Mechanical properties of rat bone marrow and circulating neutrophils and their responses to inflammatory mediators

Blood ◽  
2002 ◽  
Vol 99 (6) ◽  
pp. 2207-2213 ◽  
Author(s):  
Hajime Saito ◽  
Jean Lai ◽  
Rick Rogers ◽  
Claire M. Doerschuk
Keyword(s):  
Mechanical Properties ◽  
Bone Marrow ◽  
Plasma Membrane ◽  
Complement Protein ◽  
Magnetic Twisting Cytometry ◽  
Pulmonary Capillaries ◽  
Shape Changes ◽  
Rat Bone

Abstract Neutrophils are continuously released from the bone marrow (BM), and this release is accelerated during inflammation. This study compared the mechanical properties of mature neutrophils within the BM and the circulating blood, as well as the role of microtubule rearrangement in the release of neutrophils from the BM in rats. Neutrophils isolated from the BM were stiffer than neutrophils in the circulating blood, using magnetic twisting cytometry. BM neutrophils also contained more F-actin within the submembrane region than circulating neutrophils when examined using confocal microscopy, suggesting that mature quiescent neutrophils within the BM are stiffer than circulating neutrophils because of increased formation of F-actin beneath the plasma membrane. Complement protein 5 fragments or formylmethionyl-leucylphenylalanine (fMLP) induced a stiffening response within 2 minutes that was greater in circulating than in BM neutrophils. This stiffening required F-actin formation within the submembrane region but not microtubule rearrangement in both circulating and BM neutrophils. fMLP-induced shape changes were more pronounced in circulating than in BM neutrophils, which showed fewer and smaller pseudopods and fewer membrane irregularities. In vivo, fMLP induced neutropenia, sequestration of neutrophils within the pulmonary capillaries, and release of neutrophils from the BM. Studies using colchicine demonstrated that rearrangement of microtubules was not required for any of these processes but was required for normal trafficking of neutrophils through the pulmonary capillaries.

scholarly journals Metabolism of rat bone proteoglycans in vivo

1983 ◽  
Vol 216 (3) ◽  
pp. 589-596 ◽  
Author(s):  
C W Prince ◽  
F Rahemtulla ◽  
W T Butler
Keyword(s):  
Extraction Procedure ◽  
Total Radioactivity ◽  
Mineralized Tissue ◽  
Guanidinium Chloride ◽  
Disaccharide Composition ◽  
A Minor ◽  
Papain Digestion ◽  
Rat Bone

Former evaluations of the role of proteoglycans in mineralization have neglected to address the possibility that the metabolism of proteoglycans may be of significance in this regard. This problem was studied by using radiolabeling in vivo of rat calvaria with [35Sulphate for 2-72 h and a sequential extraction procedure to yield two pools of newly synthesized proteoglycans: one obtained from non-mineralized tissue by extraction with guanidinium chloride (GdmCl) and another obtained only after demineralization with EDTA. Total radioactivity in calvaria was maximal after 12 h of incorporation, but by 36 h had declined to a level that was about 55-65% of maximum. Radioactivity in the GdmCl extract declined steadily after 12 h, whereas that in the EDTA extract remained constant until 36 h, when it began to increase. Each extract contained a minor proteoglycan that eluted at the void volume (Vo) of a Sepharose CL-6B column. Unlike in the EDTA extract, this proteoglycan gradually disappeared from the GdmCl extract. Each extract also contained a major, smaller proteoglycan, with a Kav. of 0.24 and 0.36 in the GdmCl and EDTA extracts respectively. Papain digestion of each extract yielded glycosaminoglycan chains with Kav. values of 0.32 and 0.50 on CL-6B in the GdmCl and EDTA extracts respectively. Digestion of each extract with chondroitinase ABC and chondroitinase AC showed that the glycosaminoglycans were of similar disaccharide composition, with about 85% being 4-sulphated and the remainder 6-sulphated and/or iduronic acid-containing. These data suggest that about 45% of the newly synthesized proteoglycans are removed from the tissue during the course of mineralization.

scholarly journals Role of the Endocytic Machinery in the Sorting of Lysosome-associated Membrane Proteins

2005 ◽  
Vol 16 (9) ◽  
pp. 4231-4242 ◽  
Author(s):  
Katy Janvier ◽  
Juan S. Bonifacino
Keyword(s):  
Plasma Membrane ◽  
Coat Protein ◽  
Membrane Proteins ◽  
Adaptor Protein ◽  
The Other ◽  
Sorting Signals ◽  
Efficient Delivery ◽  
Endocytic Machinery

The limiting membrane of the lysosome contains a group of transmembrane glycoproteins named lysosome-associated membrane proteins (Lamps). These proteins are targeted to lysosomes by virtue of tyrosine-based sorting signals in their cytosolic tails. Four adaptor protein (AP) complexes, AP-1, AP-2, AP-3, and AP-4, interact with such signals and are therefore candidates for mediating sorting of the Lamps to lysosomes. However, the role of these complexes and of the coat protein, clathrin, in sorting of the Lamps in vivo has either not been addressed or remains controversial. We have used RNA interference to show that AP-2 and clathrin—and to a lesser extent the other AP complexes—are required for efficient delivery of the Lamps to lysosomes. Because AP-2 is exclusively associated with plasma membrane clathrin coats, our observations imply that a significant population of Lamps traffic via the plasma membrane en route to lysosomes.

scholarly journals Purinergic Modulation of Interleukin-1β Release from Microglial Cells Stimulated with Bacterial Endotoxin

1997 ◽  
Vol 185 (3) ◽  
pp. 579-582 ◽  
Author(s):  
Davide Ferrari ◽  
Paola Chiozzi ◽  
Simonetta Falzoni ◽  
Stefania Hanau ◽  
Francesco Di  Virgilio
Keyword(s):  
Plasma Membrane ◽  
P2x7 Receptor ◽  
Purinergic Receptor ◽  
Present Report ◽  
Physiological Role ◽  
Bacterial Endotoxin ◽  
Microglial Cells

Microglial cells express a peculiar plasma membrane receptor for extracellular ATP, named P2Z/P2X7 purinergic receptor, that triggers massive transmembrane ion fluxes and a reversible permeabilization of the plasma membrane to hydrophylic molecules of up to 900 dalton molecule weight and eventual cell death (Di Virgilio, F. 1995. Immunol. Today. 16:524–528). The physiological role of this newly cloned (Surprenant, A., F. Rassendren, E. Kawashima, R.A. North and G. Buell. 1996. Science (Wash. DC). 272:735–737) cytolytic receptor is unknown. In vitro and in vivo activation of the macrophage and microglial cell P2Z/P2X7 receptor by exogenous ATP causes a large and rapid release of mature IL-1β. In the present report we investigated the role of microglial P2Z/P2X7 receptor in IL-1β release triggered by LPS. Our data suggest that LPS-dependent IL-1β release involves activation of this purinergic receptor as it is inhibited by the selective P2Z/P2X7 blocker oxidized ATP and modulated by ATP-hydrolyzing enzymes such as apyrase or hexokinase. Furthermore, microglial cells release ATP when stimulated with LPS. LPS-dependent release of ATP is also observed in monocyte-derived human macrophages. It is suggested that bacterial endotoxin activates an autocrine/paracrine loop that drives ATP-dependent IL-1β secretion.

Abstract 11664: The Identification and Hierarchy of Bone Marrow-Derived Artery-Resident Mesodermal Progenitor Cells and Their Dynamics in Atherosclerosis

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Hyun-Jai Cho ◽  
Hyun-Ju Cho ◽  
Yoo-Wook Kwon ◽  
Young-Bae Park ◽  
Hyo-Soo Kim
Keyword(s):  
Bone Marrow ◽  
Progenitor Cells ◽  
Peripheral Circulation ◽  
Humoral Factors ◽  
Cell Property ◽  
Apoe Ko Mice ◽  
Multiplex Cytokine ◽  
Developmental Hierarchy

Background: We recently identified bone marrow (BM)-derived artery resident calcifying progenitor cells. Sca-1+PDGFRα- cells may possess bipotent (osteoblastic/osteoclastic) characteristics. However, the nature of progenitor cells remains elusive. Hypothesis: We investigated developmental hierarchy of progenitor cells and in vivo dynamics in atherosclerosis. Methods and Results: We harvested cells from BM and artery of C57 mice. In BM, Lin-CD29+Sca-1+PDGFRα- cells showed hematopoietic potential and differentiated into osteoclasts (OC). They also possessed mesenchymal stem cell property including osteoblastic (OB) differentiation, suggesting that Sca-1+PDGFRα- cells could be mesodermal progenitor cells. Interestingly, BM-derived artery-resident, clonal Sca-1+PDGFRα- cells maintained bipotency but lost hematopoietic nature. In contrast, Sca-1+PDGFRα+ cells in BM and artery only showed unipotency (OB). When we overexpressed or knocked down PDGFRα, there was no alteration in OB or OC differentiation of Sca-1+PDGFRα- cells and no effect on OB differentiation of Sca-1+PDGFRα+ cells, indicating PDGFRα as a surface marker but not a functional player. In hyperlipidemic ApoE-KO mice compared with control, Sca-1+PDGFRα- cells were less mobilized from BM to peripheral circulation and less infiltrated into atherosclerotic plaque, whereas Sca-1+PDGFRα+ cells were not significantly affected. Multiplex cytokine assay of serum and artery revealed that IL-1β was significantly increased and IL-5 was markedly decreased in atherosclerotic mice. IL-1β decreased the migration of Sca-1+PDGFRα- cells by 5 folds compared with TNFα, and IL-5 increased the migration as much as TNFα. But the migration of Sca-1+PDGFRα+ cells was not altered. These data indicate that atherosclerosis-related humoral factors mainly regulated mesodermal progenitor cells’ dynamics. Conclusion: We demonstrate that Sca-1+PDGFRα- cell is a mesodermal progenitor cell that possesses both hematopoietic and mesenchymal potentials. In atherogenesis, the mobilization and infiltration of Sca-1+PDGFRα- progenitor cells were regulated by IL-1β and IL-5. These data provide a novel mechanism regarding the role of bipotent progenitor cells in atherosclerosis.

Effects of dietary alteration of bicarbonate and magnesium on rat bone

1986 ◽  
Vol 250 (2) ◽  
pp. F302-F307 ◽  
Author(s):  
J. M. Burnell ◽  
C. Liu ◽  
A. G. Miller ◽  
E. Teubner
Keyword(s):  
Crystal Structure ◽  
Bone Formation ◽  
X Ray Diffraction ◽  
X Ray ◽  
Growing Rats ◽  
Final Composition ◽  
Rat Bone

To study the effects of bicarbonate and magnesium on bone, mild acidosis and/or hypermagnesemia were produced in growing rats by feeding ammonium chloride and/or magnesium sulfate. Bone composition, quantitative histomorphometry, and mineral x-ray diffraction (XRD) characteristics were measured after 6 wk of treatment. The results demonstrated that both acidosis (decreased HCO3) and hypermagnesemia inhibited periosteal bone formation, and, when combined, results were summative; and the previously observed in vitro role of HCO3- and Mg2+ as inhibitors of crystal growth were confirmed in vivo. XRD measurements demonstrated that decreased plasma HCO3 resulted in larger crystals and increased Mg resulted in smaller crystals. However, the combined XRD effects of acidosis and hypermagnesemia resembled acidosis alone. It is postulated that the final composition and crystal structure of bone are strongly influenced by HCO3- and Mg2+, and the effects are mediated by the combined influence on both osteoblastic bone formation and the growth of hydroxyapatite.

scholarly journals Effects of Nrf2 Deficiency on Bone Microarchitecture in an Experimental Model of Osteoporosis

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Lidia Ibáñez ◽  
María Luisa Ferrándiz ◽  
Rita Brines ◽  
David Guede ◽  
Antonio Cuadrado ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Bone Marrow ◽  
Bone Metabolism ◽  
Bone Microarchitecture ◽  
Related Factor ◽  
Mineral Density ◽  
Trabecular Bone Mineral Density ◽  
Bone Marrow Derived Cells

Objective. Redox imbalance contributes to bone fragility. We have evaluated the in vivo role of nuclear factor erythroid derived 2-related factor-2 (Nrf2), an important regulator of cellular responses to oxidative stress, in bone metabolism using a model of postmenopausal osteoporosis.Methods. Ovariectomy was performed in both wild-type and mice deficient in Nrf2 (Nrf2−/−). Bone microarchitecture was analyzed byμCT. Serum markers of bone metabolism were also measured. Reactive oxygen species production was determined using dihydrorhodamine 123.Results. Sham-operated or ovariectomized Nrf2−/−mice exhibit a loss in trabecular bone mineral density in femur, accompanied by a reduction in cortical area in vertebrae. Nrf2 deficiency tended to increase osteoblastic markers and significantly enhanced osteoclastic markers in sham-operated animals indicating an increased bone turnover with a main effect on bone resorption. We have also shown an increased production of oxidative stress in bone marrow-derived cells from sham-operated or ovariectomized Nrf2−/−mice and a higher responsiveness of bone marrow-derived cells to osteoclastogenic stimuli in vitro.Conclusion. We have demonstrated in vivo a key role of Nrf2 in the maintenance of bone microarchitecture.

scholarly journals Encapsulation of Rat Bone Marrow Derived Mesenchymal Stem Cells in Alginate Dialdehyde/Gelatin Microbeads with and without Nanoscaled Bioactive Glass for In Vivo Bone Tissue Engineering

Materials ◽  
2018 ◽  
Vol 11 (10) ◽  
pp. 1880 ◽  
Author(s):  
Ulrike Rottensteiner-Brandl ◽  
Rainer Detsch ◽  
Bapi Sarker ◽  
Lara Lingens ◽  
Katrin Köhn ◽  
...  
Keyword(s):  
Tissue Engineering ◽  
Bone Marrow ◽  
Cell Survival ◽  
Bone Tissue Engineering ◽  
Bone Tissue ◽  
Immune Reaction ◽  
Three Dimensional ◽  
High Cell ◽  
Rat Bone

Alginate dialdehyde (ADA), gelatin, and nano-scaled bioactive glass (nBG) particles are being currently investigated for their potential use as three-dimensional scaffolding materials for bone tissue engineering. ADA and gelatin provide a three-dimensional scaffold with properties supporting cell adhesion and proliferation. Combined with nanocristalline BG, this composition closely mimics the mineral phase of bone. In the present study, rat bone marrow derived mesenchymal stem cells (MSCs), commonly used as an osteogenic cell source, were evaluated after encapsulation into ADA-gelatin hydrogel with and without nBG. High cell survival was found in vitro for up to 28 days with or without addition of nBG assessed by calcein staining, proving the cell-friendly encapsulation process. After subcutaneous implantation into rats, survival was assessed by DAPI/TUNEL fluorescence staining. Hematoxylin-eosin staining and immunohistochemical staining for the macrophage marker ED1 (CD68) and the endothelial cell marker lectin were used to evaluate immune reaction and vascularization. After in vivo implantation, high cell survival was found after 1 week, with a notable decrease after 4 weeks. Immune reaction was very mild, proving the biocompatibility of the material. Angiogenesis in implanted constructs was significantly improved by cell encapsulation, compared to cell-free beads, as the implanted MSCs were able to attract endothelial cells. Constructs with nBG showed higher numbers of vital MSCs and lectin positive endothelial cells, thus showing a higher degree of angiogenesis, although this difference was not significant. These results support the use of ADA/gelatin/nBG as a scaffold and of MSCs as a source of osteogenic cells for bone tissue engineering. Future studies should however improve long term cell survival and focus on differentiation potential of encapsulated cells in vivo.

scholarly journals Stimulation of interleukin-6 production by endothelin in rat bone marrow-derived stromal cells

Blood ◽  
1994 ◽  
Vol 84 (8) ◽  
pp. 2531-2538 ◽  
Author(s):  
T Agui ◽  
X Xin ◽  
Y Cai ◽  
T Sakai ◽  
K Matsumoto
Keyword(s):  
Bone Marrow ◽  
Stromal Cells ◽  
Interleukin 6 ◽  
Hormonal Regulation ◽  
Ex Vivo ◽  
Second Messengers ◽  
Immunodeficiency Virus ◽  
Rat Bone ◽  
Stimulation Of

Abstract Endothelin (ET) produced by endothelial cells has recently been found to be a potent vasoconstricting hormone. In this report, ET is shown to be a potent stimulator of interleukin-6 (IL-6) production by rat bone marrow (BM)-derived stromal cells. It was also shown that ET increased the level of mRNA for IL-6 in these cells. The two types of ET receptor (R), ETAR and ETBR, were shown to be expressed on both BM-derived stromal cells in culture and ex vivo in BM tissue, suggesting that ET works as a physiologic stimulator of IL-6 production in the BM. It was shown that ETAR is coupled to phospholipase C activation, leading to the production of inositol 1,4,5-trisphosphate (IP3) and 1,2- diacylglycerol (DAG) as second messengers in BM-derived stromal cells. This was corroborated by data showing that IL-6 production in these cells was induced by combined stimulation with ionomycin and phorbol myristate acetate, thereby bypassing the effects of IP3 and DAG, respectively. This is the first report on the hormonal regulation of IL- 6 production by BM stromal cells, indicating that hematopoiesis is subject to endocrinologic regulation under physiologic conditions. ET has recently been reported to be produced by macrophages in response to bacterial lipopolysaccharide and human immunodeficiency virus-1 glycoprotein 120. These facts, taken together with our findings, raise the possibility that ET shares the same role of IL-1 as a local cytokine, mediating an intercellular signal between macrophages and BM stromal cells in response to bacterial or viral stimulation.

Successful in vivo purging of CD34-containing peripheral blood harvests in mantle cell and indolent lymphoma: evidence for a role of both chemotherapy and rituximab infusion

Blood ◽  
2000 ◽  
Vol 96 (3) ◽  
pp. 864-869 ◽  
Author(s):  
Michele Magni ◽  
Massimo Di Nicola ◽  
Liliana Devizzi ◽  
Paola Matteucci ◽  
Fabrizio Lombardi ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Follicular Lymphoma ◽  
Mantle Cell Lymphoma ◽  
Cell Lymphoma ◽  
Bone Marrow Involvement ◽  
Hematopoietic Stem ◽  
Mantle Cell

Abstract Elimination of tumor cells (“purging”) from hematopoietic stem cell products is a major goal of bone marrow–supported high-dose cancer chemotherapy. We developed an in vivo purging method capable of providing tumor-free stem cell products from most patients with mantle cell or follicular lymphoma and bone marrow involvement. In a prospective study, 15 patients with CD20+ mantle cell or follicular lymphoma, bone marrow involvement, and polymerase chain reaction (PCR)–detectable molecular rearrangement received 2 cycles of intensive chemotherapy, each of which was followed by infusion of a growth factor and 2 doses of the anti-CD20 monoclonal antibody rituximab. The role of rituximab was established by comparison with 10 control patients prospectively treated with an identical chemotherapy regimen but no rituximab. The CD34+ cells harvested from the patients who received both chemotherapy and rituximab were PCR-negative in 93% of cases (versus 40% of controls;P = .007). Aside from providing PCR-negative harvests, the chemoimmunotherapy treatment produced complete clinical and molecular remission in all 14 evaluable patients, including all 6 with mantle cell lymphoma (versus 70% of controls). In vivo purging of hematopoietic progenitor cells can be successfully accomplished in most patients with CD20+ lymphoma, including mantle cell lymphoma. The results depended on the activity of both chemotherapy and rituximab infusion and provide the proof of principle that in vivo purging is feasible and possibly superior to currently available ex vivo techniques. The high short-term complete-response rate observed suggests the presence of a more-than-additive antilymphoma effect of the chemoimmunotherapy combination used.

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