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04 November 2012, Volume 16 Issue 45 Previous Issue    Next Issue

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Expression of transient receptor potential vanilloid receptor 5 during osteogenic differentiation of bone marrow mesenchymal stem cells Li Fu-chun, Ma Xue-ling, Qu Yan-ping, Gu Gui-shan 2012, 16 (45):  8361-8366.  doi: 10.3969/j.issn.2095-4344.2012.45.001 Abstract ( 306 )   PDF (526KB) ( 554 )   Save BACKGROUND: At present, the calcium ion channel research is confined in the electrophysiological properties demonstrated by transient receptor potential vanilla receptor 5 in the kidney, small intestine and smooth oocytes. OBJECTIVE: To explore the expression and significance of new calcium ion channel transient receptor potential vanilla receptor 5 in rat bone marrow mesenchymal stem cells derived osteoblasts. METHODS: The rat bone marrow mesenchymal stem cells were separated, extracted and cultivated, then the rat bone marrow mesenchymal stem cells were induced to differentiate into osteoblasts, the expression of transient receptor potential vanilla receptor 5 in osteoblasts derived from rat bone marrow mesenchymal stem cells was detected by immunohistochemistry. RESULTS AND CONCLUSION: The in vitro cultured rat bone marrow mesenchymal stem cells were successfully induced to differentiate into osteoblasts. Alkaline phosphatase staining and alizarin red staining results showed that the purple-black and red mineralized nodules were visible in the cytoplasm of the osteoblasts derived from rat bone marrow mesenchymal stem cells. Immunohistochemical staining results showed the strongly positive expression of transient receptor potential vanilla receptor 5 in bone marrow mesenchymal stem cells derived osteoblasts. Results confirm that transient receptor potential vanilla receptor 5 calcium ion channel exists in rat bone marrow mesenchymal stem cells derived osteoblasts, and the transient receptor potential vanilla receptor 5 calcium ion channel may be the candidate channel which can regulate the osteoblasts proliferation and differentiation. Related Articles | Metrics

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Effect of different magnitudes of continuous tensile stress on osteogenic differentiation of rat bone marrow stromal cells Dai Qing-gang, Zhang Peng, Wu Yu-qiong, Fu Run-qing, Zhao Jing-lei, Yang Xiao, Liu Jia-qiang, Jiang Ling-yong, Fang Bing 2012, 16 (45):  8367-8373.  doi: 10.3969/j.issn.2095-4344.2012.45.002 Abstract ( 347 )   PDF (541KB) ( 451 )   Save BACKGROUND: Mechanical stimulation can promote the osteogenic differentiation of bone marrow stromal cells, but the effect of different magnitude of tensile stress on the bone marrow stromal cells is not clear. OBJECTIVE: To study the effect of different magnitudes of continuous tensile stress on the osteogenic differentiation of rat bone marrow stromal cells. METHODS: Rat bone marrow stromal cells were obtained and purified by full-blood attachment culture. The 5%, 10% and 15% continuous tensile stress (1 Hz) were strained on bone marrow stromal cells with Flexercell-4000 mechanical loading system and lasted for 48 hours, and the cells in the control group were cultured without stress. The mRNA and protein expression of osteoblast-related genes alkaline phosphatase, collagen type Ⅰ, osteocalcin and osteoblast-specific transcription factor Runx2 were analyzed at 1, 6, 12, 24 and 48 hours after strain. RESULTS AND CONCLUSION: The mRNA expression of alkaline phosphatase, collagen type Ⅰ and osteocalcin was increased in rat bone marrow stromal cells subjected to 5% and 10% continuous tensile stress when compared with the control group (P < 0.05), but the 10% tensile stress group increased earlier than the 5% tensile stress group, and the amplitude was higher. The mRNA expression of alkaline phosphatase and collagen type Ⅰ was increased at 6 hours after 15% elongation (P < 0.05), and then decreased gradually. At 48 hours after continuous tensile stress, the indicators above were lower than those in the control group (P < 0.05). The expression of Runx2 protein in bone marrow stromal cells at 6 hours in 15% elongation was higher than that in the control group (P < 0.05), so did the 5% and 10% elongation groups (P < 0.05). It indicates that 5%, 10% and 15% tensile stress can promote the osteogenic differentiation of bone marrow stromal cells, and 10% tonsile stress has the most significant effect. References | Related Articles | Metrics

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Effect of human bone marrow mesenchymal stem cells culture supernatant on related enzyme expression in hepatic stellate cells Wang Shan, Zhang Li-ting, Chen Hong 2012, 16 (45):  8374-8379.  doi: 10.3969/j.issn.2095-4344.2012.45.003 Abstract ( 329 )   PDF (534KB) ( 521 )   Save BACKGROUND: The studies have proved that bone marrow mesenchymal stem cells transplantation therapy can improve or even reverse the liver fibrosis, but its mechanism is still unclear. OBJECTIVE: To detect the effect of human bone marrow mesenchymal stem cells culture supernatant on the expression of matrix metalloproteinase 2 and tissue inhibitor of metalloproteinase 1 in hepatic stellate cells. METHODS: The density gradient centrifugation was used to isolate human bone marrow mesenchymal stem cells and collected the culture supernatant of human bone marrow mesenchymal stem cells after cultured for 7 days, the human bone marrow mesenchymal stem cells culture supernatant added with the hepatic stellate cells was considered as experimental group, a simple hepatic stellate cells group and simple human bone marrow mesenchymal stem cells group were set up as control groups. The expressions of matrix metalloproteinase-2 protein and tissue inhibitor of matrix metalloproteinase-1 in the culture supernatant of three groups were detected after cultured for 24 and 48 hours. RESULTS AND CONCLUSION: The expression of matrix metalloproteinase-2 protein and tissue inhibitor of matrix metalloproteinase-1 in the experimental group was decreased when compared with simple hepatic stellate cells group and simple human bone marnow mesenchymal stem cells group (P < 0.05 or P < 0.01). The results showed that bone marrow mesenchymal stem cells culture supernatant can inhibit the expression of matrix metalloproteinase-2 protein and tissue inhibitor of matrix metalloproteinase-1 in hepatic stellate cells. Related Articles | Metrics

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Correlation between Stro-1+ cells and bone morphogenetic protein and its receptor in proximal femur Shen Yi, Li Xiao-miao, Fu Xiao-dong, Wang Wei-li 2012, 16 (45):  8380-8384.  doi: 10.3969/j.issn.2095-4344.2012.45.004 Abstract ( 393 )   PDF (405KB) ( 536 )   Save BACKGROUND: A successful osseointegration after total hip arthroplasty relies on the bone reconstruction around the prosthesis. Bone marrow mesenchymal stem cells are recruited and the commitment of bone marrow mesenchymal stem cells to an osteoblastic differentiation pathway is apparently under the control of bone morphogenic proteins. OBJECTIVE: To analyze the correlation between Stro-1+ bone marrow mesenchymal stem cells and bone morphogenetic protein 7 and bone morphogenetic protein receptor in proximal femur. METHODS: Bone samples were obtained from the discarded metaphysis region of the proximal femur in 32 patients with primary total hip arthroplasty, and the number of bone marrow mesenchymal stem cells was measured after in vitro cultured for 14 days. Stro-1+ was used to mark the stem cells. The expression of bone morphogenetic protein 7, bone morphogenetic protein receptor 1a and bone morphogenetic protein receptor 2 was detected. RESULTS AND CONCLUSION: There were differences of Stro-1+ cells, bone morphogenetic protein 7, bone morphogenetic protein receptor 1a and bone morphogenetic protein receptor 2 expression between different years, but the differences were not significant, and there was no significant difference between genders. There was a negative correlation between the numbers of Stro-1+ cells in proximal femur and expression of bone morphogenetic protein 7 that means the cell had the function to differentiate into osteoblasts. The correlation between the number of Stro-1+ cells and expression of bone morphogenetic protein receptor 1a was positive which can influence the fusion rate and the survival rate of the hip prosthesis. Related Articles | Metrics

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Changes in T2 value and related factor expression after treatment of early osteoarthritis by bone marrow mesenchymal stem cells transplantation Hu Xu-yu, Cui Yan-an, Liu Zhao, Zhuang Chao 2012, 16 (45):  8385-8389.  doi: 10.3969/j.issn.2095-4344.2012.45.005 Abstract ( 505 )   PDF (455KB) ( 601 )   Save BACKGROUND: Serum tumor necrosis factor α and interleukin-1β as an inflammatory mediator is related to the condition of osteoarthritis. However, the reports on the treatment of osteoarthritis by bone marrow mesenchymal stem cells are rare. OBJECTIVE: To observe the T2 value, the serum tumor necrosis factor α and interleukin-1β levels in the treatment of early osteoarthritis in rabbits by bone marrow mesenchymal stem cells transplantation. METHODS: Thirty-six New Zealand white rabbits were randomly and equally divided into control group, model group and treatment group. Rabbits in the model group were only used to establish the osteoarthritis model; left knee joint cavity of rabbits in the treatment group was injected with bone marrow mesenchymal stem cells at 4 weeks after modeling; knee joint cavity of rabbits in the control group was injected with normal saline in the same dose. Left knee joint magnetic resonance imaging and the detection of serum tumor necrosis factor α and interleukin-1β levels were performed at 2 weeks, 1, 2 and 3 months after treatment. RESULTS AND CONCLUSION: Compared with the meodel group, the T2 value reduced, without arthroedema, and the serum tumor necrosis factor α and interleukin-1β levels were decreased in the treatment group. The T2 value was positive correlated with serum tumor necrosis factor α and interleukin-1β levels. MRI can reflect the early biologically change of articular cartilage, and it can be used to evaluate the curative effect in the early stages of osteoarthritis. Bone marrow mesenchymal stem cells can differentiate into chondrocytes in articular cavity and is effective for early osteoarthritis in rabbits. Related Articles | Metrics

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Bone marrow mesenchymal stem cells migrate to the zone of spinal cord injury Jia Quan-zhang, Li Dong-jun, Chen Yu-bing, Sun Jing-hai, Wu Chuan-zhen, Wang Feng-hua, Xu Shuang, Liu Li-ping, Jiang Da-wei, Gao De-xuan 2012, 16 (45):  8390-8393.  doi: 10.3969/j.issn.2095-4344.2012.45.006 Abstract ( 359 )   PDF (387KB) ( 441 )   Save BACKGROUND: How to observe the survival and prognosis of bone marrow mesenchymal stem cells after migration to the spinal cord injury zone remains controversial. OBJECTIVE: To observe the migration situation of bone marrow mesenchymal stem cells in the rat spinal cord injury zone. METHODS: Thirty-six Wistar rats were randomly divided into two groups. In the experimental group, the rats were used to make the spinal cord injury model, and after 1 week, 1 mL DAPI labeled bone marrow mesenchymal stem cells (1×109/L) were daily transplanted into the rats through the tail vein for 2 days. The rats in the control group were not subjected to spinal cord injury induction, but they received bone marrow mesenchymal stem cells transplantation at the same time with the same method. The frozen sections of injured spinal cord were prepared at 5, 10 and 15 days after transplantation, and the migration of bone marrow mesenchymal stem cells was observed under the confocal laser scanning microscope. RESULTS AND CONCLUSION: At 5 days after the transplantation in the experimental group, the fluorescence marked bone marrow mesenchymal stem cells were observed in the vessels of spinal cord injury tissue; 10 days after transplantation, bone marrow mesenchymal stem cells were dispersed in extravascular tissue; 15 days after transplantation, the fluorescence marked bone marrow mesenchymal stem cells were widely diffused in the injury tissue. While, the DAPI labeled bone marrow mesenchymal stem cells were not observed in the control group. Results show that bone marrow mesenchymal stem cells can migrate to the injured spinal tissue through spinal cord-blood barrier after transplantation through the tail vein. Related Articles | Metrics

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A modified method of isolating and culturing human bone marrow mesenchymal stem cells Liu Ji-chun, Hu Hong-tao, Xu Guo-hua, Jiang Yu-quan, Xu Ning, Ye Xiao-jian 2012, 16 (45):  8394-8397.  doi: 10.3969/j.issn.2095-4344.2012.45.007 Abstract ( 352 )   PDF (469KB) ( 597 )   Save BACKGROUND: Experiments for the isolation and culture of rat or rabbit bone marrow mesenchymal stem cells are more common, but the clinical experiments of tissue engineering are often on the basis of the seed cells of the same species. OBJECTIVE: To establish a protocol for isolating and culturing human bone marrow mesenchymal stem cells which is stable and efficient and consistent with the needs of clinical and laboratory experiments. METHODS: Bone marrow and spongy bone were obtained from the human anterior superior iliac spine, and then the bone marrow, the spongy bone was flushed by 15 mL culture medium and then the rinse solution was collected. The cells were primary screened on the basis of adhesion function by direct cultivation method; the surface antigens, including CD29, CD34, CD44, CD45 and CD105 were detected by flow cytometry. Bone marrow mesenchymal stem cells were differentiated into osteoblasts and adipocytes, and the differentiated mesenchymal stem cells were identified by morphological observation. RESULTS AND CONCLUSION: Cell colony could be seen at 5 days after cultured with spongy bone washing liquid. These cells were uniformly negative for CD34, CD45 and positive for CD29, CD44 and CD105. Calcium nodules were observed after osteogenic induction for 20 days and positive for alizarin red staining. The lipid droplets could be seen after adipogenic induction for 7 days and oil red O staining showed a large number of lipid deposition. The study shows that a large amount of mesenchymal stem cells can be isolated and cultured from adult human spongy bone in short time by direct cultivation methods, and the mesenchymal stem cells are uniform in morphology, the cells can express the antigens stably and differentiate into osteoblasts and adipocytes. References | Related Articles | Metrics

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Superparamagnetic iron oxide nanoparticles label human bone marrow and umbilical cord mesenchymal stem cells Ma Yan, Zhang De-qing, Chen Le, Wang Jian, Zhang Xue, Hou Yan, Bi Xiao-juan, Yang Rong, Hu An-hua 2012, 16 (45):  8398-8405.  doi: 10.3969/j.issn.2095-4344.2012.45.008 Abstract ( 289 )   PDF (527KB) ( 469 )   Save BACKGROUND: Nowadays, it is becoming more and more important to optimize safety of human derived cells, label cells efficiently and track cells after cells transplantation both in basic research and clinic application. OBJECTIVE: To compare the cell viability, labeling efficiency and imaging effect of the T2* weight image (WI) magnetic resonance (MR) between the human bone marrow and umbilical cord derived mesenchymal stem cells labeled with the superparamaganetic iron oxide nanoparticles, as well as to optimize their treatment efficiency. METHODS: The third generation of human bone marrow and umbilical cord derived mesenchymal stem cells were cultured, and labeled with 5-30 mg/L Feridex Ⅳ and protamine sulfate. RESULTS AND CONCLUSION: The viability of human bone marrow mesenchymal stromal cells was similar with human umbilical cord derived mesenchymal stem cells (P > 0.05). There was no significant difference of labeling rate between the bone marrow msenchymal stem cells labeled with 5-30 mg/L Feridex Ⅳ (P > 0.05); while there was significant difference of labeling rate between the umbilical cord derived mesenchymal stem cells labeled with 5 mg/L Feridex Ⅳ and 20 and 30 mg/L Feridex Ⅳ (P < 0.05); the positive labeling rate of umbilical cord derived mesenchymal stem cells was lower than that of bone marrow msenchymal stem cells after labeled with 10 mg/L Feridex Ⅳ (P < 0.05). When two sources of cells were labeled with Feridex Ⅳ more than 2 mg/L, the iron oxide particles were found in the cell suspension and could not be removed by elution and filtration. The signal intensity from 3.0T MR GRE T2*WI scan was decreased with the increasing of Feridex Ⅳ concentration in both cell types. It is safe and effective to label the two tissue-derived mesenchymal stem cells with 10 mg/L Feridex Ⅳ-protamine sulfate complex, and can be observed with T2*WI MR. Related Articles | Metrics

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Efficient method for isolation of human umbilical cord mesenchymal stem cells Dong Min, Chen Xian-jiu, Ma Yi-qiong, Zhao Jie, Xue Guo-fang, Chen Yan, Niu Bo 2012, 16 (45):  8406-8412.  doi: 10.3969/j.issn.2095-4344.2012.45.009 Abstract ( 1567 )   PDF (593KB) ( 1267 )   Save BACKGROUND: The theoretical value of the mesenchymal stem cells from human umbilical cord isolated by the traditional methods is far away from that of the mesenchymal stem cells from human umbilical cord contained in the human umbilical cord tissues, which lead to the great waste of the umbilical cord tissues and prevent the large-scale preparation and culture of mesenchymal stem cells. OBJECTIVE: To optimize and develop a new efficient method for the isolation of human umbilical cord mesenchymal stem cells in order to provide the technical solutions for the clinical application of mesencymal stem cells. METHODS: The experiment was divided into four groups. The control group was treated with traditional enzymatic method (trypsin and collagenase type II), on this basis, the collagenase type Ⅳ, hyaluronidase and DNase were added into the umbilical cord tissues for digestion. The indicators obtained through different isolation methods and different operation time were compared, including the amount of the cells, cells activity, time for the occurrence of the primary cell adherent and fusion time, as well as the proliferative capacity, surface markers and differentiation capacity of the passage 3 umbilical cord mesenchymal stem cells. RESULTS AND CONCLUSION: Collagenase type II, collagenase type IV, trypsin, hyaluronidase, and DNAase were used to digest the umbilical cord, thus cell count reached up to (12.47±0.16)×106/cm (P < 0.01) in 4 hours; cell activity was (75.00±5.07) (P < 0.01); CD105 positive cell rate was 41.1%; and CD29 positive cell rate was 83.1%, these measurement indicators in the experimental groups were higher than those in the control group; the obtained cells got adherent and associated fusion, and the time in the experimental groups was shorter than that in the control group (P < 0.01); the cells gradually got uniform shape, the cell morphology of the third-passage was spindle-shaped and swirling. The positive rates of the specific markers CD44, CD105 and CD29 of passage 3 human umbilical cord mesenchymal stem cells were higher than those in the control group, and the positive rate of CD34 in the experimental groups was lower than that in control group. After induction to fat cells for 4 weeks, differentiation results were assayed by oil red O staining, showing that a large number of lipid droplets were observed in cells; the differentiation capacity to osteoblasts was measured by alizarin red staining, and visible positive calcium deposition was observed. The experiment demonstrated that the combination of collagenase type Ⅱ, collagenase type Ⅳ, hyaluronidase, trypsin and DNAase for digestion of umbilical cord for 4 hours could significantly improve the yield, viability and proliferation ability of mesenchymal stem cells and shorten the time of isolation. In addition, the time for primary mesenchymal stem cells to adhere and grow to confluence was shortened by using the combined digestion method than common digestion method. And the ratio of mesenchymal stem cells in the primary mesenchymal stem cells and the purity of the passage three mesenchymal stem cells obtained using combined digestion method are much better than those isolated using common digestion method. Related Articles | Metrics

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Transfection of an eukaryotic expression vector expressing brain derived neurotrophic factor into human umbilical cord mesenchymal stem cells Sun Zhao-ming, Yuan Yuan, Pan Shu-mao, Zhang Ju-hua, Wang Yuan-li, Tang Guo-tai, Guan,Mao-wu, Chen Hong-guang 2012, 16 (45):  8413-8418.  doi: 10.3969/j.issn.2095-4344.2012.45.010 Abstract ( 435 )   PDF (551KB) ( 559 )   Save BACKGROUND: Stem cells transplantation is the research hot spot in the area of nerve injury repair. At present, most researches have focused on neural stem cells and bone marrow stem cells transplantation, and there are rare reports on the research of umbilical cord mesenchymal stem cells, but the umbilical cord mesenchymal stem cells have the advantages of rich source and little ethical restrictions. OBJECTIVE: To construct an eukaryotic expression vector expressing brain derived neurotrophic factor, and to transfect it into the umbilical cord mesenchymal stem cells and establish a cell model of brain derived neurotrophic factor. METHODS: Firstly, the c-DNA of brain derived neurotrophic factor was gained by reverse transcription-PCR and then inserted into multi-clone site of pcDNAⅢ vector to construct the expression vector of brain derived neurotrophic factor pcDNAⅢ. Then the vector was used to transtect umbilical cord mesenchymal stem cells. At last, the expression level of brain derived neurotrophic factor in the umbilical cord mesenchymal stem cells was tested by Western blot; the cultured PC12 cells and the embryo rat pallium neurons were added into the umbilical cord mesenchymal stem cells containing brain derived neurotrophic factor to test the biological activity of brain derived neurotrophic factor. RESULTS AND CONCLUSION: The eukaryotic expression vector expressing brain derived neurotrophic factor was successfully constructed, and this vector could highly express the brain derived neurotrophic factor protein in umbilical cord mesenchymal stem cells. In vitro testing showed the brain derived neurotrophic factor protein had the biological activity. It indicates that the umbilical cord mesenchymal stem cells can be used as the eukaryotic expression vector of brain derived neurotrophic factor for research on the nerve injury repair. Related Articles | Metrics

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Bone morphogenetic protein 2 expression in umbilical cord mesenchymal stem cells after adipogenic induction Chi Ying, Rong Li-juan, Xu Fang-yun, Han Zhi-bo, Yang Shao-guang, Wang You-wei, Ma Feng-xia, Lu Shi-hong, Han Zhong-chao 2012, 16 (45):  8419-8423.  doi: 10.3969/j.issn.2095-4344.2012.45.011 Abstract ( 317 )   PDF (597KB) ( 452 )   Save BACKGROUND: Expression of bone morphogenetic protein 2 and peroxisome proliferator-activated receptorγgene in umbilical cord mesenchymal stem cells after adipogenic induction is increased at the same time. OBEJECTIVE: To investigate the changes of bone morphogenetic protein 2 and peroxisome proliferator-activated receptorγgene expression in umbilical cord mesenchymal stem cells after adipogenic induction, and to compare the results. METHODS: With specific inductive medium, umbilical cord mesenchymal stem cells were obtained and then induced into adipogenisis. Their morphological characteristics were observed with inverted microscope. Oil red O staining was used to observe the adipogenic phenomenon, and von kossa staining and alizarin red staining were used to identify the formation of precipitation. Real-time polymerase chain reaction was used to quantify the gene expression of bone morphogenetic protein 2 and peroxisome proliferator-activated receptor γ at various time points after adipogenic induction. RESULTS AND CONCLUSION: Umbilical cord mesenchymal stem cells could be isolated and cultured successfully. After cultured in adipogenic medium, the cells differentiated into adipogenisis. The result of real-time polymerase chain reaction showed that both the expression levels of bone morphogenetic protein 2 and peroxisome proliferator-activated receptorγgene were increased after adipogenic induction, but peroxisome proliferator-activated receptorγplayed a major role in the regulation of differentiation. Related Articles | Metrics

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Construction of vascularized adipose using adipose-derived stem cells and autogeneic platelet-rich plasma carrier complex in vivo by tissue engineering Li Hong-mian, Liu Da-lie, Zhao Pei-ran, Liang Shuang-wu 2012, 16 (45):  8424-8429.  doi: 10.3969/j.issn.2095-4344.2012.45.012 Abstract ( 250 )   PDF (581KB) ( 571 )   Save BACKGROUND: Revascularization mechanism is a determinal factor of successful construction of adipose tissue by tissue engineering. OBJECTIVE: To investigate the feasibility of construction of vasuclarized adipose using adipose-derived stem cells and autogeneic platelet-rich plasma carrier complex in vivo by tissue engineering。 METHODS: Adipose-derived stem cells were isolated from the subcutaneous adipose tissue of healthy adult after liposuction, and primary culture and subculture of adipose-derived stem cells were conducted. After being induced towards adipocytes for 2 weeks, 5× 1010/L passage 3 cell suspension labeled by BrdU was prepared. Two groups were included: experimental group, in which 0.5 mL cell suspension, 100 μL platelet-rich plasma and 0.5 mL fibrin glue were implanted into the subcutaneous fascia of nude mice; control group in which 0.5 mL cell suspension, 100 μL DMEM and 0.5 mL fibrin glue were implanted into the subcutaneous fascia of nude mice. RESULTS AND CONCLUSION: At 8 weeks after surgery, neogenetic vessels grew into the scaffolds and mild fiber encapsulation was observed in the experimental group, while few vessels grew into the scaffolds and mild fiber encapsulation was also observed in the control group. The wet weight of cambium in the experimental group was higher than that in the control group (P < 0.01). Hematoxylin-eosin staining showed the formation of neogenetic adipose tissues and the growth of micrangium in the implant. The number of micro vessels in the experimental group was greater than that in the control group (P < 0.01). The immunofluorescence staining of cambium showed that the cell nucleus of regenerated adipocytes and partial capillary endothelium in both groups presented green fluorescence. It is feasible to prepare vasuclarized adipose using adipose-derived stem cells and autogeneic platelet-rich plasma carrier complex in vivo by tissue engineering. Adipose-derived stem cells and autogeneic platelet-rich plasma participate in neovascularization of neogenetic adipose tissue. The autogeneic platelet-rich plasma can promote revascularization of tissue-engineered complex and ensure survival of more seed cells. Related Articles | Metrics

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Immune regulatory effect of human adipose-derived mesenchymal stem cells Wang Jian-jun, Fan Yan, Ma Ai-qun, Gao Ming-xuan 2012, 16 (45):  8430-8434.  doi: 10.3969/j.issn.2095-4344.2012.45.013 Abstract ( 313 )   PDF (458KB) ( 535 )   Save BACKGROUND: We know little about the immune regulatory effect of adipose-derived mesenchymal stem cells. OBJECTIVE: To investigate the influence of adult adipose-derived mesenchymal stem cells on lymphocyte proliferation, cells subsets and the level of cell factors in vitro, in order to understand the immune regulatory effect. METHODS: Adipose-derived mesenchymal stem cells were isolated from adult adipose tissue. The phenotype was identified with the flow cytometry. Adipose-derived mesenchymal stem cells were induced into osteoblasts and myocardial cells, respectively, and identified with the immunocytochemistry staining. The human peripheral blood lymphocytes were separated and cultured with adipose-derived mesenchymal stem cells according to different concentrations (1:1, 1:10, 1:100), and added the plant haemagglutinin for simulation. The control group was set up, and after 3 days, the culture fluid was collected. RESULTS AND CONCLUSION: The absorbance value of adipose-derived mesenchymal stem cells in the treatment group after co-cultured with lymphocytes with three different concentrations was significantly lower than that in the control group, and 1:1 concentration had the lowest absorbance value (P < 0.05); the proportion of CD4+CD25+ Tregs in the treatment groups was significantly higher than that in the control group, and highest in the 1:1 concentration group. The cell factors of interleukin-2, interleukin-4 and γ-interferon were obviously reduced in the treatment groups, while the interleukin-10 level was increased, and showed a certain concentration-dependent manner (P < 0.05). Adipose-derived mesenchymal stem cells can significantly suppress the proliferation of lymphocyte in vitro. The inhibition effect may relate with the increasing of CD4+CD25+ Tregs subsets, and relate with the changing of cytokine secretion of lymphocyte. Related Articles | Metrics

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In vitro isolation, cultivation and identification of ecto-mesenchymal stem cells Li Bing, Fei Zhou, Hu Shi-jie, Lin Wei, Li Xia, Hu Xue-an, Wang Bing 2012, 16 (45):  8435-8439.  doi: 10.3969/j.issn.2095-4344.2012.45.014 Abstract ( 284 )   PDF (491KB) ( 424 )   Save BACKGROUND: Ecto-mesenchymal stem cells are not only a new kind of seed cells in tissue engineering research, but also an innovative therapeutic strategy for tumors in clinical nervous system. OBJECTIVE: To observe morphology and ultrastructure, identify nerve stem cell marker and test the ability of growth and the proliferation of in vitro isolated ecto-mesenchymal stem cells. METHODS: Ecto-mesenchymal stem cells were enzymatically isolated from the first branchial arch of rat embryo and maintained in an undifferentiated state with beads coated low affinity nerve growth factor receptor antibody. Then they were purified by magnetic activated cell sorting. RESULTS AND CONCLUSION: Ecto-mesenchymal stem cells after purification by magnetic activated cell sorting grew like swirls and were fibroblast-like and long spindle-shaped. The ultrastructure of ecto-mesenchymal stem cells was characterized by mesenchymal-like cells with irregular shape, undifferentiation and high activity of proliferation. Immunohistochemical staining manifested that ecto-mesenchymal stem cells expressed human natural killer cells-1, Vimentin and S-100 but not cytokeratin. Ecto-mesenchymal stem cells grew into the exponential phase after cultured for 3 days, and reached to plateau phase after cultured for 6 days. The growth curve was “S” like and the doubling time was 40.28 hours. The evidence suggested that the cells in vitro cultured and purified by magnetic activated cell sorting were undifferentiated ecto-mesenchymal stem cells with stem cell characteristics. Related Articles | Metrics

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Transplantation of non-adherent mesenchymal stem cells improves the learning and memory functions of mice with cerebral ischemia Zhang Jing-han, Ben Xiao-ming 2012, 16 (45):  8440-8444.  doi: 10.3969/j.issn.2095-4344.2012.45.015 Abstract ( 230 )   PDF (526KB) ( 399 )   Save BACKGROUND: Non-adherent mesenchymal stem cells exhibit the capacity of self-renewal and can be induce-differentiated into adipocytes, osteoblasts and chondrocytes. Non-adherent mesenchymal stem cells can survive and partial non-adherent mesenchymal stem cells can differentiate into neural cells or glial cells after transplantation into ischemic brain regions. OBJECTIVE: To investigate the effect of non-adherent mesenchymal stem cells transplanted into ischemic brain regions on recovery of neurological function. METHODS: Total bone marrow cells from β-Gal transgenic mice were separated by the whole bone marrow method. Passage 3 non-adherent mesenchymal stem cells were collected and purified by repeated transfer method. Twenty C57BL/6L mice were established into models of occlusion of middle cerebral artery and then randomized into two groups. In the cell transplantation group, 3 μL passage 3 non-adherent mesenchymal stem cells suspension was injected into the ischemic brain region at 7 days after middle cerebral artery occlusion. At the same time, equal amount of physiological saline was injected into the control group. Since 28 days after cell transplantation, the learning and memory functions of mice were determined by 8-day Morris water maze test. RESULTS AND CONCLUSION: In the place navigation test, the escape latency and swim length of mice in the cell transplantation group were significantly shortened than those in the control group (P < 0.05). In the probe trial, the number that mice passed cross the platform in the cell transplantation group was significantly greater than that in the control group (P < 0.05). Transplantation of non-adherent mesenchymal stem cells can improve the learning and memory functions of mice with cerebral ischemia. Related Articles | Metrics

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Effects of intravenous transplantation of human umbilical blood mesenchymal stem cells on the improvement of brain function after hypoxic-ischemic brain damage in neonatal rats Yan Xiao-hua, Yu Zhen, Liu Wei, Xu Xin, Zhou Zhi-qing, Zhang Li-na 2012, 16 (45):  8445-8452.  doi: 10.3969/j.issn.2095-4344.2012.45.016 Abstract ( 369 )   PDF (603KB) ( 576 )   Save BACKGROUND: We have explored the effect and the mechanism of human umbilical blood mesenchymal stem cells transplantation in repairing neural function after hypoxic-ischemic brain damage in neonatal rats from many aspects, such as the nerve cells replacement, promoting the proliferation and differentiation of endogenous neural stem cells, protecting the neurons, promoting the synaptic reconstruction and reducing the white matter damage. OBJECTIVE: To transplant human umbilical blood mesenchymal stem cells via the vein across the blood-brain barrier and then into the brain tissue, to study the effects of intravenous transplantation of human umbilical blood mesenchymal stem cells in repairing the brain function after hypoxic-ischemic brain damage in neonatal rats. METHODS: The 7 days Sprague-Dawley neonatal rats were divided into three groups: sham-operation group, hypoxic-ischemic brain damage group and transplantation group. In the sham-operation group, the left common carotid artery was isolated without ligation; rats in the hypoxic-ischemic brain damage group were used to establish the hypoxic-ischemic brain damage model; in the transplantation group, the human umbilical blood mesenchymal stem cells were transplanted via tail vein at 8 days after hypoxic-ischemic brain damage, and the rats in the sham-operation group and hypoxic-ischemic brain damage group received intravenous injection of normal saline in the same dose. RESULTS AND CONCLUSION: Immunofluorescence observations showed that the human umbilical blood mesenchymal stem cells migrated to the hippocampus at 5 weeks after transplantation. Nissl staining showed the Nissl bodies of pyramidal cells in left hippocampal dentate gyrus area was increased significantly after transplantation, it indicated that the mesenchyamal stem cells could differentiate into neurons after transplantation. Behavioral test results showed that when compared with the sham-operation group, rats in the hypoxic-ischemic brain damage group had a reduced spontaneous alternation rate in T-maze acquisition tests, the rats took more time in finding the three arms baited with water and had an increased number of working and reference memory errors in radial maze acquisition tests (P < 0.05); at 5 weeks after umbilical blood mesenchymal stem cells transplantation, the indicators above were improved (P < 0.05). Intravenous transplantation of human umbilical blood mesenchymal stem cells can improve and enhance the long-term learning and memory and spatial discrimination ability of hypoxic-ischemic brain damage rats. Related Articles | Metrics

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Treatment of early osteonecrosis of femoral head with autologous stem cells transplantation by computer navigat Cui Da-ping, Zhao De-wei 2012, 16 (45):  8453-8459.  doi: 10.3969/j.issn.2095-4344.2012.45.017 Abstract ( 423 )   PDF (577KB) ( 531 )   Save BACKGROUND: How to use the tissue engineering technique to repair the cartilage with femoral head collapse is the main research direction in the treatment of the femoral head necrosis after collapse. OBJECTIVE: To observe the therapeutic effect of in vitro cultured autologous bone marrow mesenchymal stem cells transplantation on the treatment of early osteonecrosis of femoral head by computer navigation and arthroscopy. METHODS: 104 patients with early osteonecrosis of femoral head were collected according to the international classification of osteonecrosis of Association Research Circulation Osseous, all the patients received the core decompression under the aids of computer navigation, and the necrosis debridement was performed under arthroscopy. Among them, 53 patients received phase two transplantation of 2 mL in vitro isolated and cultured bone autologous bone marrow mesenchymal stem cells at 1×109 /L. RESULTS AND CONCLUSION: After 30 months follow-up and magnetic resonance examination, the femoral head necrosis volume of all patients were reduced significantly, and the Harris score was increased significantly (P < 0.05); the femoral head necrosis volume and Harris score of the patients with autologous bone marrow mesenchymal stem cells transplantation were improved more significantly (P < 0.05), the clinical success rate and radiographic success rate was 92% and 90%, respectively; no obvious adverse reactions were observed. Core decompression and autologous bone marrow mesenchymal stem cells in vitro culture and transplantation by computer navigation and arthroscopy is a safe and effective method for the treatment of early avascular necrosis of femoral head, and the effect is better than the core decompression. Related Articles | Metrics

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Busulfan for unrelated allogeneic hematopoietic stem cell transplantation Li Xu-dong, He Yi, Wang Dong-ning, Lu Ying, Lin Dong-jun, Xiao Ruo-zhi, Huang Ren-wei 2012, 16 (45):  8460-8464.  doi: 10.3969/j.issn.2095-4344.2012.45.018 Abstract ( 281 )   PDF (422KB) ( 508 )   Save BACKGROUND: Erratic gastrointestinal absorption as a result of oral administration of busulfan not only affects the efficacy, but also increases the risk of toxicity. There are few papers about intravenous busulfan in conditioning regimen for unrelated allogeneic hematopoietic stem cell transplantation although it is used in China for several years. OBJECTIVE: To study the efficacy and associated complications of intravenous busulfan in conditioning regimens for unrelated allogeneic hematopoietic stem cell transplantation. METHODS: Fourteen patients received intravenous busulfan + cyclophosphamide before unrelated allogeneic hematopoietic stem cell transplantation, while 18 patients received oral busulfan + cyclophosphamide before related transplantation. The therapeutic indices and the adverse events including gastrointestinal tract reaction, oral mucositis, hemorrhagic cystitis, hepatic function lesion, acute graft versus-host disease in two groups were assessed. RESULTS AND CONCLUSION: All patients had hematopoietic engraftment. The hepatic damage and oral mucositis in the intravenous busulfan group were significantly lower than those in the oral busulfan group (14% vs. 67%, 7% vs. 55%, P < 0.01). There were no significant differences in gastrointestinal tract reaction, hemorrhagic cystitis, reconstruction, acute graft-versus-host disease, and chronic-versus-host disease between two groups. The intravenous busulfan combined with cyclophosphamide in the treatment of unrelated allogeneic hematopoietic stem cell transplantation acquires satisfactory efficacy with low toxicity. Related Articles | Metrics

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Four kinds of cells transplantation for reconstructing immune function of C57BL female mice Ruan Guang-ping, Wang Jin-xiang, Yao Xiang, Deng Yong-li, Pang Rong-qing, Cai Xue-min, Wang Qiang, Ma Li-hua, Pan Xing-hua 2012, 16 (45):  8465-8470.  doi: 10.3969/j.issn.2095-4344.2012.45.019 Abstract ( 303 )   PDF (656KB) ( 426 )   Save BACKGROUND: It is reported that muscle-derived cells can rebuild the hematopoietic function, whether the other cells can rebuild the hematopoietic function also needs further observation. OBJECTIVE: To compare the effect of different cells on the recovery of hematopoietic function of the C57BL female mice. METHODS: We removed the eyeballs of green fluorescent protein transgenic male mice in order to collect the peripheral blood. Then the bloods were hemolyzed to prepare white blood cells. And then, the spleen and the liver were obtained to prepare the spleen cells and liver cells suspension. The bone marrow cells were prepared. The C57BLfemale mice were irradiated by half-lethal dose and transfused with four kinds of cells. At 18, 39 and 53 days after transplantation, green fluorescence protein-positive cells in peripheral blood were detected. At the same time, the bloods were marked with CD4-PE and CD8-PE. At 100 days after transplantation, mice were sacrificed and their bone marrows were collected to detect Y chromosome-positive rate. RESULTS AND CONCLUSION: At 18, 39 and 53 days after transplantation, green fluorescence protein-positive cells were detected in peripheral blood of C57BL female mice; the percentage of green fluorescence protein-positive cells in the peripheral blood of C57BL female mice: transfused with spleen cells and marrow cells > transfused with liver cells > transfused with peripheral white blood cells. At the same time, the CD4 and green fluorescence protein double positive cells and CD8 and green fluorescence protein double positive cells were also detected in the peripheral blood of C57BL female mice after transfused with spleen cells and marrow cells, while the double positive cells were not detected in the peripheral blood of C57BL female mice after transfused with liver cells and peripheral white blood cells. At 100 days after transplantation, the Y chromosome-positive rate in bone marrow was positively correlated with the green fluorescence protein-positive rate in peripheral blood. The white blood cells, spleen cells, liver cells and effect marrow cells in the green fluorescent protein transgenic male mice can rebuild hematopoietic function, and the effect of spleen cells is similar to bone marrow cells but stronger than liver cells. The effects of liver cells are stronger than the white blood cells. Related Articles | Metrics

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Bone marrow mesenchymal stem cells transplantation promotes the motor function recovery after spinal cord injury Liu Sheng-liang, Fan Ye-wen, Li Yan, Zhang Hong-nan, Xue Meng-meng, Chen Ming-wei, Kong Jian, Liu Xin, Cui Kai-yu, Peng Yan, Wang Duo, Liu Jing-jing, Ge Xiao-ping, Wang Yong, Wu Shu-lian 2012, 16 (45):  8471-8475.  doi: 10.3969/j.issn.2095-4344.2012.45.020 Abstract ( 250 )   PDF (495KB) ( 496 )   Save BACKGROUND: The studies have confirmed that the bone marrow mesenchymal stem cells transplantation can be induced to differentiate into neural cells, reconstruct neuronal circuits, and promote axonal regeneration so as to recover of spinal cord function. OBJECTIVE: To explore the repair effect of bone marrow mesenchymal stem cells on spinal cord injury. METHODS: Forty C57BL/6 mice were divided into four groups, mice in the sham-operation group did not combat the spinal cord, and mice in the other three groups were used to establish the spinal cord injury model by weight impact method. At 7 days after injury, bone marrow mesenchymal stem cell suspension was injected in treatment group via orbital venous plexus by microinjector; mice in the control group were injected with DEME culture medium in the same dose; mice in the model group were not treated. The extent of spinal cord injury was detected by hematoxylin-eosin staining. Nerve cells differentiated from bone marrow mesenchymal stem cells were identified by immunocytochemistry staining. The growth state of transplanted cells was observed by fluorescent microscope. Motor functional recovery was evaluated by modified Tarlov score. RESULTS AND CONCLUSION: After the bone marrow mesenchymal stem cells were induced to differentiate for 7 days, positive expression of nuclear factor and glial fibrillary acidic protein could be seen in the cells. Both lower extremities of the mice in the model group were paralyzed, and the mice in the sham-operation group had normal action (P < 0.01). At 2 weeks after transplantation, the symptoms of motor functional deficits in the treatment group was gradually restored, and the recovery in the sham-operation group was not obvious (P < 0.05); at 4 weeks after transplantation, there was no significant difference of the Tarlov score between treatment group and sham-operation group (P > 0. 05). Bone marrow mesenchymal stem cells transplantation can promote the motor functional recovery after spinal cord injury. Related Articles | Metrics

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Connexin-43 expression in cardiac tissue after adipose-derived stem cells transplantation for treatment of acute myocardial infarction Jie Ling-jun, Wang Yi-qing, Ma Qun-cao, Chen Xiao-min, Luo Hong, Zhang Cheng-Wei 2012, 16 (45):  8476-8480.  doi: 10.3969/j.issn.2095-4344.2012.45.021 Abstract ( 285 )   PDF (527KB) ( 763 )   Save BACKGROUND: The therapeutic effect of adipose-derived stem cells (ADSC) as a novel kind of seed cells on myocardial infarction in rats has been rarely reported. OBJECTIVE: To investigate the effects of ADSCs transplantation on cardiac function and connexin-43 expression in rats with acute myocardial infarction. METHODS: Rat ADSCs were isolated and cultured in vitro. Twenty-four Sprague-Dawley rats were divided into sham-operated, myocardial infarction and ADSCs groups. Rats in the former two groups were prepared into acute myocardial infarction models by ligation of the left anterior descending branch. In the sham-operated group, only chest was cut open without ligation. In the ADSCs group, at 30 minutes after myocardial infarction, ADSCs were administered via four points, 25 μL per point. Rats in the sham-operated and myocardial infarction groups received identical amounts of phosphate buffered saline. At 2 weeks after ADSCs transplantation, index examination was performed. RESULTS AND CONCLUSION: Echocardiogram showed that left ventricular ejection fraction, and left ventricular fractional shortening were significantly greater in the myocardial infarction group than in the ADSCs group (P < 0.05). Masson’s trichrome staining demonstrated that fibrinous exudate level was significantly lower in the ADSCs group than in the myocardial infarction group (P < 0.05). Immunofluorescent results showed that connexin-43 expression and vascular density in the ADSCs group were significantly increased than in the myocardial infarction group (P < 0.05-0.01). Real-time PCR showed that connexin-43 mRNA expression in the ADSCs group was significantly greater than in the myocardial infarction group (P < 0.01). ADSCs transplantation can greatly improve the cardiac function, reduce fibrinous exudates and up-regulate connexin-43 expression after myocardial infarction. Related Articles | Metrics

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5 μg/L basic fibroblast growth factors promote the proliferation of adipose-derived stem cells Chen Yang, Gao Jian-hua, Cha Peng-fei, Lu Feng 2012, 16 (45):  8481-8485.  doi: 10.3969/j.issn.2095-4344.2012.45.022 Abstract ( 270 )   PDF (568KB) ( 719 )   Save BACKGROUND: Basic fibroblast growth factors have been widely used for clinical autologous fat grafting because they exhibit great potential to promote angiogenesis and adipocyte proliferation and differentiation. OBJECTIVE: To investigate the effects of different concentrations of basic fibroblast growth factors on proliferation of adipose-derived stem cells and analyze the proper concentration of basic fibroblast growth factors for promoting proliferation of adipose-derived stem cells. METHODS: Adipose-derived stem cells were isolated from adipose tissue by liposuction. Cell morphology and growth characteristics were observed under the microscope. Culture medium containing 0, 5, 10, 20, 40, 80, 60 μg/L basic fibroblast growth factors was prepared. Adipose-derived stem cells were cultured in a 96-well plate at 103 cells/well using culture medium containing basic fibroblast growth factors. The proliferation of adipose-derived stem cells was determined by MTT for successive 7 days. RESULTS AND CONCLUSION: Basic fibroblast growth factors could promote the proliferation of adipose-derived stem cells significantly. The proliferation-promoting effect was significant since the 3rd day and peaked on the 5th and 6th days. There was no obvious difference in proliferation-promoting effects between different basic fibroblast growth factor concentrations. 5 μg/L was the most appropriate concentration for basic fibroblast growth factors in promoting the proliferation of adipose-derived stem cells. Related Articles | Metrics

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Xianlinggubao affects bone mass and osteogenic differentiation of bone marrow stromal cells in rats Xing Lei, Jiao Ying-hua, Tian Fa-ming 2012, 16 (45):  8486-8490.  doi: 10.3969/j.issn.2095-4344.2012.45.023 Abstract ( 294 )   PDF (526KB) ( 634 )   Save BACKGROUND: Bone loss in rats can be achieved by bilateral ovariectomy. Xianlinggubao as a Chinese agent has been shown to promote bone formation. OBJECTIVE: To investigate the effects of Xianlinggubao on bone mass and the expression level of genes related to osteogenic differentiation of bone marrow stromal cells in rats. METHODS: Twenty-four 3-month-old Sprague-Dawley rats were randomly divided into three groups, with eight rats in each group. Rats from the ovariectomy group and Xianlinggubao group underwent ovariectomy. At 6 weeks after ovariectomy, Xianlinggubao group rats were daily administered 250 mg/kg Xianlinggubao for a total of 8 weeks. Rats from the normal control group received no intervention. RESULTS AND CONCLUSION: The bone mineral density at L2-4 vertebral body in the Xianlinggubao group was significantly higher than that in the ovariectomy group, but it was significantly lower than that in the normal control group (both P < 0.05). Serum osteocalcin, bone morphogenetic protein 2, osteocalcin mRNA expression levels in the ovariectomy group were significantly lower than those in the Xianlinggubao group and normal control group (both P < 0.05). Extracellular matrix mineralization ability in the ovariectomy group was obviously lower than that in the Xianlinggubao group and normal control group. At 14 weeks after ovariectomy, rat bone loss was obvious, and Xianlinggubao can partly prevent bone loss. The underlying mechanism is likely to be related to Xianlinggubao promotion of osteogenic differentiation of rat bone marrow stromal cells. Related Articles | Metrics

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Construction of RNA interference lentiviral vector of ribosomal protein S14 gene and the expression in SKM-1 cells Jiang Wen, Luo Jing, Liu Lin, Xiao Qing, Yang Ze-song, Wang Li 2012, 16 (45):  8491-8495.  doi: 10.3969/j.issn.2095-4344.2012.45.024 Abstract ( 518 )   PDF (524KB) ( 624 )   Save BACKGROUND: Studies have shown that decreased expression of ribosome protein S14 (RPS14) gene in hematopoietic stem/progenitor cells can lead to the dysplasia of hematopoietic cells, in particular the blocked erythroid differentiation. Normal expression of RPS14 gene can help to maintain the normal hematopoiesis. OBJECTIVE: To construct a RNA interference lentiviral vector of RPS14 gene, and to observe its expression in the human myelodysplastic syndrome cell line SKM-1 cells. METHODS: The targeting sequence of RPS14 gene which effectively silenced in RNA inference was confirmed in our previous study. The GC-shRPS14 lentiviral vector was established and sequenced. The 293T cells were co-transfected with lentiviral vector GC-shRPS14, pHelper 1.0 and pHelper 2.0. The titer of virus was tested according to the expression level of GFP. The resulting S14shRNA lentiviral vector GC-shRPS14 was used to infect human myelodysplastic syndrome cell line SKM-1. And the RPS14 gene expression in the target cells was detected by Western Blot. RESULTS AND CONCLUSION: DAN sequencing demonstrated that the lentiviral vector GC-shRPS14 of RPS14 shRNA was constructed. The titer of concentrated virus was 2×109 Tu/ml. GFP expression could be seen in the transfected SKM-1 cells under fluorescence microscope, and the transfection efficiency was 75% Western Blot showed that. RPS14 expression was significantly inhibited in the target cells after transfection. The RNA interference lentiviral vector of RPS14 was constructed successfully, which is capable of delivering the target gene RPS14 into human myelodysplastic syndrome cell line SKM-1 for its silent expression. Related Articles | Metrics

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Cloning efficiency of human endometrial cells in vitro Ma Ying, He Yuan-li 2012, 16 (45):  8496-8500.  doi: 10.3969/j.issn.2095-4344.2012.45.025 Abstract ( 477 )   PDF (499KB) ( 611 )   Save BACKGROUND: Colony forming ability is an important characteristic of stem cells. Many studies have shown that the endometrium has monoclonal characteristics. OBJECTIVE: To detect the cloning efficiency of human endometrial cells in vitro. METHODS: Human endometrial cells were isolated and dissociated mechanically and enzymatically from human endometrium. Stromal and epithelial cells were separated by two series of filters. The cell morphology and growth characteristics were observed and the expressions of vimentin and keratin were detected. Flow cytometry was used to identify the expression of CD133, CD34, CD45, CD90, CD73 and CD29 markers in endometrial cells. The clone growth characteristics were observed, the size of clones was measured and the rate of cloning efficiency was calculated. Meanwhile, human uterine muscle cell was used as a control. RESULTS AND CONCLUSION: After primary cultured for 12 days, glandular epithelial cells and stromal cells were divided into large and small clones by size and growth characteristics. The gland clone was positive for keratin and the stromal clone was positive for vimentin by immunohistochemistry. Results from flow cytometry showed that endometrial cells were positive for CD29, CD90 and CD73 but negative for CD34, CD45 and CD133. There was significant difference of the sizes and cloning effiency of the large and little gland clones between glandular epithelial cells and stromal cells (P < 0.05). Human uterine smooth muscle cells were primary cultured by limiting dilution method. The endometrial cells can form landular epithelial cells and stromal cells in vitro which included large and small clones. The proliferation efficiency of the large gland clone was higher than that of the small gland clone. These cells may be the endometrial stem cells. Related Articles | Metrics

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Influence of various volume fractions of platelet-rich plasma on the proliferation of rat hair follicle stem cells Peng Yu, Wang Yu-jie, Li Jia, Muratrixat 2012, 16 (45):  8501-8505.  doi: 10.3969/j.issn.2095-4344.2012.45.026 Abstract ( 238 )   PDF (515KB) ( 496 )   Save BACKGROUND: Recent studies have found that the platelet-rich plasma has an important role in promoting bone regeneration, soft tissue repair and proliferation and differentiation of the seed cells in tissue engineering. OBJECTIVE: To observe the effect of different methods and different volume fractions of platelet-rich plasma on the proliferation of rat hair follicle stem cells. METHODS: Combination of microdissection and two-step enzyme digestion with differential adhesion methods were used to obtain the third generation of rat hair follicle stem cells with high purity and good growth state. The second centrifugation method and the third centrifugation method were used to obtain the rat platelet-rich plasma, and then diluted into the K-SFM medium with the volume fraction of 1.0%, 2.0% and 4.0%, while the platelet-rich plasma containing different volume fractions of medium was considered as the experimental group and the platelet-rich plasma without medium was used as a control. RESULTS AND CONCLUSION: Highly purified hair follicle stem cells could be obtained by microdissection, two-step enzyme digestion and differential adhesion methods. There was no significant difference between experimental group and control group at 2 days before culture (P > 0.05); and then hair follicle stem cells in the experimental group showed higher proliferation activity after the platelet-rich plasma intervention for 2 days. After cultured for 4 days, the hair follicle stem cells grew into the growth logarithmic phase; and after 5 days, the cells reached to 70%-80% confluence, and then went into the platform after cultured for 6-7 days. The cell proliferation in the high dose platelet-rich plasma group was better than that in the low dose platelet-rich plasma group. Under the condition of the same volume fraction, the effect of the platelet-rich plasma obtained by third centrifugation method on the proliferation of hair follicle stem cells was significantly greater than that obtained by the second centrifugation method. The platelet-rich plasma prepared by the third centrifugation method with the volume fraction of 4.0% has the most significant effect on the proliferation of hair follicle stem cells. Related Articles | Metrics

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In vitro culture and identification of spinal cord neural stem cells from rats of different ages Yu Zi-chao, Guo Kai-jin 2012, 16 (45):  8506-8509.  doi: 10.3969/j.issn.2095-4344.2012.45.027 Abstract ( 234 )   PDF (419KB) ( 476 )   Save BACKGROUND: There have been few reports describing the proliferation and differentiation characteristics of neural stem cells from rats of different ages. OBJECTIVE: To investigate the in vitro isolation, culture and identification of spinal cord neural stem cells from rats of different ages. METHODS: Spinal cord from rats of different ages was taken and prepared into single cell suspension. Primary cells at 5.0×105/L were cultured with DMEM/F12 medium containing epidermal growth factor, basic fibroblast growth and B27 and then passaged once every other day. RESULTS AND CONCLUSION: Cells could be cultured from different ages of rat spinal cord. The suspending cell spheres, named neural stem cell spheres showed compact structure and were of different sizes. After passage, single cells proliferated rapidly and form neurospheres and the number increased obviously. Neural stem cell spheres can be in vitro cultured from the spinal cord of rats at different ages. Related Articles | Metrics

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Effects of X binding protein-1 gene-modified neural stem cells transplantation on expression of related neurotransmitters and nucleoprotein in the substantia nigra of rat Parkinson's disease model Zhang Yan-hua, Lu Lei, Dong Jing, Song Lei, He Chun-ke, Zhang Yuan 2012, 16 (45):  8510-8513.  doi: 10.3969/j.issn.2095-4344.2012.45.028 Abstract ( 295 )   PDF (386KB) ( 578 )   Save BACKGROUND: Previous studies have shown that X binding protein-1 gene can be transfected into the neural stem cells and is steadily expressed. X binding protein-1 gene transfected neural stem cells transplantation for cerebral infarction treatment is better than ordinary neural stem cells. OBJECTIVE: To study the treatment effect of X binding protein-1 transfected neural stem cells transplantation on Parkinson's disease through detecting the influence on monoamine neurotransmitters and α-synuclein levels in the substantia nigra of rat Parkinson's disease model. METHODS: Twenty-seven rat Parkinson's disease models were divided into three groups randomly, and 9 rats in each group. The rats in control group were transplanted with PBS through the lateral ventricle, while neural stem cells group was transplanted with blank neural stem cell suspension, and transfection group was transplanted with X binding protein-1 gene-modified neural stem cells. In 7, 14, 21 and 28 days after transplantation, neurobehavioral score of the rats was estimated respectively, α-synuclein level was examined by Western blot, and the tyrosine hydroxylase positive cells were revealed by immunohistochemical staining. RESULTS AND CONCLUSION: The expression of α-synuclein was decreased in the transfection group. The number of tyrosine hydroxylase positive cells in the transfection group was higher than that in the neural stem cells group and control group, and the rotation speed of transfection group was decreased obviously. The results show that the differentiation rate of binding protein-1 gene-modified neural stem cells differentiating into dopaminergic neurons is higher and the transfection can reduce α-synuclein content in rat substantia nigra, so as to treat Parkinson's symptoms. Related Articles | Metrics

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Design of distributed wireless multi-point temperature measuring system in blood cold chain Peng Da-ming, Jian Jun, Yu Xue-fei 2012, 16 (45):  8514-8519.  doi: 10.3969/j.issn.2095-4344.2012.45.029 Abstract ( 326 )   PDF (437KB) ( 779 )   Save BACKGROUND: Blood and blood products are different from general drugs, they need different storage temperatures for preservation of biological activity when they are in in vitro environment. In order to ensure safe, effective and reliable quality, blood cold chain is necessary to avoid a fact that too high or low temperature influences the quality of blood or blood products. OBJECTIVE: Based on the existing problems of present domestic blood cold chain management, this paper proposed an application of distributed wireless multi-point temperature measuring system. METHODS: The system used the technology of 1-wire digital sensor, wireless communication and single-chip microcomputer in the management. It can intelligently identify the abnormal temperature fluctuation and alarm. The PC monitoring software was designed by using Visual C# 2008, which can monitor many slave systems simultaneously. RESULTS AND CONCLUSION: Repeated experiments show that the system operates stably and has good reliability. Related Articles | Metrics

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Wnt signaling pathway for osteogenic differentiation of bone marrow mesenchymal stem cells induced by eucommia bark Zhang Xian, Zhu Li-hua, Qian Xiao-wei, Tan Xiang-ling 2012, 16 (45):  8520-8523.  doi: 10.3969/j.issn.2095-4344.2012.45.030 Abstract ( 283 )   PDF (388KB) ( 644 )   Save BACKGROUND: Recently, there are many reports describing Chinese medicine and the effective components of Chinese medicine for treatment of osteoporosis. But there are few reports regarding cellular osteogenic differentiation and regulation signaling. OBJECTIVE: To investigate the expression of Wnt signaling pathway related genes during osteogenic differentiation of rat bone marrow mesenchymal stem cells induced by eucommia bark. METHODS: Passage 3 Sprague-Dawley rat bone marrow mesenchymal stem cells were inoculated into 6-well plate, with 1×103 cells per well. 24 hours later, induced medium supplemented with 7.5% fetal bovine serum-containing DMEM/F12 (1:1) and 1/1 000 eucommia bark was refreshed. Normal culture medium was used in the negative control group. After induction for 8 hours, 1, 3, and 7 days, expression levels of Fzd and LRP receptors of Wnt passway, β-catenin, intranuclear Wnt target genes and Wnt inhibitory factor were determined by RT-qPCR. RESULTS AND CONCLUSION: Compared with negative control group, Fzd2 expression increased to 11.86 folds and Fzd3 expression increased to 2 folds respectively after induction for 3 days; Fzd2 expression increased to 5.12 folds and Fzd3 rebounded to normal level after induction for 7 days; β-catenin expression increased to 2 folds after induction for 3 days, and WIF1 expression decreased significantly after induction for 3 and 7 days, respectively. These findings suggest that Wnt signaling pathway may participate in the process of eucommia bark promoting in osteogenic differentiation of bone marrow mesenchymal stem cells. Related Articles | Metrics

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Research progress in proteins regulating synaptogenesis Huang Yun, Guo Wei-tao, Wang Bin 2012, 16 (45):  8524-8529.  doi: 10.3969/j.issn.2095-4344.2012.45.031 Abstract ( 550 )   PDF (531KB) ( 580 )   Save BACKGROUND: Stem cells have been shown to differentiate into neuronal cells in vitro. But how do neuronal cells form synaptic connections after stem cell differentiation to achieve the information transfer function and what is the mechanism underlying synaptogenesis remain unclear. OBJECTIVE: To summarize the effects of proteins regulating the synaptogenesis by retrieving the experimental studies describing these proteins influencing synaptogenesis since 2000. METHODS: A computer-based online retrieval of CNKI and PubMed databases was performed to search related papers using the key words synapse and synaptogenesis. After excluding repeated studies, 39 papers were included in the final analysis. RESULTS AND CONCLUSION: The synaptogenesis can be divided into three related aspects: (1) formation of synaptic structure; (2) transition of some synapses from silent to active; (3) elimination of unnecessary synapses. Synaptogenesis associated proteins and their functions have been confirmed and preliminarily studied. These proteins include thrombospondins, synapse differentiation induced gene product, synaptic cell adhesion molecule, major histocompatibility complexⅠ, myocyte enhancer factor 2, and fragile X mental retardion protein. These proteins play a vital role in synapse formation, development and maturation. Related Articles | Metrics

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Directed differentiation of hepatic stem cells from different origins Wei Chun-shan, Tang Hai-hong, Tong Guang-dong, Chen Xiao-yin 2012, 16 (45):  8530-8534.  doi: 10.3969/j.issn.2095-4344.2012.45.032 Abstract ( 287 )   PDF (575KB) ( 695 )   Save BACKGROUND: The hepatic stem cells originated from embryo or hepatic oval cells were always studied actively in basic researches. But the other stem cells from umbilical cord blood or bone marrow are paid more attention in clinical study. There is no agreement on differentiation of varied hepatic stem cells. OBJECTIVE: To summarize the potentiality of directed differentiation of hepatic stem cells from different origins. METHODS: PubMed databases and China National Knowledge Infrastructure (CNKI) were retrieved for articles published from January 1990 to October 2011 in English and Chinese respectively with the searching words of differentiation, hepatic stem cell or liver stem cell. Articles in the same research field published in the authoritative journals or recently published (past 3 years) were included. Forty-two articles were included in the final analysis. RESULTS AND CONCLUSION: The majority of basic studies of hepatic stem cells focus on embryo-derived hepatic stem cells or hepatic oval cells; while in the clinical studies, umbilical cord- and bone marrow-derived hepatic stem cells have been paid more attention. There is no agreement on differentiation potential of hepatic stem cells from various origins. This paper summarizes the difference of differentiation potential of hepatic stem cells from different origins and investigates the decisive effect of different inductors so as to provide strong evidence for further screening reliable inductors and improving directed induced differentiation. More basic studies involving hepatic stem cells are expected to meet clinical requirements. This would help to achieve agreement in origins and inductors of hepatic stem cells and establish uniform standards to collect stem cells or uniform clinical operation guideline. Related Articles | Metrics

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Stem cells for repair of traumatic nerve injury Sun Hong, Weng Li-xin 2012, 16 (45):  8535-8542.  doi: 10.3969/j.issn.2095-4344.2012.45.033 Abstract ( 365 )   PDF (719KB) ( 495 )   Save BACKGROUND: Stem cells are a kind of cells with the capacity for self-renewal and multi-lineage differentiation and in vitro proliferation and have become an ideal material for cell transplantation. At present, stem cells and regenerative medicine have become a gradually increasing area in the field of natural science. This study analyzed the advances in application of bone marrow mesenchymal stem cells, neural stem cells, embryonic stem cells, umbilical cord blood stem cells and adipose-derived stem cells in repair of traumatic nerve injury. However, the source, the oriented induction, transplantation and neurological repair function judgment of neural stem cells have still many problems. OBJECTIVE: To investigate the advances in application of stem cells in repair of traumatic nerve injury. METHODS: A computer-based online retrieval was performed by the first author in SCI (Science Citation Index) database to search papers regarding the application of stem cells in the repair of traumatic nerve injury with the key words “stem cell, bone marrow mesenchymal stem cells, neural stem cell, embryonic stem cell, adipose-derived stem cells, umbilical cord blood stem cells, adult stem cells, traumatic brain injury, spinal cord injury, traumatic nerve injury, growth factor, repair” in both Chinese and English. The advances in application of bone marrow mesenchymal stem cells, neural stem cells, embryonic stem cells, umbilical cord blood stem cells and adipose-derived stem cells in repair of traumatic nerve injury were further analyzed. RESULTS AND CONCLUSION: Stem cells are a kind of special cells with capacity for self-renewal, strong proliferation and multi-lineage differentiation. The self-renewal and multi-lineage differentiation potential are the most significant biological characteristics. At present, stem cells have been successfully isolated from many tissues or organs including embryonic stem cells, hematopoietic stem cells and bone marrow mesenchymal stem cells. In addition, the cells most studied in recent years include neura l stem cells, muscle stem cells, osteoblasts, endodermal stem cells and retinal stem cells. The multi-lineage differentiation potential of stem cells provides a new way for repair of traumatic nerve injury and great advances have been made in neural repair and function reconstruction after brain injury and spinal cord injury. Therefore, stem cells have wide application prospect. However, there still have some problems to be further solved. Related Articles | Metrics

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Stem cells transplantation for liver regeneration Yang Yi, Chen Xi, Xu Xin, Wu Tao, Li Yi-ming 2012, 16 (45):  8543-8550.  doi: 10.3969/j.issn.2095-4344.2012.45.034 Abstract ( 333 )   PDF (790KB) ( 523 )   Save BACKGROUND: Satisfactory therapeutic effects can not been acquired in treatment of end-stage liver diseases by the routine comprehensive treatments. Therefore, stem cells transplantation for treatment of liver disease has become an interesting issue and has been considered as a new method in treatment of this disease. OBJECTIVE: To clarify the experimental research and clinical application of stem cells transplantation for liver regeneration. METHODS: In vitro and in vivo experimental studies regarding liver disease treatment by transplantation of different kinds of stem cells were analyzed, including bone marrow stem cells, embryonic stem cells, adipose stem cells and peripheral blood stem cells. The biological characteristics and functions were compared between normal liver cells and transplanted cells using immunohistological staining, pathological observation and biochemical index detection to determine the treatment effects of stem cells transplantation for the treatment of liver disease. RESULTS AND CONCLUSION: Bone marrow stem cells, embryonic stem cells, adipose stem cells and peripheral blood stem cells can be induced to differentiate into hepatocyte-like cells under certain conditions, and these cells have therapeutic effects on the injured liver and can also promote the repair and regeneration of the liver. Meanwhile, these cells can be used for transgenic therapy in stem cells transplantation. In addition, stem cells transplantation can be used for treatment of various liver diseases without immunological rejections. Related Articles | Metrics