In Vivo Localization of Heterologous Anti-Erythrocyte Antibodies in the Rat Bone Marrow and Their Effect on the Proliferative Capacity of Immature Erythroid Cells

Abstract The in vivo localization of heterologous anti-erythrocyte antibodies in the rat bone marrow was determined by the I131-labeled antibody technic. I131-labeled anti-erythrocyte antibodies localized specifically in the bone marrow indicating the presence of localizing antibody. Both the localizing antibody and the incomplete antibody were thermostable, whereas hemolysins and hemagglutinins were thermolabile. Following an intravenous injection of antierythrocyte antibodies in rats, hemolysins and hemagglutinins were cleared rapidly from the plasma. The incomplete antibodies became attached to circulating red cells within 6 hours and red cell sensitization persisted for 1 week. The localizing antibody localized in the bone marrow within 30 minutes, leaving no activity in plasma. The anti-erythrocyte antibodies markedly reduced the uptake of tritiated thymidine by erythroblasts in vitro, demonstrating their inhibitory effect on the proliferative capacity of erythroblasts.

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The effect of chemotherapy on the kinetics and proliferative capacity of normal and tumorous tissues in vivo

Blood ◽

10.1182/blood.v45.1.107.107 ◽

1975 ◽

Vol 45 (1) ◽

pp. 107-118 ◽

Cited By ~ 18

Author(s):

SH Rosenoff ◽

JM Bull ◽

RC Young

Keyword(s):

Bone Marrow ◽

Dna Synthesis ◽

Colony Formation ◽

Antitumor Agents ◽

Tritiated Thymidine ◽

Chemotherapeutic Agents ◽

Proliferative Capacity

Abstract The proliferative state of a given tissue is a major determinant of its sensitivity to both phase-specific and cycle-specific chemotherapeutic agents. To study the extent of injury induced by antitumor agents to normal and tumorous tissues, a technique for following DNA synthesis as reflected in the incorporation of tritiated thymidine (3H-TdR) into DNA was compared to the conventional radioautographic technique of the labeling index (LI) and to the functional kinetic technique of granulocyte colony formation in vitro. Alterations in DNA synthesis induced by a single dose of cyclophosphamide in normal and tumorous tissues in vivo paralleled in many respects the changes seen when the more time-consuming techniques of the LI or granulocyte colony formation were employed. However, the recovery of granulocyte colony formation after cyclophosphamide therapy laged behind the recovery of DNA synthesis in the bone marrow, obscuring a kinetic event of potential therapeutic significance. The determination of DNA synthesis simultaneously in normal and tumorous tissues in vivo was easy to perform and supplied therapeutically pertinent results comparatively quickly.

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The effect of chemotherapy on the kinetics and proliferative capacity of normal and tumorous tissues in vivo

Blood ◽

10.1182/blood.v45.1.107.bloodjournal451107 ◽

1975 ◽

Vol 45 (1) ◽

pp. 107-118

Author(s):

SH Rosenoff ◽

JM Bull ◽

RC Young

Keyword(s):

Bone Marrow ◽

Dna Synthesis ◽

Colony Formation ◽

Antitumor Agents ◽

Tritiated Thymidine ◽

Chemotherapeutic Agents ◽

Proliferative Capacity

The proliferative state of a given tissue is a major determinant of its sensitivity to both phase-specific and cycle-specific chemotherapeutic agents. To study the extent of injury induced by antitumor agents to normal and tumorous tissues, a technique for following DNA synthesis as reflected in the incorporation of tritiated thymidine (3H-TdR) into DNA was compared to the conventional radioautographic technique of the labeling index (LI) and to the functional kinetic technique of granulocyte colony formation in vitro. Alterations in DNA synthesis induced by a single dose of cyclophosphamide in normal and tumorous tissues in vivo paralleled in many respects the changes seen when the more time-consuming techniques of the LI or granulocyte colony formation were employed. However, the recovery of granulocyte colony formation after cyclophosphamide therapy laged behind the recovery of DNA synthesis in the bone marrow, obscuring a kinetic event of potential therapeutic significance. The determination of DNA synthesis simultaneously in normal and tumorous tissues in vivo was easy to perform and supplied therapeutically pertinent results comparatively quickly.

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Effect of oxalate and malonate on red cell metabolism

Blood ◽

10.1182/blood.v70.5.1389.bloodjournal7051389 ◽

1987 ◽

Vol 70 (5) ◽

pp. 1389-1393

Author(s):

E Beutler ◽

L Forman ◽

C West

Keyword(s):

Pyruvate Kinase ◽

Cell Metabolism ◽

Inhibitory Effect ◽

Red Cells ◽

Red Cell ◽

Acid Analogue ◽

2,3 Dpg

The addition of oxalate to blood stored in Citrate-phosphate-dextrose (CPD) produces a marked improvement in 2,3-diphosphoglycerate (2,3-DPG) preservation; an increase in 2,3-DPG levels can also be documented in short-term incubation studies. Oxalate is a potent in vitro inhibitor of red cell lactate dehydrogenase, monophosphoglycerate mutase, and pyruvate kinase (PK). In the presence of fructose 1,6-diphosphate the latter inhibitory effect is competitive with phospho(enol)pyruvate (PEP). Determination of the levels of intermediate compounds in red cells incubated with oxalate suggest the presence of inhibition at the PK step, indicating that this is the site of oxalate action. Apparent inhibition at the glyceraldehyde phosphate dehydrogenase step is apparently due to an increase in the NADH/NAD ratio. Oxalate had no effect on the in vivo viability of rabbit red cells stored in CPD preservatives for 21 days. Greater understanding of the toxicity of oxalate is required before it can be considered suitable as a component of preservative media, but appreciation of the mechanism by which it affects 2,3-DPG levels may be important in design of other blood additives. Malonate, the 3-carbon dicarboxylic acid analogue of oxalate late did not inhibit pyruvate kinase nor affect 2,3-DPG levels.

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Use of 5-fluorouracil to analyze the effect of macrophage inflammatory protein-1 alpha on long-term reconstituting stem cells in vivo

Blood ◽

10.1182/blood.v81.6.1497.1497 ◽

1993 ◽

Vol 81 (6) ◽

pp. 1497-1504 ◽

Cited By ~ 33

Author(s):

VF Quesniaux ◽

GJ Graham ◽

I Pragnell ◽

D Donaldson ◽

SD Wolpe ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Inhibitory Effect ◽

In Vivo Model ◽

Hematopoietic Progenitors ◽

Inflammatory Protein ◽

Macrophage Inflammatory Protein

Abstract A macrophage-derived inhibitor of early hematopoietic progenitors (colony-forming unit-spleen, CFU-A) called stem cell inhibitor was found to be identical to macrophage inflammatory protein-1 alpha (MIP-1 alpha). We investigated the effect of MIP-1 alpha on the earliest stem cells that sustain long-term hematopoiesis in vivo in a competitive bone marrow repopulation assay. Because long-term reconstituting (LTR) stem cells are normally quiescent, an in vivo model was first developed in which they are triggered to cycle. A first 5-fluorouracil (5-FU) injection was used to eliminate later progenitors, causing the LTR stem cells, which are normally resistant to 5-FU, to enter the cell cycle and become sensitive to a second 5-FU injection administered 5 days later. Human MIP-1 alpha administered from day 0 to 7 was unable to prevent the depletion of the LTR stem cells by the second 5-FU treatment, as observed on day 7 in this model, suggesting that the LTR stem cells were not prevented from being triggered into cycle despite the MIP-1 alpha treatment. However, the MIP-1 alpha protocol used here did substantially decrease the number of more mature hematopoietic progenitors (granulocyte-macrophage colony-forming cells [CFC], burst- forming unit-erythroid, CFCmulti, and preCFCmulti) recovered in the bone marrow shortly after a single 5-FU injection. In vitro, MIP-1 alpha had no inhibitory effect on the ability of these progenitors to form colonies. This study confirms the in vivo inhibitory effect of MIP- 1 alpha on subpopulations of hematopoietic progenitors that are activated in myelodepressed animals. However, MIP-1 alpha had no effect on the long-term reconstituting stem cells in vivo under conditions in which it effectively reduced all later progenitors.

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Distinct, strict requirements for Gfi-1b in adult bone marrow red cell and platelet generation

Journal of Experimental Medicine ◽

10.1084/jem.20131065 ◽

2014 ◽

Vol 211 (5) ◽

pp. 909-927 ◽

Cited By ~ 37

Author(s):

Adlen Foudi ◽

Daniel J. Kramer ◽

Jinzhong Qin ◽

Denise Ye ◽

Anna-Sophie Behlich ◽

...

Keyword(s):

Bone Marrow ◽

Wide Spectrum ◽

Transcriptional Repressor ◽

Erythropoietin Receptor ◽

Red Cell ◽

Genome Wide ◽

Cytoplasmic Maturation ◽

Adult Bone

The zinc finger transcriptional repressor Gfi-1b is essential for erythroid and megakaryocytic development in the embryo. Its roles in the maintenance of bone marrow erythropoiesis and thrombopoiesis have not been defined. We investigated Gfi-1b’s adult functions using a loxP-flanked Gfi-1b allele in combination with a novel doxycycline-inducible Cre transgene that efficiently mediates recombination in the bone marrow. We reveal strict, lineage-intrinsic requirements for continuous adult Gfi-1b expression at two distinct critical stages of erythropoiesis and megakaryopoiesis. Induced disruption of Gfi-1b was lethal within 3 wk with severely reduced hemoglobin levels and platelet counts. The erythroid lineage was arrested early in bipotential progenitors, which did not give rise to mature erythroid cells in vitro or in vivo. Yet Gfi-1b−/− progenitors had initiated the erythroid program as they expressed many lineage-restricted genes, including Klf1/Eklf and Erythropoietin receptor. In contrast, the megakaryocytic lineage developed beyond the progenitor stage in Gfi-1b’s absence and was arrested at the promegakaryocyte stage, after nuclear polyploidization, but before cytoplasmic maturation. Genome-wide analyses revealed that Gfi-1b directly regulates a wide spectrum of megakaryocytic and erythroid genes, predominantly repressing their expression. Together our study establishes Gfi-1b as a master transcriptional repressor of adult erythropoiesis and thrombopoiesis.

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Effect of oxalate and malonate on red cell metabolism

Blood ◽

10.1182/blood.v70.5.1389.1389 ◽

1987 ◽

Vol 70 (5) ◽

pp. 1389-1393 ◽

Cited By ~ 11

Author(s):

E Beutler ◽

L Forman ◽

C West

Keyword(s):

Pyruvate Kinase ◽

Cell Metabolism ◽

Inhibitory Effect ◽

Red Cells ◽

Red Cell ◽

Acid Analogue ◽

2,3 Dpg

Abstract The addition of oxalate to blood stored in Citrate-phosphate-dextrose (CPD) produces a marked improvement in 2,3-diphosphoglycerate (2,3-DPG) preservation; an increase in 2,3-DPG levels can also be documented in short-term incubation studies. Oxalate is a potent in vitro inhibitor of red cell lactate dehydrogenase, monophosphoglycerate mutase, and pyruvate kinase (PK). In the presence of fructose 1,6-diphosphate the latter inhibitory effect is competitive with phospho(enol)pyruvate (PEP). Determination of the levels of intermediate compounds in red cells incubated with oxalate suggest the presence of inhibition at the PK step, indicating that this is the site of oxalate action. Apparent inhibition at the glyceraldehyde phosphate dehydrogenase step is apparently due to an increase in the NADH/NAD ratio. Oxalate had no effect on the in vivo viability of rabbit red cells stored in CPD preservatives for 21 days. Greater understanding of the toxicity of oxalate is required before it can be considered suitable as a component of preservative media, but appreciation of the mechanism by which it affects 2,3-DPG levels may be important in design of other blood additives. Malonate, the 3-carbon dicarboxylic acid analogue of oxalate late did not inhibit pyruvate kinase nor affect 2,3-DPG levels.

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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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Analysis of megakaryocyte ploidy in rat bone marrow cultures

Blood ◽

10.1182/blood.v74.6.1952.bloodjournal7461952 ◽

1989 ◽

Vol 74 (6) ◽

pp. 1952-1962

Author(s):

DJ Kuter ◽

SM Greenberg ◽

RD Rosenberg

Keyword(s):

Bone Marrow ◽

Culture Conditions ◽

Recombinant Erythropoietin ◽

Bone Marrow Cultures ◽

Ploidy Changes ◽

Macrophage Colony ◽

Vitro Model ◽

Rat Bone

Megakaryocytes undergo changes in ploidy in vivo in response to varying demands for platelets. Attempts to study the putative factor(s) regulating these ploidy changes have been frustrated by the lack of an appropriate in vitro model of megakaryocyte endomitosis. This report describes a culture system in which rat bone marrow is depleted of identifiable megakaryocytes and enriched in their precursor cells. Morphologically identifiable megakaryocytes appear when the depleted marrow is cultured in vitro. The total number of nucleated cells, as well as the number of megakaryocytes and their ploidy distribution, are quantitated very precisely by flow cytometry. Although the total number of nucleated cells declines by 35% to 40% over 3 days in culture, the number of megakaryocytes rises 10-fold. The number of nucleated cells, the number of megakaryocytes, and the extent of megakaryocyte ploidization behave as independent variables in culture and are dependent on the culture conditions. The addition of recombinant erythropoietin promotes a rise in the number of megakaryocytes and a shift in ploidy to higher values while recombinant murine granulocyte- macrophage colony stimulating factor is without effect on the cultured megakaryocytes. This in vitro system may provide a means to study those factors that affect megakaryocyte growth and ploidization.

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Human Bone Marrow-Derived Mesenchymal Stem Cell-Secreted Exosomes Overexpressing Microrna-124-3p Inhibit DLBCL Progression By Downregulating NFATc1

10.21203/rs.3.rs-117708/v1 ◽

2020 ◽

Author(s):

Xiaolei Ding ◽

Lingzhi Xu ◽

Xiuhua Sun ◽

Xiaoxuan Zhao ◽

Bing Gao ◽

...

Keyword(s):

Bone Marrow ◽

Cell Proliferation ◽

B Cell Lymphoma ◽

Inhibitory Effect ◽

Human Bone ◽

Human Bone Marrow ◽

Proliferation And Apoptosis ◽

Microrna 124

Abstract Background: Exosomes play important roles in intercellular communication by delivering microRNAs (miRNAs) that mediate tumor initiation and development, including those in diffuse large B cell lymphoma (DLBCL). To date, however, limited studies on the inhibitory effect of exosomes derived from human bone marrow-derived mesenchymal stem cells (hBMSCs) on DLBCL progression have been reported. Therefore, this study aimed to investigate the role of hBMSC-secreted exosomes carrying microRNA-124-3p in the development of DLBCL.Methods: Microarray-based expression analysis was adopted to identify differentially expressed genes and regulatory miRNAs, which revealed the candidate NFATc1. Next, the binding affinity between miR-124-3p and NFATc1 was using luciferase activity assays. The mechanism underlying NFATc1 regulation was investigated using lentiviral transfections. Subsequently, DLBCL cells were cocultured with exosomes derived from hBMSCs transfected with a miR-124-3p mimic or control. Proliferation and apoptosis were measured in vitro. Finally, the effects of hBMSC-derived miR-124-3p on tumor growth were investigated in vivo.Results: MiR-124-3p was downregulated while NFATc1 was upregulated in DLBCL cells. MiR-124-3p specifically targeted and negatively regulated the expression of NFATc1 in DLBCL cells, upregulated miR-124-3p-inhibited DLBCL cell proliferation and promoted apoptosis. In addition, we found that hBMSC-derived exosomes carrying miR-124-3p repressed DLBCL cell proliferation both in vitro and in vivo.Conclusion: hBMSC-derived exosomal miR-124-3p represses the development of DLBCL through the downregulation of NFATc1.

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