The effects of erythropoietin and recombinant cytokines (G-CSF, SCF, IL-3 and GM-CSF) on colony formation and self-renewal by erythroid burst-forming units (BFU-E) and granulocyte-macrophage progenitors (CFU-GM) from mobilized peripheral blood progenitor cells (PBPCs) of the normal donors and patients were investigated. To better understand how combinations of the cytokines may be appropriate for stem cell manipulation, a model involving replating of individual BFU-E and CFU-GM colonies has been used to study the effects of cytokines on colony formation and self-renewal. Based on granulocyte colony-stimulating factor (G-CSF) and erythropoietin(EPO) alone serve as a baseline with which to compare the effects of the combinations with stem cell factor (SCF), interleukin-3 (IL-3) or granulocyte-macrophage colony-stimulating factor (GM-CSF). The results revealed that: 1. BFU-E derived from PBPCs produced a significantly great numbers of subcolonies in EPO plus IL-3 and EPO+SCF+IL-3, and a difference in EPO+SCF compared with EPO alone. 2. Compared patients to normal donors the self-renewal ability of BFU-E was not influenced after EPO plus SCF or IL-3. 3. There was a significantly increased frequency of CFU-GM in the presence of SCF, IL-3, and GM-CSF used in conjunction with G-CSF. Comparing the frequencies of CFU-GM in patients and donors, patients' PBPCs were more sensitive to G-CSF+SCF and GMix (G-CSF+SCF+IL-3+GM-CSF), especially to G-CSF alone than donors. 4. There was a significant increase in AUC (area under the curve of subcolony distribution) in the presence of G-CSF combined with other cytokines. GMix was identified as the optimal combination of cytokines for both the expansion of CFU-GM as well as the expansion of clonogenic progenitor cells. 5. Donors have a high AUC than patients, especially there was a significant increase AUC in G-CSF alone (P = 0.0067).
Cytokines ; pharmacology ; Erythroid Precursor Cells ; drug effects ; Erythropoietin ; pharmacology ; Humans ; Leukocytes, Mononuclear ; drug effects ; physiology ; Recombinant Proteins ; pharmacology ; Stem Cells ; drug effects

Cell Differentiation ; drug effects ; Cell Separation ; Endothelial Cells ; cytology ; Fibronectins ; pharmacology ; Humans ; Leukocytes, Mononuclear ; cytology ; Stem Cells ; cytology

This study was aimed to investigate the effect of dexamethasone (Dex) on immunosuppressive ability of mesenchymal stem cells (MSC) during expansion and differentiation of MSC. MSC were cultured in 96-well flat-bottom plates. Proliferation assays were performed by using the BrdU colorimetric ELISA Kit. To explore the effect of Dex on MSC immunosuppressive ability, MSC were firstly cultured in complete culture medium for 14 d with Dex (10 nmol/L), and then, peripheral blood mononuclear cells (PBMNC) were co-cultured with MSC in 96-well flat-bottom plates for 3 d. Phytohemagglutinin A (PHA, 10 µg/ml) was used to stimulate activation of PBMNC. The concentrations of IFN-γ in culture supernatants was detected by ELISA. The results indicated that there was no obvious difference in representative phenotypes of MSC between experimental and control groups after MSC were treated with low concentration of Dex (10 nmol/L) for 14 d, but the suppression of Dex-treated MSC on lymphocyte activation in same concentration of cells was significantly reduced as compared with control group. After the Dex-treated MSC were co-cultured with IFN-γ for 12 h, the immunoregulatory ability of MSC was recovered in a certain degree. It is concluded that the Dex impairs the immunosuppressive ability of MSC, the IFN-γ can protect and reverse the immunosuppressive ability of MSC impaired by Dex, so that, when the immunoregulatory activity of MSC is investigated, it is necessary to avoid adding Dex in the culture medium.
Cells, Cultured ; Dexamethasone ; adverse effects ; Humans ; Immune Tolerance ; drug effects ; Interferon-gamma ; immunology ; Leukocytes, Mononuclear ; Lymphocyte Activation ; immunology ; Mesenchymal Stromal Cells ; cytology ; drug effects ; immunology

Intravenous immunoglobulin (IVIG) is being increasingly used to treat numerous immune-mediated diseases. However, there is a paucity of knowledge on the specific mode of action of IVIG in vivo. In this study, the in vitro effects of IVIG on peripheral blood mononuclear cell (PBMC) proliferation using phytohemagglutinin (PHA), anti-CD3 monoclonal antibody (MAb), phorbol myristate acetate (PMA), or purified protein derivatives (PPD) have been analyzed. The PBMCs were obtained from more than 10 individual donors. In all cases, IVIG almost completely inhibited PBMC proliferation at concentration above 20 mg/mL except when used in conjunction with PMA. PHA-induced proliferation of PBMCs at concentrations ranging from 1 to 15 mg/mL did not show significant differences. Anti-CD3 MAb-induced proliferation showed dose-dependent inhibition at concentrations ranging from 1 to 10 mg/mL. Interestingly, PMA-induced proliferation of PBMCs showed a dose-dependent increase at the same concentration range. PPD-induced proliferation of PBMC at concentrations ranging from 1 to 10 mg/mL did not show any statistically significant differences. These results suggest that high dose IVIG may be necessary to immune modulation in vivo and IVIG has various effects on PBMCs proliferation in limited concentration in vitro.
Cell Division/drug effects ; Cells, Cultured ; Dose-Response Relationship, Drug ; Human ; Immunoglobulins, Intravenous/*pharmacology ; Leukocytes, Mononuclear/*drug effects/physiology ; Tetradecanoylphorbol Acetate/pharmacology

OBJECTIVETo study the effect of hydroquinone on apoptosis of bone marrow mononuclear cells, and to evaluate the toxic effect of benzene on stem cells.

METHODSCell morphology was observed by HT fluorescent stain method, and DNA fragments were analyzed by agarose gel electrophoresis. Anti-Annexin V FITC plus PI staining for apoptotic and necrotic rate was examined by flow cytometer.

RESULTSAfter adding different concentrations of hydroquinone to the cells for 6 h culture, the fluorescent intensity of nucleus increased, the color of nucleus became deep and inhomogeneous, and the chromatin was condensed and distributed around the neucleus. DNA ladder was detected in all samples. Cell apoptotic rate in different concentration of hydroquinone groups was significantly higher than that in blank control group (P < 0.05). With the increase of the concentration of hydroquinone, the apoptotic and necrotic rate also increased. The optimal concentration of hydroquinone was 50 micro mol/L. When it was >or= 75 micro mol/L, the necrotic rate increased significantly. Hydroquinone-induced apoptosis was associated with culture time at the concentration of 50 micro mol/L, and the peak apoptotic time was 10 h, then the apoptotic rate decreased and necrotic rate increased.

CONCLUSIONHydroquinone can induce apoptosis of bone marrow mononuclear cells in vitro with dose-effect and time-effect relationship.

Apoptosis ; drug effects ; Bone Marrow Cells ; cytology ; Cells, Cultured ; Dose-Response Relationship, Drug ; Humans ; Hydroquinones ; pharmacology ; Leukocytes, Mononuclear ; cytology ; drug effects ; Mutagens ; pharmacology

OBJECTIVETo investigate the effect of ligustrazine (LT) on hematopoiesis in mice after bone marrow isotransplantation (iso-BMT).

METHODSThe typical model of iso-BMT was established and the model mice were randomly divided into two groups, the LT group treated with LT injection 0.2 ml and the control group treated with normal saline 0.2 ml, twice a day by gastrogavage. The following parameters were observed in the day 1, 7 and 14: peripheral blood cells, bone marrow mono-nuclear cells (BMMNC), heparin sulfate (HS) expression in bone marrow section by immunohistochemical SABC-AP method, stromal cell derived factor-1 (SDF-1) expression and CXC chemotaxis factor receptor 4 (CXCR4) expression.

RESULTSThe levels of peripheral WBC, platelet, BMMNC, CXCR4, HS, SDF-1 at the day 7 and 14 in the LT group were all higher significantly than those in the control group (P < 0.05 or P < 0.01).

CONCLUSIONLT could improve the bone marrow hematopoiesis in the early hematopoietic re-establishing stage after BMT.

Animals ; Bone Marrow Transplantation ; Female ; Hematopoiesis ; drug effects ; Leukocytes, Mononuclear ; drug effects ; Male ; Mice ; Mice, Inbred BALB C ; Pyrazines ; pharmacology ; Random Allocation ; Receptors, CXCR4 ; blood

OBJECTIVETo study the effects of palmitic acid (PA) on the proliferation of peripheral blood-derived endothelial progenitor cells (EPCs) in vitro.

METHODSThe mononuclear cells (MNCs) were isolated from the peripheral blood by Ficoll density-gradient centrifugation. The isolated EPCs were characterized by Di-LDI uptake and FITC-lectin binding assay using laser confocal microscope, and further identified by detection of CD34, CD133 and VEGFR2 expression using flow cytometry. The cultured EPCs were incubated in the presence of PA at the concentrations of 0, 50, 100, 200, 400 and 800 micromol/L for different durations (0, 12, 24, 36, 48 and 60 h). The cell morphology was observed and cell proliferation determined with CCK-8 assay.

RESULTSIncubation with 400 and 800 micromol/L of PA significantly inhibited the proliferative ability of EPCs as compared with the control group (P < 0.05). PA at 400 micromol/L had the strongest effect on the cell proliferation, and this effect was intensified with the passage of time, reaching the peak at 48 h with the growth inhibition rate of 58.59% (P < 0.05).

CONCLUSIONHigh-concentration PA can significantly inhibit the proliferation of EPCs in vitro.

Cell Differentiation ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Endothelial Cells ; cytology ; Humans ; Leukocytes, Mononuclear ; cytology ; Palmitic Acid ; pharmacology ; Stem Cells ; cytology

OBJECTIVETo study the effects of rare earth exposure on human telomerase and apoptosis of human peripheral mononuclear cells (PBMNs).

METHODSRare earth mine lot in Xunwu county, the biggest ion absorptive rare earth mine lot of China, was selected as the study site. Another village of Xunwu county, with comparable geological structure and social environment was selected as the control site. Thirty healthy adults were randomly selected from the study site as exposure group and another 30 healthy adults randomly selected from the control site as control group. The blood content of 15 rare earth elements, including La, Ce, Pr, Nd, Sm, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb, Lu and Y, were determined by inductive coupled plasma-source mass spectrometry (ICP-MS). The total contents of rare earth elements in the blood were calculated. The TRAP and FCM assays were carried out to analyse the telomerase and apoptosis of human PBMNCs respectively.

RESULTSIn the exposure group, the concentration of La, Ce, Dy and Y were significantly higher (P<0.001), and Pr, Nd, Sm, Gd and Yb were higher than those in the control group (P<0.05). The total content of rare earth in the blood of exposure group showed significant difference compared with control group (P<0.001). Telomerase activity in PBMNs of the exposure group was higher than that in the control group (P<0.05); there were 11 adults in the exposure group (30 adults) and 5 adults in control group (30 adults) showed positive telomerase activity. The average age of the exposure group was (38.69 +/- 8.02) years-old, while the control group was (40.45 +/- 9.02) years-old (P >0.05). It was found that there was a significant relationship between telomerase activity and the total content of rare earth elements (P <0.01). 3. The proportion of apoptosis was not different between the two groups (P >0.05), but the cells in the S-phase and G2-M phase were increased (P <0.01) in the exposed group.

CONCLUSIONThe telomerase activity of PBMNs in the rare earth elements exposed group was higher than that of the control group, and there is no effect on apoptotic rate of PBMNs, but may promote the diploid DNA replication, and increase the percentage of G2/M and S phase cells.

Apoptosis ; drug effects ; Environmental Exposure ; adverse effects ; Female ; Humans ; Leukocytes, Mononuclear ; cytology ; enzymology ; Male ; Metals, Rare Earth ; adverse effects ; analysis ; Telomerase ; metabolism

Adult ; Apoptosis ; drug effects ; Benzene ; poisoning ; Biomarkers ; analysis ; Bone Marrow Cells ; drug effects ; Female ; Flow Cytometry ; Humans ; Leukocytes, Mononuclear ; drug effects ; Lymphocytes ; drug effects ; metabolism ; Male ; Micronuclei, Chromosome-Defective ; drug effects ; metabolism ; Occupational Exposure ; analysis

OBJECTIVETo study the anticancer effect and mechanism of curcumin on Raji cells in vitro and compared the cytotoxicities of curcumin on Raji cells and normal human peripheral blood mononuclear cell (PBMC).

METHODSThe effects of curcumin on proliferation of Raji cells and human PBMC were tested by MTT assay, its effects on apoptosis of them were determined by Annexin-V/PI double-labeled cytometry and TUNEL, and its effects on DNA distribution in Raji cells was studied by PI single labeled cytometry.

RESULTSCurcumin showed marked inhibition on proliferation of Raji cell, could induce Raji cell apoptosis in time- and dose-dependent manner. After curcumin treatment, the cell cycle of Raji cells was blocked in G0/G1 and G2/M phase and those in the S phase decreased proportionally. But curcumin showed no significant effect on inhibiting proliferation or inducing apoptosis on human PBMC.

CONCLUSIONCurcumin could regulate the cell cycle of Raji cells and induce its apoptosis, so as to inhibit its proliferation, but with no significant cytotoxicity on human PBMC. It selectively affects the tumor cell.

Antineoplastic Agents, Phytogenic ; pharmacology ; Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Division ; drug effects ; Curcumin ; pharmacology ; Dose-Response Relationship, Drug ; Humans ; Leukocytes, Mononuclear ; cytology ; drug effects ; Lymphoma, B-Cell ; pathology ; Tumor Cells, Cultured