Haematopoietic stem cells in spleen have distinct differentiative potential for antigen presenting cells

Dendritic cells (DC) are known to develop from macrophage dendritic progenitors (MDP) in bone marrow (BM), which give rise to conventional (c)DC and monocytes, both dominant antigen presenting cell (APC) subsets in spleen. This laboratory has however defined a distinct dendritic‐like cell subset in spleen (L‐DC), which can also be derived in long‐term cultures of spleen. In line with the restricted in vitro development of only L‐DC in these stromal cultures, we questioned whether self‐renewing HSC or progenitors exist in spleen with restricted differentiative capacity for only L‐DC. Neonatal spleen and BM were compared for their ability to reconstitute mice and to give rise to L‐DC, as well as other splenic APC. Neonatal spleen cells were transplanted into allotype‐distinct lethally irradiated hosts along with host‐type competitor BM cells, and assayed over 8 to 51 weeks for haematopoietic reconstitution of L‐DC and cDC subsets, along with other lymphoid and myeloid cells. In this study, neonatal spleen showed multilineage haematopoietic reconstitution in mouse chimeras, rather than specific or restricted ability to differentiate into L‐DC. However, the representation of individual APC subsets was found to be unequal in chimeras partially reconstituted with donor cells, such that more donor‐derived progeny were seen for L‐DC than for myeloid and cDC subsets. The ability of HSC in spleen to develop into L‐DC was indicated by a strong bias in the subset size of these cells over other splenic APC subsets. This type of evidence supports a model whereby spleen represents an important site for haematopoiesis of this distinct DC subset. The conditions under which haematopoiesis of L‐DC occurs in spleen, or the progenitors involved, will require further investigation.     

Development of ramified microglia from early macrophages in the zebrafish optic tectum

Microglia, the resident macrophage precursors of the brain, are necessary for the maintenance of tissue homeostasis and activated by a wide range of pathological stimuli. They have a key role in immune and inflammatory responses. Early microglia stem from primitive macrophages, however the transition from early motile forms to the ramified mature resident microglia has not been assayed in real time. In order to provide such an assay, we used zebrafish transgenic lines in which fluorescent reporter expression is driven by the promoter of 1 (mpeg1; Ellet et al. [2011]: Blood 117(4): e49–e56,). This enabled the investigation of the development of these cells in live, intact larvae. We show that microglia develop from highly motile amoeboid cells that are engaged in phagocytosis of apoptotic cell bodies into a microglial cell type that rapidly morphs back and forth between amoeboid and ramified morphologies. These morphing microglia eventually settle into a typical mature ramified morphology. Developing microglia frequently come into contact with blood capillaries in the brain, and also frequently contact each other. Up to 10 days postfertilization, microglia were observed to undergo symmetric division. In the adult optic tectum, the microglia are highly branched, resembling mammalian microglia. In addition, the mpeg1 transgene also labeled highly branched cells in the skin overlying the optic tectum from 8–9 days postfertilization, which likely represent Langerhans cells. Thus, the development of zebrafish microglia and their cellular interactions was studied in the intact developing brain in real time and at cellular resolution. © 2012 Wiley Periodicals, Inc. Develop Neurobiol, 2013     

Identification of a novel antigen cross‐presenting cell type in spleen

Antigen-presenting cells (APC), like dendritic cells (DC), are essential for T-cell activation, leading to immunity or tolerance. Multiple DC subsets each play a unique role in the immune response. Here, a novel splenic dendritic-like APC has been characterized in mice that has immune function and cell surface phenotype distinct from other, described DC subsets. These were identified as a cell type continuously produced in spleen long-term cultures (LTC) and have an in vivo equivalent cell type in mice, namely ‘L-DC’. This study characterizes LTC-DC in terms of marker phenotype and function, and compares them with L-DC and other known splenic DC and myeloid subsets. L-DC display a myeloid dendritic-like phenotype equivalent to LTC-DC as CD11c^loCD11b^hiMHC-II⁻CD8α⁻ cells, distinct by high accessibility and endocytic capacity for blood-borne antigen. Both LTC-DC and L-DC have strong antigen cross-presentation ability leading to strong activation of CD8⁺ T cells, particularly after exposure to lipopolysaccharide. However, they have weak ability to stimulate CD4⁺ T cells in antigen-specific responses. Evidence is presented here for a novel DC type produced by in vitro haematopoiesis which has distinct antigen-presenting potential and reflects a DC subset present also in vivo in spleen.     

A New Hypothalamic Polypeptide Is a Regulator of Myelopoiesis

The effect of hypothalamic proline-rich polypeptide (PRP) against Pseudomonas aeruginosa infection in BALB/c mice with leukopenia was investigated. Mice were treated with cyclophosphamide (CPA) and were then injected with PRP 24 h after CPA treatment. The lethal doses of P. aeruginosa were injected to mice when the number of peripheral blood leukocytes reached a nadir on day 5 after CPA administration. The administration of PRP significantly increased the survival of infected mice, and had a pronounced protective effect during the period of development of the infection. The number of bacteria in internal organs of PRP-treated mice was significantly lower than that in control mice. In PRP-treated mice, the neutrophil levels in peripheral blood started to increase 7 days after CPA administration and were consistently higher, and they were more mature than those in controls. Our results may indicate the ability of PRP to stimulate recovery of myelopoiesis and enhance mature neutrophil function.     

Immune complex-trapping cells in the spleen of the chicken

Summary By means of immunohistoperoxidase techniques and the use of HRP-anti-HRP complexes, follicular dendritic cells in chicken spleen can be characterized both at the light-microscopical and ultrastructural level. In contrast to findings in mammals follicular dendritic cells in chicken spleen exhibit evident acid-phosphatase activity and possess considerable numbers of primary lysosomes. After intravenous injection of immune complexes a transient immune complex-trapping occurs in the peripheral parts of the Schweigger-Seidel sheath. The immune complextrapping cells in the Schweigger-Seidel sheath and germinal centre show an identical enzyme histochemical pattern and only minor differences in ultrastructural characteristics.Shortly after intravenous injection of immune complexes and carbon particles these compounds show an identical distribution pattern; however, in the following days these distribution patterns become divergent.     

Histochemical in situ identification of bovine embryonic blood cells reveals differences to the adult haematopoietic system and suggests a close relationship between haematopoietic stem cells and primordial germ cells

Cryostat sections of bovine embryos of exactly known age (obtained from artificial insemination), ranging from 32 to 60 days post-insemination, were treated with a wide range of antibodies directed against cell surface antigens or lineage-specific factors in order to demonstrate different types of fetal blood cells and their precursors. An antibody specific to bovine c-kit (bk-1) stained not only presumptive haematopoietic stem cells in the dorsal aorta and the embryonic liver, but also a subpopulation of putative primordial germ cells in the gonadal anlage, the latter being further characterised by a positive labelling with the lectins STA, WFA and WGA and a histochemical reaction for alkaline phosphatase. The antibody against CD 45, commonly regarded as a pan-leukocyte marker, reacted in the bovine embryo with different types of blood cells, as well as with presumptive vasculogenetic cells and a subpopulation of putative primordial germ cells. CD 61 immunoreaction proved to be a useful tool for demonstrating megakaryocytopoiesis in the embryonic liver, in addition to the lumen of blood vessels and the mesonephros. Staining with BM-2 was restricted to a single population of medium-sized, round to oval cells, forming small groups within the parenchymal strands of the liver. Characterised furthermore by a U-shaped nucleus, this BM-2-positive cell type apparently represents a developmental stage in the granulopoietic lineage. B-lymphocytopoiesis in the bovine liver was detected with antibodies directed against WC-4 and IgM, but not until day 58 post-insemination. Using antibodies to CD 14, no positive results could be obtained in embryonic tissues, although anti-CD 14-positive macrophages were easily recognised in lymph nodes of adult bovines. The antibody against CD 68, however, identified two populations of primitive macrophages in our samples. One population was located in parenchymal strands of the embryonic liver, probably acting as nursing cells for haematopoietic foci, and the other was observed intravasally in the sinusoids of the liver, most probably representing primitive Kupffer cells.     

Endothelial and Leptin Receptor+ cells promote the maintenance of stem cells and hematopoiesis in early postnatal murine bone marrow

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Age-related changes in human hematopoietic stem/progenitor cells

Adult stem cells are critical for maintaining cellular homeostasis throughout life, yet the effects of age on their regenerative capacity are poorly understood. All lymphoid and myeloid blood cell lineages are continuously generated from hematopoietic stem cells present in human bone marrow. With age, significant changes in the function and composition of mature blood cells are observed. In this study, we report that age-related changes also occur in the human hematopoietic stem cell compartment. We find that the proportion of multipotent CD34(+) CD38(-) cells increases in the bone marrow of elderly (>70 years) individuals. CD34(+) CD38(+) CD90(-) CD45RA(+/-) CD10(-) and CD34(+) CD33(+) myeloid progenitors persist at the same level in the bone marrow, while the frequency of early CD34(+) CD38(+) CD90(-) CD45RA(+) CD10(+) and committed CD34(+) CD19(+) B-lymphoid progenitors decreases with age. In contrast to mice models of aging, transplantation experiments with immunodeficient NOD/SCID/IL-2Rγ null (NSG) mice showed that the frequency of NSG repopulating cells does not change significantly with age, and there is a decrease in myeloid lineage reconstitution. An age-related decrease in the capacity of CD34(+) cells to generate myeloid cells was also seen in colony-forming assays in vitro. Thus, with increasing age, human hematopoietic stem/progenitor cells undergo quantitative changes as well as functional modifications.     

Modulation of Myelopoiesis Progenitors Is an Integral Component of Trained Immunity

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Iron particle labeling of haematopoietic progenitor cells: an in vitro study

We present a method for labeling bone marrow haematopoietic progenitor cells with iron particles. Labeling was assessed by magnetic resonance imaging and electron microscopy. Labeling with iron particles could allow the following by imaging techniques of haematopoietic cells in physiologic and pathologic conditions such as the engraftment of haematopoietic progenitor cells or the migration of myelomonocytic cells in inflammatory diseases.     

Oncostatin M production by human dendritic cells in response to bacterial products

Oncostatin M (OSM) is a pleiomorphic cytokine that belongs to the IL-6 cytokine family. It is produced by activated T cells and monocytes/macrophages and plays an important role in the process of inflammatory responses. Although dendritic cells (DCs) have been shown to secrete a variety of cytokines, it is not elucidated whether DCs are able to produce OSM. To clarify this, using human DCs derived from peripheral blood cells, we measured the protein levels of OSM in the supernatants of DC cultures by ELISA and examined the expression of OSM mRNA by RT-PCR after stimulation with lipopolysaccharide (LPS) or fixed Staphylococcus aureus (SACS). Upon stimulation with bacterial products, DCs secreted a large amount of OSM protein in a dose- and time-dependent manner. Concomitantly, the expression of OSM mRNA by DCs was markedly up-regulated. Compared the ability of DCs to produce OSM with that of monocytes, which are major producers of OSM, DCs released significantly higher amounts of OSM protein in the culture supernatants than monocytes. These findings indicate for the first time that human monocyte-derived DCs can synthesize and secrete large amounts of OSM in response to bacterial products, suggesting that OSM produced by DCs at infectious sites may play a role in modulating inflammatory responses.     

Renal ischemia/reperfusion injury inhibits differentiation of dendritic cells derived from bone marrow monocytes in rats

Dendritic cells (DCs) are impacted by surgical injury, exercise, and other physiological stressors. This study aims to determine whether renal I/R injury affects 1) the differentiation of myeloid DCs from bone marrow monocytes (BMMos) and the maturation and activation state of these DCs and 2) DC infiltration of kidney. Sprague-Dawley rats were subjected to I/R injury or sham-operated. Creatinine clearance was monitored daily during the 14 d of reperfusion that followed the ischemic insult. At 2 and 14 d of reperfusion, the following were assessed 1) properties of BMMo-derived DCs (i.e., the amount of generated DCs, differentiation state markers [CD11c, CD80, CD86, and Ia], and functional state [MLR and amount of IL-12 produced]), and 2) the presence of DCs in the kidney. Numbers of BMMo-derived DCs were significantly decreased in the I/R injured group (compared with the sham-operated group) at 2 d but not 14 d. A comparison of the their functionality found mixed lymphocyte response [MLR] and IL-12 production were similar in the two groups at both time points. Also, immunohistochemistry showed infiltrating DCs in the outer medulla of the I/R injured kidney at 2 d but not 14 d of reperfusion. Thus, I/R stress reduces the number of DCs differentiated from BMMos but not the functional activity of these DCs. This decrease may reflect a stress-induced downshift in the capacity of BMMos to differentiate into DCs and a parallel upshift in the capacity of DCs to infiltrate the kidney.     

Lactococcus lactis subsp. lactis JCM 5805 activates natural killer cells via dendritic cells

Lactococcus lactis subsp. lactis JCM 5805 (JCM5805) has been shown to stimulate plasmacytoid dendritic cells (pDC). Here, we investigated the effect of JCM5805 on NK cells. In vitro studies suggested that JCM5805 activated natural killer (NK) cells via dendritic cells including pDC. Furthermore, the oral administration of JCM5805 enhanced the cytotoxic activity of NK cells     

Recapitulative haematopoietic development of human pluripotent stem cells in the absence of exogenous haematopoietic cytokines

To improve the recapitulative quality of human pluripotent stem cell (hPSC) differentiation, we removed exogenous haematopoietic cytokines from the defined differentiation system. Here, we show that endogenous stimuli and VEGF are sufficient to induce robust hPSC-derived haematopoiesis, intensive generation of haematopoietic progenitors, maturation of blood cells and the emergence of definitive precursor cells including those that phenotypically identical to early human embryonic haematopoietic stem cells (HSCs). Moreover, the cytokine-free system produces significantly higher numbers of haematopoietic progenitors compared to the published protocols. The removal of cytokines revealed a broad developmental potential of the early blood cells, stabilized the hPSC-derived definitive precursors and led to spontaneous activation of inflammatory signalling. Our cytokine-free protocol is simple, efficient, reproducible and applicable for embryonic stem cells (ESCs) and induced PSCs. The spectrum of recapitulative features of the novel protocol makes the cytokine-free differentiation a preferred model for studying the early human haematopoietic development.     

Cancer vaccines based on dendritic cells loaded with tumor-associated antigens

Dendritic cells (DCs) can be used as an antigen presentation platform for vaccination against cancer. In this approach, DCs are expanded in vitro from monocyte-derived progenitors, and subsequently loaded with well-characterized tumor-associated antigens (TAAs). TAAs can be incubated with DCs in various forms, including peptides, recombinant proteins, plasmid DNA, formulated RNA, or recombinant viruses. Advantages and limitations of DC-based cellular vaccines against cancers, as well as preliminary results of clinical studies already performed in humans, are discussed. Importantly, significant advances in our understanding of the biology of DCs can be used to support the design of new vaccines or adjuvants in order to elicit T_H1 cellular immune responses. This revised version was published online in July 2006 with corrections to the Cover Date.     

Murine spleen contains a diversity of myeloid and dendritic cells distinct in antigen presenting function

The spleen contains multiple subsets of myeloid and dendritic cells (DC). DC are important antigen presenting cells (APC) which induce and control the adaptive immune response. They are cells specialized for antigen capture, processing and presentation to naïve T cells. However, DC are a heterogeneous population and each subset differs subtly in phenotype, function and location. Similarly, myeloid cell subsets can be distinguished which can also play an important role in the regulation of immunity. This review aims to characterize splenic subsets of DC and myeloid cells to better understand their individual roles in the immune response.