Rapamycin prevents experimental sclerodermatous chronic graft-versus-host disease in mice

Background: The most widely used mice model of chronic graft-versus-host disease (cGvHD) is an MHC-matched bone marrow transplantation model of sclerodermatous cGvHD. A limitation of that model is that mortality is relatively low, making difficult to study the impact of potentially therapeutic compounds. Aims: To develop a more severe model of cGVHD and to assess the impact of Rapamycin administration in that model. Results: Lethally irradiated Balb/C mice were injected with 10x106 bone marrow cells and 70x106 splenocytes from B10.D2 donor mice. Twenty-one days later, all mice developed cGvHD. For the severe model, donor B10.D2 mice were injected with 0.5x106 splenocytes from Balb/C twenty-one days before transplantation. All mice from the severe model (n=8) died a median of 32 days while 3 of 7 mice in the classical model survived beyond day 52. Mean survival was decreased in the severe model compared to the classical model (32 days versus 37 days; p=0.0185). Recipient mice in the severe group experienced higher weight loss, hair loss and skin fi brosis. Numbers of T lymphocytes (231.9 ± 151.4 versus 951 ± 532.8; p=0.0032) and CD4+ T cells (63.25 ± 41.93 versus 135.0 ± 14.39; p=0.0018) per microliter of blood at day 21 were lower in the severe group than in the classical model. Moreover, number of regulatory T cells (Tregs) was decreased in the severe model (1.250 ± 0.8864 versus 8.000 ± 6.753; p=0.0151). We then investigated whether rapamycin administration could prevent GVHD in the severe model. All (n=8) mice treated with PBS (placebo) died a median of 32 days after transplantation, while 6 of 8 mice given 1 mg/kg/day i.p. rapamycin survived beyond day 52 (p=0.0012). Number of Tregs/μl was higher at day 21 in rapamycin-treated mice than in mice given PBS (2.000±1.195 versus 1.250±0.8864; p=0.0796). Moreover, number of naïve CD4+T (10.00±4.192 versus 30.25±5.185; p= 0.0089) and effector memory T cells (EMT) (30.67±3.180 versus 67.33±7.881; p= 0.0125) were higher in rapamycin mice. Finally, proliferation of EMT (assessed by fl ow cytometry using Ki-67) was higher in PBS than in rapamycin mice (45.28%±4.084 versus 31.90%± 2.003; p=0.0474). Conclusion: We have developed a mice model of severe cGVHD. Interestingly, rapamycin prevented death from cGVHD in that model, perhaps through in vivo expansion of Treg

A colitogenic memory CD4+ T cell population mediates gastrointestinal graft-versus-host disease

Damage to the gastrointestinal tract is a major cause of morbidity and mortality in graft-versus-host disease (GVHD) and is attributable to T cell–mediated inflammation. In this work, we identified a unique CD4+ T cell population that constitutively expresses the β2 integrin CD11c and displays a biased central memory phenotype and memory T cell transcriptional profile, innate-like properties, and increased expression of the gut-homing molecules α4β7 and CCR9. Using several complementary murine GVHD models, we determined that adoptive transfer and early accumulation of β2 integrin–expressing CD4+ T cells in the gastrointestinal tract initiated Th1-mediated proinflammatory cytokine production, augmented pathological damage in the colon, and increased mortality. The pathogenic effect of this CD4+ T cell population critically depended on coexpression of the IL-23 receptor, which was required for maximal inflammatory effects. Non–Foxp3-expressing CD4+ T cells produced IL-10, which regulated colonic inflammation and attenuated lethality in the absence of functional CD4+Foxp3+ T cells. Thus, the coordinate expression of CD11c and the IL-23 receptor defines an IL-10–regulated, colitogenic memory CD4+ T cell subset that is poised to initiate inflammation when there is loss of tolerance and breakdown of mucosal barriers

Comparison of Mesenchymal Stromal Cells From Different Origins for the Treatment of Graft-vs.-Host-Disease in a Humanized Mouse Model

Mesenchymal stromal cells (MSCs) have potent immunomodulatory properties that make them an attractive tool against graft- vs.-host disease (GVHD). However, despite promising results in phase I/II studies, bone marrow (BM-) derived MSCs failed to demonstrate their superiority over placebo in the sole phase III trial reported thus far. MSCs from different tissue origins display different characteristics, but their therapeutic benefits have never been directly compared in GVHD. Here, we compared the impact of BM-, umbilical cord (UC-), and adipose-tissue (AT-) derived MSCs on T-cell function in vitro and assessed their efficacy for the treatment of GVHD induced by injection of human peripheral blood mononuclear cells in NOD-scid IL-2Rγnull HLA-A2/HHD mice. In vitro, resting BM- and AT-MSCs were more potent than UC-MSCs to inhibit lymphocyte proliferation, whereas UC- and AT-MSCs induced a higher regulatory T-cell (CD4+CD25+FoxP3+)/T helper 17 ratio. Interestingly, AT-MSCs and UC-MSCs activated the coagulation pathway at a higher level than BM-MSCs. In vivo, AT-MSC infusions were complicated by sudden death in 4 of 16 animals, precluding an analysis of their efficacy. Intravenous MSC infusions (UC- or BM- combined) failed to significantly increase overall survival (OS) in an analysis combining data from 80 mice (hazard ratio [HR] = 0.59, 95% confidence interval [CI] 0.32–1.08, P = 0.087). In a sensitivity analysis we also compared OS in control vs. each MSC group separately. The results for the BM-MSC vs. control comparison was HR = 0.63 (95% CI 0.30–1.34, P = 0.24) while the figures for the UC-MSC vs. control comparison was HR = 0.56 (95% CI 0.28–1.10, P = 0.09). Altogether, these results suggest that MSCs from various origins have different effects on immune cells in vitro and in vivo. However, none significantly prevented death from GVHD. Finally, our data suggest that the safety profile of AT-MSC and UC-MSC need to be closely monitored given their pro-coagulant activities in vitro

Using Classical Population Genetics Tools with Heterochroneous Data: Time Matters!

BACKGROUND:New polymorphism datasets from heterochroneous data have arisen thanks to recent advances in experimental and microbial molecular evolution, and the sequencing of ancient DNA (aDNA). However, classical tools for population genetics analyses do not take into account heterochrony between subsets, despite potential bias on neutrality and population structure tests. Here, we characterize the extent of such possible biases using serial coalescent simulations. METHODOLOGY/PRINCIPAL FINDINGS:We first use a coalescent framework to generate datasets assuming no or different levels of heterochrony and contrast most classical population genetic statistics. We show that even weak levels of heterochrony ( approximately 10% of the average depth of a standard population tree) affect the distribution of polymorphism substantially, leading to overestimate the level of polymorphism theta, to star like trees, with an excess of rare mutations and a deficit of linkage disequilibrium, which are the hallmark of e.g. population expansion (possibly after a drastic bottleneck). Substantial departures of the tests are detected in the opposite direction for more heterochroneous and equilibrated datasets, with balanced trees mimicking in particular population contraction, balancing selection, and population differentiation. We therefore introduce simple corrections to classical estimators of polymorphism and of the genetic distance between populations, in order to remove heterochrony-driven bias. Finally, we show that these effects do occur on real aDNA datasets, taking advantage of the currently available sequence data for Cave Bears (Ursus spelaeus), for which large mtDNA haplotypes have been reported over a substantial time period (22-130 thousand years ago (KYA)). CONCLUSIONS/SIGNIFICANCE:Considering serial sampling changed the conclusion of several tests, indicating that neglecting heterochrony could provide significant support for false past history of populations and inappropriate conservation decisions. We therefore argue for systematically considering heterochroneous models when analyzing heterochroneous samples covering a large time scale

Compréhension et traitement de la maladie du greffon contre l'Hôte murine de type chronique

Allogeneic hematopoietic cell transplantation (allo-HCT) remains the best treatment option for several hematological malignancies and some genetic disorders. Anti-tumoral efficacy of this approach is based not only on high dose chemo-radiotherapy given in the conditioning regimen, but also on immune-mediated graft-versus-tumor (GVT) effects, primarily mediated by donor T cells contained in the graft. Unfortunately, these donor immune cells can also target recipient tissues, causing a life-threatening complication of allo-HCT: the Graft-versus-Host Disease (GVHD). GVHD comprises two syndromes: acute GVHD (aGVHD), a deregulated inflammatory response causing skin, gastro-intestinal tract and/or liver damages, and chronic GVHD (cGVHD) generally occurring beyond day 100 after transplantation, and affecting virtually any tissue, and often inducing tissue fibrosis. In recent years, significant progress has been achieved for aGVHD prevention. However, little improvement has been made for cGVHD treatment. Sclerodermatous cGVHD (Scl-cGVHD) occurs in up to 15% of patients who develop cGVHD and is one of the most severe forms of cGVHD. As there is a lack of efficient treatment for cGVHD, the first aim of this work was to find efficient drugs for scl-cGVHD. However, reconstitution of a fully functional hematopoietic system is crucial for transplantation outcomes. Impacts of TKIs on hematopoietic stem cell engraftment and differentiation early after allo-HCT are unknown. We first demonstrated that imatinib and nilotinib had a similar impact on hematopoiesis in vitro and did not affect engraftment in immunodeficient mice. Since the PDGF receptor and TGF-beta play a significant role in the fibrosing process occurring during scl-GVHD and their signaling pathways are inhibited by imatinib, we next assessed this TKI in a murine model of scl-GVHD. Unfortunately, imatinib failed to ameliorate scl-cGVHD in a severe murine model of scl-cGVHD despite it significantly inhibited PDGF-r in vivo. In a third part, rapamycin was investigated both in vivo and in vitro for scl-cGVHD. Rapamycin inhibits conventional T cell activation and proliferation without inhibiting Treg cells by acting via mTor. This immunosuppressant also inhibit fibrosis by acting via the PI3K/Akt signaling pathway, suggesting that rapamycin is a good candidate for scl-cGVHD. Rapamycin was able to increase survival of recipient mice by decreasing skin fibrosis and decreasing homing of effector T cells in GVHD target organs

Chapter 35: Immune Reconstitution and Tolerance

peer reviewedHematopoietic cell transplantation (HCT) remains the only curative treatment option for many patients with hematological malignancies and genetic disorders. Recipients can receive either an autologous or allogeneic graft, which is often dependent upon the primary disease for which transplantation is contemplated. One of the major goals of transplantation is the development of a fully functional immune system. Patients undergoing allogeneic hematopoietic cell transplantation (alloHCT), in particular, have the highest rate of transplantrelated mortality (TRM) due to complications that can result from immunodeficiencies or immune dysregulation that occurs as a consequence of the procedure. These include, but are not limited to, infections, graft‐versus‐host disease (GvHD), autoimmune disorders, and secondary malignancies. Recovery of a fully functional immune system is thus critical for HCT success and improving immune reconstitution is of great clinical interest. This chapter will focus on immune reconstitution following allo‐HCT since immune recovery is more complicated in this setting due to the underlying HLA disparity between the donor and the recipient