Since flow cytometry was not feasible for sorting a huge amount of cells for clinical use, the method of double immunomagnetic positive sorting was used for selection of CD34(+)CD59(+) cells from bone marrow mononuclear cells in patients with paroxysmal nocturnal hemoglobinuria (PNH), which laid the groundwork for clinical ABMT/APBSCT of patients with PNH. Immunomagnetic positive selection was used for two times, the microbeads were removed from the CD34(+) cells selected firstly by means of overnight culture, then the sufficient CD34(+)CD59(+) cells were used for ex vivo expansion. The results showed that the survival, proliferation and colony-forming units of the selected CD34(+)CD59(+) cells by double immunomagnetic positive sorting had no significant difference as compared with that of CD34(+)CD59(+) cells selected by flow cytometry technique. It is suggested that the double immunomagnetic positive sorting promotes the use for separation and purification hematopoietic stem/progenitor cells and other cells with double or multiple markers cells for autologous hematopoietic stem cell transplantation in PNH patients.

The object of this study was to explore the effects of Panax notoginosides (PNS) on proliferation and differentiation of human CD34(+) stem/progenitor cells. CD34(+) cells were isolated from human bone marrow by using immune beads of Dynal M- 450 system. The cells were exposed to PNS at different concentrations in both liquid and semi-solid culture for 14 days. The cells were marked with monoclonal antibodies and analyzed by flow cytometry after culture. The CFU-Mix colony formation from CD34(+) cells was assayed. The results showed that: (1) The yield of CD34(+) cells after being selected by immune beads were (1.03 +/- 0.74)% out of bone marrow nuclear cells with purity of 86% - 93%. (2) PNS (10 - 25 mg/L) stimulated the proliferation of CD34(+) cells, and raised the colony numbers of CFU-Mix obviously in vitro. PNS 25 mg/L was the optimal concentration to promote proliferation of CD34(+) cells, the increasing rate of CFU-Mix colony was (34.7 +/- 16.0)%. (3) The differentiation of CD34(+) cells was induced by exposure to PNS (25, 50 and 100 mg/L) in liquid culture for 14 days. The percentages of CD33(+) and CD15(+) cells were increased after PNS exposure, which were significantly higher than those of control (P < 0.01), however CD71(+) and G-A(+) cells were no obviously difference after PNS treatment. In conclusion, Panax notoginosides not only promote the proliferation of CD34(+) cells, but also induce the differentiation committed to granulocytes.

To study the biological role of human cultured bone marrow mesenchymal stem cell (BM-MSC) in hematopoiesis by investigation of its expression of multiple hematopoietic growth factors, RT-PCR was used to analyze the expression of SCF, Flt3-ligand, TPO, LIF, G-CSF, GM-CSF, IL-3, IL-6 and IL-11 at mRNA level for human BM-MSC from healthy donors and patients with leukemia and lymphoma. BM-MSC were incubated with or without hydrocortison (HC). The results clearly showed that the cultured BM-MSC expressed mRNA of SCF, Flt3-ligand, TPO, LIF, IL-6 and IL-11 at passages 3 up to 15, but did not express G-CSF, GM-CSF and IL-3. The same expression pattern of above cytokines was seen also for the patient's BM-MSC. HC was able to induce BM-MSC to express G-CSF but not to express GM-CSF. BM-MSC seemed not to change morphologically after incubation with HC for up to 21 days. In conclusion, both normal and patient BM-MSC should be potential to promote hematopoiesis according to their expression of multiple hematopoietic cytokines, and HC is able to induce hematopoietic growth factor expression.

The purpose of this study was to observe the bone marrow endothelial cell-conditioned medium (BECM) and cytokines, i.e. vascular endothelial growth factor (VEGF), stem cell factor (SCF) and EPO promoting the generation of hematopoietic precursor cells from mouse embryonic stem cells (ESC) in vitro. Day 4 embryoid body (4dEB) cells were derived from ESC-D3 cell line, a murine ESC line, and then induced with BECM and/or cytokines. Four groups, i.e. BECM, BECM + VEGF + SCF + EPO, VEGF + SCF + EPO and control (spontaneous differentiation), were designed. Immunochemistry staining and flow cytometry were adopted to observe the antigen expression, RT-PCR to detect hematopoietic transcription factors, and hematopoietic progenitor assay to examine hematopoietic differentiation. The results showed that the cells induced from ESC expressed hematopoietic precursor cell antigens (c-kit, Sca-1, Thy-1 and CD34), transcription factors (c-myb, SCL and beta-H1) and generated HPP-CFC and BFU-E. The effect of BECM + VEGF + SCF + EPO was the most potent in the inducing groups according to the numbers of hematopoietic precursor cells and colonies. It is concluded that BECM promotes the differentiation of ESC into hematopoietic precursor cells in vitro, and this effect is the strongest when BECM combining with VEGF + SCF + EPO.

To isolate MSCs from adult human bone marrow cells and to induce them into adipocytes.

To investigate the effects of endothelial cells on the proliferations of granulocytic, erythrocytic and megakaryocytic progenitor cells.

To study the effect of PF4 and relative peptide PF4 17-70 on the chemoattract ability, the expression of adhesion molecules and CXCR4 on the flesh cord blood CD34+ cells.

To observe the influence of tumor necrosis factor-alpha (TNF-alpha) on differentiation of rat mesencephalic neural stem cells (NSCs), the numbers of neurons, astrocytes and oligodendrocytes generated from NSCs were analyzed after differentiation for 3 days by using immunocytochemistry technique. The results show that: (1) TNF-alpha enhanced the proportions of neurons and oligodendrocytes in progeny of NSCs; and (2) TNF-alpha induced the proliferation of oligodendrocytes derived from NSCs, but the proliferation of astrocytes was not influenced by TNF-alpha. We conclude that the TNF-alpha could influence the application of NSCs.

Bone marrow mesenchymal stem cells (MSCs) are multipotent tissue stem cells that can be induced in vitro to differentiate into a variety of cells such as osteoblasts, chondrocytes and adipocytes. MSCs are useful vehicles for both cell and gene therapy for a variety of diseases. Here, we injected human MSCs with enhanced green fluorescent protein (EGFP) into the striatum of Parkinson disease (PD) rat and examined their survival, migration, differentiation, and the behavior changes in PD rats, which will provide a theoretical foundation and technical method for clinic PD therapy by stem cells. The results showed that human bone marrow MSCs can survive in rat brain for a long time (exceeding 70 d). MSCs were found in multiple areas of the rat brain including the striatum, the corpus callosum, contralateral cortex and even the brain vascular wall. Immunocytochemical staining suggested that implanted cells expressed human neurofilament (NF), neuron-specific enolase (NSE) and glial fibrillary acid protein (GFAP). At the same time, remission in abnormal behavior of the PD rats appeared. Rotation scores decreased gradually from 8.86+/-2.09 r/min pre-transplantation to 4.87+/-2.06 r/min 90 d post-transplantation (statistic result showed P<0.05).

Breast cancer resistance protein (BCRP) is a new multidrug resistance-related transmembrane transporter. BCRP is a 655-amino acid, 72.6 kDa protein, localized in the plasma membrane. As a member of the ATP-binding cassette family of drug transporters, BCRP has only one ATP-binding cassette and six putative transmembrane domains, suggesting that BCRP is a half-transporter, which may function as a homodimer or heterodimer. The BCRP-overexpressing tumor cells are resistant to mitoxantrone, adriamycin, daunorubicin, etoposide, topotecan and irinotecan, but lack resistance to paclitaxel and vincristine. Fumitremorgin C and GF120918 can effectively reverse multidrug resistance in BCRP-overexpressing tumor cells, associated with an increase in drug accumulation. In normal human tissues, low to high expressions of BCRP in placental syncytiotrophoblasts, in the epithelium of the small intestine and colon, in the liver canalicular membrane, in ducts of the breast, in endothelium of the blood vessel and in stem cells were reported. This expression profile allows speculation on a role of BCRP in protection of the fetus and in the regulation of transport of chemicals through the epithelium of the gastrointestinal tract. BCRP can account for chemoresistance of some clinical cancers such as acute myeloid leukemia, non-small cell lung cancer,and breast cancer.

To analyze the incidence and the effective prevention and treatment of graft-versus-host disease (GVHD) for the allogeneic peripheral blood stem cell transplantation (allo-PBSCT) for the treatment of leukemia.

To explore the significance of nonmyeloablative allogeneic peripheral blood hematopoietic stem cell transplantation in the treatment of hematological diseases.

To explore the ex vivo expansion characteristics of selected CD(34)(+) cells and mononuclear cells (MNC).

To investigate the clinical value of glycosylated G-CSF combined with middle-high dose cyclophosphamide (Cy) or conventional chemotherapy with increased dose of Cy for mobilizing peripheral blood progenitor cells in patients with tumor.

To study the effect of ex vivo expansion on the adhesion activities of umbilical cord blood hematopoietic stem and progenitor cells (HSPC).

To monitor the variety of lymphocytes and CD34(+) cell subsets in peripheral blood (PB) and peripheral blood progenitor cell (PBPC) during mobilization and collection, and determine the optimal time for PBPC collection.

To establish a strain of human cervical carcinoma cell line and to provide a cervical carcinoma animal model.

To investigate the feasibility of differentiation of bone marrow mesenchymal stem cells (MSCs) into vascular endothelial cells and the mechanism of its involvement in wound healing.