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AU A subpopulation: Pleaseconfirmthatallheadinglevelsarerepresentedcorrectly of deeply quiescent, so-called dormant hematopoietic :stem cells (dHSCs) resides at the top of the hematopoietic hierarchy and serves as a reserve pool for HSCs. The state of dormancy protects the HSC pool from exhaustion throughout life; however, excessive dormancy may prevent an efficient response to hematological stresses. Despite the significance of dHSCs, the mechanisms maintaining their dormancy remain elusive. Here, we identify CD38 as a novel and broadly applicable surface marker for the enrichment of murine dHSCs. We demonstrate that cyclic adenosine diphosphate ribose (cADPR), the product of CD38 cyclase activity, regulates the expression of the transcription factor c-Fos by increasing the release of Ca2+ from the endoplasmic reticulum (ER). Subsequently, we uncover that c-Fos induces the expression of the cell cycle inhibitor p57Kip2 to drive HSC dormancy. Moreover, we found that CD38 ecto-enzymatic activity at the neighboring CD38-positive cells can promote human HSC quiescence. Together, CD38/cADPR/Ca2+/c-Fos/ p57Kip2 axis maintains HSC dormancy. Pharmacological manipulations of this pathway can provide new strategies to improve the success of stem cell transplantation and blood regeneration after injury or disease., SCOPUS: ar.j, info:eu-repo/semantics/published

Precision medicine can significantly improve outcomes for patients with cancer, but implementation requires comprehensive characterization of tumor cells to identify therapeutically exploitable vulnerabilities. Here, we describe somatic biallelic TET2 mutations in an elderly patient with acute myeloid leukemia (AML) that was chemoresistant to anthracycline and cytarabine but acutely sensitive to 5'-azacitidine (5'-Aza) hypomethylating monotherapy, resulting in long-term morphological remission. Given the role of TET2 as a regulator of genomic methylation, we hypothesized that mutant TET2 allele dosage affects response to 5'-Aza. Using an isogenic cell model system and an orthotopic mouse xenograft, we demonstrate that biallelic TET2 mutations confer sensitivity to 5'-Aza compared with cells with monoallelic mutations. Our data argue in favor of using hypomethylating agents for chemoresistant disease or as first-line therapy in patients with biallelic TET2-mutated AML and demonstrate the importance of considering mutant allele dosage in the implementation of precision medicine for patients with cancer.

Myelodysplastic syndromes (MDS) belong to the most common hematological neoplasms in the elderly population, characterized by ineffective hematopoiesis, peripheral cytopenia and the risk of transformation into acute myeloid leukemia. A dysregulated innate immune response and pro-inflammatory bone marrow microenvironment play a crucial role in the MDS pathogenesis by providing chronic inflammation which makes those pathways the perfect candidate for future therapeutics. Specifically, it has been shown that the alarmin S100A9, an important ligand for dri-ving inflammation and promoting tumor progression, is elevated in MDS patients. Previous expe-riments performed in the Stem Cell Lab 2 provided evidence that mesenchymal stromal cells (MSCs), an important component of the BM niche with immunomodulatory capacity, can be tar-geted by the novel oral small molecular drug Tasquinimod (TASQ, Active Biotech) which has demonstrated S100A9 inhibitory activity. The inhibition of inflammation-related molecules such as IL-1b, IL-18, PD-L1, resulted in a significant improvement of the hematopoietic support by MSCs. However, almost nothing is known about potential effects of TASQ in the context of immunomo-dulation. Therefore, we aimed in this project to understand the mechanisms of S100A9+/- TASQ concerning the immunomodulatory capacity of MDS-MSCs in response to T cell-mediated in-flammation by analyzing adhesion (ICAM1, VCAM1), immune checkpoint (PDL1, PDL2), anti-inflammatory cytokine (COX2, IDO1), chemokines (CCL2, IL8) and extracellular matrix-related (COL4A2, COL1A1) gene expression with quantitative real-time PCR. We observed a general de-crease in the aforementioned genes except for COL4A2 and COL1A1 upon treatment with TASQ, though T cell-mediated inflammation and activity remained unaffected, suggesting that inhibition of S100A9 reduces the inflammation-mediated immunomodulatory potential of MDS-MSCs.:Motivation Aim Methods Result Conclusion

Myelodysplastic neoplasms (MDS) are a group of heterogeneous, clonal disorders characterised by ineffective haematopoiesis and peripheral blood cytopenia. MDS is highly progressive, difficult to treat, and is one of the most common blood cancers, affecting 4-5/100.000 people below the age of 70 and many more thereafter. Single or multiple driver gene mutations and chromosomal abnormalities in the haematopoietic compartment lead to MDS. These somatic gene mutations account for the dysregulation of epigenetic, DNA repair, cohesion complex, and spliceosome pathways. The International prognostic scoring system (IPSS) that was developed in 1997, revised (IPSS-R) in 2016 and updated in 2022 (IPSS-M) classifies MDS into low risk (LR-), intermediate (Int-), and high risk (HR-) groups. The haematopoietic disorder is accompanied by changes in the bone marrow microenvironment (BMME) and especially in mesenchymal cells (MSCs). BMME provides a supportive milieu for haematopoiesis and can be targeted by clinically available drugs such as AZA. The non cellular component of the BMME, the extracellular matrix (ECM), is a framework providing structural and biochemical support via cell-ECM interactions and the maintenance of growth factor gradients. To date, studies of bone marrow interactions in homeostasis and disease have focused largely on soluble and membrane-associated factors, while the involvement of the ECM in MDS and its response to therapy is underexplored. Therefore, this study aimed to characterise the MDS MSC derived ECM of both LR- and HR-MDS in comparison to that from healthy age matched donors in terms of composition, biophysical properties and functional haematopoietic support. This study also aimed to evaluate the impact of in vivo and in vitro AZA treatment on MDS MSC derived ECM. To investigate this, in vitro ECMs were generated by culturing of MSC monolayers on chemically prepared coverslips followed by decellularization using NH4OH and DNase-1 solution. The biop, Myelodysplastische Neoplasien (MDS) sind eine Gruppe heterogener, klonaler Erkrankungen, die durch ineffektive Hämatopoese und Zytopenie des peripheren Blutes gekennzeichnet sind. MDS sind hochgradig progressiv, schwer zu behandeln und gehören zu den häufigsten Blutkrebserkrankungen, von denen 4-5/100.000 Menschen unter 70 Jahren betroffen sind. Die Inzidenz steigt mit zunehmendem Alter deutlich an. MDS wird durch einzelne oder mehrfache Mutationen von Treibergenen und Chromosomenanomalien im hämatopoetischen Kompartiment verursacht. Diese somatischen Genmutationen sind für die Dysregulation von epigenetischen, DNA-Reparatur-, Kohäsionskomplex- und Spleißosomen-Signalwegen verantwortlich. Das Internationale Prognosesystem (IPSS) wurde 1997 entwickelt, 2016 überarbeitet (IPSS-R) und 2022 aktualisiert (IPSS M), um MDS in Gruppen mit niedrigem Risiko (LR-), mittlerem (Int ) und hohem Risiko (HR-) einzuteilen. Die hämatopoetische Erkrankung geht mit Veränderungen in der Mikroumgebung des Knochenmarks (BMME) einher, insbesondere bei mesenchymalen Zellen (MSCs). Das BMME bietet ein unterstützendes Milieu für die Hämatopoese und kann durch klinisch verfügbare Medikamente wie AZA beeinflusst werden. Die nichtzelluläre Komponente der BMME, die extrazelluläre Matrix (ECM), ist ein Gerüst, das durch Zell-ECM-Interaktionen und die Aufrechterhaltung von Wachstumsfaktorgradienten strukturelle und biochemische Unterstützung bietet. Bislang haben sich Studien über die Interaktionen im Knochenmark bei Homöostase und Krankheit hauptsächlich auf lösliche und membranassoziierte Faktoren konzentriert, während die Beteiligung der ECM an MDS und ihre Reaktion auf die Therapie noch nicht ausreichend erforscht ist. Daher zielte diese Studie darauf ab, die aus MDS-MSCs abgeleitete ECM sowohl bei LR- als auch bei HR-MDS im Vergleich zu der von gesunden, altersgleichen Spendern zu charakterisieren, und zwar hinsichtlich der Zusammensetzung, der biophysikalischen Eigenschaften und der funktione

A subpopulation of deeply quiescent, so-called dormant hematopoietic stem cells (dHSCs) resides at the top of the hematopoietic hierarchy and serves as a reserve pool for HSCs possessing the greatest long-term blood repopulation capacity. The state of dormancy protects the HSC pool from exhaustion throughout life, however excessive dormancy may block an efficient response to hematological stresses. The mechanisms of HSC dormancy remain elusive, mainly due to the absence of surface markers that allow dHSC prompt isolation. Here, we identify CD38 as a novel surface marker for murine dHSCs that is broadly applicable. Moreover, we demonstrate that cyclic adenosine diphosphate ribose (cADPR), the product of CD38 cyclase activity, regulates the expression of the transcription factor c-Fos by increasing cytoplasmic Ca2+ concentration. Strikingly, we uncover that c-Fos drives HSCs dormancy through the induction of the cell cycle inhibitor p57Kip2. Moreover, we found that CD38 ecto-enzymatic activity at the neighboring CD38-positive cells can promote human HSC quiescence. Together, CD38/cADPR/Ca2+/cFos/p57Kip2 axis maintains HSC dormancy. Pharmacological manipulations of this pathway can provide new strategies to expand dHSCs for transplantation or to activate them during hematological stresses., info:eu-repo/semantics/published

Myelodysplastic syndromes (MDS) comprise a heterogeneous group of hematologic malignancies characterized by clonal hematopoiesis, one or more cytopenias such as anemia, neutropenia, or thrombocytopenia, abnormal cellular maturation, and a high risk of progression to acute myeloid leukemia. The bone marrow microenvironment (BMME) in general and mesenchymal stromal cells (MSCs) in particular contribute to both the initiation and progression of MDS. However, little is known about the role of MSC-derived extracellular matrix (ECM) in this context. Therefore, we performed a comparative analysis of in vitro deposited MSC-derived ECM of different MDS subtypes and healthy controls. Atomic force microscopy analyses demonstrated that MDS ECM was significantly thicker and more compliant than those from healthy MSCs. Scanning electron microscopy showed a dense meshwork of fibrillar bundles connected by numerous smaller structures that span the distance between fibers in MDS ECM. Glycosaminoglycan (GAG) structures were detectable at high abundance in MDS ECM as white, sponge-like arrays on top of the fibrillar network. Quantification by Blyscan assay confirmed these observations, with higher concentrations of sulfated GAGs in MDS ECM. Fluorescent lectin staining with wheat germ agglutinin and peanut agglutinin demonstrated increased deposition of N-acetyl-glucosamine GAGs (hyaluronan (HA) and heparan sulfate) in low risk (LR) MDS ECM. Differential expression of N-acetyl-galactosamine GAGs (chondroitin sulfate, dermatan sulfate) was observed between LR- and high risk (HR)-MDS. Moreover, increased amounts of HA in the matrix of MSCs from LR-MDS patients were found to correlate with enhanced HA synthase 1 mRNA expression in these cells. Stimulation of mononuclear cells from healthy donors with low molecular weight HA resulted in an increased expression of various pro-inflammatory cytokines suggesting a contribution of the ECM to the inflammatory BMME typical of LR-MDS. CD34+ hemato

Myelodysplastic syndromes (MDS) comprise a heterogeneous group of hematologic malignancies characterized by clonal hematopoiesis, one or more cytopenias such as anemia, neutropenia, or thrombocytopenia, abnormal cellular maturation, and a high risk of progression to acute myeloid leukemia. The bone marrow microenvironment (BMME) in general and mesenchymal stromal cells (MSCs) in particular contribute to both the initiation and progression of MDS. However, little is known about the role of MSC-derived extracellularmatrix (ECM) in this context. Therefore, we performed a comparative analysis of in vitro deposited MSC-derived ECM of different MDS subtypes and healthy controls. Atomic force microscopy analyses demonstrated that MDS ECM was significantly thicker and more compliant than those from healthy MSCs. Scanning electron microscopy showed a dense meshwork of fibrillar bundles connected by numerous smaller structures that span the distance between fibers in MDS ECM. Glycosaminoglycan (GAG) structures were detectable at high abundance in MDS ECM as white, sponge-like arrays on top of the fibrillar network. Quantification by Blyscan assay confirmed these observations, with higher concentrations of sulfated GAGs in MDS ECM. Fluorescent lectin staining with wheat germ agglutinin and peanut agglutinin demonstrated increased deposition of N-acetyl-glucosamine GAGs (hyaluronan (HA) and heparan sulfate) in low risk (LR) MDS ECM. Differential expression of Nacetyl- galactosamine GAGs (chondroitin sulfate, dermatan sulfate) was observed between LR- and high risk (HR)-MDS. Moreover, increased amounts of HA in the matrix of MSCs from LR-MDS patients were found to correlate with enhanced HA synthase 1 mRNA expression in these cells. Stimulation of mononuclear cells from healthy donors with low molecular weight HA resulted in an increased expression of various pro-inflammatory cytokines suggesting a contribution of the ECM to the inflammatory BMME typical of LR-MDS. CD34+ hematop

Functional impairment of the bone marrow (BM) niche has been suggested as a major reason for prolonged cytopenia and secondary graft failure after allogeneic hematopoietic cell transplantation (alloHCT). Because mesenchymal stromal cells (MSCs) serve as multipotent progenitors for several niche components in the BM, they might play a key role in this process. We used collagenase digested trephine biopsies to directly quantify MSCs in 73 patients before (n = 18) and/or after alloHCT (n = 65). For the first time, we demonstrate that acute graft-versus-host disease (aGvHD, n = 39) is associated with a significant decrease in MSC numbers. MSC reduction can be observed even before the clinical onset of aGvHD (n = 10). Assessing MSCs instantly after biopsy collection revealed phenotypic and functional differences depending on the occurrence of aGvHD. These differences vanished during ex vivo expansion. The MSC endotypes observed revealed an enhanced population of donor-derived classical dendritic cells type 1 and alloreactive T cells as the causing agent for compartmental inflammation and MSC damage before clinical onset of aGvHD was ascertained. In conclusion, MSCs endotypes may constitute a predisposing conductor of alloreactivity after alloHCT preceding the clinical diagnosis of aGvHD.