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

The aim of this study was to assess the involvement of multipotential progenitor cells in the pathogenesis of Mooren's ulcer using immunohistochemical staining techniques. Tissue specimens were collected from 3 Mooren's ulcer patients who underwent lamellar keratectomy. Immunohistochemical staining patterns were analyzed using antibodies: CD34, c-kit, STRO-1, CD45RO, VEGF and alpha-SMA. Strong positive CD34, c-kit and STRO-1 cells were revealed in Mooren's ulcer specimens, especially in the superficial stroma. A few weakly expressed CD34 stroma cells were seen in normal limbal cornea but no immunoreactivity for c-kit and STRO-1 could be found. CD45RO positive T cells were found to have infiltrated in Mooren's ulcer. The immunostaining pattern of VEGF and yen a- SMA was closely correlated with the degree of expression and the number of CD34 positive cells. Bone marrow-derived multipotential progenitor cells may be involved in the pathogenesis of Mooren's ulcer by synergizing with other factors to amplify autoimmune destructive reactions and to contribute to the regeneration process. Specific therapeutic strategies that target the role of these cells in the disease are warranted.
Cornea/*pathology ; Corneal Ulcer/*pathology ; Hematopoietic Stem Cells/*pathology ; Humans ; Multipotent Stem Cells/*pathology ; Research Support, Non-U.S. Gov't

OBJECTIVESevere newborn hypoxic-ischemic encephalopathy (HIE) has a very high rate of disability and no effective treatment is available. The present study aimed to preliminarily evaluate the effects of human neural stem cell transplantation in treatment of severe neonatal HIE.

METHODSThe patient was a 75-day old male infant with sequelae of severe HIE who had highly delayed development of intelligence and movement and myotonia. MRI showed multiple cerebromalacia and encephalatrophy. Cells obtained from the forebrain of an 11-week old fetus were cultured and amplified for 15 days. And then the human fetal neural stem cells were injected into cerebral ventricle of this infant.

RESULTSTwenty eight days after transplantation, remarkable improvement occurred not only in his myotonia but also in his intelligence and movement, which became similar to those of the normal infants of the same age. Positron emission tomography (PET) showed significantly increased radioactivity at temporal and occipital lobes which suggested that the cellular metabolism had increased greatly.

CONCLUSIONThe short-term effect of NSCs transplantation on the infant with severe HIE sequelae was significant. PET suggested that the implanted NSCs survived. Many more studies are needed to evaluate long-term effects of NSC transplantation in treatment of HIE.

Asphyxia Neonatorum ; complications ; Brain ; pathology ; physiopathology ; Female ; Humans ; Hypoxia-Ischemia, Brain ; etiology ; pathology ; physiopathology ; therapy ; Infant ; Infant, Newborn ; Injections, Intraventricular ; Multipotent Stem Cells ; transplantation ; Neurons ; Positron-Emission Tomography ; Prognosis ; Stem Cell Transplantation ; methods ; Time Factors ; Treatment Outcome

OBJECTIVETo establish a neonatal rat model of hypoxic-ischemic encephalopathy and clarify the changing features of neural stem cells (NSCs) in episodes of hypoxic-ischemic encephalopathy (HIE) so as to provide experimental and theoretical evidences for treating HIE by applying NSCs at the appropraite time.

METHODSTotally 210 neonatal rats aged 7 d were divided randomly into three groups, normal control group, hypoxic group and hypoxic-ischemic group with 70 rats in each. According to the time of sacrefice, 70 rats of every group were further divided randomly into seven groups including third hour (3 h), the sixth hour (6 h), first day (1 d), third day (3 d), the seventh day (7 d), the fourteenth day (14 d) and the twenty-first day (21 d), with 10 rats in each subgroup. The left common carotid artery of the neonatal rats in hypoxic-ischemic group was ligated and those in the hypoxic group were subjected to inhalation of 8% oxygen for 2.5 h. NSCs from brain tissues of the rats of the three groups were determined with HE staining and immunohistochemical method under light microscope.

RESULTSMost of neonatal rats in hypoxic-ischemic group behaved turning to the left stably 1 h after normal concentration of oxygen was given. In hypoxic-ischemic group, slight brain injury occurred at 3 h, severe brain injury appeared at 1 d, glial cells proliferated at 3 d and 7 d, brain atrophy was found at 14 d and 21 d. NSCs existed in brain tissues of rats in all the three groups. NSCs in normal control and the hypoxic group mainly distributed in hippocampus, subventricular tissues, striatum and cortex. But NSCs in hippocampus located in layers of molecule, cone cell and inner granular cell. NSCs in hypoxic-ischemic group showed obvious regional distribution, less in the regions with pathological changes. At 3 h, 6 h and 14 d, there was no difference in the number of NSCs between hypoxi and hypoxic-ischemic group. At 1 d, 3 d and 7 d, there was a highly significant difference in the number of NSCs between hypoxic and hypoxic-ischemic group. At 21 d, there was a significant difference in the number of NSCs between hypoxic and hypoxic-ischemic group, meanwhile, there was a significant difference in the number of NSCs between control and hypoxic group. At 3 d, there was a very significant difference in the number of NSCs between control and hypoxic-ischemic group. At 7 d and 21 d points, there was a highly significant difference in the number of NSCs between control and hypoxic group.

CONCLUSIONThe neonatal rat model of HIE was successfully established. NSCs increased in earlier period and decreased in later period of HIE, ultimately, NSCs located in the injured regions died one after anotner. Hypoxia induces NSCs' proliferation.

Animals ; Animals, Newborn ; Atrophy ; Brain ; pathology ; Carotid Artery, Common ; surgery ; Disease Models, Animal ; Hypoxia-Ischemia, Brain ; pathology ; Immunohistochemistry ; Ligation ; Multipotent Stem Cells ; pathology ; Neuroglia ; pathology ; Neurons ; pathology ; Rats ; Rats, Sprague-Dawley ; Stem Cell Transplantation ; methods ; Time Factors

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