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    07 May 2014, Volume 18 Issue 19 Previous Issue    Next Issue
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    In vitro transfection of triple-point mutants of hypoxia-inducible factor 1 alpha into bone marrow mesenchymal stem cells
    Hui Chun-ying, Xiao Hong-yan, He Xin-ling, Wang Wei
    2014, 18 (19):  2953-2960.  doi: 10.3969/j.issn.2095-4344.2014.19.001
    Abstract ( 302 )   PDF (1145KB) ( 445 )   Save

    BACKGROUND: The incidence of spinal cord injury is increasing year by year in China so that the construction of effective vascular network in local injury as soon as possible is the guarantee of metabolism and nutritional support to differentiation of all kinds of cells and healing of injury is also promoted by vascular network.
    OBJECTIVE: To study the effect of triple-point mutants of hypoxia-inducible factor 1α (HIF1α) to promote angiogenesis after spinal cord injury in normoxic condition.
    METHODS: Site-directed mutagenesis of 402, 564 and 803 amino acids in human HIF1α coding sequence area was completed by PCR, and the adenovirus pAdEasy-1 system was recombined with post-mutation HIF1α gene. Packaging viral and titration determination of experimental group was completed and the same was done to non mutation group and control virus group. The future experiment was continued with three virus groups and blank group (A group: including mutation HIF1α gene virus liquid; B group: including non mutation HIF1α gene virus liquid; C group: including control virus liquid; D group: blank group). Then, virus liquid was transferred into rat bone marrow mesenchymal stem cells. We observed transfection efficiency of virus by enhanced green fluorescent protein and to detect mRNA and protein expression of HIF1α gene in all transfection cells. We also detected protein expression of vascular endothelial growth factor acting as downstream angiogenesis gene of HIF1α in four groups by Western blot.
    RESULTS AND CONCLUSION: Three adenoviral recombinants were successfully constructed and the packaging and identification were accomplished. The site-directed mutations of 402, 564 and 803 amino acids in coding sequence area were successful and all of them were changed to alanine. The level of HIF1α mRNA expression in both A group and B group were significantly higher than that in the C group and D group (P < 0.05). The expression levels of HIF1α and vascular endothelial growth factor proteins in A group was significantly higher than those in the other three groups (P < 0.05). These findings indicate that the HIF1α gene largely and effectively express in normoxic condition after triple-point mutation and the high-efficiency expression of vascular endothelial growth factor which is a downstream angiogenesis gene of HIF1α is promoted so that it is maybe a new therapeutic way of angiogenesis in the treatment of spinal cord injury.


    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


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    miR-378 effects on the proliferation and apoptosis of bone marrow mesenchymal stem cells under hypoxic-ischemic condition
    Guo Tian-zhu, Xing Yue, Hou Jing-ying, Zhou Chang-qing, Zhong Ting-ting, Zheng Shao-xin, Wang Tong
    2014, 18 (19):  2961-2967.  doi: 10.3969/j.issn.2095-4344.2014.19.002
    Abstract ( 368 )   PDF (907KB) ( 422 )   Save

    BACKGROUND: To investigate the influence of miR-378 on bone marrow mesenchymal stem cells proliferation and apoptosis under hypoxic-ischemic condition will provide a new method for further improving the survival of bone marrow mesenchymal stem cells in the infarcted myocardium.
    OBJECTIVE: To observe the viability of bone marrow mesenchymal stem cells and their ability of angiogenesis under hypoxic-ischemic condition after miR-378 transfection.
    METHODS: The bone marrow mesenchymal stem cells of Sprague-Dawley rat were cultured in vitro and divided into the non-transfection group and the transfection group. Mesenchymal stem cells were transfected with miR-378 mimic in the transfection group while the non-transfection group was taken as the control group. After transfection, the two groups were incubated under hypoxic-ischemic condition (serum-free medium,1% O2, 5% CO2, 94% N2) at 37 ℃ for 24 hours. Viability and proliferation of bone marrow mesenchymal stem cells were evaluated by the trypan-blue exclusion assay and MTS assay respectively. The cell apoptosis was detected by TUNEL assay. The culture supernatant of the two groups was collected separately and used as the conditioned medium after their exposure to hypoxic-ischemic environment. Endothelial cells were then co-cultured with the conditioned medium for the vasculature formation assay.
    RESULTS AND CONCLUSION: After the experience of hypoxia-ischemia for 24 hours and 72 hours, there was a larger number of the living cells in the transfection group in contrast to the non-transfection group (both P < 0.01). Compared with the non-transfection group, miR-378-MSCs group presented a stronger ability of proliferation, and their proliferation rates were obviously higher at 24 and 48 hours (P < 0.01 and P < 0.05, respectively). The percentage of apoptotic cells in the transfection group was significantly declined under the hypoxic-ischemic condition (P < 0.05). The vasculature formation assay indicated that the lumen-like structures were found in both of the two groups, however, the number of lumen-like structures was remarkably increased in the transfection group (P < 0.01). miR-378 could promote the proliferation of bone marrow mesenchymal stem cells and suppress their apoptosis under hypoxic-ischemic condition. It could also enhance their ability of vasculogenesis.


    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


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    Melatonin inhibits adipogenic differentiation of bone marrow mesenchymal stem cells through retinoid-related orphan receptor alpha
    Zhang Liang-ming, Wang Xuan, Gao Wen-jie, Huang Dong-sheng, Rong Li-min
    2014, 18 (19):  2968-2974.  doi: 10.3969/j.issn.2095-4344.2014.19.003
    Abstract ( 315 )   PDF (949KB) ( 740 )   Save

    BACKGROUND: Our previous studies have demonstrated that melatonin inhibits the adipogenic differentiation of bone marrow mesenchymal stem cells. However, the mechanism underlying the effect of melatonin on adipogenesis is still unknown.
    OBJECTIVE: To determine whether melatonin can inhibit the adipogenesis of bone marrow mesenchymal stem cells through retinoid-related orphan receptor α.
    METHODS: Bone marrow mesenchymal stem cells were isolated and purified by density gradient centrifugation combined with cell attachment culture. The phenotype was investigated by flow cytometry. Then, cells were induced for adipogenic differentiation with melatonin, CGP52608 and normal adipose tissue, respectively. The levels of retinoid-related orphan receptor α mRNA and protein were investigated by real-time RT-PCR and western blot assay, respectively. Further, the effect of retinoid-related orphan receptor α on the dipogenic differentiation of bone marrow mesenchymal stem cells was investigated by CGP52608.
    RESULTS AND CONCLUSION: The primary isolated bone marrow mesenchymal stem cells were spindle-shaped fibroblast-like cells. These cells did not express hematopoietic stem cells markers: CD34 and CD45; and highly expressed MSC markers: CD29, CD44, and CD10. The result of RT-PCR demonstrated that melatonin nuclear receptor, retinoid-related orphan receptor α, was highly expressed in bone marrow mesenchymal stem cells and the expression of retinoid-related orphan receptor α was further enhanced by melatonin in a dose-dependent manner, which was confirmed at protein level by western blot assay. During adiogenesis, the expression of retinoid-related orphan receptor α mRNA was up-regulated in the early stage, but maintained at a low level in the mild-later stage. While the retinoid-related orphan receptor α was activated by agonist CGP52608, the adipogenic differentiation of bone marrow mesenchymal stem cells was inhibited, which was similar to the inhibitory effect of melatonin. Therefore, melatonin inhibited the adipogenic differentiation of bone marrow mesenchymal stem cells through retinoid-related orphan receptor α, suggesting that melatonin plays an important role in the differentiation of adipocytes.


    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


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    Cultivation of myocardial tissue by using c-kit+ bone marrow mesenchymal stem cells and decellularized heart matrix
    Zhang Guang-wei, Gu Tian-xiang, Guan Xiao-yu, Zhang Yu-hai
    2014, 18 (19):  2975-2980.  doi: 10.3969/j.issn.2095-4344.2014.19.004
    Abstract ( 384 )   PDF (770KB) ( 429 )   Save

    BACKGROUND: Studies have found that c-kit+ bone marrow mesenchymal stem cells can differentiate into myocardial cells specifically, which may be the ideal seed cells.
    OBJECTIVE: To investigate the feasibility of cultivation of myocardial tissue by using c-kit+ bone marrow mesenchymal stem cells and decellularized heart matrix.
    METHODS: Heart tissues harvested from adult rats were decellularized for the following experiments. Primary rat bone marrow-derived mesenchymal stem cells were cultured in vitro. Until passage 8, bone marrow mesenchymal stem cells were enriched for c-kit and induced by 5 μmol/L 5-azacytidine for 2 weeks, and a second enrichment for the dihydropyridine receptor subunit α2δ1 was performed before analysis of cardiac differentiation or implantation into decellularized heart matrix for cultivation of myocardial tissue. Six weeks later, myocardial differentiation was identified by specific cardiac protein and action potential. Immunofluorescence staining was used to analysis neonatal myocardial tissue.
    RESULTS AND CONCLUSION: Six weeks after the second enrichment, 60% bone marrow mesenchymal stem cells expressed cardiac troponin T, GATA binding protein 4, and connexin 43, and these cells could be induced to yield cardiac action potential, which was identified as cardiac differentiation. And when implanted into decellularized heart matrix, these cells could form myocardial tissue arranged regularly.


    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


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    Differentiation of mesenchymal stem cells into cardiomyocyte-like cells induced by H9C2 cell culture medium
    Huang Lv, Zhang Hui, Xie Li, Li Yong-lin, Zhang Xiao-gang
    2014, 18 (19):  2981-2986.  doi: 10.3969/j.issn.2095-4344.2014.19.005
    Abstract ( 421 )   PDF (1855KB) ( 679 )   Save

    BACKGROUND: Mesenchymal stem cells can differentiate into cardiomyocytes in vitro induced by different methods, such as chemical drug induction, autologous transplantation in vivo, and in vitro simulation of cardiac-like microenvironment, but these inducible methods show low induction rate, complex operation, and toxic side effects.
    OBJECTIVE: To investigate the role of H9C2 cell culture medium in the differentiation of mesenchymal stem cells into cardiomyocyte-like cells.
    METHODS: Mesenchymal stem cells were obtained by the whole bone marrow adherent culture and H9C2 cell culture medium was prepared as a culture medium. Then mesenchymal stem cells were co-cultured with H9C2 cell culture medium for 1, 3, 5, 7 days. H9C2 cells cultured in 10% Dulbecco’s modified Eagle’s medium/Ham’s nutrient mixture F-12 served as positive control groups, while mesenchymal stem cells cultured in 10% Dulbecco’s modified Eagle’s medium/Ham’s nutrient mixture F-12 as negative control group. Immunofluorescence and western blot assay were used to detect expression of myocardial cell junction protein (desmin) and troponin T. Real-time quantitative PCR was applied to detect mRNA expression of myocardial cell trait genes, α-cardiac myosin heavy chain and β-myosin heavy chain.
    RESULTS AND CONCLUSION: After co-culture of H9C2 cell culture medium and mesenchymal stem cells for 7 days, the troponin T positive cells were up to (16±7)%, which was significantly higher than that of non-induced mesenchymal stem cells. Compared with the negative control group, the expression of troponin T protein and desmin after induction were significantly increased (P < 0.05) by western-blot detection; real-time PCR showed that the mRNA expression of α-cardiac myosin heavy chain and β-myosin heavy chain in differentiated cells were both up-regulated (P < 0.05). These findings suggest that H9C2 cell culture medium may induce the differentiation of mesenchymal stem cells into cardiomyocyte-like 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


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    Osteogenic induction of human adipose-derived stem cells
    Guo Yan-ping, Tao Chang-bo, Zhang Ai-jun, Li Xue-yang, Shen Cai-qi, Li Zheng-dao, Jin Pei-sheng
    2014, 18 (19):  2987-2992.  doi: 10.3969/j.issn.2095-4344.2014.19.006
    Abstract ( 539 )   PDF (1097KB) ( 626 )   Save

    BACKGROUND: A great amount of mesenchymal stem cells can be successfully derived from fat tissue and induced to differentiate into osteoblasts, chondrocytes, adipocytes and myocardial cells.
    OBJECTIVE: To establish the method of isolating, culturing and osteogenic differentiation of adipose-derived stem cells in vitro, and to explore the potential of adipose-derived stem cells as seed cells for bone tissue engineering.
    METHODS: Collagenase enzymatic digestion was used to isolate adipose-derived stem cells from human fat tissue which were then cultured in vitro. Flow cytometry was used to detect cell surface markers. Cell counting kit-8 assay was performed to examine cell viability. Adipose-derived stem cells were induced by osteogenic induced reagent to differentiate into bone cells. In addition, we also performed BCIP/NBT method to detect alkaline phosphatase activity. Alizarin red staining was used to detect the formation of calcium node. RT-PCR was performed to examine alkaline phosphatase and osteopontin expression.
    RESULTS AND CONCLUSION: We successfully obtained adipose-derived stem cells from fat aspirated liquid. Adipose-derived stem cells obtained could passage stably and proliferate highly. Flow cytometry data showed the expression of stem cell surface markers. Adipose-derived stem cells showed typical osteoblast morphology after osteogenic differentiation. Alkaline phosphatase staining was positive and alizarin red staining displayed the formation of calcium node. Furthermore, we found that alkaline phosphatase and osteopontin mRNA was expressed after differentiation 0, 3, 7, 14, 21, 28 days. These findings indicate that adipose-derived stem cells can be obtained from fat tissue through enzymatic digestion, differentiate towards bone cells, and express alkaline phosphatase and osteopontin, which can become potential seed cells for bone tissue engineering.


    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


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    Human adipose-derived stem cells promote angiogenic capability of random pattern flaps in mice
    Cao Jing, Jiang Nan, Xu Yang-yang, Zhu Meng-lin, Yang Liu
    2014, 18 (19):  2993-2998.  doi: 10.3969/j.issn.2095-4344.2014.19.007
    Abstract ( 330 )   PDF (2388KB) ( 398 )   Save

    BACKGROUND: Adipose-derived stem cells are a population of multilineage cells isolated from adipose tissue, which may have a positive effect on the treatment of ischemic diseases.
    OBJECTIVE: To explore the therapeutic and angiogenic effects of adipose-derived stem cells via local transplantation on random pattern skin flaps in mice.
    METHODS: Human adipose-derived stem cells were isolated, cultured and passaged in vitro. On the back of the SPF mice, random pattern skin flaps were performed. After the operation, the adipose-derived stem cells were injected into the pedicle, central, and distal end of the flaps in the experimental group, while only PBS was injected into the control flaps. Seven days later, the survival rate of flaps was evaluated. Immunofluorescence assay was preformed to observe the distribution of microvessels in the flaps and trace the CM-Dil labeled adipose-derived stem cells, while the level of vascular endothelial growth factor was tested by ELISA and the protein expression of stromal cell-derived factor 1 tested by western blot at day 14 after adipose-derived stem cells transplantation.
    RESULTS AND CONCLUSION: Compared with the control group, there was a significant increase in the flap survival rate in the experimental group, and along with the sharply increased number of microvessels, the secretions of vascular endothelial growth factor and stromal cell-derived factor 1 were also obviously raised in the experimental group (P < 0.05). After local transplantation of adipose-derived stem cells into random skin flaps, it could intervene the secretion of vascular endothelial growth factor and stromal cell-derived factor 1 and promote angiogenesis of the skin flap.


    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


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    Effect and mechanism of mesenchymal stem cells on the aging kidney
    Li Yan-ju, Liu Yang, Wang Ji-shi, Meng Wei, Huang Yi
    2014, 18 (19):  2999-3004.  doi: 10.3969/j.issn.2095-4344.2014.19.008
    Abstract ( 289 )   PDF (721KB) ( 424 )   Save

    BACKGROUND: Bone marrow mesenchymal stem cells can differentiate into renal tubular epithelial cells. It is speculated that mesenchymal stem cells may have a therapeutic role in chronic renal dysfunction.
    OBJECTIVE: To study the effect of bone marrow mesenchymal stem cells on the aging kidney of rats and its possible mechanism.
    METHODS: Aging models were established in rats. Rat bone marrow mesechymal stem cells cultured in vitro were labeled by 5,6-carboxyfluorescein diacetate, succinimidyl ester, and then injected into the rats via the tail vein. Under a fluorescence microscope, the cell homing was observed. The contents of urea nitrogen and creatinine in serum of rats were tested by automatic biochemical analyzer. The overlaying of blue fluorescent 2-(4-amidinophenyl)-6-indolecarbamidine dihydrochloride and the specific protein of red fluorescent tubular epithelial cells (keratin protein) was detected by indirect immunofluorescence and computer image stacking technique.
    RESULTS AND CONCLUSION: Bone marrow mesenchymal stem cells could arrive the kidney of rats. Compared with the model group, the contents of urea nitrogen and creatinine in serum were decreased in the treated group (P < 0.05). The treated group showed a small amount of double positive cells. These findings indicate that rat bone marrow mesenchymal stem cells may differentiate into renal tubular epithelial cells, which may be one of mechanisms that improve the aging kidney of rats.


    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


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    Systematic establishment for directed differentiation of induced pluripotent stem cells into hematopoietic progenitor cells in vitro
    Shen Jie, Li Jin-hui, Ruan Jing, Qiu Yun, Zhu Yin, Cui Da-xiang, Fan Zhi-hong, Wang Zheng
    2014, 18 (19):  3005-3011.  doi: 10.3969/j.issn.2095-4344.2014.19.009
    Abstract ( 362 )   PDF (1279KB) ( 630 )   Save

    BACKGROUND: There have been a large number of reports on establishing induced pluripotent stem cell lines, but studies concerning large-scale in vitro induced differentiation of induced pluripotent stem cells into hematopoietic progenitor cells still have a lack of in-depth discussion.
    OBJECTIVE: To develop methods to induce differentiation of induced pluripotent stem cells into hematopoietic progenitor cells in vitro.
    METHODS: Using the method of infection with lentivirus particles containing four transcriptionfactor genes, which are Oct4, Sox2, Nanog and Lin28, human skin fibroblasts are transduced into induced pluripotent stem cells. In the induced differentiation system, Y-27632, a kind of tyrosine kinase inhibitor-ROCK (p160-Rho-associated coiled-coil kinase), was added, which obviously suppressed apoptosis of cells. Based on conditioned medium with OP9 cells, a differentiation system of inducing induced pluripotent stem cells differentiating into hematopoietic progenitor cells was established.
    RESULTS AND CONCLUSION: (1) Apoptosis of induced pluripotent stem cells at the first three passages was very obvious, and the cells were difficult in a large-scale expansion. After Y-27632 was added, the apoptosis of embryonic stem cells was obviously inhibited. (2) During embryoid body differentiation, induced pluripotent stem cells cultured in OP9 conditional growth medium differentiated into hematopoietic progenitor cells in vitro that were positive for CD34.


    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


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    Human umbilical cord mesenchymal stem cells: isolation, identification and multi-directional differentiation
    Li Mao, Huang Wen
    2014, 18 (19):  3012-3016.  doi: 10.3969/j.issn.2095-4344.2014.19.010
    Abstract ( 444 )   PDF (2047KB) ( 534 )   Save

    BACKGROUND: The umbilical cord mesenchymal stem cells can be obtained conveniently, noninvasively, without ethical restrictions, which are more original that stem cells from other sources and have little immunogenicity. Therefore, umbilical cord mesenchymal stem cells are a kind of ideal seed cells with promising application prospects.
    OBJECTIVE: To induce the differentiation of mesenchymal stem cells derived from human umbilical cord into adipocytes and osteoblasts.
    METHODS: The umbilical cord mesenchymal stem cells were isolated by tissue explants method. The morphology, proliferation and immunophenotype of the 3rd passage cells were analyzed, and then cells were induced to adipocytes and osteoblasts in vitro. The expressions of CD90, CD105, CD34 and CD15 were detected by flow cytometry.
    RESULTS AND CONCLUSION: The umbilical cord mesenchymal stem cells isolated by tissue explants method could be cultured and proliferated in vitro. Flow cytometry analysis revealed that the cells were strongly positive for CD90 and CD105, but negative for CD34 and CD45. The umbilical cord mesenchymal stem cells could be induced to adipocytes and osteoblasts in vitro. These findings indicate that mesenchymal stem cells can be successfully obtained from human umbilical cord by tissue explants method and have the potential of multi-directional differentiation.


    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


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    Growth factors promote the therapeutical effects of autologous bone marrow mesenchymal stem cell transplantation on acute myocardial infarction
    Wen Ti, Qi Xun
    2014, 18 (19):  3017-3022.  doi: 10.3969/j.issn.2095-4344.2014.19.011
    Abstract ( 215 )   PDF (2139KB) ( 429 )   Save

    BACKGROUND: The growth factor is a potent mobilization agent for stem cells, which can increase adhesion and proliferation of injected cells, induce stem cells to migrate to the infarct zone, proliferate, differentiate, as well as participate in myocardiac repair.
    OBJECTIVE: To investigate the effect of hepatocyte growth factor (HGF) and insulin-like growth factor (IGF) on transplantation of bone marrow mesenchymal stem cells (BMSCs) after acute myocardial infarction. 
    METHODS: Primary rabbit BMSCs were cultured in vitro and labeled by red fluorescence dye CM-Dil for transplantation. After the mid third of left anterior descending was ligated, model rabbits were grouped into four groups: control group, BMSCs group, HGF+IGF group, and HGF+IGF+BMSCs group (n=6 in each group). Different interventional agents were injected into the myocardium at four sites within the ischemic region. Masson trichrome staining was performed to determine viable myocardium, and immunofluorescence staining was used to identify BMSCs differentiation. The cardiac function was assessed with Doppler echocardiography.
    RESULTS AND CONCLUSION: Four weeks after treatment, CM-Dil/cTNT+ cells significantly increased in the HGF+IGF+BMSCs group, compared with BMSCs group. Consequently, viable myocardial tissues significantly increased, left ventricular ejection fraction was significantly improved, and left ventricular end-diatolic volume significantly decreased in the HGF+IGF+BMSCs group, relative to the other three groups. Combination of HGF and IGF that promotes differentiation of transplanted autologous BMSCs into cardiomyocytes, thus increasing viable myocardium, improving left ventricular function, and inhibiting left ventricular remodeling, may be a new method for the cell treatment of acute myocardial 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


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    Biological characteristics of bone marrow versus adipose-derived mesenchymal stem cells from C57 mice
    Zhang Rong-yao, Bi Xiao-juan, Ma Yan, Duan Xian-lin, Xue Wen-jing, Wang Yi-chun, Jiang Ming
    2014, 18 (19):  3023-3029.  doi: 10.3969/j.issn.2095-4344.2014.19.012
    Abstract ( 477 )   PDF (2181KB) ( 729 )   Save

     

     


    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


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    BACKGROUND: Adipose-derived stem cells have similar characteristics of bone marrow mesenchymal stem cells, including adhesion and fibroblast-like clone formation. In addition, adipose-derived stem cells can differentiate into bone, adipose, cartilage.
    OBJECTIVE: To compare the biological characteristics of C57 mouse adipose-derived and bone marrow mesenchymal stem cells.
    METHODS: Mesenchymal stem cells were obtained respectively from the adipose and bone marrow of C57 mice under sterile condition. Adipose-derived and bone marrow mesenchymal stem cells were isolated and purified in vitro to determine cell morphology, suface marker, growth kinetics, differentiation potential and Notch signal related gene.
    RESULTS AND CONCLUSION: Morphology was similar between adipose-derived and bone marrow mesenchymal stem cells under an optical microscope. Adipose-derived and bone marrow mesenchymal stem cells at passage 3 expressed CD29, CD105, Sca-1, but not expressed CD34, CD133. In addition, bone marrow mesenchymal stem cells also expressed CD45. The growth kinetics and cell clone analysis indicated that the proliferation rate of adipose-derived mesenchymal stem cells was significantly faster than that of bone marrow mesenchymal stem cells. Both of adipose-derived and bone marrow mesenchymal stem cells were induced to osteoblasts, adipocytes and chondrocytes, but the adipose-derived mesenchymal stem cells were easier to the osteogenetic differentiation than bone marrow mesenchymal stem cells. Notch signal related gene detection shown that adipose-derived mesenchymal stem cells expressed lower levels of Jagged-1 mRNA, but higher levels of Hes-1 mRNA compared with bone marrow mesenchymal stem cells. These findings suggest that the proliferation rate of adipose-derived mesenchymal stem cells is significantly faster than that of bone marrow mesenchymal stem cells, and the former is apt to the osteogenetic differentiation and may be related with the expressed levels of Hes-1 mRNA.

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    Comparative study of proliferative and neurosphere differentiation capacities of adipose-derived mesenchymal stem cells at different passages
    Zhang En-feng, Cao Rui, Xu Tao, Wang Xiao-yan, Maierdan Maimaiti, Wang Guo-qi, Sheng Wei-bin
    2014, 18 (19):  3030-3035.  doi: 10.3969/j.issn.2095-4344.2014.19.013
    Abstract ( 259 )   PDF (839KB) ( 545 )   Save

    BACKGROUND: Adipose-derived mesenchymal stem cells are a group of pluripotent stem cells, and under certain conditions can differentiate into neural stem cells in vitro.
    OBJECTIVE: To investigate the proliferative and differentiation ability of different passage mesenchymal stem cell from adipose tissue into neurospheres.
    METHODS: The adipose-derived mesenchymal stem cells from Sprague-Dawley rats were separated and cultured in vitro, and morphology and proliferation rate of cells were observed and compared respectively at passages 3, 6, 10 and 20. The cell surface antigens and cells cycle were identified by flow cytometry. Furthermore, adipose-derived mesenchymal stem cells were induced into neurospheres, and the neurosphere rate was counted.
    RESULTS AND CONCLUSION: Adipose-derived mesenchymal stem cells were mainly in long spindle shape, and cells at different passages had highly proliferative capacity in vitro. Except passage 3, adipose-derived mesenchymal stem cells strongly expressed CD29, CD44, CD73 and lowly expressed CD45 and CD34. The proportion of G0/G1 phase in cell cycle was 93.4% at passage 3, 92.7% at passage 6, 92.4% at passage 10, 86.0% at passage 20. Adipose-derived mesenchymal stem cells at passages 6 and 10 were easier to differentiate into neurospheres than those at passage 20 (P < 0.05), but cells at passage 3 were difficult to differentiate into neurospheres. Therefore, when using adipose-derived mesenchymal stem cells as seed cells, we should pay attention to choose the appropriate amplification passage in order to obtain the cells with best differentiation potential and cell purity.


    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


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    Subarachnoid block has no influence on neural differentiation potential of mononuclear cells in the cord blood
    Zhang Yu-wen, Zhang Xiao-qing, Zhou Ning, Liu Xing-yuan
    2014, 18 (19):  3036-3041.  doi: 10.3969/j.issn.2095-4344.2014.19.014
    Abstract ( 360 )   PDF (1033KB) ( 374 )   Save

    BACKGROUND: Some scholars believe that anesthesia can improve the quality of cord blood separation and collection, and subarachnoid block is commonly used in cesarean section, but there is no research on the influence of subarachnoid block on differentiation potential of mononuclear cells into neural stem cells and astrocytes. 
    OBJECTIVE: To study the influence of subarachnoid block on differentiation potential of mononuclear cells in the cord blood of neonates into neural stem cells and astrocytes. 
    METHODS: Mononuclear cells isolated from 20 neonates delivered spontaneously and 20 neonates born by cesarean delivery undergoing subarachnoid block were cultured in vitro, acting as control group and study group. After delivery, cord blood samples were taken to isolate and culture mononuclear cells in vitro. After 3 days of routine culture, the cells were subject to non-induced culture and induced differentiation into neural stem cells. The cultured cells were identified by the cell makers Nestin and glial fibrillary acidic protein with immunohistochemistry identification. 
    RESULTS AND CONCLUSION: The Nestin and glial fibrillary acidic protein showed no difference between the two groups after induced differentiation (P > 0.05), indicating subarachnoid block has no impact on the differentiation of mononuclear cells in the cord blood from neonates into neural stem cells and astrocytes.


    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


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    Culture methods for primary human umbilical cord mesenchymal stem cells
    Wang Fei, Zhou Hong, Guo Yu-cheng, Su Xiao-xia, Li Ang, Zhao Xiao-peng
    2014, 18 (19):  3042-3047.  doi: 10.3969/j.issn.2095-4344.2014.19.015
    Abstract ( 876 )   PDF (806KB) ( 895 )   Save

    BACKGROUND: How to get a lot of stable and dynamic human umbilical cord mesenchymal stem cells is still a difficulty.
    OBJECTIVE: To investigate the best culture way of human umbilical cord mesenchymal stem cells by comparing tissue explant, enzyme digestion, enzymolysis methods.
    METHODS: Ten fresh umbilical cords were isolated to human umbilical cord mesenchymal stem cells by using tissue explant, enzyme digestion, and enzymolysis methods, respectively. Hereafter, we compared cells harvested using three culture methods in terms of the time for the primary cells to creep, successful rates of cell-culturing, and curves of cell growth. Surface markers of cells were detected by flow cytometry and multiple differentiations of cells were detected.
    RESULTS AND CONCLUSION: The time for the primary cells to creep out in the enzymolysis and enzyme digestion groups had no significant difference, but both of them were significantly shorter than that in the tissue explant group (P < 0.01). The successful rate of primary cell culture in the enzymolysis group was significantly higher than that in the other two groups. The cell proliferations of three groups had no significant difference. The cell surface markers and differentiation ability of the third generation of human umbilical cord mesenchymal stem cells cultured by enzymolysis method met the characteristics of mesenchymal stem cells. In conclusion, the enzymolysis method can shorten the time for primary human umbilical cord mesenchymal stem cells to creep out and increase the successful rates of primary 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


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    Clinical observation of cardiac function in patients with acute myocardial infarction complicated with heart failure undergoing stem cell transplantation: a 2-year follow-up visit
    Chen Xiao-ming, Yang Zhi-yong, Cui Dan-yang, Zhang Ming
    2014, 18 (19):  3048-3053.  doi: 10.3969/j.issn.2095-4344.2014.19.016
    Abstract ( 318 )   PDF (325KB) ( 380 )   Save

    BACKGROUND: Many animal experiments and clinical trials have demonstrated that stem cell transplantation can improve heart function and reduce ventricular dilatation and ventricular remodeling, which has shown an incomparable superiority over traditional therapies in the treatment of myocardial infarction complicated by heart failure.
    OBJECTIVE: To observe the clinical effects of single autologous peripheral blood stem cell transplantation in acute myocardial infarction patients with heart failure.
    METHODS: Since 2006 August to 2010 June, 23 patients who were diagnosed to have acute ST elevation myocardial infarction complicated with heart failure were selected and divided into two groups: cell transplantation group (n=11) and control group (n=12). All patients underwent emergency coronary angiography and percutaneous coronary intervention with drug eluting stent implantation. In the stem cell transplantation group, peripheral blood stem cells positive for CD34 (about 1×108) were collected mobilized by granulocyte colony stimulating factor at 5 days after stent implantation, and then the cells were injected into infarcted vessels using coronary angiography method. Two-year follow-up was performed after cell transplantation to observe the cardiac function and adverse reactions in patients.
    RESULTS AND CONCLUSION: After 6 months of follow-up, the cardiac function in the cell transplantation group was improved significantly compared with that before cell transplantation (P < 0.05), and the left ventricular ejection fraction was increased by (6.2±0.2)% and the left ventricular end diastolic volume was reduced by (4.7±2.9) mm. However, there was no difference in follow-up results by the end of 1 and 2 years after cell transplantation (P > 0.05), as well as no adverse reaction occurred. In the control group, after 6 months of follow-up, the left ventricular ejection fraction was reduced by (0.5±0.1)% and the left ventricular end diastolic volume was increased by (0.4±0.3) mm, which were deteriorated year after year. Percutaneous coronary intervention with autologous peripheral blood stem cell transplantation can significantly improve the left ventricular function, reduce left ventricular volume, and delay or prevent left ventricular remodeling in patients with acute myocardial infarction, which is safe and effective. But up to 2 years after cell transplantation, the cardiac function shows no further improvement.


    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


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    Co-transplantation of fetal rat pancreatic stem cells and islets of langerhans to preserve viability of islets
    Zhang Yu-sen, Bao Shi-yun, Zhang Yue, Li Fu-rong
    2014, 18 (19):  3054-3059.  doi: 10.3969/j.issn.2095-4344.2014.19.017
    Abstract ( 304 )   PDF (821KB) ( 346 )   Save

    BACKGROUND: Pancreatic stem cells can maintain the islet structure in vitro, reduce necrosis and apoptosis of islet cells, in vitro prolong islet survival, and keep islet activity.
    OBJECTIVE: To observe the possibility of preserving viability of islets in vivo by co-transplanting fetal rat pancreatic stem cells and islets so as to improve the outcome of islet transplantation.
    METHODS: Thirty-five adult rats were randomly divided into five groups, including co-transplantation, islet transplantation alone, pancreatic stem cell transplantation alone, diabetes control and normal control groups. Diabetic models were established in the former four groups by intraperitoneal injection of streptozotocin-citrate buffer. Pancreatic stem cells from the fetal rats at pregnant 16 days or islets from adult Sprague-Dawley rats.
    RESULTS AND CONCLUSION: In the co-transplantation group, the levels of blood glucose and plasma insulin returned to the normal after 5 days of co-transplantation, and the survival time of islets was (18.2±2.4) days. In the islet transplantation alone group, the level of blood glucose was reduced to normal after 1 week of transplantation, and the survival time of islets was (14.4±2.1) days. Significant different was found between the survival time of islets between this two groups (P < 0.05). However, the level of blood glucose was still abnormal in the other groups. These findings indicate that the co-transplantation of fetal rat pancreatic stem cells and islets can prolong the survival time of islets in vivo, protect viability of islets and improve the outcome of islet transplantation.


    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


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    Wnt/beta-catenin signaling pathway mediates oriented differentiation of hair-follicle-generating stem cells induced by keratinocyte growth factor and lithium chloride
    Yang Bin, Deng Li-huan, Li Bing-hang, Wu Xiao-ying, Ding Yu-de, Dong Xue (Plastic Surgery Hospital
    2014, 18 (19):  3060-3068.  doi: 10.3969/j.issn.2095-4344.2014.19.018
    Abstract ( 512 )   PDF (3520KB) ( 387 )   Save

    BACKGROUND: The proliferation and differentiation of hair-follicle-generating stem cells are influenced by the joint action of their own genes and external signals. Wnt/β-catenin signaling pathway plays an important role in the development of hair follicles, but the detailed mechanisms are not yet clear.
    OBJECTIVE: To investigate, with interruption of keratinocyte growth factor and lithium chloride, the function and the interrelationship of Wnt/β-catenin signaling pathway with other signal factors when human hair-follicle-generating stem cells differentiate into dermal papilla cells or epidermal cells.
    METHODS: Hair-follicle-generating stem cells were isolated from the bulge and cultivated. Then the growth curve of hair-follicle-generating stem cells was tested and formed in order to observe the cell proliferation ability cultivated at different densities (1×106/L, 1×107/L, 1×108/L, 1×109/L) at each time. Immunoflurorescene staining was performed to identify hair-follicle-generating stem cells and their differentiated cells. Lithium chloride (0, 0.5, 1.5, 10, 25 mmol/L individually) and keratinocyte growth factor (0, 10, 25, 50, 100 μg/L individually) were used to induce the differentiation of hair-follicle-generating stem cells. Then, we contrasted and analyzed the proliferation ability in each case, thereby investigating the most appropriate concentration of keratinocyte growth factor and lithium chloride to spur the differentiation of hair-follicle-generating stem cells. At days 3, 5, 7 and 9, we tested and compared the mRNA expressions of β-catenin, APC, GSK-3β, Axin and Lef1 from cells in control group, 10 mmol/L lithium chloride group and 10 μg/L keratinocyte growth factor group.
    RESULTS AND CONCLUSION: Isolating cultured hair-follicle-generating stem cells still had a great reproductive activity and multi-lineage potential even after various times subculture in vitro. With higher lithium chloride concentration, the proliferation ability of hair-follicle-generating stem cells declined; while it increased when keratinocyte growth factor concentration increased. In K-SFM medium which contained lithium chloride, the transformation of hair-follicle-generating stem cells was obvious, showing distinct differences among groups. Especially, the level of β-catenin reached the peak when lithium chloride > 10 mmol/L. However, in K-SFM medium which contained keratinocyte growth factor, hair-follicle-generating stem cells differentiated into epidermal cells and the level of β-catenin changed slightly. We found that, while spurring the differentiation of hair-follicle-generating stem cells, lithium chloride could activate Wnt/β-catenin signal pathway and inhibit GSK-3β, a vital component of degradation compound. This facilitated β-catenin expressing in the cytoplasm to translocate into the nucleus. As a result, the transcription of target gene increased. It is the most appropriate concentration to spur hair-follicle-generating stem cells differentiation when lithium chloride level is > 10 mmol/L, but the proliferation ability declines correspondingly. Keratinocyte growth factor, which can facilitate hair-follicle-generating stem cells differentiated into epidermal cells, is a key factor to accelerate proliferation ability and migration of hair-follicle-generating stem cells, re-epithelialization and healing of wound. The mechanisms of hair-follicle-generating stem cells oriented differentiation induced by lithium chloride and keratinocyte growth factor are activating Wnt/β-catenin signal pathway, inducing change of β-catenin expression, and activating the transcription of target gene related to Wnt/β-catenin 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


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    Sodium-calcium exchanger promoter promotes differentiation of mouse induced pluripotent stem cells into cardiomyocytes in vitro
    Dai Bo, Li Hong-tao, Liu Ze, Xiao Ding-zhang, Yu Xi-yong, Chen Bin, He Jian-xin, Xiang Ding-cheng, Qiu Jian, Wang Yi-gang
    2014, 18 (19):  3069-3074.  doi: 10.3969/j.issn.2095-4344.2014.19.019
    Abstract ( 382 )   PDF (2492KB) ( 422 )   Save

    BACKGROUND: Inducing pluripotent stem cells has been considered as a promising treatment for ischemic heart disease. However, an ideal inducing method has not been found yet.
    OBJECTIVE: To investigate the role of sodium-calcium exchanger (NCX1) promoter in the differentiation of mouse induced pluriptent stem cells (miPS) into cardiomyocytes.
    METHODS: The pLVX-IRES-ZsGreen1 vectors which contain NCX1 promoter constructed by recombinant DNA technology were co-transfected to 293FT cells with ViraPowerTM Lentiviral Packaging Mix. The recombinant lentiviruses infected with miPS were selected and purified by puromycin. miPS were recovered and passaged to form embryoid bodies. The embryoid bodies were induced by differentiation medium containing various concentrations of the virus titer. The number of beating embryoid bodies were calculated. The expression profiles of the myocardial intra-markers were tested to determine the differentiation efficiency of iPSC by RT-PCR and immunofluorescence analysis.
    RESULTS AND CONCLUSION: pLVX-IRES-ZsGreen1 vectors which contain NCX1 promoter were constructed and confirmed by PCR. Virus could be packaged, purified and concentrated successfully. The recombinant lentivirus to transduce miPS was sorted by flow cytometry. In contrast to NCX1-/GFP- cells, NCX1+/GFP+ cells were differentiated and developed prominent beating areas with sustained contractile activity for additional 4 days, and demonstrated positive expression of gap communication marker CX43 and cardiac troponin. The expressions of GATA4, MEF2c and Nkx2.5 in the NCX1+ cells were 4.2, 7.5, and 2.5 times those in NCX1- cells. Results showed the NCX1 promoter can promote the cardiac differentiation of miPS.


    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


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    Bone morphogenetic protein 2 stimulated osteo-chondrogenic differentiation of patellar tendon-derived stem cells isolated from a failed tendon-healing animal model of tendinopathy
    Lin Yu-cheng, Wang Chen, Rui Yun-feng, Cheng Xin-kun, Ma Liang-yu
    2014, 18 (19):  3075-3081.  doi: 10.3969/j.issn.2095-4344.2014.19.020
    Abstract ( 343 )   PDF (3412KB) ( 413 )   Save

    BACKGROUND: The pathogenesis of tendinopathy remains unclear and hence treatment of tendinopathy is usually palliative.
    OBJECTIVE: To investigate the effects of bone morphogenetic protein 2 on the osteogenic and chondrogenic differentiation of patellar tendon-derived stem cells isolated from collagenase-induced tendinopathy rats in vitro. 
    METHODS: Patellar tendon-derived stem cells were isolated from patellar tendons of collagenase-induced tendinopathy rats. The multi-differentiation potential of patellar tendon-derived stem cells at passage 3 was identified by osteogenic, adipogenic and chondrogenic differentiation assays. The patellar tendon-derived stem cells were cultured to the 3rd passage in complete culture medium, and then the cells were divided into two groups with (experimental group) or without recombinant human bone morphogenetic protein 2 (control group) until the cells reached confluence for 7 days. Their osteogenic response to bone morphogenetic protein 2 in vitro  was examined by alizarin red S staining of calciumnodule formation and quantification assay. The patellar tendon-derived stem cell pellets were cultured in complete culture medium with (experimental group) or without bone morphogenetic protein 2 (control grup) for 21 days. Chondrogenic differentiation of the cell pellets was evaluated by hematoxylin-eosin staining, alcian blue staining, immunohistochemical staining for Sox9 and collagen type II.
    RESULTS AND CONCLUSION: Primary patellar tendon-derived stem cells from the tendinopathy rats cultured in vitro showed clonal growth; after passage, spindle fibroblast-like and flat-like cells were detectable. The cells were positive for oil red O staining at 10 days after adipogenic induction, positive for alizarin red staining at 7 days after osteogenic induction, and positive for hematoxylin-eosin staining and immunohistochemical staining of collagen type II at 14 days after chondrogenic induction. After patellar tendon-derived stem cells were induced with recombinant human bone morphogenetic protein 2 for 7 days, the result of alizarin red staining was positive in the experimental group, but negative in the control group without recombinant human bone morphogenetic protein 2. The difference in the result of alizarin red staining between the two groups was statistically significant. After patellar tendon-derived stem cells were induced with recombinant human bone morphogenetic protein 2 for 21 days, the results of hematoxylin-eosin staining, alcian blue staining, immunohistochemical staining for Sox9 and collagen type II were all positive. In conclusion, bone morphogenetic protein 2 could stimulate the osteogenic and chondrogenic differentiation of patellar tendon-derived stem cells isolated from collagenase-induced tendinopathy rats in vitro, which can help to better understand the pathogenesis of tendinopathy. 


    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


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    Biological characters of mesenchymal stem cells separated from different components of human placenta
    Hong Yan, Huo Si-wei, Lu Yao, Zhang Yi
    2014, 18 (19):  3082-3087.  doi: 10.3969/j.issn.2095-4344.2014.19.021
    Abstract ( 448 )   PDF (2144KB) ( 613 )   Save

    BACKGROUND: Human placenta is a stable source for mesenchymal stem cells, which is becoming a cell source in the regenerative medicine that attracts widespread attentions.
    OBJECTIVE: To compare the biological characters of mesenchymal stem cells that separated from different components of human placenta (human chorion and villous trophoblast).
    METHODS: The amniotic membrane of placenta surface was detached surgically. Human chorion and villous trophoblast in the fetal side was cut into pieces. After digestion with PBS containing collagenase II, mononuclear cells were separated and cultured in Dulbecco’s modified Eagle’s medium containing 10% fetal bovine serum in 37 ℃ and 5% CO2, 95% saturated humidity, after 48 hours the full amount in liquid, dislodge suspension cells. Forty-eight hours later, the medium was changed completed, and non-adherent cells were removed. When cell fusion reached about 90%, trypsin digestion was employed for cell passage. Biological characters of mesenchymal stem cells separated from different components of human placenta were compared through observation of total number of primary cells, cell morphology, and surface markers expression (CD90, CD73 and CD105).
    RESULTS AND CONCLUSION: The flow cytometric analysis revealed that the cells separated from the human chorion and villous trophoblast were over 90% strongly positive for CD90, CD73, CD105. These two sources of cells showed typical fibroblast morphology, suggesting that the cells have the characteristics of mesenchymal stem cells. Under the same enzyme digestion time, enzyme concentration, and shaking speed, more cells are visible from the chorion, and the subsequent culture is easier to harvest 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


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    Establishment and characterization of CD-1 neonatal female germline stem cell lines
    Bi Hai-wei, Wu Ji
    2014, 18 (19):  3088-3093.  doi: 10.3969/j.issn.2095-4344.2014.19.022
    Abstract ( 374 )   PDF (2593KB) ( 581 )   Save

    BACKGROUND: In traditional view, the germ cells in mammal ovaries were considered to lose the capacity of self-renew after birth. However, recent studies have showed the existence of female germline stem cells (FGSCs) which take the responsibility for regenerating oocytes in the ovaries of postnatal mammals.
    OBJECTIVE: To establish the FGSC line and to explore the biological characterization of the FGSC line from neonatal CD-1 mouse ovaries.
    METHODS: Using two-step enzymatic digestion and immunomagnetic purification, Fragilis-positive cells were isolated from neonatal CD-1 mice ovaries and subjected to long-term culture. Then their biological characteristics were studies with RT-PCR, fluorescent immunocytochemistry and cytogenetic analysis.
    RESULTS AND CONCLUSION: FGSCs from neonatal CD-1 mice aged 3-5 days were isolated and long-term cultured for 70 passages to establish a cell line. This cell line expressed Mvh, Dazl, Oct-4, Stella, Blimp1, Fragilis, and was proven to be mitotically active germ cells by BrdU/Fragilis dual immunofluorescence analysis. Moreover, cytogenetic analysis by Gimsa staining confirmed their normal karyotype. In all, these results indicate the existence of FGSCs with mitotic activity in neonatal CD-1 mouse ovaries.


    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


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    Umbilical cord mesenchymal stem cell transplantation for the treatment of diabetes mellitus in rats
    Li Li, Zhao Bo
    2014, 18 (19):  3094-3099.  doi: 10.3969/j.issn.2095-4344.2014.19.023
    Abstract ( 411 )   PDF (833KB) ( 450 )   Save

    BACKGROUND: Umbilical cord mesenchymal stem cells have strong proliferation and differentiation capacities, and can be induced to differentiate into pancreatic β cells, thereby playing a therapeutic effect on diabetes mellitus.
    OBJECTIVE: To study the therapeutic effects of transplantation of umbilical cord mesenchymal stem cells for treatment of diabetes mellitus in rats.
    METHODS: Thirty male Sprague-Dawley rats were randomly divided into control group (n=6), transplantation group (n=12) and diabetic group (n=12). Rats in the control group were given normal saline injection. Rats in the other two groups were injected with streptozotocin at a dose of 45 mg/kg to establish diabetic models. After modeling, transplantation of umbilical cord mesenchymal stem cells via tail vein was given in the transplantation group.
    RESULTS AND CONCLUSION: Thirty days after modeling, the fasting blood glucose was maintained at a higher level in comparison with the control group (P < 0.05). However, compared with the diabetic group, the level of fasting blood glucose was significantly reduced (P < 0.05) and the body mass was increased in the transplantation group (P < 0.05). After 45 days, the fasting blood glucose level and body mass in the transplantation group had no significant difference from those in the control group (P > 0.05), but in the diabetic group, the fasting blood glucose level was still higher and the body mass continued to decrease. These findings indicate that the transplantation of umbilical cord mesenchymal stem cells can be effective in the treatment of diabetes mellitus in rats.


    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


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    Expression of recombinant retroviral vector carrying nerve growth factor gene in neural stem cells
    He Jun-feng, Gu Guo-mei, Long Da-hong
    2014, 18 (19):  3100-3104.  doi: 10.3969/j.issn.2095-4344.2014.19.024
    Abstract ( 430 )   PDF (658KB) ( 484 )   Save

    BACKGROUND: Nerve growth factor (NGF) belongs to a biological macromolecule that is difficult to pass through the blood-brain barrier. However, a retroviral vector carrying exogenous gene can be stably inserted and integrated into the host cell genome, which is suitable for gene therapy.
    OBJECTIVE: To study the gene expression of recombinant retroviral vector carrying rat NGF gene in neural stem cells.
    METHODS: The rat NGF gene was inserted into a retroviral vector pLEGFP-N1, which was transferred into packaging cells PT67 by Lipofectamine 2000. The positive clones in virus supernatant were collected by G418 selection and used to infect neural stem cells. After that, the NGF expression was tested by enzyme linked immunosorbent assay and the biological competence by PC12 cells, and then morphological change of neural stem cells and cell typing were examined by fluorescent microscope.
    RESULTS AND CONCLUSION: The neural stem cells could express extrinsic source NGF protein after the recombinant plasmid was infected into neural stem cells. The PC12 cells increased in the experimental group and stretched out long neurites. And the NGF protein could maintain the neural stem cell survival and stimulate their differentiation. These findings suggest that the neural stem cells carrying extrinsic source NGF gene could express NGF successfully, and the NGF protein induced the survival and differentiation 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


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    Biological axis of CXC chemokine ligand and expression of stem cell markers in human colorectal cancer tissues as well as their relationship with clinical and pathological characteristics
    Huang Zhe
    2014, 18 (19):  3105-3110.  doi: 10.3969/j.issn.2095-4344.2014.19.025
    Abstract ( 336 )   PDF (635KB) ( 395 )   Save

    BACKGROUND: The biological axis of CXC chemokine ligand 12 (CXCL12)/CXC chemokine receptor 4 (CXCR4) plays an important role in the specific metastasis of tumors, whereas the mRNA expression of glycoprotein hormone receptor 5, a stem cell marker, exerts an important effects on tumor proliferation, invasion and metastasis.
    OBJECTIVE: To investigate the expressions of CXCL12/CXCR4 and glycoprotein hormone receptor 5 in human colorectal cancer tissues as well as their relationship with clinical and pathological characteristics.
    METHODS: Totally 100 colorectal cancer patients admitted at Liaoning Cancer Hospital & Institute from January to June 2013 were enrolled as experimental group, and another 100 healthy volunteers served as control group. Immunohistochemical SP method was used to detect the expression of CXCL12, CXCR4 and glycoprotein hormone receptor 5 mRNA in human colorectal cancer tissues. We analyzed the correlation of CXCL12, CXCR4 and glycoprotein hormone receptor 5 mRNA expression with colorectal cancer patients’ age, sex, tumor size and site, lymphatic metastasis and prognosis.
    RESULTS AND CONCLUSION: There was a higher expression of CXCR4 and glycoprotein hormone receptor 5 mRNA, but lower expression of CXCL12 mRNA in the human colorectal cancer tissues. The mRNA expression of CXCR4, CXCL12 and glycoprotein hormone receptor 5 showed no correlation with age, gender, tumor size and site of colorectal cancer patients, but was related to lymphatic metastasis. For colorectal cancer patients with lymphatic metastasis, the expression of CXCR4 and glycoprotein hormone receptor 5 mRNA was higher, but no change was found in the expression of CXCL12 mRNA. The expression of CXCR4 was increased with the degree of tumor malignancy as well as glycoprotein hormone receptor 5 expressed on the surface of gastrointestinal cancer and brain tumor stem cells. These findings indicate that the growth and metastasis of human colorectal cancer tissues are promoted by increasing the expression of CXCR4 and glycoprotein hormone receptor 5 in human colorectal cancer tissues; the CXCL12/CXCR4 biological axis and glycoprotein hormone receptor 5 are expected to become a new target therapy for tumor diagnosis and 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


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    Relationship between serum levels of stem cell factors and uremic pruritus in maintenance hemodialysis patients
    Qiu Fang-xin, Song Lei, Su Jun, Liu Xue-mei
    2014, 18 (19):  3111-3116.  doi: 10.3969/j.issn.2095-4344.2014.19.026
    Abstract ( 434 )   PDF (719KB) ( 402 )   Save

    BACKGROUND: Currently the pathogenesis of skin lesions in chronic renal failure patients has not yet been fully elucidated. Stem cell factor is a multifunctional cytokine, which may play a crucial role in the process of complications of chronic kidney disease. Changes related to stem cell factor levels in the peripheral blood of maintenance hemodialysis patients have been rarely reported.
    OBJECTIVE: To investigate the changes of serum stem cell factor levels in maintenance hemodialysis patients and their relationships with uremic pruritus.
    METHODS: Serum levels of stem cell factors in 86 maintenance hemodialysis patients were detected by enzyme-linked immunosorbent assay and compared with statistical methods. Visual analogue scale was to assess the severity of pruritus in patients with uremia. Then based on the scores, patients were divided into four groups: group A, 0-2 scores (n=23); group B, 3-5 scores (n=21); group C, 6-8 scores (n=24); and group D, > 8 scores (n=18). The differences in serum stem cell factor levels among the four groups were determined by Fisher’s least significant difference test. The correlation between stem cell factor levels and the severity of pruritus, blood hemoglobin and intact parathyroid hormone were analyzed by Pearson correlation analysis.
    RESULTS AND CONCLUSION: Gender, age, body mass index and blood pressure were all not statistically different among different groups (P > 0.05). With the progression of uremic pruritus, serum levels of stem cell factor in the peripheral blood increased (P < 0.05), blood hemoglobin levels gradually decreased (P < 0.05), and levels of intact parathyroid hormone increased (P < 0.05). Serum levels of stem cell factors in the peripheral blood was negatively related to blood hemoglobin (r=-0.60, P < 0.01), but positively correlated to intact parathyroid hormone (r=0.70, P < 0.01). These findings indicate that serum levels of stem cell factors in the peripheral blood may play an important role in the occurrence and development of uremic pruritus.


    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


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