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06 August 2015, Volume 19 Issue 32 Previous Issue    Next Issue

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Regulating osteogenic and adipogenic differentiation of bone marrow mesenchymal stem cells by extracellular adenosine triphosphate Li Wen-kai, Zhang Ying-chi, Wei Sheng, Yang Yong, Wu Hua 2015, 19 (32):  5085-5091.  doi: 10.3969/j.issn.2095-4344.2015.32.001 Abstract ( 395 )   PDF (1176KB) ( 622 )   Save BACKGROUND: Impaired balance between osteogenesis and adipogenesis of bone marrow mesenchymal stem cells is a crucial pathological mechanism of osteoporosis. Mechanical loads applied to bone tissue can increase bone formation and improve bone strength, and meanwhile lead to the release of extracellular nucleotides, such as adenosine triphosphate. OBJECTIVE: To determine the effects of adenosine triphosphate on the osteogenic and adipogenic differentiation of bone marrow mesenchymal stem cells and to investigate the underlying mechanisms. METHODS: The effect of adenosine triphosphate (10, 50, 250 μmol/L) on differentiation of bone marrow mesenchymal stem cells were measured by osteogenic and adipogenic related genes expression, alizarin red staining and oil red O staining. The activation of ERK1/2 signaling pathway by adenosine triphosphate was tested using western blot assay. RESULTS AND CONCLUSION: Incubation of bone marrow mesenchymal stem cells with adenosine triphosphate resulted in the dose-dependent increase of osteogenic genes expression and calcium deposition, and inhibition of adipogenic genes expression and lipid droplet formation, but had no effects on cell proliferation. Adenosine triphosphate activated ERK1/2 signaling pathway, and U0126 as an ERK1/2 inhibitor restrained the effect of adenosine triphosphate on the differentiation of bone marrow mesenchymal stem cells.   中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Adrenomedullin effect on expression of apoptosis-related protein in bone marrow mesenchymal stem cells under hypoxia Yao Yue-xian, Wang Jun-sheng, Su Jin-zi, Jiang Jin-feng, Cai Wen-qin 2015, 19 (32):  5092-5096.  doi: 10.3969/j.issn.2095-4344.2015.32.002 Abstract ( 275 )   PDF (872KB) ( 646 )   Save BACKGROUND: Adrenomedullin gene transfection can strength the anti-apoptotic ability of bone marrow mesenchymal stem cells under ischemia and hypoxia, but its mechanism is not yet clear. OBJECTIVE: To investigate the effect of adrenomedullin on the expression of apoptosis-related proteins, Bax, Bcl-2 and Caspase-3, in bone marrow mesenchymal stem cells under hypoxia and ischemia. METHODS: Bone marrow mesenchymal stem cells of Sprague-Dawley rats were isolated, cultured and purified, and then cultured in serum-free medium under hypoxic condition for 0, 3, 6, 9, 12 hours. Then, western blot assay was employed to detect the expression of Bax, Bcl-2 and Caspase-3 so as to determine the optimal hypoxia time that was determined at 6 hours of hypoxia. Depending on whether adrenomedullin pretreatment was done, the cells were divided into control group (with no adrenomedullin pretreatment before hypoxia and ischemia) and adrenomedullin groups with different concentrations (1, 10, 100 μg/L). Afterwards, the expression of Bax, Bcl-2 and Caspase-3 was detected by using western blot assay. RESULTS AND CONCLUSION: (1) After cultured in serum-free medium under hypoxia for 0, 3, 6, 9, 12 hours, the expression of Bax, Bcl-2 and Caspase-3 in bone marrow mesenchymal stem cells were increased (P < 0.05);  at 6 hours of hypoxia, the Bax/Bcl-2 ratio and Caspase-3 expression reached the minimum value (P < 0.05). (2) At 6 hours of hypoxia, the expression of Bax and Caspase-3 protein as well as Bax/Bcl-2 ratio became the lowest in the 100 μg/L group compared with the 1 and 10 μg/L groups, but the expression of Bcl-2 protein reached the peak (P < 0.05). These findings indicate that adrenomedullin can reduce the expression of Bax/Bcl-2 ratio and Caspase-3 protein in bone marrow mesenchymal stem cells cultured in serum-free medium under hypoxic conditions, which is in a dose-dependent manner.  中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Mechanical stretch promotes mesenchymal stem cell-osteoblast lineage migration through activation of mammalian target of rapamycin/matrix metalloproteinases signaling pathway Yang Zi-hui, Wu Bao-lei, Jia Sen, Yang Xin-jie, Shan Chun, Liu Xiao-chang, Wang Lei, Lei De-lin 2015, 19 (32):  5097-5102.  doi: 10.3969/j.issn.2095-4344.2015.32.003 Abstract ( 499 )   PDF (6478KB) ( 431 )   Save BACKGROUND: Distraction osteogenesis is one of the most important tissue engineering technologies. However, the exact signaling pathway controlling mesenchymal stem cell-osteoblast lineage (MSC-OB) migration during distraction osteogenesis has not yet been elucidated. More efforts should be paid to make a full understanding of the mechanism on MSC-OB lineage migration, which can improve the clinical efficacy of distraction osteogenesis. OBJECTIVE: To evaluate the effects of mechanical stretch on the ability of MSC-OB mobility and expression of mammalian target of rapamycin (mTOR) signaling pathway as well as matrix metalloproteinases (MMPs) in MSC-OB, and to make clear the mechanism by which controls MSC-OB migration during distraction osteogenesis. METHODS: Twelve Sprague-Dawley rats were randomized into two groups: experimental group (n=6), an in vivo  rat mandibular distraction osteogenesis model was established on the right side of rats; non-stretch group (n=6), only the mandibular resection was done but with no distraction osteogenesis. Immunohistochemical staining was used to detect phosphorylated mTOR expression in new osteotylus at 15 days after operation. In addition, an in vitro cell stretch model was made in the mandibular mesenchymal stem cells from healthy Sprague-Dawley rats under resting tension force (6%, 4 hours); no distraction was done in control group. The ability of MSC-OB mobility, the expression of mTOR, Raptor, p70S6K and MMPs were evaluated using experiment methods including immunohistochemistry staining, real-time PCR and scratch assay. RESULTS AND CONCLUSION: The expression of phosphorylated mTOR in MSC-OB was upregulated in the mandibular bone callus of the stretch group than the non-stretch group (P < 0.05). In the in vitro experiments, MSC-OB applied with mechanical stretch (6%, 4 hours) showed elevated gene expression levels of mTOR, Raptor, p70S6K, MMP-2, MMP-9 and MMP-13 compared with the control group (0%, 4 hours). Meanwhile, MSC-OB in the experiment group (6%, 4 hours) showed a greater ability of mobility, as demonstrated by a farther distance after 48 hours of observation (P < 0.05). The present study suggests that the enhancement of MSC-OB mobility correlates with increase of the gene expression of MMPs and mTOR signaling pathway. Mechanical stretch may promote MSC-OB migration through activation of mTOR/MMPs signaling pathway.   中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Osteogenic and adipogenic differentiation of bone marrow mesenchymal stem cells under directional induction  Zhao Qin-peng 2015, 19 (32):  5103-5107.  doi: 10.3969/j.issn.2095-4344.2015.32.004 Abstract ( 363 )   PDF (3830KB) ( 793 )   Save BACKGROUND: Bone marrow mesenchymal stem cells are non-hematopoietic stem cells from the bone marrow and can differentiate into chondrocytes, osteocytes and adipocytes under different induction conditions, which are the most promising seed cells for tissue engineering. OBJECTIVE: To evaluate the osteogenic and adipogenic differentiation of bone marrow mesenchymal stem cells under directional induction. METHODS: Bone marrow samples from Japanese rabbits were extracted aseptically to isolate and purify bone marrow mesenchymal stem cells using density gradient centrifugation and cell adherent method. Then, the cells were cultured in osteogenic and adipogenic induction media as experimental group. Another cells cultured in DMEM medium acted as control group. Sudan IV staining, Von Kossa staining and alkaline phosphatase activity detection were performed at different time after culture to compare the osteogenic and adipogenic rates. RESULTS AND CONCLUSION: After 7 days of adipogenic induction, small lipid droplets were visible and the cells arranged disorderly; at 21 days of induction, see lipid droplets with high refractivity were seen in the cytoplasm. Sudan IV staining showed that there were a large amount of red lipid droplets in the cloning center of bone marrow mesenchymal stem cells, but only 5% bone marrow mesenchymal stem cells differentiated into  adipocytes in the control group. At 7 days of osteogenic induction, induced cells were confluent in a flagstone shape, and there were many nodules; at 14 days of culture, the center of brown mineralized nodules were visible; at 21 days of culture, small pieces of mineralized nodules formed. The osteogenic and adipogenic rates were 40% and 20% in the experimental group, respectively, which were higher than those in the control group (5%, 5%). These findings indicate that under certain conditions, bone marrow mesenchymal stem cells can partially differentiate into fat cells part, and another part differentiate into osteocytes, suggesting there is a certain link between them, that is, more adipocytes and fewer osteoblasts, or vice versa. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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MHC antigen expression on the surface of bone marrow stromal stem cells after directional induction in vitro  Tong Yan-xiang, Feng Wei, Suyilaqimuge, Lv Hui-cheng, Wu Yi-min, Jiang Dian-ming 2015, 19 (32):  5112-5117.  doi: 10.3969/j.issn.2095-4344.2015.32.005 Abstract ( 421 )   PDF (3891KB) ( 565 )   Save BACKGROUND: Allogeneic bone marrow stromal stem cell transplantation for treatment of bone diseases is a hot topic. To seek effective methods for prevention of post-transplantation immune rejection is urgent to be solved. OBJECTIVE: To explore the expression of MHC antigen on the surface of bone marrow stromal stem cells after osteogenic induction in vitro. METHODS: Bone marrow samples were extracted from rabbits to in vitro isolate and culture bone marrow stromal stem cells. Then, the cells were cultured in IMDM medium containing bone morphogenetic protein-2. Flow cytometry was used to analyze the expression of MHC antigen on the osteoblasts differentiated from bone marrow stromal stem cells. RESULTS AND CONCLUSION: There was highly expressed MHC I antigen but no MHCII antigen on the osteoblasts differentiated from bone marrow stromal stem cells. After osteogenic induction, no immune rejection was found. These findings indicate that allogeneic or xenogeneic bone marrow stromal cell transplantation can be used in the treatment of bone defects. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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miR-155 is conductive to chondrogenic differentiation of bone marrow mesenchymal stem cells Liu Bo, Wang Xi-ming, Pan Qi, Tian Qi-jun 2015, 19 (32):  5113-5117.  doi: 10.3969/j.issn.2095-4344.2015.32.006 Abstract ( 308 )   PDF (953KB) ( 564 )   Save BACKGROUND: It is discovered recently that miRNA is a new regulator that is able to have an impact on gene expression and miRNA contributes to proliferation, differentiation and self-renewal of pluripotent stem cells. OBJECTIVE: To investigate the mechanism by which miR-155 regulates chondrogenic differentiation of bone marrow mesenchymal stem cells. METHODS: Sixty healthy Sprague-Dawley aged 12 weeks were randomized into study group and control group, with 30 in each group. Under anesthesia, rats were sacrificed to harvest bone marrow of the lower limbs. Then bone marrow mesenchymal stem cells were isolated, cultured, and transfected with miR-155 mimics in the study group and a negative control sequence in the control group. After chondrogenic induction, RT-PCR was used to detect the expressions of Sox9, Collagen II, Aggrecan and Collagen X gene, and western blot assay to detect the expression of Sox9 and Runx2 proteins. RESULTS AND CONCLUSION: Compared with the control group, the mRNA expressions of Sox9, Collagen II and Aggrecan were higher, but the mRNA expression of Collagen X was lower in the study group (P < 0.05); the protein expression of Sox9 was higher, but the protein expression of Runx2 was lower in the study group (P < 0.05). These findings indicate that miR-155 promotes the chondrogenic differentiation of bone marrow mesenchymal stem cells and moreover, it can suppress the hypertrophy trend of bone marrow mesenchymal stem cells differentiating into chondrocytes.   中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Hepatocyte proliferation and apoptosis under regulation of human telomerase reverse transcriptase gene-modified bone marrow mesenchymal stem cells Bai Dong, Zhou Zhong-xiao, Zhang Jian 2015, 19 (32):  5118-5122.  doi: 10.3969/j.issn.2095-4344.2015.32.007 Abstract ( 293 )   PDF (835KB) ( 633 )   Save BACKGROUND: Studies have shown that human telomerase reverse transcriptase gene (hTERT) transfection can significantly extend the life cycle of bone marrow mesenchymal stem cells so that the cells can continue to maintain pluripotency. OBJECTIVE: To investigate the effects of hTERT gene-modified bone marrow mesenchymal stem cells on hepatocyte proliferation and apoptosis. METHODS: Bone marrow mesenchymal stem cells from rats were isolated and cultured using direct adherent method. Then, hTERT eukaryotic expression plasmid, pCIneo-hTERT, was transferred into the cells using liposome transfection method. The hTERT-modified bone marrow mesenchymal stem cells were co-cultured with hepatocytes at 1:1 (observation group), and meanwhile, non-transfected bone marrow mesenchyam stem cells were co-cultured with hepatocytes at 1:1 (control group), and hepatocytes cultured alone served as single culture group. Effects of bone marrow mesenchymal stem cells on hepatocyte proliferation and apoptosis were observed by MTT assay and immunofluorescence staining. RESULTS AND CONCLUSION: The proliferative rate of hepatocytes was significantly higher in the observation group than the control and single culture groups (P < 0.05), and the survival rate of hepatocytes was significantly higher in the observation group than the single culture group (P < 0.05). Experimental findings suggest  hTERT-modified bone marrow mesenchymal stem cells can inhibit hepatocyte apoptosis but promote hepatocyte proliferation, so as to improve hepatocyte function. Figures and Tables | References | Related Articles | Metrics

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Macrophage migration inhibitory factor in lipopolysaccharide-induced peri-implant inflammation of bone marrow mesenchymal stem cells Xia Dong-jing, Pei Hao 2015, 19 (32):  5123-5128.  doi: 10.3969/j.issn.2095-4344.2015.32.008 Abstract ( 346 )   PDF (1048KB) ( 450 )   Save BACKGROUND: Currently, a large number of studies have confirmed that macrophage migration inhibitory factor plays an important role in a variety of biological activities, such as tumor development. In recent years, it also plays an important role in the inflammatory process, and has achieved a lot of results. However, it is unclear whether and how the macrophage migration inhibitory factor plays a role around the oral implant under oral environment. OBJECTIVE: To investigate the effect of macrophage migration inhibitory factor on the inflammation of bone marrow mesenchymal stem cells growing around the titanium implant. METHODS: First, bone marrow mesenchymal stem cells were seeded onto titanium cell culture disks to simulate the peri-implant environment in the mouth, and then, the cells were divided into four groups: control group, without any stimulation; lipopolysaccharide group, lipopolysaccharide-induced inflammation of bone marrow mesenchymal stem cells; non-specific small interfering RNA (siRNA)+lipopolysaccharide group, non-specific siRNA-transfected and lipopolysaccharide-induced cells; macrophage migration inhibitory factor siRNA+lipopolysaccharide group, cells under the stimulation of lipopolysaccharide were transfected with  macrophage migration inhibitory factor. RESULTS AND CONCLUSION: Using flow cytometry, the cells expressing over 95% CD29 and CD90 as well as less than 5% CD 45 were selected in the experiment. Cell counting kit-8 test showed that macrophage migration inhibitory factor siRNA+lipopolysaccharide had no influence on the proliferation of bone marrow mesenchymal stem cells. Lipopolysaccharide significantly stimulated the inflammatory reactions of bone marrow mesenchymal stem cells, which was 15-20 times of the control group (P < 0.01). However, compared with the lipopolysaccharide group, the levels of interleukin-1β, interluekin-6 and tumor necrosis factor-α were increased significantly after transfection with macrophage migration inhibitory factor siRNA+lipopolysaccharide stimulation (P < 0.01). These findings indicate that lipopolysaccharide can promote inflammation of bone marrow mesenchymal stem cells around the oral implant, but macrophage migration inhibitory factor siRNA can, to some extent, inhibit the occurrence of inflammation.   中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Correlation between functional status of bone marrow mesenchymal stem cells and disease progression in osteoarthritis patients  Liu Jian 2015, 19 (32):  5129-5133.  doi: 10.3969/j.issn.2095-4344.2015.32.009 Abstract ( 317 )   PDF (3772KB) ( 669 )   Save BACKGROUND: There is a certain relationship between bone marrow mesenchymal stem cells and articular cartilage, and its cellular functional status is one of the causes of osteoarthritis. OBJECTIVE: To explore the correlation between functional status of bone marrow mesenchymal stem cells and disease progression in patients with osteoarthritis. METHODS: The clinical data of 45 patients were collected, including 15 cases undergoing arthroscopic knee cruciate ligament reconstruction (control group), 15 cases of mild osteoarthritis (mild osteoarthritis group), 15 cases of severe osteoarthritis (severe osteoarthritis group). Bone marrow mesenchymal stem cells from three groups were cultured in vitro to observe the morphological changes. Proliferative and differentiation ability was detected and compared between the three groups. RESULTS AND CONCLUSION: At 24 hours of primary culture, the bone marrow mesenchymal stem cells were arranged in regular pattern and spindle-shaped in the three groups with no morphological differences. At 14 days of chondrogenic induction, toluidine blue staining was positive for bone marrow mesenchymal stem cells, but there was still no difference in the chondrogenic ability in the three groups. Cell counting kit-8 results showed that the proliferative ability of cells was strongest in the control group and weakest in the severe osteoarthritis group. These findings indicate that there is a certain correlation between the functional status of bone marrow mesenchymal stem cells and the progress of the disease. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程   Figures and Tables | References | Related Articles | Metrics

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Breviscapine combined with bone marrow mesenchymal stem cell transplantation promotes neurological recovery of rats with cerebral infarction  Zhao Wei, Liu Xue-zheng, Yang Ting-shuang 2015, 19 (32):  5134-5139.  doi: 10.3969/j.issn.2095-4344.2015.32.010 Abstract ( 419 )   PDF (4935KB) ( 681 )   Save BACKGROUND: Breviscapine treatment of cerebral infarction has curative effects, few side effects, stable long-term efficacy and few side effects, which can improve the micro-environment of damaged central nervous system after cerebral infarction. OBJECTIVE: To investigate the effects of breviscapine injection combined with bone marrow mesenchymal stem cell transplantation on neurological recovery and growth-associated protein 43 expression in rats after cerebral infarction. METHODS: Sixty Sprague-Dawley rats undergoing middle cerebral artery occlusion were randomized into cerebral infarction group, cell transplantation group and combined group. At 6 hours after modeling, 1 mL PBS, 1 mL bone marrow mesenchymal stem cell suspension (2.5×106), and 1 mL bone marrow mesenchymal stem cell suspension (2.5×106)+75 mg/kg breviscapine injection were respectively injected via the tail vein in the three groups, once a day, continuously for 5 days. RESULTS AND CONCLUSION: At 2 weeks after transplantation, BrdU-positive bone marrow mesenchymal stem cells were mainly gathered in the peri-infarction region, and the number of positive cells was higher in the combined group than the other two groups (P < 0.01). At 1, 2, 3 weeks after transplantation, the neurological deficit scores were significantly lower in the combined group than the other two groups (P < 0.05). At 2 weeks  after transplantation, the combined group had smaller infarct size, milder edema, and higher expression of growth-associated protein 43 as compared with the other two groups (P < 0.05). Under light microscope, glial cells proliferated dramatically and brain edema significantly reduced in the combined group. These findings indicate that breviscapine injection combined with bone marrow mesenchymal stem cell transplantation can significantly reduce infarct size and brain edema, promote neurological recovery and increase the expression of growth-associated protein 43 in rats with cerebral infarction. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Osteogenic differentiation of adipose-derived stem cells transfected by lentivirus vector carrying LIM mineralization protein-1 and hypoxia-inducible factor-1alpha Pan Wei-min, Liu Min, Yang Jian-chang, Duan Chun-guang, Huang Yue 2015, 19 (32):  5140-5147.  doi: 10.3969/j.issn.2095-4344.2015.32.011 Abstract ( 317 )   PDF (6925KB) ( 354 )   Save BACKGROUND: LIM mineralization protein-1 (LMP-1) and hypoxia-inducible factor-1α (HIF-1α) as intracellular proteins can induce osteogenic differentiation and promote angiogenesis, respectively. Therefore, their combination is of great significance for effectively inducing the osteogenic differentiation of adipose-derived stem cells. OBJECTIVE: To study the osteogenic differentiation of adipose-derived stem cells transfected by lentivirus vector carrying LMP-1 and HIF-1α.  METHODS: Reverse transcription-PCR technology was employed to clone LMP-1 and HIF-1α genes, and the genes were cloned to lentivirus vectors pLVX-EF1α-DsRed-Hyg and pLVX-EF1α-IRES2-AcGFP1 to construct main lentiviral vectors pLVX-EF1α-DsRed-Hyg-RLMP-1 and pLVX-EF1α-IRES2-AcGFP1-RHIF-1α. Then, Lenti-X 293T cells were transfected with main vectors pLVX-EF1α-DsRed-Hyg-RLMP-1 and pLVX-EF1α-IRES2-AcGFP1-RHIF-1α, packaging plasmid and coated plasmid. After that, lentiviral vectors were packaged to transfect adipose-derived stem cells from rats that were obtained by tissue explants culture and enzyme digestion methods. At 3, 7, 14 days after transfection, reverse transcription-PCR technology was adopted to detect the expression of osteogeic genes, such as bone morphogenetic protein 2, Runx-2, alkaline phosphatase, osteocalcin as well as to detect the expression of vascular endothelial growth factor. RESULTS AND CONCLUSION: Lentiviral vectors pLVX-EF1α-DsRed-Hyg-RLMP-1 and pLVX-EF1α-IRES2-AcGFP1-RHIF-1α were effectively transfected into adipose-derived stem cells. Reverse transcription-PCR results showed that from the 7th day to the 14th day after lentivirus transfection, bone morphogenetic protein 2, Runx-2, alkaline phosphatase and osteocalcin all over-expressed. These findings indicate that the combination of LMP-1 and HIF-1α can enhance the osteogenic activity of adipose-derived stem cells. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Wnt signaling promotes the differentiation of adipose mesenchymal stem cells into type II alveolar epithelial cells Shi Li, Jie Jing, Wang Fang, Zhao Dan, Zhang Xiu-fang, Peng Li-ping 2015, 19 (32):  5148-5154.  doi: 10.3969/j.issn.2095-4344.2015.32.012 Abstract ( 342 )   PDF (1665KB) ( 646 )   Save BACKGROUND: Ability of adipose mesenchymal stem cells differentiating into type II alveolar epithelial cells and the regulating mechanism have not been fully elucidated. OBJECTIVE: To study the ability of adipose mesenchymal stem cells differentiating into type II alveolar epithelial cells in vitro and the function of Wnt pathway in the regulation of differentiation. METHODS: Adipose mesenchymal stem cells were obtained from fat tissue of rats and identified by flow cytometry. The adipose mesenchymal stem cells were divided into control group, small airway growth medium (SAGM) group and Wnt3a group. Control group was treated with normal DMEM medium; SAGM and Wnt3a groups were both treated with small airway growth medium, and additionally, the Wnt3a group was treated with Wnt3a, a Wnt signaling pathway agonist. After 10 days, quantitative RT-PCR and immunofluorescence detection were used to test the expression of surfactant proteins B, C, D, type II alveolar epithelial markers. Phosphorylated β-catenin and GSK-3β were detected using western blot after 5 and 10 days of induction. RESULTS AND CONCLUSION: Adipose mesenchymal stem cells with high purity could be successfully isolated from the adipose tissue of rats, and expressed CD44 and CD29, but not CD11b and CD45. After SAGM treatment, protein and mRNA expressions of surfactant proteins B, C, D were all increased in adipose mesenchymal stem cells (P < 0.01), indicating the ability of adipose mesenchymal stem cells to be induced into type II epithelial cells. Surfactant proteins B, C, D expressions at protein and mRNA levels were significantly higher in the Wnt3a group than the SAGM group (P < 0.01). During the induction progress, the expression of phosphorylated β-catenin gradually increased, but GSK-3β expression gradually decreased in the Wnt3a group (P < 0.01). These findings indicate that Wnt signaling pathways are involved in differentiation of adipose mesenchymal stem cells into type II alveolar epithelial cells. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程   Figures and Tables | References | Related Articles | Metrics

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Human subcutaneous adipose-derived stem cells: osteoblastic/adipogenic differentiation and identification   Xiao Jian-hong, Zhang Yang-chun, Zhang Chang-ran, Yang Xing 2015, 19 (32):  5155-5161.  doi: 10.3969/j.issn.2095-4344.2015.32.013 Abstract ( 713 )   PDF (1391KB) ( 521 )   Save BACKGROUND: Adipose-derived stem cells are a kind of mesenchyam stem cells with multipotent differentiation capacity, which have more advantages than bone marrow mesenchymal stem cells in tissue engineering research. OBJECTIVE: To establish a method to isolate and purify adipose-derived stem cells from human subcutaneous adipose tissues followed by in vitro amplification and osteoblastic/adipogenic differentiation. METHODS: Adipose-derived stem cells were isolated from human subcutaneous adipose tissue and cultured by density gradient centrifugation and adherent culture. Cell morphology and growth features were observed under inverted microscope. Adipose-derived stem cells at passages 2 and 5 were selected for viability measurement using cell counting kit-8 method, and then cell growth curves were drawn. The immunophenotype identification was analyzed by flow cytometry. Passage 5 cells underwent osteoblastic/adipogenic induction to confirm the multi-differentiation potential. RESULTS AND CONCLUSION: (1) Using density gradient centrifugation and adherent culture method, high-purity human adipose-derived stem cells can be successfully isolated from human adipose tissues. (2) The growth process of human adipose-derived stem cells includes stagnant phase, logarithmic phase and plateau phase, which meets the growth rhythm of normal cells. Moreover, the population doubling time is shorter. (3).Human adipose-derived stem cells are positive for stem cell-related antigens, with low immunogenicity and the multi-differentiation potential. (4) Labeling human adipose-derived stem cells with DAPI is a simple efficient labeled method, and the labeling rate is high but the cytotoxicity is low. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Allogeneic adipose-derived stem cells combined with nano collagen-based bone for repair of ulna bone defects  Shen Ling, Wang Xi-you, Chen Ping, Yu Tong 2015, 19 (32):  5162-5166.  doi: 10.3969/j.issn.2095-4344.2015.32.014 Abstract ( 270 )   PDF (1019KB) ( 475 )   Save BACKGROUND: With the continuous development of medical model, high-level treatment and rehabilitation for bone defects are required. Therefore, bone tissue engineering technique based on stem cell culture has become a more important component of bone defect repair. OBJECTIVE: To explore and analyze the feasibility of allogeneic adipose-derived stem cells combined with nano collagen-based bone for repair of ulna bone defects. METHODS: Seventy adult female New Zealand rabbits were selected, 10 of which were used to isolate and culture adipose-derived stem cells, another 60 were randomized into scaffold group, scaffold+bone autograft group, scaffold+allogeneic adipose-derived stem cells group, with 20 rabbits in each group. The middle part of the ulna was resected to construct 12-mm bone defect models in the three groups, and then, corresponding materials were implanted. At 6, 9 and 12 weeks after implantation, bone mineral density was detected; at 14 weeks after implantation, the rates of solid fusion rate, non-solid fusion, and false joint were observed. RESULTS AND CONCLUSION: Compared with the other two groups, the rate of solid fusion was significantly higher and the rates of non-solid fusion and false joint were lower in the scaffold+allogeneic adipose-derived stem  cells group (P < 0.05). Compared with the scaffold group, the rate of solid fusion was higher and the rates of non-solid fusion and false joint were significantly lower in the scaffold+bone autograft group (P < 0.05). The bone mineral density increased dramatically at 6, 9, 12 weeks after implantation of scaffold+allogeneic adipose-derived stem cells, which was significantly higher than that in the scaffold group and scaffold+bone autograft group (P < 0.05). Compared with the scaffold group, the bone mineral density in the scaffold+bone autograft group was significantly higher (P < 0.05). These findings indicate that allogeneic adipose-derived stem cells combined with nano collagen-based bone has exact fusion effects with a higher bone mineral density.  中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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EdU-labeled human umbilical cord mesenchymal stem cells in vitro: 5 and 10 µmol/L are the optimal concentrations of EdU  Wang Wu, Zhang Fei, Li Gui-cai, Qi Bao-jun, Wu Zhong-yan, Cao Peng 2015, 19 (32):  5167-5171.  doi: 10.3969/j.issn.2095-4344.2015.32.015 Abstract ( 336 )   PDF (1969KB) ( 681 )   Save BACKGROUND: EdU is a new nuclear marker, and currently, it is rarely reported. OBJECTIVE: To determine the optimal concentration of EdU to label human umbilical cord mesenchymal stem cells. METHODS: Human umbilical cord mesenchymal stem cells were isolated, purified and subcultured. Cell morphology and growth were observed under inverted microscope. Flow cytometry was used to identify cell surface markers, as well as adipogenic identification. EdU at concentrations of 5, 10, 20, 50, 100 µmol/L was used to label human umbilical cord mesenchymal stem cells for 24 hours. The optimal concentration that resulted in the highest labeling efficiency was selected, and then cell proliferation curve was drawn. RESULTS AND CONCLUSION: Under the inverted microscope, cells grew adherently in a long spindle shape, and EdU-labeled cells had the same morphology. Flow cytometry showed that cells were positive for CD44, and had adipogenic differentiation ability. When the concentration of EdU was 5 and 10 µmol/L, the labeling efficiency was the highest, indicating that 5 and 10 µmol/L are the optimal concentrations of EdU to label human umbilical cord mesenchymal stem cells.  中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Changes of lung cancer stem cells in patients with lung squamous carcinoma during primary culture Liu Zhe-liang, Wu Jiao, Wang Lin-xian, Chen Yue-jun, Wu Guan-yu, Xiao Gao-ming 2015, 19 (32):  5172-5176.  doi: 10.3969/j.issn.2095-4344.2015.32.016 Abstract ( 310 )   PDF (1296KB) ( 690 )   Save BACKGROUND: Studies have shown that lung cancer stem cells can be isolated from the lung cancer cell lines, But there are few reports on in vitro isolation, culture and identification of lung cancer stem cells in patients with lung squamous carcinoma. OBJECTIVE: To establish the feasible methods of harvesting lung cancer stem cells from fresh lung cancer tissues in patients with lung squamous carcinoma, and to investigate the alterations in cell number and function during primary culture.  METHODS: Side population cells were isolated by collagenase digestion, Ficoll density gradient centrifugation and Hoechst 33342 efflux properties. The isolated cells were isolated and cultured in conditioned medium. Flow cytometry method was used to detect lung cancer stem cells based on the cell surface markers CD133 and CD44, and the positive rates of CD133+, CD44+ and CD133+/CD44+ were recorded. The single cell clones assay, flat colony formation assay and the cell sphere formation assay were used to identify the stem-like characteristics of lung cancer stem cells between the first and fourth generations. RESULTS AND CONCLUSION: The positive rates of CD133+, CD44+ and CD133+/CD44+ cells at the fourth generation were increased significantly, and the positive rates of CD133+ and CD133+/CD44+ cells at passage 4 were significantly higher than those at the first generation. The abilities of single cell clone formation, the flat colony formation and the cell sphere formation in the fourth-generation cells were greatly enhanced compared with the first-generation cells. Experimental findings showed that stem cell-like lung cancer cells were obtained from fresh lung cancer tissue in patients with lung squamous carcinoma, which stably and rapidly amplified in vitro, laying the foundation for the further study of the heterogeneity and drug resistance of lung cancer stem cells. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Optimal induction conditions for adipose-derived stem cells differentiating into lymphatic endothelial cells Chen Xiao-hu, Chen Tian-he, Xu Xue-zhan, Guo Qiang 2015, 19 (32):  5177-5181.  doi: 10.3969/j.issn.2095-4344.2015.32.017 Abstract ( 318 )   PDF (5090KB) ( 611 )   Save BACKGROUND: Our previous studies have shown that adipose-derived stem cells under vascular endothelial growth factor C (VEGF-C) can be induced to differentiate into lymphatic endothelial cells that are confirmed by lymphatic vascular endothelial hyaluronan receptor-1 staining. However, its optimal induction program is not clear. OBJECTIVE: To investigate the best condition for the differentiation of adipose-derived stem cells into lymphatic endothelial cells under induction of VEGF-C156s. METHODS: Adipose tissues from healthy adults were collected to isolate adipose-derived mesenchymal stem cells using trypsin digestion method. Flow cytometry was employed to detect cell surface markers, and in vitro differentiation capacity was identified by adipogenic and osteogenic induction. Passage 3 cells at good growth state were selected and divided into six groups: cells in control group were cultured in low-glucose DMEM, and those in the rest five groups were treated with 25, 50, 100, 200, 300 μg/L VEGF-C156s, respectively. RESULTS AND CONCLUSION: Adipose-derived stem cells were successfully obtained by trypsin digestion and purification, and then differentiated into lymphatic endothelial cells under the induction of VEGF-C156s, basic fibroblast growth factor and other growth factors. No cells were positive for lymphatic vascular endothelial  hyaluronan receptor-1 in the control group. After 8 days of induction, few cells were positive in the 25 μg/L VEGF-C156s group; a great amount of positive cells were visible in the 50 and 100 μg/L VEGF-C156s groups; 200 and 300 μg/L VEGF-C156s resulted in a large number of deaths in the cells. These findings indicate that it is optimal for adipose-derived stem cells to differentiate into lymphatic endothelial cells under 8-day induction of 50 μg/L VEGF-C156s. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Adipose-derived stem cells: isolation, culture and differentiation into endothelial progenitor cells   Liu Zi-qi, Sun Tong-wen, Wan You-dong, Yao Rui, Kan Quan-cheng 2015, 19 (32):  5182-5187.  doi: 10.3969/j.issn.2095-4344.2015.32.018 Abstract ( 281 )   PDF (2191KB) ( 803 )   Save BACKGROUND: Adipose-derived stem cells are regarded as the potential seed cells for tissue engineering. Collagenase digestion is used to isolate adipose-derived stem cells from fat pads currently. However, there are some problems, such as cumbersome operation and high cost. OBJECTIVE: To study the basic biological characteristics of adipose-derived stem cells by tissue explants culture and to explore the differentiation potential into osteoblasts, adipocytes and endothelial progenitor cells in vitro. METHODS: Adipose-derived stem cells were isolated by tissue explants technique from the bilateral groin fat pads of rats under aseptic conditions, and cultured in vitro. Cell counting kit-8 was used to detect the proliferative activity, and flow cytometry was employed to analyze the expression of cell surface markers. Passage 4 adipose-derived stem cells were cultured in osteogenic medium, adipogenic medium and endothelial progenitor cell medium for 2-3 weeks, and then the cells were identified. RESULTS AND CONCLUSION: Adipose-derived stem cells that were isolated by tissue explants culture were easily cultured, and after subculture, cells were mainly spindle-shaped and grew in clone-like manner with swirling arrangement. Cells that experienced repeated subcultures still kept stronger proliferative ability and the cell growth curve was shaped as a parabola. Immunochemical staining analysis revealed that adipose-derived  stem cells were positive for CD44, CD90 and CD29, but negative for CD31, CD45. After adipogenic/osteogenic induction, the cells were respectively positive for oil red O staining and alizarin red staining. Induced endothelial progenitor cells were identified with CD34 and the ability to uptake Dil-ac-LDL and FITC-UEA. These findings indicate using the using tissue explants culture, high-purity adipose-derived stem cells easy to proliferate can be harvested, highly express stem cells-related antigens, and have the ability to differentiate into osteoblasts, adipocytes and endothelial progenitor cells, which meet the needs of seed cells in tissue engineering research. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Stomach cancer stem cells: isolation, identification and characteristics Li Yang, Zhao Yong-fu 2015, 19 (32):  5188-5191.  doi: 10.3969/j.issn.2095-4344.2015.32.019 Abstract ( 576 )   PDF (4088KB) ( 727 )   Save BACKGROUND: With the in-depth study of cancer stem cells, increasing evidence has shown that cancer stem cells are the reason for cancer metastasis and recurrence, and therefore it is of great significance to isolate and identify cancer stem cells that can elucidate the pathogenesis of cancer and development of anticancer drugs. OBJECTIVE: To isolate and culture gastric cancer stem cells and then to detect its biological characteristics. METHODS: Cancer tissues from 16 patients with gastric cancer were cultured using adherent method and enzymic digestion methods to isolate gastric cancer stem cells. The cell morphology was observed under inverted microscope to drawn out growth curves. Osteogenic and adipogenic ability of cells were also detected. RESULTS AND CONCLUSION: Gastric cancer stem cells were isolated successfully by both two methods. Under the microscope, the cells were fusiform- or polygon-shaped. At confluence, the cells grew in a vortex or radial manner. The cell growth curve showed that 1-3 days were latent period, 4-9 days were logarithmic growth phase, and over 10 days were growth plateau phase. Flow cytometry results showed that passage 3 gastric cancer stem cells highly expressed CD90, CD29, CD44, but lowly expressed CD34, CD45 and HLA-DR. After osteogenic induction, calcified nodules were clearly visible in the gastric cancer stem cells at passage 3; after adipogenic induction, bright and tiny fat droplets were seen in the cytoplasm. These findings indicate that cancer stem cells exist in the gastric cancer tissues, and have similar morphology, biological property and multipotent differentiation ability to normal cells, which are probably involved in gastric cancer occurrence and development. Figures and Tables | References | Related Articles | Metrics

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Biological characteristics of CD90+ tumor stem cells in ovarian cancer cells Jiang Xiao-mang, Zhao Na, Long Min 2015, 19 (32):  5193-5198.  doi: 10.3969/j.issn.2095-4344.2015.32.020 Abstract ( 337 )   PDF (1115KB) ( 578 )   Save BACKGROUND: There is a close connection between the occurrence and development of tumor stem cells and ovarian cancer. CD90+ is an important tumor stem cell marker. OBJECTIVE: To explore the biological characteristics of CD90+ tumor stem cells in ovarian cancer cells. METHODS: The CD133 and CD90 positive rate of SKOV3 and primary ovarian cancer cells were detected by flow cytometry. The CD90+ and CD90- relative expression in stem cells and epithelium was detected by RT-PCR. Transwell invasion assay was employed to observe the cell invasion ability, clone formation test was done to observe cell proliferation and differentiation capacity, suspension ball test was adopted to observe pluripotent stem cells. The tumor formation time and tumor formation rate were observed by limited tumor dilution in immunodeficient mice. RESULTS AND CONCLUSION: The positive rates of CD133 and CD90 in SKOV3 were significantly lower than those in primary ovarian cancer cells. The expression of CD133 and OCT4 in CD90+ cells of SKOVS was significantly higher than that in CD90- cells of SKOVS. The expression of CD44, CD133, acetaldehyde dehydrogenase-1 and OCT4 in CD90+ stem cells of primary ovarian cancer cells was significantly higher than that in CD90- stem cells of primary ovarian cancer cells. There were significant differences in the  epithelial-mesenchymal related gene expressions between CD90- and CD90+ stem cells of SKOV3 and primary ovarian cancer cells. With the increase of inoculated cells, the tumor formation rate of CD90- and CD90+ cells was increased continuously, but the tumor formation time was decreased. The tumor rate of CD90- cells was lower than that of CD90+ cells. The number of transmembrane cells, cell clones and suspended cell balls was significantly higher in the CD90+ stem cells than the CD90- stem cells. These findings indicate that in ovarian cancer cells, CD90+ stem cells can highly express stem cell-related genes and epithelial-mesenchymal related genes, which have a higher invasion, proliferation and differentiation ability, as well as tumorigenic and pluripotent ability. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Neurologic function and expression of angiopoietin-1 and its receptor at recovery stage of cerebral hemorrhage after neural stem cell transplantation in rats Ren Hong-yu, Li Ming-xuan, He Cheng, Fan Yong-li, Ma Jian-gong, Si Hao-tian, Fang Shu-min, 2015, 19 (32):  5199-5203.  doi: 10.3969/j.issn.2095-4344.2015.32.021 Abstract ( 351 )   PDF (1012KB) ( 825 )   Save BACKGROUND: Recent studies have found that bone marrow mesenchymal stem cells that cultured in vitro for a long time can naturally differentiate into neural stem cells, which then differentiate into neurons and glial cells, thereby providing a new therapeutic thinking for Parkinson’s disease, sequela of cerebral infarction, cerebellar atrophy and brain dysplasia. OBJECTIVE: To discuss the influence of neural stem cell transplantation on neurologic function of rats with cerebral hemorrhage at recovery stage and the relevant mechanism of action. METHODS: Sixty male Sprague-Dawley rats were randomly divided into normal group (n=18), cerebral hemorrhage group (n=21) and transplantation group (n=21). Cerebral hemorrhage models were established in the latter two groups using VII type collagen enzyme induction method. At 21 days of modeling, rats in the transplantation group were injected neural stem cells via the tail vein, and those in the other two groups received the same volume of normal saline. At 7, 14, 21 days after cell transplantation, modified adhesive removal test (MST) was employed to evaluate the neurologic function of rats, and then the rats were killed. RT-PCR was used to detect angiopoietin-1 mRNA expression in the bleeding tissues, and western blot assay was employed to measure tyrosine kinase receptor-2 protein expression.  RESULTS AND CONCLUSION: Compared with the normal group, the MST scores in the cerebral hemorrhage group and transplantation group were significantly decreased (P < 0.05). From the 7th day after transplantation, MST scores in the transplantation group were significantly higher than those in the cerebral hemorrhage group (P < 0.05). At 7, 14, 21 days after transplantation, expressions of angiopoietin-1 mRNA and tyrosine kinase receptor-2 protein were ranked as follows: transplantation group > cerebral hemorrhage group > normal group, and there was a significant difference among the three groups (P < 0.05). These findings indicate that neural stem cell transplantation can effectively promote the neurologic recovery of rats with cerebral hemorrhage at recovery stage, and the concrete mechanism may be related to the increase of angiopoietin-1 mRNA and tyrosine kinase receptor-2 protein in the bleeding tissues.    中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Calcium alginate/bone marrow mesenchymal stem cells combined with degradation membrane for repair of skin defects Wang Nan, Sun Pei-jie, Su Qiu-xiang, Fu Ling, Zhou Zhe, Li Na-ran, Yang Xiao-xia 2015, 19 (32):  5204-5209.  doi: 10.3969/j.issn.2095-4344.2015.32.022 Abstract ( 311 )   PDF (2581KB) ( 604 )   Save BACKGROUND: Skin transplantation is regarded as the most effective therapy for large-area skin defects, which is limited by donor sources and immune rejection. Therefore, it is extremely accelerate the construction of the dermis in skin tissue engineering. OBJECTIVE: To investigate the effects of bone marrow mesenchymal stem cells/calcium alginate gel, basic fibroblast growth factor and degradation membrane on the repair of full-thickness skin defects. METHODS: Bone marrow mesenchymal stem cells were isolated from 15 New Zealand rabbits, and then cultured, amplified, subcultured and purified. Three full-thickness skin defects were made on the back of every rabbit, and randomly treated with bone marrow mesenchymal stem cells/calcium alginate gel, basic fibroblast growth factor and degradation membrane as experimental group, bone marrow mesenchymal stem cells/calcium alginate gel as control 1 group, and calcium alginate gel as control 2 group. The wounds were all covered with amniotic membrane. After 7, 14, 21 days, new wound tissues were taken for hematoxylin-eosin staining, immunohistochemistry staining and image analysis. RESULTS AND CONCLUSION: Dermis tissues in the experimental group were obviously thicker than those in control 1 and control 2 groups; there were more fibroblasts, vessels and collagen fibers in the experimental group. Especially at 14 and 21 days after operation, epidermal hyperplasia was faster with a larger coverage area in the experimental group, and at 21 days, the new epidermal tissues mainly exhibited multi-layered structure, which was superior to the control 1 and 2 groups. It follows that the combination of bone marrow mesenchymal stem cells/calcium alginate gel, basic fibroblast growth factor and degradation membrane for skin defects can accelerate the repair and regeneration of the dermis, and thus promote the epidermis regeneration and reconstruction.   中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Bone marrow mesenchymal stem cells combined with theaflavin for steroid-induced avascular necrosis of femoral head Zhao Chuan-yong, Ding Yan-fang, Zhang Wen-zhi, Yao Yu-long 2015, 19 (32):  5210-5214.  doi: 10.3969/j.issn.2095-4344.2015.32.023 Abstract ( 314 )   PDF (957KB) ( 553 )   Save BACKGROUND: Pathological mechanism of femoral head necrosis is not fully known, and high-dose corticosteroids are prone to induce femoral head necrosis. OBJECTIVE: To investigate the clinical effect of bone marrow mesenchymal stem cells combined with theaflavin on steroid-induced avascular necrosis of femoral head in rats. METHODS: Rat bone marrow mesenchymal stem cells were isolated and cultured in vitro and compounded onto gelatin sponge, and then, divided into four groups: model group, decompression group, cell transplantation group and combined group. Models of steroid-induced avascular necrosis of femoral head were established using liquid nitrogen method. In the combined group, gelatin sponge carrying bone marrow mesenchymal stem cells was transplanted and 250 mg theaflavin was given daily after modeling. RESULTS AND CONCLUSION: At 4 weeks after modeling, the femoral head samples in the four groups were round with exfoliation of articular cartilage that was pale in color. Then, the exfoliation of articular cartilage was aggravated in the model group, and some specimens appeared to collapse; the femoral head specimens in the combined group were round and pale. Until the 8th week after modeling, bone necrosis was increased in the model group; osteoblasts were found in the decompression group with fibrous callus formation; a few of empty bone lacunae appeared with irregular medullary cavity in the cell transplantation group; a large amount of new bone tissues formed in the combined group, and intramedullary adiocytes were in regular array. At 4 and 8 weeks after modeling, the number of empty bone lacunae was significantly lower in the combined group than the other groups (P < 0.05). These findings indicate that bone marrow mesenchymal stem cells combined with theaflavin is an ideal therapy for steroid-induced avascular necrosis of femoral head. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Application of bone marrow mesenchymal stem cells in hematopoietic stem cell transplantation for sensitized and non-sensitized BALB/c mice Guo Xiao-wei, Guan Chen-xia 2015, 19 (32):  5215-5219.  doi: 10.3969/j.issn.2095-4344.2015.32.024 Abstract ( 321 )   PDF (764KB) ( 443 )   Save BACKGROUND: Hematopoietic stem cell transplantation has therapeutic effects on many diseases, but its application has some limitations, such as cell harvesting and age-limited number of cells. OBJECTIVE: To investigate the application value of bone marrow mesenchymal stem cells in hematopoietic stem cell transplantation for sensitized and non-sensitized BALB/c mice. METHODS: Bone marrow cells derived from BALB/c mice were isolated and cultured in vitro to harvest mesenchymal stem cells using adherent method. The cell surface markers were detected by flow cytometry. A murine model of sensitization was established by transfusion of allogeneic spleen cells. Mesenchymal stem cells labeled with green fluorescent dye were transplanted into non-sensitized and sensitized recipient mice, and the homing of mesenchymal stem cells in vivo was monitored at different time points post transplantation. Additionally, under irradiation pretreatment, sensitized BALB/c mice under irradiation were subjected to combined transplantation of allogeneic bone marrow cells and syngeneic mesenchymal stem cells. Survival rate of BALB/c mice was monitored daily. RESULTS AND CONCLUSION: At 48 hours after transplantation, mesenchymal stem cells in sensitized and non-sensitized recipients were homing to the spleen and bone marrow, respectively. In the experiment of hematopoietic stem cell transplantation, the sensitized recipients died at 12-15 days after combined  transplantation, with a median of 14 days; however, the sensitized recipients only undergoing allogeneic bone marrow cell transplantation had a survival median of 13 days. These findings indicate that the transplanted mesenchymal stem cells in sensitized recipients are mainly homing to the spleen and bone marrow, but the combination transplantation cannot enhance the transplantation of allogeneic hematopoietic stem/progenitor cells in sensitized recipients.  中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Rapid production of tissue-engineered bone based on two kinds of adult stem cells in bone fusion  Yin Jian, Yang Yi, Yang Xiao-feng, Xiao Wei, Chai Li-ming, Yuan Hong, Jin Ge-le 2015, 19 (32):  5220-5224.  doi: 10.3969/j.issn.2095-4344.2015.32.025 Abstract ( 265 )   PDF (1099KB) ( 432 )   Save BACKGROUND: Despite tissue repair with a kind of stem cells has achieved great outcomes, but the combination of two or more kinds of stem cells to construct tissue-engineered bone is still rare. OBJECTIVE: To evaluate the efficacy of tissue-engineered bone constructed based on combining adipose-derived stem cells with bone marrow mesenchymal stem cells and allogenic bone in rabbit posterior intertransverse lumbar interbody fusion models. METHODS: Seventy-five New Zealand rabbits were randomly divided into five groups. L5,6 interbody fusion models were made in each group. Then, these five groups were respectively treated as follows (1) group A: bone marrow mesenchymal stem cells/allograft iliac bone with adipose-derived stem cells; (2) group B: bone marrow mesenchymal stem cells/allograft iliac bone with normal saline; (3) group C: allograft iliac bone with adipose-derived stem cells; (4) group D: allograft iliac bone with normal saline; group E: autologous iliac bone alone. Each group was detected by PET/CT to contrast the standard uptake value at 1, 3, 5 weeks postoperatively. RESULTS AND CONCLUSION: PET/CT images showed varying degrees of bone fusion and bone metabolism enhancement in the five groups. With time, the standard uptake value was increased in each group, and except the group D, the standard uptake value was higher in all the groups at 3, 5 weeks than at 1 week (P < 0.05). There was no difference in the standard uptake value of all the five groups at weeks 3 and 5 after operation (P > 0.05). The standard uptake value of the group A was better than that in the group B and group E (P < 0.05). Taken together, the combination of adipose-derived stem cells and bone marrow mesenchymal stem cells for rapid production of tissue-engineered bone has good osteogenesis and vascularization.   中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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In vitro labeled neural stem cells of fetal rats: MRI observation Zheng Zhao-feng, Wang Rong-fang, Wang Qi 2015, 19 (32):  5225-5230.  doi: 10.3969/j.issn.2095-4344.2015.32.026 Abstract ( 297 )   PDF (1522KB) ( 683 )   Save BACKGROUND: It is necessary to dynamically monitor the survival, recognition and migration of neural stem cells after implantation. OBJECTIVE: To in vitro label fetal rat neural stem cells using MRI technology so as to provide applied evidence of neural stem cells in nervous system repair. METHODS: Fetal rat neural stem cells were isolated, cultured and labeled followed by identification and cell viability detection. A rat model of cerebral ischemia-reperfusion injury was established. Fetal neural stem cells labeled by superparamagnetic iron oxide particles in vitro were transplanted into the left brain of model rats, and unlabeled fetal rat neural stem cells transplanted into the right brain. Prussian blue staining was used to observe the colonization and migration of implanted neural stem cells. MRI tracing was employed to monitor the signal changes of neural stem cells dynamically after in vivo transplantation. RESULTS AND CONCLUSION: Over 95% fetal rat neural stem cells were labeled successfully by superparamagnetic iron oxide particles, and under electron microscope, there were iron particles in labeled neural stem cells, which were concentrated in the lysosome and endosome. MRI results showed that the labeled neural stem cells had a changing trend of low signals. No difference was found in the cell viability between labeled and unlabeled cells, but T2WI and T2*WI signals were reduced in labeled neural stem cells. These findings confirm that superparamagnetic iron oxide-labeled fetal rat neural stem cells can highly express, and MRI tracing can be used for in vivo monitoring of neural stem cells.     中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Bone marrow mesenchymal stem cells/collagen/chitosan combined with bone transport for tibial defect repair: study protocol for a randomized controlled trial  Piao Cheng-zhe, Liu Jun, Liu Xin, Ma Yong, Cai Zhen-cun, Shi Dan 2015, 19 (32):  5231-5235.  doi: 10.3969/j.issn.2095-4344.2015.32.027 Abstract ( 235 )   PDF (833KB) ( 521 )   Save BACKGROUND: Bone marrow mesenchymal stem cells play an osteogenic role under the assistance of scaffold materials. The scaffold cannot only deliver the cells to the bone defect area, but also act as a new bone growth framework. Collagen-chitosan composite is one of ideal scaffold materials in bone tissue engineering, which has osteoinductive ability and better osteogenic ability than conventional scaffolds. Bone transport technology has been widely used in the clinical repair of long bone defects, but it has some deficiencies, such as slow osteogenesis, long  time for external fixation and nonunion. How to further accelerate bone formation and reduce complications has become the current problem to be solved. Here, it is hypothesized that bone marrow mesenchymal stem cells/ collagen/chitosan composite scaffold can increase the therapeutic effect of bone transport in the repair of tibial bone defects. METHODS/DESIGN: This study is a randomized controlled animal experiment, including in vitro and in vivo tests. In vitro test: Bone marrow mesenchymal stem cells are isolated from the bone marrow of New Zealand rabbits aged 1-2 months, and passaged to the third generation. Then, cell suspension is added onto the collagen-chitosan scaffold to construct the bone marrow mesenchymal stem cells/collagen/chitosan composite scaffold. In vivo test: Twenty-four New Zealand rabbits at 3-4 months are selected and randomly assigned to receive bone transport, scaffold implantation, bone transport+scaffold implantation, respectively. The primary outcome measures are the growth of implant materials and bone defect interface, X-ray detection of bone defect repair, hematoxylin-eosin staining and scanning electron microscope observation of bone formation in the bone defect region, immunohistochemical detection of type I collage expression in the osteogenic region, scanning electron microscope observation of interface bonding between implant materials and host bone, ultrastructure and bone formation. DISCUSSION: The results from this animal experiment will help to determine the feasibility of bone marrow mesenchymal stem cells/collagen/chitosan composite scaffold to accelerate bone repair during bone defect repair using bone transport technology. References | Related Articles | Metrics

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Application of placental mesenchymal stem cells Niu Ting, Li Ai-bin, Cao Jing-yun, Shen Lan, Chen Li 2015, 19 (32):  5236-5242.  doi: 10.3969/j.issn.2095-4344.2015.32.028 Abstract ( 377 )   PDF (949KB) ( 731 )   Save BACKGROUND: Placental mesenchymal stem cells are becoming a new source of seed cells because of wide range of sources, low immunogenicity and not involving ethical issues. OBJECTIVE: To elaborate the sources, biological characteristics and latest application of placental mesenchymal stem cells. METHODS: Literature search was performed in PubMed, ScienceDirect, OvidSP, CNKI databases for relevant literatures published from 2003 to 2015. The key words were “placenta, mesenchymal stem cells, placenta mesenchymal stem cells, cell transplantation, application mechanism” in Chinese and English, respectively. Then, 57 papers were further analyzed and reviewed in line with the theme. RESULTS AND CONCLUSION: Placental mesenchymal stem cells have been isolated and cultured successfully, and confirmed to have multi-differentiation potential. A large number of placental mesenchymal stem cells have been used in the experimental animal and clinical researches, and they have a great potential in bone tissue engineering, revascularizaion and nerve repair. However, the specific mechanism underlying the application of placental mesenchymal stem cells is not clear. In order to ensure the safety and effectiveness, there are still many problems to be further studied before placental mesenchymal stem cells are widely used in clinic.  中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Biological characteristics of different sources of mesenchymal stem cells Fang Hong-song, Zhou Jian-lin, Peng Hao, Deng Shuang, Weng Jin-qing, Liu Feng, Chen Sen, Zhou Guan-jin 2015, 19 (32):  5243-5248.  doi: 10.3969/j.issn.2095-4344.2015.32.029 Abstract ( 642 )   PDF (968KB) ( 765 )   Save BACKGROUND: Mesenchymal stem cells, under in vivo or in vitro specific induction conditions, can differentiate into the cartilage, muscle, tendons and so on. Clinical trials concerning mesenchymal stem cells mainly include tissue repair (such as bone, cartilage and joint repair) and treatment of heart, liver, spinal cord injury and nervous system diseases. OBJECTIVE: To compare the biological characteristics of mesenchymal stem cells from different sources. METHODS: PubMed and CNKI databases were retrieved for articles related to sources of mesenchymal stem cells and biological characteristics of mesenchymal stem cells published from 1987 to 2015. The retrieved articles were summarized and analyzed in the following aspects: cell surface marker, proliferation, differentiation, migration, and function, so as to explore the merits and demerits of mesenchymal stem cells from different sources. RESULTS AND CONCLUSION: A difference in the proliferation ability and surface markers is found between different sources of mesenchymal stem cells. Immunological competence of mesenchymal stem cells from different sources may be correlated with their activation status, species differences, tissue sources and culture  conditions, resulting the immunological competence of mesenchymal stem cells from different sources is not exact the same. In-depth understanding of the factors and mechanisms by which influence the migration of mesenchymal stem cells from different sources can enhance the migration ability of different sources of mesenchymal stem cells, and increase their efficiency in wound healing, tissue repair and regeneration treatment.  中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics