scholarly journals Systemic prostaglandin E2 increases cancellous bone formation and mass in aging rats and stimulates their bone marrow osteogenic capacity in vivo and in vitro

2001 ◽  
Vol 168 (1) ◽  
pp. 131-139 ◽  
Author(s):  
S Keila ◽  
A Kelner ◽  
M Weinreb
Keyword(s):  
Bone Marrow ◽  
Alkaline Phosphatase ◽  
Bone Formation ◽  
Cancellous Bone ◽  
Nodule Formation ◽  
Anabolic Effect ◽  
Aging Rats ◽  
Old Rats

Prostaglandin E(2) (PGE(2)) has been shown to exert a bone anabolic effect in young and adult rats. In this study we tested whether it possesses a similar effect on bone formation and bone mass in aging rats. Fifteen-month-old rats were injected daily with either PGE(2) at 5 mg/kg or vehicle for 14 days. PGE(2) treatment stimulated the rate of cancellous bone formation (a approximately 5.5-fold increase in bone formation rate), measured by the incorporation of calcein into bone-forming surfaces at the tibial proximal metaphysis. This effect resulted in increased cancellous bone area (+54%) at the same site. Since PGE(2) treatment resulted in a much higher proportion of bone surface undergoing bone formation and thus lined with osteoblasts, we tested the hypothesis that PGE(2) stimulates osteoblast differentiation from bone marrow precursor cells both in vivo and in vitro. We found that ex vivo cultures of bone marrow stromal cells from rats injected for 2 weeks with PGE(2) at 5 mg/kg per day yielded more ( approximately 4-fold) mineralized nodules and exhibited a greater (by 30-40%) alkaline phosphatase activity compared with cultures from vehicle-injected rats, attesting to a stimulation of osteoblastic differentiation by PGE(2). We also compared the osteogenic capacity of bone marrow from aging (15-month-old) versus young (5-week-old) rats and its regulation by PGE(2) in vitro. Bone marrow stromal cell cultures from aging rats exhibited a greatly diminished osteogenic capacity, reflected in reduced nodule formation ( approximately 6% of young animals) and lower alkaline phosphatase activity ( approximately 60% of young animals). However, these parameters could be stimulated in both groups of animals by incubation with 10-100 nM PGE(2). The magnitude of this stimulation was greater in cultures from aging rats (+550% vs +70% in nodule formation of aging compared with young rats). In conclusion, we demonstrate here that PGE(2) exerts a bone anabolic effect in aging rats, similar to the effect we and others have reported in young, growing rats. The PGE(2)-stimulated bone formation, which augments bone mass, most likely results from recruitment of osteoblasts from their bone marrow stromal precursors.

scholarly journals Inhibition of cyclo-oxigenase-2 activity does not abolish the anabolic effect of prostaglandin E2 in vivo or in vitro

2002 ◽  
Vol 174 (1) ◽  
pp. 127-135 ◽  
Author(s):  
M Weinreb ◽  
A Kelner ◽  
S Keila
Keyword(s):  
Bone Formation ◽  
Bone Tissue ◽  
Receptor Expression ◽  
Male Rats ◽  
Nodule Formation ◽  
High Dose ◽  
Anabolic Effect ◽  
Cox 2

It was previously reported that the expression of cyclo-oxigenase-2 (COX-2) is induced by prostaglandin E(2) (PGE(2)) in vitro in an osteogenic cell line and organ culture, suggesting an autoamplification mechanism. In this study, we first tested whether this phenomenon also occurs in bone tissue in vivo and found that a single anabolic dose of PGE(2) (5 mg/kg) induced (between 30 and 120 min) in rat tibiae, an increase in the mRNA level of COX-2 (2.5- to 9-fold) but not that of COX-1. Secondly, to test whether COX-2 activity in generating endogenous prostaglandins (PGs) is required for the in vivo anabolic properties of PGE(2), young male rats were injected daily with either vehicle (8% ethanol) or 5 mg/kg PGE(2) for 21 days. PGE(2)-injected rats received, 45 min prior to PGE(2), either dimethyl sulphoxide (as vehicle) or one of two doses of NS-398, a selective COX-2 inhibitor: a low dose (3 mg/kg) or a high dose (10 mg/kg). PGE(2) increased bone formation (measured as cancellous mineralizing surface, mineral apposition rate and bone formation rate) and bone mass (measured as cancellous bone area and surface and cortical width). None of these increases was suppressed by pre-administration of NS-398. In contrast, the high dose of NS-398 effectively suppressed an increase in rat hind-paw volume induced by a local carrageenan injection. Furthermore, since COX-2 inactivation may affect PG receptor expression, we found that pre-administration of NS-398 did not abolish the induction in EP(4) receptor mRNA levels, caused by PGE(2) in rat bone tissue. For in vitro testing, rat femoral bone marrow stromal cell cultures were initiated and were incubated in the absence or presence of PGE(2) at 100 nM (as an inducer) and with increasing concentrations of NS-398 (10(-8) M to 10(-5) M) for 21 days, after which time mineralized (Von-Kossa positive) nodules were counted. PGE(2) increased nodule formation as previously reported; however, NS-398 reduced nodule formation in both control and PGE(2)-treated cultures to the same extent. We conclude that while the level of COX-2 mRNA is increased in vivo by administration of PGE(2), inhibition of its activity (i.e. generation of endogenous PGs) does not abolish the anabolic effect of PGE(2).

scholarly journals Ibudilast Mitigates Delayed Bone Healing Caused by Lipopolysaccharide by Altering Osteoblast and Osteoclast Activity

2021 ◽  
Vol 22 (3) ◽  
pp. 1169
Author(s):  
Yuhan Chang ◽  
Chih-Chien Hu ◽  
Ying-Yu Wu ◽  
Steve W. N. Ueng ◽  
Chih-Hsiang Chang ◽  
...  
Keyword(s):  
Cell Wall ◽  
Bone Formation ◽  
Cancellous Bone ◽  
Bone Healing ◽  
Bone Bridge ◽  
Cell Wall Components ◽  
Osteoclast Activity ◽  
Wall Components

Bacterial infection in orthopedic surgery is challenging because cell wall components released after bactericidal treatment can alter osteoblast and osteoclast activity and impair fracture stability. However, the precise effects and mechanisms whereby cell wall components impair bone healing are unclear. In this study, we characterized the effects of lipopolysaccharide (LPS) on bone healing and osteoclast and osteoblast activity in vitro and in vivo and evaluated the effects of ibudilast, an antagonist of toll-like receptor 4 (TLR4), on LPS-induced changes. In particular, micro-computed tomography was used to reconstruct femoral morphology and analyze callus bone content in a femoral defect mouse model. In the sham-treated group, significant bone bridge and cancellous bone formation were observed after surgery, however, LPS treatment delayed bone bridge and cancellous bone formation. LPS inhibited osteogenic factor-induced MC3T3-E1 cell differentiation, alkaline phosphatase (ALP) levels, calcium deposition, and osteopontin secretion and increased the activity of osteoclast-associated molecules, including cathepsin K and tartrate-resistant acid phosphatase in vitro. Finally, ibudilast blocked the LPS-induced inhibition of osteoblast activation and activation of osteoclast in vitro and attenuated LPS-induced delayed callus bone formation in vivo. Our results provide a basis for the development of a novel strategy for the treatment of bone infection.

4:51146. Nell-1 Induces Osteoblastic Differentiation of Bone Marrow Mesenchymal Cells In Vitro and Bone Formation In Vivo

2006 ◽  
Vol 6 (5) ◽  
pp. 74S
Author(s):  
Tara Aghaloo ◽  
Xinquan Jiang ◽  
Xinli Zhang ◽  
Zhang Zhiyuang ◽  
Jeffrey C. Wang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Formation ◽  
Osteoblastic Differentiation ◽  
Mesenchymal Cells ◽  
Bone Marrow Mesenchymal Cells

Prospective In Vivo Identification of Osteogenic Stem/Progenitor Cells from Bone Marrow-Derived Lin−/Sca-1+/CD45− Cells.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1409-1409
Author(s):  
Zhuo Wang ◽  
Junghun Jung ◽  
Magdalena Kucia ◽  
Junhui Song ◽  
Yusuke Shiozawa ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Bone Formation ◽  
Cultured Cells ◽  
Fluorescent Microscopy ◽  
Micro Ct ◽  
Mineralized Tissue

Abstract We previously developed an in vivo prospective assay for identification of non-cultured cells with MSC potential. Using this assay we identified a population of cells that were slowly cycling and of low density that were capable of multilineage differentiation both in vitro and in vivo (Z. Wang et al, Stem Cells. 2006 24(6):1573). Further characterization of these cells suggested that they resemble a homogenous population of rare Lin−/Sca-1+/CD45− cells that have the morphology and express several markers of undifferentiated embryonic-like stem cells. In vitro the Lin−/Sca-1+/CD45− cells may differentiate into cells from all three germ-layers (M. Kucia et al, Leukemia. 2007 21(2):297). To determine the in vivo fate of this population, we transplanted 500 or 5,000 Lin−/Sca-1+/CD45− cells from a GFP mouse into SCID mice in each group (n=3) immediately after cell sorting to evaluate tissue generation in vivo. At 4 weeks the regenerative potential of these populations was evaluated by micro-CT and histology, and cells were tracked by gross examination of the harvested tissues by fluorescent microscopy. The results showed that a large number of GFP+ cells are located in the implants, indicating that the transplanted cells maintain the ability to contribute to the generation of new tissue. Bone-like tissue was observed in the Lin−/Sca-1+/CD45− group with as low as 500-cells/implant, while 5,000 Lin−/Sca-1+/CD45− cells generated significantly larger mineralized tissue volume, which was confirmed by micro-CT. Lin−/Sca-1+/CD45+ cell only implantation did not form any mineralized tissue, however, while mixed with 2x106 whole bone morrow cells, positive mineralized tissue occurred. Whole bone marrow mixture also improve bone formation in Lin−/Sca-1+/CD45− cell implants compared the actual bone volumes measured by micro-CT. This study demonstrates that non-cultured BM-derived Lin−/Sca-1+/CD45− cells exhibit the capacity to form bone in vivo with as low as 500 cells/implant. Whole bone marrow mixtures can enhance the bone formation, presumably through the interaction of other populations cells. Based on these findings, it is proposed that non-cultured BM-derived Lin−/Sca-1+/CD45− cells are enriched osteogenic cells that can be applied to bone regeneration in vivo.

Effects of in vitro chondrogenic priming time of bone-marrow-derived mesenchymal stromal cells on in vivo endochondral bone formation

2015 ◽  
Vol 13 ◽  
pp. 254-265 ◽  
Author(s):  
Wanxun Yang ◽  
Sanne K. Both ◽  
Gerjo J.V.M. van Osch ◽  
Yining Wang ◽  
John A. Jansen ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Formation ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Endochondral Bone Formation ◽  
Endochondral Bone

STIMULATION OF BONE MARROW CELLS AND BONE FORMATION BY NACRE. IN VIVO AND IN VITRO STUDIES

Bioceramics ◽  
1999 ◽  
Author(s):  
M. Lamghari ◽  
S. Berland ◽  
A. Laurent ◽  
H. Huet ◽  
M.J. Almeida ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Formation ◽  
Bone Marrow Cells ◽  
In Vitro Studies ◽  
Stimulation Of ◽  
Marrow Cells

scholarly journals Characterization of a 5-fluorouracil-enriched osteoprogenitor population of the murine bone marrow

Blood ◽  
1993 ◽  
Vol 82 (12) ◽  
pp. 3580-3591
Author(s):  
N Falla ◽  
Vlasselaer Van ◽  
J Bierkens ◽  
B Borremans ◽  
G Schoeters ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Cells ◽  
Collagen Matrix ◽  
Collagen Type I ◽  
Nodule Formation ◽  
Mouse Bone Marrow ◽  
Type I ◽  
Murine Bone Marrow

In the presence of beta-glycerophosphate and vitamin C, cultures of normal mouse bone marrow cells form three-dimensional structures that stain positive with the Von Kossa technique and express alkaline phosphatase (ALP), collagen type I, and osteocalcin. Little is known about the characteristics and frequency of the cells that contribute to this phenomenon. Most likely, mature osteoblastic cells do not contribute to the nodule formation because no osteocalcin expressing cells are detected in the flushed marrow by in situ hybridization. Limiting dilution analysis shows that, in normal bone marrow, 1 of 2.2 x 10(5) cells has the potency to form a bone nodule and to express ALP, collagen, and osteocalcin in a temporal fashion. Upon in vivo treatment with 5-fluorouracil (5-FU), this frequency increases 12-fold, eg, 1 in 1.75 x 10(4) cells shows osteogenic activity. In comparison, fibroblast colony forming cells occur at a frequency of 1 of 2.5 x 10(4) or 1 of 5 x 10(3) plated cells in normal or 5-FU-treated marrow, respectively. Using density centrifugation, the majority of the osteoprogenitor cells in 5-FU marrow are found in the low-density (1.066 to 1.067 g/mL) fractions. In addition, these cells bind to nylon wool but not to plastic and aggregate in the presence of wheat germ agglutinin and soybean agglutinin. Scanning and transmission electron microscopy shows that the bone nodules in 5-FU marrow cultures are composed of fibroblastoid cells embedded in a mineralized collagen matrix. In conclusion, our results show that a quiescent cell population in the murine bone marrow with fibroblastoid characteristics contributes to the formation of bone-like nodules in vitro.

scholarly journals CD13 is a Critical Regulator of Cell-cell Fusion in Osteoclastogenesis

2020 ◽  
Author(s):  
Mallika Ghosh ◽  
Ivo Kalajzic ◽  
Hector Leonardo Aguila ◽  
Linda H Shapiro
Keyword(s):  
Bone Marrow ◽  
Bone Formation ◽  
Progenitor Cells ◽  
Cell Fusion ◽  
Formation Rate ◽  
Giant Cells ◽  
Bone Formation Rate ◽  
Cell Cell

AbstractIn vertebrates, bone formation is dynamically controlled by the activity of two specialized cell types: the bone-generating osteoblasts and bone-degrading osteoclasts. Osteoblasts produce the soluble receptor activator of NFκB ligand (RANKL) that binds to its receptor RANK on the surface of osteoclast precursor cells to promote osteoclastogenesis, a process that involves cell-cell fusion and assembly of molecular machinery to ultimately degrade the bone. CD13 is a transmembrane aminopeptidase that is highly expressed in cells of myeloid lineage has been shown to regulate dynamin-dependent receptor endocytosis and recycling and is a necessary component of actin cytoskeletal organization. In the present study, we show that CD13-deficient mice display a normal distribution of osteoclast progenitor populations in the bone marrow, but present a low bone density phenotype. Further, the endosteal bone formation rate is similar between genotypes, indicating a defect in osteoclast-specific function in vivo. Loss of CD13 led to exaggerated in vitro osteoclastogenesis as indicated by significantly enhanced fusion of bone marrow-derived multinucleated osteoclasts in the presence of M-CSF and RANKL, resulting in abnormally large cells with remarkably high numbers of nuclei with a concomitant increase in bone resorption activity. Similarly, we also observed increased formation of multinucleated giant cells (MGC) in CD13KO bone marrow progenitor cells stimulated with IL-4 and IL-13, suggesting that CD13 may regulate cell-cell fusion events via a common pathway, independent of RANKL signaling. Mechanistically, while expression levels of the fusion-regulatory proteins dynamin and DC-STAMP are normally downregulated as fusion progresses in fusion-competent mononucleated progenitor cells, in the absence of CD13 they are uniformly sustained at high levels, even in mature multi-nucleated osteoclasts. Taken together, we conclude that CD13 may regulate cell-cell fusion by controlling expression and localization of key fusion proteins that are critical for both osteoclast and MGC fusion.

Platelet-Derived Factors Allow Human Mesenchymal Stem Cells to Spontaneously Undergo Endochondral Bone Differentiation and Provide Bone Marrow Support in a Xenogenic In Vivo Model

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1322-1322
Author(s):  
Nathalie Etchart ◽  
Andreas Reinisch ◽  
Anna Ortner ◽  
Margareta Frühwirth ◽  
Rokhsareh Rohban ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Formation ◽  
Human Platelet ◽  
Human Bone ◽  
Mouse Bone Marrow ◽  
Bone Differentiation ◽  
Nsg Mice ◽  
Ectopic Bone

Abstract Abstract 1322 Introduction. Mesenchymal Stem/Progenitor Cells (MSPC) are regarded as the universal skeletal progenitor, theoretically capable of differentiating into cartilage, bone, tendon and muscle. Their functions as pericytes as well as key bone marrow stromal cells are also well documented. Tremendous benefits could therefore be achieved by cell therapy with MSPC. Unfortunately, the very successful research aimed at isolating, expanding and differentiating these cells in vitro has so far failed to translate into significant clinical advances. In particular, bone regeneration studies are disappointing, because application of MSPC in vivo requires their osteogenic pre-differentiation in vitro and/or their co-implantation with bone chips, and because results are highly donor-dependant. In this context, recent studies indicate that MSPC expanded from human Bone Marrow (BM) in media supplemented by Human Platelet Lysate (HPL) in lieu of Fetal Bovine Serum (FBS) have a higher rate of bone differentiation. Here, we use a new, simplified mouse model of ectopic bone formation requiring neither pre-differentiation nor bone matrix. We show that human BM-MSPC expanded in HPL-supplemented medium from all donors spontaneously form bone through an endochondral mechanism. Importantly, the ossicles generated from MSPC from almost half of the donors become the site of ectopic bone marrow development. Further experiments suggest that maintenance of MSPC stemness by platelet-derived factors during cell expansion is paramount to this effect. Methods and Results. MSPC are expanded from human bone marrow following standard protocols, with culture media supplemented either with 10% FBS or with 10% HPL. Cultured MSPC are resuspended in a non-mineral collagen/laminin matrix (Matrigel®) and injected subcutaneously into immune-deficient NSG mice. Bone formation is monitored non-invasively by osteosensitive near-infrared imaging and/or by histology on paraffin-embedded ossicles. HPL-derived MSPC from 13 out of 13 donors form bone in vivo compared to only 2 out of 10 FBS-derived MSPC. Extensive cartilage formation is observed as early as one week after implantation, while signs of ossification appear from the third week onwards. In addition, ossicles generated by HPL-MSPC from 6 out of 13 donors become colonised by mouse bone marrow, indicating that platelet-derived factors maintain the capacity of MSPC to reconstitute a functional bone marrow niche. Bone formation in vivo is considerably delayed, but not blocked, by pre-treatment of the cells with Cholera Toxin, indicating that a Gα protein-coupled receptor (GPCR-α) ligand is partly responsible for the biological effect of HPL. In vitro, HPL and FBS-derived cells display remarkably little phenotypic differences, with the notable exception of the stemness-associated surface marker SSEA-4 which is consistently more expressed on HPL- than on FBS-derived MSPC. Interestingly, serial passage in FBS medium of cells first expanded in HPL medium leads to the rapid loss of surface SSEA-4 expression, paralleled by a loss of osteogenic and bone marrow support potential in vivo. Loss of SSEA-4 expression is also observed in cells serially passaged in HPL medium containing the PDGF receptor-β inhibitor Imatinib, and ossicles derived from these cells are no longer able to attract mouse bone marrow in vivo. These data suggest that human platelet-derived factors allow MSPC to retain their stem cell potential in culture as well as in the ectopic bone microenvironment. This hypothesis is further supported by the observation that human MSPC can be re-isolated and re-expanded from bone marrow-infiltrated ossicles and, remarkably, build bone again when re-injected into NSG mice. Conclusion. Our data suggest that platelet-derived factors, including PDGF and an (as yet unidentified) GPCR-α ligand, contribute in vitro to maintain BM-MSPC stemness. In vivo, they efficiently drive BM-MSPC to differentiate along the chondrogenic and osteogenic lineages, while preserving MSPC bone marrow-support function. We conclude that therapeutic approaches using MSPC for skeletal regeneration should preferentially use early passage BM-MSPC expanded in HPL-supplemented medium. Furthermore, the observed correlation between surface SSEA-4 expression and multipotency in BM-MSPC can be exploited to monitor the quality of the cell preparations. Disclosures: No relevant conflicts of interest to declare.

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