The fate of rat bone marrow, spleen and periosteum cultivated in vivo in the diffusion chamber, with special reference to bone formation*1

AbstractBone marrow-derived mesenchymal stem cells (BM-MSCs), the common progenitor cells of adipocytes and osteoblasts, have been recognized as the key mediator during bone formation. Herein, our study aim to investigate molecular mechanisms underlying circular RNA (circRNA) AFF4 (circ_AFF4)-regulated BM-MSCs osteogenesis. BM-MSCs were characterized by FACS, ARS, and ALP staining. Expression patterns of circ_AFF4, miR-135a-5p, FNDC5/Irisin, SMAD1/5, and osteogenesis markers, including ALP, BMP4, RUNX2, Spp1, and Colla1 were detected by qRT-PCR, western blot, or immunofluorescence staining, respectively. Interactions between circ_AFF4 and miR-135a-5p, FNDC5, and miR-135a-5p were analyzed using web tools including TargetScan, miRanda, and miRDB, and further confirmed by luciferase reporter assay and RNA pull-down. Complex formation between Irisin and Integrin αV was verified by Co-immunoprecipitation. To further verify the functional role of circ_AFF4 in vivo during bone formation, we conducted animal experiments harboring circ_AFF4 knockdown, and born samples were evaluated by immunohistochemistry, hematoxylin and eosin, and Masson staining. Circ_AFF4 was upregulated upon osteogenic differentiation induction in BM-MSCs, and miR-135a-5p expression declined as differentiation proceeds. Circ_AFF4 knockdown significantly inhibited osteogenesis potential in BM-MSCs. Circ_AFF4 stimulated FNDC5/Irisin expression through complementary binding to its downstream target molecule miR-135a-5p. Irisin formed an intermolecular complex with Integrin αV and activated the SMAD1/5 pathway during osteogenic differentiation. Our work revealed that circ_AFF4, acting as a sponge of miR-135a-5p, triggers the promotion of FNDC5/Irisin via activating the SMAD1/5 pathway to induce osteogenic differentiation in BM-MSCs. These findings gained a deeper insight into the circRNA-miRNA regulatory system in the bone marrow microenvironment and may improve our understanding of bone formation-related diseases at physiological and pathological levels.

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Effects of dietary alteration of bicarbonate and magnesium on rat bone

AJP Renal Physiology ◽

10.1152/ajprenal.1986.250.2.f302 ◽

1986 ◽

Vol 250 (2) ◽

pp. F302-F307 ◽

Cited By ~ 4

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.

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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 ◽

10.3390/ma11101880 ◽

2018 ◽

Vol 11 (10) ◽

pp. 1880 ◽

Cited By ~ 10

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.

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Radioautographic Studies of Bone Marrow Lymphocytes in Vivo and in Diffusion Chamber Cultures

Blood ◽

10.1182/blood.v23.1.1.1 ◽

1964 ◽

Vol 23 (1) ◽

pp. 1-17 ◽

Cited By ~ 89

Author(s):

D. G. OSMOND ◽

N. B. EVERETT

Keyword(s):

Bone Marrow ◽

Large Scale ◽

Bone Marrow Cells ◽

Tritiated Thymidine ◽

Diffusion Chamber ◽

Blood Stream ◽

Turnover Time ◽

Diffusion Chambers ◽

Transitional Cells

Abstract Radioautography with tritiated thymidine has been utilized to examine the turnover rate and origin of small lymphocytes in the bone marrow of the guinea-pig. Very few marrow lymphocytes were initially labeled by a single injection of tritiated thymidine, but thereafter the number of labeled lymphocytes rapidly increased to high maximum levels at 3 days. Analysis of the labeling curves and grain counts indicates that the population of marrow lymphocytes is maintained in a dynamic steady state with an average turnover time of 3 days or less. Suspensions of bone marrow cells were isolated from the circulation within intraperitoneal diffusion chambers after short-term labeling with tritiated thymidine in vivo. Although very few small lymphocytes were labeled when introduced into the diffusion chambers, a considerable percentage became labeled during the subsequent culture period. Tritiated thymidine was also administered intravenously whilst excluded from one hind limb by the application of an occlusive compression bandage for 20 minutes. Very few labeled small lymphocytes were found after 72 hours in the tibial marrow of the initially occluded limb, whereas the normal high percentage was labeled in the control tibial marrow. These experiments do not demonstrate any large-scale influx of small lymphocytes from the blood stream into the marrow parenchyma. They suggest that newly formed small lymphocytes appear in the marrow as a result of the division of locally situated precursor cells, but the mechanism of intramedullary lymphocytopoiesis is uncertain. "Transitional" cells, intermediate in morphology between blast cells and small lymphocytes, synthesize DNA and are actively proliferative, but they do not appear to account fully for the rate of lymphocyte production. Certain large, undifferentiated labeled cells appeared in the bone marrow as a result of hematogenous migration. Some implications of these findings are discussed.

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In vivo bone formation by human bone marrow stromal cells: Effect of carrier particle size and shape

Biotechnology and Bioengineering ◽

10.1002/1097-0290(20010105)72:1<96::aid-bit13>3.0.co;2-a ◽

2000 ◽

Vol 72 (1) ◽

pp. 96-107 ◽

Cited By ~ 136

Author(s):

Mahesh H. Mankani ◽

Sergei A. Kuznetsov ◽

Bruce Fowler ◽

Albert Kingman ◽

Pamela Gehron Robey

Keyword(s):

Bone Marrow ◽

Particle Size ◽

Bone Formation ◽

Stromal Cells ◽

Bone Marrow Stromal Cells ◽

Marrow Stromal Cells ◽

Human Bone ◽

Particle Size And Shape ◽

In Vivo Bone Formation

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Effect of Zinc Supplementation on Renal Anemia in 5/6-Nephrectomized Rats and a Comparison with Treatment with Recombinant Human Erythropoietin

International Journal of Molecular Sciences ◽

10.3390/ijms20204985 ◽

2019 ◽

Vol 20 (20) ◽

pp. 4985 ◽

Cited By ~ 2

Author(s):

Hui-Lin Feng ◽

Yen-Hua Chen ◽

Sen-Shyong Jeng

Keyword(s):

Bone Marrow ◽

Recombinant Human Erythropoietin ◽

Zinc Supplementation ◽

Bone Marrow Cells ◽

Human Erythropoietin ◽

Post Surgery ◽

Rat Bone ◽

Marrow Cells

Anemia is a severe complication in patients with chronic kidney disease (CKD). Treatment with exogenous erythropoietin (EPO) can correct anemia in many with CKD. We produced 5/6-nephrectomized rats that became uremic and anemic at 25 days post surgery. Injection of the anemic 5/6-nephrectomized rats with 2.8 mg zinc/kg body weight raised their red blood cell (RBC) levels from approximately 85% of the control to 95% in one day and continued for 4 days. We compared the effect of ZnSO4 and recombinant human erythropoietin (rHuEPO) injections on relieving anemia in 5/6-nephrectomized rats. After three consecutive injections, both the ZnSO4 and rHuEPO groups had significantly higher RBC levels (98 ± 6% and 102 ± 6% of the control) than the saline group (90 ± 3% of the control). In vivo, zinc relieved anemia in 5/6-nephrectomized rats similar to rHuEPO. In vitro, we cultured rat bone marrow cells supplemented with ZnCl2, rHuEPO, or saline. In a 4-day suspension culture, we found that zinc induced erythropoiesis similar to rHuEPO. When rat bone marrow cells were supplement-cultured with zinc, we found that zinc stimulated the production of EPO in the culture medium and that the level of EPO produced was dependent on the concentration of zinc supplemented. The production of EPO via zinc supplementation was involved in the process of erythropoiesis.

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Two classes of murine granulocyte progenitor cells expressed in plasma clot diffusion chamber cultures

Blood ◽

10.1182/blood.v59.4.838.838 ◽

1982 ◽

Vol 59 (4) ◽

pp. 838-843 ◽

Cited By ~ 6

Author(s):

HN Steinberg ◽

PL Page ◽

SH Robinson

Keyword(s):

Bone Marrow ◽

Progenitor Cells ◽

Cytosine Arabinoside ◽

Diffusion Chamber ◽

Velocity Sedimentation ◽

Mouse Bone Marrow ◽

Plasma Clot ◽

Residual Concentration

Abstract Two distinct classes of granulocyte progenitor cells present in normal mouse bone marrow are expressed sequentially in the vivo plasma clot diffusion chamber culture system. By several criteria, progenitor cells giving rise to granulocyte colonies on day 4 of culture (CFU-D4) are different from those giving rise to colonies on day 7 (CFU-D7). These differences include: cell cycle activity as measured by in vitro incubation with cytosine arabinoside, residual concentration in the bone marrow after in vivo treatment of donor mice with cytosine arabinoside or methotrexate, resistance to osmotic lysis, size as determined by velocity sedimentation, and the morphology of the granulocyte colonies to which these cells give rise. The CFU-D7 appears to represent an earlier progenitor cell than the CFU-D4 in the differentiation pathway of the granulocyte and is analagous in many respects to the BFU-E in the erythroid pathway.

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