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Energy production by means of ATP synthesis in cancer cells has been investigated frequently as a potential therapeutic target in this century. Both (an)aerobic glycolysis and oxidative phosphorylation (OXPHOS) have been studied. Here, we review recent literature on energy production in glioblastoma stem cells (GSCs) and leukemic stem cells (LSCs) versus their normal counterparts, neural stem cells (NSCs) and hematopoietic stem cells (HSCs), respectively. These two cancer stem cell types were compared because their niches in glioblastoma tumors and in bone marrow are similar. In this study, it became apparent that (1) ATP is produced in NSCs and HSCs by anaerobic glycolysis, whereas fatty acid oxidation (FAO) is essential for their stem cell fate and (2) ATP is produced in GSCs and LSCs by OXPHOS despite the hypoxic conditions in their niches with FAO and amino acids providing its substrate. These metabolic processes appeared to be under tight control of cellular regulation mechanisms which are discussed in depth. However, our conclusion is that systemic therapeutic targeting of ATP production via glycolysis or OXPHOS is not an attractive option because of its unwanted side effects in cancer patients.

The shape and spatial organization -the anatomy- of a tissue profoundly influences its function. Knowledge of the anatomical relationships between parent and daughter cells is necessary to understand differentiation and how the crosstalk between the different cells in the tissue leads to physiological maintenance and pathological perturbations. Blood cell production takes place in the bone marrow through the progressive differentiation of stem cells and progenitors. These are maintained and regulated by a heterogeneous microenvironment composed of stromal and hematopoietic cells. While hematopoiesis has been studied in extraordinary detail through functional and multiomics approaches, much less is known about the spatial organization of blood production and how local cues from the microenvironment influence this anatomy. Here, we discuss some of the studies that revealed a complex anatomy of hematopoiesis where discrete local microenvironments spatially organize and regulate specific subsets of hematopoietic stem cells and/or progenitors. We focus on the open questions in the field and discuss how new tools and technological advances are poised to transform our understanding of the anatomy of hematopoiesis.

The lung cancer stem cell (LuCSC) model comprises an attractive framework to explore acquired drug resistance in non-small cell lung cancer (NSCLC) treatment. Here, we used NSCLC cell line model to translate cellular heterogeneity into tractable populations to understand the origin of lung cancers and drug resistance. The epithelial LuCSCs, presumably arising from alveolar bipotent stem/progenitor cells, were lineage naive, noninvasive, and prone to creating aggressive progeny expressing AT2/AT1 markers. LuCSC-holoclones were able to initiate rimmed niches, where their specialization created pseudo-alveoli structures. Mechanistically, LuCSC transitioning from self-renewal (β-catenin and Nanog signaling) to malignant lineage differentiation is regulated by EGFR activation and the inverse inhibition of tumor suppressor MIG6. We further identified the functional roles of endogenous EGFR signaling in mediating progeny invasiveness and their ligands in LuCSC differentiation. Importantly, drug screening demonstrated that EGFR driving progeny were strongly responsive to TKIs; however, the LuCSCs were exclusively resistant but sensitive to AMPK agonist Metformin, antibiotic Salinomycin and to a lesser degree Carboplatin. Our data reveals previously an unknown mechanism of NSCLC resistance to EGFR-TKIs, which is associated with LuCSCs bearing a silenced EGFR and inversely expressed MIG6 suppressor gene. Taken altogether, successful NSCLC treatment requires development of a novel combination of drugs, efficiently targeting both LuCSCs and heterogeneous progeny.

Hematopoietic stem cells (HSCs) are responsible for life-long production of all mature blood cells. Under homeostasis, HSCs in their native bone marrow niches are believed to undergo asymmetric cell divisions (ACDs), with one daughter cell maintaining HSC identity and the other committing to differentiate into various mature blood cell types. Due to the lack of key niche signals, in vitro HSCs differentiate rapidly, making it challenging to capture and study ACD. To overcome this bottleneck, in this study, we used interferon alpha (IFN&alpha, ) treatment to &rdquo, pre-instruct&rdquo, HSC fate directly in their native niche, and then systematically studied the fate of dividing HSCs in vitro at the single cell level via time-lapse analysis, as well as multigene and protein expression analysis. Triggering HSCs&rsquo, exit from dormancy via IFN&alpha, was found to significantly increase the frequency of asynchronous divisions in paired daughter cells (PDCs). Using single-cell gene expression analyses, we identified 12 asymmetrically expressed genes in PDCs. Subsequent immunocytochemistry analysis showed that at least three of the candidates, i.e., Glut1, JAM3 and HK2, were asymmetrically distributed in PDCs. Functional validation of these observations by colony formation assays highlighted the implication of asymmetric distribution of these markers as hallmarks of HSCs, for example, to reliably discriminate committed and self-renewing daughter cells in dividing HSCs. Our data provided evidence for the importance of in vivo instructions in guiding HSC fate, especially ACD, and shed light on putative molecular players involved in this process. Understanding the mechanisms of cell fate decision making should enable the development of improved HSC expansion protocols for therapeutic applications.

Various services based on sharing have recently emerged in cities across the globe. They entail promises of a more sustainable mode of transportation. Through an in-depth case study, this paper follows the undertakings within a recent demonstration project aimed at implementing a residential bike-sharing service. The involved actors succeeded in presenting a functional service but failed to make the service commercially viable. Consequently, the configuration did not spread beyond the niche level. Based on this finding, the paper identifies a risk in enforcing progressive solutions to becoming commercialised, and urges public actors to play a greater role in steering the transition towards a more sustainable future transport system. By shedding light on the process of failing, the paper contributes important lessons concerning the setup of future bike-sharing systems that can challenge the automobility regime. © 2020 The Authors

11 p.-4 fig., Bacterial resistance to antibiotics poses enormous health and economic burdens to our society, and it is of the essence to explore old and new ways to deal with these problems. Here we review the current status of multi-resistance genes and how they spread among bacteria. We discuss strategies to deal with resistant bacteria, namely the search for new targets and the use of inhibitors of protein-protein interactions, fragment-based methods, or modified antisense RNAs. Finally, we discuss integrated approaches that consider bacterial populations and their niches, as well as the role of global regulators that activate and/or repress the expression of multiple genes in fluctuating environments and, therefore, enable resistant bacteria to colonize new niches. Understanding how the global regulatory circuits work is, probably, the best way to tackle bacterial resistance., While writing this review, authors were supported by Grant BIO2016-76412- C2-2-R AEI/FEDER (to A.B.) from the Ministry of Science,Innovation and Universities.

International audience; The emergence of carbapenem-resistant enterobacterial species poses a serious threat to public health worldwide. OXA-48-type carbapenem-hydrolyzing class D β-lactamases are widely distributed among Enterobacteriaceae, with significant geographical differences. To date, 11 OXA-48-like variants have been identified, with classical OXA-48 being the most widespread. These enzymes show high-level hydrolytic activity against penicillins and low-level hydrolysis towards carbapenems. Since the first description of the OXA-48 carbapenemase in Turkey, bacterial strains producing the enzyme have been extensively reported in nosocomial and community outbreaks in many parts of the word, particularly in the Mediterranean area and European countries. The rapid spread of Enterobacteriaceae producing OXA-48-like enzymes in different ecosystems has become a serious issue recently. The number of reservoirs for such organisms is increasing, not only in hospitals, but also in the community, among animals (e.g., livestock, companion animals, and wildlife) and in the environment. This review aims to summarize the main characteristics of the OXA-48-type carbapenemases, covering genetic and enzymatic traits, their epidemiology, clonality and associated genes, correlation with extended-spectrum β-lactamases (ESBLs) or plasmidic AmpC (pAmpC) in different bacterial species worldwide.

Glioblastoma is the most aggressive and malignant primary brain tumor in adults and has a poor patient survival of only 20 months after diagnosis. This poor patient survival is at least partly caused by glioblastoma stem cells (GSCs), which are slowly-dividing and therefore therapy-resistant. GSCs are localized in protective hypoxic peri-arteriolar niches where these aforementioned stemness properties are maintained. We previously showed that hypoxic peri-arteriolar GSC niches in human glioblastoma are functionally similar to hypoxic peri-arteriolar hematopoietic stem cell (HSC) niches in human bone marrow. GSCs and HSCs express the receptor C-X-C receptor type 4 (CXCR4), which binds to the chemoattractant stromal-derived factor-1α (SDF-1α), which is highly expressed in GSC niches in glioblastoma and HSC niches in bone marrow. This receptor−ligand interaction retains the GSCs/HSCs in their niches and thereby maintains their slowly-dividing state. In acute myeloid leukemia (AML), leukemic cells use the SDF-1α−CXCR4 interaction to migrate to HSC niches and become slowly-dividing and therapy-resistant leukemic stem cells (LSCs). In this communication, we aim to elucidate how disruption of the SDF-1α−CXCR4 interaction using the FDA-approved CXCR4 inhibitor plerixafor (AMD3100) may be used to force slowly-dividing cancer stem cells out of their niches in glioblastoma and AML. Ultimately, this strategy aims to induce GSC and LSC differentiation and their sensitization to therapy.

The adaptive immune system has the unique ability to remember and rapidly mount protective response against previously encountered pathogen. This feature of the immune system is termed immunological memory and the functional duty is carried out by differentiated T and B cells. The maintenance of memory cells is important to confer long-lasting protection for the organism. Also, strategic positioning of these memory cells throughout the organism in crucial to ensure timely response against recurrent antigenic stimulation. While the tissue distribution and maintenance of memory T and plasma cells has been described, the lifestyle of memory B cells (Bmem) has not been well studied so far. In my doctoral thesis I investigated the tissue organization and lifestyle of memory B cells in mice, which would serve as the starting point for further translational studies in humans. To determine the tissue distribution, isotype-switched Bmem of spleen, bone marrow (BM), peripheral blood, and lymph nodes were enumerated under different immunization and infection protocols. The majority of isotype-switched Bmem were localized in the spleen, but a significant population was also contained within the BM. Comparison of the repertoire of B cell receptor (BCR), a unique identifier of each individual B cell, the repertoires of isotype-switched Bmem of spleen and BM revealed limited overlap of B cells with same BCR (clonotypes) generated during a specific immune response. The majority of Bmem clonotypes are expressed exclusively in either organ, demonstrating that isotype-switched Bmem of the two organs represent distinct resident populations with minimal exchange between them via blood circulation. Phenotypically, isotype-switched Bmem of the two organs differ in surface protein expression of CD21 (complement receptor) and CD62L (L-selectin) with subsets of CD21low and CD21high populations in the BM but not in spleen, also, isotype-switched Bmem of BM express higher levels of CD62L compared to those in spleen. Isotype-switched Bmem of BM and spleen are resting in the G0 of cell cycle as determined by the expression of the proliferative marker Ki67, and are refractory to in vivo treatment with cyclophosphamide (a DNA alkylating agent which kills proliferating cells). In the BM, isotype-switched Bmem are located in close proximity to reticular stromal cells expressing VCAM-1. To further understand the role of BM stromal cells in organization of survival niches for memory cells, the biology and functional properties of VCAM-1+ stromal cells were analyzed. Next generation sequencing single cell mRNA transcriptomes profiling of directly ex vivo isolated BM VCAM-1+ stromal cells revealed distinct subpopulation of stromal cells defined by the expression of cytokines and chemokines which have been described to be important for the maintenance and survival of subsets of hematopoietic and immune cells subsets. Altogether, the findings of this thesis demonstrate that murine Bmem are residing in the BM as distinct population of B cell memory and that distinct subsets of BM stromal cells organize survival niches for different hematopoietic cells, including memory cells., Das adaptive Immunsystem hat die einzigartige Fähigkeit sich an zuvor vorgefundene Krankheitserreger zu erinnern und schnell gegen diese schützend zu reagieren. Diese Fähigkeit des Immunsystems heißt immunologisches Gedächtnis und wird von differenzierten T- und B-Zellen ausgeübt. Die Aufrechterhaltung der Gedächtniszellen ist wichtig um dem Organismus einen dauerhaften Schutz zu verleihen. Weiterhin ist eine strategische Positionierung dieser Gedächtniszellen im gesamten Organismus entscheidend um eine zeitnahe Reaktion gegen wiederkehrende antigenische Stimulierungen sicherzustellen. Während die Gewebeverteilung und -erhaltung von Gedächtnis T-Zellen und Plasma Zellen bereits gut beschrieben wurde, wurde der Lebensstil von Gedächtnis-B-Zellen (Bmem) bislang nicht weiter untersucht. In meiner Doktorarbeit habe ich die Gewebeorganisation und den Lebensstil von Gedächtnis B-Zellen in Mäusen untersucht, welche als Ausgangspunkt für weitere translationale Studien am Menschen dient. Um die Gewebeverteilung zu untersuchen, wurden Isotyp-veränderte Bmem der Milz, des Knochenmarks, von peripherem Blut und von Lymphknoten unter verschiedenen Immunisierungs- und Infektionsprotokollen ausgezählt. Die Mehrheit der Isotyp-veränderten Bmem wurden in der Milz lokalisiert, es ist aber auch eine signifikante Population im Knochenmark enthalten. Der Vergleich des Repertoires des B-Zell-Rezeptors (BCR), einer einzigartigen Bezeichnung jeder individuellen B-Zelle, des Repertoires der Isotyp-veränderten Bmem der Milz und des Knochenmarks hat gezeigt, dass B-Zellen mit derselben BCR (Klonotypen), welche während einer spezifischen Immunreaktion generiert wurden, sich kaum überlappen. Die Mehrheit der Bmem Klonotypen wird ausschließlich in einem der beiden Organe exprimiert, was beweist, dass Isotyp-veränderte Bmem zweier Organe verschiedene Populationen mit nur minimalen Austausch über den Blutkreislauf repräsentieren. Phänotypisch, Isotyp-veränderte Bmem der beiden Organe unterscheiden sich in der Oberflächenproteinexpression von CD21 (Komplementrezeptor) und CD62L (L-Selektin) mit Untergruppen von CD21low und CD21high Populationen im Knochenmark, aber nicht in der Milz. Weiterhin beinhalten Isotyp-veränderte Bmem des Knochenmarks einen höheren Spiegel von CD62L im Vergleich zur Milz. Isotyp-veränderte Bmem des Knochenmarks und der Milz ruhen im G0 des Zellzyklus, wie durch die Expression des proliferativen Markers Ki67 bestimmt wurde, und sind widerstandsfähig gegen in vivo Behandlungen mit Cyclophosphamid (einem DNA alkylierenden Wirkstoff, der wuchernde Zellen tötet). Im Knochenmark befinden sich Isotyp-veränderte Bmem in unmittelbarer Nähe zu retikulären Stromazellen, die VCAM-1 exprimieren. Um die Rolle der Knochenmark Stromazellen bei der Organisation von Überlebensnischen für Gedächtniszellen genauer zu verstehen, wurden die Biologie und die funktionellen Eigenschaften von VCAM-1+ Stromazellen analysiert. Die Sequenzierung von Einzelzell-mRNA-Transkriptomen von direkt ex vivo isolierten Knochenmark VCAM-1+ Stromazellen ergab eindeutige Subpopulationen von Stromazellen, die durch die Expression von Cytokinen und Chemokinen definiert wurden, welche als wichtig für die Aufrechterhaltung und das Überleben von Subpopulationen von hämatopoetischen Zellen und Immunzellen beschrieben wurden. Zusammenfassend beschreiben die Ergebnisse der Dissertation, dass murine Bmem im Knochenmark als ausgeprägte ansässige Population des B-Zell-Gedächtnisses vorkommen und das Subpopulationen von Knochenmark Stromazellen Überlebensnischen für verschiedene hämatopoetische Zellen, inklusive den Gedächtniszellen des Immunsystems, organisieren.

Glioblastoma is the most aggressive primary brain tumor. Slowly dividing and therapy-resistant glioblastoma stem cells (GSCs) reside in protective peri-arteriolar niches and are held responsible for glioblastoma recurrence. Recently, we showed similarities between GSC niches and hematopoietic stem cell (HSC) niches in bone marrow. Acute myeloid leukemia (AML) cells hijack HSC niches and are transformed into therapy-resistant leukemic stem cells (LSCs). Current clinical trials are focussed on removal of LSCs out of HSC niches to differentiate and to become sensitized to chemotherapy. In the present study, we elaborated further on these similarities by immunohistochemical analyses of 17 biomarkers in paraffin sections of human glioblastoma and human bone marrow. We found all 17 biomarkers to be expressed both in hypoxic peri-arteriolar HSC niches in bone marrow and hypoxic peri-arteriolar GSC niches in glioblastoma. Our findings implicate that GSC niches are being formed in glioblastoma as a copy of HSC niches in bone marrow. These similarities between HSC niches and GSC niches provide a theoretic basis for the development of novel strategies to force GSCs out of their niches, in a similar manner as in AML, to induce GSC differentiation and proliferation to render them more sensitive to anti-glioblastoma therapies.

Summary We earlier showed that outside-in integrin signaling through POSTN-ITGAV interaction plays an important role in regulating adult hematopoietic stem cell (HSC) quiescence. Here, we show that Itgav deletion results in increased frequency of phenotypic HSCs in fetal liver (FL) due to faster proliferation. Systemic deletion of Postn led to increased proliferation of FL HSCs, albeit without any loss of stemness, unlike Vav-Itgav−/− HSCs. Based on RNA sequencing analysis of FL and bone marrow HSCs, we predicted the involvement of DNA damage response pathways in this dichotomy. Indeed, proliferative HSCs from Postn-deficient FL tissues showed increased levels of DNA repair, resulting in lesser double-strand breaks. Thus POSTN, with its expression majorly localized in the vascular endothelium of FL tissue, acts as a regulator of stem cell pool size during development. Overall, we demonstrate that the duality of response to proliferation in HSCs is developmental stage dependent and can be correlated with DNA damage responses., Graphical Abstract, Highlights • Interruption of POSTN-ITGAV interaction leads to HSC expansion in fetal liver • HSC from fetal liver in comparison with adult BM excel in their DNA damage responses • POSTN is a potential component of the vascular niche for HSCs in the fetal liver, Khurana and colleagues demonstrate here a role of outside-in integrin signaling in regulating the size of hematopoietic stem cell (HSC) pool in mouse fetal liver. Using Postn−/− and Vav-Itgav−/− mice, they show that HSCs lacking POSTN-ITGAV interaction proliferate faster without any functional loss. The study proposes a possible role of increased levels of DNA damage responses in tolerating proliferation stress during fetal development.

Hematopoietic stem cells (HSC) are the most powerful type of adult stem cell found in the body. Hematopoietic stem cells are multipotent and capable of giving rise to all other types of hematopoietic cells found in the organism. A single HSC is capable of regenerating a functional hematopoietic system when transplanted into a recipient. Hematopoietic stem cells reside in the bone marrow in specific multicellular structures called niches. These niches are indispensable for maintaining and regulating HSC numbers and function. It has become increasingly clearer that HSC and their niches can also be regulated by migrating leukocytes. Here we will discuss the composition of murine bone marrow niches and how HSC and their niches are regulated by different types of leukocytes that traffic between the periphery and the niche. Unless otherwise indicated all the studies discussed below were performed in mouse models.

Cell-based tissue engineering offers great promise to regenerative therapy, but so far it has been restricted due to insufficient number of cells obtained from donors and the lack of efficient ways of delivering them to target sites. This study shows, for the first time, the ability of a composite scaffold of gelatin and poly(methyl vinyl ether- alt-maleic anhydride) (GP-2) as a niche for expansion and multilineage differentiation ability of human umbilical cord–derived mesenchymal stem cells. First, the in vivo biocompatibility of scaffolds was checked by subcutaneous implantation of scaffolds in male Wistar rats for up to 45 days. Hematological parameters and histology of skin near implanted region rule out the probability of any adverse effects due to the scaffolds. The isolated human umbilical cord–derived mesenchymal stem cells were seeded on to pre-optimized scaffolds and induced to differentiate into osteogenic and adipogenic lineages by culturing in respective induction media. The human umbilical cord–derived mesenchymal stem cells were found to be viable and proliferated well on scaffolds when assessed with live/dead and PicoGreen assay. The biochemical assays such as alkaline phosphatase activity and triglycerides estimation confirmed the differentiation of cells toward particular lineages when cultured on scaffolds with appropriate inductive media. The study exhibited the proficiency of scaffold GP-2 for mesenchymal stem cells’ adherence, proliferation, and differentiation and also showed its engraftment efficiency. Taken together, our study establishes the in vivo biocompatibility of composite scaffold and, importantly, indicates its potential for stem cell–based therapy.

The microenvironment of malignant gliomas is described according to its definition in the literature. Beside tumor cells, a series of stromal cells (microglia/macrophages, pericytes, fibroblasts, endothelial cells, normal and reactive astrocytes) represents the cell component, whereas a complex network of molecular signaling represents the functional component. Its most evident expressions are perivascular and perinecrotic niches that are believed to be the site of tumor stem cells or progenitors in the tumor. Phenotypically, both niches are not easily recognizable; here, they are described together with a critical revision of their concept. As for perinecrotic niches, an alternative interpretation is given about their origin that regards the tumor stem cells as the residue of those that populated hyperproliferating areas in which necroses develop. This is based on the concept that the stem-like is a status and not a cell type, depending on the microenvironment that regulates a conversion of tumor non-stem cells and tumor stem cells through a cell reprogramming.

Pigs are one of the most important economic livestock. Gut microbiota is not only critical to the health but also the production efficiency of pigs. Manipulating gut microbiota relies on the full view of gut microbiome and the understanding of drive forces shaping microbial communities. 16s rDNA sequencing was used to profile microbiota along the longitudinal and radical axes to obtain the topographical map of microbiome in different intestinal compartments in young pigs. Alpha and beta-diversities revealed distinct differences in microbial compositions between the distal ileum and cecum and colon, as well as between the lumen and mucosa. Firmicutes and Proteobacteria dominated in the ileum, constituting 95 and 80% of the luminal and mucosa-attached microbiome. Transitioning from the small intestine to the large intestine, luminal Bacteroidetes increased from 1.69 to 45.98% in the cecum and 40.09% in the colon, while mucosal Bacteroidetes raised from 9 to 35.36% and 27.96%. Concurrently, luminal Firmicutes and Proteobacteria and mucosal-attached Proteobacteria remarkably decreased. By co-occurrence network analyses, Prevotellaceae, Ruminococcaceae, Lachnospiraceae and Veillonellaceae were recognized as the central nodes of luminal microbial network, and Prevotellaceae and Enterobacteriaceae, Caulobacteraceae, Enterococcaceae, Xanthomonadaceae, Pseudomonadaceae were identified as mucosal central nodes. Co-abundance was uncovered among Prevotellaceae, Lachnospiraceae, and Veillonellaceae in the luminal and mucosal microbiome, while opportunistic pathogens from γ-Proteobacteria in the mucosa. Strong co-exclusion was shown between Enterobacteriaceae with Prevotellaceae-centered microbial groups in the lumen. Redundancy analysis found bile acids and short chain fatty acids explained 37.1 and 41% of variations in the luminal microbial composition, respectively. Primary bile acid, taurine- and glycine- conjugated bile acids were positively correlated with Lactobacillaceae, Enterobacteriaceae, Clostridiaceae_1, Peptostreptococcaceae, whereas secondary bile acids, acetate, propionate, butyrate, and valerate were positively correlated with Prevotellaceae, Acidaminococcaceae, Ruminococcaceae, Lachnospiraceae, Desulfovibronaceae, Veillonellaceae. Functional analyses demonstrated that Prevotella, Veillonellaceae, Lachnospiraceae, and Ruminococcaceae were positively correlated with gene functions related to amino acids, energy, cofactors and vitamins metabolism, which are indispensable for the hosts. These results suggested site specific colonization and co-occurrence of swine gut microbiome closely relate to the microenvironment in each niche. Interactions of core gut microbiome greatly contributed to metabolism and/or immunity in the swine intestine.

Dental pulp stem/progenitor cells guarantee tooth homeostasis, repair and regeneration throughout life. The decision between renewal and differentiation of these cells is influenced by physical and molecular interactions with stromal cells and extracellular matrix molecules forming the specialized microenvironment of dental pulp stem cell niches. Here we study the activation of putative pulp niches after tooth injury through the upregulation of Notch signaling pathway. Notch1, Notch2, and Notch3 molecules were used as markers of dental pulp stem/progenitor cells. Upon dental injury, Notch1 and Notch3 are detected in cells related to vascular structures suggesting a role of these proteins in the activation of specific pulpal perivascular niches. In contrast, a population of Notch2-positive cells that are actively proliferative is observed in the apical part of the pulp. Kinetics of these cells is followed up with a lipophilic DiI labeling, showing that apical pulp cells migrate toward the injury site where dynamic regenerative/repair events occur. The knowledge of the activation and regulation of dental pulp stem/progenitor cells within their niches in pathologic conditions may be helpful for the realization of innovative dental treatments in the near future.

In mammals, megakaryocytes (MKs) in the bone marrow (BM) produce blood platelets, required for hemostasis and thrombosis. MKs originate from hematopoietic stem cells and are thought to migrate from an endosteal niche towards the vascular sinusoids during their maturation. Through imaging of MKs in the intact BM, here we show that MKs can be found within the entire BM, without a bias towards bone-distant regions. By combining in vivo two-photon microscopy and in situ light-sheet fluorescence microscopy with computational simulations, we reveal surprisingly slow MK migration, limited intervascular space, and a vessel-biased MK pool. These data challenge the current thrombopoiesis model of MK migration and support a modified model, where MKs at sinusoids are replenished by sinusoidal precursors rather than cells from a distant periostic niche. As MKs do not need to migrate to reach the vessel, therapies to increase MK numbers might be sufficient to raise platelet counts., Megakaryocyte maturation is thought to occur as the cells migrate from a vessel-distant (endosteal) niche to the vessel within the bone. Here, the authors show that megakaryocytes represent largely sessile cells in close contact with the vasculature and homogeneously distributed in the bone marrow.

Glioblastoma multiforme (GBM) harbors are not only rapidly dividing cells but also small populations of slowly dividing and dormant cells with tumorigenesity, self-renewal, and multi-lineage differentiation capabilities. Known as glioblastoma stem cells (GSCs), they are resistant to conventional chemo- and radiotherapy and may be a causative factor in recurrence. The treatment outcome in patients with GBM remains unsatisfactory and their mean survival time has not improved sufficiently. We studied clinical evidence and basic research findings to assess the possibility of new treatment strategies that target GSCs and their specific microenvironments (GBM niches) and raise the possibility of adding new treatments to eradicate GSCs and GBM niches.

Stem cells guarantee tissue repair and regeneration throughout life. The decision between cell self-renewal and differentiation is influenced by a specialized microenvironment called the ‘stem cell niche’. In the tooth, stem cell niches are formed at specific anatomic locations of the dental pulp. The microenvironment of these niches regulates how dental pulp stem cell populations participate in tissue maintenance, repair, and regeneration. Signaling molecules such as Notch proteins are important regulators of stem cell function, with various capacities to induce proliferation or differentiation. Dental injuries often lead to odontoblast apoptosis, which triggers activation of dental pulp stem cells followed by their proliferation, migration, and differentiation into odontoblast-like cells, which elaborate a reparative dentin. Better knowledge of the regulation of dental pulp stem cells within their niches in pathological conditions will aid in the development of novel treatments for dental tissue repair and regeneration.

Stroke affects one in every six people worldwide, and is the leading cause of adult disability. After stroke, some limited spontaneous recovery occurs, the mechanisms of which remain largely unknown. Multiple, parallel approaches are being investigated to develop neuroprotective, reparative and regenerative strategies for the treatment of stroke. For years, clinical studies have tried to use exogenous cell therapy as a means of brain repair, with varying success. Since the rediscovery of adult neurogenesis and the identification of adult neural stem cells in the late nineties, one promising field of investigation is focused upon triggering and stimulating this self-repair system to replace the neurons lost following brain injury. For instance, it is has been demonstrated that the adult brain has the capacity to produce large numbers of new neurons in response to stroke. The purpose of this review is to provide an updated overview of stroke-induced adult neurogenesis, from a cellular and molecular perspective, to its impact on brain repair and functional recovery.

Leukemic stem cells (LSCs) reside within bone marrow niches that maintain their relatively quiescent state and convey resistance to conventional treatment. Many of the microenvironmental signals converge on RAC GTPases. Although it has become clear that RAC proteins fulfill important roles in the hematopoietic compartment, little has been revealed about the downstream effectors and molecular mechanisms. We observed that in BCR-ABL-transduced human hematopoietic stem/progenitor cells (HSPCs) depletion of RAC2 but not RAC1 induced a marked and immediate decrease in proliferation, progenitor frequency, cobblestone formation and replating capacity, indicative for reduced self-renewal. Cell cycle analyses showed reduced cell cycle activity in RAC2-depleted BCR-ABL leukemic cobblestones coinciding with an increased apoptosis. Moreover, a decrease in mitochondrial membrane potential was observed upon RAC2 downregulation, paralleled by severe mitochondrial ultrastructural malformations as determined by automated electron microscopy. Proteome analysis revealed that RAC2 specifically interacted with a set of mitochondrial proteins including mitochondrial transport proteins SAM50 and Metaxin 1, and interactions were confirmed in independent co-immunoprecipitation studies. Downregulation of SAM50 also impaired the proliferation and replating capacity of BCR-ABL-expressing cells, again associated with a decreased mitochondrial membrane potential. Taken together, these data suggest an important role for RAC2 in maintaining mitochondrial integrity.

A great challenge when conducting ex vivo studies of leukaemia is the construction of an appropriate experimental platform that would recapitulate the bone marrow (BM) environment. Such a 3D scaffold system has been previously developed in our group [1]. Additionally to the BM architectural characteristics, parameters such as oxygen and glucose concentration are crucial as their value could differ between patients as well as within the same patient at different stages of treatment, consequently affecting the resistance of leukaemia to chemotherapy. The effect of oxidative and glucose stress—at levels close to human physiologic ones—on the proliferation and metabolic evolution of an AML model system (K-562 cell line) in conventional 2D cultures as well as in 3D scaffolds were studied. We observed that the K-562 cell line can proliferate and remain alive for 2 weeks in medium with glucose close to physiological levels both in 20 and 5 % O2. We report interesting differences on the cellular response to the environmental, i.e., oxidative and/or nutritional stress stimuli in 2D and 3D. Higher adaptation to oxidative stress under non-starving conditions is observed in the 3D system. The glucose level in the medium has more impact on the cellular proliferation in the 3D compared to the 2D system. These differences can be of significant importance both when applying chemotherapy in vitro and also when constructing mathematical tools for optimisation of disease treatment.

Upon transplant, functional HSC clones preferentially expand in certain skeletal locations, exhibiting only limited migration toward other niches., Hematopoietic stem cells (HSCs) are able to migrate through the blood stream and engraft bone marrow (BM) niches. These features are key factors for successful stem cell transplantations that are used in cancer patients and in gene therapy protocols. It is unknown to what extent transplanted HSCs distribute throughout different anatomical niches in the BM and whether this changes with age. Here we determine the degree of hematopoietic migration at a clonal level by transplanting individual young and aged mouse HSCs labeled with barcoded viral vector, followed by assessing the skeletal distribution of hundreds of HSC clones. We detected highly skewed representation of individual clones in different bones at least 11 mo after transplantation. Importantly, a single challenge with the clinically relevant mobilizing agent granulocyte colony-stimulating factor (G-CSF) caused rapid redistribution of HSCs across the skeletal compartments. Old and young HSC clones showed a similar level of migratory behavior. Clonal make-up of blood of secondary recipients recapitulates the barcode composition of HSCs in the bone of origin. These data demonstrate a previously unanticipated high skeletal disequilibrium of the clonal composition of HSC pool long-term after transplantation. Our findings have important implications for experimental and clinical and stem cell transplantation protocols.

The vertical distribution of pollinators is an important component in the foraging pattern of plants strata, and it influences the reproductive system (pollen/ovule ratio) and seed/ovule ratio. Niches in two different strata from Adesmia tristis Vogel were evaluated in these aspects. This plant is an endemic shrub from the Campos de Cima da Serra in Southern Brazil. The studies were carried out from January 2010, to January 2011, at Pró-Mata/PUCRS (Catholic University of Rio Grande do Sul) (29°27′-29°35′S and 50°08′-50°15′W), São Francisco de Paula, sate of Rio Grande do Sul, Brazil. Breeding system of A. tristis is mandatory allogamy. The vertical profile in A. tristis has differentiated foraging niches among the most common pollinators. Bees of Megachile genus forage in the upper stratum, and representative bees of the Andrenidae family explore the lower stratum. The upper stratum of the vertical profile had more contribution to seed production. Adesmia tristis showed evidence of pollination deficit. A distribuição vertical dos polinizadores é um importante componente no padrão de forrageamento nos estratos das plantas e influencia o sistema reprodutivo (relação pólen/óvulo) e a razão semente/óvulo. Nichos em dois estratos diferentes de Adesmia tristis Vogel foram avaliados quanto a esses aspectos. Essa planta é um arbusto endêmico dos campos de Cima da Serra no Sul do Brasil. Os estudos ocorreram de janeiro de 2010 a janeiro de 2011, no Pró-Mata/PUCRS (Pontifícia Universidade Católica do Rio Grande do Sul) (29°27′-29°35′S e 50°08′-50°15′W), São Francisco de Paula, estado do Rio Grande do Sul, Brasil. O sistema reprodutivo de A. tristis é alogamia obrigatória. O perfil vertical em A. tristis possui diferentes nichos de forrageamento entre os polinizadores mais comuns. Abelhas do gênero Megachile forrageiam no estrato superior e as abelhas representantes da família Andrenidae exploram o estrato inferior. O estrato superior do perfil vertical contribui mais na produção de sementes. Adesmia tristis apresentou evidências de déficit de polinização.

We report a reliable strategy to perform automated image cytometry of single (non-adherent) stem cells captured in microfluidic traps. The method rapidly segments images of an entire microfluidic chip based on the detection of horizontal edges of microfluidic channels, from where the position of the trapped cells can be derived and the trapped cells identified with very high precision (>97%). We used this method to successfully quantify the efficiency and spatial distribution of single-cell loading of a microfluidic chip comprised of 2048 single-cell traps. Furthermore, cytometric analysis of trapped primary hematopoietic stem cells (HSC) faithfully recapitulated the distribution of cells in the G1 and S/G2-M phase of the cell cycle that was measured by flow cytometry. This approach should be applicable to automatically track single live cells in a wealth of microfluidic systems.