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01 July 2012, Volume 16 Issue 27 Previous Issue    Next Issue

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Isolation, cultivation and osteogenic induction of guinea pig bone marrow mesenchymal stem cells Shi Xiao-lu, Duan Li-fu, Ma Yan, Bi Xiao-juan, Liu Li-zhong 2012, 16 (27):  4941-4946.  doi: 10.3969/j.issn.2095-4344.2012.27.001 Abstract ( 312 )   PDF (579KB) ( 315 )   Save BACKGROUND: The culture technique and osteogenic induction of bone marrow mesenchymal stem cells would be further investigated based on flexible and easy harvesting of bone marrow mesenchymal stem cells. OBJECTIVE: To investigate the practical methods of isolation and cultivation of guinea pig bone marrow mesenchymal stromal cells in vitro and to investigate the phenotype characteristics and multilineage differentiation potential of bone marrow mesenchymal stem cells. METHODS: Guinea pig bone marrow mesenchymal stem cells were isolated and purified by adhesion method. After subculturing and proliferation, expression of cell surface molecule CD29, CD44, CD45 was detected by flow cytometry. The multilineage differentiation capability of bone marrow mesenchymal stem cells was examined by culturing cells under conditions favorable for adipogenic and osteogenic differentiation. RESULTS AND CONCLUSION: The primary cultured bone marrow mesenchymal stem cells adhered to plastic surface after 96 hours and exhibited an oval, asteroid or fusiform shape. After 8 days, the cells reached 90 % confluence in a single layer. After subculturing and amplification, the cells were further purified with a uniform firbroblast-like morphology. Flow cytometry analysis showed that the positive rate of CD29 and CD44 was 95.7% and 65.7% respectively. After culture in adipogenic and osteogenic medium respectively, passage 3 cells were successfully induced to differentiate into adipogenic and osteogenic lineages, as identified by oil red O, alizarin red S, alkaline phosphatase staining and immunohistochemical type Ⅰ collagenase staining. These findings suggest that by adherence method, in vitro isolated, cultured guinea pig bone marrow mesenchymal stem cells can greatly proliferate and maintain the phenotypic and functional properties of mesenchymal stem cells. Related Articles | Metrics

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骨髓间充质干细胞体外调控肝星状细胞死亡受体5的表达 Yang Wen, Qin Shan-yu, Jiang Hai-xing, Zhang Jun-hong, Ning Lin, Meng Yun-chao 2012, 16 (27):  4947-4952.  doi: 10.3969/j.issn.2095-4344.2012.27.002 Abstract ( 281 )   PDF (476KB) ( 393 )   Save 背景：肿瘤坏死因子相关凋亡诱导配体可以通过上调死亡受体5蛋白的表达诱导大鼠肝星状细胞凋亡。 目的：观察大鼠骨髓间充质干细胞对肝星状细胞死亡受体5和Caspase-3蛋白表达的影响，探讨骨髓间充质干细胞诱导肝星状细胞凋亡及其机制。 方法：贴壁筛选法培养、纯化SD大鼠骨髓间充质干细胞，传至第3代使用；大鼠原代肝星状细胞和肝纤维原细胞系冻融后传代使用。应用6孔培养板，建立上下双层细胞共培养体系，常规培养。将细胞分为4组：①空白对照组：肝星状细胞和骨髓间充质干细胞分别单独培养。②阴性对照组：肝纤维原细胞与肝星状细胞共培养(模拟星状细胞体内生长环境)。③骨髓间充质干细胞与肝星状细胞共培养组。④实验组：1.5 mg/L TRAIL多克隆抗体与骨髓间充质干细胞作用6 h后，不换液，再与肝星状细胞共培养。 结果与结论：在共培养组中肝星状细胞的增殖降低，凋亡增加，且肝星状细胞死亡受体5、Caspase-3蛋白的表达均显著升高，并呈时间依赖性，且与其他组比较差异有显著性意义(P < 0.01)。实验组Caspase-3和死亡受体5蛋白的表达在共培养的各个时间段均低于共培养组，与共培养组比较差异有显著性意义(P < 0.01)。提示骨髓间充质干细胞与肝星状细胞共培养能抑制肝星状细胞的增殖，促进肝星状细胞的凋亡，其机制可能为骨髓间充质干细胞旁分泌肿瘤坏死因子相关凋亡诱导配体通过上调Caspase-3和死亡受体5蛋白的表达实现的。 Related Articles | Metrics

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Bone morphogenetic protein-2 induces rat bone marrow mesenchymal stem cells to differentiate into cardiomyocyte-like cells in vitro Gao Chen-wei, Wang Hai-ping, Wu Zhi-gang, Lü Yang, Huo Yan-li, Wang Hao-yu 2012, 16 (27):  4953-4958.  doi: 10.3969/j.issn.2095-4344.2012.27.003 Abstract ( 226 )   PDF (566KB) ( 344 )   Save BACKGROUND: Many kinds of cytokines, traditional Chinese medicine monomers and tissue lysate have been used for the differentiation process by investigators. And they try to treat myocardial infarction. OBJECTIVE: To explore the effect of bone morphogenetic protein-2 in the differentiation of rat bone marrow mesenchymal stem cells into cardiomyocyte-like cells in vitro. METHODS: Bone marrow mesenchymal stem cells were isolated from SD rat bone marrow and cultured for 72 hours with the medium containing bone morphogenetic protein-2 (the control group was cultured with normal medium), and then cultured with normal medium for 4 weeks. RESULTS AND CONCLUSION: After purification and culture, the cells showed CD29 positive and CD34 negative, which met the characteristics of bone marrow mesenchymal stem cells, the growth curve showed the cells had similar growtj patterns. Bone marrow mesenchymal stem cells exhibited a narrow morphology and tightly packed in the same direction after induced by bone morphogenetic protein-2. The differentiated cells were positive for C-TnI, C-TnT, desmin, α-sarcomeric actin and P38MAPK. There was no positive staining in control group. Bone morphogenetic protein-2 can induce the bone marrow mesenchymal stem cells to differentiate into cardiomyocyte-like cells in vitro. It is a new and effective inductor for bone marrow mesenchymal stem cells to differentiate into cardiomyocyte-like cells in vitro. Related Articles | Metrics

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Co-culture of rabbit bone marrow mesenchymal stem cells and human amniotic membrane Zhang Zhi-jin, Guo Lin, Zhao De-wei 2012, 16 (27):  4959-4962.  doi: 10.3969/j.issn. 2095-4344.2012.27. 004 Abstract ( 152 )   PDF (392KB) ( 298 )   Save BACKGROUND: In recent years, great progress has been made in repair of cartilage defects using tissue engineering technique. Bone marrow mesenchymal stem cells (BMSCs) as common seed cells have been widely used in the tissue engineering of cartilage tissue. OBJECTIVE: Human amniotic membrane was used as cell scaffold to load rabbit BMSCs and the biocompatibility of human amniotic membrane was observed. METHODS: Rabbit BMSCs were inoculated onto human acellular amnion. After co-culture, cell morphology was observed. The proliferation of BMSCs was investigated by MTT assay, and the histocompatibility of human amniotic membrane was evaluated. Cell survival and apoptosis was determined by 4’, 6-diamidino-2-phenylindole. RESULTS AND CONCLUTION: BMSCs grew well on the surface of human amniotic membrane and cell viability increased. DAPI staining results showed that only a small number of cells were apoptotic. These findings suggest that human amniotic membrane is a scaffold material with good biocompatibility and can promote the proliferation of BMSCs. Related Articles | Metrics

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Effects of tissue homogenate from injured liver on induced differentiation of bone marrow mesenchymal stem ce Hou Jun-xia, Mei Zhe-chuan, Jiang Ming-de 2012, 16 (27):  4963-4967.  doi: 10.3969/j.issn.2095-4344.2012.27.005 Abstract ( 219 )   PDF (499KB) ( 342 )   Save BACKGROUND: Whether bone marrow mesenchymal stem cells can differentiate into hepatocytes is closely related to microenvironment. OBJECTIVE: To explore the effects of the acute alcoholic liver injury tissue homogenate on induced differentiation of bone marrow mesenchymal stem cells. METHODS: BMSCs were isolated from Sprague-Dawley rats, cultured, purified and identified in vitro. Rat models of alcohol-induced acute liver injury were prepared. Tissue homogenate of normal and injured liver was used to induce differentiation of bone marrow mesenchymal stem cells. The α-fetoprotein and albumin expression was detected at 0, 3, 7, 10, 14, and 21 days using RT-PCR and western blot methods, respectively. RESULTS AND CONCLUSION: After induced by tissue homogenate of normal and injured liver, bone marrow mesenchymal stem cells exhibit epithelioid change. After induced by each tissue homogenate, α-fetoprotein expression was detected from the 3rd day, peaked at about 7 days, decreased thereafter, and exhibited a very low level by 14 days; albumin expression was detected from the 7th day, and the expression level was gradually increased with time going. The mRNA and protein expression level was consistent. These findings demonstrate that hepatocytes gradually developed till mature. There was no significant difference in α-fetoprotein or albumin expression after induced by tissue homogenate of normal and injured liver (P > 0.05). The liver tissue homogenates of acute alcoholic liver injury can induce bone marrow mesenchymal stem cells to differentiate into hepatocytes, with similar effects to normal liver tissue homogenate. Related Articles | Metrics

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Biological characteristics and chondrogenic differentiation of bone marrow-derived multipotent adult progenitor cells Wen Yu, Fan Shi-xiang, Yi Xian-hong, Li Bin, Pan Jun 2012, 16 (27):  4968-4973.  doi: 10.3969/j.issn.2095-4344.2012.27.006 Abstract ( 334 )   PDF (477KB) ( 328 )   Save BACKGROUND: Bone marrow-derived multipotent adult progenitor cells have stronger differentiation and expansion potential than bone marrow mesenchymal stem cells and exhibit the multiple differentiation potential of chondroblast-like cells. But the reports on the differentiation into chondroblast-like cells are rare. OBJECTIVE: To study the culture and characteristics of bone marrow-derived multipotent adult progenitor cells and the potential to differentiate into chondroblast-like cells. METHODS: The bone marrow-derived multipotent adult progenitor cells were cultured in vitro, and then the morphology and the growth conditions were observed and identified. The differentiation medium was used to introduce the bone marrow-derived multipotent adult progenitor cells to differentiate into chondroblast-like cells. RESULTS AND CONCLUSION: Observation under the light microscope showed that bone marrow-derived multipotent adult progenitor cells were large and grew in long polygonal with several slender pseudopodia, the cells were with large nucleus and affluent cytoplasma and had strong proliferation ability; the expression of transcription factor 4 and stage-specific embryonic antigen 1 could be seen, the expressions of T-cell surface adhesion molecules of CD44, CD45 and CD133 and major organizations compatibility antigen systems Ⅱ were not shown. After induction and differentiation, toluidine blue stained differentiated cells showed metachromasia and could express collagen type Ⅱ. The bone marrow-derived multipotent adult progenitor cells were successfully cultured in vitro, and the bone marrow-derived multipotent adult progenitor cells could differentiate into the chondroblast-like cells in a certain induction conditions. Related Articles | Metrics

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Telomerase expression during induced differentiation of bone marrow mesenchymal stem cells into neural stem cells Xu Tao, Yang Ming-kun, Sheng Wei-bin, Huang Kai 2012, 16 (27):  4974-4978.  doi: 10.3969/j.issn.2095-4344.2012.27.007 Abstract ( 227 )   PDF (433KB) ( 320 )   Save BACKGROUND: Telomerase plays an important role in cell differentiation and activation. OBJECTIVE: To detect the expression of telomerase during induced differentiation of bone marrow mesenchymal stem cells into neural stem cells. METHODS: The expression of telomerase in the former six generations of SD rat bone marrow mesenchymal stem cell subcultures was detected using the method of PCR-ELISA. Passage 3 SD rat bone marrow mesenchymal stem cells were induced to differentiate into neural stem cells and subpassaged till passage 6, and the expression of telomerase in each generation was detected. The protein extract of 293 cells of human embryo kidney cell line was used as the positive control, and protein extract from inactivated cells was used as the negative control. RESULTS AND CONCLUSION: Telomerase showed a positive expression in the former six generations of bone mesenchymal stem cell subcultures, which was 9.4%-30.9% of the positive control groups, and the highest telomerase expression appeared in the third generation. Telomerase showed a positive expression in the former five generations of subcultures after induced-differentiation into neural stem cells, which was 5.4%-33.1% of the positive control groups. Telomerase expression peaked in the third and fourth generations of subcultures. These results suggest that telomerase is an important factor of cell differentiation and proliferation. Related Articles | Metrics

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Adipose derived mesenchymal stem cells differentiate into endothelial cells and construction of tissue-engineered heart valves in vitro Zhou Hong, Zhang Tao 2012, 16 (27):  4979-4984.  doi: 10.3969/j.issn.2095-4344.2012.27.008 Abstract ( 371 )   PDF (591KB) ( 428 )   Save BACKGROUND: Tissue-engineered heart valves are the hotspot of artificial heart valve replacement studies. It utilizes tissue-engineering technology which can construct artificial valves by planting seed cells in valve scaffolds. The current domestic and foreign studies mainly focus on the selection of seed cells and valve scaffolds. OBJECTIVE: To explore the feasibility of constructing tissue-engineered heart valves in vitro by seeding endothelial cells differentiated from adipose derived mesenchymal stem cells in decellularized porcine aortic valve scaffolds. METHODS: The human adipose tissue was collected by liposuction. Then the adipose derived mesenchymal stem cells were isolated and cultured, and the cell phenotype was identified by flow cytometry. The cell differentiation markers were detected by immunocytochemistry and RT-PCR. The decellularized porcine aortic valve scaffolds were made by Triton X-100 combined with trypsin digestion procedure. Tissue-engineered heart valves were constructed by seeding the endothelial cells differentiated from adipose derived mesenchymal stem cells on the decellularized porcine aortic valve scaffolds. Morphological characteristics of tissue-engineered heart valves were observed under light and electron microscopes. RESULTS AND CONCLUTION: The endothelial cells differentiated from adipose derived mesenchymal stem cells could express endothelial markers, such as CD31, CD34, CD144 and Ⅷ factor and endothelial nitric oxide synthetase (eNOS). Porcine cells could no longer be detected on decellularized porcine aortic valve scaffolds with elastic fibers and collagenous fibers were well preserved. Endothelial cells formed a continuous monolayer on the surface of the tissue-engineered heart valves. Results indicate that the endothelial cells differentiated from adipose derived mesenchymal stem cells have some endothelial function and can proliferate well on the decellularized porcine aortic valve scaffolds. It is feasible to construct tissue-engineered heart valves in vitro by planting the endothelial cells differentiated from adipose derived mesenchymal stem cells on the decellularized porcine aortic valve scaffolds. Related Articles | Metrics

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Osteogenic induction of adipose derived stromal cells in bone tissue engineering in vivo Li Xiao-yu, Yao Jin-feng, Liu Zheng-hua, Cai Jian-bo 2012, 16 (27):  4985-4990.  doi: 10.3969/j.issn.2095-4344.2012.27.009 Abstract ( 155 )   PDF (577KB) ( 331 )   Save BACKGROUND: Osteogenic induction has been regarded as an indispensable step for adipose-derived stromal cells (ADSCs) to transform into osteoblasts, thus to secret bone matrix to repair bone defects. However, this conclusion has not been confirmed. OBJECTIVE: Tissue-engineered constructs were prepared from osteogenically induced ADSCs and scaffolds. The constructs were implanted into bone defects and non-bone areas. Whether osteogenically induced ADSCs were transformed into osteoblasts was determined according to bone formation. METHODS: ADSCs were prepared by collagenase Ⅰ digestion of subcutaneous fat from the back of dogs aged 12 months. Passage 3 ADSCs were harvested and seeded on the biphasic calcium phosphate ceramics to form a cell-scaffold construct. Box-like bone defects, 20 mm in length and 10 mm in height, were created in the mandibular bilaterally in each dog along the alveolar bone where teeth had been extracted. Then the cell-scaffold constructs were implanted in dog mandibular bone defects. An empty defect group was used as control. The tissue-engineered constructs and osteoinductive calcium phosphate ceramics were implanted in muscle. After implantation for 6 and 12 weeks, bone formation was analyzed using histomorphometry. RESULTS AND CONCLUSION: ADSCs-scaffold construct promoted bone formation in bone defect area, but not in muscle area, however, osteoinductive calcium phosphate ceramics promoted bone formation in muscle area. ADSCs were not transformed into osteoblasts after osteogenic induction. The precise mechanism needs further investigation. Related Articles | Metrics

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Cardiac-specific gene expression in cardiomyocytes differentiated by rabbit adipose-derived stem cells under 5-azacytidine induction Song Yan-qin, Zhang Yan-mei, Wang Ni-ni, Xiu Jin, Zhang Rui-ping 2012, 16 (27):  4991-4995.  doi: 10.3969/j.issn.2095-4344.2012.27.010 Abstract ( 239 )   PDF (432KB) ( 308 )   Save BACKGROUND: Intramyocardial transplantation of adipose-derived stem cells using biological technique for repair of injured myocardial tissue is a gradually increasing research field of cardiovascular disease. There has been no report describing cardiac-specific gene expression in cardiomyocytes induce-differentiated by rabbit adipose-derived stem cells. OBJECTIVE: To investigate cardiac-specific gene expression in cardiomyocytes differentiated by rabbit adipose-derived stem cells under 5-azacytidine induction. METHODS: Passage 2 adipose-derived stem cells were treated with 2.5, 5, 10, 20, 40, 80 μmo1/L 5- azacytidine for 24 hours and cultured with complete medium without 5-azacytidine. Four weeks later, cellular growth and morphological changes were observed. At the first, second, third and fourth weeks, the mRNA expression of atrial natriuretic peptide, brain natriuretic peptide and α-skeletal actin in adipose-derived stem cells was detected by RT-PCT. RESULTS AND CONCLUSION: After 5-azacytidine induction for 1 week, the majority of cells were arranged in parallel, with increased soma, and the proportion of shuttle-shaped cells was decreased, in particular in the 40, 80 μmol/L 5- azacytidine groups. At the fourth week, cell refraction was attenuated, cell activity was weakened, and more cells died in the 40, 80 μmol/L 5-azacytidine groups. The mRNA expression of atrial natriuretic peptide, brain natriuretic peptide and α-skeletal actin in adipose-derived stem cells increased gradually. The mRNA expression of atrial natriuretic peptide, brain natriuretic peptide and α-skeletal actin in the 10 μmol/L 5-azacytidine group was significantly higher than that in the other groups (P < 0.05). 5-azacytidine induced rabbit adipose-derived stem cells show cardiac-specific gene expression. Related Articles | Metrics

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Rapid accumulation of autologous adipose-derived stem cells promotes early vascularization of implanted allogeneic bone Yin Jian, Yang Yi, Yang Xiao-feng, Cao Xin, Deng Wei, Deng Qiang, Chai Li-ming, Jin Ge-le 2012, 16 (27):  4996-5000.  doi: 10.3969/j.issn.2095-4344.2012.27.011 Abstract ( 207 )   PDF (497KB) ( 444 )   Save BACKGROUND: It is not yet available to construct tissue-engineered bone quickly for repair of bone defects in the clinic, and early vascularization of the implants remains to be paid increasing attention. OBJECTIVE: The rapidly accumulated autologous adipose-derived stem cells were directly placed around the implants to investigate the effects on early vascularization of implants using positron emission tomography PET/CT and to search a method to rapidly construct tissue-engineered bone in the clinic. METHODS: Forty-five male New Zealand rabbits were randomly and evenly divided into three groups. These groups were implanted respectively in the lumbar 5,6 intertransverse space with allograft iliac bone (allograft group), allograft iliac bone composited with rapidly accumulated adipose-derived stem cells (compound group), and autologous iliac bone (autograft group). Each group was injected with 18F-NaF at 1, 3, 5 weeks post-surgery and detected by positron emission tomography PET/CT to contrast the standard uptake value. RESULTS AND CONCLUSION: At 1, 3, 5 weeks post-surgery, the distribution of radioactivity in the middle-segment of allogeneic bone was low, with poor improvement in blood supply and metabolism. The standard uptake value of 18F-NaF in the autograft group and compound group was significantly higher than that in the allograft group respectively (P < 0.05). It is demonstrated that rapid accumulation of autologous adipose-derived stem cells may promote early vascularization of allogeneic bone and improve the blood supply and metabolism in the implanted area. It is also a novel idea to rapidly construct tissue-engineered bone in the clinic. Related Articles | Metrics

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Effects of high glucose and high lipid on proliferation and apoptosis of human umbilical cord mensenchymal stem cells Xu Wei, Yu Wen-long, Gao Hong, Wang Li, Liu Xiang-ping, Wang Yan-gang 2012, 16 (27):  5001-5005.  doi: 10.3969/j.issn.2095-4344.2012.27.012 Abstract ( 209 )   PDF (509KB) ( 414 )   Save BACKGROUND: Several studies have confirmed that the curative effects of mesenchymal stem cell transplantation into diabetes patients in vivo are influenced by many factors, in which blood glucose and lipids play a key role. OBEJECTIVE: To simulate in vivo environment of high glucose and high lipid and further validate the effects of high glucose and high lipid on umbilical cord mesenchymal stem cells cultured in vitro. METHODS: Human umbilical cord mesenchymal stem cells were cultured using cell culture medium containing different concentrations of glucose and palmitic acid. Then the cells were divided into seven groups: control, low glucose, low lipid, low glucose+low lipid, high glucose, high lipid, and high glucose+high lipid. After intervention for 48 hours, cell proliferation was measured by CCK-8 method and cell apoptosis was detected by Annexin-V/PI method. RESULTS AND CONCLUSION: Under the condition of high glucose and high lipid, cell proliferation was significantly decreased (P < 0.01), while cell apoptosis was significantly increased (P < 0.01) after short-term culture compared with the control group. With increasing blood glucose and blood lipid concentration, cell proliferation was significantly inhibited, and cell apoptosis was significantly increased. In the low glucose + low lipid group and high lipid group, cell proliferation was inhibited (both P < 0.05) and cell apoptosis was increased (both P < 0.01). High glucose promoted the growth of human umbilical cord mesenchymal stem cells (P < 0.01). Results showed that diabetes patients exhibited a high glucose and high lipid in vivo environment, which inhibit the growth of human umbilical cord mesenchymal stem cells. Prior to transplantation of mesenchymal stem cells, control of patient’s blood glucose and lipid level would benefit for the functioning of transplanted stem cells. Related Articles | Metrics

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Detection of BCR/ABL fusion gene in bone marrow-derived Flk1+CD31-CD34- mesenchymal stem cells from chronic myelogenous leukemia patients Zhu Xi-shan, Zhang Ge, Shi Wei, Tai Wei-ping 2012, 16 (27):  5006-5010.  doi: 10.3969/j.issn.2095-4344.2012.27.013 Abstract ( 245 )   PDF (392KB) ( 427 )   Save BACKGROUND: The in vivo and in vitro experiment has confirmed that Flk1+CD31-CD34- mesenchymal stem cells can differentiate into hematopoietic and endothelial cells at the single cells level. OBJECTIVE: To observe the expression of BCR/ABL fusion gene from the bone marrow of chronic myelogenous leukemia patients. METHODS: The primitive bone marrow-derived mesenchymal stem cells from chronic myelogenous leukemia patients were cultured and amplified in vitro. The growth curve was detected and the cell cycle and immune phenotype were analyzed. The expression of BCR/ABL fusion gene was detected by RT-PCR and FISH. RESULTS AND CONCLUSION: The primitive bone marrow-derived mesenchymal stem cells from chronic myelogenous leukemia patients showed a fibroblast-like morphology, most of these cells were in the G0/G1 phases and had high expansion of Flk1, CD13, CD29, CD44. The expression of BCR/ABL fusion gene could be identified by RT-PCR and FISH. The transformation of leukemia gene in chronic myelogenous leukemia may occur on blood cells and endothelial cells which were higher than hemangioblastic cells. Related Articles | Metrics

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Hyperbaric oxygenation for theraputic angiogenesis acquired by transplantation of vascular endothelial growth factor gene modified bone marrow mesenchymal stem cells in rats with myocardial infarction Gao Feng, Zhang Jin-bao, Zhou Kai, Ouyang Hui, Tang Ke, Wu Xiao-chen, He Chun-yang, Liu Wei-yong 2012, 16 (27):  5011-5016.  doi: 10.3969/j.issn.2095-4344.2012.27.014 Abstract ( 246 )   PDF (592KB) ( 408 )   Save BACKGROUND: The pathological changes of bone marrow mesenchymal stem cells after transplantation into peri-infarct myocardium including ischemia, hypoxia, inflammation, and cell apoptosis greatly influence the survival of transplanted bone marrow mesenchymal stem cells, while hyperbaric oxygenation can significantly improve the hypoxic state. OBJECTIVE: To investigate the effects of hyperbaric oxygenation for therapeutic angiogenesis acquired by transplantation of vascular endothelial growth factor gene modified bone marrow mesenchymal stemcells in rats with myocardial infarction. METHODS: Rat bone marrow mesenchymal stem cells transfected with eukaryotic expression vector pcDNA3.1(-)/VEGF165 were transplanted into the ischemic tissue of a rat model of myocardial infarction. Prior to transplantation, rat bone marrow mesenchymal stem cells were labeled with CM-Dil. Within 2 weeks after transplantation, hyperbaric oxygenation was performed daily. At 1 month after transplantation, the left ventricular ejection fraction was measured by ultrasonic cardiogram, the density of newly formed vessels was determined by histochemical, hematoxylin-eosin, and Ⅷ factor staining. RESULTS AND CONCLUSION: CM-DiI could label nearly 100% bone marrow mesenchymal stem cells. At 1 month after transplantation, the left ventricular ejection fraction and the density of newly formed vessels in rats subjected to hyperbaric oxygenation were significantly increased (P < 0.05). Hyperbaric oxygenation after cell transplantation results in remarkable therapeutic angiogenesis and improves cardiac function by transplantation of vascular endothelial growth factor gene-modified bone marrow mesenchymal stem cells. Related Articles | Metrics

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Persistent expression of pIRES-bone morphogenetic protein 2 plasmid after transfection into rat bone marrow mesenchymal stem cells Sun Li1, Tian Xiao-bin1, Hu Ru-yin, Wang Yuan-zheng, Tian Jia-liang, Han Wei, Li Bo, Yuan Jun, Nie Ying-jie 2012, 16 (27):  5017-5021.  doi: 10.3969/j.issn.2095-4344.2012.27.015 Abstract ( 180 )   PDF (424KB) ( 397 )   Save BACKGROUND: Recombinant human bone morphogenetic protein has been widely used in bone tissue engineering for treatment of bone defects or bon nonunion. But direct application of exogenous bone morphogenetic protein for repair of bone defects yields unsatisfactory effects. OBJECTIVE: To investigate persistent secretion and expression of bone morphogenetic protein in rat bone marrow mesenchymal stem cells transfected with pIRES-bone morphogenetic protein. METHODS: Rat bone marrow mesenchymal stem cells were isolated and cultured by the whole bone marrow adherence method. Under the liposome induction, eukaryotic expression plasmid pIRES-bone morphogenic protein 2 was introduced into rat bone marrow mesenchymal stem cells. RESULTS AND CONCLUSION: ELISA results showed that with the prolonged transfection time of pIRES-bone morphogenetic protein, bone morphogenetic protein 2 secreted by rat bone marrow mesenchymal stem cells was gradually increased, peaked at 10-12 days after transfection, and was relatively much at 15 days after transfection. These findings suggest that after transfected by pIRES-bone morphogenetic protein 2, rat bone marrow mesenchymal stem cells can persistently and stably secrete bone morphogenetic protein 2. Related Articles | Metrics

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Culture of human peripheral blood endothelial progenitor cells by autologous serum Zhang Peng, Ren Yu-sheng, Liang Chun, Leng Bing, Cao Zhi-yong, Wu Zong-gui 2012, 16 (27):  5022-5027.  doi: 10.3969/j.issn.2095-4344.2012.27.016 Abstract ( 198 )   PDF (594KB) ( 296 )   Save BACKGROUND: Traditional method for culture of endothelial progenitor cells from human peripheral blood is complex to operate, costs high, and acquires a small number of cells. OBJECTIVE: To culture endothelial progenitor cells from human peripheral blood using autologous serum and identify their functions. METHODS: Peripheral blood was collected from volunteers. Mononuclear cells were separated by density centrifugation and were induced to differentiate into endothelial progenitor cells in vitro. The influences of different culture conditions (EGM-2MV, M199+10% autologous serum+insulin like growth factor, M199+10% fetal bovine serum+vascular endothelial growth factor+basic fibroblast growth factor+insulin-like growth factor+epidermal growth factor) on the proliferation and migration of endothelial progenitor cells were observed. The cultured endothelial progenitor cells were identified by cell morphology observation, fluorescent staining and flow cytometry. RESULTS AND CONCLUSION: Endothelial progenitor cells with higher proliferation and migration ability were obtained under EGM-2MV and M199+10% autologous serum + insulin-like growth factor conditions (P < 0.05). Furthermore, the percentage of double positive cells stained by Dil-labeled acetylated low density lipoprotein and FITC-labeled Ulex europaeus agglutinin 1 was higher than 80%, and these cells also expressed CD34, CD133 and KDR. Addition of autologous serum to M199 culture medium is a simple, highly efficient method for culturing endothelial progenitor cells. Related Articles | Metrics

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Expression of replication-deficient adenovirus expressing rat interleukin-10 in bone marrow mesenchymal stem cells Li Shuang, Bi Liang-kuan, Zhu Bin, Tang Bing, Zhu Jia-yuan 2012, 16 (27):  5028-5033.  doi: 10.3969/j.issn.2095-4344.2012.27.017 Abstract ( 197 )   PDF (672KB) ( 519 )   Save BACKGROUND: Studies have demonstrated that interleukin-10 (IL-10) plays an important role in anti-inflammatory reaction and promoting prognosis of diseases such as sepsis. OBJECTIVE: A recombinant adenovirus containing rat IL-10 was constructed and bone marrow mesenchymal stem cells were as gene vectors to investigate exogenous gene expression in cells. METHODS: Total RNA was extracted from SD rat fresh spleen tissue by reverse transcription-polymerase chain reaction and rat IL-10 gene was cloned. After package, amplification and purified by AdEasy system, recombinant adenovirus particles containing target gene were harvested. Rat bone marrow mesenchymal stem cells were isolated and amplified using density gradient centrifugation and adherence. Phenotype identification was performed through the use of flow cytometer. Bone marrow mesenchymal stem cells were infected using different multiplicities of infection of Ad.rIL-10/Ad.GFP. Then efficiency of infection was observed under fluorescent microscope. The expression of target gene in rat bone marrow mesenchymal stem cells was detected by reverse transcription-polymerase chain reaction and western blot method. RESULTS AND CONCLUSION: The recombinant adenovirus Ad.rIL-10 was successfully constructed. The titer was calculated as 6.0×1010 pfu/mL after amplification and purification of adenovirus. Ad.rIL-10 effectively infected 70% of rat bone marrow mesenchymal stem cells with the multiplicity of infection value of 200. These findings suggest that rat bone marrow mesenchymal stem cells with rat IL-10 recombinant adenovirus can express target gene effectively and persistently. Rat bone marrow mesenchymal stem cells are optimal and promising vector for gene therapy. Related Articles | Metrics

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Biological characteristics of epidermal stem cells and follicle stem cells Yang Qing, Li Xiu-lan, Zhang Yang, Guo Yue, Sun Xiao-lei 2012, 16 (27):  5034-5038.  doi: 10.3969/j.issn.2095-4344.2012.27.018 Abstract ( 288 )   PDF (503KB) ( 592 )   Save BACKGROUND: More appropriate skin tissue engineering seed cells can better repair skin function. OBEJECTIVE: To isolate and culture epidermal stem cells and follicle stem cells, and compare their biological characteristics. METHODS: Epidermal stem cells and follicle stem cells of 2-month-old New Zealand rabbits were isolated and cultured in vitro. The biological characteristics of passage 2, 3, 6 cells which grew well were observed. RESULTS AND CONCLUSIONS: Follicle stem cells attached faster and had higher proliferative capacity than epidermal stem cells. Immunohistochemical staining and fluorescent quantitative PCR showed that the follicle stem cells showed higher protein and mRNA expression of β1 integrin and K19 than epidermal stem cells. These findings suggest that follicle stem cells, as seed cells of skin tissue engineering, are superior to epidermal stem cells. Related Articles | Metrics

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Role of transforming growth factor β1 during transplantation of rat bone marrow mesenchynmal stem cells for repair of myocardial infarction Lin Chu-wei, Zhou Sheng-hua, Dai Hai-ying, Deng Ping, Yin Zhi-lan, Guan Xian-song, Xia Xin 2012, 16 (27):  5039-5045.  doi: 10.3969/j.issn.2095-4344.2012.27.019 Abstract ( 155 )   PDF (708KB) ( 470 )   Save BACKGROUND: Studies have demonstrated that bone marrow mesenchymal stem cells promote the accumulation of a large number of myofibroblasts in the site of transplantation by paracrine action, secrete much collagen, which contributes to repair of myocardial infaction and improvement of cardiac systolic and diastolic function. OBJECTIVE: To investigate the role of transforming growth factor β1 during transplantation of rat bone marrow mesenchynmal stem cells for repair of myocardial infarction. METHODS: (1) In vitro experiments: rat bone marrow mesenchymal stem cells were cultured in simulated micro-environment of myocardial infarction. The levels of tumor necrosis factor α, platelet derived growth factor, and transforming growth factor β1 were detected. (2) In vivo experiments: BrdU-labeled rat bone marrow mesenchymal stem cells, transforming growth factor β1 and PBS were transplanted into the infracted myocardium. The difference of collagen Ⅰ, Ⅲ expression in rat myocardium post-transplantation was determined by immunohistochemical staining, PCR, western blot methods. RESULTS AND CONCLUSION: After rat bone marrow mesenchymal stem cells were cultured in simulated micro-environment for 14 days, high level of transforming growth factor β1 could be detected. These findings suggest that transplantation of rat bone marrow mesenchymal stem cells into infarct myocardium can promote the generation of collagen Ⅰ, Ⅲ and improve rat cardiac systolic and diastolic function, and transforming growth factor β1 may play an important role. Related Articles | Metrics

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Effect of stromal cell derived factor-1α preconditioning on migration of rat bone marrow mesenchymal stem cells Shu Bo, Liu Zhi-jiang, Fan Fang 2012, 16 (27):  5046-2051.  doi: 10.3969/j.issn.2095-4344.2012.27.020 Abstract ( 190 )   PDF (487KB) ( 343 )   Save BACKGROUND: During transplantation of bone marrow mesenchymal stem cells (BMSCs), only a few BMSCs can migrate into injured tissue. Therefore, how to increase the number of migrated BMSCs is a key to BMSC transplantation. OBJECTIVE: To investigate the effect of stromal cell derived factor-1α (SDF-1α) preconditioning on the migration of rat BMSCs. METHODS: BMSCs were in vitro isolated, cultured, and passaged. Using Transwell inserts technique, the effects of SDF-1α at different concentrations on the migration of BMSCs were observed and the optimal concentration of SDF-1α was determined. After preconditioned by SDF-1α, SDF-1α+AMD3100 and SDF-1α+LY294002 respectively, the migration of BMSCs was observed, and the effect of SDF-1α preconditioning on mRNA and protein level of chemokine receptor CXCR 4 in BMSCs and AKT phosphorylation were investigated. RESULTS AND CONCLUSION: Incubation of 0.2 mg/L SDF-1α for 10 hours could produce the optimal migration of BMSCs. The role of SDF-1α in inducing the migration of BMSCs increased with the increasing concentration of SDF-1α used for 6 hours of preconditioning. AMD3100 and LY294002 could reverse the effect of SDF-1α preconditioning. The chemokine receptor CXCR 4 expression and Akt phosphorylation were increased after SDF-1α preconditioning. SDF-1α preconditioning can enhance the migration of BMSCs through increasing the expression of chemokine receptor CXCR 4. Akt signaling pathway is correlated with this migration. Related Articles | Metrics

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Influence of secreted cytokines of bone marrow mesenchymal stem cells on the stability of rat peritoneal mast cell membrane Gao Ming-min, Liu Xiao-ping, Yu Ye-jun, Jia Yue-wei, Wang Jia-hui, Li Wei 2012, 16 (27):  5052-5056.  doi: 10.3969/j.issn.2095-4344.2012.27.021 Abstract ( 260 )   PDF (437KB) ( 301 )   Save BACKGROUND: Mesenchymal stem cells secreted cytokines has immunomodulatory effects on specific and nonspecific immune cells. Whether the cytokines have an effect on mast cells requires further studies. OBJECTIVE: To explore the influence of the cytokines secreted by bone marrow mesenchymal stem cells on the degranulation of specific and nonspecific induced mast cells and secretion of inflammatory mediators. METHODS: Rat bone marrow mesenchymal stem cells were isolated and purified by whole bone marrow adherent method. The stem cell cytokines were obtained from the supernatants of the third generation of bone marrow mesenchymal stem cells. The mast cells were isolated from rat peritoneal mast cells. Mast cells were specifically sensitized by rat antiovalbumin serum and nonspecifically stimulated by Compound48/80. RESULTS AND CONCLUSION: Compared with the control group, cytokines from bone marrow mesenchymal stem cells and ketotifen could significantly inhibit the degranulation of the specific and nonspecific mast cells and release tryptase, and there was significant difference (P < 0.05), however, the effect of cytokines from bone marrow mesenchymal stem cells was weaker than that from the ketotifen (P < 0.05). Cytokines from bone marrow mesenchymal stem cells could stable mast cells membrane, inhibit the degranulation of mast cells and the release inflammatory mediators in mast cells. Related Articles | Metrics

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Effect of curcumin on osteogenic differentiation of rat bone marrow mesenchymal stem cells Gu Qiao-li, Cai Yan, Huang Chen, Yang Hui-lin 2012, 16 (27):  5057-5061.  doi: 10.3969/j.issn.2095-4344.2012.27.022 Abstract ( 259 )   PDF (404KB) ( 407 )   Save BACKGROUND: Curcumin can significantly reduce the bone resorption of osteoblasts, inhibit osteoclastogenesis and induce apoptosis in ostebclasts. OBJECTIVE: To investigate the effects of different concentrations of curcumin on osteoblast differentiation of rat bone marrow mesenchymal stem cells. METHODS: Rat bone marrow mesenchymal stem cells were isolated by adherent method. After subcultured to the third generation, osteogenic induction was performed with osteogenic culture medium. 0 (control), 10 and 15 μmol/L curcumin was added into the osteogenic medium at 3 days before induction. RESULTS AND CONCLUSION: After seeded and cultured for 7 days, scattered small colonies of bone marrow mesenchymal stem cells could be seen under the inverted microscope, and the colonies were in radial growth. As the culture time prolonged, the cell colonies were increased and the morphology of the cells was spindle shaped. At 7 days after addition of osteoblast medium, cells changed from spindle-shaped to cuboid and were more apparent by days. After addition of different concentrations of curcumin, there was no difference in the proliferation of bone marrow mesenchymal stem cells. Curcumin could increase the activity of alkaline phosphatase in bone marrow mesenchymal stem cells and the expression of Runx-2 and heme monooxygenase 1 in a dose-dependent manner. These findings demonstrate that curcumin can promote osteogenic differentiation of rat bone marrow mesenchymal stem cells. The effect of curcumin on osteogenic differentiation of rat bone marrow mesenchymal stem cells is correlated with heme monooxygenase 1 expression. Related Articles | Metrics

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Establishment of Dami cell model of SP600125-induced polyploidization which was reversed by curcumin Li Yan-ping, Zhou Fan, Ma Dong-chu, Jiang Zhi-ming, Liu Yan-qin, Liu Jing-hua, Wang Ji-gang 2012, 16 (27):  5062-5066.  doi: 10.3969/j.issn.2095-4344.2012.27.023 Abstract ( 210 )   PDF (527KB) ( 380 )   Save BACKGROUND: Cell polyploidization is an important biological phenomenon. It not only influences the normal physiological function of tissue and hematopoietic system, but also is the compensatory performance under stress condition. It is closely related to the occurrence of some pathological processes, in particular malignant tumorigenesis. OBJECTIVE: To optimize the Dami cell model of SP600125-induced polyploidization and to investigate the mechanism by which curcumin reversed the polyploidization. METHODS: 15, 60, 60 μmol/mL SP600125 was added to the cell suspension to induce the polyploidization of Dami cells. The optimal induction time and concentration were determined. Then 5, 10, 20 mmol/L curcumin was added to the cell suspension containing SP600125. Cells were collected after 72 hours of culture. Cell ploidy changes were determined by flow cytometry using Annexin V-PI staining. Cell cycle-related protein expression changes were detected by Western blot method. RESUITS AND CONCLUSION: After treated with 30 mmol/L SP600125 for 72 hours, the number of polyploid Dami cells was increased and the expression of cyclin D3 was highest. The ployploidization of Dami cells was significantly reversed by curcumin, and the expression of cyclin D3 was gradually decreased, showing a significant time- and dose-dependent manner. A Dami cell model of polyploidization was successfully established and the optical induction time and concentration for SP600125 were determined at 72 hours and 30 mmol/L. The polyploidization of Dami cells may be reversed through inhibiting cyclinD3 expression. Related Articles | Metrics

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Survival of human adipose-derived mesenchymal stem cells transplanted via the tail vein in rat brain with cerebral ischemia Gao Xu, Dong Tao, Li Zhi-qing, Jiang Wei, Wei Xue-zhong, Cao Peng, Liang Guo-biao 2012, 16 (27):  5067-5071.  doi: 10.3969/j.issn.2095-4344.2012.27.024 Abstract ( 213 )   PDF (463KB) ( 296 )   Save BACKGROUND: Mesenchymal stem cells transplantation can improve the neurological function of ischemic cerebrovascular disease. OBJECTIVE: To explore the effect of human adipose-derived mesenchymal stem cells transplanted via the tail vein on functional recovery in rats with focal cerebral ischemia. METHODS: Focal cerebral ischemia rat models were established, and then rat models were divided into experimental group and control group. In the experimental group, 5×106 adipose-derived mesenchymal stem cells were injected into rat models via the tail vein, and the rat models in the control group were treated without cell transplantation. RESULTS AND CONCLUSION: Behavioral score of neurological defects in two groups was gradually reduced with the time prolonged after cell transplantation. At 14 and 21 days after transplantation, the behavioral score of rats in the experimental group was significantly lower than that in the control group (P < 0.05) and the improvement of experimental group was significantly better than that of the control group (P < 0.01); transplanted adipose-derived mesenchymal stem cells were gathered and survived surrounding the blood vesselss and periischemic areas in rats with focal cerebral ischemia. Adipose-derived mesenchymal stem cells transplanted via the tail vein can survive in the host brain and improve neurological function. Related Articles | Metrics

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Long term intranasal delivery of bone marrow stromal cells improves rat brain injury caused by ischemia reperfusion Wang Yang, Shen Hui-juan, Zhu Qun-e, Wang Fei, Xu Wei-zhen, Yu Qian, Zeng Xian-zhi 2012, 16 (27):  5072-5075.  doi: 10.3969/j.issn.2095-4344.2012.27.025 Abstract ( 186 )   PDF (1199KB) ( 293 )   Save BACKGROUND: Nasal delivery is a highly effective way to deliver drug into the brain, which can overcome the blockage of blood-brain barrier and peripheral side-effects. OBJECTIVE: To explore the feasibility of long term intranasal delivery of bone marrow stromal cells for treatment of brain injury caused by ischemia reperfusion. METHODS: Bone marrow stromal cells were sorted from rat whole bone marrow by adherent culture. Brain ischemia reperfusion rats were randomly divided into experimental group or control group. Bone marrow stromal cells or phosphate buffer saline were dropped into the nasal cavity of experimental group and control group respectively, once every 2 days for a total of 4 weeks. Rat behaviors were assessed once a week. Brain pathological examination was performed after the last behavior assessment. RESULTS AND CONCLUSION: Compared with the control group, the results of modified neurological severity score and Morris water maze test showed that rat behaviors of experimental group were gradually improved at 2 weeks after treatment (P < 0.05, P < 0.01). It corresponded to behavior improvement, and ischemia reperfusion resulted in 66% cell loss in hippocampal CA1 region in the control group; however, in the experimental group, the cell loss in hippocampal CA1 region was significantly lower to 25% (P < 0.01). Our data indicated that long term intranasal delivery of bone marrow stromal cells can improve the brain injury caused by ischemia reperfusion. Related Articles | Metrics

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Effects of ulinastatin application combined with umbilical cord mesenchymal stem cells transplantation on changes in motor evoked potential and motor function of hindlimbs in rats with spinal cord injury Zhang Zhan-xiu, Li Zhi-yuan, Shen Yong 2012, 16 (27):  5076-5080.  doi: 10.3969/j.issn.2095-4344.2012.27.026 Abstract ( 182 )   PDF (440KB) ( 279 )   Save BACKGROUND: Ulinastatin can reduce the inflammatory response, and get rid of oxygen free radicals. Ulinastatin has a protective effect on the central nervous system injury, and can effectively increase the survival rate of transplanted cells after spinal cord injury. OBJECTIVE: To investigate the effects of umbilical cord mesenchymal stem cells transplantation and ulinastatin on the repair of motor function of hindlimbs in rats with spinal cord injury. METHODS: Wistar rats were used to establish the spinal cord injury models and the models were randomly divided into four groups: control group, umbilical cord mesenchymal stem cells group, ulinastatin group and combination group. The models in the control group underwent tail vein injection of culture medium and intraperitoneal injection of normal saline. The umbilical cord mesenchymal stem cells group was injected with umbilical cord-matrix stem cells via the tail vein. The ulinastatin group underwent intraperitoneal injection of ulinastatin and the combination group underwent tail vein injection of umbilical cord mesenchymal stem cells and intraperitoneal injection of ulinastatin. RESULTS AND CONCLUSION: At 4 weeks after transplantation, the function of rat hindlimbs in the combination group was superior to that in the umbilical cord mesenchymal stem cells transplantation group and the ulinastatin group (P < 0.05), and the hindlimb function was superior in the umbilical cord mesenchymal stem cells transplantation group and the ulinastatin group to that in the control group (P < 0.05). The number of PKH26-positive cells in the combination group was greater than that in the umbilical cord mesenchymal stem cells transplantation group, and the number was also significantly greater in the umbilical cord mesenchymal stem cells transplantation group than that in the ulinastatin group and control group (P < 0.01). At 8 weeks after transplantation, the latency and amplitude of somatosensory evoked potential and motor evoked potential in the combination group were significantly better than those in the other three groups (P < 0.05, P < 0.01). Umbilical cord mesenchymal stem cells transplantation combined with the ulinastatin can promote the regeneration of nerve synapses in rats with spinal cord injury, and can improve their motor function and electrophysiological function. The umbilical cord mesenchymal stem cells transplantation combined with the ulinastatin produces better effects than single use in repair of motor function of hundlimbs of rats with spinal cord injury. Related Articles | Metrics

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Autologous bone marrow mesenchymal stem cells for myocardial renewal and repair Xiao Wen-tao, Gao Chuan-yu, Dai Guo-you, Li Mu-wei, Wang Xian-pei, Liu Hong-zhi, Qi Da-tun 2012, 16 (27):  5081-5086.  doi: 10.3969/j.issn.2095-4344.2012.27.027 Abstract ( 221 )   PDF (592KB) ( 403 )   Save BACKGROUND: Current clinical trials of stem cell transplantation focus on the research field of myocardial infarction. Few studies are about primary cardiomyopathy, and there have been no reports describing the comparison between myocardial infarction and primary cardiomyopathy. OBJECTIVE: To assess the efficacy and safety of intracoronary transplantation of autologous bone marrow mesenchymal stem cells for treatment of non-functional myocardium caused by ischemic or non-ischemic diseases. METHODS: 38 patients who underwent selective percutaneous coronary intervention due to myocardial infaction were divided into trial A group and control A group randomly. 36 patients with dilated cardiomyopathy were divided into trial B group and control B group randomly. Equal amounts of normal saline or bone marrow mesenchymal stem cells were infused into corresponding coronary artery through the guiding catheter post-operation in control groups and trial groups respectively. RESULTS AND CONCLUSION: At 1 month after transplantation, compared with prior to transplantation and control groups, only left ventricular ejection fraction in the trial groups increased significantly (P < 0.05). At 3 months after transplantation, compared with prior to transplantation and control groups, the left ventricular ejection fraction in the trial groups increased significantly while myocardial perfusion defects in the trial groups were significantly reduced (P < 0.05); however, at this time point, there was significant difference in the left ventricular ejection fraction between trial groups (P < 0.05); the left ventricular end-diastolic diameter in the trial A group decreased significantly compared with prior to transplantation, control A group and trial B group (P < 0.05), while the trail B group showed significantly decreased left ventricular end-diastolic diameter only compared with prior to transplantation (P < 0.05). There was no significant difference in incidence of malignant cardiovascular events between trail groups and control groups during follow-up period (P > 0.05). Intracoronary transplantation of bone marrow mesenchymal stem cells is safe and effective in renewal and repair of non-functional myocardium and its efficacy in acute myocardial infarction patients is better than dilated cardiomyopathy patients. Related Articles | Metrics

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Construction of lentivector containing alpha-smooth muscle actin promoter by multisite Gateway technology Yuan Xiao-feng, Xiang Peng, Li Wei-qiang, Hu Xiao-jun, Peng Chao-quan 2012, 16 (27):  5087-5091.  doi: 10.3969/j.issn.2095-4344.2012.27.028 Abstract ( 233 )   PDF (490KB) ( 415 )   Save BACKGROUND: Alpha-smooth muscle actin gene is one of several genes expressed in smooth muscle cells and has been recognized as the marker for smooth muscle cell phenotype transformation. OBJECTIVE: To construct a recombinant pLVpuro/αSMA-hrGFP lentiviral vector by multisite Gateway technology. METHODS: Primers containing attB sites were designed and used to amplify the alpha-smooth muscle actin (αSMA) promoter fragment by PCR from the plasmid containing the mouse αSMA promoter sequence (SMP8-Cre). By the BP recombination reaction, the attB flanked PCR product containing αSMA promoter sequence was cloned to an attP-containing pDONR P4P1r donor vector to create an entry clone, pUp-αSMA. Finally, pUp-αSMA and pDown-hrGFP were shuttled into the destination vector pDEST-puromycin by LR recombination reaction to generate pLVpuro/αSMA-hrGFP. The expression vector was confirmed by PCR and gene sequencing. Then this expression vector was transferred into the C2C12 cell line, and the gene expression was confirmed by immunofluorescent staining. RESULTS AND CONCLUSION: The pLVpuro/αSMA-hrGFP expression vector was successfully constructed with the right αSMA promoter fragment confirmed by sequencing. The activity of αSMA promoter was verified by cell transfection and immunofluorescent staining. The recombinant pLVpuro/αSMA-hrGFP lentivector was successfully constructed. And it offers an important tool to monitor the differentiation process from embryonic stem cells to vascular smooth muscle cells, and for cell tracing and gene function studies. Related Articles | Metrics

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Immune function detection of umbilical cord blood from newborns of different genders between cesarean section and vaginal birth Liu Ke-qin, Liu Chen, Yin Wei-dong, Zhang Bin 2012, 16 (27):  5092-5096.  doi: 10.3969/j.issn.2095-4344.2012.27.029 Abstract ( 230 )   PDF (412KB) ( 478 )   Save BACKGROUND: There have been many factors that influence neonatal immune function. In vaginal birth, uterine contraction makes the newborn child in a relative ischemic hypoxic status, and the type of delivery can influence the immune status of the newborn children. The newborn children delivered by cesarean section have low immunity and predispose to secondary infection. OBJECTIVE: To explore the influence of cesarean section versus vaginal birth on immune function of umbilical cord blood of newborn children of different genders. METHODS: 100 cases of women who had full-term single birth were selected, including 50 cases subjected to vaginal delivery (30 male babies and 20 female babies) and 50 cases subjected to cesarean section (30 male babies and 20 female babies). Umbilical cord blood immunoglobulin (IgG, IgA, IgM), the complement (C3, C4), C-reactive protein of newborn children were detected by transmission immune turbidity method using automatic biochemical analyzer. T lymphocyte antigen (CD4, CD8) was determined by flow cytometry. RESULTS AND CONCLUSION: In the cesarean section group, the level of umbilical cord blood lymphocyte CD4 antigen and the mixed antigen CD4 and CD8 in female babies was significantly higher (P < 0.01), but CD8 antigen level was significantly lower (P < 0.01), than in the vaginal birth group. In the cesarean section group, umbilical cord IgA and IgM level in female babies was significantly higher (P < 0.01), but IgG level was significantly lower (P < 0.05) than in male babies. Umbilical cord IgG, IgM, C-reactive protein levels of female babies in the cesarean section group were significantly lower than in the vaginal birth group (P < 0.01). In the cesarean section group, umbilical cord IgM level of male babies was significantly lower (P < 0.01), but C-reactive protein level of male babies was significantly higher (P < 0.01), than in the vaginal birth group (P < 0.01). These findings suggest that the type of delivery produces different influences on immune function of newborn children of different genders, and cesarean section can decrease the immune function of newborn children. Related Articles | Metrics

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Effect of quality and quantity of blood taken from unilateral and bilateral iliac on the concentration of bone marrow mononuclear cells and the efficacy of bone marrow mononuclear cells transplantation for the treatment of early femoral head necrosis Shi Shao-hui, Li Zi-rong, Sun Wei, Wang Bai-liang, Xu Shu-qing, Wang Ran-dong 2012, 16 (27):  5097-5102.  doi: 10.3969/j.issn.2095-4344.2012.27.030 Abstract ( 279 )   PDF (330KB) ( 257 )   Save BACKGROUND: Whether the quality and quantity of blood taken from unilateral and bilateral iliac would affect the concentration of bone marrow mononuclear cells? OBJECTIVE: To observe the efficacy of mononuclear cells concentration on early femoral head necrosis treated by autologous bone marrow mononuclear cells transplantation. METHODS: The clinical data of 120 patients of 168 hips with femoral head necrosis were retrospectively analyzed, and all the patients were processed with core decompression plus autologous mononuclear cells transplantation. This study included 27 cases (39 hips), who were taken blood about 200 mL from the unilateral iliac, 29 cases (40 hips), who were taken blood about 300 mL from the unilateral iliac, 30 cases (42 hips), who were taken blood about 200 mL from the bilateral iliac, 34 cases (47 hips), who were taken blood about 300 mL from the bilateral iliac. After the blood was taken from lilac, the bone marrow blood gradient centrifugation was performed by cell segregating, and the mononuclear cells count was observed before and after separation; the mononuclear cells were injected into the bone necrosis area after core decompression. All the patients were followed-up for 12-36 months, and the CT imaging and the Harris score of the hips in the patients before and after treatment were observed. RESULTS AND CONCLUSION: There was no significant difference of the mononuclear cells count between subgroup of 200 mL and 300 mL taken from unilateral and bilateral iliac (P > 0.05), but the mononuclear cells count taken from the bilateral iliac was significantly higher than that taken from the unilateral iliac (P < 0.01). There was no significant difference of the Harris score between groups before transplantation (P > 0.05), and compared with before transplantation, the Harris score and the hip fineness rate was significantly increased after transplantation (P < 0.05, P < 0.01). In short, core decompression plus autologous bone marrow mononuclear cell transplantation is an easy and safe method for the treatment of early femoral head necrosis. The concentration of mononuclear cells after centrifugation when taken from bilateral iliac is significantly higher than that taken from unilateral iliac, and the better follow-up the higher concentration. Related Articles | Metrics

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Mechanism underlying endogenous neural stem cells migration following ischemic brain damage and the regulatory effect of traditional Chinese medicine Su Xiao-hui, Kong Xiang-ying, Lin Na 2012, 16 (27):  5103-5107.  doi: 10.3969/j.issn.2095-4344.2012.27.031 Abstract ( 223 )   PDF (614KB) ( 365 )   Save BACKGROUND: Migration of endogenous neural stem cells (NSCs) caused by ischemic brain injury has been a spot light in neuroscience. After ischemic brain injury, neuron becomes swelling and necrotic due to ischemia and hypoxia. NSCs that migrated to the necrosis area driven by a variety of factors can repair the brain tissue and make neural function recovered partially. OBJECTIVE: To review the mechanism underlying NSCs migration caused by ischemic brain injury and the regulatory effects of traditional Chinese medicine. METHODS: A computer-based retrieval of PubMed Database, Chinese Journal Full-text Database, China Doctor Dissertations Full-text Database and China National Knowledge Infrastructure to search articles describing the mechanism underlying NSCs migration caused by ischemic brain injury and the regulatory effects of traditional Chinese medicine published from 2000 to 2011 using the key words “cerebral ischemia, endogenous neural stem cells, migration, traditional Chinese medicine”. The repetitive studies were excluded. The retrieved literatures contained original articles, thesis and reviews. RESULTS AND CONCLUSION: A great number of articles have proven that NSCs migrate to the lesion site after brain injury. Factors such as cytokines, growth factors, chemotactic factors and inflammatory factors are involved in the migration of NSCs. Also, many studies have also confirmed that traditional Chinese medicine plays an important role in regulating the migration of endogenous NSCs. Related Articles | Metrics

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Identification of neural stem cells Zhang Jun-hu, Wang Jian-jiao, Zheng Yong-ri 2012, 16 (27):  5108-5112.  doi: 10.3969/j.issn.2095-4344.2012.27.032 Abstract ( 295 )   PDF (494KB) ( 478 )   Save BACKGROUND: Due to the less number of neural stem cells, no specific morphology and lack of surface antigen, the identification and purify of the neural stem cells are difficult. OBJECTIVE: To summarize the main method to identify the neural stem cells. METHODS: The PubMed database and Wanfang database form 1990 to 2011 were used to search the articles on the characteristics of neural stem cells, main identification method and application prospects. RESULTS AND CONCLUSION: At present, the main method to identify the neural stem cells was in vivo dye identification method, surface marker identification method, molecular biology identification method and nuclide notation. The markers included Nestin, vimentin, Musashi1, Musashi2, CD133 (AC133), sialic acid-neural cell adhesion molecule, Lewis X LeX/SSEA-1, p75 neurotrophin receptor and epidermal growth factor. The surface marker identification method, molecular biology identification method and the combination of two methods were the main methods to identify the neural stem cells. It is a possible research tools to obtain breakthrough. Related Articles | Metrics

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Adult stem cells in the endometrium Liu Zhi-zhen, Liu Dan, Xie Jun 2012, 16 (27):  5113-5117.  doi: 10.3969/j.issn.2095-4344.2012.27.033 Abstract ( 274 )   PDF (533KB) ( 378 )   Save BACKGROUND: In recent years, the studies on uterine endometrial stem cells, especially endometrial stem cells, have made great progress, many scholars have indentified the existence of endometrial stem cells through different ways. OBJECTIVE: To review the basis for the existence, location and surface markers of endometrial stem cells. METHODS: Wanfang database was used to retrieve the articles published from January 1995 to December 2010 with the key words of “endometrium stem cell” in Chinese. The PudMed database was also used to search the articles published from January 1995 to December 2010 with the key words of “endometrium stem cell” in English. A total of 233 articles were retrieved. Finally, 30 articles were selected according to the inclusion criteria. RESULTS AND CONCLUSION: In recent years, the study on the isolation and identification of endometrial stem cells has made great progress, but the specific markers of endometrial stem cells were still need to be explored. Here, we will review our current understanding of the possibility existence, the possible location and the possible surface markers of endometrial stem cells through colony-forming ability, cell mark retention experiments and tissue reconstruction capacity. Related Articles | Metrics

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Transplantation of stem cells from different sources for treatment of diabetic foot Song Ming-xian 2012, 16 (27):  5118-5125.  doi: 10.3969/j.issn.2095-4344.2012.27.034 Abstract ( 265 )   PDF (839KB) ( 342 )   Save BACKGROUND: Diabetic foot patients often exhibit limb distal ischemia and the clinical outcomes are not very satisfactory. Autologous stem cells transplantation is a safe and effective treatment method in recent years. OBJECTIVE: To explore the literature trends of stem cells transplantation for the treatment of diabetic foot. METHODS: The CNKI database from January 2003 to December 2011 was searched for the literatures related to stem cells for the treatment of diabetic foot. The key words were “stem cells, diabetic foot” and the characteristics of the data were analyzed by analysis capabilities of the database and the chart function of Excel software. The autologous bone marrow stem cells, autologous hematopoietic stem cells or umbilical cord blood stem cell transplantation were used for the treatment of diabetic foot; the cells were injected into the lower limb ischemia site through intramuscular injection or vascular infusion. The physical pain, cold sensation, skin temperature, intermittent claudication, ulcer healing degree and angiographic results were compared after treatment, and the effect of cells transplantation was assessed. RESULTS AND CONCLUSION: Among the literatures included in the CNKI database from 2003 to 2011, a total of 125 literatures related to the stem cells for the treatment of diabetic foot were screened out. The amount of the literatures showed an increasing trend, and the main content of the literatures containing the diabetic foot, stem cell transplantation and transplantation. Stem cells for the transplantation mainly include autologous cells, including bone marrow stem cells and hematopoietic stem cells. Stem cells transplantation can alleviate the symptoms of diabetic foot and release the pain, and during the transplantation, lower limb angiography shows varying degrees of neonatal collateral blood vessel formation which plays an important role in improving the lower limb ischemia of diabetic foot patients. Related Articles | Metrics

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Epidermal stem cells in wound repair by tissue engineering Lü Shu-kun 2012, 16 (27):  5126-5130.  doi: 10.3969/j.issn.2095-4344.2012.27.035 Abstract ( 226 )   PDF (571KB) ( 473 )   Save BACKGROUND: With rapid development of cell biology, molecular biology technique, and tissue engineering, increasing studies focus on seed cells for constructing skin wound surface by tissue engineering. OBJECTIVE: To summarize the biological characteristics of epidermal stem cells and investigate their regenerative and clinical application value in repair of skin wound surface. METHODS: A computer-based online retrieval of SNKI and PubMed database for searching papers describing epidermal stem cells for repair of skin injury using the key words of “epidermal stem cells, tissue engineering skin, wound surface” in English and Chinese. Papers related to research progress in epidermal stem cells for repair of skin injury were selected. Papers that were published recently in the same research field or in high-impact journals were selected. A total of 129 papers were initially retrieved. According to inclusion criteria, 25 papers were included for final analysis. RESULTS AND CONCLUSION: Epidermal stem cells support for epidermal generation, differentiation and wound healing. Epidermal stem cells provide the condition for maintaining skin normal structure and intracellular environment due to their normal proliferation and differentiation and are the ideal seed cells of skin tissue engineering. They show satisfactory prospects in treatment of large-area skin defects, cell therapy of skin disease, and gene therapy. In vitro culture of epidermal stem cells is a prerequisite for studying tissue engineering of artificial skin. As the isolation, purification and culture technology of epidermal stem cells continues to improve, we can quickly build epidermis. Nevertheless, application of epidermal stem cells requires further investigation. Related Articles | Metrics