To select the binding-peptide of the cell surface marker CD133 of cancer stem cells from phage peptide library, and to find a new tool for research on stem cells, tumor therapy and anti-metastasis of cancer.

To explore stem cells phenotype of human epidermal growth factor receptor 2 (HER2)-overexpressing breast cancer cells, MDA-MB-453 by isolation of CD44+CD24(-/low) and side population(SP). Analyze the relationship between SP cells and the gene amplification levels of the key proteins, beta-catenin and Notch-1 in Wnt and Notch signal pathways. And study the correlation of SP cells and the gene amplification of HER2 and human epidermal growth factor receptor 3 (HER3).

To explore whether the CD44+/CD24(-/low)/ABCG2(-) (ATP binding cassette superfamily G member 2) cells are associated with prognosis and clinical response in breast cancer patients.

To study the effects of hepatitis B virus (HBV) infection in vitro on differentiation of mesenchymal stem cells (MSCs) to liver cells.

To observe whether murine bone marrow mesenchymal stem cells (MSCs) implantation improves the survival of hepatocellular carcinoma (HCC)-bearing mice,and to investigate whether MSCs can differentiate to hepatocytes in HCC microenvironment in a mouse model of orthotopic HCC and its effects on tumor cells.

Cysteine synthase is a key enzyme for restricting plant cysteine synthesis. Cysteine synthase gene, designated PcCSase1 (GenBank accession no. EU597481), was successfully isolated from Polygonum sibiricum Laxm. by RACE technique. This gene was 1 260 bp in full-length and encoded a peptide of 382 amino acids. Based on bioinformatic analysis, PcCSase1 is a cytoplasm cysteine synthesis and contains a 16 amino-terminal (N-terminal) signal peptide, which led the PcCSase1 to go to the cytoplasm. The results obtained through homologous sequence analysis indicated that PcCSase1 mature protein was highly conserved in plants, which shared approximate 90% in the amino acid sequence. Expression analysis by RT-PCR showed that PcCSase1 gene was expressed in leaf, stem and root with the largest expression in leaf. Under 3% NaHCO3 stress, the largest expression of PcCSase1 gene was detected in leaf, stem and root at the second day following stress. PcCSase1 gene was inserted into pYES2 and transformed into yeast cells (Saccharomyces cerevisiae). The contents of the glutathione in the recombinant yeast and the cysteine in the medium were increased. INVSc1-pYES2-PcCSase1 was more tolerant to salt treatment than INVSc1-pYES2 and the former survival rate was higher than that of the later under the stress of 10% NaHCO3 and 5 mol/L NaCl. These results proved that PcCSase1 gene may confer high salt-tolerance.

Ex vivo expansion of hematopoietic progenitor cells (HPCs) is valuable for clinical application, however, traditional ex vivo culture negatively affects long-term hematopoietic reconstitution ability. In the hematopoietic system, the expression of Notch receptors and their ligands has been widely reported. Active Notch signal inhibits the differentiation of HSCs while promotes their expansion, suggesting that ex vivo expansion of hematopoietic progenitor cells could be enhanced by manipulating Notch signal pathways. In this article the Notch signal pathways, Notch signal and maintenance of hematopoietic progenitor cells, Notch signal and expansion of hematopoietic progenitor cells and molecular mechanism of Notch signal maintaining undifferentiation of hematopoietic progenitor cells were reviewed.

This study was purposed to investigate immune reconstitution at 12 months after allogeneic peripheral blood stem cell transplantation (all-PBSCT) and its relation with the influencing factors such as age, HLA compatibility, graft versus host disease and viral infection. The T lymphocyte subgroups (CD3(+), CD4(+), CD8(+)), B lymphocyte (CD19(+)) and NK (CD16(+)CD56(+)) cells in peripheral blood and serum immunoglobulin concentrations (IgG, IgA and IgM) of 37 patients were analyzed by flow cytometry and scatter turbidimetry, respectively at 1, 3, 6 and 12 months after transplantation. The results showed that CD3(+) cell percentage was (47.5 +/- 23.2)% at 1 month, (75.1 +/- 6.4)% at 3 months, (69.7 +/- 12)% at 6 months and (71.7 +/- 4.2)% at 12 months. CD4(+) cell percentage was (13.3 +/- 6.4)% at 1 month, (20.2 +/- 11.4)% at 3 months, (46.9 +/- 10.3)% at 6 months and (29.1 +/- 18.7)% at 12 months. CD8(+) cell percentage was (43.1 +/- 23.2)% at 1 month, (42.6 +/- 16.9)% at 3 months, (69.7 +/- 12)% at 6 months and (47 +/- 5.6)% at 12 months. CD16(+)56(+) cell percentage was (14.4 +/- 8.4)% at 1 month, (15.9 +/- 7.6)% at 3 months, (14.7 +/- 6.6)% at 6 months and (13.6 +/- 3.4)% at 12 months. CD19(+) cell percentage was (6.4 +/- 5.6)% at 1 month, (11.7 +/- 2.4)% at 3 months, (13.3 +/- 7.3)% at 6 months and (16.7 +/- 5.7)% at 12 months. The serum concentration of IgA was (0.37 +/- 0.14) g/L at 1 month, (0.28 +/- 0.21) g/L at 3 months, (0.42 +/- 0.18) g/L at 6 months and (0.53 +/- 0.34) g/L at 12 months. The serum concentration of IgG was (12.7 +/- 3.8) g/L at 1 month, (16.3 +/- 5.2) g/L at 3 months, (14.3 +/- 6.2) g/L at 6 months and (15.4 +/- 6.9) g/L at 12 months. The serum concentration of IgM was (0.56 +/- 0.24) g/L at 1 month, (0.64 +/- 0.16) g/L at 3 months, (1.1 +/- 0.35) g/L at 6 months and (1.2 +/- 0.28) g/L at 12 months. There were no significant differences between percentage of T lymphocyte subgroups in peripheral blood and serum immunoglobulin concentrations of the patients > or = 45 years old and the patients < 45 years old. The CD19(+) cell percentage of the patients with chronic GVHD at 12 month was less than that of the other ones at 12 months after transplantation. CD4(+) and CD19(+) cell percentage recovery in the patients of haploidentical transplantation was later than that in patients of HLA complete identical transplantation. The CD4(+)/CD8(+) cell ratio and CD4(+) cell percentage of those patients infected with herpes zoster were significantly lower than those without herpes zoster. It is concluded that the CD3(+) cell percentage begins to recover at 3 months after allo-PBSCT. CD4(+) cell percentage begins to recover at 6 months after allo-PBSCT. CD8(+) cell percentage begins to recover at 1 month after allo-PBSCT. B cell percentage recovers at 3 to 6 months after allo-PBSCT. NK cell percentage recovers at 1 to 3 months after allo-PBSCT. The serum concentration of IgG recovers to normal at 1 month after transplantation which is associated with routine infusion of immunoglobulin. The concentration of IgM gradually recovers to normal at 3 months after transplantation. The concentration of IgA does not recover to normal at 12 months after transplantation. The function of B cells recovers slowly in patients with cGVHD. The CD4(+) cell absolute value and CD4(+)/CD8(+) ratio significantly decrease in patients with herpes zoster.

Depletion of T and B cells from the graft is prerequisite for haploidentical transplantation to decrease the risk of GVHD and EBV-associated lymphoproliferative disease. This study was aimed to investigate the performance of T-cell and B-cell simultaneous depletion from mobilized peripheral blood stem cells (PBSCs) for the first time in China, using anti-CD3 and anti-CD19 antibodies conjugated to magnetic microbeads by the CliniMACS device. The depletion efficiency of T-cell and B-cells was analyzed by flow cytometry; the function of the stem cells after depletion was evaluated using colony assays. The results indicated that the mononuclear cell count prior to T- and B-cell depletion was 4.88 x 10(10). After depletion, the percentage of T cells was 0.02% with a log (10) depletion of 4.4. The percentage of B cells was less than 0.01% with a log (10) depletion of at least 3.3. The product contained not only CD34(+) stem cells, but also NK cells, monocytes and granulocytes. After T- and B-cell depletion the purity of CD34(+) cells was 0.98%, the number of CD34 cells was 1.84 x 10(8) and their recovery rate was 69.7%. The number of NK cells was 2.54 x 10(9) and the recovery rate of NK cells was 71.7%. In vitro colony assays showed no negative impact on function of the hematopoietic stem cells. In conclusion, the CliniMACS system can be used to efficiently deplete T and B cells from PBSCs simultaneously, without adverse effect on biological function of hematopoietic stem cells. This study provides technical platform for haploidentical hematopoietic stem cell transplantation.

This study was purposed to investigate the biological characteristics and immunogenicity changes of ex vivo expanded megakaryocyte progenitors from human umbilical cord blood CD34(+) cells in order to provide experimental basis for clinical application of ex vivo expanded umbilical cord blood megakaryocyte progenitor cells. Mononuclear cells (MNCs) were obtained from umbilical cord blood by Ficoll-Hyapaque density gradient separation. CD34(+) cells were enriched by magnetic cell sorting (MACS). The selected CD34(+) cells were seeded in serum-free medium stimulated with thrombopoietin (TPO, 50 ng/ml), interleukin 11 (IL-11, 50 ng/ml), and heparin (25 U/ml) for 14 days. The immunophenotyping (CD34(+), CD41a(+), CD61(+), CD34(+) CD41a(+) and CD34(+) CD61(+) cells) of amplificated products, matured megakaryocyte apoptosis, and expression of human leukocyte antigen (HLA) class I and class II molecules were measured by fluorescence-activated cell sorter (FACS). The number of colony-forming units-megakaryocyte (CFU-Mk) was also evaluated by CFU-Mk assay. The results showed that the umbilical cord blood CD34(+) mononuclear cells could be effectively differentiated into megakaryocytes. The peak expression ratios of CD41a(+) and CD61(+) cells were all at 14th days, while that of CD34(+) CD41(+) and CD34(+) CD61(+) cells were at 7th day [(3.41 +/- 2.80)% and (1.89 +/- 1.43)%, respectively]. The expansion times of large and small CFU-Mk reached peak at 7th day (20.66 +/- 32.79) and 10th day (435.62 +/- 482.65), respectively. The apoptotic rates of megakaryocytes at 7th, 10th, 14th day were (19.48 +/- 9.64)%, (26.87 +/- 9.03)%, and (52.46 +/- 11.74)%, respectively. The apoptotic rate of megakaryocytes had no significant difference in 7 and 10 days culture (p > 0.05), while that significantly increased in culture for 14 day culture, compared with culture for 7 and 10 days (p < 0.05, respectively). The expression of HLA class I and class II molecules on megakaryocytes decreased along with the prolongation of expansion time and sharply decreased in 0 to 10 days. It is concluded that the cytokines of TPO, IL-11, and heparin can promote the expansion of megakaryocyte progenitors from umbilical cord blood CD34(+) mononuclear cells effectively in vitro. The peaked expansion times of large CFU-Mk, the peaked expression ratios of CD34(+) CD41(+) and CD34(+) CD61(+) cells were all at 7th day. So the culture for 7 days appeared to be the optimal duration of expanding megakaryocyte progenitors.

This study was purposed to explore the immunoregulatory effects of human bone marrow mesenchymal stem cells (MSCs) on active T lymphocytes in vitro and the new strategy to prevent graft-versus-host disease (GVHD) in allogeneic hematopoietic stem cell transplantation (allo-HSCT). Mononuclear cells from human peripheral blood cells were isolated and cultured in the presence of phytohemagglutinin (PHA) (final concentration was 10 microg/ml) for different times. The ability of T lymphocyte proliferation and activation was measured by (3)H-Thyramine incorporation. The expressions of CD3(+)CD4(+), CD3(+)CD8(+), CD4(+)CD25(+) and CD4(+)CD152(+) on T cells were detected by FCM after coculture for 72 hours. Experiment was divided into 4 groups: A group as control (no added MSCs), B group (actived T cells + 2 x 10(4) MSCs), C group (actived T cells + 4 x 10(4) MSCs), D group (actived T cells + 8 x 10(4) MSCs). The results showed that the ability of T lymphocyte proliferation in the same PHA concentration increased with prolonging of time. ability of T lymphocyte proliferation was strongest when culturing for 48 hours (p < 0.01); the expressions of CD44, CD105, CD29 and FIK1 of MSCs were positive, expressions of CD33, CD34, CD45 and HLA-DR were negative. MSCs inhibited T lymphocyte proliferation and the inhibitory effect depended on the amount of MSCs. CD3(+)CD8(+), CD4(+)CD25(+) and CD4(+)CD152(+) T cells cocultured with MSCs increased obviously and CD3(+)CD4(+) expression significantly decreased, as compared with control group (p < 0.01). It is concluded that the MSCs inhibit T lymphocyte proliferation induced by mitogen (PHA), and perform their immunosuppressive function by up-regulation of CD3(+)CD8(+), CD4(+)CD25(+) and CD4(+)CD152(+) expressions and down-regulation of CD3(+)CD4(+) expression.

The study was purposed to investigate the differentiation ability of mesenchymal stem cells (MSCs) into myocardial cells in vitro. Rat bone marrow-derived MSCs were labeled and co-cultured with neonatal rat cardiomyocytes (CM) for 5 - 7 days. The expression of cell surface antigens was detected by flow cytometry, and the expression of muscle-specific marker myosin and troponin T in labeled cells was detected by immunofluorescence. The results showed that in vitro cultured MSCs expressed CD90, CD44, CD105, CD54, not expressed CD34, CD45, CD31. After co-cultured with neonatal rat CM, labeled MSCs differentiated into cardiomyocyte-like cells expressing myosin and troponin T. It is concluded that MSCs can differentiate into cardiomyocyte-like cells when co-cultured with neonatal myocardial cells in vitro. In co-culture of two kind of cells in ratio of four to one showed obvious efficacy differentiating MSCs into CMs.

The objective of this study was to investigate the expressions of hematopoietic cytokines EPO, SCF and GM-CSF in bone marrow cells of patients with chronic aplastic anemia (CAA) and their significance. The mRNA expressions of EPO, SCF and GM-CSF from 35 CAA patients and 10 healthy individuals were detected by semi-quantitative RT-PCR. The results showed that the levels of EPO, SCF and GM-CSF in CAA patients were obviously lower than those in the healthy individuals (p < 0.01). It is concluded that the immunity disorder mediated by hematopoietic cytokines is one of the pathogenesis in the CAA patients, it provided theoretical basis for guiding clinical treatment.

This study was aimed to explore the effect of bortezomib on proliferation and apoptosis of acute monocytic leukemic cells SHI-1 and the function of Bcl-2 gene family including Bcl2l12, Bcl-2 and Bax in its apoptosis. SHI-1 cells were cultured and treated with bortezomib of different concentrations for different time. MTT assay was used to detect the proliferation and apoptosis, Annexin-V staining, mitochondrial transmembrane potential (DeltaPsim) and DNA aga-rose gel electrophoresis were used to investigate apoptosis of SHI-1 cells. RT-PCR was used to analyze the levels of Bcl2l12, Bcl-2 and bax mRNA in SHI-1 cells treated with bortezomib for 0, 6, 12 and 24 hours. The results showed that bortezomib inhibited the proliferation of SHI-1 cells in time-and doze-dependent manners, the IC(50) at 24 and 48 hours were 54.13 nmol/L and 5.45 nmol/L respectively. Bortezomib could induce apoptosis of SHI-1 cells in time-dependent manner, increase expression of Annexin-V positive cells, decrease DeltaPsim of SHI-1 cells and result in DNA fragmentation and morphologic changes of apoptosis. RT-PCR showed that Bcl2l12 mRNA expression was up-regulated, bcl-2 mRNA expression was down-regulated and bax mRNA expression was not changed obviously. It is concluded that bortezomib inhibits the proliferation of SHI-1 and induces apoptosis in which Bcl2l12 and Bcl-2 gene can be ones of the main genes taking part in.

Recently, eloquent studies from some solid tumors have provided proofs that cancers originate from cancer stem cells (CSC). The discovery of CSC has changed our view of carcinogenesis and chemotherapy. The aim of this study is to identify and characterize the CSC population that drives and maintains lung adenocarcinoma growth and metastasis.

To explore the mechanism of asthma, by evaluating the changes of CCR3 and EOS expression in the lung tissues and bone marrow of Guinea pig asthmatic models at different times.

To investigate the effects of adipose-derived stem cell (ADSC) transplantation on the angiogenesis in the brain post focal cerebral ischemia in rats.

To study the hematopoietic reconstitution ability of fresh and cultured cord blood CD34(+) cells by sublethally irradiated NOD/SCID mice.

To explore the effects of graft composition on hematopoietic reconstitution and graft-versus-host disease (GVHD) in HLA-identical related donor peripheral blood stem cell transplantation (PBSCT) for hematological malignancies.

To evaluate the prognostic implication of the killer-immunoglobulin like receptor (KIR) ligand mismatch in HLA mismatched hematopoietic stem cell transplantation (HSCT).