OBJECTIVETo observe whether ginsenoside Rg1 could reduce the infarcted area and improve the heart function by path of promoting bone marrow stem cells differentiated to vascular endothelial cells (VECs).

METHODSBone marrow was drawn from rabbit's ilium and labelled with red fluorochrome DiI, then it was transferred again into the rabbit's body. The rabbits was then made into myocardiac infarction model. The model rabbits were divided into the control group and the ginsenoside Rgl treated group (treated group). The infracted area at two weeks, and the left ventricular function at one and two weeks after infarction were determined respectively. The DiI positive cell rate of myelogenetic cells in ischmia area and CD31 positive cell rate of VECs were determined by confocal microscopy. Myocardial interstitial granulocyte colony-stimulating factor(GCSF) levels during ischemia and reperfusion period were determined also.

RESULTSDiI positive rate of CD31 staining positive cells in the treated group was obviously increased, and the concentration of G-CSF in myocardium interstitial obviously increased, accompanied with obviously improving of heart function and obviously reducing of infarcted area.

CONCLUSIONGinsenoside Rgl could stimulate the G-CSF secretion in local myocardiac tissues, thus to induce bone marrow mononuclear cells migrate to myocadial tissue and further differentiate to VECs. The regeneration of endothelium cells show certain direct action in promoting capillary regeneration of infarcted myocardium tissue and maintaining the blood supply.

Animals ; Bone Marrow Cells ; cytology ; Cell Differentiation ; drug effects ; Cell Movement ; drug effects ; Coronary Circulation ; drug effects ; Endothelial Cells ; cytology ; Ginsenosides ; pharmacology ; Granulocyte Colony-Stimulating Factor ; biosynthesis ; Male ; Multipotent Stem Cells ; cytology ; Myocardial Infarction ; metabolism ; pathology ; Rabbits

OBJECTIVECytomegalovirus (CMV) infection was greatly common in the world. CMV infection produces usually mild or asymptomatic infections in individuals with normal immune responses, whereas it may cause serious disease in immunosuppressive patients. Clinical manifestations include suppression of myelopoiesis, a mononucleosis like syndrome, hepatosplenomegaly, lymphadenopathy, thrombocytopenia, and hemolytic anemia. In patients undergoing bone marrow transplantation CMV remains the most common infectious causes of morbidity and mortality. But the treatment drugs with specific effect for CMV was fewer at the present. This study was to investigate the effect of CMV on proliferation of colony forming unit granulocyte-macrophage (CFU-GM), CFU-erythroid (CFU-E), brust forming unit-erythroid (BFU-E), CFU-multipotential (CFU-Mix) and CFU-megakaryocyte (CFU-Mk) progenitor cells of cord blood (CB) with the presence of ganciclovir (GCV) and astragalus membranaceus in vitro.

METHODSTwenty CB samples were collected from fetal umbilical vein of normal term spontaneous delivery neonates. The colony forming unit-assay was applied to observe the suppression effect of CMV-AD169 strain on CFU-GM, CFU-E, BFU-E, CFU-Mix and CFU-Mk of CB with the presence of GCV and astragalus membranaceus in vitro. The technique of PCR was used to demonstrate the existence of CMV-AD169 DNA in the colony cells of cultured CFU-GM, CFU-E, BFU-E, CFU-Mix and CFU-Mk.

RESULTS(1) The numbers of CFU-GM, CFU-E, BFU-E, CFU-Mix and CFU-Mk colonies in CMV infection groups were significantly less than those in blank and mock group, respectively. The last time of colonies in groups with CMV infection was significantly shorten compared with the blank and mock group. (2) CMV-DNA was positively detected in the colony cells of CMV infection groups by PCR, while negative in the control groups. (3) The lasting time of CFU-GM, CFU-E, BFU-E, CFU-Mix and CFU-Mk colonies infected with CMV extended significantly with the presence of astragalus membranaceus and GCV, and the numbers of those increased significantly compared with the CMV infection group, respectively. The increasing rate of colonies was 27.2%, 45.2%, 49.1%, 39.0% and 11.9% with astragalus membranaceus group, 37.4%, 74.2%, 71.7%, 67.4% and 38.9% with GCV group, 53.6%, 83.8%, 88.7%, 87.8% and 61.5% with astragalus membranaceus and GCV group, respectively.

CONCLUSIONSThe differentiation and proliferation of CFU-GM, CFU-E, BFU-E, CFU-Mix and CFU-Mk were significantly inhibited after infected with CMV-AD169 strain. The suppression effect of CMV-AD169 on CFU-GM, CFU-E, BFU-E, CFU-Mix and CFU-Mk was inhibited with the presence of GCV and astragalus membranaceus in vitro. This suggested that CMV-AD169 may be inhibited or killed by GCV and Astragalus Membranaceus in vitro.

Antiviral Agents ; pharmacology ; Astragalus membranaceus ; chemistry ; Cell Division ; drug effects ; Cytomegalovirus ; drug effects ; Cytomegalovirus Infections ; drug therapy ; Drugs, Chinese Herbal ; pharmacology ; Erythroid Precursor Cells ; cytology ; drug effects ; metabolism ; Fetal Blood ; cytology ; drug effects ; metabolism ; Ganciclovir ; pharmacology ; Hematopoietic Stem Cells ; cytology ; drug effects ; metabolism ; Humans ; Multipotent Stem Cells ; cytology ; drug effects ; metabolism

Human mesenchymal stem cells (MSCs) have emerged as attractive cellular vehicles to deliver therapeutic genes for ex-vivo therapy of diverse diseases; this is, in part, because they have the capability to migrate into tumor or lesion sites. Previously, we showed that MSCs could be utilized to deliver a bacterial cytosine deaminase (CD) suicide gene to brain tumors. Here we assessed whether transduction with a retroviral vector encoding CD gene altered the stem cell property of MSCs. MSCs were transduced at passage 1 and cultivated up to passage 11. We found that proliferation and differentiation potentials, chromosomal stability and surface antigenicity of MSCs were not altered by retroviral transduction. The results indicate that retroviral vectors can be safely utilized for delivery of suicide genes to MSCs for ex-vivo therapy. We also found that a single retroviral transduction was sufficient for sustainable expression up to passage 10. The persistent expression of the transduced gene indicates that transduced MSCs provide a tractable and manageable approach for potential use in allogeneic transplantation.
Adolescent ; Animals ; Cell Death/drug effects ; Cell Line, Tumor ; Cell Proliferation/drug effects ; Cell Transformation, Neoplastic/drug effects/pathology ; Child ; Cytosine Deaminase/*genetics/therapeutic use ; Fluorouracil/pharmacology ; Genetic Therapy ; Genomic Instability/drug effects ; Humans ; Karyotype ; Mesenchymal Stromal Cells/*cytology/drug effects/metabolism ; Mice ; Multipotent Stem Cells/cytology/drug effects/metabolism ; Neoplasms/therapy ; Retroviridae/*metabolism ; Time Factors ; *Transduction, Genetic

OBJECTIVE: To explore the differentiation potential of QY1 bone marrow mesenchymal stem cell (MSCs) line cells into cardiacmyocytes and vascular endothelial cells in vitro, to optimize the suitable conditions of MSCs differentiating into cardiomyocytes in vitro, and to examine the potentials of MSCs differentiating into cardiomyogenesis and vasculogenesis. METHODS: Specifically committed differentiation inductive medium was employed, including 5-azacytidine for cardiomyogenesis and vascular endothelial growth factor for vasculogenesis in culture respectively in vitro. The differentiated cells were identified by immunohistochemistry and molecular biology. RESULTS: MSCs line cells had been cultured in the normal culture medium for 72 hours, then the differentiation inductive medium including 10 micromol/L 5-azacytidine was added into the normal culture dishes for 24 hours only. After that the culture medium was changed back to the normal culture medium. Normal culture medium was changed every 7 days. The second induction was performed after 14 days. The differentiated cells treated with 5-azacytidine could beat spontaneously and formed myotube structures in the optimal induction conditions, and the differentiation rate was (39.47+/-0.56)%. The differentiated cells expressed specific cardiomyocytic proteins identified by the positive immunohistochemistry staining with anti-alpha-sarcomeric antibody and anti-Cx-43 antibody, and also expressed the alpha-myosin heavy chain examined by RT-PCR. The differentiated cells began to appear as the lined up vascular endothelial cells after 48 hour treatment with vascular endothelial growth factor. Some of the differentiated cells connected each other to form vascular endothelial web-like structure after 7 day treatment with vascular endothelial growth factor. On 14 d after treating with vascular endothelial growth factor, the differentiated cells were identified by immunohistochemistry staining. The expressions of both specific surface antibody CD31 and factor VIII for vascular endothelial cells were positive. CONCLUSION The cells of QY1 bone marrow mesenchymal stem cell line may differentiate into cardiomyocytes or vascular endothelial cells in vitro under specific condition.
Azacitidine ; pharmacology ; Bone Marrow Cells ; cytology ; Cell Differentiation ; drug effects ; physiology ; Cell Line ; Dose-Response Relationship, Drug ; Endothelial Cells ; cytology ; metabolism ; Factor VIII ; biosynthesis ; Humans ; Immunohistochemistry ; Mesenchymal Stem Cells ; cytology ; Multipotent Stem Cells ; cytology ; Myocytes, Cardiac ; cytology ; Platelet Endothelial Cell Adhesion Molecule-1 ; biosynthesis

The objective of present study is to investigate the effect of human cytomegalovirus (HCMV) infection on human hippocampus neural stem cells NSCs differentiation in vitro, Fetal hippocampus tissue was dissociated mechanically and then cultured in proliferation medium with EGF and bFGF. Immunofluorescence method was used to detect the expression of NSCs marker-Nestin within these cells. Cultured in 10% FBS, NSCs began to differentiate. On the onset of the differentiation, HCMV AD169 (MOI=5) was added into the differentiation medium. After 7 days differentiation, the effect of HCMV infection on NSCs differentiation was observed by detecting the rate of nestin, GFAP and HCMV immediate-early (IE) positive cells with confocal microscopy and immunofluorescence method. The resucts showed most of the cells (passage 4-6 ) were Nestin positive and could differentiate into NSE-positive neurons and GFAP-positive astrocytes. On day 7 postinfection, 86% +/- 12% of infected cells were IE positive. The percentage of Nestin-positive cells was 50% +/- 19% and 93% +/- 10% (t= 6.03, P<0.01)and those of GFAP-positive cells was 81% +/- 11% and 55 +/- 17% (t=3.77, P<0.01) in uninfected and infected cells respectively. These findings indicated that NSCs were HCMV permissive cell and HCMV AD 169 infection suppressed the differentiation of Hippocampus-genetic human neural stem cells into astrocytes.
Astrocytes ; cytology ; Cell Differentiation ; drug effects ; Cells, Cultured ; Cytomegalovirus ; growth & development ; physiology ; Epidermal Growth Factor ; pharmacology ; Fibroblast Growth Factor 2 ; pharmacology ; Hippocampus ; cytology ; Humans ; Intermediate Filament Proteins ; metabolism ; Microscopy, Fluorescence ; Multipotent Stem Cells ; cytology ; drug effects ; metabolism ; virology ; Nerve Tissue Proteins ; metabolism ; Nestin ; Neurons ; cytology

Neural stem cells (NSCs) have mainly been applied to neurodegeneration in some medically intractable neurologic diseases. In this study, we established a novel NSC line and investigated the cytotoxic responses of NSCs to exogenous neurotoxicants, glutamates and reactive oxygen species (ROS). A multipotent NSC line, B2A1 cells, was established from long-term primary cultures of oligodendrocyte-enriched cells from an adult BALB/c mouse brain. B2A1 cells could be differentiated into neuronal, astrocytic and oligodendroglial lineages. The cells also expressed genotypic mRNA messages for both neural progenitor cells and differentiated neuronoglial cells. B2A1 cells treated with hydrogen peroxide and L-buthionine-(S,R)-sulfoximine underwent 30-40% cell death, while B2A1 cells treated with glutamate and kainate showed 25-35% cell death. Cytopathologic changes consisting of swollen cell bodies, loss of cytoplasmic processes, and nuclear chromatin disintegration, developed after exposure to both ROS and excitotoxic chemicals. These results suggest that B2A1 cells may be useful in the study of NSC biology and may constitute an effective neurotoxicity screening system for ROS and excitotoxic chemicals.
Animals ; Brain/*cytology ; Buthionine Sulfoximine/pharmacology ; Cell Differentiation ; Cell Line ; Cell Lineage ; Cytokines/pharmacology ; Enzyme Inhibitors/pharmacology ; Excitatory Amino Acid Agonists/pharmacology ; Glutamic Acid/pharmacology ; Humans ; Hydrogen Peroxide/pharmacology ; Intercellular Signaling Peptides and Proteins/pharmacology ; Kainic Acid/pharmacology ; Mice ; Mice, Inbred BALB C ; Multipotent Stem Cells/cytology/*drug effects/physiology ; Neuroglia/cytology/drug effects/physiology ; Neurons/cytology/*drug effects/physiology ; Neurotoxins/*pharmacology ; Oxidants/pharmacology ; Phenotype ; Reactive Oxygen Species/metabolism