A kinetic model of bone marrow neutrophil production that characterizes late phenotypic maturation

Acute inflammatory stimuli rapidly mobilize neutrophils from the bone marrow by shortening postmitotic maturation time and releasing younger neutrophils; however, the kinetics of this change in maturation time remains unknown. We propose a kinetic model that examines the rate of change in neutrophil average age at exit from the bone marrow during active mobilization to quantify this response and use this model to examine the temporal profile of late neutrophil phenotypic maturation. Total and CD10(-)/CD16(low) circulating neutrophils were quantified in cardiac surgery patients during extracorporeal circulation (ECC). Net growth in the circulating neutrophil pool occurred during the procedural (0.04 +/- 0.02 x 10(9) x l(-1) x min(-1)), warming (0.14 +/- 0.02 x 10(9) x l(-1) x min(-1)), and weaning (0.12 +/- 0.06 x 10(9) x l(-1) x min(-1)) phases of ECC. When applied to our differential equation mathematical model, these results predict that neutrophil average age at exit from the bone marrow decreased continually during ECC, resulting in average neutrophil release 8.44 +/- 2.20 h earlier during the weaning phase than at the beginning of ECC sampling. Modeling of concurrent changes in CD10(-)/CD16(low) neutrophil numbers indicates that CD10 expression is directly related to neutrophil mean age and predicts that the proportion of mobilizable postmitotic neutrophils that are CD10(+) increases from 64 to 81% during these sampled 8.4 h of maturation.     

Suppressive effects of swainsonine on C6 glioma cell in vitro and in vivo

Swainsonine, an extract from Astragalus membranaceus, is known for its anti-cancer effects and could prevent metastases. In order to investigate the effects and mechanisms of swainsonine in C6 glioma cells, we carry out correlated experiments in vitro and in vivo. After treatment with swainsonine, the effective dose and IC₅₀ value of swainsonine in the C6 glioma cell were examined using the MTT assay. Cell cycle distribution and apoptotic rates were analyzed using FCM and [Ca²⁺]_i was measured by LSCM. Expressions of p16 and p53 protein were evaluated by immunocytochemical methods. Simultaneously, glioma-bearing rats were administered swainsonine at doses of 2, 4 and 8 mg/kg body wt. The inhibition rate was calculated and pathological sections were observed. The results indicated that the growth of C6 glioma cells is inhibited by swainsonine in vitro, with an IC₅₀ value within 24 h of 0.05 μg/ml. Increases in swainsonine correlate with S phase percentages of 11.3%, 11.6% and 12.4%, respectively. Moreover, the expression of apoptosis inhibiting p53 and p16 protein decreases gradually. Tumor weight in vivo decreased clearly and HE dyeing of tumor tissue showed gray, its texture was soft, with necrosis and hemorrhagic concentrated inward. Swainsonine could inhibit the proliferation of C6 glioma cells in vitro and the growth of C6 glioma in vivo. The mechanisms of swainsonine-induced apoptosis may relate with the expression of apoptosis-related genes and overloading-[Ca²⁺]_i-induced endoplasmic reticulum stress.     

Ultrastructural localization of peroxidase activity in developing neutrophil granulocytes from human bone marrow

Summary Developing neutrophil granulocytes of normal human bone marrow were investigated with the diaminobenzidine technique to determine the ultrastructural localization of peroxidase activity. Neutrophil granulocytes have three types of granule: nucleated, azurophil, and specific granules. These granules are produced consecutively during the eomyelocyte stage, the promyelocyte stage, and the myelocyte stage, respectively.The organelles involved in the production of granules, i.e., the nuclear envelope, rough endoplasmic reticulum, and Golgi apparatus, are peroxidase positive during the eomyolocyte and promyelocyte stages and peroxidase negative thereafter. This pattern differs for the granules themselves: nucleated granules are negative in the eomyelocyte and become positive in the promyelocyte. Azurophil granules become positive in the promyelocyte. Specific granules are negative.Our observations highly suggest that small golgi-derived peroxidase-positive vesicles are involved in the maturation of both nucleated granules and azurophil granules.In honour of Prof. P. van Duijn     

Ultrastructural localization of peroxidase activity in developing neutrophil granulocytes from human bone marrow

Developing neutrophil granulocytes of normal human bone marrow were investigated with the diaminobenzidine technique to determine the ultrastructural localization of peroxidase activity. Neutrophil granulocytes have three types of granule: nucleated, azurophil, and specific granules. These granules are produced consecutively during the eomyelocyte stage, the promyelocyte stage, and the myelocyte stage, respectively. The organelles involved in the production of granules, i.e., the nuclear envelope, rough endoplasmic reticulum, and Golgi apparatus, are peroxidase positive during the eomyelocyte and promyelocyte stages and peroxidase negative thereafter. This pattern differs for the granules themselves: nucleated granules are negative in the eomyelocyte and become positive in the promyelocyte. Azurophil granules become positive in the promyelocyte. Specific granules are negative. Our observations highly suggest that small Golgi-derived peroxidase-positive vesicles are involved in the maturation of both nucleated granules and azurophil granules.     

Formation and proliferative effects of lipoxins in human bone marrow

Lipoxins A4 and B4 together with the all-trans lipoxin (LX) isomers were produced by normal human bone marrow cell suspensions after incubation with ionophore A23187. Both LXA4 and LXB4 enhanced the growth of myeloid progenitor cells in semisolid agar in the presence of suboptimal concentrations of recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF). Lipoxin A4 at 10(-10) M stimulated the colony formation in 13 out of 15 tested human bone marrows with a mean (+/- SEM) increase of 47 +/- 11% (p = 0.001). A similar stimulatory effect was observed after addition of LXB4 (10(-10) M). The monohydroxyeicosatetraenoic acids 5-, 12- and 15-HETE did not affect colony growth. In addition, LXA4 (10(-8) M) efficiently counteracted the increased colony formation induced by leukotriene C4 (10(-10) M), suggesting an antagonistic relationship between these lipoxygenase products. The results support a role for lipoxins in the regulation of human myelopoiesis.     

Understanding heterogeneity of human bone marrow plasma cell maturation and survival pathways by single-cell analyses

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Radioautographic studies on the formation and maturation of bone marrow lymphocytes in mice

Abstract. DNA labelling by [³H]thymidine and the sandwich radioimmunolabelling method were used to characterize marrow lymphoid cells and to study the kinetics of production and maturation of small lymphocytes in the bone marrow of adult mice. Marrow lymphoid cells consisted of non-proliferating small lymphocytes, 30–40% of which had detectable surface immunoglobulin (SmIg), and proliferating large lymphoid cells lacking SmIg. Double-labelling experiments employing [³H]thymidine in vivo followed by sandwich radioimmunolabelling in vitro indicated that marrow small lymophocytes lack detectable SmIg when they are formed but develop SmIg within the first few days after production. Marrow lymphocytopoiesis includes; (1) praliferation of large lymphoid cells, which are presumptive small lymphocyte progenitors, which have a cell cycle time of 14–15 hr, and (2) a 3–5-day intramyeloid stage when many newly formed small lymphocytes undergo maturational changes towards the B cell lineage.     

Human granulocyte colony stimulating factor: effects on human long-term bone marrow cultures

Human granulocyte colony stimulating factor (G-CSF) was previously shown to support the survival and proliferation of early myeloid progenitors (pre-CFU) that are capable of generating more mature CFU-GM progenitor cells. To evaluate the scope of action of G-CSF in the hierarchy of hematopoietic stem cells, we studied the effects of recombinant G-CSF (rhG-CSF) on long-term cultures of normal human bone marrow cells (LTBMC). We found that rhG-CSF predominantly influenced initial cell proliferation and expansion of CFU-GM progenitor cells in LTBMC before establishment of a confluent adherent layer. In rhG-CSF-treated LTBMC, the stromal cell layer was associated with a higher proliferative capacity and progenitor cell content as compared to control cultures. This effect was pronounced early after layer confluence and was gradually lost with culture time. rhG-CSF did not alter the duration of the productive phase of LTBMC, suggesting that it may not be active on the hematopoietic stem cells responsible for LTBMC propagation. Alternatively, stromal cells may exert tight regulatory control over progenitor cells, even in the presence of rhG-CSF.     

Cytogenetic effects of ribavirin on mouse bone marrow

The micronucleus test and mitotic chromosome analysis were used to study the in vivo mutagenic activity of ribavirin on bone marrow cells of Swiss albino mice. To determine the incidence of micronuclei, mice were injected i.p. twice, at an interval of 24 h. with the drug at doses of 20, 100 and 200 mg/kg. Animals were killed 6 h after the second dose and bone marrow was examined for the presence of micronuclei in developing erythrocytes. Ribavirin significantly (P less than 0.05) induced micronuclei in polychromatic erythrocytes at all doses. A study was conducted to investigate the cytogenetic effect of the drug on mitotic chromosomes. Ribavirin at 200 mg/kg/day was administered to mice for 3 and 5 days. Repeated treatment with the high dose of ribavirin produced a highly significant (P less than 0.02) increase in abnormal metaphase spreads. The results indicate that ribavirin is mutagenic to bone marrow cells of mice as evaluated by the micronucleus test and by chromosome analysis.     

AcSDKP抑制体外培养条件下人骨髓间充质干细胞的增殖

N-乙酰基-丝氨酰-天冬氨酰-赖氨酰-脯氨酸(N-acetyl-seryl-aspartyl-lysyl-proline,AcSDKP)是一种具有生理调控活性的四肽因子,对造血干/祖细胞增殖具有抑制作用。本研究采用集落形成实验、甲基偶氮唑盐(MTT)比色法、细胞分裂指数测定等方法,考察了AcSDKP对体外培养的人骨髓间充质干细胞(mesenchymal stem cell,MSC)增殖的影响。结果显示,在AcSDKP浓度为1×10-12mol／L-1×10-9mol／L的培养体系中,人骨髓MSC集落生成率和大小、活力细胞数和分裂指数均降低,最大效应浓度为1×10-11mol／L。以上实验结果表明,在体外培养条件下,一定浓度的AcSDKP对人骨髓MSC 的增殖具有抑制作用。     

Lipid composition of human bone marrow

1. A modified method for the analysis of phospholipid mixtures by selective hydrolysis is described. 2. The phospholipid compositions of normal human bone marrow and of the bone marrows of patients who died with anaemia or various forms of leukaemia were investigated. 3. Phospholipids from normal bone marrow comprised about 44% of lecithin, 4% of choline plasmalogen, 7% of glyceryl ether phospholipid (choline base), 10% of sphingomyelin, 22% of phosphatidylethanolamine plus phosphatidylserine, 8% of ethanolamine plasmalogen and 5% of glyceryl ether phospholipid (ethanolamine base). 4. The proportion of kephalin (i.e. phosphatidylethanolamine plus phosphatidylserine) in the pathological bone marrows tended to be lower than normal. No other consistent differences were observed between the normal and pathological samples. 4. A ceramide dihexoside was isolated from normal bone marrow.