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Mesenchymal (stromal) stem cells (MSCs) constitute populations of mesodermal multipotent cells involved in tissue regeneration and homeostasis in many different organs. Here we performed comprehensive characterization of the transcriptional and epigenomic changes associated with osteoblast and adipocyte differentiation of human MSCs. We demonstrate that adipogenesis is driven by considerable remodeling of the chromatin landscape and de novo activation of enhancers, whereas osteogenesis involves activation of preestablished enhancers. Using machine learning algorithms for in silico modeling of transcriptional regulation, we identify a large and diverse transcriptional network of pro-osteogenic and antiadipogenic transcription factors. Intriguingly, binding motifs for these factors overlap with SNPs related to bone and fat formation in humans, and knockdown of single members of this network is sufficient to modulate differentiation in both directions, thus indicating that lineage determination is a delicate balance between the activities of many different transcription factors. Transcriptional and epigenomic profiling of osteoblast and adipocyte differentiation shows that adipogenesis is driven by de novo activation of enhancers, whereas osteogenesis involves preestablished enhancers and depends on the activation of pro-osteogenic and antiadipogenic transcription factors.

Endothelial cells (ECs) contribute to haematopoietic stem cell (HSC) maintenance in bone marrow, but the differential contributions of EC subtypes remain unknown, owing to the lack of methods to separate with high purity arterial endothelial cells (AECs) from sinusoidal endothelial cells (SECs). Here we show that the combination of podoplanin (PDPN) and Sca-1 expression distinguishes AECs (CD45 − Ter119 − Sca-1 bright PDPN − ) from SECs (CD45 − Ter119 − Sca-1 dim PDPN + ). PDPN can be substituted for antibodies against the adhesion molecules ICAM1 or E-selectin. Unexpectedly, prospective isolation reveals that AECs secrete nearly all detectable EC-derived stem cell factors (SCF). Genetic deletion of Scf in AECs, but not SECs, significantly reduced functional HSCs. Lineage-tracing analyses suggest that AECs and SECs self-regenerate independently after severe genotoxic insults, indicating the persistence of, and recovery from, radio-resistant pre-specified EC precursors. AEC-derived SCF also promotes HSC recovery after myeloablation. These results thus uncover heterogeneity in the contribution of ECs in stem cell niches. Endothelial cells (EC) are known to contribute to haematopoietic stem cell (HSC) maintenance in the bone marrow (BM). Here the authors demonstrate that arterial ECs can be distinguished from sinusoidal ECs by podoplanin and Sca-1 expression, and that specifically arterial, but not sinusoidal ECs maintain HSCs by secreting SCF.

Several cell types have been proposed to create niches for haematopoietic stem cells (HSCs). However, the expression patterns of HSC maintenance factors have not been systematically studied and no such factor has been conditionally deleted from any candidate niche cell. Thus, the cellular sources of these factors are undetermined. Stem cell factor (SCF; also known as KITL) is a key niche component that maintains HSCs. Here, using Scf gfp knock-in mice, we found that Scf was primarily expressed by perivascular cells throughout the bone marrow. HSC frequency and function were not affected when Scf was conditionally deleted from haematopoietic cells, osteoblasts, nestin -cre - or nestin- creER -expressing cells. However, HSCs were depleted from bone marrow when Scf was deleted from endothelial cells or leptin receptor ( Lepr )-expressing perivascular stromal cells. Most HSCs were lost when Scf was deleted from both endothelial and Lepr -expressing perivascular cells. Thus, HSCs reside in a perivascular niche in which multiple cell types express factors that promote HSC maintenance. The cellular sources of stem cell factor, a major haematopoietic stem cell (HSC) niche cytokine required for HSC maintenance, are identified. The anatomy of a stem-cell niche The haematopoietic niche is a microenvironment that maintains and regulates haematopoietic stem cells (HSCs). Using a systematic conditional genetic approach, Sean Morrison and colleagues identify the cellular sources of stem cell factor (SCF), a major niche cytokine required for maintenance of haematopoietic stem cells. They found SCF to be expressed by endothelial cells and leptin receptor-expressing perivascular stromal cells, but not by previously described niche cells, such as osteoblasts. These findings demonstrate that haematopoietic stem cells reside in a perivascular niche in which multiple cell types express factors that promote their maintenance.

The total number of acquired melanocytic nevi on the skin is strongly correlated with melanoma risk. Here we report a meta-analysis of 11 nevus GWAS from Australia, Netherlands, UK, and USA comprising 52,506 individuals. We confirm known loci including MTAP , PLA2G6 , and IRF4 , and detect novel SNPs in KITLG and a region of 9q32. In a bivariate analysis combining the nevus results with a recent melanoma GWAS meta-analysis (12,874 cases, 23,203 controls), SNPs near GPRC5A, CYP1B1 , PPARGC1B , HDAC4 , FAM208B, DOCK8 , and SYNE2 reached global significance, and other loci, including MIR146A and OBFC1, reached a suggestive level. Overall, we conclude that most nevus genes affect melanoma risk ( KITLG an exception), while many melanoma risk loci do not alter nevus count. For example, variants in TERC and OBFC1 affect both traits, but other telomere length maintenance genes seem to affect melanoma risk only. Our findings implicate multiple pathways in nevogenesis. Melanocytic nevus count is associated with melanoma risk. In this study, a meta-analysis of 11 nevus GWAS studies identifies novel SNPs in KITLG and 9q32, and bivariate analysis with melanoma GWAS meta-analysis reveals that most nevus genes affect melanoma risk, while melanoma risk loci do not alter the nevus count.

The host tissue microenvironment influences malignant cell proliferation and metastasis, but little is known about how tumor-induced changes in the microenvironment affect benign cellular ecosystems. Applying dynamic in vivo imaging to a mouse model, we show that leukemic cell growth disrupts normal hematopoietic progenitor cell (HPC) bone marrow niches and creates abnormal microenvironments that sequester transplanted human CD34? (HPC-enriched) cells. CD34? cells in leukemic mice declined in number over time and failed to mobilize into the peripheral circulation in response to cytokine stimulation. Neutralization of stem cell factor (SCF) secreted by leukemic cells inhibited CD34? cell migration into malignant niches, normalized CD34? cell numbers, and restored CD34? cell mobilization in leukemic mice. These data suggest that the tumor microenvironment causes HPC dysfunction by usurping normal HPC niches and that therapeutic inhibition of HPC interaction with tumor niches may help maintain normal progenitor cell function in the setting of malignancy.

Hematopoietic insufficiency is the hallmark of acute myeloid leukemia (AML) and predisposes patients to life-threatening complications such as bleeding and infections. Addressing the contribution of mesenchymal stromal cells (MSC) to AML-induced hematopoietic failure we show that MSC from AML patients ( n =64) exhibit significant growth deficiency and impaired osteogenic differentiation capacity. This was molecularly reflected by a specific methylation signature affecting pathways involved in cell differentiation, proliferation and skeletal development. In addition, we found distinct alterations of hematopoiesis-regulating factors such as Kit-ligand and Jagged1 accompanied by a significantly diminished ability to support CD34+ hematopoietic stem and progenitor cells in long-term culture-initiating cells (LTC-ICs) assays. This deficient osteogenic differentiation and insufficient stromal support was reversible and correlated with disease status as indicated by Osteocalcin serum levels and LTC-IC frequencies returning to normal values at remission. In line with this, cultivation of healthy MSC in conditioned medium from four AML cell lines resulted in decreased proliferation and osteogenic differentiation. Taken together, AML-derived MSC are molecularly and functionally altered and contribute to hematopoietic insufficiency. Inverse correlation with disease status and adoption of an AML-like phenotype after exposure to leukemic conditions suggests an instructive role of leukemic cells on bone marrow microenvironment.

Accumulated evidence suggests a physiological relationship between the transcription factor NRF3 (NFE2L3) and cancers. Under physiological conditions, NRF3 is repressed by its endoplasmic reticulum (ER) sequestration. In response to unidentified signals, NRF3 enters the nucleus and modulates gene expression. However, molecular mechanisms underlying the nuclear translocation of NRF3 and its target gene in cancer cells remain poorly understood. We herein report that multiple regulation of NRF3 activities controls cell proliferation. Our analyses reveal that under physiological conditions, NRF3 is rapidly degraded by the ER-associated degradation (ERAD) ubiquitin ligase HRD1 and valosin-containing protein (VCP) in the cytoplasm. Furthermore, NRF3 is also degraded by β-TRCP, an adaptor for the Skp1-Cul1-F-box protein (SCF) ubiquitin ligase in the nucleus. The nuclear translocation of NRF3 from the ER requires the aspartic protease DNA-damage inducible 1 homolog 2 (DDI2) but does not require inhibition of its HRD1-VCP-mediated degradation. Finally, NRF3 mediates gene expression of the cell cycle regulator U2AF homology motif kinase 1 (UHMK1) for cell proliferation. Collectively, our study provides us many insights into the molecular regulation and biological function of NRF3 in cancer cells.

Genetic manipulation of NOD/SCID (NS) mice has yielded numerous sub-strains with specific traits useful for the study of human hematopoietic xenografts, each with unique characteristics. Here, we have compared the engraftment and output of umbilical cord blood (UCB) CD34+ cells in four immune-deficient strains: NS, NS with additional IL2RG knockout (NSG), NS with transgenic expression of human myeloid promoting cytokines SCF, GM-CSF, and IL-3 (NSS), and NS with both IL2RG knockout and transgenic cytokine expression (NSGS). Overall engraftment of human hematopoietic cells was highest in the IL2RG knockout strains (NSG and NSGS), while myeloid cell output was notably enhanced in the two strains with transgenic cytokine expression (NSS and NSGS). In further comparisons of NSG and NSGS mice, several additional differences were noted. NSGS mice were found to have a more rapid reconstitution of T cells, improved B cell differentiation, increased levels of NK cells, reduced platelets, and reduced maintenance of primitive CD34+ cells in the bone marrow. NSGS were superior hosts for secondary engraftment and both strains were equally suitable for experiments of graft versus host disease. Increased levels of human cytokines as well as human IgG and IgM were detected in the serum of humanized NSGS mice. Furthermore, immunization of humanized NSGS mice provided evidence of a functional response to repeated antigen exposure, implying a more complete hematopoietic graft was generated in these mice. These results highlight the important role that myeloid cells and myeloid-supportive cytokines play in the formation of a more functional xenograft immune system in humanized mice.

Oxidative stress has a critical role in the pathogenesis of vitiligo. However, the specific molecular mechanism involved in oxidative stress-induced melanocyte death is not well characterized. Given the powerful role of microRNAs (miRNAs) in the regulation of cell survival as well as the fact that the generation of miRNAs can be affected by oxidative stress, we hypothesized that miRNAs may participate in vitiligo pathogenesis by modulating the expression of vital genes in melanocytes. In the present study, we initially found that miR-25 was increased in both serum and lesion samples from vitiligo patients, and its serum level was correlated with the activity of vitiligo. Moreover, restoration of miR-25 promoted the H 2 O 2 -induced melanocyte destruction and led to the dysfunction of melanocytes. Further experiments proved that MITF, a master regulator in melanocyte survival and function, accounted for the miR-25-caused damaging impact on melanocytes. Notably, other than the direct role on melanocytes, we observed that miR-25 inhibited the production and secretion of SCF and bFGF from keratinocytes, thus impairing their paracrine protective effect on the survival of melanocytes under oxidative stress. At last, we verified that oxidative stress could induce the overexpression of miR-25 in both melanocytes and keratinocytes possibly by demethylating the promoter region of miR-25. Taken together, our study demonstrates that oxidative stress-induced overexpression of miR-25 in vitiligo has a crucial role in promoting the degeneration of melanocytes by not only suppressing MITF in melanocytes but also impairing the paracrine protective effect of keratinocytes. Therefore, it is worthy to investigate the possibility of miR-25 as a potential drug target for anti-oxidative therapy in vitiligo.

Group 3 innate lymphoid cells (ILC3s) have emerged as an important player in the pathogenesis of neutrophilic asthma. However, the regulatory mechanism supporting ILC3 responses in the lung remains largely unclear. Here, we demonstrated that stem cell factor (SCF) expression is significantly increased and positively correlated with IL-17A and MPO expression in asthmatic patients. Notably, we identified ILC3 as a major IL-17A-producing responder to SCF in the lung. In mice, SCF synergized with IL-1β/IL-23 to enhance pulmonary ILC3 activation and neutrophilic inflammation. Mechanistically, SCF promoted ILC3 proliferation and cytokine production. Transcriptomic analysis revealed that SCF treatment upregulated the genes related to proliferation and Th17 differentiation, associated with increased AKT and STAT3 signaling. In contrast, deficiency of SCF receptor c-Kit reduced ILC3 proliferation and IL-17A production, resulting in the amelioration of airway hyperreactivity (AHR) and neutrophilic inflammation in mouse neutrophilic asthma model. Furthermore, genetic deletion of SCF in fibroblasts revealed fibroblasts as the primary source of SCF for ILC3 activation in the lung. Moreover, administration of imatinib, a c-Kit inhibitor, alleviated LPS, air pollution or ovalbumin/LPS-induced AHR and neutrophilic inflammation. Our findings elucidated a positive modulatory role of SCF/c-Kit signaling in ILC3 responses during neutrophilic inflammation, offering a potential therapeutic target for neutrophilic asthma.