Embryonic Regulation of the Mouse Hematopoietic Niche

Hematopoietic stem cells (HSCs) can differentiate into several types of hematopoietic cells (HCs) (such as erythrocytes, megakaryocytes, lymphocytes, neutrophils, or macrophages) and also undergo self-renewal to sustain hematopoiesis throughout an organism's lifetime. HSCs are currently used clinically as transplantation therapy in regenerative medicine and are typically obtained from healthy donors or cord blood. However, problems remain in HSC transplantation, such as shortage of cells, donor risks, rejection, and graft-versus-host disease (GVHD). Thus, increased understanding of HSC regulation should enable us to improve HSC therapy and develop novel regenerative medicine techniques. HSC regulation is governed by two types of activity: intrinsic regulation, programmed primarily by cell autonomous gene expression, and extrinsic factors, which originate from so-called “niche cells” surrounding HSCs. Here, we focus on the latter and discuss HSC regulation with special emphasis on the role played by niche cells

Intra-Aortic Clusters Undergo Endothelial to Hematopoietic Phenotypic Transition during Early Embryogenesis

Intra-aortic clusters (IACs) attach to floor of large arteries and are considered to have recently acquired hematopoietic stem cell (HSC)-potential in vertebrate early mid-gestation embryos. The formation and function of IACs is poorly understood. To address this issue, IACs were characterized by immunohistochemistry and flow cytometry in mouse embryos. Immunohistochemical analysis revealed that IACs simultaneously express the surface antigens CD31, CD34 and c-Kit. As embryos developed from 9.5 to 10.5 dpc, IACs up-regulate the hematopoietic markers CD41 and CD45 while down-regulating the endothelial surface antigen VE-cadherin/CD144, suggesting that IACs lose endothelial phenotype after 9.5 dpc. Analysis of the hematopoietic potential of IACs revealed a significant change in macrophage CFC activity from 9.5 to 10.5 dpc. To further characterize IACs, we isolated IACs based on CD45 expression. Correspondingly, the expression of hematopoietic transcription factors in the CD45(neg) fraction of IACs was significantly up-regulated. These results suggest that the transition from endothelial to hematopoietic phenotype of IACs occurs after 9.5 dpc

Antiviral Activity and Underlying Action Mechanism of Euglena Extract against Influenza Virus

It is difficult to match annual vaccines against the exact influenza strain that is spreading in any given flu season. Owing to the emergence of drug-resistant viral strains, new approaches for treating influenza are needed. Euglena gracilis (hereinafter Euglena), microalga, used as functional foods and supplements, have been shown to alleviate symptoms of influenza virus infection in mice. However, the mechanism underlying the inhibitory action of microalgae against the influenza virus is unknown. Here, we aimed to study the antiviral activity of Euglena extract against the influenza virus and the underlying action mechanism using Madin–Darby canine kidney (MDCK) cells. Euglena extract strongly inhibited infection by all influenza virus strains examined, including those resistant to the anti-influenza drugs oseltamivir and amantadine. A time-of-addition assay revealed that Euglena extract did not affect the cycle of virus replication, and cell pretreatment or prolonged treatment of infected cells reduced the virus titer. Thus, Euglena extract may activate the host cell defense mechanisms, rather than directly acting on the influenza virus. Moreover, various minerals, mainly zinc, in Euglena extract were found to be involved in the antiviral activity of the extract. In conclusion, Euglena extract could be a potent agent for preventing and treating influenza

ロシア語の移動動詞における接頭辞の付加可能性とその実態および傾向について ―接頭辞iz- / vz-を中心に―

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Expression of CD45 by mouse and human IACs.

<p>Transverse sections of AGM region were made from ICR mouse embryos at 9.5 and 10.5 dpc and from human embryos at 32 day-old, according to the Carnegie classification, stained with antibodies and observed by confocal microscopy. Arrowheads indicate IACs. (<b>A</b>) Mouse IACs in the omphalomesenteric artery (OMA) at 9.5 dpc expressed c-Kit, but not CD45. CD45 (green) and c-Kit (red). Magnified view of IACs is shown at right upper panel in Merge panel. Original magnification is 40x. (<b>B-D</b>) Mouse IACs in the dorsal aorta (DA) (B), OMA (C) and umbilical artery (UA) (D) at 10.5 dpc expressed c-Kit, and some expressed CD45. CD45 (green) and c-Kit (red). Original magnification is 40x. (<b>E</b>) All human IACs in the DA expressed CD34, and some expressed CD45. CD34 (green), CD45 (red) and TOTO-3 (blue). NT (Neural Tube); Ao (Aorta); Mn (Mesonephros). Original magnification is 20x.</p

Gene expression analysis in CD31⁺/CD34⁺/c-Kit⁺ AGM cells separated by CD45 expression.

<p>(<b>A</b>) Single cell suspensions of the caudal portion of embryos containing the AGM region at 10.5 dpc were prepared and analyzed by flow cytometry. Cells expressing CD31 and CD34, IAC markers, were first gated. The profile shows expression of c-Kit (x-axis) and CD45 (y-axis) in CD31⁺/CD34⁺ AGM cells (left). Based on intensity of CD45 expression, CD31⁺/CD34⁺/c-Kit⁺ AGM cells were separated into three fractions, CD45-negative (under 10² of CD45-fluorescence, same as negative control), -low positive (from 10².⁵ to 10³.⁵ of CD45-fluorescence), and -high positive (approximately over 10⁴ of CD45-fluorescence). Isotype control and compensation samples of flow cytometric analysis are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035763#pone.0035763.s004" target="_blank">Figure S4</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035763#pone.0035763.s001" target="_blank">S5</a>. (<b>B</b>) The percentage of CD45-negative, -low positive, and -high positive c-Kit⁺/CD31⁺/CD34⁺ AGM cells was calculated both at 9.5 dpc (white bars) and 10.5 dpc (black bars). (<b>C-H</b>) Gene expression of <i>CD45</i> (C), <i>Runx1</i> (D), <i>c-Myb</i> (E), <i>Evi-1</i> (F), <i>SCL</i> (G) and <i>Gata2</i> (H) was analyzed in sorted CD45-negative, -low positive and -high positive c-Kit⁺/CD31⁺/CD34⁺ AGM cells. Expression levels of <i>CD45</i> mRNA are up-regulated as c-Kit⁺/CD31⁺/CD34⁺ cells express CD45 surface protein. Expression levels of <i>Runx1</i>, <i>c-Myb</i>, <i>Evi-1</i>, <i>SCL</i> and <i>Gata2</i> were highest in CD45-low positive c-Kit⁺/CD31⁺/CD34⁺ cells, whereas that of <i>Evi-1</i> was highest in CD45-negative c-Kit⁺/CD31⁺/CD34⁺ cells. RQ represents relative quantity of template in the original sample.</p

Confocal images of IACs expressing CD31/CD34/c-Kit in the AGM region.

<p>Transverse sections of AGM region from ICR mouse embryos at 9.0 and 10.5 dpc were stained with antibodies and observed by confocal microscopy. (<b>A</b>) IACs were observed in the omphalomesenteric artery (OMA) at 9.0 dpc (left; magnified view of IACs in upper right panel) and in the OMA, dorsal aorta (DA) and umbilical artery (UA) at 10.5 dpc (right). CD31 (red), c-Kit (green), and TOTO-3 (blue). Arrows indicate IACs. Original magnification is 20x. (<b>B-D</b>) IACs were observed in the DA (B), OMA (C) and UA (D) at 10.5 dpc. Left panel shows staining for CD31 (red), c-Kit (green), and TOTO-3 (blue), and right panel shows staining for CD34 (red), c-Kit (green), and TOTO-3 (blue) staining. Images were taken at 40x and zoom was used to show a detail at right lower panel. Another IAC in the DA is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035763#pone.0035763.s001" target="_blank">Figure S1</a>. (<b>E</b>) IACs expressing Ki-67, a marker of proliferation, were observed in the DA (left), OMA (middle) and UA (right). Ki-67 (red), c-Kit (green), and TOTO-3 (blue). Images were taken at 40x and zoom was used to show a detail.</p