ALBERT

Description: Adipose-derived circulating miRNAs regulate gene expression in other tissues Nature 542, 7642 (2017). doi:10.1038/nature21365 Authors: Thomas Thomou, Marcelo A. Mori, Jonathan M. Dreyfuss, Masahiro Konishi, Masaji Sakaguchi, Christian Wolfrum, Tata Nageswara Rao, Jonathon N. Winnay, Ruben Garcia-Martin, Steven K. Grinspoon, Phillip Gorden & C. Ronald Kahn Adipose tissue is a major site of energy storage and has a role in the regulation of metabolism through the release of adipokines. Here we show that mice with an adipose-tissue-specific knockout of the microRNA (miRNA)-processing enzyme Dicer (ADicerKO), as well as humans with lipodystrophy,

Description: Corrigendum: Adipose-derived circulating miRNAs regulate gene expression in other tissues Nature 545, 7653 (2017). doi:10.1038/nature22319 Author: Thomas Thomou, Marcelo A. Mori, Jonathan M. Dreyfuss, Masahiro Konishi, Masaji Sakaguchi, Christian Wolfrum, Tata Nageswara Rao, Jonathon N. Winnay, Ruben Garcia-Martin, Steven K. Grinspoon, Phillip Gorden & C. Ronald Kahn Nature542, 450–455 (2017); doi:10.1038/nature21365In this Article, in Fig. 4f–h, the x-axis labels ‘WT + exoKO’ and ‘ADicerKO + exoKO’ were inadvertently reversed, and in Fig. 5c, f, the y-axis labels should have been labelled

Description: Introduction: Definitive hematopoietic stem cells (HSCs) sustain blood production from fetal development throughout life. In mice, most of steady state, young adult HSCs are in the G0 phase of cell cycle (quiescence), and are estimated to divide roughly once a month. Daily hematopoietic production is thus mainly sustained by highly proliferative downstream hematopoietic progenitor cells (HPCs). Aged haematopoiesis was demonstrated to be distinct from young haematopoiesis in various aspects such as i) a shift from lymphopoiesis to myelopoiesis, ii) functional decline of HSCs (self-renewal, homing), and iii) HSCs pool expansion. While several studies attempted to address whether changes in HSCs turnover during aging can account for the distinct aging associated phenotype and function, it remained to be determined whether aged HSCs overall cycle more or less frequently than young HSCs. Methods: To construct data-based, quantitative models, we measured turnover rates and compartment sizes of populations of HSCs, HSPCs and granulopoiesis/granulocytes, i.e. a post-mitotic mature hematopoietic linage with a short half-life. We examined four age groups: 3 week, 2 month, 1 year and 2 year old mice. Mice in each group were i.p. injected every 4 hours with 1 mcg EdU up to a maximum time of 48 hours. HSC, HSPC and granulopoiesis/granoulocyte compartment sizes and snapshot cell-cycle analysis was performed by FACS at multiple sampling points in BM and peripheral blood (PB), respectively. Based on this data, we built a mathematical model of HSC turn-over and HSPC differentiation during ageing. Moreover, we evaluated HSC cycling by CFSE dilution in steady-state transplantation experiments (as described before; Takizawa et al., J Exp Med 2011). Results: In line with previous reports, the HSCs compartment size gradually increased with age from 3wk old mice to 2 year old mice. In sharp contrast, cycling activity of HSCs as determined by EdU incorporation decreased gradually and significantly with increasing age. This was driven by decreased activation from the quiescent state, while the time that actively cycling HSCs require to progress through cell-division remains constant with age. Multipotent Progenitor (MPP) cycling showed a non-significant trend towards slower turn-over. These results were confirmed by complementary CFSE-dilution experiments. Mathematical modeling of HSC proliferation and differentiation revealed a higher probability of self-renewing divisions in 3 week old mice as compared to 2 month, 1 and 2 year old mice, with the latter both having nearly equal chances of self-renewing versus differentiating divisions. Conclusions: Our data clarifies the long-standing question, how the HSC pool increases with age. Instead of an increase in active cycling, an increase in HSC quiescence is responsible for the increased size of the HSCs pool in aging. Disclosures No relevant conflicts of interest to declare.

Description: JAK2-V617F is the most frequently recurring somatic mutation in patients with myeloproliferative neoplasm (MPN), but it can also be found in healthy individuals with clonal hematopoiesis of indeterminate potential (CHIP) with a frequency much higher than the incidence of MPN. This suggests that the acquisition of the JAK2-V617F is not the rate-limiting step and other factors might be required for the expansion of the JAK2 mutated clone and initiation of MPN disease. Chronic inflammation is a hallmark of advanced MPN and is associated with progression to myelofibrosis and AML. Interleukin-1β (IL-1β) is one of the master regulators of the inflammatory state and its aberrant activity has been implicated in various pathological diseases including MPN. Here we focused on the early stages of MPN disease initiation and examined the role of IL-1β in this context. We hypothesized that IL-1β mediated inflammation may promote early expansion of the JAK2 mutant clone to reach a critical clone size capable of initiating MPN. We used a genetic approach and crossed IL-1β knockout (IL1β-/-) mice with our tamoxifen inducible SclCreER;JAK2-V617F (VF) mice, generating a triple mutant SclCreER;JAK2-V617F;IL-1β-/-(VF;IL1β-/-) line. We then transplanted two million bone marrow (BM) cells from VF and VF;IL1β-/- mice into lethally irradiated wildtype (WT) orIL1β-/- recipients. Complete blood counts monitored every 4 weeks for up to 32 weeks post transplantation showed reduced platelet, neutrophil, leukocyte and monocyte counts in mice transplanted with VF;IL1β-/-as compared to VF. Furthermore, terminal analysis at week 16 and 32 revealed reduced splenomegaly and bone marrow fibrosis in the mice receiving VF cells lacking IL1β. This experiment shows that IL1β plays an important role in MPN pathogenesis in this mouse model. To test the hypothesis that IL1β favors clonal expansion during MPN disease initiation, we performed competitive dilution assays by mixing BM cells from VF or VF;IL1β-/-mice that also co-express the GFP protein as a reporter (VF;GFP or VF;IL1β-/-;GFP) with BM cells from IL1β-/- mice in 1:100 ratio and transplanted into lethally irradiated WT recipients (Figure 1A). Successful engraftment was defined by presence of 〉1% GFP+ cells within Gr-1+ granulocytes in peripheral blood (PB) at week 18 after transplantation. In mice transplanted with VF;GFP, we found engraftment in 25 of 29 (86%) recipients whereas in mice transplanted with VF;IL1β-/-;GFP, only 18 of 29 (62%) recipients showed engraftment. Moreover, 10 of 25 (40%) mice engrafted with VF;GFP developed MPN at 24 weeks after transplantation as compared to only 2 of 18 (11%) mice engrafted with VF;IL1β-/-;GFP cells. GFP chimerism measured every 6 weeks in peripheral blood (PB) from erythroid (Ter119+), megakaryocytic (CD61+) and granulocytic lineages (Gr-1+) was significantly reduced in mice transplanted with VF;IL1β-/-;GFP compared to mice transplanted with VF;GFP cells (Figure 1A), suggesting the capacity to produce IL-1β protein by the VF cells was promoting the expansion of the clone and MPN manifestation.To define the relative contributions of hematopoietic and non-hematopoietic cell derived IL-1β in promoting MPN initiation, we performed competitive dilution assays in IL1β-/-recipients (Figure 1B). We detected engraftment in 27 of 30 (90%) IL1β-/-recipients transplanted with VF;GFP and 27 of 33 (82%) mice transplanted with VF;IL1β-/-;GFP. Furthermore, 9 of 27 (33%) mice engrafted with either VF;GFP or VF;IL1β-/-;GFP developed MPN at 24 weeks after transplantation. However GFP chimerism in Ter119, Gr-1 and CD61 was lower in mice transplanted with VF;IL1β-/-;GFP compared to mice transplanted with VF;GFP (Figure 1B). We further looked at plasma IL-1β protein levels by ELISA (Figure 1C). Interestingly, we found that IL-1β protein levels were also reduced in WT mice transplanted with VF;IL1β-/-;GFP donor cells, indicating that the non-hematopoietic WT cell cannot compensate for the deficiency of IL-1β in the VF clone. Overall, our results demonstrate that IL-1β favors early clonal expansion and show that IL-1β produced by the JAK2 mutant cells is required for optimal MPN disease initiation. Disclosures No relevant conflicts of interest to declare.

Description: Acute graft versus host disease (aGVHD) remains a major complication in patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT), the only curative treatment for many malignant hematologic diseases. After initial priming in secondary lymphoid organs, alloreactive donor T cells efficiently migrate to the intestinal tract, liver and skin. We observed that alloreactive effector T cells infiltrating and attacking the lamina propria of the small and large intestines closely interact with intestinal myeloid cells of host origin. Here we asked whether these intimate interactions regulate alloreactive effector T cell responses and how they impact intestinal aGvHD. To address these questions, we employed non-invasive bioluminescence imaging, fluorescence and confocal microscopy, clinical and histopathologic scoring, flow cytometry and single cell RNA sequencing in murine models of myeloablative, MHC-mismatched allo-HSCT. In the intestinal lamina propria, we observed that allogeneic T cells closely interacted with CD11b+CD11c+CD103- radio-resistant host type hematopoietic myeloid antigen presenting cells. Selective depletion of intestinal CD11chi or CX3CR1+CD11chi host cells 3 or 8 days after allo-HSCT accelerated alloreactive T cell infiltration, increased T cell mediated inflammatory cytokine production and exacerbated tissue damage resulting in hyperacute lethal aGvHD. These results suggested a strong immunoregulatory effect of these intestinal host-type myeloid cells. Single cell RNA-Seq analysis and flow cytometry (e.g. MHC II, CD11c, F4/80, CD26, CD64, CCR2, CX3CR1), lineage reporter- and defined knockout mice determined these cells as non-classical monocyte derived macrophages as the development and differentiation of these cells did not depend on Flt3, Zbtb46, and CCR2 but rather on CSF-1R and CX3CR1. Adoptive transfer, bone marrow chimeras and parabiosis experiments revealed that these non-classical monocyte derived macrophages differentiated from non-circulating non-classical monocytic precursors. Finally, PD-L1 expression on these intestinal host non-classical monocyte derived macrophages but not on stroma or other hematopoietic cells regulated alloreactive T cell responses in the intestinal tract. Based on these findings we postulate that a specialized and persistent subpopulation of host non-classical monocyte derived macrophages can potently suppress alloreactive T cells in the lamina propria of the intestinal tract. Fostering the differentiation and function of these tissue resident cells may represent an attractive therapeutic strategy to prevent intestinal aGvHD. Disclosures No relevant conflicts of interest to declare.