Isolation of Murine Bone Marrow Scavenging Sinusoidal Endothelial Cells

2014 ◽  
pp. 59-71 ◽  
Author(s):  
Peter A. G. McCourt ◽  
Ana Oteiza ◽  
Benjamin Cao ◽  
Susan K. Nilsson
Keyword(s):  
Bone Marrow ◽  
Endothelial Cells ◽  
Sinusoidal Endothelial Cells ◽  
Murine Bone Marrow

scholarly journals A VCAM-like adhesion molecule on murine bone marrow stromal cells mediates binding of lymphocyte precursors in culture.

1991 ◽  
Vol 114 (3) ◽  
pp. 557-565 ◽  
Author(s):  
K Miyake ◽  
K Medina ◽  
K Ishihara ◽  
M Kimoto ◽  
R Auerbach ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Endothelial Cells ◽  
Cell Adhesion ◽  
Endothelial Cell ◽  
Adhesion Molecule ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Cell Adhesion Molecule ◽  
B Lymphocyte ◽  
Murine Bone Marrow

Two new mAbs (M/K-1 and M/K-2) define an adhesion molecule expressed on stromal cell clones derived from murine bone marrow. The protein is similar in size to a human endothelial cell adhesion molecule known as VCAM-1 or INCAM110. VCAM-1 is expressed on endothelial cells in inflammatory sites and recognized by the integrin VLA-4 expressed on lymphocytes and monocytes. The new stromal cell molecule is a candidate ligand for the VLA-4 expressed on immature B lineage lymphocytes and a possible homologue of human VCAM-1. We now report additional similarities in the distribution, structure, and function of these proteins. The M/K antibodies detected large cells in normal bone marrow, as well as rare cells in other tissues. The antigen was constitutively expressed and functioned as a cell adhesion molecule on cultured murine endothelial cells. It correlated with the presence of mRNA which hybridized to a human VCAM-1 cDNA probe. Partial NH2 terminal amino acid sequencing of the murine protein revealed similarities to VCAM-1 and attachment of human lymphoma cells to murine endothelial cell lines was inhibited by the M/K antibodies. All of these observations suggest that the murine and human cell adhesion proteins may be related. The antibodies selectively interfered with B lymphocyte formation when included in long term bone marrow cultures. Moreover, they caused rapid detachment of lymphocytes from the adherent layer when added to preestablished cultures. The VCAM-like cell adhesion molecule on stromal cells and VLA-4 on lymphocyte precursors may both be important for B lymphocyte formation.

In Situ Visualization of Human MSC Engraftment in Bone Marrow: Integration into Murine Microenvironment and Interaction with Human HSC.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1283-1283
Author(s):  
Yukari Muguruma ◽  
Takashi Yahata ◽  
Hiroko Miyatake ◽  
Kiyoshi Ando ◽  
Tomomitsu Hotta
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Endothelial Cells ◽  
Stromal Cells ◽  
Fluorescent Protein ◽  
Hematopoietic Cells ◽  
Human Mscs ◽  
Murine Bone Marrow ◽  
Visual Evidence

Abstract Bone marrow is a complex organ system composed of two distinct lineages of cells: the hematopoietic cells and the supporting stromal cells, often referred as hematopoietic microenvironment (HME). Mesenchymal stem cells (MSCs) in bone marrow are shown to give rise to some of the components of HME, including osteoblasts, adipocytes and stromal fibroblasts in vitro, and to endothelial cells in vivo. It is a well accepted, but not definitely proven, concept that the HME provides structural niches, where dormant hematopoietic stem cells (HSCs) reside, and controls their renewal and differentiation. Although cotransplantation of human MSCs together with human HSCs resulted in increased chimerism of HSCs in animal models, existence of donor MSCs could only be detected using sensitive PCR-based analysis. Until this date, there is no physical evidence that transplanted MSCs have indeed engrafted in bone marrow and directly participated in that biological effect. In this study, we present the visual evidence for the sustained integration of human MSCs in murine bone marrow. Furthermore, we are able to delineate the physical interaction of injected human MSCs and cord blood derived CD34-positive HSCs (CBCD34). In order to assess the spatial distribution, lineage commitment and interaction of MSCs and HSCs in situ, we transplanted green fluorescent protein (GFP)-transduced MSCs and yellow fluorescent protein (YFP)-transduced CBCD34 into tibia of NOD/SCID mice. Ten weeks after intramedullary injection, longitudinal sections of mouse tibiae were made and stained with various antibodies for multicolor immunofluorescent analysis using a confocal microscope. We detected not only the existence of GFP-expressing MSCs in bone marrow, but also differentiation into several cell lineages. GFP-expressing cells exhibited phenotype and morphplogy of N-cadherin-positive bone lining osteoblasts, osteocalcin-positive osteocytes in bone, cells lining abluminal surface of vasculature, and in rare occasion, CD34 and CD31-positive endothelial cells. We then quantitatively evaluated the proportion of GFP-MSCs interacted with primitive YFP-CD34 and lineage committed YFP-CD15 and -Glycophorin-expressing cells as well as the proportion of above mentioned hematopoietic cells interacted with GFP-MSC. Approximately 50% of MSCs associated with CD34-posititive stem cells compared to only 2% and 3% of those with CD15 and Glycophorin-positive cells, respectively. It was also evident that the frequency of CD34-positive cells interacted with MSCs was significantly higher than those with CD15 and Glycophorin-positive cells. The results were consistent with a long appreciated notion that more primitive cells closely interact with hematopoietic supporting stromal cells. Furthermore, we quantitatively proved that the majority of YFP-CD34-positive HSCs were found close proximity to the bone. By transplanting GFP-MSCs together with YFP-HSCs, this study provided direct visual evidence that transplanted human MSCs engrafted in murine bone marrow and integrated into HME, which physically interacted with human HSC.

Bone Marrow Sinusoidal Endothelium Undergoes DNA Damage and Repair Following Lethal Irradiation.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4866-4866
Author(s):  
Xiao-Miao Li ◽  
Zhongbo Hu ◽  
Marda L. Jorgenson ◽  
John R. Wingard ◽  
William B. Slayton
Keyword(s):  
Bone Marrow ◽  
Dna Damage ◽  
Endothelial Cells ◽  
Bone Marrow Transplant ◽  
Caspase 3 ◽  
Marrow Transplant ◽  
Sinusoidal Endothelial Cells ◽  
Dna Damage And Repair ◽  
Post Irradiation ◽  
Lethal Irradiation

Abstract In the light of the possibility that adult bone marrow cells possess hemangioblast ability, work from our laboratory demonstrates that the bone marrow sinusoids remain predominantly host-derived following bone marrow transplant when ionizing irradiation is used as the conditioning regimen. To determine the effect of lethal irradiation to the host sinusoidal endothelial cells, we performed four apoptosis related assays and two cell proliferation assays on bone marrow sections at various time points during the first two weeks post-irradiation. We found: Phosphorylated H2AX was present in both hematopoietic and sinusoidal endothelial cells. However, only hematopoietic cells showed caspase-3 dependent apoptosis. Three days after radiation, some sinusoidal endothelial cells became TUNEL (Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay) positive, but were activated caspase-3 and ISOL (in situ oligo ligation assay) negative, suggesting non-apoptotic DNA fragmentation. TUNEL positive endothelial cells were present in non-transplanted irradiated bone marrow 7–13 days post-irradiation while after 7 days, there were almost no TUNEL positive endothelial cells in transplanted animal, demonstrating that donor cells support sinusoidal endothelial survival. In some endothelial cells, TUNEL signal was concentrated in discrete areas of the nucleus, suggesting a repair process that involves the localization and removal of damaged DNA fragments. Very few sinusoidal endothelial cells were Ki67 positive and even fewer were BrdU positive, demonstrating that endothelial cell division is not a major mechanism for the survival of bone marrow sinusoidal system after irradiation on the short term. These results demonstrate that sinusoidal endothelial cells undergo DNA damage and repair after lethal irradiation for bone marrow transplant. These results may explain, in part, why patients with impaired DNA damage/repair mechanisms have engraftment defects.

scholarly journals Uptake of chemically modified low density lipoproteins in vivo is mediated by specific endothelial cells.

1985 ◽  
Vol 100 (1) ◽  
pp. 103-117 ◽  
Author(s):  
R E Pitas ◽  
J Boyles ◽  
R W Mahley ◽  
D M Bissell
Keyword(s):  
Bone Marrow ◽  
Endothelial Cells ◽  
Kupffer Cells ◽  
Low Density Lipoproteins ◽  
Low Density ◽  
Coated Pits ◽  
Sinusoidal Endothelial Cells ◽  
Modified Lipoproteins

Acetoacetylated (AcAc) and acetylated (Ac) low density lipoproteins (LDL) are rapidly cleared from the plasma (t1/2 approximately equal to 1 min). Because macrophages, Kupffer cells, and to a lesser extent, endothelial cells metabolize these modified lipoproteins in vitro, it was of interest to determine whether endothelial cells or macrophages could be responsible for the in vivo uptake of these lipoproteins. As previously reported, the liver is the predominant site of the uptake of AcAc LDL; however, we have found that the spleen, bone marrow, adrenal, and ovary also participate in this rapid clearance. A histological examination of tissue sections, undertaken after the administration of AcAc LDL or Ac LDL (labeled with either 125I or a fluorescent probe) to rats, dogs, or guinea pigs, was used to identify the specific cells binding and internalizing these lipoproteins in vivo. With both techniques, the sinusoidal endothelial cells of the liver, spleen, bone marrow, and adrenal were labeled. Less labeling was noted in the ovarian endothelia. Uptake of AcAc LDL by endothelial cells of the liver, spleen, and bone marrow was confirmed by transmission electron microscopy. These data suggest uptake through coated pits. Uptake of AcAc LDL was not observed in the endothelia of arteries (including the coronaries and aorta), veins, or capillaries of the heart, testes, kidney, brain, adipose tissue, and duodenum. Kupffer cells accounted for a maximum of 14% of the 125I-labeled AcAc LDL taken up by the liver. Isolated sinusoidal endothelial cells from the rat liver displayed saturable, high affinity binding of AcAc LDL (Kd = 2.5 X 10(-9) M at 4 degrees C), and were shown to degrade AcAc LDL 10 times more effectively than aortic endothelial cells. These data indicate that specific sinusoidal endothelial cells, not the macrophages of the reticuloendothelial system, are primarily responsible for the removal of these modified lipoproteins from the circulation in vivo.

scholarly journals Stabilins are expressed in bone marrow sinusoidal endothelial cells and mediate scavenging and cell adhesive functions

2009 ◽  
Vol 390 (3) ◽  
pp. 883-886 ◽  
Author(s):  
Hong Qian ◽  
Sophie Johansson ◽  
Peter McCourt ◽  
Bård Smedsrød ◽  
Marja Ekblom ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Endothelial Cells ◽  
Sinusoidal Endothelial Cells ◽  
Cell Adhesive

Stabilin-1 and Stabilin-2 Are Expressed in Bone Marrow Sinusoidal Endothelial Cells and Mediate Scavenging and Cell Adhesive Functions

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1368-1368
Author(s):  
Hong Qian ◽  
Sophie Johansson ◽  
Peter McCourt ◽  
Bård Smedsrød ◽  
Marja Ekblom ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Endothelial Cells ◽  
Cell Trafficking ◽  
Tissue Remodelling ◽  
Sinusoidal Endothelial Cells ◽  
Recognition Mechanism ◽  
Hek 293 ◽  
Human Embryonic Kidney Cells ◽  
Transfected Cells ◽  
Specific Recognition

Abstract Stabilin-1 and stabilin-2 have been identified as scavenging receptors in sinusoidal capillaries in some tissues, including liver and spleen. They function as endocytic receptors for SPARC and hyaluronan, respectively, and for several other ligands. In the present study, we show by real-time PCR, Western blot and immunohistochemistry that bone marrow (BM) sinusoidal endothelial cells (SECs) ubiquitously express stabilin-1 and stabilin-2. To test the ability of BM SECs to function as a scavenging endothelium, we analyzed the uptake of specific scavenger receptor ligands after intravenous injection. Accumulation of TRITC labelled formaldehyde-treated serum albumin (FSA) was observed one hour after the injection in the BM SECs. Injection of unlabelled FSA together with TRITC-FSA reduced the fluorescence in the SECs, indicating that the uptake was due to specific recognition of FSA. Likewise, FITC-conjugated advanced glycation end products (AGEs)-modified BSA accumulated in BM SECs. These two ligands are primarily cleared by the stabilins. These results suggested that bone marrow SECs have a scavenging function. Stabilin-2 has been identified as a major receptor for hyaluronan. Hyaluronan is synthesized by primitive hematopoietic cells and has been shown to influence stem and progenitor (HSPC) functions, including mobilization and homing into BM (Nilsson et al., Blood.2003;101:856). Consequently, it is possible that adhesion of hyaluronan on HSPCs to stabilin-2 is a recognition mechanism in BM SECs, directing circulating HSPCs into BM. To investigate this, we studied adhesion of mouse BM lin-Sca-1+Kit+ (LSK) HSPCs to stabilin-1 or stabilin-2 transfected human embryonic kidney cells (HEK 293 cells). We found increased adhesion of LSK cells to stabilin-2 expressing cells, as compared to stabilin-1 expressing or non-transfected cells. Notably, hyaluronidase treatment abolished the increased adhesion of the LSK cell to stabilin-2 transfected cells. These findings indicate that stabilin-2 mediates adhesion of HSPCs to bone marrow SECs. In conclusion, this study shows a novel function for BM SECs as a scavenging endothelium expressing stabilin-1 and stabilin-2. The specific function of stabilins in recognition and endocytosis of extracellular matrix molecules, including hyaluronan and SPARC (Kzhyshkowska J et al., J Cell Mol Med.2006;10:635), suggests that these receptors are involved in tissue remodelling and cell trafficking in BM. Importantly, hyaluronan-mediated binding of HSPCs to SEC stabilin-2 may be a specific recognition mechanism for HSPCs in BM sinusoids. Similarly, binding of tumour cell-associated hyaluronan by stabilin-2 might cause tumour cells in blood to halt in bone marrow, thereby increasing the likelihood for metastasis at this site.

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