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05 February 2010, Volume 14 Issue 6 Previous Issue    Next Issue

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Isolation, purification and osteoinduction differentiation of canine bone marrow mesenchymal stem cells: Feasibility of in vitro isolation using Ficoll density gradient centrifugation Xie Fang, Teng Li, Cai Lei, Xu Jia-jie, Jin Xiao-lei, Xiao Ran, Cao Yi-lin 2010, 14 (6):  951-956.  doi: 10.3969/j.issn.1673-8225.2010.06.001 Abstract ( 315 )   PDF (723KB) ( 541 )   Save BACKGROUND: Classic isolation method of bone marrow mesenchymal stem cells (BMSCs) is Percoll density gradient centrifugation. Blood cell component was removed. However, this method is complicated. Preparation density was needed when isolating dog bone marrow. Moreover, centrifugation was frequent, which had a great damage to cells. OBJECTIVE: To establish methods of the isolation, proliferation, culture and osteoinduction of canine BMSCs, and observe the in vitro proliferation and ability to osteoinduction differentiation. METHODS: 10 mL bone marrow was extracted from dog posterior superior iliac spine, heparin anticoagulation, diluted using Hanks juice, treated with 1.077 g/mL Ficoll solution 3 mL, and centrifuged at 2 000 r/min for 20 minutes. Karyocytes were absorbed to form white cloudlike layering interface, and then centrifuged twice using DMEM supplemented with fetal bovine serum, incubated at 12×104/cm2 at 37 ℃ in a 5% CO2 incubator. Following subculture, cells were incubated in DMEM containing dexamethasone, β-sodium phosphoglycerol and ascorbic acid 2-phosphate. Immunocytochemical staining and immunofluorescence staining were utilized to detect osteocalcin, osteopontin and type Ⅰ collagen expression in osteoblasts. Alkaline phosphatase staining and alizarin red staining were performed. RESULTS AND CONCLUSION: 1.077 g/mL Ficoll density gradient centrifugation was used to isolate karyocytes that were significant compared with Percoll solution. Obtained BMSCs had high purity, good growth and the mean doubling time was 24 hours. Following in vitro osteogenic incubation of dog BMSCs, osteocalcin, osteopontin and type Ⅰ collagen showed positive expression. Alkaline phosphatase staining demonstrated bluish-green cytoplasm. Alizarin red staining showed red nodes in extracellular matrix, and could differentiate into osteoblasts in vitro. Related Articles | Metrics

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Proliferation, metabolism, and osteogenic differentiation of mesenchymal stem cells under hypoxia: Cell sources of placenta amniotic versus bone marrow Chen Ting, Zhou Yan, Zhang Zhi-ping, Tan Wen-song 2010, 14 (6):  957-961.  doi: 10.3969/j.issn.1673-8225.2010.06.002 Abstract ( 418 )   PDF (436KB) ( 517 )   Save BACKGROUND: Effect of hypoxia on osteogenic differentiation of bone marrow mesenchymal stem cells (BMMSCs) has been differently reported. Those differences might cause by varying volume fraction of oxygen and varying source of BMMSCs. OBJECTIVE: To compare the biological differences between placenta and BMMSCs under hypoxia. METHODS: Human placenta amniotic mesenchymal stem cells (hAMSCs) and rabbit BMMSCs were isolated by two step proteinases and whole bone marrow adhesion, respectively. hAMSCs and BMMSCs at the same passage were seeded in 12-well plates at an initial cell density of 2 × 104 cells per well with α-MEM containing 10% FBS. Then, the cells were cultured under 5% O2 or 20% O2 for 12 days. hAMSCs and BMMSCs at the same passage were seeded in 12-well plates at an initial cell density of 1 × 105 cells per well with osteogenic medium. Then, the cells were cultured under 5% O2 or 20% O2 for 14 days. Cell growth curve, the specific glucose consumption rates and specific lactate production rates, and osteogenic differentiation were detected. RESULTS AND CONCLUSION: Compared to normal oxygen, hypoxia promoted the proliferation and osteogenic differentiation of MSCs. When compared to BMMSCs, statistically significant enhancement of the growth of hAMSCs by hypoxia was observed. hAMSCs cultured under hypoxia exhibited lower glucose consumption and lactate production in contrast with BMMSCs. Furthermore, comparison between hAMSCs and BMMSCs showed that the alkaline phosphatase expression of BMMSCs was significantly enhanced by hypoxia and was markedly higher compared with hAMSCs. The amount of calcium deposition was also enhanced by hypoxia, but there were no statistically significant differences between hAMSCs and BMMSCs. Related Articles | Metrics

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Preparation, cultivation and identification of neural stem cells in fetal rat cerebral cortex: Do they have multiple differentiation potency? He Yue-qiu, Chen Hui-jin, Qian Long-hua, Chen Guan-yi 2010, 14 (6):  962-966.  doi: 10.3969/j.issn.1673-8225.2010.06.003 Abstract ( 267 )   PDF (496KB) ( 504 )   Save BACKGROUND: Periventricular leukomalacia is a major syndrome of premature infant brain injury, which has been not prevented and cured yet. Theoretically, neural stem cells which were transplanted into white matter with an absence of oligodendroglial cells might be an ideal method to cure periventricular leukomalacia. OBJECTIVE: To prepare the multi-lineage potential of neural stem cells for the use of intraventricular transplantation.  METHODS: Cerebral cortex was obtained from 12-14-day fetal rats and sectioned into 1.0-mm3 sections. The single cell suspension was separated and purified. The neurospheres were incubated with DMEM/F12 culture medium containing fetal bovine serum to observe primary and passage culture of neural stem cells. The differentiation of neural stem cells was determined using immunohistochemical method. RESULTS AND CONCLUSION: The viability of cultured neural stem cells was (94.3±2.2)%. The neurosphere was formed at day 3 after primary culture. The proliferation of neurosphere slowed down after 10-passage culture, and some cells became old. All neurospheres were positively Nestin-staining, thus they were considered as neural stem cells. A further incubation of 4-passage neurospheres, immunohistochemical method indicated that the neurosphere was positively GFAP, β-tublin, and O4 staining, respectively. This suggested that cultured neural stem cells are able to self-renew, proliferate, and differentiate into neurons, astrocytes and oligodendroglial cells. Related Articles | Metrics

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Differentiation of bone marrow stromal cells into neuron-like cells induced by brain tissue extract from ischemia/reperfusion rats in vitro Han Wei, Gong De-zheng, Li Xiao-mei, Lu Qiong1, Yin Li, Wu Wei-hua 2010, 14 (6):  967-972.  doi: 10.3969/j.issn.1673-8225.2010.06.004 Abstract ( 219 )   PDF (489KB) ( 540 )   Save BACKGROUND: Differentiation of bone marrow stromal cells (BMSCs) into neurons requires two processes: orientation and differentiation. Orientation and differentiation are results from different gene expression in cells with the same gene bank. Gene expression requires a certain condition. Changes in extracellular matrix can induce changes in cell morphology and gene expression manner. OBJECTIVE: To explore the possibility of BMSC differentiation into neuron-like cells under tissue extract from rat damaged brain. METHODS: The fifth passage of green fluorescent protein (GFP)-transfected BMSCs was induced to differentiate in brain tissue extract from ischemia/reperfusion rats or normal rats. A blank control was set. Cell morphology change was observed under the phase contrast microscope, and then evaluated using immunohistochemical staining. RESULTS AND CONCLUSION: Primarily cultured BMSCs were purified and amplified, and then showed even spindle shape. The third passage of BMSCs was positive for CD44 and CD106, but negative for CD34. Under the fluorescence microscope, BMSCs showed fluorescence expression, but the strength was weak 24 hours following GFP transfection. Numerous cells presented significant green fluorescence 48 hours later. Following adding brain tissue extract from ischemia/reperfusion rats. Induced cells presented neuron-like feature, but neuron specific enolase specific antibody presented positive expression. Compared with the blank control group, the differentiation rate of BMSCs was significantly increased in the ischemia/reperfusion group and normal group (P < 0.05). The increased range was significantly greater in the ischemia/reperfusion group than the normal group (P < 0.05). These results indicated that brain tissue extract from ischemia/reperfusion rats can successfully induce the differentiation of BMSCs into neuron-like cells. Related Articles | Metrics

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Human amniotic epithelial cells-secreted neurotrophic factors induces the differentiation of human umbilical cord blood mesenchymal stem cells into neuron-like cells: Possibility verification Zhang Xiao-ming, Sun Hai-mei, Yang Hui, Ji Feng-qing 2010, 14 (6):  973-978.  doi: 10.3969/j.issn.1673-8225.2010.06.005 Abstract ( 278 )   PDF (472KB) ( 489 )   Save BACKGROUND: Group pre-test has confirmed that amnion endothelial cell conditioned medium can induce human umbilical cord blood mesenchymal stem cells into dopaminergic neuron-like cells. In this process, neurotrophic factors and their receptors may play an important role. OBJECTIVE: To study the function of neurotrophic factors secreted by amniotic epithelial cells in the differentiation of human umbilical cord blood mesenchymal stem cells into neurons. METHODS: P1 human umbilical cord blood mesenchymal stem cells at 2×108 /L were incubated and assigned to 3 group. Control group was added with HG-DMEM medium. Induction group received human amniotic epithelial cell medium. Blocking agent group underwent blocking agent K252a fluid, and the incubated was conducted at 36 ℃ for 40 minutes, and then amniotic epithelial cell medium was added. Immunofluorescence chemistry was used to determine neuron specific enolase and dopamine transporter expression in human umbilical cord blood mesenchymal stem cells. Real-time quantitative PCR was employed to detect neuron specific enolase, dopamine transporter and tyrosine hydroxylase expression in human umbilical cord blood mesenchymal stem cells. RESULTS AND CONCLUSION: Nerve growth factor and brain-derived neurotrophic factor were observed in human amniotic supernatant. P1 human umbilical cord blood mesenchymal stem cells expressed Trka and Trkb. Forty-eight hours following induction, compared with the control group, positive expression of neuron specific enolase and dopamine transporter was significantly increased in the induction and blocking agent groups (P < 0.05), especially in the induction group (P < 0.05). Neuron specific enolase, dopamine transporter and tyrosine hydroxylase mRNA levels were significantly greater in the induction and blocking agent groups compared with the control group (P < 0.01), and each gene mRNA levels were significantly greater in the induction group than in the blocking agent group (P < 0.01). Results verified that neurotrophic factor in the human amniotic epithelial cells plays important effects on differentiation of human umbilical cord blood mesenchymal stem cells into neurons. The promotion effects are mediated by activating Trk receptor. Related Articles | Metrics

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Microglia activation stimulates bone marrow mesenchymal stem cells to release gliocyte-derived neurotrophic factor for protection of dopaminergic neurons Fan Dong-yan, Wang Ping, Liu Ran, Niu Feng-lan, Du Bo 2010, 14 (6):  979-984.  doi: 10.3969/j.issn.1673-8225.2010.06.006 Abstract ( 344 )   PDF (453KB) ( 508 )   Save BACKGROUND: Studies are very few regarding the specific reaction of bone marrow mesenchymal stem cells (BMSCs) to activated microglia. Moreover, it remains unclear how MSCs maintain dopaminergic neuronal survival under specific microenvironment. OBJECTIVE: To explore the effect of BMSCs stimulated by activated microglia on dopaminergic neuron survival. METHODS: BMSCs were isolated from Wistar rats by attachment method, and in vitro cultured; microglia was activated, and dopaminergic neurons were cultured by enzyme digestion method. The experiment included 5 groups: BMSCs, microglia, lipopolysaccharide (LPS)+microglia; BMSCs+LPS+microglia groups, in which the dopaminergic neurons were cultured with corresponding culture medium; the dopaminergic neurons alone group was cultured with 10% fetal bovine serum+ DMEM/F12. The effect of different microenvironment on dopaminergic neuron survival and gliocyte-derived neurotrophic factor released from BMSCs were detected by immunofluorescence technique. RESULTS AND CONCLUSION: The release of gliocyte-derived neurotrophic factor in groups involving BMSCs was greater than corresponding control group. Tyrosine hydroxylase immunofluorescence showed that neuronal survival of dopaminergic neurons alone group was 15%, microglia group was 10%, LPS+microglia was 5%, but BMSCs+LPS+microglia group was 28%, significantly greater than the other groups (P < 0.05). In addition, survival of in vitro cultured dopaminergic neurons was decreased with increasing culture duration, but the survival of dopaminergic neurons in group involving BMSCs was significantly greater than corresponding control group. This indicates that microglia activation stimulated BMSCs to upregulate gliocyte-derived neurotrophic factor to prevent dopaminergic neurons from toxic injury, and inhibit delayed death of dopaminergic neurons. Related Articles | Metrics

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Survival and migration of transplanted neural stem cells: Can it elevate the efficiency of transplantation by cerebellar fastigial nucleus stimulation? Huang Zuo-yi, Wu Cheng-ji, Zhu Xiao-feng, Dong Shu-xin, Wei Chun-jie 2010, 14 (6):  985-991.  doi: 10.3969/j.issn.1673-8225.2010.06.007 Abstract ( 287 )   PDF (481KB) ( 517 )   Save BACKGROUND: The discrepancy of neural stem cells (NSCs) transplantation is low cell survival rate and poor differentiation, which can not relieve the large apoptosis of neuron and endothelial cells in ischemic penumbra. This paper proposes a principle of overall intervention to provide an optimal environment for transplanted cells.  OBJECTIVE: To observe the effect of cerebellar fastigial nucleus electrical stimulation (FNS) on survival and migration of NSCs. METHODS: The EGF-responsive hippocampal NSCs of neonate Wistar rats were isolated and cultured in vitro. They were labeled by Brdu and induced by embryo cattles blood serum. The multi-differentiation potential was studied. Totally 80 rats were assigned into 4 groups. FNS+NSCs transplantation (n=32): at 24 hours FNS, right middle cerebral artery occlusion (MCAO) was prepared and received NSCs transplantation. FNS group (n =8): received the same procedure as FNS+NSCs transplantation except substitute PBS for NSCs. NSCs transplantation group (n=32): concentric electrode was inserted without electrify, the remained procedure was the same to FNS+NSCs transplantation group. Control group (n=8): concentric electrode was inserted without electrify, the remained procedure was the same to FNS group. The neural functions were recorded at hours 6, 24 and days 3, 7, 14, 28 after infarction. Rats were sacrificed at days 3, 7, 14 and 28 after transplantation, and the survival and migration of NSCs were investigated by Brdu immunocytochemical staining. RESULTS AND CONCLUSION: Isolated and purified epidermal growth factor-responsive hippocampal NSCs were Nestin-positive and had ability of self-renewing and multi-directional differentiation. More than 85% NSCs expressed the antigen of Brdu after Brdu labeled. The functional scores of the FNS+NSCs transplantation group were significant better than those of the other 3 groups at 28 days after transplantation (P < 0.05-0.01). The number of survival cells in the FNS+NSCs transplantation group was significantly greater than that of NSCs transplantation group at days 14 and 28 after transplantation. FNS can improve the survival rate of transplanted NSCs and the functional scores following MCAO. The grading-up global environment can improve the substitution role of transplanted NSCs at local cerebral infarction damage. Related Articles | Metrics

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Effects of folic acid on pERK1/2 expression of neural stem cells from neonatal rats under hypoxia condition in vitro Yang Yang, Huang Guo-wei, Zhang Xu-mei, Zhao Lin, You Qing-ju, Liu Jia-jie 2010, 14 (6):  992-995.  doi: 10.3969/j.issn.1673-8225.2010.06.008 Abstract ( 283 )   PDF (435KB) ( 400 )   Save BACKGROUND: Previous studies have verified that under normal culture of neural stem cells (NSCs), folic acid can accelerate proliferation of NSCs by phosphorylation of mitogen activated protein kinase path activation ERK1/2. OBJECTIVE: To investigate the effect of folic acid on NSC extracellular signal regulatory protein kinase pERK1/2 under hypoxic condition. METHODS: NSCs from Neonatal rats were cultured in vitro by serum-free culture method, and incubated in a flask at 1×108/L. Except normal control group, self-made hypoxia equipment was used in the hypoxia model, folic acid deficiency and folic acid supplemented groups at day 3. At 37 ℃, hypoxia culture was conducted in the thermostat for 6 hours. The contents of folic acid were 4 mg/L, 4 mg/L, 0.65 mg/L, 8 mg/L in the four groups. Cells following 6 days were collected to count the density using trypan blue. RT-PCR was utilized to detect pERK1/2 mRNA expression. Western blot assay was employed to determine pERK1/2 protein expression. RESULTS AND CONCLUSION: Compared with the normal control group, the proliferation of NSCs and the expression of ERK1/2mRNA and pERK1/2 protein were decreased significantly in the hypoxia model group. Compared with the hypoxia model group, the proliferation of NSCs and the expression of ERK1/2mRNA and pERK1/2 protein were increased in the folic acid supplemented group, whereas decreased in the folic acid deficiency group. There were significant differences among groups (P < 0.001). Above-described results verified that folic acid supplementation can activate ERK1/2 phosphorylatin and accelerate proliferation of NSCs under hypoxia condition. Related Articles | Metrics

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Human umbilical cord mesenchymal stem cells under atomic force microscope: Correlation between biological characteristics and ultrastructure Chen Li, Wu Ben-qing, Zhu Hua-min 2010, 14 (6):  996-1001.  doi: 10.3969/j.issn.1673-8225.2010.06.009 Abstract ( 233 )   PDF (871KB) ( 435 )   Save BACKGROUND: The relationship between cellular morphosis and function is undetachable, but there is little investigation about ultrastructure of human umbilical cord mesenchymal stem cells (hUCMSCs). OBJECTIVE: To study the relationship between the functions of hUCMSCs and their ultrastructure obtained by atomic force microscope (AFM). METHODS: hUCMSCs were isolated, cultured, expanded after enzyme digestion. P3 cells were observed under AFM. Immunophenotype and cell cycle were analyzed by flow cytometry, as well as induction of the adipogenic, osteogenic differentiation of hUCMSCs were identified using Oil red O staining and alkaline phosphatase staining. RESULTS AND CONCLUSION: hUCMSCs at passage 3 were strongly positive for CD44 and CD29, weakly positive for CD106, but negative for hematopoietic marker CD34. Cells in G0/G1 phase accounted for 80%. Proliferation index was 19.9%. Following adipogenic induction, alkaline phosphatase staining demonstrated brown cytoplasm in cube and polygonal cells. AFM showed hUCMSCs were spindle shape, obvious cytoskeletal filament that connected into nets, which fit for the strong capacities for proliferation, migration and differentiation. Related Articles | Metrics

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Isolation, culture and multiple differentiations of rabbit bone marrow-derived mesenchymal stem cells in vitro Xu Cheng-feng, Hu Da-hai, Zhao Zhou-ting, Zhang Wan-fu, Bai Xiao-zhi, Cai Wei-xia 2010, 14 (6):  1002-1005.  doi: 10.3969/j.issn.1673-8225.2010.06.010 Abstract ( 281 )   PDF (447KB) ( 509 )   Save BACKGROUND: Mesenchymal stem cells (MSCs), with low immunogenicity, can regulate cellular immunity and mitigate graft rejection, which has a good prospect in tissue engineering. However, it is rarely present in bone marrow.  OBJECTIVE: To explore an isolation and culture method of the rabbit bone marrow-derived MSCs, to observe the biological characteristics and differentiation potential of bone marrow-derived MSCs. METHODS: MSCs were isolated from rabbit tibia bone marrow by combination of gradient centrifugation and different adherent method, then proliferation in vitro. Morphology was examined by phase contrast microscopy, and the growth curve of cultured MSCs was drawn via MTT results. MSCs were treated with osteogenetic inductor (L-DMEM/F12, 10% fetal bovine serum, 0.1 μmol/L dexamethasone, 200 μmol/L vitamin C, 10 mmol/L β-phosphoglycerol), adipose inductor (L-DMEM/F12, 10% FBS, 1 μmol/L dexamethasone, 200 μmol/L antifani, 0.5 mmol/L IBMX, 10 μg/mL insulin), and chondrocytes inductor (L-DMEM/F12, 10% FBS, 10 μg/L TGF-β1, 0.1 μmol/L dexamethasone, 50 μmol/L vitamin C, 6.25 mg/L insulin) to differentiated into osteoblast, dipocytes and chondrocytes. And the differentiated cells were identified by alkaline phosphatase staining, oil red O staining, and toluidine blue staining, respectively. RESULTS AND CONCLUSION: Bone marrow-derived MSCs can be isolated and cultured by the combination of gradient centrifugation and different adherent method in vitro, which have the better potentiality of proliferation and multi-directional differentiation. Mostly of the primary and passaged cells were spindle-shaped. After osteogenetic induction, cells were positive to alkaline phosphatase staining. Oil red O staining showed that red lipid droplet existed in adipose cells, and toluidine blue staining showed that toluidine blue was positive after chondrocytes induction. Related Articles | Metrics

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Effect validation of bone marrow-derived mesenchymal stem cells cultured by adherent method in vitro Zhang Yan, Chen Xi-hai, Ji Yan-chao, Zhai Zhe, Wu Bo 2010, 14 (6):  1006-1008.  doi: 10.3969/j.issn.1673-8225.2009.06.011 Abstract ( 294 )   PDF (384KB) ( 432 )   Save BACKGROUND: A small number of mesenchymal stem cells (MSCs) present in bone marrow, which would gradually drop with age.  OBJECTIVE: To verify the effect of adherent method on culture of bone marrow-derived MSCs. METHODS: Under anaesthesia, bone marrow cells were obtained from femur and tibia of rats, cultured by DMEM containing calf serum, placed in an incubator containing 5% CO2 at 37 ℃. The culture medium was renewed after 24 hours, and remained periodical medium change with once per week. The weakly adherent cells were passaged. The cell morphology, growth curve, and the expression of cell-surface markers were identified by flow cytometry and immunocytochemical staining. RESULTS AND CONCLUSION: After 24 hours of culture, the cells could adhere to the walls with fusiform or triangle shapes, proliferated faster after 2-3 days, and presented whirlpool-like or clustering. The cells reached a logarithmic growth phase after 2 days, and into the late stationary phase after 12 days, which covered the bottle after 15 days. The cultured cells were positive to CD90 and CD54. The results verified that bone marrow-derived MSCs can be isolated by adherent method. This method is easy operation, and can maintain cell activity preferably. Related Articles | Metrics

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Co-culture of prostate carcinoma cells with alginate and bone marrow mesenchymal stem cells: To observe the effect of stem cells on proliferation speed and clustering size of prostate carcinoma cells Xie Jie, Chen An-min, Guo Feng-jin, Wang Jian-chao, Liao Hui, Liu Hao, Chen Chao 2010, 14 (6):  1009-1014.  doi: 10.3969/j.issn.1673-8225.2010.06.012 Abstract ( 310 )   PDF (528KB) ( 510 )   Save BACKGROUND: Models concerning tumor external environment mainly concentrated on laboratory two-dimensional culture and in vitro animal experiment, which lack of three-dimensional stereo. OBJECTIVE: To establish in vitro bone metastasis stereo models of human prostate carcinoma, and to investigate the effect of stem cells on proliferation rate and clustering size of prostate carcinoma cells. METHODS: Bone marrow mesenchymal stem cells (BMMSCs) were extracted from 2 clean grade SD rats. Alginate was used to simulate medullary microenvironment, where prostate carcinoma cells and BMMSCs were co-culturedd. Growth of the cells in the three-dimensional model was observed through microscope and histological sections. The carcinoma cells were transfected with green fluorescent protein. The proliferation of monoclonal cells clustering was observed under light microscope and fluorescence microscope.  RESULTS AND CONCLUSION: In the co-culture group, the clustering speed, clustering amount and tumor formation rate were greater that those of the control group. The monoclonal cells clustering was formed at 7.75 days and 6.00 days in the control and co-culture groups, respectively, with cell counts of (95.13±11.63) and (112.53±14.67) after 10 days. The formation rate of fluorescent cell clones was (77.10±6.85)% in the control group and (64.55±6.21)% in the co-culture group, the difference had significance. The results suggested that: the alginate microenvironment is conductive to proliferation and clustering of prostate carcinoma cells and BMMSCs. Related Articles | Metrics

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Tumorigenicity of rat bone marrow-derived liver stem cells Hou Jian-bin, Liu Chao, Yu Xian-huan, Xu Lei-bo 2010, 14 (6):  1015-1018.  doi: 10.3969/j.issn.1673-8225.2010.06.013 Abstract ( 340 )   PDF (418KB) ( 579 )   Save BACKGROUND: Mobilizing autologous or extraneous bone marrow-derived liver stem cells may promote liver regeneration, however, its safety before the large scale clinical application needs further evaluation. OBJECTIVE: Bone marrow-derived liver stem cells (BDLSCs) were induced by culturing the rat bone marrow mesenchymal cells in the medium containing 5% cholestatic sera, and then were implanted into nude mice to observe the tumorigenicity. METHODS: Rat bone marrow mesenchymal cells (BMSCs) were isolated and incubated in the medium containing 5% cholestatic sera. Immunofluorescent stain was used to detect the expression of albumin, alpha-fetoprotein and cytokeratin18 by the cultured cells. Glycogen and urea synthesis by these cells were analyzed, respectively. BDLSCs following 14 days of culture were incubated in the skin of nude mice to observe neoplasia in local site. RESULTS AND CONCLUSION: Rat BMSCs survived in the medium containing 5% cholestatic serum and formed into small colonies on the fourth day after culture. Seven days later, the colonies expanded and there appeared some polygonal cells in the peripheral area. About 14 days later, these polygonal cells were confluent and presented the shape of cobblestone. Immunofluorescent stain showed that these cells expressed cytokeratin18, albumin and alpha-fetoprotein. Staining for glycogen displayed that glycogen granules were seen in cells. From 12 to15 days after culture, urea nitrogen concentrations in the medium were gradually increased. Rat BDLSCs were incubated in the skin of nude mice. Thirty days later, no neoplasia was found in the local site, and the tissue structure was normal. This result indicated that rat BDLSCs induced with the medium containing 5% cholestatic serum might have not tumorigenicity. Related Articles | Metrics

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Differentiation of bone marrow mesenchymal stem cells into the cells of skin appendages in diabetic wound Zhong Xiao-hong, Wang Ming-gang, Zhao Li-ping, Gao Xin-yu 2010, 14 (6):  1019-1022.  doi: 10.3969/j.issn.1673-8225.2010.06.014 Abstract ( 229 )   PDF (450KB) ( 496 )   Save BACKGROUND: Bone marrow mesenchymal stem cells (MSCs) have multi-differentiation potential. In the acute wound, MSCs have been demonstrated to have the potential for differentiating into skin cells. However, there are few reports regarding its differentiation in diabetic wound. OBJECTIVE: To observe the feasibility of differentiation of MSCs into the cells of skin appendages under the microenvironment of diabetic wound. METHODS: MSCs were isolated from the bone marrow of rats, purified and cultured. Third or fourth passage MSCs were selected and labeled with 5-bromodeoxyuridine (5-BrdU). The rats were injected intraperitoneally with single administration of streptozocin to establish diabetes model. After 2 weeks, a round skin wound was made on the dorsal back of rats. BrdU-labeled MSCs at a density of 1×109/L were injected into the wound of the rats. The specimens were harvested from the wound tissues to prepare sections at 2 and 3 weeks after transplantation, followed by immunohistochemical staining with BrdU or keratin. RESULTS AND CONCLUSION: BrdU positive cells aggregated in the epidermis, dermis and hypodermia. Some positive cells appeared in the sebaceous glands and sebaceous duct cells and expressed keratin simultaneously in serial sections. During diabetic wound healing, MSCs have the potential to differentiate into the cells of the skin appendages. Related Articles | Metrics

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Retrorsine effects on regeneration and repair of injured liver in mice undergoing partial hepatectomy Liao Zhi-ling, Chen Jia-ling, Kuang Xiao-cong, Zhu Ming-yi, Huang Ying-hua, Cai Jie 2010, 14 (6):  1023-1026.  doi: 10.3969/j.issn.1673-8225.2010.06.015 Abstract ( 327 )   PDF (481KB) ( 460 )   Save BACKGROUND: In many studies, rats were commonly used as models of retrorsine-induced hepatic injury. Some reports have confirmed that retrorsine cannot inhibit proliferation of mouse hepatic cells. Other reports have shown that retrorsine has inhibitory effects on proliferation of mouse hepatic cells. OBJECTIVE: To study the liver regeneration after hepatic injury by creating mouse models treated with partial hepatectomy combination with retrorsine. METHODS: A total of 40 C57BL/6J mice were equally and randomly assigned to 2 groups. In the partial hepatectomy combined with retrorsine group, intraperitoneal injection of retrorsine 70 mg/kg was conducted, twice, within an interval of 2 weeks. Four weeks later, 2/3 hepatectomy was performed. In the partial hepatectomy group, intraperitoneal injection of saline 70 mg/kg was performed, twice, with an interval of 2 weeks. Four weeks later, 2/3 hepatectomy was performed. At 14 days after partial hepatectomy, the restoration of the livers was observed. The liver cell injury was observed at 3, 7 days with hematoxylin-eosin staining. The hepatocyte proliferation was observed at 3 days with BrdU staining. Oval cell proliferation was observed at 3, 7and 14 days with CK19 and C-kit antibody immunohistochemistry. RESULTS AND CONCLUSION: In the partial hepatectomy group, the damaged liver nearly restored to normal at 14 days after partial hepatectomy, and the result was contrary to partial hepatectomy combined with retrorsine group. Hematoxylin-eosin staining demonstrated that significant degeneration changes in hepatic cells in the partial hepatectomy combined with retrorsine group. BrdU staining showed that hepatocyte proliferation at day 3 was significantly determined in the partial hepatectomy group, but few in the partial hepatectomy combined with retrorsine group. CK19 and C-kit immunohistochemistry demonstrated that visible oval cell proliferation was seen in mice of partial hepatectomy combined with retrorsine group. First of all, hepatic oval cells appeared in portal area and differentiated into hepatic cells and bile duct cells, and then grew into the hepatic lobule gradually. These indicated that retrorsine can obviously inhibit hepatocyte regeneration after liver injury in mice. The model of mice treated with retrorsine and partial hepatectomy could induce oval cell proliferation. Related Articles | Metrics

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Four kinds of extracellular matrixes support the growth of human embryonic stem cells: Effect differences Hu Zhi-xing, Luo Min, Zhou Yi-ping, Liang Dao-ming 2010, 14 (6):  1027-1030.  doi: 10.3969/j.issn.1673-8225.2010.06.016 Abstract ( 245 )   PDF (412KB) ( 498 )   Save BACKGROUND: Extracellular matrixes play important roles in the maintenance of undifferentiated state and self-renewal of human embryonic stem cells (hESCs). Establishment of a defined serum- and feeder-free culture system is a prerequisite for the application of hESCs in cell-replacement therapy. OBJECTIVE: The present study was designed to compare the effects of four kinds of extracellular matrixes on the support the growth of hESCs. METHODS: We set four groups using conventionally recovered hESCs BGO2. BG02 was cultured on Matrigel-, laminin-, fibronectin-, or collagen-coated plates in serum-free medium containing basic fibroblast growth factor (bFGF), transforming growth factor beta 1(TGFβ1), Insulin-Transferrin-Selenium (ITS). The attachment rate and differentiated rate of BG02 clumps in various conditions were measured. The proportions of Oct-4 and Nanog positive cells in various groups were assessed by flow cytometry. The extracellular matrix receptor expression was analyzed by RT-PCR. RESULTS AND CONCLUSION: The attachment rates of BG02 cell clumps in Matrigel and laminin groups were significantly higher than that of fibronectin and collagen groups (P < 0.05), but differentiated rate was significantly reduced (P < 0.05). The percentage of Oct-4+ and Nanog+ BG02 cells in Matrigel and laminin groups were higher than that of fibronectin and collagen groups (P < 0.05). No significant difference in each index was determined between the fibronectin and collagen groups (P > 0.05). BG02 cells in Matrigel and laminin groups expressed higher levels of integrin α5, integrin α6 and integrin β1 mRNA than that in fibronectin and collagen groups. Above-mentioned results indicated that Matrigel and laminin could support the proliferation of BG02 cells well, while fibronectin and collagen could not support the proliferation of BG02 cells. These variations might be attributed to the activation of integrin subunits in BG02 cells. Related Articles | Metrics

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Effects of methylprednisolone on the secreted function of Schwann cells Qu Wei, Fei Liang-jian, Jiang Hua-jun, Fu Chong-yang, Zhang Wei-guo, Lü De-cheng 2010, 14 (6):  1031-1036.  doi: 10.3969/j.issn.1673-8225.2010.06.017 Abstract ( 266 )   PDF (644KB) ( 473 )   Save BACKGROUND: Secretion of various neurotrophic factors by Schwann cells plays important roles in neural regeneration. However, the secretion capability is affected by many factors. To seek a feasible method for promoting nerve growth factor secretion by Schwann cells is a key of regeneraion following neurologic defect. OBJECTIVE: To explore the effects of methylprednisolone(solu-medrol) on the secreted function of Schwann cells of cultured rats.   METHODS: Schwann cells were isolated and cultured by enzyme digestion method. Cell growth was observed under an inverted phase contrast microscope. Following passage, purity of some Schwann cells was identified using S-100 protein immunity. Other Schwann cells were regulated using cell counting plate into 1×109/L, and incubated in a 6-well culture plate (15 wells) for further incubation. Following 4 days of culture, different concentrations of solu-medrol (10-3, 10-4, 10-6, 10-8 mol/L) were administrated to the cell, while blank control group (1 well) was given no drug. 24, 48 and 72 hours after administration, reverse trancription-polymerase chain reaction (RT-PCR) was used in the detection of the levels of nerve growth factor mRNA. RESULTS AND CONCLUSION: Number of primarily cultured cells was significantly increased at day 7, and 80% cells were confluent. Subcultured cells were spindle-shaped, with 2 thin long processes, showing positive fluorescence staining. Fibroblasts were round or flat, showing negative reaction of fluorescence staining. Reserve transcription-polymerase chain reaction demonstrated that nerve growth factor number at 72 hours affected by 10-8 mol/L radiosone was increased compared with the blank control group and other concentrations and other time points (P < 0.05). Number of nerve growth factor was reduced following treatment of 10-3 mol/L radiosone compared with the blank control group and other concentrations (P < 0.05). These results suggested that high concentration of solu-medrol prohibits secreted function of Schwann’s cells, but long time and low dosage solu-medrol promotes secreted function of Schwann’s cells. Related Articles | Metrics

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In vivo magnetic resonance imaging tracking of bone marrow-derived mesenchymal stem cells via intracoronary administration: Consistency to pathohistological results Dou Xing-kui, Guo Tao, Yu Zhuo, Zhao Xin-xiang, Sun Hai-mei, Pu Shun-hua, Kang Bo 2010, 14 (6):  1037-1042.  doi: 10.3969/j.issn.1673-8225.2010.06.018 Abstract ( 269 )   PDF (551KB) ( 452 )   Save BACKGROUND: Recent trials and clinical studies have shown that intracoronary transplantation of bone marrow-derived mesenchymal stem cells (MSCs) improves cardiac function following acute myocardial infarction (AMI). However, whether homing of MSCs into the infarcted myocardium or not is still unknown. OBJECTIVE: To study the homing of MSCs intracoronary administration in porcine myocardial infarction model using in vivo magnetic resonance imaging tracking. METHODS: Porcine MSCs were isolated and cultured by the whole bone marrow method. Following labeling by superparamagnetic iron oxide (SPIO), MSCs were treated with trypsinization to adjust the concentration at 1010/L. Myocardial infarction was induced in all 10 pigs. At one week after modeling, the labeled MSCs were delivered via intracoronary infusion with standard over-the-wire (OTW) balloon angioplasty catheters. Prussian blue staining was used to evaluate labeling efficiency, and double echo steady state was used to scan four-chamber and cor biloculare at long axis view, which was considered as locating phase to obtain image of left ventricle at short axis view. RESULTS AND CONCLUSION: MSCs could be efficiently and safely labeled with SPIO. Intracoronary transplantation of MSCs is able to home the sites of myocardial injury and the border between infarcted and normal tissue. MRI can track SPIO-labeled MSCs delivered through intracoronary and were confirmed on pathology. After 5 weeks the injected labeled cells could still be detected with MRI. Related Articles | Metrics

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Differentiation and survival of autologous bone marrow mesenchymal stem cells following transplantation into the myocardium Cai Hong-yan, Nie Jun, Chen Li-xing, Zhao Ling, Guo Tao, Xiao Jian-ming 2010, 14 (6):  1043-1047.  doi: 10.3969/j.issn.1673-8225.2010.06.019 Abstract ( 298 )   PDF (498KB) ( 454 )   Save BACKGROUND: It was uncertain that the migration, differentiation and survival of transplanted bone marrow mesenchymal stem cells (BMSCs) into myocardium after the acute myocardial infarction. OBJECTIVE: To investigate the migration, differentiation and survival of rabbit transplanted autologous BMSCs in myocardium after the acute myocardial infarction. METHODS: Rabbit BMSCs were isolated and labeled by DAPI in vitro. Rabbit left anterior descending branch was ligated to establish acute myocardial infarction models. Following successful model establishment, 30 New Zealand rabbits were assigned to BMSC and control groups (n = 15). In the BMSC group, autologous BMSCs were infused into the surrounding sites of the infracted region by 4 points 1 hour following coronary artery ligation. In the control group, the same region was injected with an equal volume of saline. Injection volume was 30 μL in each point. Five animals from each group were sacrificed 10 minutes, 3 days and 4 weeks following transplantation. The heart was obtained to undergo frozen sections. The distribution of DAPI-labeled BMSCs was observed using fluorescence microscope. Immunofluorescence method was used to examine the troponin Ⅰ and α-actin. RESULTS AND CONCLUSION: DAPI-labeled BMSCs with blue nuclei were distributed extensively in the myocardium of the cell transplantation group, ovoid in shape and arranged in parallel with the cardiac muscle fibers. Troponin Ⅰ and α-actin were positive immunofluorescently in the cytoplasm of the labeled BMSCs. Results indicated that transplanted BMSCs in the ischemic myocardium could differentiate into myocardial cells under stimulation of local microenvironment. Related Articles | Metrics

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Immunomodulatory effects of xenogenous umbilical cord mesenchymal stem cells on rat heart transplantation Liu Kui-li, Liu De-zhong, Jin Jian-gang, Li Hai-bin, Shi Ying-chang, Li Li, Han Yong, Xu Xiao-guang, Shi Bing-yi 2010, 14 (6):  1048-1052.  doi: 10.3969/j.issn.1673-8225.2010.06.020 Abstract ( 531 )   PDF (488KB) ( 452 )   Save BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) can prolong the survival time of mice and baboons’ alloskin graft and degrade acute and chronic graft-versus-host disease (GVHD) incidence after hematopoietic stem cell transplantation. But, at present there is no report that rat umbilical cord mesenchymal stem cells (UC-MSCs) reduced rejection response following heart transplantation. OBJECTIVE: To study the immunomodulatory effects of rat UC-MSCs on a model of allogeneic heart transplantation. METHODS: A total of 20 DA rats served as donors, and 20 Lewis rats as recipients. They were equally and randomly assigned to 2 groups: drug intervention and control groups (n=10). Using double cannulation, the left pulmonary artery and innominate artery of rat donors and external jugular vein and common carotid artery of rat recipients received end-to-end anastomosis under a microscope to establish heterotopic cardiac transplantation. One Wistar pregnant rat was selected to harvest UC-MSCs by collagenase digestion method. Following model establishment, rats in the cell transplantation group received UC-MSCs via caudal vein. Rats in the control group received sodium chloride. Survival time of the transplanted heart was determined. The transplanted heart received histopathology score using the acute rejection diagnosis criteria. Lymphocyte infiltration of transplanted heart was observed using hematoxylin-eosin staining. RESULTS AND CONCLUSION: Compared with the control group, the survival time of the transplanted heart was significantly longer in the cell transplantation group (P = 0.001), and the pathological score of acute rejection was significantly reduced (P = 0.000 4). There were lots of lymphocyte and monocyte infiltration in the myocardium in the control group. Little lymphocyte infiltration was detected in the myocardium in the cell transplantation group, with the presence of mild edema of myocardial interstitial substance. Results verified that rat UC-MSCs can induce immune tolerance of heart transplantation, soften immunological rejection and prolong xenograft survival. Related Articles | Metrics

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Histopathological changes of rat injured spinal cord following olfactory ensheathing cell transplantation Wang Guo-yu, He Xi-jing, Yuan Pu-wei, Li Hao-peng, Chang Rui 2010, 14 (6):  1053-1057.  doi: 10.3969/j.issn.1673-8225.2010.06.021 Abstract ( 252 )   PDF (761KB) ( 425 )   Save BACKGROUND: There are no effective treatments for spinal cord injury. Transplantation of olfactory ensheathing cells (OECs) has achieved great progress in repairing spinal cord injury. OBJECTIVE: To observe the effect of OECs transplantation on pathological and ultrastructural alterations of spinal cord, and the role in spinal cord injury developing. METHODS:A total of 60 SD rats were randomly divided into blank, model, transplantation and DF12 groups, with 15 animals in each group. The entire vertebral plate of T10, and partial vertebral plate of T9 and T11 of blank group were cut open, and gelatin sponge was used for hemostasis. In the model group, the spinal cord was excised. In the transplantation and DF12 groups, OECs and DF12 culture solution were injected following spinal cord excision. The incision was sutured. Two rats from each group were anesthetized 1, 3, 7, 14, 28, 42, and 56 days following injury, and injured areas were observed by light microscopy and electron microscopy. RESULTS AND CONCLUSION: Following spinal cord injury, pathological and ultrastructural changes occurred, such as hemorrhage, edema, degeneration, necrosis, cavitation, gliacyte proliferation and nerve fiber regeneration. OECs transplantation attenuated neuronal and nerve fiber necrosis, relieved degree of pathological reaction, protected injured neurons, prevented gliacyte proliferation and increased nerve fiber regeneration. Results show that OECs transplantation ameliorated pathological reactions and promoted spinal cord injury repair. Related Articles | Metrics

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Flk-1+ bone marrow mesenchymal stem cell transplantation upregulates interleukin-6 level: Whether it simultaneously aggravates collagen-induced arthritis in mice? Chen Bin, Huang Shan, Hu Jian-li, Sun Zhao, Han Qin, Song Zeng-xuan, Zhao Chun-hua 2010, 14 (6):  1058-1063.  doi: 10.3969/j.issn.1673-8225.2010.06.022 Abstract ( 276 )   PDF (619KB) ( 465 )   Save BACKGROUND: Immunoloregulation of mesenchymal stem cells (MSCs) is commonly approved. Previous studies have confirmed the ability of Flk-1+ bone marrow MSCs (BMSCs) to inhibit T/B lymphocyte proliferation in vitro. OBJECTIVE: To investigate the therapeutic effect of Flk-1+ BMSCs in collagen-induced arthritis mice. METHODS: A total of 18 healthy male DBA-1(H-2Kq) mice aged 10 weeks were randomly divided into 3 groups. All the mice were injected at the base of the tail with bovine type II collagen (CII), and received a booster injection of CII on day 21 to establish the CIA mice model. DBA-1(H-2Kq)mouse Flk-1+ BMSCs were isolated in vitro by the density gradient centrifugation and adherence screening. Following initial immunity, mice in the cell transplantation group were infused with Flk-1+ BMSCs (1-2)×106 cells/mouse via the caudal vein. Mice in the cell transplantation group were injected with the same volume of Flk-1+ BMSCs during booster. Mice in the model control group were injected with an equal volume of saline 0 or 21 days following initial immunity. Following initial immunity and booster immunization, claw pad thickening and clinical score were observed, changes of joint pathology and dynamic changes in serum factor mass concentration were determined in mice.  RESULTS AND CONCLUSION: Compared with the model control group, no significant difference in claw pad thickening and mean clinical score was detected in the cell transplantation group following initial immunity (P > 0.05), with the presence of obvious damage to synovial membrane and inflammatory cell infiltration. Mass concentration of each serum cell factor was similar. The claw pad was significantly thickened (P < 0.01), mean clinical score reached 3.35 points, with severe damage to synovial membrane, proliferation of blood capillary in the cell transplantation group following booster immunization. Interleukin-6 levels were greatly increased at day 28 following initial immunity (P < 0.1), but decreased at day 35 following initial immunity (P < 0.1). Results indicated that in the collagen-induced arthritis mouse models, Flk-1+ BMSC transplantation did not obtain prospective therapeutic efficacy, but aggravation of arthritis was observed in the cell transplantation group following booster immunization. Upregulation of interleukin-6 concentration could aggravate the behavior symptom of rheumatoid arthritis mice. Related Articles | Metrics

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Intravenous transplantation of bone marrow mesenchymal stem cells in combination with recombinant human growth hormone repairs myocardium and vascular tissues in rats with congestive cardiomyopathy Yao Wei, Wang Feng-zhi 2010, 14 (6):  1064-1067.  doi: 10.3969/j.issn.1673-8225.2010.06.023 Abstract ( 256 )   PDF (449KB) ( 495 )   Save BACKGROUND: It is controversial whether bone marrow mesenchymal stem cells can retain in cardiac injured position, or differentiate into cardiomyocytes or not. OBJECTIVE: To study the effects of recombinant human growth hormone and bone marrow mesenchymal stem cells (BMSCs) intravenous transplantation on myocardium and angiogenesis in rats with congestive cardiomyopathy. METHODS: BMSCs were collected from rats by density gradient centrifugation and adhesive-screening method. Models of cardiac failure were established using adriamycin induction in the model, cell transplantation, growth hormone and combination groups. Following model establishment, cell transplantation group received BrdU-labeled BMSCs (8×1013/L) via vein. Growth hormone group underwent subcutaneous injection of human growth hormone 2 U/kg per day, for 14 consecutive days. Combination group received injection of human growth hormone and BMSC transplantation. At week 4, samples were collected. Immunohistochemical staining for BrdU+MHC and BrdU+Actin was used to determine homing of BMSCs to evaluate the differentiation of transplanted cells into cardiomyocytes and vascular endothelia cells. Hematoxylin-eosin staining was utilized to detect vascular density. RESULTS AND CONCLUSION: Compared with the cell transplantation group, positive rate of Brdu immunohistochemistry was increased in the combination group (P < 0.001). The number of cardiomyocytes and vascular endothelia cells was significantly increased following Brdu+MHC and Brdu+Actin staining (P < 0.001). Compared with the model group, total vascular density, microvessel density and capillary density were significantly increased in the growth hormone, cell transplantation and combination groups (P < 0.001). No significant difference was determined among growth hormone, cell transplantation and combination groups (P > 0.05). Intravenous transplantation of BMSCs could repair cardiomyocytes and vascular endothelial cells by homing into the heart. BMSCs could survive in damaged area and differentiate into cardiomyocytes or vascular endothelial cells and increase the vascular density significantly. Growth hormone could improve microenvironment and raise rates of differentiating into cardiomyocytes or vascular endothelial cells. Related Articles | Metrics

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Best time for mobilization and collection of peripheral blood stem cell from healthy donors Chen Xiao-xia, Wang Zhi-ming, Luo Xian-sheng, Xu Dan-dan, Li Xing, Lei Mei-qing 2010, 14 (6):  1068-1071.  doi: 10.3969/j.issn.1673-8225.2010.06.024 Abstract ( 369 )   PDF (334KB) ( 582 )   Save BACKGROUND: Effective mobilization and collection of hemopoietic stem cells are initial factors for peripheral stem cell transplantation, while they make sure a permanent reconstruction of hemopoiesis. Mobilization and collection have been developed; however, the collection is more, and the yield of hemopoietic stem cells is various. OBJECTIVE: To investigate the best time of mobilization and collection of peripheral blood stem cells from healthy donors. METHODS: A total of 16 donors who were selected from Haikou People’s Hospital between January 2003 and December 2008 were randomly divided mobilization group (A, n=6) and mobilization + collection group (B, n=10). A group was subcutaneously injected with 5.0-10.0 μg/(kg·d) recombinant human granulocyte colony-stimulating factor (rhG-CSF) (Filgrastim), and B group was treated with rhG-CSF and intravenously injected with 10 mg dexamethasone. Peripheral stem cells were collected twice in each group. The two groups were collected at the fourth and 5th day of mobilization after the second or 4th hour subcutaneous injection of rhG-CSF. The collection was 4.0-5.0 mL, the manhandled volume was 3.0-5.0 mL, and the total blood volume was 6.7-10.1 L. RESULTS AND CONCLUSION: Number of mononuclear cells was (4.0-8.0)×108 kg-1 in the two groups. The cell concentration of fourth hour was higher than the second hour after rhG-CSF treatment (P < 0.05). The mononuclear cell concentration of fourth hour was higher than the second hour after the fourth and 5th day of rhG-CSF treatment. Our research showed that we could collect sufficient amount of cells (to a high concentration) by one time, which had reasonable collection time – value-effectiveness relationship, when the cycle blood volume was 1.8-2.2 times of circulating blood volume. Related Articles | Metrics

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Ligustrazine induces rat bone morrow mesenchymal stem cells to differentiate into neuron-like cells: Screening of the optimal inductive concentration Chen Bing, Yin Yan-qing, Ke Jun-long, Zou Xin-hui, Peng Hao, Tan Shan-feng, Xu Zhi-en 2010, 14 (6):  1072-1077.  doi: 10.3969/j.issn.1673-8225.2010.06.025 Abstract ( 280 )   PDF (558KB) ( 528 )   Save BACKGROUND: There are numerous inducers used in inducing bone marrow mesenchymal stem cells (BMMSCs) differentiate into neuron-like cells, however, due to poisonous, most chemical inducers can not be used in human. OBJECTIVE: To investigate the effect of ligustrazine on differentiation of rat BMMSCs into neuron-like cells in vitro, and to search for the optimal inductive concentration. METHODS: After SD rats were anesthetized, bone marrow was obtained from the femoral and tibial bones, centrifuged, and the supernatant was discarded. The extracted cells were cultured in L-DMEM supplemented with 15% fetal bovine serum. The expression of CD44 and CD45 of the 5th passage of BMMSCs were identified by immunocytochemical technique. Serum-free L-DMEM medium contains 1.00, 1.25 and 1.50 g/L ligustrazine concentrations were used to induce the 5th passage of BMMSCs in vitro. Morphology changes of BMMSCs were observed under an inverted phase microscope. Expression of nestin, neuron-specific enolase and glial fibrillary acidic protein were identified by immunocytochemical technique, and the expression ratio of neuron-like cells’ surface antigens induced by different concentrations of ligustrazine were compared.  RESULTS AND CONCLUSION: ①Most primarily cultured BMMSCs adhered to the wall at 3 days after culture, which proliferated faster after passaged, and the 5th passage of cells were mostly purified into BMMSCs, spread radially or vortex-likely. ②The 5th passage of BMMSCs was positive expressed (98.02±0.81)% CD44, but negative for CD45. ③Neuron-like cells with prominence and bifurcation could be seen after induction. The immunocytochemical method showed that nestin and neuron-specific enolase in most induced cells were positive expressed, especially received a highest ration of neuron-specific enolase expressing in the induced group with 1.25 g/L concentration of ligustrazine. It revealed that ligustrazine can induce BMMSCs differentiated into neuron-like cells, and 1.25 g/L is the optimal inductive concentration. Related Articles | Metrics

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Effects of three kinds of Mongalian medicine axillary choerospondias fruit extracts on differentiation of human umbilical cord blood stem cells in vitro Sa Rengaowa, Bulin Baiyila, Chen Ji-ming 2010, 14 (6):  1078-1081.  doi: 10.3969/j.issn.1673-8225.2010.06.026 Abstract ( 312 )   PDF (353KB) ( 487 )   Save BACKGROUND: Human umbilical cord blood stem cells have been widely used in the study of spinal cord injury, but in vitro differentiation of human umbilical cord blood stem cells (hUCBSCs) has been limited by various factors. Mongalian medicine axillary choerospondias fruit extract has protective effects on neural cells, but the action mechanisms are unclear. OBJECTIVE: To observe promoting effects of 3 kinds of Mongalian medicine axillary choerospondias fruit extracts on in vitro differentiation of hUCBSCs. METHODS: Fresh umbilical cord blood was obtained from healthy puerperants to prepare hUCBSC suspension. The purified hUCBSCs were incubated in 40 petri dishes. The Mongalian medicine axillary choerospondias fruit extracts were divided into: sample 1 group: ethanol extraction, ethyl acetate extraction, crude drug mass concentration was 8.25 g/mL; sample 2 group: ethanol extraction, NKA resin isolation, 10% ethanol eluting concentration, crude drug mass concentration was 1.72 g/mL; sample 3 group: ethanol extraction, NKA resin isolation, 70% ethanol eluting concentration, crude drug mass concentration was 2.41 g/mL; control group: incubation of 80% DMEM containing 20% calf serum. Effects of various mass concentrations of Mongalian medicine axillary choerospondias fruit extract on hUCBSCs proliferation were observed. Proportion in S phase was measured using flow cytometry at 24 and 72 hours. RESULTS AND CONCLUSION: The proliferation of hUCBSCs was not significant in the sample 3 group. At day 10, the proliferation was significantly greater in the sample 1 and 2 groups compared with the sample 3 and control groups (P < 0.01). The number of hUCBSCs was significantly increased at 24 and 72 hours in S phase in the sample 1 and 2 groups. Mongalian medicine axillary choerospondias fruit extract (crude drug mass concentration 8.25, 1.72 g/mL) could promote in vitro proliferation of hUCBSCs. Related Articles | Metrics

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Signal molecules related to self-renewal of hemopoietic stem cells: Effects and pathways Li Na, Shi Zeng-li, Wang Yue-si 2010, 14 (6):  1084-1087.  doi: 10.3969/j.issn.1673-8225.2010.06.028 Abstract ( 306 )   PDF (418KB) ( 492 )   Save BACKGROUND: There are small amount of hematopoietic stem cells, which are easy to divide in vitro and show the difficulty in applying to transplantation. OBJECTIVE: This paper has focused on the role and means of the Wnt, Notch, Bmi_1, Shh, HOXB4 signaling molecule in the maintenance of hematopoietic stem cell self-renewal and regulation. METHODS: With the key words of “HSC, Wnt, Notch, Bmi_1, Shh, HOXB4” for the search, we searched PubMed database (2002-01/2008-12) in English. Literatures closely related to the hematopoietic stem cell self-renewal related signaling molecules were included. Repetitive research and Meta analysis were excluded. RESULTS AND CONCLUSION: The computer initially retrieved 216 documents, of which 30 documents for research. Hematopoietic stem cells are self-renewing, have strong differentiation and growth and regeneration capacities, can produce various types of blood cells ancestor cells, are widely used to treat blood diseases, but the hematopoietic stem cell differentiation in vitro demonstrated that the difficulties used in transplantation. How to make hematopoietic stem cells in vitro amplification and processing, while maintaining hematopoietic stem cell self-renewal characteristics is of a key issue. In recent years, different signaling pathways to enhance the ability of hematopoietic stem cell self-renewal signaling molecule have been a research hotspot. The article focused on the role and means of the Wnt, Notch, Bmi_1, Shh, HOXB4 in the maintenance of hematopoietic stem cell self-renewal and regulation and found that both the above-mentioned five signaling molecules can enhance hematopoietic stem cell self-renewal function. There are also a number of factors playing an important role in the maintenance of hematopoietic stem cell self-renewal process, such as endogenous factors, a series of transcription factors Oct-4, Ehox, Nanog, SCL, Runx1 and so on, to explore how their regulatory networks formed by the interaction control self-renewal of hematopoietic stem cells will become a key point in the research of self-renewal of hematopoietic stem cells. Related Articles | Metrics

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Stem cells and exercise-induced skeletal muscle cell apoptosis Qu Hong-lin, Wang Yong 2010, 14 (6):  1088-1091.  doi: 10.3969/j.issn.1673-8225.2010.06.029 Abstract ( 322 )   PDF (445KB) ( 507 )   Save BACKGROUND: Exercise-induced skeletal muscle apoptosis has been referred to as the current focus of sports medicine, and the application of stem cells in athletic injury rehabilitation and prevention has been reported. However, the role of stem cells in cell apoptosis remains unclear. OBJECTIVE: To summarize the effect and mechanism of stem cells in preventing exercise-induced skeletal muscle apoptosis to provide references for scientific sports training and physical activity. METHODS: A computer-based online search of PubMed (1991-01/2009-10) and CNKI (1994-01/2009-10) was performed for related articles with the keywords “"Exercise Training, Sports, Skeletal Muscle, Apoptosis" in English and "stem cells, exercise, skeletal muscle, apoptosis," in Chinese. Inclusion criteria: ① studies on stem cells and their apoptosis in skeletal muscle cells; ②articles in the same field published recently or in the authoritative journals. Exclusion criteria: ① Repetitive articles; ② Meta analysis. RESULTS AND CONCLUSION: A total of 360 articles were collected from the summary of literature through the stem cells and its application in the field of sports medicine research, and its skeletal muscle apoptosis in the changes and development trend of the application. Finally, 31 articles were included, including 21 reviews and 10 clinical or experimental studies. High-intensity exercise can cause apoptosis in skeletal muscle cells, while the use of stem cell technology can prevent apoptosis, to a certain extents by regulating Bcl-2 and Bax protein expression, thereby promoting the early recovery of skeletal muscle. Related Articles | Metrics

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Epithelial cells-stem cells interactions in wound healing Chen Su-li, Liu Liu 2010, 14 (6):  1092-1096.  doi: 10.3969/j.issn.1673-8225.2010.06.030 Abstract ( 287 )   PDF (530KB) ( 555 )   Save BACKGROUND: At the process of wound healing, the epithelial cells and epidermal stem cells interactions is complicated which promoting the mechanisms of wound healing. The abnormities of any fine phase can be result in delayed healing or scarring. OBJECTIVE: To summarize the characterization of epithelial cells and epidermal stem cells and their interaction in wound healing so that we can recognize the mechanisms of scarring from histomorphology, biochemistry and molecular variations. METHODS: A computer-based online search of Pubmed Database was undertaken to identify the relevant articles on epithelial cell and epidermal stem cells interactions in wound healing and results in scarring published from 1974 to 2009 with the key words of “epithelial cells, epithelial stem cells, wound healing, keratinocytes, fibroblasts, hypertrophic scar, keloid” in English. At the same time, Chinese relevant articles were searched in China Journal Full-text Database (CJFD) published between 1999 and 2009 with the same key words in Chinese. A total of 43 articles were collected about the cell structure (3 articles), the cell interactions in wound healing and scarring (16 articles), the relevant on epithelial stem cells (24 articles). RESULTS AND CONCLUSION: The effect of different cells in epithelial tissue is known to all. The keratinocytes and fibroblasts had been on studying frequently, and their closely correlated to scarring. At present, the epithelial stem cells has been confirmed, but the relevant on their location, quality and specific epidermal markers are unclear which need to study and explore. With the rapidly development of science and technology, we maybe control the expression of cells by genic regulation not only culture the cells of our need but to prevent the scarring. Related Articles | Metrics

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Whether autologous bone marrow mesenchymal stem cell transplantation is safe, feasible, and effective to the treatment of intracerebral hemorrhage: A 32-case analysis Zhu Jian-xin, Li Zhong-min, Xiao Tai-wu, Chen Shuang-feng, Geng Feng-yang, Fu Qiang, Guo Chuan-jun 2010, 14 (6):  1097-1100.  doi: 10.3969/j.issn.1673-8225.2010.06.031 Abstract ( 294 )   PDF (421KB) ( 587 )   Save BACKGROUND: Previous animal studies demonstrated that bone marrow mesenchymal stem cells could differentiate into nerve cells under a certain condition; however, the clinical application for treating nervous system disease has been less reported. OBJECTIVE: To observe a short-term effect of autologous bone marrow mesenchymal stem cell transplantation on treating cerebral hemorrhage. METHODS: A total of 32 patients with cerebral hemorrhage who were selected from the Department of Neurosurgery, Liaocheng Brain Hospital between November 2007 and January 2009 were considered as a treatment group. According to general data and the amount of hematoma, they were treated by drilling drainage or hematoma evacuation. Drainage tubes were detained into hematoma cavity, and 3.5 mL autologous bone marrow mesenchymal stem cell suspension was injected through drainage tube. A total of 40 additional patients who did not treated with stem cell transplantation were considered as a control group. Neurologic impairment (NIHSS) and activities of daily living (Barthel index) were performed before and 6 months after transplantation; meanwhile, the brain MRI, serum biochemical and tumor marker were evaluated to detect security of stem cell transplantation. RESULTS AND CONCLUSION: The NIHSS score and Barthel index in the treatment group were similar to those in the control group before transplantation. Compared with control group, NIHSS scores were significantly decreased in the treatment group (P < 0.01), but Barthel index was significantly increased 6 months after transplantation (P < 0.01). Compared with before transplantation, NIHSS score were significantly decreased (P < 0.01), but Barthel index was significantly increased in the treatment group 6 months after transplantation (P < 0.01). Two patients in the treatment group had febrile, which was recovered after treatment. The following-up 6 months after transplantation demonstrated that brain MRI and biochemical indicators were normal except an increasing of CA-153 caused by lung cancer in one patient. The autologous bone marrow mesenchymal stem cell transplantation for treatment of cerebral hemorrhage is safe and effective in a short-term period; however the long-term effect still needs to be further studied. Related Articles | Metrics

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Changes of vascular cell adhesion molecule-1 in serum and marrow stroma cell supernatant of patients with relapsed acute leukemia Li Xue-gang, Hou Li-jun 2010, 14 (6):  1101-1104.  doi: 10.3969/j.issn.1673-8225.2010.06.032 Abstract ( 234 )   PDF (374KB) ( 447 )   Save BACKGROUND: Relapse of acute leukemia is possibly correlated with abnormal expression of cellular adhesion molecule-1 (VCAM-1). OBJECTIVE: To observe the level of VCAM-1 in serum and bone marrow stromal cell (MSC) supernatant of patients with relapsed acute leukemia. METHODS: Samples of serum and MSC supernatant were collected from 17 patients with remission-phase and relapse-phase acute leukemia hospitalized in the Department of Hematology, the Fifth Affiliated Hospital of Sun Yat-sen University between June 2006 and March 2008. The levels of VCAM-1 were measured with ELISA in remission and relapse phases. RESULTS AND CONCLUSION: Compared with the remission phase, the VCAM-1 level of serum was significantly increased in the relapse phase (P < 0.05). At 1 and 3 weeks after in vitro culture, the VCAM-1 level in the MSC supernatant was not changed (P > 0.05), but it was significantly increased on the fourth week (P < 0.05). The results demonstrated that VCAM-1 expression of serum and MSC supernatant in the relapse phase was greater than that in the remission phase, suggesting that abnormal expression of VCAM-1 was possibly correlated with relapse of acute leukemia. Related Articles | Metrics

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Rat bone marrow masenchymal stem cells differentiate into neuron-like cells and glial-like cells under hippocampal neuron conditioned medium in vitro Li Zhao-hui, Cai Zhi-ping, Cui Hui-xian, Li Sha, Xie Guo-sheng, Li Nan, Xue Lei 2010, 14 (6):  1105-1110.  doi: 10.3969/j.issn.1673-8225.2010.06.033 Abstract ( 209 )   PDF (399KB) ( 634 )   Save BACKGROUND: There are few reports addressing the differentiation of bone marrow mesenchymal stem cells (BMSCs) into neurons, and the uncertainties mainly focused on the differentiated neurons had neuron morphology, but did not have neuron function. OBJECTIVE: To investigate the feasibility of rat bone marrow mesenchyma stem cells (BMSCs) differentiation into neuron-like cells and glial-like cells under rat hippocampal neuron’s conditional medium. METHODS: Rat BMSCs at passage 5 were divided into 4 groups. The medium of hippocampal neurons and glial cells was added in the conditioned medium group. The Dulbecco's modified Eagle's medium containing bFGF was added in the basic fibroblast growth factor (bFGF) group. The serum-free medium containing Neurobasal and B27 was added in the serum-free medium group. The DMEM supplemented with fetal bovine serum was added in the negative control group. 12 and 24 hours following induction, neuron specific enolase (NSE), microtubule-associated protein-2 (MAP-2) and glial fibrillary acidic protein (GFAP) were detected using immunocytochemical staining in each group. NSE, MAP-2 and GFAP expression was determined using Western-blot assay. RESULTS AND CONCLUSION: 12 and 24 hours following induction, BMSCs were positive for MAP-2, GFAP and NSE in the conditioned medium, bFGF and serum-free medium groups, but negative in the negative control group. Compared with the negative control group, MAP-2 expression was significantly enhanced in the conditioned medium, bFGF and serum-free medium groups 24 hours following induction (P < 0.05), and the increased range was significantly greater in the conditioned medium group compared with other two groups (P < 0.05). No significant difference in NSE and GFAP expression was detected in the conditioned medium, bFGF and serum-free medium groups. Results suggested that hippocampal neuron conditioned medium can in vitro induce the differentiation of rat BMSCs into neuron-like cells and glial cell-like cells. Compared with the bFGF medium and serum-free medium, positive rate was greatest in the hippocampal neuron conditioned medium-induced neurons and glial cells. Related Articles | Metrics

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Feeder-free growth of human embryonic stem cells supported by basic fibroblast growth factor Xu Hui-fang, Zhang Su-ming 2010, 14 (6):  1111-1116.  doi: 10.3969/j.issn.1673-8225.2010.06.034 Abstract ( 273 )   PDF (317KB) ( 469 )   Save BACKGROUND: Human embryonic stem cells (hESCs) are pluripotent cells which may differentiate into tissues of all three germ layers. Such research as the feeder-free growth of hESCs is few in China. Fibroblast growth factor (FGF) is a major factor to maintain the undifferentiated state of hESCs. OBJECTIVE: To evaluate the ability of FGF at different concentrations in maintaining the undifferentiated state and pluripotency of hESC lines in the long-term culture. METHODS: Two cell lines of hES-8 and hES-18 were cultured with mouse embryonic fibroblast condition medium for 3 passages and then transferred into mouse embryonic fibroblast condition medium containing different concentrations of FGF: 100, 160, 250 μg/L for 8 passages. The hESCs were removed from the petri dish, cell clusters were digested with collagenase IV and gathered. Cell differentiation and pluripotency were observed. The eighth generation of the hESCs were collected and incubated into severe combined immunodeficiency mice, so as to observe teratoma formation. Morphologies of the cells were evaluated. Alkaline phosphatase staining, surface labeling immunocytochemical analysis and RT-PCR assay method were utilized to determine the OCT-4 expression and tumorigenesis in vivo. RESULTS AND CONCLUSION: Cultured in mouse embryonic fibroblast condition medium containing 160 and 250 μg/L FGF, two cell lines of hESC could maintain undifferentiated state: Clones were round with a high ratio of nucleus to cytoplasm. Large areas in the center of clones were undifferentiated cells, while surrounding the clones were differentiated cells; Strong positive expression for alkaline phosphatase staining was observed; Two cell lines showed high levels of OCT-4 transcription factor protein; The surface markers SSEA-4, TRA-1-60, TRA-1-81 were all positive on both two lines; The hESC clusters could form embryoid body in vivo 10 days later; 3 germ layers of teratomas were also obtained after implanted into severe combined immunodeficiency mice. Mouse embryonic fibroblast condition medium containing 100 μg/L FGF was not sufficient to maintain the long-term proliferation of hESCs, and most of the cells differentiated and died after 4 passages. Alone with concentration 160 μg/LbFGF or more could maintain two hESC lines undifferentiated stably in vitro, has no influence on the differentiation and totipotency of two cell lines. Related Articles | Metrics

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Extraction and purification of neonatal versus adult rat Schwann cells Liu Zhi-xin, Song Bao-hui, Zhao Fu-sheng, Li Yue-zhen, Liang Jun 2010, 14 (6):  1115-1119.  doi: 10.3969/j.issn.1673-8225.2010.06.035 Abstract ( 264 )   PDF (402KB) ( 487 )   Save BACKGROUND: Schwann cells are the seed cells of neural repair, and it is a key to harvest a large number of Schwann cells with high purity and activity. OBJECTIVE: To compare the in vitro culture, purification, and morphology of Schwann cells between neonatal and adult rats, and investigate a simple and feasible culture method to harvest high-purity Schwann cells. METHODS:Totally 30 Sprague-Dawley rats, comprising 20 neonatal (1-3 days after birth, neonatal group) and 10 adult (weighing 150-200 g, adult group) rats, were included. Following double-enzyme digestion and two incubations, Schwann cells were isolated and purified by differential attachment. Cell morphology and attaching speed were determined through the use of inverted microscope. Cells were counted and cell purity was calculated. Cell proliferative ability was detected by MTT microcolorimetry. Curves of cell proliferation in each group were depicted to determine proliferative speed. Schwann cells were identified by S-100 immunochemistry. RESULTS AND CONCLUSION: Compared with fibroblasts, neonatal rat Schwann cells exhibited faster, while adult rat Schwann cells showed slower, attaching speed. Both neonatal and adult groups yielded over 96% cell purity. MTT microcolorimetry results revealed that Schwann cells proliferated actively in neonatal and adult groups. Cell proliferative curves show that neonatal rat Schwann cells proliferated faster than adult rat Schwann cells (P < 0.05). S-100 immunochemistry results showed positive results in both groups. All these findings suggest that double-enzyme digestion and two incubations followed by differential attachment is a satisfactory method to harvest considerable Schwann cells with high purity and activity. Neonatal rat Schwann cells show stronger proliferative, attaching capacities than adult rat Schwann cells. Related Articles | Metrics

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Human umbilical cord blood-derived mesenchymal stem cells inhibit cardiomyocyte apoptosis under co-culture conditions A safety and efficacy assessment Yang Shui-xiang, Huang Jing-ling 2010, 14 (6):  1120-1124.  doi: 10.3969/j.issn.1673-8225.2010.06.036 Abstract ( 267 )   PDF (387KB) ( 452 )   Save BACKGROUND: Umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) have been shown to lead to new tissue formation after homing and engrafting to the heart. But the safety of UCB-MSCs engrafting remains to be further investigated. OBJECTIVE: To study the safety and apoptosis inhibition of the UCB-MSCs under co-culture conditions on human cardiomyocytes.  METHODS: UCB was collected at delivery with informed consent obtained from 10 donors. The UCB-MSCs were treated with 5-azaserine to induce differentiation into cardiomyocytes. The in vitro cultured cells of the 3rd-5th passages and dividing cells were taken to detect telomerase activity, tumor-related gene expression, G-banding patterns of chromosomal karyotupes, cell surface antigen expression, tumor formation in nude mice, and inhibited apoptosis under co-culture conditions. RESULTS AND CONCLUSION: Prior to and after 5-azaserine induction, telomerase activity and tumor-related gene expression (p53, cyclin A, cdk2, β-actin, C-fos, h-TERT, c-myc) of UCB-MSCs were similar, no abnormal chromosomal karyotupes were observed, immunophenotype exhibited no change, CD34 was negative, but CD44 and CD90 (Thy-1) were positive. At 10 weeks after inoculation of UCB-MSCs, nude mice still survived healthily and no formed tumor in vivo was observed. Hematoxylin-eosin staining suggested normal subcutaneous tissue. Compared with simple cardiomyocytes, UCB-MSCs could significantly inhibit cardiomyocyte apoptosis under co-culture conditions (P < 0.05), indicating that human UCB-MSCs are a valuable, safe, and effective source of cell transplantation treatment. Related Articles | Metrics

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Role of insulin-like growth factor-1 in proliferation, migration and differentiation of neural stem cells in cerebral infarction rats Ye Fei, Xi Gang-ming, Chen Tao, Bao Yu-hua, Wang Jia-ning 2010, 14 (6):  1125-1129.  doi: 10.3969/j.issn.1673-8225.2010.06.037 Abstract ( 353 )   PDF (342KB) ( 475 )   Save BACKGROUND: Insulin-like growth factor-1 (IGF-1) is a peptide hormone, it has been proved a promotion role on the proliferation of precursor cells. OBJECTIVE: To explore the intravenous injection of IGF-1 on the proliferation, migration and differentiation of neural stem cells in rats after cerebral ischemia. METHODS: Eight adult male SD rats were randomly divided into control group and experimental group, with 40 rats in each group. The rats in two groups were used to prepare models of focal cerebral ischemia using modified suture method, the rats in the experimental group were treated with tail vein injection of IGF-1, according to 100 μg/kg computation, the injection was given for 6 continuous days; in the control group, rats were given equal volume of saline. The rats were decapitated at 7, 14, 21, 28 days following intervention, respectively, and rats in each group were given intraperitoneal injection of the BrdU at 1 day before death. Immunohistochemistry and double staining were applied to detect the expressions of BrdU-positive cells, PSA-NCAM-positive cells, BrdU + PSA-NCAM double-positive cells, and BrdU + MAP2 double-positive cells. RESULTS AND CONCLUSION: The number of BrdU-positive cells and PSA-NCAM positive cells reached the peak at 7 days after ischemia; BrdU + PSA-NCAM double-labeled-positive cells could be detected in ischemic bilateral subependymal zone and dentate gyrus, the number was the most at 7 days, then followed by a gradual decrease; the BrdU + MAP2 double-positive cells began to increase from 14 days, and then gradually increased along with the decrease of BrdU + PSA-NCAM double-positive expression, showing a reverse trend. Intravenous injection of IGF-1 can induce the proliferation, differentiation and migration of neural stem cells in rats following ischemic brain injury. Related Articles | Metrics

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Expression of vascular cell adhesion molecule 1 and intercellular adhesion molecule 1 in rat bone marrow mesenchymal stem cells Xu Yun, Hu Yan-lai, Zhang Zhao-lin, Liang Yan-hong, Tian Hua, Tian Guang-ping 2010, 14 (6):  1130-1133.  doi: 10.3969/j.issn.1673-8225.2010.06.038 Abstract ( 272 )   PDF (337KB) ( 454 )   Save BACKGROUND: Following bone marrow mesenchymal stem cells (BMMSCs) infusion therapy, which factor promotes BMMSCs migrated to correct position is a key point, currently, adhesion molecule is thought to be playing an important role in mediating BMMSCs migration. OBJECTIVE: To investigate the expression of vascular cell adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1) in rat BMMSCs. METHODS: BMMSCs were in vitro separated from rat bone marrow by directly adherence method. The expression of VCAM-1 and ICAM-1 were identified by using immunocytochemical staining, and the expression rates of antigen were tested by flow cytometry, in addition, their mRNA expressions were measured by RT-PCR. RESULTS AND CONCLUSION: Immunocytochemistry demonstrated that BMMSCs weakly expressed VCAM-1, but strong expressed ICAM-1. Flow cytometry showed that the expression rate of VCAM-1 was 6%, and the expression rate of ICAM-1 was 100%. RT-PCR showed that BMMSCs expressed a low level of VCAM-1 mRNA but a high level of ICAM-1 mRNA. It revealed   under physiological condition, BMMSCs expressed a low level of VCAM-1, whereas they expressed a high level of ICAM-1. Related Articles | Metrics

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Glycosylphosphatidilinoditol-specific phospholipase    D expression in bone marrow mononuclear cells derived from acute leukemia patients Xiao Guang-fen, Chen Fang-ping, Wang Guang-ping, Fu Bin, Xie Jun-ming, Cheng Ying-ni, Li Qun, Jian Zai-fu 2010, 14 (6):  1134-1137.  doi: 10.3969/j.issn.1673-8225.2010.06.039 Abstract ( 254 )   PDF (395KB) ( 486 )   Save BACKGROUND: The correlation of gycosylphosphatidilinoditol-specific phospholipase D (GPI-PLD) activity, mRNA expression to leukemia type, hepatosplenomegaly and/or lymphadenopathy has been rarely reported. OBJECTIVE: To explore the correlation of GPI-PLD expression to leukemia type and hepatosplenomegaly and/or lymphadenopathy of acute myeloid leukemia (AML) patients.  METHODS: Fresh bone marrow specimens were obtained from 43 newly diagnosed AML patients, 28 acute lymphocytic leukemia (ALL) patients, and 21 normal persons. Bone marrow mononuclear cells were harvested by density gradient centrifugation. GPI-anchored human placent alkaline phosphatase was used as substrate. GPI-PLD activity was determined bytriton-X114 phase partitioning procedure. GPI-PLD mRNA expression was detected by semi-quantitative RT-PCR. The relationship of GPI-PLD activity, mRNA expression and leukemia type, hepatosplenomegaly and/or lymphadenopathy was analyzed. RESULTS AND CONCLUSION: Compared with control group, GPI-PLD activity and mRNA expression in bone marrow mononuclear cells were significantly higher in AML group (P < 0.01), while they were significantly lower in the ALL group (P < 0.01). Of 43 patients with AML patients, 13 patients had hepatosplenomegaly and/or lymphadenopathy. The GPI-PLD activity (%) and mRNA expression were significantly higher in AML patients without hepatosplenomegaly and lymphadenopathy than those patients with hepatosplenomegaly and/or lymphadenopathy (P < 0.05). These results demonstrated that GPI-PLD activity alteration is consistent with GPI-PLD mRNA expression in AML patients, and the expression levels correlate to leukemia type and hepatosplenomegaly and/or lymphadenopathy of AML patients. Related Articles | Metrics

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Rituximab combined with autologous hematopoietic stem cell transplantation for treatment of non-Hodgkin lymphoma in 6 patients Sun Zhi-qiang, Wang Ji-shi, Lu Ying-hao, Xie Run-lan, Long Zheng-mei 2010, 14 (6):  1138-1140.  doi: 10.3969/j.issn.1673-8225.2010.06.040 Abstract ( 253 )   PDF (254KB) ( 442 )   Save BACKGROUND: Rituximab single or in combination with CHOP regimen for treatment of CD20-positive non-Hodgkin lymphoma has achieved good curative effects. Autologous hematopoietic stem cell transplantation (AHSCT) has been shown to improve the curative effects and increase survival rate of patients with non-Hodgkin lymphoma. However, the curative effects of these two methods remain disputed. OBJECTIVE: To investigate the efficiency of rituximab in combination with AHSCT on CD 20-positive non-Hodgkin lymphoma. METHODS: Six patients with CD 20-positive non-Hodgkin lymphoma (stage IV) underwent AHSCT and rituximab administration. 375 mg/m2 rituximab was intravenously administered 2-4 times prior to AHSCT, twice prior to and after peripheral blood stem cells mobilization and preprocessing, respectively, as well as once every 3 months after AHSCT. RESULTS AND CONCLUSION: The mean number of mononuclear cells and CD 34-positive cells was 5.13×10-8/kg and 4.75×10-6/kg, respectively. Following AHSCT, all 6 patients presented normal hematopoietic functions, neutrophils exceeded 0.5×10-9/L at 9-15 days and blood platelet counts exceeded 20×10-9/L at 12-19 days. Hemorrhagic cystitis, interstitial pneumonia, cytomegalovirus infection, or hepatic venous obstruction was not observed during the whole process of AHSCT in each patient. At 6-32 months, patients completely recovered. These results indicate that rituximab in combination with AHSCT is a good method for treatment of CD20-positive non-Hodgkin lymphoma and rituximab maintenance therapy could prevent disease recurrence. Related Articles | Metrics