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PubMedDate not supplied

[Modification of a method for determining the cytotoxic activity of lymphocytes--natural killers in space experiments].

A space modification of a method for determining the cytotoxic activity of natural killers (NK) uses cultures of human lymphocytes and re-inoculated suspension of tumorous myeloblasts K-562. The 3H-uridine labeled target-cells fixated in 0.1 % formalin solution proved to be best suited to the conditions prohibiting regular recharge of cultural medium and utilization of radioactive agents. The following practical conclusions were made from the space-flown experiment: pre-launch procedures (lymphocyte isolation and delivery to the launch site) and time period till dock must not be longer than 5 days (1) and during transportation to the orbiting vehicle cell cultures must be kept at 18-20 degrees C (2). The modified method can be employed to study various aspects of cell interaction such as the NK ability to destroy target-cells in suspension cultured exposed in microgravity.

Antimicrobial Cationic Peptides↗
PubMed2004

The dynamics of precursor cells in the olfactory epithelium of juvenile and adult guinea pigs.

The dynamics of precursor cells in the olfactory epithelium of juvenile and adult guinea pigs were examined by immunohistochemical double staining using bromodeoxyuridine (BrdU), the neural cell adhesion molecule (N-CAM) and the protein gene product 9.5 (PGP9.5). Expression of apoptotic cells in the olfactory epithelium with the use of the TdT-mediated dUTP biotin nick end labeling (TUNEL) method was also observed. BrdU was given to healthy guinea pigs at the ages of 2 weeks and 6 months old. Tissue specimens were serially collected 1 h to 28 days after administration. BrdU-labeled cells were seen above the basal cell layer after 1 h and migrated to the middle layer of the olfactory epithelium, after 1 day in juveniles and 5 days in adults with expression of N-CAM. PGP9.5 was observed in BrdU-labeled cells after 5 days in juvenile guinea pigs and 7 days in adult. At 14 days after administration, BrdU-labeled cells in the epithelium appeared to decrease. However, a few of these cells were recognized above the basal cell layer after 28 days. The number and location of TUNEL-positive cells did not significantly differ between the juvenile and adult olfactory epithelium. Therefore, we conclude that the division speed from stem cells in juveniles is faster than that in adults, and apoptosis is unaffected by aging in the normal olfactory epithelium.

Age Factors↗
PubMed2005

Gene expression profiling of human promyelocytic cells in response to infection with Anaplasma phagocytophilum.

Anaplasma phagocytophilum (Rickettsiales: Anaplasmataceae) causes human, equine and canine granulocytic anaplasmosis and tick-borne fever of ruminants. The rickettsia parasitizes granulocytes and bone marrow progenitor cells, and can be propagated in human promyelocytic and tick cell lines. In this study, microarrays of synthetic polynucleotides of 21,329 human genes were used to identify genes that are differentially expressed in HL-60 human promyelocytic cells in response to infection with A. phagocytophilum. Semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) of selected genes confirmed the results of the microarray analysis. Six genes in the A. phagocytophilum-infected cells were found to be upregulated greater than 30-fold, while expression of downregulated genes most often did not change more than sixfold. Genes that were found to be differentially regulated in infected cells were those essential for cellular mechanisms including growth and differentiation, cell transport, signalling and communication and protective response against infection, some of which are most likely necessary for infection and multiplication of A. phagocytophilum in host cells. The differentially regulated genes described herein provide new information on the gene expression profiles in A. phagocytophilum-infected HL-60 cells, thus expanding in a global manner the existing information on the response of mammalian cells to A. phagocytophilum infection.

Anaplasma phagocytophilum↗
PubMed1988

Hydrocortisone modulates colony-stimulating activity produced by human bone marrow-derived adherent cells.

In an attempt to clarify the significance of hydrocortisone (HC) in human long-term bone marrow cultures, the production of colony-stimulating activity (CSA) and colony-enhancing activity (CEA) by human bone marrow-derived adherent cells (MDAC) and the modulation by HC were examined. The CSA production by MDAC was demonstrated using bilayer agar cultures. After treatment of MDAC with 10(-6) mol/l HC, the CSA production was markedly enhanced, and after treatment of macrophages with 10(-6) mol/l HC, the CSA production was inhibited. When added to a granulocyte-macrophage precursor cells (CFU-GM) assay system, HC inhibited colony formation. These results suggest that HC treatment directly stimulates CSA production by nonmacrophage cells of MDAC. The conditioned medium of the confluent layer of MDAC contained CEA, which was not influenced by the HC treatment of MDAC. Thus, HC plays an essential role in granulopoiesis in vitro, enhancing the CSA production by MDAC and inhibiting the differentiation of CFU-GM.

Bone Marrow↗
PubMed1990

[Methods of isolating hematopoiesis islets from the bone marrow].

The effectiveness of the enzymatic and mechanical methods for isolation of hemopoietic islets (HI) from rat bone marrow was comparatively studied. It has been shown that the use of collagenase brings higher yields of HI than the mechanical treatment of bone marrow, mainly at the expense of associations containing no central phagocytizing macrophages. When the mechanical method of HI isolation was used, erythroid islets associated with mononuclear phagocytes prevailed, while with the enzymatic method mainly erythro-granulocytic islets associated with macrophages or reticular cells were isolated. It has been suggested that granulocytic and erythro-granulocytic associations are less resistant to the mechanical treatment.

Animals↗
PubMed1990

Reversal of azidothymidine-induced bone marrow suppression by 2',3'-dideoxythymidine as studied by hemopoietic clonal culture.

The hematological toxicity of the antiviral drug azidothymidine (AZT) was studied in vitro by measuring the cell growth of hematopoietic progenitor cells obtained from normal bone marrow. AZT inhibited the growth of granulocyte-macrophage (CFU-GM) and early and late erythroid (BFU-E and CFU-E) progenitors in a dose-dependent manner. The hemopoietic inhibition of AZT was significantly reduced by the addition of 2',3'-dideoxythymidine (2',3'-ddT). The results suggest that 2',3'-ddT may be of potential value in reducing AZT toxicity.

Bone Marrow↗
PubMed1994

Granulocyte colony stimulating factor: from laboratory bench to clinical use.

A great deal has been learned about haematopoiesis since the discovery, purification and cloning of haemopoietic growth factors. The function of these growth factors is to promote survival and enhance mitosis (at higher concentrations), and influence differentiation and development of stem cells and progenitor cells. The type of mature cells produced reflects the combinations of growth factors to which the stem cells are exposed. Growth factors may also control the functional activity of mature cells. Treatment with granulocyte colony stimulating factor (G-CSF) produces neutrophils that are at least equivalent to those produced during normal haematopoiesis and does not appear to deplete the haematopoietic stem or progenitor cell populations.

Animals↗
PubMed2004

c-Kit receptor (CD117) expression on myeloblasts and white blood cell counts in acute myeloid leukemia.

BACKGROUND: The c-Kit receptor is considered to play a crucial role in hematopoiesis. Induction of mobilization of hematopoietic cells in the bone marrow requires cooperative signaling through c-Kit and c-Kit ligand pathway, and these interactions are important in the retention of stem cells within the bone marrow. Therefore, we analyzed c-Kit density on the leukemic myeloblasts of patients with acute myeloid leukemia (AML) in relation to white blood cell count (WBC) in the peripheral blood. METHODS: Bone marrow aspirates collected from patients with AML and bone marrow aspirates and leukapheresis products after granulocyte colony-stimulating factor blood mobilization from adult volunteers were studied. To determine the level of c-Kit receptor expression, we applied quantitative (relative fluorescence intensity and antibody binding per cell) cytometric methods. RESULTS: Our data showed negative correlation between the level of c-Kit expression intensity on myeloblasts and the number of leukocytes in blood of AML patients. The c-Kit receptor density on myeloblasts in patients with low WBC was significantly stronger than that on myeloblasts in patients with high WBC. In the latter patient group, the density c-Kit receptor on myeloblasts was similar to that on CD34(+) cells in mobilized peripheral blood. CONCLUSIONS: The obtained data suggest an involvement of c-Kit receptor in the regulation of leukemic myeloblasts egress to the peripheral blood.

Female↗
PubMedDate not supplied

[The granulocyte-macrophage colony stimulating factor and the radiation protective effect of yeast mannan].

The effect of mannan polysaccharide on the haemopoiesis recovery in irradiated mice has been investigated. Mannan has been shown to exert both the protective and the stimulatory effect: it accelerates restoration of femur bone marrow cellularity and nucleate cell number in the peripheral blood and causes a larger initial yield and subsequent more rapid postirradiation dynamics of pluripotent haemopoietic stem cells and precursor cells of granulocytes and macrophages.

Animals↗
PubMed1976

Inhibition of murine granulopoiesis in vitro by dexamethasone.

The effect of dexamethasone on mouse bone marrow granulocyte-monocyte precursor cells (CFU-C) was studied in vitro. Dexamethasone inhibited colony formation in the presence of maximally stimulating concentrations of colony-stimulating activity. A mean colony reduction of 55% occurred at 10(-9) M dexamethasone and inhibition was observed at concentrations as low as 10(-12)M. The dexamethasone suppression could be abrogated by progesterone. These studies provide evidence that the committed granulocyte-monocyte stem cell has a glucocorticoid receptor mechanism that when activated results in inhibition of cellular proliferation in vitro.

Agar↗
PubMedDate not supplied

Expansion and differentiation of human hematopoietic cells from static cultures through small-scale bioreactors.

Maintenance of the progenitor cells responsible for hematopoiesis has generally been accomplished using a feeder layer of stromal cells in stationary culture. Here, we compared the expansion of the total cell and progenitor cell populations using low-density mononuclear cells (LDMCs) obtained from human bone marrow in static culture (T-flasks) and in different cell culture bioreactors designed for the scale-up of mammalian cells. Static cultures were performed without the presence of a previously established stromal cell layer. Expansion of marrow in all cases was accomplished through the use of added cytokines such as IL-3, GM-CSF, and c-kit ligand. The results for the total cell expansion in static culture ranged from 4.4- to 32-fold. The cell number increase was affected by such factors as patient to patient variability, freeze-thawing, and the combination of cytokines used. Due to widespread use and the small amount of marrow needed, static cultures were used as a basis for comparison with other expansion systems. The cell culture systems used to evaluate the scale-up of marrow cultures included suspension, microcarrier, airlift, and hollow fiber bioreactors. Using identical media, cytokines, and feed schedules, LDMCs in the suspension bioreactor expanded to a value of 1.6 compared to a normalized value of 1.0 for static cultures for the two runs investigated. Expansion results for microcarrier cultures averaged 0.75 when compared to static cultures. A cell number increase in the airlift bioreactor resulted in an expansion which was 0.70 of the control static culture. Granulocyte-macrophage and erythroid progenitor assay data were also evaluated for the suspension, microcarrier, and airlift bioreactors.(ABSTRACT TRUNCATED AT 250 WORDS)

Biotechnology↗
PubMed2005

Myeloid enzymes profile related to the immunophenotypic characteristics of blast cells from patients with acute myeloid leukemia (AML) at diagnosis.

The purpose of this study was to assess the possible relationship between the cytochemical enzyme profile and immunophenotypic characteristics of distinct acute myeloid leukemia (AML) subtypes in discrete stages of leukemic cells maturation. As the proportion of leukemic blast cells is critical for exact cytochemical analysis, study was restricted to the evaluation of 48 adult and pediatric patients with newly diagnosed AMLs with 80% or more blasts in analyzed samples. The cytochemical investigation of myeloperoxidase (MPO), Sudan black B (SBB), chloroacetate esterase (CAE), alpha-naphthyl butyrate esterase (ANBE), alpha-naphthyl acetate esterase (ANAE) and acid phosphatase (AP) in peripheral blood and/or bone marrow was performed. The immunophenotype was examined for the maturation dependent myeloid antigens CD13, CD33, CD11b, CD14, CD15, CD65, CD36, cytoplasmic MPO, non-lineage associated CD34 and HLA-DR antigens, lymphoid- associated antigens CD7, CD4, CD38 as well as natural killer cell associated marker CD56. Flow cytometry by double marker staining and visualization of pathologic cells in dot plots reflected immunophenotypic aberrancy and degree of cell maturation. The patients were classified into AML subtypes M0- M2, M3, M4 and M5 according to the main morphological, cytochemical and immunophenotypical features. The variable combinations of MPO, SBB, CAE and ANBE were identified in relation to immunophenotype. The cytochemical profile of blasts was in concordance with immunophenotype, particularly in more differentiated AML subtypes, M3, M4 and M5. The findings of myeloid antigens expression and cytochemical features in poorly differentiated AML subtypes showed no practical relevance of cytochemical analysis. Notwithstanding that the cytochemical analysis of AML subtypes not sufficiently identifies the distinct aberrancies in heterogeneous leukemic blast cell populations, evaluation of the cytochemical profile in connection with immunophenotyping may help to classify the AML patients to relevant subtypes with more accuracy.

Acute Disease↗
PubMed1994

Marked and reproducible increase in trilineage blood cell counts by administration of granulocyte colony-stimulating factor in a patient with refractory anaemia with excess blasts in transformation.

We report a patient with refractory anaemia with excess blasts in transformation (RAEB-t) who presented with severe pancytopenia and received four intermittent series of granulocyte colony-stimulating factor (G-CSF) treatment over 1.5 years. In addition to the increase in mature neutrophils, platelet count and haemoglobin level were dramatically increased. These haematological improvements were dependent on G-CSF during these treatment series. Bone marrow colony assay revealed that G-CSF increased both CFU-E- and BFU-E-derived colonies in vitro. Clinical usage of G-CSF in myelodysplastic syndrome (MDS) is discussed, with particular emphasis on mechanisms of trilineage response.

Anemia, Refractory, with Excess of Blasts↗
PubMed1985

Effect of D-penicillamine on human granulopoiesis in vitro.

The effect of D-penicillamine on human bone marrow granulocyte-monocyte precursor cells (CFU-C) is studied in vitro. Bone marrow samples were obtained from 47 donors (40 patients and 7 healthy volunteers) and cultured in semisolid agar in the presence of various concentrations of the drug. We found an inhibitory, possibly dose-dependent, effect of D-penicillamine on colony formation. The pathophysiological mechanism and the clinical relevance of this effect remain poorly understood. These findings, however, emphasize the need for careful monitoring of the granulocyte counts during D-penicillamine therapy.

Adult↗
PubMed2000

Prostaglandin E2 receptor antagonist (SC-19220) treatment restores the balance to bone marrow myelopoiesis after burn sepsis.

BACKGROUND: Although prostaglandin E2 (PGE2) has been shown to be immunosuppressive, its role in the development of specific bone marrow myeloid lineages after thermal injury and sepsis has yet to be elucidated. The purpose of this study was to demonstrate that alterations in bone marrow progenitor proliferation favoring monocytopoiesis in burn sepsis can be restored by blocking the cellular interactions of PGE2. METHODS: A murine model of burn sepsis with and without treatment with SC-19220, a PGE2 receptor antagonist, was used to determine peripheral monocyte and neutrophil counts as well as the colony forming potential of colony-stimulating factor responsive bone marrow progenitors. RESULTS: Burn sepsis augmented the growth of the early colony-forming unit granulocyte-macrophage and monocyte progenitors and the number of circulating monocytes, whereas granulocyte progenitors and circulating neutrophils demonstrated an opposite response. Treatment with SC-19220 nearly reversed these alterations. CONCLUSION: These data indicate that abrogating PGE2's actions during burn sepsis can restore the balance in bone marrow granulocyte and monocyte production, further consolidating the pivotal role PGE2 plays in the pathogenesis of burn sepsis.

Animals↗
PubMed2005

Intranucleolar translocation of AgNORs in early granulocytic precursors in chronic myeloid leukaemia and K 562 cells.

The present study was undertaken to provide missing information on the distribution of AgNORs in large nucleoli of human leukaemic early granulocytic precursors in vivo as well as in vitro. In vivo, the distribution of AgNORs was studied in early granulocytic precursors of patients suffering from chronic myeloid leukaemia who were both untreated and treated with imatinib mesylate. AgNORs were visualized by silver reaction under conditions which facilitated to see their distribution by light microscopy. In vitro, the distribution of AgNORs was studied in proliferating and ageing K 562 cells which originated from chronic myeloid leukaemia. In vitro, the ageing of K 562 cells produced intranucleolar translocation of AgNORs to the nucleolar periphery. Such translocation was also observed in some leukaemic early granulocytic precursors in vivo, e.g. in bone marrow myeloblasts and promyelocytes of leukaemic patients. As was expected, the intranucleolar translocation of AgNORs in early granulocytic precursors was more frequent in patients treated with the cytostatic therapy--imanitib mesylate. The abovementioned findings suggest that myeloblasts and promyelocytes with AgNORs translocated to the periphery of large nucleoli might be in the ageing state, similarly as blastic cells of leukaemic myeloid origin (K 562 cells) in ageing cultures. Thus, the translocation of AgNORs might be a useful marker of premature ageing in the future and might contribute to the evaluation of the single cell state under various experimental as well as clinical conditions. However, more clinically oriented studies are required in this direction.

Cell Line, Tumor↗
PubMed2005

Influence of telomere length on short-term recovery after allogeneic stem cell transplantation.

OBJECTIVE: Telomeres shorten in somatic cells during aging and states of increased turnover, including hematopoietic stem cell transplantation. Fast hematopoietic recovery is critical for the patients' course after hematopoietic stem cell transplantation. It is unknown whether telomere length in hematopoietic stem cells (HSCs) predicts short-term hematopoietic recovery. METHODS: We quantified telomere length by flow fluorescence in situ hybridization analysis in HSCs and granulocytes of healthy stem cell donors and monitored time to peripheral blood cell recovery in transplanted hosts. Furthermore, we measured in vitro repopulation potency of HSCs by assaying for colony-forming units granulocyte-macrophage (CFU-GM). RESULTS: Telomere length in HSC shortens continuously in vivo and is comparable to telomere length in granulocytes from the same individual. Numbers of in vitro formed CFU-GM per HSC show an inverse relationship to age and telomere length. However, telomere length in HSCs was not correlated with short-term recovery after HSC transplantation. CONCLUSION: These findings suggest that healthy stem cell donors have sufficient telomere length reserve to repopulate a myeloablatively treated host, despite continuous aging of HSCs in vivo and decreased repopulation ability of HSCs from older donors in vitro.

Adolescent↗