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03 September 2010, Volume 14 Issue 36 Previous Issue    Next Issue

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Bone marrow stromal cells differentiation potentials in vitro from mdx mice versus C57 mice Leng Yan, Zhang Wei-xi, Zhou Chen, Zheng Zhen-yang, Zhang Cheng 2010, 14 (36):  6651-6656.  doi: 10.3969/j.issn.1673-8225.2010.36.001 Abstract ( 159 )   PDF (457KB) ( 320 )   Save BACKGROUND: Duchenne’s muscular dystrophy (DMD) is a fatal recessive X-linked form of muscular dystrophy characterized by progressive muscular degeneration with no certain treatment. However, the proliferation and multipotent differentiation potential of bone marrow stromal cells (BMSCs) from DMD model mice (mdx mice) and the effectiveness of BMSCs self-transplantation are needed to be further studied. OBJECTIVE: To investigate the proliferation and multipotent differentiation potential of BMSCs from mdx mice during in vitro culture. METHODS: Both C57 and mdx mice were killed to obtain the BMSCs for culturing in vitro. After Giemsa’s staining, the colony forming unit-fibroblast (CFU-f) of BMSCs assay was performed. With specific inductive mediums, we succeeded to induce the BMSCs to differentiate into osteogenesis, adipogenesis, myogenesis respectively. Their morphological characteristics were observed with microscope. Von kossa staining, oil red O staining and immunofluorescence for desmin were utilized to identify the differentiated BMSCs respectively and quantify their differentiation efficiency. After the BMSCs were induced for 1 week, the cellular total RNA of differentiated BMSCs was extracted, and then reverse transcription was performed. Real-time polymerase chain reaction (PCR) was used to quantify the gene expression of differentiated BMSCs. RESULTS AND CONCLUSION: Compared with C57 mice, both number and volume of CFU-f of BMSCs from mdx mice were smaller. Compared to C57 mice, the efficiency of osteogenesis and myogenesis of BMSCs from mdx mice was significantly lower (P < 0.01). However, the efficiency of adipogenesis of BMSCs from both groups had no statistic difference (P > 0.05). Real-time PCR showed that both of osteogenic and myogenic gene expression of BMSCs from mdx mice decreased respectively compared to C57 mice. However, adipogenic gene expression from two groups had no difference. The results indicated that proliferation and multipotent differentiation potentials of BMSCs from mdx mice declined compared with C57 mice. The defection of Dystrophin gene may contribute to it. Therefore, the autoplastic BMSCs transplantation of mdx mice will be interfered. Related Articles | Metrics

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Biological characteristics of early and late endothelial progenitor cells derived from mouse bone marrow  Deng Zheng-ming, Sun Xi-tai, Zhu Hai-tao, Ding Yi-tao 2010, 14 (36):  6657-6661.  doi: 10.3969/j.issn.1673-8225.2010.36.002 Abstract ( 97 )   PDF (345KB) ( 451 )   Save BACKGROUND: The unified identification, biological characteristics and exact role of endothelial progenitor cells (EPCs) in angiogenesis are controversial. OBJECTIVE: To explore the conditions which were suitable to isolate and culture the early and late EPCs derived from mouse bone marrow, and compare the different biological characteristics of the early and late EPCs using the method of sequence of differential adhesion. METHODS: Mononuclear cells were isolated from the mouse bone marrow by Ficoll density gradient centrifugation, and then were grown in culture flask coated fibronectin. The early and late EPCs were isolated and purified using the method of sequence of differential adhesion. We compared the morphology, uptake DiI-LDL and FITC-UEA-1, cell immunophenotype, proliferation ability and capacity to form the cavity structure of the early and late EPCs in vitro. RESULTS AND CONCLUSION: Early EPCs showed colony-like growth at early time, whereas late EPCs exhibited cobblestone-shaped morphology. Both of the early and late EPCs could phagocytize DiI-LDL and bind to FITC-UEA-1. Late EPCs had greater proliferative potential and formed vascular-like structure whereas early EPCs could not form vascular-like structure. The results suggested that there are early and late EPCs with different characteristics in bone marrow, and late EPCs had greater proliferation potential and formed vascular-like structure. Related Articles | Metrics

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Culture and identification of sheep bone marrow-derived endothelial progenitor cells Yuan Jian-ming, Guan Song-hui, Dang Rui-shan, Yang Xiang-qun, Shen Man-ru, Zhang Chuan-sen 2010, 14 (36):  6662-6666.  doi: 10.3969/j.issn.1673-8225.2010.36.003 Abstract ( 87 )   PDF (478KB) ( 408 )   Save BACKGROUND: Nowadays, the study of tissue-engineered venous valve has just started, the choice of seed cells is the key, endothelial progenitor cells are optimal seed cells for tissue-engineered venous valve construction in vitro. OBJECTIVE: To study the culture method of sheep bone marrow-derived endothelial progenitor cells (EPCs) and provide experimental basis for seed cells selection of sheep tissue engineering by culture and identification of sheep bone marrow-derived endothelial progenitor cells in vitro. METHODS: Sheep bone marrow mononuclear cells were obtained after whole bone marrow culturing selectively by using bone marrow condition medium. After primary cells were passaged, CD133-positive cells were sorted by microbeds instrument and cultured continually. Flow cytometry confirmed CD133 expression in cells before separation. The cell growth curve was drawn after separation and passage to observe cell growth capacity. Cell specific molecule CD133, D34, VWF expression were detected by immunocytochemistry. FITC labeled BS-1-lectin and DiI labeled ac-LDL were used to label cells and detect cell phagocytosis. RESULTS AND CONCLUSION: The sheep bone marrow mononuclear cells began to adhere on day 2 in primary culture and fully integrated on day 7. Cells started to went into the logarithmic growth phase on day 2 after passage and cell morphology formed typical paving stone-like on 3-5 days after passage; cells went into platform phase on day 7 after passage. Cells microbead sorting rate was 12.6% after passage. Flow cytometry showed CD133-positive cells rate was 12.64%. Immunocytochemistry showed that cells presented CD133, CD34, VWF-positive expression. FITC-labeled BS-1-lectin and DiI-ac-LDL double positive cells rate was 85.3%. This study successfully isolated and cultured EPCs from sheep bone marrow mononuclear cells. Related Articles | Metrics

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Bone marrow mesenchymal stem cells-induced transformation of cardiac fibroblasts into myofibroblasts: Optimal microenvironment of myocardial infarction at different stages Lin Chu-wei, Zhou Sheng-hua, Liu Qi-ming, Du You-you, Yao Li-qun, Zhang Liang, Cai Xiang-ping 2010, 14 (36):  6667-6671.  doi: 10.3969/j.issn.1673-8225.2010.36.004 Abstract ( 116 )   PDF (385KB) ( 343 )   Save BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) can effectively improve heart function following myocardial infarction. Its action mechanism is associated with aggregation of a large number of myofibroblasts in the transplant region, but few reports have verified that gathered myofibroblasts are transformed from BMSCs or from fibroblasts. OBJECTIVE: To explore the impact of microenvironment of myocardial infarction in different periods in rat BMSCs inducing cardiac fibroblasts into myofibroblasts. METHODS: The culture group was assigned to three groups: rat BMSCs group, cardiac fibroblasts group and combination group. Myocardial homogenate was added at different times after myocardial infarction for 7-28 days. A blank control group was set, without myocardial homogenate. Immunocytochemical staining was used to determine the differences of α-smooth muscle actin (α-SMA) expression and the number of fibroblasts into myofibroblasts between the two groups. RESULTS AND CONCLUSION: In the combination group, the positive rate of cardiac fibroblasts into myofibroblasts is highest, when BMSCs and cardiac fibroblasts were cultured together and intervened by myocardial homogenate on day 14 after myocardial infarction. Results suggest that in the microenvironment of post-myocardial infarction, BMSCs can induce cardiac fibroblasts into myofibroblasts, while the microenvironment of post-myocardial infarction on day 14 is the best one. Related Articles | Metrics

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Effect of bone marrow-derived mesenchymal-like stem cells versus bone marrow-derived mononuclear cells on bleomycin-induced pulmonary fibrosis Chen Juan, Liu Wei-wei, Liu Yi-mi, Li Su-mei, Yu Wei, Chen Jia-yu, Zhang Cheng, Lin Chen-li, Tang Li-juan, Fang Mao,Zhong Xue-yun 2010, 14 (36):  6672-6676.  doi: 10.3969/j.issn.1673-8225.2010.36.005 Abstract ( 128 )   PDF (443KB) ( 479 )   Save BACKGROUND: Previous studies demonstrated that many diseases can be effectively treated by using both bone marrow-derived mesenchymal-like stem cells (BM-MSCs) and bone marrow-derived mononuclear cells (BM-MNCs). BM-MSCs also have been used in lung fibrosis models.  OBJECIIVE: To compare the curative effect of BM-MSCs and BM-MNCs for treating bleomycin A5-induced alveolitis and early fibrosis. METHODS: BM-MSCs and BM-MNCs from Wistar male young rats were cultured and marked with 4, 6-diamidino-2-phenylindole (DAPI). A total of 21 female Wistar rats were intratracheally injected with bleomycin A5 to establish pulmonary fibrosis models. The rats were randomly divided into BM-MSCs group, BM-MNCs group and model group. The rats were sacrificed on day 7 following model induction. Pathological sections of pulmonary tissues were obtained to observe inflammation and fibrosis. Fluorocyte marked with DAPI was detected by fluorescence microscopy. The levels of hydroxyproline and tumor necrosis factors-α (TNF-α) were measured by enzyme linked immunosorbent assay (ELISA).   RESULTS AND CONCLUSION: ①In BM-MSCs group and BM-MNCs group, BM-MSCs and BM-MNCs marked with DAPI were detected in lung frozen section. ②Pulmonary alveolitis was most serious in model group, pulmonary alveolitis and fibrosis in BM-MNCs group was more serious than BM-MSCs group, and significant differences were detected among groups (P < 0.05). ③Hydroxyproline and TNF-α levels were highest in model group, and lowest in the BM-MSCs group, and significant differences were detected among groups (P < 0.05). Results suggest that both BM-MSCs and BM-MNCs can be planted in the impaired lung tissue, and effectively prevent the development of bleomycin-induced pulmonary alveolitis and early fibrosis. The curative effect of BM-MSCs is better than that of BM-MNCs. Related Articles | Metrics

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Induced differentiation of human bone marrow mesenchymal stem cells into dopaminergic neurons Jin Hui, Zhang Lei, Li Wei-wei 2010, 14 (36):  6677-6680.  doi: 10.3969/j.issn.1673-8225.2010.36.006 Abstract ( 155 )   PDF (345KB) ( 308 )   Save BACKGROUND: In vivo and vitro studies have confirmed that the ratio of differentiation of human bone marrow mesenchymal stem cells (hBMSCs) into neurons was obviously lower compared with glial cells. Moreover, these neurons will gradually die. Finally, the number of neurons in living cells is low. OBJECTIVE: To explore the potential of hBMSCs differentiation into dopaminergic neuron-like cells in vitro.  METHODS: The hBMSCs were isolated, purified and amplified. After pretreatment with basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) in vitro, the hBMSCs were cultured at the medium containing glial cell line-derived neurotrophic factor (GDNF) and Angiotensin II (AngⅡ) to induce hBMSCs differentiation into neurons and dopaminergic neurons. Changes in cell morphology were observed during differentiation. Marker expression of neurons and dopaminergic neurons were detected by immunohistochemistry. RESULTS AND CONCLUSION: Following induction, hBMSCs presented dipolar, multipolar and pyramidal typical neuron morphology, significantly expressed anti-human nerve nestin [(55.7±4.3)%] and neuron specific enolase [(78.2±6.7)%]. Moreover, a majority of hBMSCs expressed tyrosine hydroxylase [(48.5±5.6)%], but did not express glial fibrillary acidic protein. These indicated that under suitable conditions, hBMSCs can differentiate into neurons and dopaminergic neuron-like cells. Related Articles | Metrics

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Isolation, cultivation and chondrogenic differentiation of adipose-derived stromal cells in vitro Zhao Da-qing, Li Qing, Ma Yu, Deng Zhi-hong, Qiao Li, Lu Lian-jun, Qiu Jian-hua 2010, 14 (36):  6681-6684.  doi: 10.3969/j.issn.1673-8225.2010.36.007 Abstract ( 84 )   PDF (344KB) ( 430 )   Save BACKGROUND: Tissue engineering has been investigated as a potential means for generation of replacement cartilage. But the ideal seed cell which is still a problem has to be further solved. OBJECTIVE: To observe the solation and cultivation of adipose-derived stromal cells (ADSCs), and to investigate its chondrogenic differentiation ability in vitro. METHODS: Subcutaneous fatty tissue from New Zealand rabbits were digested with collagenase. The obtained ADSCs were cultured in the chondrogenic induced medium. Incubation and passage of ADSCs were observed under inverted microscope. The feasibility of ADSCs cartilage induction was detected by immunohistochemistry, Alcian Blue stain, and picrosirius red stain. Proliferation of ADSCs was observed by MTT methods. RESULTS AND CONCLUSION: ADSCs could be isolated and cultured in vitro. After cultured in chondrogenic medium, the cells differentiated toward cartilage cells and secreted the specific cartilaginous matrices acid mucopolysaccharide and type Ⅱ collagen. Primary cultured ADSCs were spindle-shaped, with oval nucleus. At day 3, ADSCs were in the exponential growth phase, and the population doubling time was about 55 hours. At day 4, cell growth was stable. At day 9, the cells were observed aging symptoms. Cartilage-induced cell proliferation rate increased significantly, while with the incubation time prolonging, cell volume increased significantly, from long fusiform shape into large, multi-polygonal pseudopodia, furthermore nuclei were also larger, and nucleolus was clear. At day 2 after incubation, ADSCs were in the exponential growth phase, and the population doubling time was about 30 hours. At days 8, the cell growth was stable. After the 15th generation, cell proliferation slowed down and cells were observed signs of aging. This suggested that ADSCs differentiated into chondrocytes under a certain degree; therefore, ADSCs could be served as seed cell for cartilaginous tissue engineering. Related Articles | Metrics

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Biological characteristics and phenotypes of adipose tissue-derived stem cells Cai Zhen, Pan Bo, Lin Lin, Jiang Hai-yue, Zhuang Hong-xing 2010, 14 (36):  6685-6688.  doi: 10.3969/j.issn.1673-8225.2010.36.008 Abstract ( 156 )   PDF (364KB) ( 609 )   Save BACKGROUND: Adipose-derived stem cells (ADSCs) are capable of multi-directional differentiation and exist in human processed lipoaspirate. However, no in-depth studies have addressed biological characteristics of adipose tissue-derived stem cells (ADSCs) in vitro, and there are some disputes on the results from phenotype studies. OBJECTIVE: To investigate biological characteristics and phenotypes of human ADSCs. METHODS: Human adipose tissue was collected from persons undergoing fat extraction. The ADSCs were digested with collagenase, cultured and subcultured in vitro. Cell morphology was observed and growth curves were drawn. The phenotypes of ADSCs were identified by immunohistochemistry and flow cytometry. RESULTS AND CONCLUSION: ADSCs grew well, presented fibroblast-like growth. No significant change in cell morphology was detected within passage 15. The results from immunohistochemistry showed that cells were positive for CD29, CD44, CD105, but negative for CD34, CD45. Flow cytometry results have shown that cells were positive for CD29, CD44, CD105, MHC-Ⅰ, but negative for CD3, CD14, CD19, CD31, CD33, CD34, CD45, CD106, CD117, CD184. Results have suggested that ADSCs can be abundantly harvested and have stable proliferation in poorly differentiated status in vitro. Stem cell-related antigens were highly expressed in ADSCs. Related Articles | Metrics

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Amplification of human adipose-derived stem cells using tissue culture technique: An evaluation of biological properties Peng Zhi, Chen Qi, Jia Zhen-hua, Tang Hong-wei 2010, 14 (36):  6689-6694.  doi: 10.3969/j.issn.1673-8225.2010.36.009 Abstract ( 86 )   PDF (566KB) ( 806 )   Save BACKGROUND: Tissue culture technique was used to isolate human adipose-derived stem cells (ADSCs) for in vitro culture. This method is simple, can save time, and keeps biological properties of stem cells. According to this, the multiple-directional differentiation provides theoretic evidences and reference line for clinical application of tissue engineered seed cells.  OBJECTIVE: To investigate the basic biological characteristics of human ADSCs by tissue culture techniques and their surface protein expression, and to explore their osteogenic and adipogenic potential in vitro. METHODS: Human adipose tissue was obtained from the abdominal subcutaneous adipose tissue of health patients, and then ADSCs were isolated with tissue culture techniques and cultured in vitro. Morphology of ADSCs was observed. Cell cycle and surface proteins of the third passage of ADSCs were analyzed by flow cytometry technique. ADSCs from the third passages were induced into osteogenic and adipogenic lineages by different revulsants. Cells were examined by von Kossa staining and Oi1 Red O staining. RESULTS AND CONCLUSION: Primary and passage ADSCs were fibroblast-like and could rapidly expand. The majority of the third passage of cells was in G0/G1 phase (88%). The third passage of ADSCs was positive for CD29, CD44, CD90 and CD105, while negative for CD34, CD45 and CD106. Human ADSCs were positive for Oi1 Red O staining at 21 days of adipogenic induction, and positive for Alizarin Reds staining and Von Kossa staining after osteogenic induction. Therefore, human ADSCs have the potential to differentiate into osteoblasts and adipocytes. Related Articles | Metrics

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Transfection of human adipose-derived stem cells with lentiviral vector containing enhanced-green fluorescent protein: A validity study Wang Wei-wei, Zhang Jin-yuan 2010, 14 (36):  6695-6698.  doi: 10.3969/j.issn.1673-8225.2010.36.010 Abstract ( 104 )   PDF (777KB) ( 346 )   Save BACKGROUND: Before transplantation, the ability of stem cells was obviously increased by the transfection of target gene. The key of gene therapy is that target gene was transferred effectively and was expressed appropriately in stem cells. The choice of carrier system will affect the treatment effectiveness. OBJECTIVE: To observe the transfection efficiency of lentiviral vector transfecting the human adipose-derived stem cells (hASCs) and to detect the vitality of hASCs transfected. METHODS: The recombinant lentiviral supernatant was obtained by standard lipofectamine reagent. The hASCs were transfected by lentiviral vector containing enhanced-green fluorescent protein (EGFP) with different multiplicity of infection (MOI=1, 5, 10, 15, 20). Transfection efficiency was assessed by flow cytometer for EGFP expression under fluorescence microscope. The multi-directional differentiation of hASCs after transfection was evaluated by alizarin red staining and oil red O staining. The proliferation of hASCs after transfection was detected by MTT assay. RESULTS AND CONCLUSION: Transfection efficiency was 60% at 3 day and 90%-95% at 5-7 day after infection under MOI=20. The alizarin red staining and oil red O staining showed hASCs after transfected hASCs were induced to osteogenic and adipogenic lineages, which suggested that viral infection could not affect multi-directional differentiation of hASCs. The proliferation of transfected hASCs had no marked change. This showed that hASCs transfected by lentiviral vectors carrying EGFP can effectively infect hASCs. Related Articles | Metrics

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Hypoxic preconditioning stimulates proliferation of endogenous neural stem cells in brain hippocampus Zhang Yan-bo, Yang Ming-feng, Sun Bao-liang, Niu Jing-zhong 2010, 14 (36):  6699-6702.  doi: 10.3969/j.issn.1673-8225.2010.36.011 Abstract ( 93 )   PDF (341KB) ( 457 )   Save BACKGROUND: Ischemic, hypoxic, hypoxia and other stimuli can lead to endogenous neural stem cell proliferation and differentiation, which play a role in brain repair, but it is still not clear whether hypoxic preconditioning can affect the proliferation of endogenous neural stem cells. OBJECTIVE: To explore the proliferation of endogenous neural stem cells in hippocampus of hypoxic preconditioning mice. METHODS: Balb/c clean mice were randomly divided into three groups. Mice in the hypoxia control group were placed in a wide-mouthed bottle, blocked by rubber stopper. Animals with the first asthmoid respiration marked tolerance limit of hypoxia, and once hypoxia exposure was completed. Above-mentioned procedures were repeated four times in the hypoxic preconditioning group. Mice in the normal control group were not exposed to hypoxia. The number and fluorescence intensity of BrdU-labeled cells were counted and observed by immunofluorescence and confocal laser scanning microscope. RESULTS AND CONCLUSION: Compared with hypoxia control group, the tolerance time was significantly longer in the hypoxic preconditioning group (P < 0.01). Fluorescence intensity of BrdU-labeled endogenous neural stem cells was faint, and a few BrdU-positive cells were visible in the hippocampus in the normal control group. The fluorescence intensity and BrdU-positive cell number were significantly increased in the hypoxia control and hypoxic preconditioning groups (P < 0.01). The increased range was greater in the hypoxic preconditioning group compared with the hypoxia control group (P < 0.01). Results suggest that proliferation of endogenous neural stem cells is seen obviously in hippocampus of hypoxic preconditioning mice, which is perhaps involved in cerebral protective mechanism of hypoxic preconditioning. Related Articles | Metrics

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In vivo magnetic resonance imaging tracking of transplanted Feridex-labeled adult neural stem cells in rats   Ge Feng, Tang Zhi-fang, Xu Jie, Fang Wen-feng, Zhu Ai-hua, Wu Wei-jiang, Lu Hua, Wu Xing 2010, 14 (36):  6703-6706.  doi: 10.3969/j.issn.1673-8225.2010.36.012 Abstract ( 123 )   PDF (360KB) ( 469 )   Save BACKGROUND: At present, brain section examination was needed to judge whether neural stem cells (NSCs) migrated towards damaged region following transplantation by sacrificing recipient animals, and multiple points and dynamic observation cannot be performed. OBJECTIVE: To study the feasibility of magnetic resonance imaging (MRI) tracking of transplanted Feridex-labeled adult NSCs in vivo in rats with traumatic brain injury model. METHODS: Traumatic brain injury model was established in the left hemisphere of the rats. In vitro adult NSCs were labeled by Feridex. 2 weeks later, Feridex-labeled adult NSCs were implanted into the right hemisphere. All rats received MRI at 1, 3 days, 1 and 2 weeks after implantation. 2 weeks after implantation, all the rats were killed and their brains were taken out to undergo Prussian blue staining to track the presence of labeled-cells. RESULTS AND CONCLUSION: Efficiency of labeled adult NSCs was about 85%. At implanted sites, low signal intensity could be observed in MRI examination with the scanning sequences of FSE T2WI and GRE T2. Low signal intensity lines were visible targeted to the contralateral lesion areas in labeled stem cell group. However, no signal line was found in the control group. The results from MRI were in accordance with the results from section with Prussian blue staining. These results indicated that MRI in vivo monitoring transplanted Feridex-labeled adult NSCs is feasible and effective in rats with traumatic brain injury model. Related Articles | Metrics

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Establishment of two-dimensional gel electrophoresis technique for embryonic rat neural stem cells Liu Huan, Huang Guo-wei, He Bing, Yang Yang, Zhao Lin 2010, 14 (36):  6707-6710.  doi: 10.3969/j.issn.1673-8225.2010.36.013 Abstract ( 147 )   PDF (510KB) ( 345 )   Save BACKGROUND: Neural stem cells are the current hot topic, and two-dimensional electrophoresis is the basic technique for proteome, it is necessary to establish a stable two-dimensional gel electrophoresis technology system for neural stem cells. OBJECTIVE: To establish a stable, reproducible two-dimensional gel electrophoresis separation method for neural stem cells through analysis of several key steps of two-dimensional electrophoresis. METHODS: Embryonic rat neural stem cells were cultured in serum-free cell culture. Proteins were extracted from samples and lysed. After passive rehydration, samples were isolated by different isoelectric focusing (IEF) parameters. Protein maps were analyzed by PDQuest8.0. Quality of two-dimensional gel electrophoresis maps under different IEF parameters, number and reproducibility of protein spots were detected. RESULTS AND CONCLUSION: Quality of two-dimensional gel electrophoresis diagrams differs with IEF parameters. Low-voltage step could remove the cross striations. Ideal electrophoresis could be obtained under 80 000 Vh. Proper two-dimensional gel electrophoresis methods can be constructed by optimizing IEF parameters. Related Articles | Metrics

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Changes in cardiac function following transplantation of 5-azacytidine-induced versus non-induced bone marrow mesenchymal stem cells into infarcted myocardium Chen Li-xing, Nie Jun, Cai Hong-yan, Zhao Ling, Guo Tao, Xiao Jian-ming 2010, 14 (36):  6711-6714.  doi: 10.3969/j.issn.1673-8225.2010.36.014 Abstract ( 98 )   PDF (316KB) ( 283 )   Save BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) have the potential to differentiate into cardiomyocyte-like cells following induction of 5-azacytidine in vitro. Whether transplanted stem cells could improve cardiac function was unclear. OBJECTIVE: To compare the changes in cardiac function following transplantation of BMSCs with or without induction of 5-azacytidine into tissues of acute myocardial infarction (AMI). METHODS: New Zealand rabbits were developed into AMI models by ligating left anterior descending artery. Phosphate buffered saline, 5-azacytidine-induced or non-induced BMSCs were separately injected into the surrounding regions of myocardial infarct tissues. Three days or four weeks after transplantation, echocardiogram was used to measure left ventricular end diastolic volume, ventricular end-systolic volume and ejection fraction to evaluate changes in cardiac function. RESULTS AND CONCLUSION: Following anterior descending branch ligation, the cardiac function was significantly decreased. At 3 days following transplantation, no significant difference was determined in left ventricular end diastolic volume, ventricular end-systolic volume and ejection fraction in each group. These suggested that transplantation of 5-azacytidine-induced and non-induced BMSCs did not affect rabbit cardiac function in a short phase. At 4 weeks following transplantation, compared with control group, transplanted BMSCs could decrease left ventricular end diastolic volume, ventricular end-systolic volume and ejection fraction. However, there were no obvious differences of cardiac function between 5-aza+BMSCs and BMSCs transplantation. Related Articles | Metrics

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Survival and migration of neural stem cells surrounding the lesion sites of traumatic brain injury Zhang Li, Zhang Hui, Sun Li, Luo Qiang, Hao Xiu-qing, Liang Feng, Wang Xin-sheng, Bo Ai-hua 2010, 14 (36):  6715-6719.  doi: 10.3969/j.issn.1673-8225.2010.36.015 Abstract ( 63 )   PDF (401KB) ( 351 )   Save BACKGROUND: It is difficult to regenerate in the central nervous system of adults. Following traumatic brain injury, the number of surviving cells surrounding the lesion sites directly affects the patients’ prognosis. How to make the transplanted neural stem and progenitor cells (NSPC) survival, migration and differentiation around the cortex damage in rats models of traumatic brain injury effectively are the keys of nerve regenerated and repair research. OBJECTIVE: To study survival, migration and differentiation of NSPC around the cortex damage in rats models of traumatic brain injury. METHODS: Using serum-free culture technique, epidermal growth factor and basic fibroblast growth factor were added to stimulate the origin of embryonic rat forebrain NSPC growth and proliferation. Moreover, in vitro clone culture was conducted. BrdU labeling, immunohistochemistry and immunofluorescence method were used to confirm the proliferation and multi-directional differentiation potential properties of NSPC. Survival and migration of transplanted NSPC were observed around the cortex damage in rat models of Fenney’s traumatic brain injury. RESULTS AND CONCLUSION: Neural stem and progenitor cell clones were detected by immunohistochemistry and immunofluorescence method, which showed nestin-positive and BrdU-positive reaction. They displayed NSE-positive, GFAP-positive and MAP-2-positive reaction after NSPC differentiation in vitro. BrdU-positive cells were interspersed and GFAP-positive cells number was increased surrounding the cortex damage at 7 and 14 days posttransplantation. These suggested that rat forebrain neural stem and progenitor cells are able to proliferate and differentiate into neurons and glial cells, and can survive and migrate surrounding cortex damage, showing the characteristics of integrated into brain tissue in morphological observation. Related Articles | Metrics

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Acupoint transplantation of allogenic bone marrow mesenchymal stem cells: Angiogenesis and relevant cytokine changes following acute myocardial infarction Zhang Zhe, Li Zheng, Yang Guan-lin, Chen Yan 2010, 14 (36):  6721-6723.  doi: 10.3969/j.issn.1673-8225.2010.36.016 Abstract ( 101 )   PDF (339KB) ( 339 )   Save BACKGROUND: The bone marrow mesenchymal stem cell (BMSC) transplantation ways mainly contain peripheral vein injection transplantation, coronary artery transplantation and intramyocardial injection transplantation, but there are limitations such as lack of target and high price. OBJECTIVE: To observe effects of allogenic BMSC transplantation on angiogenesis and changes in related cytokines in the infarct region following rabbit acute myocardial infarction. METHODS: The rabbit bone marrow was extracted to isolate, culture, purify, amplify, identify and label BMSCs. A total of 55 rabbits were randomly assigned to normal and operation groups. In the operation group, at 1 week following acute myocardial infarction, the rabbits were randomly divided into three groups: acupoint injection stem cell group, model control group and acupoint injection saline group. Five weeks later, myocardial tissue was obtained for immunohistochemistry BrdU staining and cardiac specific troponin T immunohistochemistry staining. Immunohistochemistry was used to detect platelet/endothelial cell adhesion molecule-1 levels in infarct myocardial tissue. Immunohistochemistry, sandwich enzyme-linked immunosorbent assay, reverse transcription-polymerase chain reaction and Western blot assay were utilized to determine vascular endothelial cell growth factor. Immunohistochemistry was employed to detect basic fibroblast growth factor. RESULTS AND CONCLUSION: BMSCs by acupoint injection could migrate towards infarct region, and differentiate into cardiac-like cells. Compared with model control group and acupoint injection saline group, platelet/endothelial cell adhesion molecule-1 expression upregulated in and surrounding the infarct tissue in the acupoint injection stem cell group. The upregulation of platelet/endothelial cell adhesion molecule-1 expression induced angiogenesis in and surrounding the infarct region. Its mechanism is to stimulate upregulation of basic fibroblast growth factor expression in myocardial tissue. Vascular endothelial cell growth factor expression did not exhibit upregulatory tendency in the infarct myocardial tissue and serum in the acupoint injection stem cell group. Thus, angiogenesis is induced by basic fibroblast growth factor and unknown factors. Related Articles | Metrics

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Apoptotic protease activating factor-1 expression in the heart, liver and lung of rabbits with multiple organ dysfunction syndrome following bone marrow mesenchymal stem cells transplantation Ling Bin, Chen Jing, Wang Ting-hua, Pan Xing-hua, Sun Jie, Liu Ping 2010, 14 (36):  6724-6728.  doi: 10.3969/j.issn.1673-8225.2010.36.017 Abstract ( 69 )   PDF (432KB) ( 311 )   Save BACKGROUND: The multiple organ dysfunction syndrome (MODS) onset and organ tissue injury are associated with cell apoptosis and necrosis. However, bone marrow mesenchymal stem cells (BMSCs) as seed cells are effective in repair field. OBJECTIVE: To observe BMSCs transplantation effects on apoptotic protease-activating factor 1 (Apaf-1) expression in involved organs of rabbits with MODS. METHODS: Two-hit animal model of MODS was established in the experimental group. Rabbit BMSCs were cultured, amplified and identified in vitro, and then labeled with GFP by lentivirus. BMSCs were transplanted via the femoral artery. We also set model control and normal groups. The survival of BMSCs in model of MODS was observed. Apaf-1 gene expression was detected in the heart, liver and lung prior to and following transplantation using RT-PCR. RESULTS AND CONCLUSION: Lentiviral transgenic GFP marker presence of BMSCs was visible in the rabbit liver and lung in the experimental group. RT-PCR results showed that Apaf-1 mRNA expression was determined in the heart, liver and lung tissues of normal group, and significantly increased in the heart, spleen and kidney of model control group. Apaf-1 mRNA expression was decreased in the rabbit heart, spleen and kidney of experimental group. These indicated that BMSCs have good biological activity. Transplanted BMSCs could implant in the damaged organs of rabbits with MODS, downregulate Apaf-1 mRNA expression, which suggested that BMSCs transplantation can provide a new therapeutic method for MODS. Related Articles | Metrics

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Allogeneic bone marrow mesenchymal stem cells transplantation for spinal cord injury in rats Ruan Zhi, Huang Hui, Sun Jian-hua, Shi Chen-hui, Dong Jin-bo, Wang Yong-ming 2010, 14 (36):  6729-6732.  doi: 10.3969/j.issn.1673-8225.2010.36.018 Abstract ( 117 )   PDF (327KB) ( 499 )   Save BACKGROUND: There is no good therapeutic tool for repairing spinal cord injury (SCI). Cell transplantation can contribute to axonal regeneration and the recovery of spinal cord function, and provide possibility for treating SCI. However, the SCI models and transplanted manners are various, and the therapeutic efficiency is different. OBJECTIVE: To investigate the therapeutic effects of allogeneic bone marrow mesenchymal stem cells (BMSCs) transplantation on rat SCI. METHODS: BMSCs were isolated from rats by the whole bone marrow method. Healthy Sprague Dawley rats were randomly assigned to cell transplantation, control and sham surgery groups. In the cell transplantation and control groups, rat SCI models were established by the modified Allen’s method. In the sham surgery group, the spinal cord was exposed only. At 4 weeks following surgery, locomotive function of SCI rats was graded weekly. Enzyme linked immunosorbent assay was used to detect expression of brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) in injured spinal cord. Immunofluorescent staining was employed to detect the expression of NF200 and glial fibrillary acidic protein (GFAP).  RESULTS AND CONCLUSION: Compared with control group, movement function was significantly improved, and BDNF and NGF protein contents were significantly increased in the spinal cord in the cell transplantation group (P < 0.05). The spinal cord capsular space was small; NF200 expression was significantly increased; GFAP expression was reduced in the rats from the transplantation group. Results indicate that allogeneic BMSCs transplantation could increase NGF levels, inhibit the glial scar formation and promote axonal regeneration and improve functional recovery in rats after SCI. Related Articles | Metrics

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Allogeneic bone marrow mesenchymal stem cells for the treatment of diabetic foot ulcers in rats and vascular endothelial growth factor expression   Cai Qian, Dong Fang, Liu Yi 2010, 14 (36):  6733-6737.  doi: 10.3969/j.issn.1673-8225.2010.36.019 Abstract ( 99 )   PDF (397KB) ( 489 )   Save BACKGROUND: Autologous bone marrow stem cells and autologous peripheral blood stem cells are commonly used to treat diabetic foot ulcer in clinic. Few studies have addressed basic research of bone marrow mesenchymal stem cells (BMSCs) for diabetic foot ulcer. OBJECTIVE: To investigate the effect of local transplantation of allogeneic BMSCs in the treatment of rats with diabetic foot ulcers and the systemic and local expression of vascular endothelial growth factor (VEGF). METHODS: A total of 90 male Wistar rats were randomly divided into three groups: control group (normal foot ulcers), BMSCs treatment group and diabetic control group. Rat models of type 2 diabetic foot ulcers were established in the BMSCs treatment group and diabetic control group. Following model induction, allogenic BMSCs and stem cell medium DMEM were respectively injected. At 1, 4 and 8 days following model establishment, rat ulcer area was observed in each group. Nucleus staining tracing and pathological examination were conducted. Enzyme linked immunosorbent assay was utilized to measure VEGF concentration in peripheral blood. Western blot assay was employed to detect VEGF concentration in local region. RESULTS AND CONCLUSION: Compared with diabetic control group, ulcer healed faster in the BMSCs treatment group, but slower than control group. Peripheral blood VEGF expression increased, but did not reach normal levels. In early days of allogenic BMSCs treatment (1-4 days), VEGF concentration was significantly elevated, but slightly elevated in late days (8 days). In the BMSCs treatment group, healing was slow; epidermis was not completely covered. The healing was significantly early compared with diabetic control group, and epidermis was obviously covered. Nucleus staining demonstrated that transplanted stem cells gathered in the regions surrounding ulcers. These indicated that allogenic BMSCs can promote the healing of diabetic foot ulcers in rats. The mechanism may be that it increased the VEGF expression in systemic condition, especially local condition. Related Articles | Metrics

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Effects of bone marrow mesenchymal stem cell transplantation on growth-associated protein-43 expression and infarction volume following permanent middle cerebral artery occlusion in rats Zhang Qi-mei, Peng Yu, Zhuang Wei-hua, Lan Jing, Li Cheng-yan 2010, 14 (36):  6738-6743.  doi: 10.3969/j.issn.1673-8225.2010.36.020 Abstract ( 76 )   PDF (410KB) ( 302 )   Save BACKGROUND: There are numerous studies on bone marrow mesenchymal stem cells (BMSCs) transplanted in the brains of ischemia rats survive and migrate through blood-brain barrier, partially differentiate into neurons, promote the secretion of various neurotrophic factors, significantly improve neurological impairment, and have long therapeutic time window. OBJECTIVE: To explore the effect of BMSC transplantation on the expression of growth-associated protein-43 and the infarct volume following permanent middle cerebral artery occlusion in rats. METHODS: Adult Sprague-Dawley rats were randomly assigned to model control group, sham operation group and stem cell transplantation group. An additional four adult Sprague Dawley rats were used to prepare BMSCs. BMSCs were labeled with 5-bromodeoxyuridine. Rats in the sham operation group were utilized to deligate right common carotid artery. The remaining rats were employed to establish right middle cerebral artery models. Following model induction, BMSCs were infused into the rats from the stem cell transplantation group. An equal volume of phosphate buffer saline was injected into the rats in the model control group. Neurological impairment was graded prior to transplantation, at 7, 14, 21 and 28 days following transplantation. Expression of growth associated protein-43 was determined surrounding the infarct region using the immunohistochemistry. RESULTS AND CONCLUSION: In the stem cell transplantation group, 5-bromodeoxyuridine-positive cells were detected in the infarct site at 7 day following transplantation, peaked at 14 days, reduced and disappeared at 28 days. Immunological activity of growth associated protein-43 was significantly higher in the stem cell transplantation group than in the model control group at 7 and 14 days following transplantation (P < 0.05). No nerve injury was tested in rats from the sham operation group, and the score of neural function was 0 point. With time prolonged, the score of neural function was gradually reduced in the model control and stem cell transplantation groups. The score was significantly lower in the stem cell transplantation group than in the model control group at 14 days following transplantation (P < 0.05). Compared with model control group, cerebral infarct volume was significantly diminished in the stem cell transplantation group (P < 0.05). Results indicated that BMSC transplantation can upregulate growth associated protein-43 expressions surrounding the infarct region after focal cerebral ischemia, can significantly decrease infarct volume. Related Articles | Metrics

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Effects of bone marrow mesenchymal stem cells transplantation on alpha-smooth muscle actin expression in acute radiation-induced liver injury rats Ji Wei-zheng, Ma Yan, Wen Hao 2010, 14 (36):  6744-6750.  doi: 10.3969/j.issn.1673-8225.2010.36.021 Abstract ( 71 )   PDF (638KB) ( 478 )   Save BACKGROUND: Radiation-induced liver injury has greatly limited the clinical application of radiotherapy. Research demonstrated that liver injury environment is suitable for the differentiation of bone marrow mesenchymal stem cells (BMSCs), which suggested that it is possible to treat radiation-induced liver injury with BMSCs transplantation. OBJECTIVE: To test effect of BMSCs transplantation on ɑ-smooth muscle actin (SMA) expression level in acute radiation-induced liver injury rats. METHODS: BMSCs were isolated from male rats, and subcultured. The third passage of BMSCs was selected as transplanted cells. Female Sprague Dawley rat models of acute radiation-induced liver injury were established, and randomly assigned to two groups. Within 4 hours following radiotherapy, BMSCs suspension was infused through caudal vein in the intervention group. An equal volume of saline was infused into the control group. ɑ-SMA expression was detected by immunohistochemistry in the liver. Sex determining gene Y (SrY) was determined using in situ hybridization technique. Pathomorphological change in the liver was observed under an optical microscope. RESULTS AND CONCLUSION: The positive expression rate of ɑ-SMA in right lobe of liver of intervention group was lower than control group (P < 0.05). The existence of SrY was more in the right lobe of liver than that of in the left lobe of liver in intervention group(P < 0.05). In control group, no SrY gene can be found. The injury degree of right lobe liver in intervention group was lower than control group. Results have verified that acute radiation-induced liver injury can lead BMSCs’ homing in male rats. BMSCs can down-regulate the expression level of ɑ-SMA induced by acute radiation in liver injury rats. Related Articles | Metrics

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Spinal cord injury-associated protein expression following neurotrophin-3 gene modified neural stem cells transplantation   Deng Xing-li, Liang Yuan-xin, Yang Zhi-yong, Song Xiao-bin, Li Yu, Yang Shu-hua 2010, 14 (36):  6751-6754.  doi: 10.3969/j.issn.1673-8225.2010.36.022 Abstract ( 80 )   PDF (443KB) ( 350 )   Save BACKGROUND: Studies have confirmed that neural stem cells (NSCs) and neurotrophic factor-3 (NT-3) gene modified neural cell combination transplantation can survive following transplantation and effectively promote functional recovery following spinal cord transsection. However, whether NT-3 gene-modified NSCs exert effects in damaged spinal cord and contribute to functional recovery of the spinal cord injury (SCI) rats remains poorly understood. OBJECTIVE: To investigate the behavioral recovery of SCI rats that received transplantation of NT-3 gene-modified fetal spinal cord-derived NSCs, and the gene expression in injury location. METHODS: Spinal cords of 30 Sprague-Dawley rats were hemisected at T9 level. The rats were randomly assigned to three groups. Cell culture medium, NSCs and NT-3 gene-modified NSCs were transplanted into the lesion site of rats of each group respectively. An additional 10 rats served as blank control group, which only received laminectomy. Following transplantation, behavior test was utilized to evaluate spinal cord functional recovery. Reverse transcription-polymerase chain reaction and Western blot assay were used to detect NT-3 and myelin basic protein expression in lesion site. RESULTS AND CONCLUSION: Rats in NT-3 gene -NSCs group got best results in behavioral test. Behavioral test outcomes were worst in the cell culture medium group. Compared with cell culture medium group, NT-3gene and myelin basic protein gene mRNA expression significantly upregulated in spinal cord tissue of rats from the NSCs and NT-3 gene-modified NSCs groups. Protein levels obtained similar results. Moreover, the outcomes were obvious in the NT-3 gene -modified NSCs group. These indicate that transplantation of NT-3 gene -modified NSCs can promote the differentiation of cells into oligodendrocytes in lesion site, and strongly express NT-3. Related Articles | Metrics

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Freeze-thaw by liquid nitrogen promotes platelet-derived growth factor-AA and transforming growth factor-β1 release  Li Hong-tao, Duan Jian-min, Zhou Mou, Ma Yu-xia, Katayama Tadashi 2010, 14 (36):  6755-6758.  doi: 10.3969/j.issn.1673-8225.2010.36.025 Abstract ( 175 )   PDF (297KB) ( 329 )   Save BACKGROUND: Platelet is a rich source of autologous growth factors. Bovine thrombin is the most common used activator to promote growth factors release from platelet. The clinical use of thrombin has some risks. OBJECTIVE: To observe the effect of freeze-thaw by liquid nitrogen to promote platelet-derived growth factor-AA and transforming growth factor-β1 release from platelet in comparison with bovine thrombin. METHODS: Platelet rich plasma and platelet poor plasma were manufactured from venous blood of five healthy volunteers by secondary centrifugation. The platelet rich plasma was then washed to get rid of the plasma and to manufacture the washed platelet. Washed platelet was activated by freeze-thaw method using liquid nitrogen to promote growth factor release and platelet rich plasma was activated by 1 000, 500, 250, 125, 62.5, 31.25 U/mL bovine thrombin/calcium chloride. Mass concentrations of platelet-derived growth factor-AA and transforming growth factor-β1 in washed platelet, platelet rich plasma and platelet poor plasma were assayed using enzyme linked immunosorbent assay.  RESULTS AND CONCLUSION: The concentrations of platelet-derived growth factor-AA and transforming growth factor-β1 in the washed platelet activated by freeze-thaw were (8.973±1.213) and (27.445±2.273) μg/L. There was no significant difference from the concentration in the bovine thrombin of 500-1 000 U/mL activated platelet rich plasma (P > 0.05). These suggested that freeze-thaw can be used as a safe and effective technique to active washed platelet instead of bovine thrombin. Related Articles | Metrics

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Effects of hypoxia inducible factor-1 alpha small interfering RNA on hypoxia inducible factor-1 alpha, stromal derived factor 1 alpha and vascular endothelial growth factor gene expression in bone marrow mesenchymal stem cells  Zhang Wen-wen, Lin Ming, Xu Chang-sheng 2010, 14 (36):  6759-6763.  doi: 10.3969/j.issn.1673-8225.2010.36.024 Abstract ( 90 )   PDF (428KB) ( 424 )   Save BACKGROUND: The cytokines secreted by bone marrow mesenchymal stem cells (BMSCs) can be affected by hypoxia. Its mechanism may exert effects via hypoxia inducible factor-1 α (HIF-1 α). OBJECTIVE: To observe HIF-1 α, stromal derived factor 1 α (SDF-1 α) and vascular endothelial growth factor (VEGF) gene expression affected by HIF-1 α small interfering RNA (siRNA) in BMSCs. METHODS: BMSCs were cultured by adherence method. The third to fifth passages of BMSCs were selected for experiments. Inverted microscope was used to observed morphology. Flow cytometry was used to detect the expression of surface markers CD34, CD44 and CD90. BMSCs were divided into four groups, normal control group (without any treatment), hypoxia group (hypoxia for 24 hours), liposome control group (BMSCs transfected with liposome then hypoxia for 24 hours), RNA interference (BMSCs transfected with RNA interference sequences then hypoxia for 24 hours). Reverse transcription-polymerase chain reaction was utilized to determine HIF-1 α, SDF-1 α and VEGF mRNA expression in BMSCs. Enzyme linked immunosorbent assay was employed to detect HIF-1 α, SDF-1 α and VEGF protein expression in BMSCs. RESULTS AND CONCLUSION: Compared with the normal control group, HIF-1 α, SDF-1 α and VEGF mRNA and protein expression increased in the hypoxia group (P < 0.05). Compared with the lipsome control group, HIF-1 α, SDF-1 α and VEGF mRNA and protein expression decreased in the RNA interference group (P < 0.05). These verified that hypoxia increased SDF-1 and VEGF gene expression. Inhibition of HIF-1 α expression decreased SDF-1 α and VEGF expression. HIF-1 α may be a regulatory factor for cytokine secretion following stem cell transplantation. Related Articles | Metrics

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Freeze-thaw by liquid nitrogen promotes platelet-derived growth factor-AA and transforming growth factor-β1 release  Li Hong-tao, Duan Jian-min, Zhou Mou, Ma Yu-xia, Katayama Tadashi 2010, 14 (36):  6764-6767.  doi: 10.3969/j.issn.1673-8225.2010.36.025 Abstract ( 86 )   PDF (331KB) ( 413 )   Save BACKGROUND: Platelet is a rich source of autologous growth factors. Bovine thrombin is the most common used activator to promote growth factors release from platelet. The clinical use of thrombin has some risks. OBJECTIVE: To observe the effect of freeze-thaw by liquid nitrogen to promote platelet-derived growth factor-AA and transforming growth factor-β1 release from platelet in comparison with bovine thrombin. METHODS: Platelet rich plasma and platelet poor plasma were manufactured from venous blood of five healthy volunteers by secondary centrifugation. The platelet rich plasma was then washed to get rid of the plasma and to manufacture the washed platelet. Washed platelet was activated by freeze-thaw method using liquid nitrogen to promote growth factor release and platelet rich plasma was activated by 1 000, 500, 250, 125, 62.5, 31.25 U/mL bovine thrombin/calcium chloride. Mass concentrations of platelet-derived growth factor-AA and transforming growth factor-β1 in washed platelet, platelet rich plasma and platelet poor plasma were assayed using enzyme linked immunosorbent assay.  RESULTS AND CONCLUSION: The concentrations of platelet-derived growth factor-AA and transforming growth factor-β1 in the washed platelet activated by freeze-thaw were (8.973±1.213) and (27.445±2.273) μg/L. There was no significant difference from the concentration in the bovine thrombin of 500-1 000 U/mL activated platelet rich plasma (P > 0.05). These suggested that freeze-thaw can be used as a safe and effective technique to active washed platelet instead of bovine thrombin. Related Articles | Metrics

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Optimization of preparing feeder layer cells of embryonic stem cells Zhang Wan-li, Li Wei, Lin Xue-ke, Li Hang, Zhai Rong-lin, Wang Guo-bin, Tao Kai-xiong 2010, 14 (36):  6768-6771.  doi: 10.3969/j.issn.1673-8225.2010.36.026 Abstract ( 86 )   PDF (351KB) ( 713 )   Save BACKGROUND: Studies have demonstrated that many factors may influence mouse embryonic fibroblasts (MEF) separation. Current researches on exploring these factors are based on semi-empirical results, which are lack of objective and quantitative data support.   OBJECTIVE: To investigate the optimal conditions in establishing MEF as mouse embryonic stem cells’ feeder layer cells. METHODS: Two different methods were conducted to isolate MEF from 13.5 days post coitum (dpc) Kunming mouse embryos. Trypan blue staining and cell counting of MEF were performed to study the relationship between cell livability and quantity. The MEF was digested by 0.05% trypsin at room temperature at various time points to analyze effects of dissociation adherent cell ability and cell viability at various digestion time points. Cells in feeder layer were inactivated using mitomycin for 2 and 2.5 hours. The cell growth curve was drawn. The optimal concentration and disposal time of mitomycin were explored. By various cryopreservation methods, the livability of MEF was compared after thawing. The optimal cryopreservation methods of MEF were screened. RESULTS AND CONCLUSION: MEF was sensitive to trypsin. The shorter trypsinase digestion time and multiple times method obviously elevated cell viability and quantity. At room temperature, 4-6 minutes digestion time contributed to cell viability preservation. 10 mg/L 2.5 hours and 20 mg/L 2 hours mitomycin inactivation was suitable for establishing cells in feeder layer. Progressive temperature reduction made MEF adaptive to temperature changes, which significantly increased MEF livability and quantity after thawing. Related Articles | Metrics

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HOXB4 gene expression during differentiation and proliferation of hematopoietic stem cells into lymphocyte progenitor cells Zou Yan, Zhai Xue-song, Chen Hong-ying 2010, 14 (36):  6772-6775.  doi: 10.3969/j.issn.1673-8225.2010.36.027 Abstract ( 106 )   PDF (340KB) ( 318 )   Save BACKGROUND: Homeobox B4 (HOXB4) gene not only promotes amplification of hematopoietic stem cells (HSCs) and activation and expression of its function, but also in vivo test suggested that HOXB4 cannot induce leukemia. Therefore, to study HOXB4 changes and effects in proliferation and differentiation of HSCs provide theoretical basis for HSCs amplification. OBJECTIVE: To observe the expression of HOXB4 gene in the proliferation and differentiation of HSCs into lymphocyte progenitor cells, and analyze the effects of all-train retinoic acid (ATRA) on HOXB4 gene. METHODS: Lymphocyte hematopoietic progenitor cells were divided into two groups. In ATRA group, 6×10-8 mol/L ATRA was added in the culture system. In normal group, an equal volume of 1640 medium was added in the culture system. The colony growth of colony forming unit-lymphocyte in the differentiation progress of the HSCs to lymphocyte progenitor cells induced by phytohemagglutinin was observed on days 3, 7 and 12 following culture. Expression of HOXB4 gene in the proliferation and differentiation of the HSCs into lymphocyte progenitor cells in vitro was detected using real-time fluorescence quantitive polymerase chain reaction. RESULTS AND CONCLUSION: HOXB4 gene has a regulatory function in the differentiation progress of the HSCs into lymphocyte progenitor cells. The expression of HOX B4 gene was diminished gradually in normal and ATRA groups over time. Compared with the normal group, HOXB4 gene was up-regulated in the ATRA group. Related Articles | Metrics

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Immunosuppressive action of human placenta mesenchymal stem cells in mixed lymphocyte reaction system   Xue Yi-feng, Huang Jian-rong, Sun Hong-li, Miao Zong-ning 2010, 14 (36):  6776-6779.  doi: 10.3969/j.issn.1673-8225.2010.36.028 Abstract ( 156 )   PDF (339KB) ( 488 )   Save BACKGROUND: Placenta comprises trophoblasts, blood vessels and plenty of mesenchyme that derived from the mesoderm. Thus, there are mesenchymal stem cells (MSCs) in placenta.  OBJECTIVE: To explore the effect of placenta MSCs (PMSCs) on the proliferation of T lymphocytes and secretion of cytokines in the mixed lymphocyte reaction system, and to evaluate the immunological characteristics of PMSCs. METHODS: The two-way mixed lymphocyte reaction (MLR) system both for human and rabbits were established. After adding PMSCs which was treated by the mitomycin with different ratios (1:5, 1:10, 1:20, 1:50, 1:100, 1:500) in the MLR system, the lymphocyte proliferation rate were measured by 3H incorporation, and the content of interleukin-2 and interferon-γ in the supernatant was detected by ELISA. RESULTS AND CONCLUSION: The proliferation of T lymphocytes could be inhibited to different extents by adding different ratios of PMSCs to human or rabbit PBMCs, and the inhibition rate increased as the ratio of PMSCs to PBMCs increased. Meanwhile, PMSCs could reduce cytokines interleukin-2 and interferon-γ secretion in the mixed lymphocyte reaction system. The PMSCs have the immune regulation ability and inhibit xenogeneic immune cells. Related Articles | Metrics

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Berberine suppresses COX-2 mRNA and protein expression in human peripheral blood mononuclear cells: c-JUN terminal kinase pathway Jiang Xin, Guo Yi, Wang Qi-zhang, Liu Hua-dong, Pang Xin-li 2010, 14 (36):  6780-6784.  doi: 10.3969/j.issn.1673-8225.2010.36.029 Abstract ( 142 )   PDF (470KB) ( 368 )   Save BACKGROUND: When the cells were stimulated by various cytokines and the environment, c-JUN terminal kinase signaling pathway can regulate cell growth, differentiation, apoptosis, cancer, inflammation and the immune response by activating different receptors. OBJECTIVE: To investigate the inhibition of berberine (BBR) on COX-2 mRNA and protein expression via c-JUN terminal kinase signaling cascade pathways in human peripheral blood mononuclear cells. METHODS: Mononuclear cells were isolated and cultured from peripheral vein blood and divided into five groups treated with blank control, lipopolysaccharide (LPS), LPS+BBR 25 µmol/L, LPS+BBR 50 µmol/L, LPS+BBR 100 µmol/L respectively. Monocytes were extracted at 30 minutes, 6 hours, 12 hours and 24 hours following culture. Reverse transcription-polymerase chain reaction (RT-PCR) was utilized to examine COX-2 mRNA levels. Western blot analysis was used to measure c-JUN terminal kinase and COX-2 protein levels. Simultaneously, selectivity c-JUN terminal kinase inhibitor was added to examine COX-2 mRNA and protein expression. RESULTS AND CONCLUSION: Compared with the blank control group, COX-2 mRNA and protein expression of LPS group significantly increased (P < 0.01). COX-2 mRNA and protein expression significantly decreased after different concentrations of BBR treatment (P < 0.05). With the increased concentration of BBR, the COX-2 expression decreased progressively. After the administration of 12 hours, the COX-2 mRNA and protein expression reduced more prominently than that of the other time points. However, there was no significant change in the level of c-JUN terminal kinase activity (P > 0.05). Following the treatment of LPS+ BBR at the concentration of 100 µmol/L, c-JUN terminal kinase activity levels were significant (P < 0.05). COX-2 mRNA and protein expressions were inhibited significantly following incubated with c-JUN terminal kinase inhibitor (P < 0.05). Results have confirmed that BBR inhibits COX-2 mRNA and protein expression in human peripheral blood mononuclear cells in a dose-dependent manner. c-JUN terminal kinase active protein expression can be significantly inhibited by BBR at a high dose. BBR inhibits COX-2 mRNA and protein expression in human peripheral blood mononuclear cells perhaps via c-JUN terminal kinase pathway. Related Articles | Metrics

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Relationship of microvillus-like protrusions to extracellular membrane particles and tunneling nanotubes on the surfaces of human fetal and hepatocellular carcinoma cells revealed by a scanning electron microscope Zhao Xiang, Hu Bai-he, Zhao Xin-rong, Zhang Ye 2010, 14 (36):  6785-6789.  doi: 10.3969/j.issn.1673-8225.2010.36.030 Abstract ( 127 )   PDF (427KB) ( 533 )   Save BACKGROUND: Liver-derived extracellular membrane particles can mediate cell-cell communication and regulate proliferation/differentiation of hepatocytes, and therefore have important application values in diagnosis of liver diseases, in prevention and treatment of liver cancers, and in development of techniques for liver regeneration and cell transplantation. However, progress in the field is hampered by limited knowledge about the origination and typological classification of liver-derived membrane particles. Tunneling nanotube (TNT) is anther newly discovered structure that also mediates cell-cell communication. However, the question whether TNT exists in hepatocytes remains unanswered. OBJECTIVE: By comparing ultrastructural characteristics of the cell surfaces of human normal fetal liver and hepatocellular carcinoma cells, the study is aimed to find the differences between normal and cancerous cells, to uncover the biogenesis of membrane particles so as to accurately classify the particles, and to assure the existence of TNT in liver cells. METHODS: A scanning electron microscope (SEM) was used to probe the samples prepared from cycling adhered fetal hepatocytes L02 and hepatocellular carcinoma BEL-7402 cells to analyze the commonness and difference of cell surface ultrastructure. RESULTS AND CONCLUSION: Extensive microvillus-like protrusions were revealed for the first time on the cell surfaces of both L02 and BEL-7402 cells. The protrusions were characteristically different with the typical microvilli on resting cells. Compared with the BEL-7402 cells, however, the average length and density of the microvillus-like protrusions were longer and higher on the L02 cells, which could be used as a guide to differentiate the two cells. Microvillus-like protrusions from the two cells could either generate nanoparticles or transform into TNT. Two types of nanoparticle biogenesis, budding and pearl-forming constriction, were observed. Taken together, high-resolution SEM images can be served as gold standard to characterize the surface ultrastructures on liver-derived cells, and to pinpoint the sources of extracellular membrane particles. Related Articles | Metrics

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Analysis of tumor stem cells-related side population in intestine cancer cell line SW-620  Gao Gang-long, Zhang Xue-li, Liu Wen-yong, Zhang Ping, Ye Dong-xia 2010, 14 (36):  6790-6793.  doi: 10.3969/j.issn.1673-8225.2010.36.031 Abstract ( 147 )   PDF (302KB) ( 327 )   Save BACKGROUND: In some malignant tumors, side population (SP) cells have the property of tumor stem cells. The studies of human cerebroma and breast carcinoma used this method. At present, studies concerning intestine cancer stem cells are in exploration stage. There is no report on successful isolation and identification of intestine cancer stem cells. OBJECTIVE: To identify the SP of cancer stem-like cells in intestine cancer cell line SW-620. METHODS: SW-620 tumor cells suspension was prepared and stained with Hoechst33342 and propidium iodide. In the experimental group, Hoechst33342 was added until the final concentration was 5 mg/L. In the verapamil group, an additional verapamil was added until the final concentration was 5 mg/L. SP was analyzed using flow cytometry. Results from three tests were selected. RESULTS AND CONCLUSION: Using Hoechst33342 blue and red light double reference graph. In the experimental group, SP cell line located at the lower left quarter region where two kinds of fluorescence were negative or very weak. Following three tests, SP proportion in SW620 was respectively 0.1%, 1.0% and 0.5%. Following verapamil treatment, the SP cell proportion was 0. These indicated that SW620 cell line contains SP cells; i.e. the possibility of SW620 cell line existence. Verapamil can inhibit dye, and obviously decrease the proportion of SP cells. Related Articles | Metrics

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Biological behavior of mouse Lewis lung cancer cells co-cultured with embryonic stem cells in vitro  Meng Long, Zhang Yang-de 2010, 14 (36):  6794-6797.  doi: 10.3969/j.issn.1673-8225.2010.36.032 Abstract ( 174 )   PDF (334KB) ( 351 )   Save BACKGROUND: From a genetic perspective, the tumor cells and embryonic stem cells (ESCs) are regulated by the proto-oncogene and have amazing ability to divide. Embryonic cancer cells were transplanted into blastocysts of normal development mice and produced chimeric mice without the malignant phenotype. The fate of tumor cells can change with embryonic existence. OBJECTIVE: To study the effect of mouse ESCs on the changes in biological behaviors of Lewis lung cancer cells. METHODS: Mouse ESCs were cocultured with Lewis lung cancer cells using Transwell chamber in vitro. In the experimental group, fibroblasts and ESCs were added in the Transwell chamber, whereas tumor cells were in the 12-well plate. In the control group, fibroblasts and fibrocytes were added in the Transwell chamber. The total number of cells were in the chamber did not change, and tumor cells were in the 12-well plate. RESULTS AND CONCLUSION: Compared with the control group, the experimental group shows significant difference in morphology of Lewis lung cancer cells; some cells appear signs of aging and apoptosis. The proliferation of Lewis lung cancer cells was significantly decreased (P < 0.05), and the number of Lewis lung cancer cells that could traverse DB gel biomembrane was significantly reduced in the experimental group (P < 0.05). Results indicate that ESCs can inhibit the growth, proliferation and invasiveness of Lewis lung cancer cells. Related Articles | Metrics

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Application progress of mesenchymal stem cells in tissue engineering    Wang Fu-ke, Liu Liu, Li Jun-nan, Zhao De-ping, Dai Xiao-ming, Li Yi-song, He Xiao-guang 2010, 14 (36):  6800-6804.  doi: 10.3969/j.issn.1673-8225.2010.36.034 Abstract ( 111 )   PDF (363KB) ( 396 )   Save BACKGROUND: Mesenchymal stem cells (MSCs) form connective tissue and organ interstitial in the body. MSCs are characterized by strong proliferative ability, multi-differentiation potential and immunomodulation. With advantages such as convenient source, easy to separate, culture, amplify and purificate, MSCs play an important role in tissue engineering and gene therapy research. OBJECTIVE: To review biological proterty of different sources of MSCs, support role for other cells, immunogenicity, immune regulation and their application. METHODS: The relevant article published between March 2001 and March 2010 were searched for in PubMed database by computer with the key words “mesenchymal stem cells, tissue engineering” . Also searched related article in Wanfang Database with the key words of “mesenchymal stem cells, tissue engineering”. A total of 182 documents were retrieved. RESULTS AND CONCLUSION: MSCs are derived from embryonic mesoderm, have the potential of multi-directional differentiation, and can be applied to tissue engineering. Moreover, MSCs provide support for cell growth, regulate immune state of the body, and have been extensively utilized in medical field. At present, the studies of MSCs function have achieved some successes, but these studies are at early stage and there are many theoretical and technical problems. Further studies are needed to determine how to effectively control the MSCs directional differentiation, proliferation, migration and integration in vivo and how to participate in host cell tissue function. Related Articles | Metrics

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Inducement factors and signal transduction regulation in the osteogenic process of bone marrow mesenchymal stem cells     Li Rui-dong, Huang Wei 2010, 14 (36):  6805-6808.  doi: 10.3969/j.issn.1673-8225.2010.36.035 Abstract ( 120 )   PDF (343KB) ( 414 )   Save BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) can be induced to differentiate into various cells under suitable conditions. In addition, BMSCs, with the characteristics of stromal cells, can secrete various cytokines, support hematopoietic function, and BMSCs are a source of ideal cells for tissue remodeling and function repair. OBJECTIVE: To review various factors that can influence osteogenic process of BMSCs and common signal transduction pathway. METHODS: A computer-based search was performed in PubMed database (1998-01/2009-05) with the key words of “bone mesenchymal stem cell, osteogenesis”. Vip database (1998-01/2009-05) was searched with the key words of “osteogenesis, bone morphogenetic protein, bone marrow mesenchymal stem cells”. RESULTS AND CONCLUSION: A total of 108 articles concerning bone marrow mesenchymal stem cells and bone morphogenetic protein were retrieved, including 31 Chinese articles and 77 English articles. Those published earlier or with poor experimental design and with poor scientific evaluation were excluded, and finally 15 articles were included. Osteogenesis procedure of BMSCs mostly depends on the induced differentiation situation including the basic adjuvants, bone morphogenetic protein, some other cytokines and hormones. In addition, a series of signal pathways plays an important role in the control and interaction of this process. Heat shock protein is worthy of paying attention on affecting the process of proliferation, differentiation and the protective mechanism of signaling adjustment for stimulation outside BMSCs. Related Articles | Metrics

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Regulatory mechanisms and influential factors of mesenchymal stem cell differentiation Hua Yan-long, Li Cai-ping 2010, 14 (36):  6809-6812.  doi: 10.3969/j.issn.1673-8225.2010.36.036 Abstract ( 95 )   PDF (346KB) ( 541 )   Save BACKGROUND: Along with the thorough understanding of mesenchyma stem cells (MSCs) that present widespread application in tissue engineering research. For example, the treatments of anemic cerebrovascular disease, body ischemia and so on. OBJECTIVE: To understand the research progress of the ability of bone marrow mesenchymal stem cell (BMSC) differentiation. METHODS: A computerized search through the PubMed database was undertaken from August 2003 to August 2009. The key words were “mesenchymal stem cell differentiation, Wnt, ß-catenin, bone morphogenetic protein, vascular endothelial growth factor”. Totally, 213 literatures were collected. Articles concerning regulatory mechanisms and influential factors of MSC differentiation were included;Repetitive studies and Meta analysis were excluded.Finally,15 articles were used for the review. RESULTS AND CONCLUSION: With the development of bioengineering disciplines, people know more about the regulation of BMSCs differentiation and influential factors. Many factors found in the survival of MSC microenvironment such as bone morphogenetic protein and vascular endothelial growth factors, which play an important role in regulating cell differentiation. The external factors also play an important role, such as the membrane potential. MSC senescence may affect the ability of cell differentiation. Related Articles | Metrics

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Bone marrow mesenchymal stem cells and sports spinal cord injury Luo Jian-da, Liu Rui-lian, Shi Dian-hui 2010, 14 (36):  6813-6816.  doi: 10.3969/j.issn.1673-8225.2010.36.037 Abstract ( 78 )   PDF (363KB) ( 371 )   Save BACKGROUND: Bone marrow mesenchymal stem cell transplantation is an effective method to treat exercise-induced spinal cord injury (SCI). However, the study concerning SCI is not enough, and lacks of exploration of its mechanism and good therapeutic methods. OBJECTIVE: To provide theoretical basis and scientific support for rehabilitation treatment programs of exercise-induced SCI by analysis of pathological features of exercise-induced SCI, physiological characteristics of bone marrow mesenchymal stem cells and its application to SCI. METHODS: We searched Pubmed database (1991/2010) with the key words of “mesenchymal stem cell, sports knee injuries”. VIP database was searched (1991/2010) with the key words of “bone marrow mesenchymal stem cell, sports knee injuries”. RESULTS AND CONCLUSION: A total of 85 literatures were retrieved, and those published early or with poor quality were excluded. Finally, 25 literatures were included. Bone marrow mesenchymal stem cells were divided into neuron-like cells, reactive secretion of various nutrition factors and growth factors to promote neuroprotective effect and promote local microvessel regeneration, resulting in treating SCI. However, related problems that affected the differentiation and its results of bone marrow mesenchymal stem cells such as ischemia, hypoxia and production of sports free radical and excitatory amino acids following exercise-induced SCI in the injury region require further investigations. Related Articles | Metrics

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Application of stem cells in orthopedics Du Zhi-po, Bei Kang-sheng 2010, 14 (36):  6817-6820.  doi: 10.3969/j.issn.1673-8225.2010.36.038 Abstract ( 104 )   PDF (375KB) ( 508 )   Save BACKGROUND: Stem cells are immature cells that can proliferate and differentiate into multi-lineage adult cells under certain conditions in vitro. Stem cells for treating osteochondral defect, femoral head necrosis and spinal nerve injury have obtained many achievements in clinic. OBJECTIVE: To summarize the new progress of stem cells and its applications in orthopedics in recent years. METHODS: A compute-based online search of Pubmed database from January 2000 to January 2010 and China National Knowledge Infrastructure from January 2000 to January 2010 was performed using the key words of “stem cells, differentiation, bone formation, cytokine, bone fracture, cartilage defect, tissue engineering” in English and “stem cells, cytokine, bone fracture, bone defect, tendon injury, skin destruction, tissue engineering” in Chinese. The experimental studies about stem cells and clinical applications in orthopedics were included. Repeated or out-of-date studies were excluded. Totally, 190 articles were collected and finally 24 were included according to inclusion and exclusion criteria.  RESULTS AND CONCLUSION: As seed cells for tissue engineering, stem cells have extensive usage in vitro, which involves many domains along medical science. Amounts of experiments of stem cells for orthopaedic diseases have succeeded in vitro. Stem cells with multiple-directional differentiation potential can differentiate into osteoblasts, chondroblasts, myoblasts and adipocytes. Combined with corresponding biomaterials, stem cells can repair bone, cartilage, tendon and muscle. Many not only limited in experimental study, but also fit for clinical treatment and obtained satisfactory outcomes. Related Articles | Metrics

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Embryonic origins of hematopoiesis Xie Yi-fan, Wu Yan 2010, 14 (36):  6821-6824.  doi: 10.3969/j.issn.1673-8225.2010.36.039 Abstract ( 1021 )   PDF (306KB) ( 1854 )   Save BACKGROUND: The process consists of primitive hematopoiesis and definitive hematopoiesis in mammalian. It has been approved that primitive hematopoiesis emergence from yolk sac. However, the original sites of definitive hematopoiesis have been a controversial issue in the field of hematopoiesis. OBJECTIVE: To explore embryonic origins of definitive hematopoiesis. METHODS: The first author retrieved PubMed database for articles published from January 2000 to March 2010. The key words were “hematopoietic stem cell, hematopoiesis, yolk sac, aorta-gonad-mesonephros (AGM) region”. Simultaneously, China National Knowledge Infrastructure database was also retrieved for relevant articles published from January 2000 to March 2010. The key words were “hematopoietic stem cell, hematopoiesis, yolk sac, AGM region”. A total of 682 articles were obtained, and the literature of repetitive content was excluded. Finally, 42 articles were included. RESULTS AND CONCLUSION: Traditional opinions thought the definitive hematopoiesis origins from yolk sac of extra-embryo. However, a series of studies showed that the definitive hematopoiesis come from AGM region of intra-embryo first. The placenta is a newly discovered site which may generate hematopoietic cells during period from AGM region to fetal liver. In the yolk sac and PSp/AGM region, the progenitor cell of hematopoietic cell presents two different models. The hematopoietic cells and endothelial cells are normally derived from the hemangioblast in the yolk sac. However, in the AGM region, hematopoietic stem cells come from haemogenic endothelium of the dorsal aorta ventral wall. In addition, hematopoitic development process is regulated by several signal pathways. Related Articles | Metrics

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Effects and possible molecular mechanisms of Schwann cells for the repair of peripheral nerve damage    Gao Wei, Shao Shui-jin, Hu Lin-na, Liu Yan-xiang 2010, 14 (36):  6825-6827.  doi: 10.3969/j.issn.1673-8225.2010.36.040 Abstract ( 188 )   PDF (270KB) ( 733 )   Save BACKGROUND: The recovery of neural function following peripheral nerve damage has been the focus. Schwann cells have very important effects on the recovery of neural function. OBJECTIVE: To summarize the effect and possible molecular mechanism of Schwann cells in promoting neural function recovery. METHODS: The databases of China National Knowledge Infrastructure, VIP and Offshore Vessel Inspection Database were searched for articles about peripheral nerve damage published from January 1993 to January 2010. The key words were “Schwann cells, peripheral nerve, nerve regeneration and review”, which were searched by inputting in titles and abstracts. The articles addressing Schwann cells effects on peripheral nerve damage were selected. The literatures in the same field that published recently or in authorized journals were included. There were 256 articles after the initial survey. Finally, 24 articles concerning action mechanism of Schwann cells according to inclusion criteria were included for this review.  RESULTS AND CONCLUSION: Schwann cells affect the damaged peripheral nerve through various channels and promote poor functional recovery. How to accelerate proliferation of Schwann cells will be a breakthrough point for the recovery of neural function. By synthesizing various factors, electroacupuncture for peripheral nerve injury will have a better future. Related Articles | Metrics

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Cardiomyogenesis of human umbilical cord mesenchymal stem cells induced by conditional culture in the presence of sphingosine-1-phosphate Zhao Zhen-qiang, Chen Zhi-bin, Cai Mei-hua, Wang Shu-rong, Chen Rong, Wang Tan，Yuan Kun-xiong, Rong Qiong-wen 2010, 14 (36):  6828-6832.  doi: 10.3969/j.issn.1673-8225.2010.36.041 Abstract ( 114 )   PDF (464KB) ( 405 )   Save BACKGROUND: Several studies have demonstrated that sphingosine-1-phosphate (S1P) can induce the differentiation of adipose-derived stem cells differentiation into smooth muscle cells. Whether S1P, rather than 5-azacytidine, can be used as an inducer of mesenchymal stem cells differentiation into cardiomyocytes remains unclear.  OBJECTIVE: To investigate the possibility that S1P promotes human umbilical cord mesenchymal stem cells (HUMSCs) differentiation into cardiomyocytes in the presence of different culture media. METHODS: HUMSCs were cultured with cardiomyocyte conditional medium (CMCM) and/or S1P media. At 1, 5, and 10 days of culture, morphological changes of HUMSCs were observed. After culture for 10 days, the induced cells were confirmed by immunocytochemical analysis and patch clamp in terms of cell phenotype and function. RESULTS AND CONCLUSION: During the induction, some cells gradually enlarged, elongated, connected with adjacent cells, and formed myotube-like structures, and some cells congregated into cell clusters in the CMCM and CMCM+S1P groups. In the CMCM+S1P group, cells exhibited special perpendicular terrace-shaped, intercalated disc-like arrangement. Immunohistochemistry results revealed that some cells strongly express specific antibodies against sarcomeric myosin and α-actinin in the CMCM and CMCM+S1P groups. These findings suggest that HUMSCs can be induced to differentiate into cardiomyocytes. Through the use of patch clamp technique, a rapid ascending, but without plateau phase, action potential, a voltage dependent inward current, and a voltage dependent outward current were recorded in some cells from the CMCM+S1P group. These findings indicate that S1P plays a key role in promoting cardiomyogenic differentiation of HUMSCs and functional integration. Related Articles | Metrics

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Equol reverses the inhibition of cyclosporin A on the proliferation and osteoblastic differentiation of bone marrow mesenchymal stem cells through estrogen receptor/ nitric oxide / cyclic guanosine monophosphate signal pathway Song Li-hua, Huang Yan, Wu Cui-ling, Shi Bian-hua 2010, 14 (36):  6833-6836.  doi: 10.3969/j.issn.1673-8225.2010.36.042 Abstract ( 150 )   PDF (427KB) ( 340 )   Save BACKGROUND: Long term taking cyclosporin A (CsA) can inhibit osteoblastic differentiation and induce osteoporosis. Equol, which has greater binding affinity to estrogen receptors, can stimulate the proliferation and osteoblastic differentiation. OBJECTIVE: To investigate whether equol may protect against the proliferation and osteoblastic differentiation inhibited by CsA in mouse bone marrow mesenchymal stem cells (BMSCs) cultures and analyze signal pathway of protection. METHODS: Primary mouse BMSCs were cultured by using attachment method and assigned to five groups, which respectively treated with equol or/and CsA in the presence or absence of ICI182780, an estrogen receptor antagonist, and Nω-nitro-L-arginine methyl ester. Under an inverted microscope, morphological changes and mineralization ability of BMSCs were observed. The cell proliferation was measured by [3H]-thymidine incorporation. The osteoblastic differentiation and mineralization of extracellular matrix in BMSCs was assessed by measuring alkaline phosphatase activity and calcium deposition, respectively. Nitric oxide production in the conditioned media and cyclic guanosine monophosphate (cGMP) content in BMSCs were determined by using commercial nitric oxide and cGMP kit, respectively. RESULTS AND CONCLUSION: Equol reversed the decreased [3H] thymidine incorporation (P < 0.05), alkaline phosphatase activity (P < 0.05) and calcium deposition (P < 0.01) of CsA, which was accompanied with the changes of nitric oxide production (P < 0.01) and cGMP content (P < 0.01). The group by co-treatment with equol and CsA possessed higher cells growth density and small mineralized nodes than CsA group on day 12 under an inverted microscope. Moreover, the equol-reversed effect was abolished by ICI182780 and Nω-nitro-L-arginine methyl ester. These indicated that equol can reverse the inhibition of CsA on the proliferation and osteoblastic differentiation of mouse BMSCs through estrogen receptor/ nitric oxide/cGMP signal pathway. Related Articles | Metrics

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Effects of erythropoietin on nestin expression in neural stem cells of neonatal rats with hypoxia-ischemia brain damage Jiang Hong, Xu Feng, Zhou Chun-qing, Li Xiang-hong, Shu Zhi-rong 2010, 14 (36):  6837-6840.  doi: 10.3969/j.issn.1673-8225.2010.36.043 Abstract ( 109 )   PDF (268KB) ( 278 )   Save BACKGROUND: Nestin is a specific antigen of neural stem cells which widely expressed in lesion of nervous system and brain regeneration. Thus, nestin expression is commonly used to assess whether lesion or damage of the nervous system can promote neural regeneration. OBJECTIVE: To investigate the effects of erythropoietin (Epo) on nestin expression in neural stem cells after hypoxia-ischemia brain damage (HIBD) in neonatal rats from the angles of neural regeneration and activation of neural stem cells. METHODS: HIBD model was established by ligation of the right common carotid artery along with 2-hour 8% hypoxia exposure in neonatal rats. The control group was not subjected to hypoxia-ischemia, and the right common carotid artery was dissociated. The treatment group received an intraperitoneal injection of recombinant human erythropoietin (rh-Epo, 5 000 IU/kg) once a day for three days after hypoxia/ischemia, while the two other groups intraperitoneally received normal saline at the same time. In each group, rats were randomly executed immediately, at 4, 7, 14 days after operation (n = 8). The nestin expression in hippocampal dentate gyrus region was examined by immunohistochemical staining and image quantitative analysis respectively. RESULTS AND CONCLUSION: The number of nestin-positive cells was significantly increased in HIBD group compared to control group at all time points (P < 0.05), and it was also significantly increased in treatment group than the other two groups at all time points (P < 0.05). The numbers of nestin-positive cells in hippocampal dentate gyrus region were significantly increased, and peaked on day 7 after operation in the three groups. The results showed that exogenous rh-Epo could enhance the expression of nestin in hippocampal dentate gyrus region of neonatal rats with HIBD, and promote the proliferation of neural stem cells. rh-Epo plays an important role in the regeneration and repair of neurocytes damaged by hypoxia-ischemia. Related Articles | Metrics