cGVHD response as a prevention treatment. Flow cytometric analysis revealed a significant reduction in the total number of macrophages and dendritic cells with MLN4924 treatment starting on day 20, while no significant changes were noticed in T cell subpopulations. Similarly, MLN4924 can successfully inhibit mixed lymphocyte reactions (MLR) in vitro. Importantly, a reduction in the total number of plasma cells, a major source of autoantibody production, was also observed. Together these data suggest that inhibition of neddylation with small molecules, such as MLN4924, can be used as a new approach for the treatment of patients with sclerodermatous cGVHD, while potentially maintaining graft versus tumor effects through preserving T cell populations. DisclosuresNo relevant conflicts of interest to declare.

Evaluation of Blood Group Conversion Following ABO-Incompatible Hematopoietic Stem Cell Transplantation (HCT)

Title: Evaluation of blood group conversion following ABO-incompatible Hematopoietic Stem Cell Transplantation (HCT).Background:While there have been a number of reports concerning the influence of AB0 incompatibility on hematopoietic transplantation (HCT) outcomes, data evaluating post-HCT blood group conversions following ABO incompatible transplants remains extremely limited.Methods:We performed a single centre retrospective analysis of 97 patients undergoing major or minor or bidirectional ABO-mismatched bone marrow (BM), peripheral blood (PB), and cord blood (CB) hematopoietic stem cell transplantation (HCT) between January2005 – April 2014, in whom engraftment had occurred and blood group data was available. The aim of the study was to analyse the red cell ABO phenotype (forward grouping) and levels of anti-A/B antibodies (reverse grouping) in these patients following the ABO-mismatched transplants.Results:97 patients transplanted between January 2004 and April 2014 were included in the analysis. There were 40 (41%) matched sibling, 32 (33%) matched unrelated and 25 (26%) CB transplants. Indications for HSCT included both haematological malignancies (n=87) and benign conditions (n=10). Of these donor recipient pairs, there were 44 (45%) minor, 39 (40%) major and 14 (15%) bidirectional ABO mismatches. All patients had achieved full donor type chimerism at the time of analysis.Analysis of post –HCT blood group conversion was performed in each of the three groups. Amongst the major mismatch patients (n=39), all had conversion of RBC phenotype to donor ABO phenotype and loss of their host derived anti-donor ABO antibodies. Interestingly, in the minor mismatch patients who were tested (n=44), all the patients had conversion of their RBC phenotype, but 43 out of 44 patients failed to produce recipient directed anti-ABO antibodies. Similarly in the bidirectional mismatch group, all 14 patients converted to donor ABO phenotype, but none produced recipient directed anti-ABO antibodies.Conclusion:In one of the largest study looking at this issue, our study demonstrated the lack of a development of recipient directed anti A/B antibodies despite complete donor chimerism in the majority (1 out of 58) of patients undergoing minor or bidirectional ABO-mismatched HSCT. We postulate that this phenomenon may be due to the donor B cell tolerance against host ABO antigen due to the presence of host A and B antigens within other body tissues. DisclosuresNo relevant conflicts of interest to declare.

Physical exercise reduces transplant arteriosclerosis in a mouse aorta transplantation model

Transplant arteriosclerosis limits long-term outcome after heart transplantation. The underlying mechanism of transplant arteriosclerosis remains alloreactivity, but it is also influenced by nonimmunologic cofactors. Physical exercise has well-established effects on the prevention of native arteriosclerosis. We hypothesized that physical exercise would reduce the development of transplant arteriosclerosis in an allogeneic transplantation setting. Segments of the thoracic aorta from C57.Bl6 (H2b) or C3H.HeJ (H2k) mice were transplanted into the abdominal aortas of CBA.Ca mice (H2k), representing a major or minor alloantigen mismatch, respectively. Three days after surgery, recipient mice were assigned to either the control or physical exercise (consisting of 2×45 minutes of treadmill training per day) groups. Transplant arteriosclerosis was assessed and quantified by histology on day 28 after vessel transplantation. Endothelial cell integrity and function in histology sections and peripheral blood was assessed. All animals developed transplant arteriosclerosis with more severe luminal occlusion in the major alloantigen mismatch setting. Animals undergoing physical exercise developed significantly less severe transplant arteriosclerosis in both the major (P<.0001) and minor (P<.0001) antigen mismatches than their respective control groups without physical exercise. CD31(+) endothelial cells were present in significantly higher numbers in the grafts and circulating in peripheral blood in the exercise groups compared with their respective control. Above that, we found enhanced endothelial nitric oxide synthase-positive cells in both exercise groups compared with the untreated groups (P=.01 and P=.02, respectively). Physical exercise has a protective effect against the development of transplant arteriosclerosis. This could be due to enhanced endothelial cell regeneration and function in the graft.

Brain MRI, SPECT and PET in Early Alzheimer’s Disease: A Minor Mismatch between Volumetric and Functional Findings

Background: Limited data are available in the literature comparing brain SPECT, PET and volumetric MRI in the same cohort of patients with Alzheimer’s disease. Objectives: The main goal was to show a correlation between two different functional imaging substrates, as determined by FDG-PET and SPECT, with structural MRI findings as defined by voxel-based volumetric measurements (VBM), in the same cohort of patients with AD. Methods: 20 patients (9 male) with confirmed clinical diagnosis of Alzheimer’s disease (AD) and mild dementia according to NINCDS/ADRDA and DSM-III-R criteria were selected to be included in the present study. Twenty normal elderly volunteers constituted the Control Group. The two groups were paired for age, sex and educational level (p>0.05). The same clinical protocol was applied in patients and controls. Brain SPECT, PET and MR were performed with an interval among them shorter than 4 weeks. Results: Reduced brain glucose metabolism (MRglc) by PET occurred predominantly in the posterior cingulum and precuneus, followed by bilateral posterior parietal, lower temporal and frontal cortices (p<0.001). Similar findings were shown on SPECT, albeit with lesser statistical significance compared to FDG-PET (Z score of 3.89 vs. 5.32). SPECT depicted subtle blood flow reduction in mesial temporal lobes on SPM, a finding not reproduced by PET. MRI showed significant atrophy in hippocampus and superior temporal cortex, bilaterally, but not in the posterior cingulate gyrus, parietal or frontal lobes. Conclusion: A not perfect correlation was seen among the findings seen on SPECT, PET and MRI in the same cohort of patients with AD, suggesting that structural and functional methods reveal distinct substrates of the disease. Keywords: Alzheimer's disease, dementia, FDG, PET, SPECT, volumetric MRI.

Therapeutic Benefit of Bortezomib on Acute Graft-versus-Host Disease Is Tissue Specific and Is Associated with Interleukin-6 Levels

Bortezomib, a proteasome inhibitor capable of direct antitumor effects, has been shown to prevent acute graft-versus-host disease (GVHD) when administered in a short course immediately after bone marrow transplantation (BMT) in mice. However, when bortezomib is given continuously, CD4+ T cell–mediated gastrointestinal tract damage increases GVHD mortality. To investigate the protective effects of bortezomib on other organs, we used a CD8-dependent acute GVHD (aGVHD) model of C3H.SW donor T cells engrafted into irradiated C57BL/6 recipients (minor MHC mismatch), which lack significant gut GVHD. Our data in this model show that bortezomib can be given continuously to prevent and treat aGVHD mediated by CD8+ T cells, but this effect is organ specific, such that only skin, and not liver, protection was observed. Despite the lack of hepatic protection, bortezomib still significantly improved survival, primarily because of its skin protection. Reduced skin GVHD by bortezomib was correlated with reduced serum and skin IL-6 levels. Administration of a blocking IL-6 antibody in this model also resulted in similar cutaneous GVHD protection. These results indicate that bortezomib or blockade of IL-6 may prevent CD8+ T cell–mediated cutaneous acute GVHD.

Recipient NK cell inactivation and intestinal barrier loss are required for MHC-matched graft-versus-host disease.

Previous studies have shown a correlation between pretransplant conditioning intensity, intestinal barrier loss, and graft-versus-host disease (GVHD) severity. However, because irradiation and other forms of pretransplant conditioning have pleiotropic effects, the precise role of intestinal barrier loss in GVHD pathogenesis remains unclear. We developed GVHD models that allowed us to isolate the specific contributions of distinct pretransplant variables. Intestinal damage was required for the induction of minor mismatch [major histocompatibility complex (MHC)-matched] GVHD, but was not necessary for major mismatch GVHD, demonstrating fundamental pathogenic distinctions between these forms of disease. Moreover, recipient natural killer (NK) cells prevented minor mismatch GVHD by limiting expansion and target organ infiltration of alloreactive T cells via a perforin-dependent mechanism, revealing an immunoregulatory function of MHC-matched recipient NK cells in GVHD. Minor mismatch GVHD required MyD88-mediated Toll-like receptor 4 (TLR4) signaling on donor cells, and intestinal damage could be bypassed by parenteral lipopolysaccharide (LPS) administration, indicating a critical role for the influx of bacterial components triggered by intestinal barrier loss. In all, the data demonstrate that pretransplant conditioning plays a dual role in promoting minor mismatch GVHD by both depleting recipient NK cells and inducing intestinal barrier loss.

CD4+ T Cells and Not Th17 Cells Are Required for Lung Transplant Obliterative Bronchiolitis.

Obliterative bronchiolitis (OB) is immune mediated fibrous obliteration of the small airways and limits long term lung transplant survival. Previous studies in a mouse model of minor MHC mismatch orthotopic left lung transplant (C57Bl/10, H-2b®C57Bl/6, H-2b) have suggested a role for IL-17A, a pro-inflammatory and pro-fibrotic cytokine, in OB development. However, the mechanisms by which IL-17A induce OB are not known. Here, we have investigated the cellular sources of IL-17A and determined their contribution to OB development in this model using antibody mediated depletion of T cells and conditional knockout mice. We found increased infiltration of IL-17A+ lymphocytes in left lung allografts at 21 days after transplant. The infiltration of IL-17A+ lymphocytes coincided with OB development. IL-17A was produced by CD4+ T cells (Th17) and gδT cells. IL-17A+ T cell subsets were higher in allografts than isografts. Depletion of CD4+ T cells abrogated OB development and led to impaired IL-17A+ lymphocyte responses in allografts. To further test if IL-17A+ CD4 T cells (Th17) were required for OB development, left lungs were transplanted from C57Bl/10 mice to conditional knockout C57Bl/6 STAT3fl/fl mice with a CD4-Cre transgene (STAT3CD4-/-). The CD4+ T cells from STAT3CD4-/- mice are not able to differentiate into Th17 cells because of lack of STAT3. Contrary to expectations, the lung allografts from STAT3CD4-/- mice demonstrated OB lesions. The frequency of IL-17A+ lymphocytes was not decreased in allografts from Th17 deficient mice and gδ T cells remained a major source of IL-17A. Our data suggest that CD4+ T cells are required for OB whereas Th17 cells are not necessary for OB. Our data may support the use of immunosuppressive strategies targeting CD4+ T cells to extend lung allograft survival in clinical settings. Supported by NIH (R01HL109310).

Induction of immune tolerance by CGEN-15001, a novel immunomodulatory Fc fusion protein of the B7 family (THER5P.823)

Abstract We discovered a novel immune checkpoint protein based on characteristics of the B7/CD28 family. CGEN-15001 is a fusion protein composed of the extracellular domain of this protein fused to IgG Fc. CGEN-15001 inhibits T cell activation, demonstrates immunomodulation by inhibiting pro-inflammatory Th1 and Th17 responses while promoting anti-inflammatory IL-10 and Th2 responses, and enhances iTreg differentiation. CGEN-15001 is efficacious in animal models of autoimmunity such as multiple sclerosis (MS), rheumatoid arthritis, type1 diabetes (T1D) and psoriasis. In the EAE model of MS and in the NOD model of T1D, short 2-week treatment with CGEN-15001 resulted in long term inhibition of disease. In the T1D model, mice treated with CGEN-15001 retained normal blood glucose levels18 weeks after cessation of treatment. To further study tolerance induction, the H-Y minor Ag mismatch bone marrow transplantation model was used. CGEN-15001 treatment resulted in graft survival and increase in Tregs, indicating donor specific tolerance induction. The prolonged therapeutic responses in models of autoimmunity and the prevention of graft rejection, imply establishment of immune tolerance by CGEN-15001. Taken together, the mode of action of CGEN-15001 that combines inhibition of autoreactive cells, immunomodulation and restoration of immune tolerance, suggests it should provide long standing and safe therapy for autoimmune diseases.

The role of myeloid cells in graft rejection (TRAN3P.893)

Abstract Acceptance of a mismatched graft is achieved through immunosuppression that requires long-term treatment. A variety of methods have been explored to prevent rejection and achieve transplant tolerance in mouse models. The combination of LPS from gram-negative Bacteria, and tissue damage from the transplant procedure has been suggested to prevent rejection and deserves further investigation. We implemented a mouse skin transplant model to determine the effects of LPS and endogenous danger signals released during engraftment, on myeloid cell functions and study their ability to induce rejection or tolerance in transplantation. We used the spontaneous single minor histocompatibility mismatch model of skin graft rejection and performed skin grafts from male mice onto syngeneic female recipients then treated the mice with LPS or PBS. Control mice completely rejected the graft between 24-34 days, while mice treated with LPS did not show graft rejection until an average of 64 days, with 50% graft acceptance. When we studied the graft immune infiltrate, we found that within 48 hours post transplantation, there was an influx of GR-1+ CD11b+ double positive cells. LPS-treated recipients showed a significant decrease in the recruitment of these GR-1+ CD11b+ double positive cells at the graft site. These results indicate that GR-1+ CD11b+ myeloid cells migrate to the site of transplantation following trauma and suggest they play a key role in graft rejection, warranting further study.

Bloodstream Infection after Stem Cell Transplantation in Children with Idiopathic Aplastic Anemia

Bloodstream infection (BSI) is the most common infectious complication of hematopoietic stem cell transplantation (HSCT) and can cause substantial morbidity and mortality. Identification of risk factors for BSI might be helpful in efforts to reduce transplantation-related death. This study analyzed the incidence of BSI and risk factors for BSI after HSCT in pediatric patients with aplastic anemia (AA). BSI occurred in 39 of the 351 patients with AA (11.1%). Onset of BSI occurred at a median of 8 days after HSCT (range, 0 to 92 days). The 5-year overall survival rate was lower in patients with BSI than in patients without BSI (63.32% ± 7.90% versus 93.35% ± 1.44%; P < .0001). Univariate analysis identified the following variables as associated with BSI: history of immunosuppressive therapy with antithymocyte globulin (ATG), transplantation from an unrelated donor, frequent blood transfusion before transplantation, major or major plus minor ABO type mismatch, graft-versus-host disease prophylaxis with tacrolimus and without cyclosporine, and long interval from diagnosis to transplantation. Among these factors, long interval from diagnosis to transplantation was the sole statistically significant risk factor for BSI on multivariate analysis. In patients who underwent HSCT from a related donor, age ≥14 years at transplantation was risk factor for BSI. In contrast, history of immunosuppressive therapy with ATG, frequent blood transfusion before HSCT, graft failure, and major or major plus minor ABO type mismatch were risk factors for BSI in patients who underwent HSCT from an unrelated donor. Because the overall 5-year survival rate without BSI was >90%, even in patients who were received a transplant from an unrelated donor, control of BSI is very important for successful HSCT in pediatric patients with AA.