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28 November 2017, Volume 21 Issue 33 Previous Issue    Next Issue

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Tetracycline hydrochloride promotes rat bone marrow mesenchymal stem cells proliferation in vitro Zhang Jue, Xue Si-liang, Luo Yuan, Zhi Wei 2017, 21 (33):  5249-5254.  doi: 10.3969/j.issn.2095-4344.2017.33.001 Abstract ( 276 )   PDF (1327KB) ( 304 )   Save BACKGROUND: Tetracycline hydrochloride is shown to promote viability and proliferation of human embryonic osteoblasts cultured in vitro, increase the production of type I collage and osteocalcin. OBJECTIVE: To study the proliferation activity of bone marrow mesenchymal stem cells (BMSCs) cultured by tetracycline hydrochloride in vitro. METHODS: Passage 3 BMSCs from Sprague-Dawley rats were cultured and divided into two groups: cells in control group were cultured in complete medium containing fetal bovine serum, and those in experimental group were cultured in complete medium containing tetracycline hydrochloride. After days 1-8 of culture, MTT and cell counting assay were used to detect cell viability and proliferation in the two groups. RESULTS AND CONCLUSION: (1) Cell counting assay: with time, the number of cells in both groups was increased, but the cell number in the experimental group was higher than that in the control group at days 4-8 of culture (P < 0.05). (2) MTT assay: With time, the number of cells in both groups was increased: incubation period was at day 1, logarithmic growth period was within days 3-6, and plateau period began at day 7. Then, the cells continued to proliferate, and the cell proliferation was higher in the experimental group than the control group at days 5-8 of culture (P < 0.05). To conclude, tetracycline hydrochloride can significantly enhance BMSCs proliferation. 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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Effect of low-temperature preparation on the biological characteristics of adipose-derived mesenchymal stem cells Zhang Feng-li, Shao Xiao-hu, Ren Huai-juan, Chen Yan-tian, Qi Nian-min 2017, 21 (33):  5255-5261.  doi: 10.3969/j.issn.2095-4344.2017.33.002 Abstract ( 276 )   PDF (1705KB) ( 257 )   Save BACKGROUND: Adipose-derived mesenchymal stem cells (ADSCs) that are manufactured in good manufacturing practice (GMP) clean rooms should be made into stem cell preparations before administration. Low-temperature preparation has many advantages over cryopreservation preparation; however, little is reported on the effect of short-term low-temperature storage on the biological characteristics of stem cells. OBJECTIVE: To evaluate the effect of 24-hour low-temperature storage using multiple electrolytes containing 5%  human serum albumin on the biological characteristics of ADSCs. METHODS: ADSCs at passages 3-6 at a concentration of 5×109/L were suspended in multiple electrolytes containing 5% human serum albumin. Cell suspension was transferred into cryogenic vials, and then these vials were placed in a cold chain shipping box for 2-8 ℃ low-temperature storage for 24 hours. Cell morphology, adhesion ability, cell viability, cell diameters and cell immunophenotyping before and after the storage were observed. RESULTS AND CONCLUSION: (1) After low-temperature storage of ADSCs for 24 hours, the number of dead cells increased. Although cell viability decreased significantly, it was still higher than 80%. Cell diameters of living cells increased significantly. (2) After low-temperature storage of ADSCs for 24 hours, few cells which were circle-shaped lost adhesion ability, and most cells could adherently grow, with the spindle-shaped morphology similar to the cells before preservation. (3) After low-temperature storage of ADSCs for 24 hours, HLA-DR, CD34 and CD45 were negatively expressed with a positive rate lower than 2%; CD29, CD73 and CD105 were positively expressed with a positive rate higher than 95%. However, the cell cluster was clearly divided into two parts after the preservation. Cells with enlarged diameters moved right in the FSC/SSC dot-plot. These results show that low-temperature preparation storage has no significant effect on the stemness of ADSCs, such as adhesion ability, cell viability and cell immunophenotype. 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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Effect of Compound Jiegu Tablets and Zhuanggu Tablets on thedifferentiation of bone marrow mesenchymal stem cells in vitro Wu Bo, Liu Ye, Ma Xu, Liu Su-yuan, Wang Yi-han, Wang Jia-yuan, Tan Cheng-bo, Yang Zheng-bo 2017, 21 (33):  5262-5267.  doi: 10.3969/j.issn.2095-4344.2017.33.003 Abstract ( 258 )   PDF (5345KB) ( 232 )   Save BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) are a kind of pluripotent stem cells in the bone marrow. Drug-containing serum treatment may be followed by a variation in the expression of genes involved in multiple signaling pathways. OBJECTIVE: To study the effect of compound Jiegu Tablets and Zhuanggu Tablets on the differentiation of BMSCs in vitro.  METHODS: Sixty Sprague-Dawley rats were randomized into Jiegu group, Zhuanggu group, combined group and blank control group. Different drug-containing sera were prepared and used to culture BMSCs isolated from the rat bone marrow. RT-PCR and western blot assay were used to detect the expression of BMP-2, Runx2, VEGF and Akt at mRNA and protein levels, respectively, at 24 hours after culture. RESULTS AND CONCLUSION: Compared with the blank control group, the expression levels of Akt and VEGF mRNA and protein were significantly increased in the Jiegu group and combined group (P < 0.05 or P < 0.01), but showed no difference in the Zhuanggu group (P > 0.05). Compared with the blank control group, the expression levels of BMP-2 and Runx2 mRNA and protein were significantly increased in the Jiegu group (P < 0.05) and very significantly increased in the Zhuanggu group and combined group (P < 0.01). These findings indicate that compound Jiegu Tablets and Zhuanggu Tablets containing sera lead to synergistic effects on theBMSCs differentiation into vascular endothelial cells and osteoblasts in vitro.  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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Co-transplantation of bone marrow mesenchymal stem cells and human umbilical vein endothelial cells transfected with vascular endothelial growth factor 165 gene promotes vascularization Xu Rong-sheng, He Yun, Li Biao, Xian De-bin, Xia De-lin 2017, 21 (33):  5268-5273.  doi: 10.3969/j.issn.2095-4344.2017.33.004 Abstract ( 291 )   PDF (5336KB) ( 243 )   Save BACKGROUND: Vascularized bone tissue engineering is a hotspot of current research and early vascularization of tissue-engineered bone also becomes an urgent problem to be solved. Addition of bioactive factors and cell co-culture method both contribute to promoting the early vascularization.  OBJECTIVE: To investigate the pro-angiogenic effect of human umbilical vein endothelial cells (HUVECs) transfected with vascular endothelial growth factor 165 (VEGF165) co-cultured with bone marrow mesenchymal stem cells (BMSCs).  METHODS: HUVECs were transfected with VEGF165 and then co-cultured with BMSCs which were purified from mouse femoral bone marrow. There were six groups in this experiment: (1) AdVEGF165-HUVECs+BMSCs, (2) AdVEGF165-HUVECs, (3) AdGFP-HUVECs+BMSCs, (4) AdGFP-HUVECs, (5) HUVECs+BMSCs, and (6) HUVECs (blank control). Cell counting kit-8 detection was applied to analyze the proliferative ability of HUVECs. The abilities of HUVECs migration and vascularization were then detected by crystal violet staining and matrigel determination, respectively.  RESULTS AND CONCLUSION: Compared with the blank control group, the proliferation of HUVECs was significantly increased in the other groups except for AdGFP-HUVECs group (P < 0.05). Compared with the blank control group, the migration and vascularization abilities of HUVECs were significantly stronger in the group of AdVEGF-HUVECs+BMSCs (P < 0.05). To conclude, the co-culture of HUVECs transfected with VEGF165 and BMSCs could promote early vascularization effectively. 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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Biocompatibility of bone marrow stromal stem cells with vascular endothelial growth factor/bone morphogenetic protein 2-poly(lactic-co-glycolic acid) copolymer/gelatin nanofibrous scaffold An Gang, Lu Bin, Zhang Wen-bo, Jiang Yu-dong 2017, 21 (33):  5274-5279.  doi: 10.3969/j.issn.2095-4344.2017.33.005 Abstract ( 260 )   PDF (6781KB) ( 270 )   Save BACKGROUND: Using bone tissue engineering methods for reconstruction of bone repair is an ideal treatment for bone defects, and it is crucial to combine biological scaffolds carrying growth factors with seed cells. OBJECTIVE: To observe the biocompatibility of rat bone marrow stromal stem cells (BMSCs) seeded onto vascular endothelial growth factor (VEGF)/bone morphogenetic protein 2 (BMP-2)-poly(lactic-co-glycolic acid) (PLGA) copolymer/ gelatin nanofibrous scaffolds. METHODS: Rat BMSCs were divided into five groups: blank scaffold group, BMP-2 group, VEGF group, VEGF/BMP-2 groups (1.2:1, 0.8:1, 0.4:1). In each group, the nanofibrous scaffold was placed at the bottom of the cell culture plate, and then rat BMSCs were seeded into the culture plate. Cell adhesion was observed under scanning electron microscope; cell proliferation was detected by cell counting kit-8; and ALP, RUNX-2 and OCN expression was determined by RT-PCR. RESULTS AND CONCLUSION: Under the scanning electron microscope, the BMSCs adhered to the scaffold and further grew into the scaffold. The adherent cells grew best in the 0.4:1 group, in which the cells were tightly integrated with the scaffold to form a cell-scaffold complex. Compared with the blank scaffold group, the cell proliferation and expression of ALP, RUNX-2 and OCN were both higher in the other groups, which was highest in the 0.4:1 group. To conclude, the VEGF/BMP-2-loaded PLGA copolymer/gelatin nanofibrous scaffold can promote cell adhesion, proliferation, osteogenic differentiation of rat BMSCs because of a good cytocompatibility. Moreover, the concentration ratio of VEGF/BMP-2 is optimized at 0.4:1.  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 differentiation of rat adipose-derived mesenchymal stem cells induced by rat lung epithelial-T-antigen negative cell line Chen Shang-ya, Cui Guan-qun, Bo Cun-xiang, Zhang Yu, Zhang En-guo, Yang Ye, Du Zhong-jun, Shao Hua 2017, 21 (33):  5280-5286.  doi: 10.3969/j.issn.2095-4344.2017.33.006 Abstract ( 360 )   PDF (5087KB) ( 247 )   Save BACKGROUND: Studies have shown that bone marrow mesenchymal stem cells have the potential of differentiation into alveolar epithelial cells in vitro, but so far no study has indicated that adipose-derived mesenchymal stem cells (ADSCs) can be differentiated into alveolar epithelial cells through long-term Transwell co-culture. OBJECTIVE: To observe whether rat lung epithelial-T-antigen negative cell lines (RLE-6TN) can induce rat ADSCs to differentiate into type II alveolar epithelial cells by long-term Transwell co-culture.  METHODS: Three SPF health female Sprague-Dawley rats were used as donors to separate, extract, culture and identity ADSCs. The experimental group was subjected to the Transwell co-culture of ADSCs and RLE-6TN, while the control group was subjected to the culture of ADSCs alone. The morphological changes of ADSCs were observed by the inverted phase contrast microscope at 21 days after co-culture. Immunofluorescence staining using surfactant protein C (SP-C) was performed on the co-cultured ADSCs. The fluorescence staining was observed using the inverted fluorescence microscope. Integral optical density (IOD) analysis was conducted by Image pro plus 6.0 software.  RESULTS AND CONCLUSION: RLE-6TN cells were identified by fluorescence staining with stable expression of SP-C protein (red fluorescence) in the experimental group, and there was no red fluorescence in the control group. After 21-day co-culture, the cell shape in the experimental group was transformed from the long spindle shape into oval or polygon shape gradually, while the cell shape in the control group remained fibroblast-like. These results show that RLE-6TN can induce ADSCs to differentiate into type II alveolar epithelial cells after a long-term (21 days) co-culture.  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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Pregnancy influences the proliferation and hepatic differentiation of rat adipose-derived stem cells Li Jun-nan, Chen Jia-yong, Zhang Yi, Lu Xiao-tao 2017, 21 (33):  5287-5292.  doi: 10.3969/j.issn.2095-4344.2017.33.007 Abstract ( 265 )   PDF (4995KB) ( 220 )   Save BACKGROUND: Pregnancy exhibits a significant association with stem cell activity. However, there is yet no report on the effects of pregnancy on adipose-derived stem cells. OBJECTIVE: To investigate the effect of pregnancy on the proliferation and differentiation of rat adipose-derived stem cells into hepatocyte-like cells.  METHODS: Adipose-derived stem cells were isolated and cultured from normal and pregnant Sprague-Dawley rats, and identified. The cell proliferation was detected by cell counting kit-8. All harvested adipose-derived stem cells were cultured in hepatocyte-induced medium and induced to differentiate into hepatocyte-like cells. Then the expressions of hepatocyte markers were detected at 0, 7, 14, 21 days of induction by fluorescent quantitative PCR; albumin secretion and urea synthesis were detected by using ELISA method. RESULTS AND CONCLUSION: Adipose-derived stem cells were successfully isolated from normal and pregnant rats, and CD45 negative staining and CD44/CD90 positive staining were observed, which was consistent with the characteristics of stem cells. Compared to the normal cells, adipose-derived stem cells from pregnant rats showed faster proliferation as well as osteogenic, adipogenic and hepatocyte-like differentiation, with the better liver function of hepatocyte-like cells. To conclude, pregnancy can promote the proliferation and differentiation of adipose-derived stem cells, which may be related to the estrogen level.   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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Liver and cardiac iron overload is harmful to HLA-identical hematopoietic stem cell transplantation in thalassemia children: an MRI detection Yang Wen-jing, Liao Jian-yun, Wen Jian-yun, Ruan Yong-sheng, Chen Li-bai, He Yue-lin, Li Chun-fu,Wu Xue-dong 2017, 21 (33):  5293-5298.  doi: 10.3969/j.issn.2095-4344.2017.33.008 Abstract ( 290 )   PDF (1084KB) ( 229 )   Save BACKGROUND: The majority of children with β-thalassemia major have iron overload, and iron overload may have negative effects on hematopoietic stem cell transplantation. OBJECTIVE: To assess the effects of liver and cardiac iron overload detected by magnetic resonance imaging (MRI) T2* on HLA-identical allogeneic hematopoietic stem cell transplantation in children with β-thalassemia major. METHODS: Eighty-one children with β-thalassemia major who were over 3 years of age and could cooperate with MRI detection were subjected to liver and heart MRI T2* tests before or after HLA-identical allogeneic hematopoietic stem cell transplantation. According to the test results, we calculated the liver and cardiac iron content, defined as an indicator of liver and heart iron overload. Then, there was a correlation analysis between the liver and cardiac iron content and serum ferritin, time of hematopoietic reconstitution, mortality rate, implantation rate and the morbidity of transplantation related complications, such as graft-versus-host disease, infections, autoimmune hemolysis, pancytopenia, hepatic veno-occlusive disease, septicemia. RESULTS AND CONCLUSION: The liver iron content was positively correlated with the time of hemoglobin implantation (r=0.229, P=0.043), and the cardiac iron content were positively correlated with the mortality rate (r=0.266, P=0.017); the serum ferritin level was negatively correlated with the implantation rate (r=-0.289, P=0.009), and positively correlated with the morbidity of septicemia (r=0.251, P=0.024) and pancytopenia (r=0.276, P=0.013). Therefore, iron overload exerts negative effects on HLA-identical allogeneic hematopoietic stem cell transplantation in β-thalassemia major children, and it is necessary to detect serum ferritin level and assess liver and cardiac iron overload before cell 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 Figures and Tables | References | Related Articles | Metrics

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Behavior improvement and inflammatory regulation in Parkinson’s disease rats after neural stem cell transplantation Zhou Jing, Bao Li-wen, Liang Jing 2017, 21 (33):  5299-5304.  doi: 10.3969/j.issn.2095-4344.2017.33.009 Abstract ( 352 )   PDF (1035KB) ( 242 )   Save BACKGROUND: Stem cell transplantation can inhibit the loss of dopaminergic neurons and improve behavioral symptoms in an animal model of Parkinson’s disease, which has a certain therapeutic effect. OBJECTIVE: To observe the changes of behavior symptoms and inflammatory factors in Parkinson’s disease rats after neural stem cell transplantation. METHODS: Totally 108 Sprague-Dawley rats were randomly divided into normal control group, model group and stem cell transplantation group, 36 rats in each group. In the model and stem cell transplantation groups, 6-hydroxydopamine was injected into the striatum of rats to establish Parkinson’s disease models, immediately followed by normal saline (5 μL) and 1×109/L neural stem cell suspension (5 μL), respectively. Six rats from each group was taken at each time point (1, 2, 4 weeks after cell transplantation) and subjected to intraperitoneal injection of 0.5 mg/kg apomorphine, followed by a 5-minute rotation test at 10 minutes after injection. ELISA was used to detect the levels of tumor necrosis factor α, interleukin-1β, interferon γ and interleukin-4 in the brain. RESULTS AND CONCLUSION: Rats in the normal control group had no rotation, while those in the stem cell transplantation and model groups presented with rotation behaviors. Moreover, the number of rotations was dramatically reduced in the stem cell transplantation group at 2 weeks after transplantation, which was significantly lower than that in the model group at 2 and 4 weeks after transplantation (P < 0.05). In both model and stem cell transplantation groups, the number of rotations was reduced significantly at 2 and 4 weeks as compared with that at 1 week after transplantation (P < 0.05). Compared with the normal control group, the levels of tumor necrosis factor α and interleukin-1β were significantly increased in the model group at different time after cell transplantation (P < 0.05); compared with the model group, these levels were significantly decreased at different time after stem cell transplantation (P < 0.05). Compared with the normal control group, the levels of interferon γ and interleukin-4 were significantly increased in the model group at different time after transplantation; compared with the model group, these levels were significantly increased at 1 and 4 weeks after stem cell transplantation (P < 0.05). To conclude, neural stem cell transplantation can improve the behavior symptoms of Parkinson’s disease rats, decrease the levels of tumor necrosis factor α and interleukin 1β (pro-inflammatory factors), and increase the levels of interferon γ and interleukin-4 (anti-inflammatory factors).   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-15b suppresses glioma stem cell migration and invasion by targeting ABCG2 signaling pathway Liu Yi-feng, Zhang Bao-chao, Wen Chang-ming, Wen Gong-ling, Zhou Guo-ping, Zhang Jing-wei, He Hai-fa, Wang Ning, Li Wei 2017, 21 (33):  5305-5312.  doi: 10.3969/j.issn.2095-4344.2017.33.010 Abstract ( 255 )   PDF (7511KB) ( 255 )   Save BACKGROUND: miR-15b plays an important role in the initiation and development of tumors, based on which, we speculate that miR-15b may be involved in the migration and invasion of glioma stem cells (GSCs). However, there is no relevant report and the mechanism of action is also unclear. OBJECTIVE: To identify the effects of miR-15b on the migration and invasion of GSCs and the mechanisms involved in this process.  METHODS: Quantitative PCR was performed to evaluate the expression of miR-15b in the gliomas tissues, normal brain tissues, GSCs and non-GSCs. After knockdown of ATP-binding cassette superfamily G member 2 (ABCG2) by ABCG2 specific siRNA, Transwell assay was performed to determine the effect of ABCG2 on GSCs migration and invasion. Additionally, the GSCs were transfected with miR-15b mimics or inhibitor to up-regulate or down-regulate the expression of miR-15b. At 48 hours after transfection, Transwell assay was used to detect the effect of miR-15b on GSCs migration and invasion; ELISA and gelatin zymography assays were performed to determine the matrix metalloproteinase-2/-9 (MMP-2/-9) expression and activity after treatment with miR-15b. CD133-positive or non-CD133-positive cells were directly injected subcutaneously into nude mice. Tumor formation was observed within 30 days after injection.  RESULTS AND CONCLUSION: miR-15b was significantly down-regulated in gliomas tissues compared with normal brain tissues, which was negatively correlated with the stage of gliomas. In addition, miR-15b was significantly down-regulated in GSCs compared with non-GSCs. Up-regulation of miR-15b significantly reduced the migration and invasion ability of GSCs (P < 0.01), and down-regulation of miR-15b significantly enhanced the cell migration and invasion of GSCs (P < 0.01). By target prediction analysis, we obtained that ABCG2 was a potential target gene of miR-15b. Luciferase assay confirmed that miR-15b targeted ABCG2 directly, and migration and invasion of GSCs were dramatically reduced by ABCG2 siRNA (P < 0.01). ELISA results showed that up-regulation miR-15b significantly inhibited MMP-2/-9 expression. ELISA and gelatin zymography assay results showed that ABCG2 siRNA did not affect MMP-2/-9 expression, but significantly inhibited the activity of MMP-2/-9. In the in vivo tumor model, GSCs were more tumorigenic as compared with non-GSCs from the same tumor in vivo. To conclude, miR-15b regulates the migration and invasion of GSCs by targeting the ABCG2 signaling pathway, and up-regulation of miR-15b can suppress the migration and invasion of GSCs.   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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Long-term serum-free culture of cancer stem/progenitor cells from basal-like breast carcinoma Dong Hua-ying, Wang Wei, Chen Yuan-wen, Bao Jun-jie, Chen Xin, Wu Cheng-yi 2017, 21 (33):  5313-5319.  doi: 10.3969/j.issn.2095-4344.2017.33.011 Abstract ( 300 )   PDF (4877KB) ( 251 )   Save BACKGROUND: Breast cancer stem cells, a subgroup of breast cancer cells, have self-renewal and multilineage differentiation potential and are characterized as resistance to chemotherapy, radiotherapy and hypoxia, and high propensity for invasiveness and metastasis, which play an important role in the recurrence and metastasis of breast cancer. The breast cancer stem cell theory can elucidate the pathogenesis of breast cancer to a certain extent, and has great values in the selection of treatment options, the discovery of new therapeutic targets and the prognosis of breast cancer. OBJECTIVE: To isolate the breast cancer stem/progenitor cells from basal-like breast carcinoma and study their self-renewal ability, phenotype and differentiation potential over long-term serum-free suspension culture in vitro.  METHODS: Twenty patients with basal-like breast cancer, including 10 patients without any treatment and 10 with neoadjuvant chemotherapy (≥ 4 cycles), were enrolled. A 1 cm3 fresh tumor tissue sample from each patient was taken and immediately sent to the laboratory to mechanically isolate tumor cells. Then, the stem/progenitor cells of basal-like breast carcinoma were enriched in ultra low attachment plates in serum free media as nonadherent mammospheres. Serial sphere formation assay was performed to determine colony formation and self-renewal ability of mammosphere-derived cells. Differentiation was induced by culturing mammosphere-derived cells in DMEM-F12 supplemented with serum but without growth factors. The proportion of CD44+/CD24- and ALDH1+ cell population was evaluated by flow cytometry.  RESULTS AND CONCLUSION: The mammospheres formed after inoculation of primary basal-like breast cancer cells isolated from the tumor tissues of breast cancer patients who did not receive neoadjuvant chemotherapy cultured in the serum-free medium with growth factors, while the mammospheres could be directly isolated from the tumor specimens of patients with neoadjuvant chemotherapy. The mammosphere-derived cells could be passaged continuously to form new mammospheres. With the increase of subculture frequency, the number of adherent cells was increased, but the number of new mammospheres decreased, and the spheres became smaller and easier to adhere. The mammosphere-derived cells could be induced to differentiate in the medium supplemented with serum. The CD44+/CD24- and ALDH1+ cells were enriched in mammosphere-derived cells, and these two phenotypic cells were decreased sharply in number or absent after cell differentiation. Most of the mammosphere-derived cells after chemotherapy had stronger potentials of self-renewal and differentiation, with higher proportion of CD44+/CD24- and ALDH1+ cells. In summary, cancer cells with self-renewal and multilineage differentiation potentials exist in the basal-like breast cancer tissues. Neoadjuvant chemotherapy can enrich breast cancer stem/progenitor cells to form spheroid-like cell clusters. Some differences in biological behaviors exist between the stem/progenitor cells from different breast cancer samples, which can vary with environmental factors.   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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Umbilical cord mesenchymal stem cell transplantation increases microvessel density in surrounding areas of acute traumatic brain injury in rats Xu Hai-huan, Dong Hua-jiang, Shang Chong-zhi, Zhao Ming-liang 2017, 21 (33):  5320-5324.  doi: 10.3969/j.issn.2095-4344.2017.33.012 Abstract ( 280 )   PDF (4448KB) ( 270 )   Save BACKGROUND: Umbilical cord mesenchymal stem cells (UC-MSCs) are a kind of pluripotent stem cells capable of relieving inflammation and edema in injured areas, promoting microvascular regeneration and improving wound healing. OBJECTIVE: To explore the therapeutic effects of UC-MSCs transplantation on acute traumatic brain injury (TBI) in rats and the effect on microvessel density of the rat brain tissue.  METHODS: Thirty Sprague Dawley rats were randomly divided into three groups, control group, TBI model group and UC-MSCs treatment group (n=10 per group). TBI models were made in the rats by free fall method. US-MSCs (1.5×106) were infused through the cerebral ventricle in the rats. Neurological severity scores were assessed at 24 hours after cell transplantation. The microvessel density in the injured brain tissue was detected using immunohistochemistry method at 14 days after cell transplantation.  RESULTS AND CONCLUSION: Compared with the TBI model group, the microvessel density value was significantly increased after UC-MSCs transplantation (P < 0.05), and the neurological severity score also became better in the UC-MSCs treatment group (P < 0.05). In summary, UC-MSCs transplantation can effectively improve became vascular remodeling and have a good neuroprotection in acute TBI 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 Figures and Tables | References | Related Articles | Metrics

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Therapeutic effect of rosiglitazone combined with adipose-derived stem cell transplantation in rats with type 2 diabetes mellitus Kang Li-li, Tian Meng 2017, 21 (33):  5325-5331.  doi: 10.3969/j.issn.2095-4344.2017.33.013 Abstract ( 318 )   PDF (1380KB) ( 241 )   Save BACKGROUND: Studies have shown that high glucose environment has a significant inhibitory effect on the survival of adipose-derived stem cells, which significantly affects the therapeutic efficacy of adipose-derived stem cells on type 2 diabetes mellitus. OBJECTIVE: To investigate the therapeutic effect of rosiglitazone combined with adipose-derived stem cell transplantation in rats with type 2 diabetes mellitus. METHODS: A total of 108 male Sprague-Dawley rats were chosen, 20 of which were randomly taken as normal controls and the rest 88 rats were used to make type 2 diabetes mellitus models through intraperitoneal injection of low-dose streptozotocin combined with high-sugar/high-fat diet. At 20 days after modeling, the model rats were randomized into model, rosiglitazone, cell transplantation and combined groups, followed by administration of normal saline (by gavage and via tail vein), rosiglitazone (by gavage), adipose-derived stem cells (via tail vein), and rosiglitazone (by gavage)+adipose-derived stem cells (via tail vein), once a day, for consecutive 7 days. Three weeks after treatment, fasting blood glucose level, insulin level, C-peptide level, body mass changes were detected; distribution and survival of adipose-derived stem cells were observed; the changes of pancreas histomorphology was observed, the apoptosis in islet cells was detected, and the CXCL1, CXCL2 and CXCR2 protein expression levels in the pancreas tissue were determined. RESULTS AND CONCLUSION: Compared with the normal control group, the fasting blood glucose level was increased (P < 0.05), while the insulin level, C-peptide level and body mass were significantly decreased in the model group (P < 0.05); the number of apoptotic islet cells was increased (P < 0.05); the expression levels of CXCL1, CXCL2 and CXCR2 in the pancreas tissue were elevated (P < 0.05); and the pancreas tissue morphologies were in disorder, with few islet cell masses. Compared with the model group, the fasting blood glucose level was decreased (P < 0.05), while the insulin level, C-peptide level and body mass were significantly increased in the rosiglitazone, cell transplantation and combined groups (P < 0.05); the number of apoptotic islet cells was decreased (P < 0.05); the expression levels of CXCL1, CXCL2 and CXCR2 in the pancreas tissue were declined (P < 0.05); and the pancreas tissue morphologies were improved to different extents, and the combined group showed the best improvement. There were many transplanted adipose-derived stem cells in the cell transplantation and combined groups, especially in the latter one. Overall findings indicate that rosiglitazone combined with adipose-derived stem cell transplantation is an effective treatment for type 2 diabetes mellitus in rats, which is better to inhibit inflammations.    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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Multiple imaging evaluation on the therapeutic efficacy of coronary artery bypass graft combined with autologous stem cell transplantation for myocardial infarction Lu Guo-xiu, Hao Shan-hu, Wang Zhi-guo, Zhang Tong, Wang Hui-shan, Zhang Guo-xu 2017, 21 (33):  5332-5338.  doi: 10.3969/j.issn.2095-4344.2017.33.014 Abstract ( 345 )   PDF (1229KB) ( 256 )   Save BACKGROUND: Stem cells are still controversial for the treatment of old myocardial infarction. Multimodal imaging evaluation is one of the key points in the study of stem cell transplantation, which can evaluate the therapeutic efficacy of stem cell transplantation from the perspective of molecular imaging. OBJECTIVE: To evaluate the therapeutic efficacy of coronary artery bypass graft (CABG) with different stem cell transplantation in patients with old myocardial infarction using multimodal imaging technology.  METHODS: Sixty patients with old myocardial infarction were enrolled and randomly divided into three groups to receive CABG, CABG+autologous bone marrow stem cell transplantation (CABG+BMC) or CABG+autologous peripheral blood stem cell transplantation (CABG+PBSC), respectively. All the patients were scanned with gated PET/CT (13N-NH3•H2O/18F-FDG), echocardiography and coronary angiography at different time points orderly (at baseline, 1, 12 and 24 months after treatment). We compared the degree of coronary stenosis (%), left ventricular ejection fraction (LVEF), percentage of defect size with myocardial perfusion/metabolic abnormal radioactive distribution (A) and the ratio of defect area (R).  RESULTS AND CONCLUSION: In the diagnosis of survival myocardial segments, the sensitivity, specificity, positive predictive value and negative predictive value for the gated PET/CT were 92.1%, 85.6%, 93.4% and 78.4%, respectively. After the above treatments, the extent of coronary stenosis decreased significantly in the three groups (P < 0.05), which was improved most at 1 month after treatment (P < 0.05). In the CABG+BMC and CABG+PBSC groups, the LVEF value increased significantly after treatment (P < 0.05). In the CABG+BMC group, the A value decreased significantly at 1 and 24 months after treatment as compared with the baseline (P < 0.05), and the A value was further decreased, indicating a significant difference at 12 and 24 months after treatment (P < 0.05). In the CABG+BMC group, the R value significantly decreased at 1 month after treatment compared with the baseline (P=0.019). To conclude, the multimodal imaging is better to evaluate the prognosis of patients undergoing CABG with different stem cell transplantation, which is beneficial for the selection of treatment and therapeutic evaluation in myocardial infarction patients. CABG combined with stem cell transplantation can improve the left ventricular function of patients in a short time, and CABG+BMC is superior to CABG+PBSC to improve the survived myocardial function in patients.    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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Cartilage repair using induced pluripotent stem cell derived chondrocytes in osteoarthriti Wu Zhang-song, Zhu Hong-xia, Luo Zhi-qiang, Wu Xiao-min, Lin Guang-yao, Zhang Ming-yu, Gu Hong-sheng,Zhu Yan-xia 2017, 21 (33):  5339-5347.  doi: 10.3969/j.issn.2095-4344.2017.33.015 Abstract ( 529 )   PDF (3710KB) ( 246 )   Save BACKGROUND: Stem cell-based therapy has been proposed for the treatment of osteoarthritis (OA) and induced pluripotent stem cells (iPSCs) are becoming a promising stem cell source as they have strong proliferation and differentiation potentials and no ethics problem.  OBJECTIVE: To explore an effective method of iPSCs differentiating into chondrocytes and to study the therapeutic effect of iPSCs derived chondrocytes on osteoarthritis. METHODS: In this study, three steps were developed to induce human iPSCs to differentiate into chondrocytes which were then transplanted into rat OA models induced by monosodium iodoacetate (MIA). There were four groups in the experiment: control group with normal saline injection, model group with MIA injection, iPSCs group with iPSCs transplantation following MIA injection, and differentiated iPSCs group with transplantation of iPSCs derived chondrocytes following MIA injection. At 15 weeks after transplantation, micro-CT was used and histological analysis of the knee joint was performed. RESULTS AND CONCLUSION: Compared with the iPS group, the expression of chondrocyte specific genes and proteins (Col2A1, GAG and Sox9) were significantly increased in the differentiated iPSCs group after 6 days of embryoid formation and after 2 weeks of cell differentiation. At 15 weeks after cell transplantation, no immune responses were observed, micro-CT showed an improvement in subchondral bone integrity, and histological examinations demonstrated the production of articular cartilage matrix. iPSCs derived chondrocytes showed better effects on articular cartilage repair than the iPSCs. To conclude, iPSCs derived chondrocytes can be effective via transplantation approach for cartilage tissue regeneration in OA 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 Figures and Tables | References | Related Articles | Metrics

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Biochemical changes in the serum and placenta of rats with gestational diabetes after transplantation of human placental mesenchymal stem cells Yang Zuo-feng, Wang Ying, Pei Ling-ling, Shen Xiao-tong, Chen Meng-ge, Zhao Jia, Zhang Nan, Liu Wei 2017, 21 (33):  5348-5353.  doi: 10.3969/j.issn.2095-4344.2017.33.016 Abstract ( 354 )   PDF (5467KB) ( 255 )   Save BACKGROUND: Human placental mesenchymal stem cells can improve the blood glucose level of diabetes mellitus rats and gestational diabetes rats, but little is reported on its effect on glucagon, adiponectin, and tumor necrosis factor-α in the serum and placental tissues. OBJECTIVE: To investigate the effects of human placental mesenchymal stem cells on the levels of glucagon, adiponectin and tumor necrosis factor-α in the serum and placental tissues in gestational diabetes rats. METHODS: A rat model of gestational diabetes was made by high-fat and high-sugar diet plus low-dose injection of streptozotocin. Passage 3 human placental mesenchymal stem cell suspension (1×1010 cells/L, 0.5 mL) was injected into gestational diabetes rats at gestational days 4 and 11 (4- and 11-day intervention groups). Meanwhile, control rats were given the same amount of normal saline. At 20 days of gestation, blood samples from the abdominal aorta were extracted, and then cesarean section was made to remove the placenta in the gestational diabetes rats. ELISA and real-time PCR were used to detect the levels of glucagon, adiponectin and tumor necrosis factor-α in the serum and placental tissues, respectively.  RESULTS AND CONCLUSION: (1) The serum and placental levels of glucagon, adiponectin and tumor necrosis factor-α showed no differences between the 4- and 11-day intervention groups (P > 0.05). (2) Compared with the control group, significantly increased serum adiponectin level and significantly decreased placental glucagon mRNA expression were found in the 4-day intervention group (P < 0.05). (3) Compared with the control group, the serum adiponectin level and the placental glucagon level both had a significant decrease in the 11-day intervention group (P < 0.01), while the serum level of tumor necrosis factor-α was significantly decreased (P < 0.01). To conclude, transplantation of human placental mesenchymal stem cells can vary the adiponectin and glucagon levels, which provides a new research idea and basis for the further study on the possible mechanism of placental mesenchymal stem cells to improve blood glucose level in gestational diabetes rats. Additionally, it is worthy while to notice that gestational diabetes rats given placental mesenchymal stem cells in the early or late pregnancy show no effects on the above indicators.   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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Dental pulp repair with dental pulp stem cells by construction of tissue-engineered dentin-pulp complex Wang Lu, Ma Xin, Han Yao-lun, Li Ya-dong, Luo Shu-wen 2017, 21 (33):  5354-5359.  doi: 10.3969/j.issn.2095-4344.2017.33.017 Abstract ( 301 )   PDF (5704KB) ( 256 )   Save BACKGROUND: Studies have shown that dental pulp stem cells have high proliferation and multi-directional differentiation abilities and can differentiate into a variety of cells under certain conditions. At present, the use of dental pulp stem cells to construct tissue-engineered dentin-pulp complex is expected to become a new strategy for human dental defect repair . OBJECTIVE: To observe the effect of dental pulp stem cells on the repair of rat tooth defects by construction of tissue-engineered dentin-pulp complex. METHODS: Twenty-four Sprague-Dawley rats were used to make animal models with dental pulp removal, and then model rats were randomly divided into model group and transplantation group. Rats in the transplantation group were subjected to tissue-engineered dentin-pulp complex transplantation, and those in the model group given no treatment. Tooth samples were collected at 3, 5, 7 weeks post transplantation and observed using hematoxylin-eosin staining and immunofluorescence staining. The dentin thickness of rats was measured by Image Pro Plus 6.0 image software system. RESULTS AND CONCLUSION: (1) Dental pulp cells was mostly spindle/oval-shaped and partially polygonal. The third generation of cells with long spindle shape showed fibrous growth and uniform morphology. Findings from immunohistochemical staining showed spindle-shaped deep-colored cells with oval nuclei stained as dark blue were identified as fibroblast-like cells, and were positire for vimtin. (2) Findings from hematoxylin-eosin staining showed vacuolar degeneration of the cells, and hbdestroyed pulp tissue and debris, irregular cord-like tissue, and a large amount of red blood cells and inflammatory cells in the pulp cavity, accompanied by clearly visible vascular dilation. Seven weeks after transplantation, a bundle of odontoblasts were visible in the matrix-like tissues of the dentin, and there was a distinct boundary between the original dentin and regenerated dentin. (3) Findings from immunofluorescent staining showed that after dentin-pulp complex transplantation, the number of cells in the pulp cavity increased significantly at 3 weeks, and there was also a substantial increase in dental pulp cells at 5 weeks that were distributed on the wall of the pulp cavity. Compared with the model group, the dentin thickness in the transplantation group was significantly higher at each time after transplantation (P < 0.05), and in the transplantation group, there was also a significant difference in the dentin thickness at different time points (P < 0.05). To conclude, the tissue-engineered dentin-pulp complex can promote dentin regeneration and repair.        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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Functional differentiation of dopaminergic neurons derived from human embryonic stem cells Peng Ya-nan, Hu Lan, Wang Tan, Li Ke, Yang Liu, Chen Li, Chen Xiao-wu, Chen Zhi-bin,Zhao Zhen-qiang 2017, 21 (33):  5360-5368.  doi: 10.3969/j.issn.2095-4344.2017.33.018 Abstract ( 506 )   PDF (2806KB) ( 372 )   Save BACKGROUND: The in vitro differentiation methods of stem cell-derived dopaminergic neurons that serve as a cell source for the replacement therapy of Parkinson’s disease are continuously optimized and improved, as well as the subsequent identification methods and testing indicators. OBJECTIVE: To observe the morphological development and electrophysiological characteristics of dopaminergic neurons differentiated from human embryonic stem cells so as to identify whether these differentiated cells have mature morphology and function under the current differentiation program.  METHODS: Monolayer adherent method combined with dual-SMAD signaling inhibition was used to induce the directional differentiation of human embryonic stem cells into dopaminergic neurons. Then the cells were identified by light microscopy, electron microscopy and immunofluorescence, and the electrophysiological properties of dopaminergic neurons were detected by patch clamp electrophysiological technique. Herein, we evaluated the electrophysiological functions of dopaminergic neurons differentiated in vitro, with reference to the evaluation standard of dopaminergic neuron in vivo. RESULTS AND CONCLUSION: In this study, we successfully obtained dopaminergic neurons with mature morphology and functions differentiated from human embryonic stem cells in vitro. Findings from the subsequent electrophysiological test confirmed that dopaminergic neurons we acquired had electrophysiological properties in accordance with the evaluation standards of dopaminergic neurons in vivo. To conclude, the monolayer adherent method combined with dual-SMAD signaling inhibition can successfully induce the directional differentiation of human embryonic stem cells into dopaminergic neurons with mature morphology and functions.    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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Protective role of serum-free supernatant of human placental fetal mesenchymal stem cells against oxidative stress injury to lung epithelial cells Fu Xue, Liu Guo-pan, Zhang Yu-jie, Yan Xiu-rui, Ma Xiao-na, Liu Xiao-ming, Wei Jun 2017, 21 (33):  5369-5374.  doi: 10.3969/j.issn.2095-4344.2017.33.019 Abstract ( 406 )   PDF (4678KB) ( 227 )   Save BACKGROUND: Preliminary experimental studies have shown that the supernatant of human placental fetal mesenchymal stem cells (fPMSCs) has a certain ability to scavenge reactive oxygen species and itself has a certain antioxidant enzyme activity. OBJECTIVE: To investigate the protective role and mechanism of fPMSCs supernatant in serum-free culture on oxidative stress-injured lung epithelial cells. METHODS: Different concentrations of hydrogen peroxide produced oxygen stimulation to lung epithelial cell lines A549 for 6, 12, 24 hours, and the survival rate of lung epithelial cells was detected using cell counting kit-8 method. When the survival rate of lung epithelial cells was 50%, the concentration of hydrogen peroxide was most suitable to make an oxidative damage model. The validity of the model was verified using Hocheest33258 staining and western blot. fPMSCs were cultured in serum-free culture medium, and the supernatant of passage 3 cells was collected. Afterwards, the injured lung epithelial cells were cultured in the fPMSCs cell supernatant for 24 hours. Meanwhile, injury group (oxidative damage only) and vitamin C group (100 μmol/L vitamin C was added in the medium) were established. In the three groups, cell apoptosis was detected by flow cytometry; and western blot was used to detect apoptosis-related proteins and proteins related to the Nrf2-Keap1-ARE signaling pathway. RESULTS AND CONCLUSION: After oxygen stimulation by 600 μmol/L hydrogen peroxide for 24 hours, the survival rate of A549 cells was (56.41±3.31)% as ascertained by the cell counting kit-8 assay. Findings from Hocheest33258 staining and western blot further confirmed the reliability of this model. Flow cytometry results showed that the apoptosis rate in the vitamin C group and the supernatant group decreased to some extent compared with the injury group, and the difference between the supernatant group and the injury group was statistically significant (P < 0.05). In addition, the expression of Bax significantly decreased and the expression of Bcl-2 significantly increased in the vitamin C group and the supernatant group as detected by western blot assay, in comparison with the injury group (P < 0.05). Compared with the injury group, the expression of Nrf2 protein increased and the expression of Keap1 decreased in the vitamin C group and the supernatant group (P < 0.05). These findings suggest that fPMSCs supernatant has a certain antioxidant capacity, and may attenuat oxidative damage and inhibit apoptosis in A549 cells. The mechanism is probably related to the Nrf2-Keap1-ARE 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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Hypoxic microenvironment promotes the proliferation of human olfactory mucosa mesenchymal stem cells and its associated mechanism Zhuo Yi, Li Xuan, Duan Da, Ge Li-te, Yuan Ting, Wu Pei, Wang Hao, Long Lang, Lu Ming 2017, 21 (33):  5375-5381.  doi: 10.3969/j.issn.2095-4344.2017.33.020 Abstract ( 398 )   PDF (6202KB) ( 212 )   Save BACKGROUND: Human olfactory mucosa mesenchymal stem cells (hOM-MSCs) not only have the basic characteristics of mesenchymal stem cells, but also originate from the ectoderm and are prone to differentiate into neurons, which are a kind of ideal seed cells for nerve repair and regeneration. Cells are conventionally cultured in about 21% in vitro, while only 3%-9% oxygen is found in the human body and tissue space. There is still no report on the effect of hypoxia on the proliferation and activity of hOM-MSCs. OBJECTIVE: To explore whether hypoxic microenvironment can promote hOM-MSCs proliferation and activity and the related mechanism.  METHODS: hOM-MSCs were isolated, cultured and identified by flow cytometry and immunofluorescence. The passage 4 hOM-MSCs were divided into three groups: 21% O2 group, 3% O2 group and 3% O2+20 µmol/L YC-1 (HIF-1α inhibitors) group. Proliferation and apoptosis of hOM-MSCs was detected by flow cytometry after 72 hours of culture. The proliferating cell nuclear antigen protein expression was detected by western blot. The mRNA and protein expression of HIF-1α and TWIST were detected by Q-PCR and western blot.  RESULTS AND CONCLUSION: The purity of hOM-MSCs was up to 97%, as defined by flow cytometry. The proliferation index of 3% O2 group was higher than the 21% O2 group (P < 0.05), and cell survival and apoptosis ratio (apoptotic cells included mechanical death + early apoptosis + late apoptosis) between the two groups had no significant difference (P > 0.05). Western blot results showed that the proliferating cell nuclear antigen protein expression in the 3% O2 group was significantly higher than that in the other groups (P < 0.05). The HIF-1α and TWIST expressions at mRNA and protein levels in the 3% O2 group were significantly higher than those in the other groups (P < 0.05). To conclude, hypoxic microenvironment can promote the hOM-MSCs proliferation and has no effect on the apoptosis, and the HIF-TWIST signal pathway plays an important role in this progress.   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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Enhanced tenogenic differentiation by Scleraxis overexpression in human amniotic mesenchymal stem cells Zhu Xi-zhong, Liu Zi-ming, Wu Shu-hong, Xiong Hua-zhang, Yang Ji-bin, Li Yu-wan, You Qi, Jin Ying,Zuo Chen, Liu Yi 2017, 21 (33):  5382-5387.  doi: 10.3969/j.issn.2095-4344.2017.33.021 Abstract ( 227 )   PDF (5474KB) ( 263 )   Save BACKGROUND: Human amniotic mesenchymal stem cells (hAMSCs) are adult stem cells with multipotential differentiation, which can be induced to differentiate into bone, cartilage and other connective tissues. Meanwhile, as a highly specific marker of tenocytes, Scleraxis is involved in aggregation and differentiation of tendon progenitor cells as well as the formation of tendon extracellular matrix. OBJECTIVE: To investigate whether hAMSCs have the ability of differentiation into tenocytes by ectopic expression of Scleraxis. METHODS: Agreed by puerpera, the amniotic membrane from the full-term placenta was separated, and hAMSCs were isolated by a two-step enzyme digestion, observed under inverted phase contrast microscope, and identified by flow cytometry. Passage 3 cells were induced via plasmid-mediated Scleraxis overexpression in overexpression group. Untransfected cells cultured in normal medium served as blank control group, and those with empty plasmid transfection were defined as empty plasmid group. Cell proliferation was tested in each group using cell counting kit-8 within 7 days of culture. Real-time quantitative PCR and western blot were used to assess the tenogenic differentiation of hAMSCs in each group at 3 and 7 days of culture. RESULTS AND CONCLUSION: Findings from the cell counting kit-8 indicated that the cell viability had no significant differences among the groups within 7 days of culture (P > 0.05). Western blot results showed the protein expression of Scleraxis in the treatment group was significantly higher than that in the other two groups (P < 0.05). Real-time PCR results showed, at 3 days of culture, the expression of collagen type I, collagen type III, Fibronectin and Tenascin-C in the overexpression group was significantly higher than that in the empty plasmid group (P < 0.05), but the expression of Tenomodulin had no difference (P > 0.05); at 7 days of culture, the expressions of collagen type I, collagen type III, Fibronectin, Tenascin-C and Tenomodulin in the overexpression group were significantly higher than those in the empty plasmid group (P < 0.05). In summary, hAMSCs can be differentiated into tenocytes by ectopic expression of Scleraxis.   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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Local administration of mononuclear cell, platelet-rich plasma and zoledronic acid for the prevention and treatment of early femoral head osteonecrosis and collapse: study protocol for a prospective, randomized, parallel, controlled clinical trial Ma Ning, Wang Hong-xia, Lu Qiang, Chen Si, Wan Yi-qun, Liu Ying-ying, Wang Xin, Peng Jiang, Guo Quan-yi 2017, 21 (33):  5388-5393.  doi: 10.3969/j.issn.2095-4344.2017.33.022 Abstract ( 359 )   PDF (1877KB) ( 201 )   Save BACKGROUND: There are various treatment methods for osteonecrosis of the femoral head (ONFH) and collapse, but conservative treatment is invalid. Once femoral head collapse occurs, the development is irreversible. Our previous research has shown that local administration of zoledronic acid can prevent necrotic femoral head collapse. Moreover, bone marrow mononuclear cells obtain satisfactory short-term efficacy in the treatment of ONFH. OBJECTIVE: To investigate the curative efficacy of local administration of mononuclear cell, platelet-rich plasma and zoledronic acid for the prevention and treatment of early ONFH and collapse. METHODS: This prospective, single-center, randomized, parallel, controlled clinical trial was conducted at the Chinese PLA General Hospital, Beijing, China. One hundred patients with ONFH (stages I-II by Ficat and Arlet classification) were enrolled and randomly assigned into either the treatment group or control group (n=50 per group). Patients were given an injection of mononuclear cell, platelet-rich plasma and zoledronic acid into the necrotic femoral head, or drilling decompression at the necrotic area. Patients in both groups were then followed up for 4, 8, 12, and 18 months. The primary outcome measures were the blood supply, osteogenesis and appearance of the necrotic femoral head observed on hip perfusion by dynamic MRI, CT restruction of the hip joint and radiography of the hip joint, as well as Harris hip scores and numerical rating scale scores. Secondary outcome measures included SF-36 Health Survey and Activities of Daily Living scores.  DISCUSSION: The outcomes of this trial have provided quantitative data for analyzing the effectiveness of local administration of mononuclear cell, platelet-rich plasma and zoledronic acid on ONFH and collapse. Written approval for this protocol was obtained from the Ethics Committee of the Chinese PLA General Hospital in China (approval No. S2015-082-01). Participants and their families are informed of the study protocol and procedures, and signed an informed consent. The study was in accordance with the guidelines of the Declaration of Helsinki, formulated by the World Medical Association. Trial began in January 2015 and will be completed in December 2017. Trial results will be published in scientific reports, or in peer-reviewed journals. This trial was registered with the ClinicalTrials.gov identifier: NCT02721940. Patient recruitment is ongoing.   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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The expression and effect of LINGO-1 during the differentiation of spinal cord derived neural stem cells Xu Gang, Zhao Chen-guang, Sun Wei, Ju Fen, Yuan Hua, Mou Xiang 2017, 21 (33):  5394-5399.  doi: 10.3969/j.issn.2095-4344.2017.33.023 Abstract ( 276 )   PDF (1531KB) ( 366 )   Save BACKGROUND: It is of great significance to explore the expression and effect of LINGO-1 in the differentiation of spinal cord derived neural stem cells (SpNSCs) for regulating neural stem cell differentiation and repairing spinal cord injury. OBJECTIVE: To investigate the expression features and biological effects of LINGO-1 in the differentiation of SpNSCs.  METHODS: SpNSCs were isolated from the rat spinal cord and cultured in vitro. The expression characteristics of LINGO-1 was observed through double immunofluorescence staining of LINGO-1 and Nestin (neural stem cells), β-Tubulin III (neurons), GFAP (astrocytes) and O4 (oligodendrocyte) at 0-5 days of differentiation. SpNSCs isolated from the rat spinal cord were cultured in vitro and divided into siRNA group and control group. The siRNA group was transfected with LINGO-1 shRNA lentiviral vector to down-regulate the expression of LINGO-1, and the control group was transfected with Scramble-shRNA lentiviral vector. The growth of neurites was detected by immunofluorescence staining at 5 days after transfection.  RESULTS AND CONCLUSION: The SpNSCs could differentiate into neurons, astrocytes and oligodendrocytes. LINGO-1 was expressed in SpNSCs, neurons and oligodendrocytes, but not in astrocytes. The neurite length of the siRNA group was significantly longer than that of the control group (P < 0.05). In summary, the SpNSCs have the potential of multi-directional differentiation, and LINGO-1 has a negative effect on the neurite growth.   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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The role of hypoxia induced factor-1alpha/apelin/APJ pathway in cardiac stem cell proliferation and cardiogenic differentiation after hypoxia preconditioning Wang Lei, Hou Jing-ying, Long Hui-bao, Wu Hao, Zhou Chang-qing, Guo Tian-zhu, Wu Quan-hua, Zhong Ting-ting, Chen Xu-xiang, Wang Tong 2017, 21 (33):  5400-5406.  doi: 10.3969/j.issn.2095-4344.2017.33.024 Abstract ( 448 )   PDF (1408KB) ( 377 )   Save BACKGROUND: Our previous studies demonstrated that cardiac stem cells (CSCs) transplantation could improve cardiac function in rats with myocardial infarction (MI). However, the overall survival and cardiac differentiation of CSCs were low.   OBJECTIVE: To investigate the effect of hypoxia preconditioning on CSCs proliferation and cardiogenic differentiation and the role of hypoxia induced factor-1alpha (HIF-1α)/apelin/putative receptor protein related to the angiotensin receptor AT1 (APJ) pathway in the procedure. METHODS: Cells cultured in vitro experienced exposure to hypoxia (1% O2) for 24 hours. Cardiogenic differentiation was induced by using 5-azacytidine for another 24 hours. Then, cells were cultured in normal condition for 2 weeks. Normoxia (20% O2) was used as a negative control during the whole process. Cell proliferation was detected using MTS method and expressions of HIF-1α, apelin, cTnT and APJ were detected using western blot assay after 24 hours of preconditioning and 2 weeks after the induction of differentiation; the percentage of cTnT-positive cardiomyocyte-like cells was observed by immunofluorescence staining. RESULTS AND CONCLUSION: Compared with the normoxia group, the hypoxia group presented a higher proliferation rate and a higher absorbance value at 490 nm (P < 0.01); the protein expressions of HIF-1α, apelin and APJ were all enhanced after hypoxia exposure for 24 hours and 2 weeks after the induction of differentiation (P < 0.01); the percentage of cTnT-positive cells was greatly increased in the hypoxia group (P < 0.01), and the expression of cTnT was also significantly intensified (P < 0.01). To conclude, hypoxia preconditioning could promote the proliferation and cardiogenic differentiation of CSCs, and the activation of HIF-1α/Apelin/APJ pathway might be involved in this process.   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 umbilical cord mesenchymal stem cell transplantation by different pathways and stem cell tracing by bioluminescence imaging in the treatment of ischemic brain injury Wang Li-xin, Li Hui-ping 2017, 21 (33):  5407-5412.  doi: 10.3969/j.issn.2095-4344.2017.33.025 Abstract ( 353 )   PDF (1183KB) ( 203 )   Save BACKGROUND: Transplantation of human umbilical cord mesenchymal stem cells can effectively promote the recovery of neurological function in animal models of ischemic brain injury. However, it is unclear on the mechanisms underlying the therapeutic actions of human umbilical cord mesenchymal stem cells and the optimal transplantation way. Bioluminescence imaging has been widely used in the field of stem cell transplantation for real-time dynamic monitoring of the proliferation, migration and survival of transplanted stem cells. OBJECTIVE: To review the studies of human umbilical cord mesenchymal stem cells in the treatment of ischemic brain injury and the recent research progress of bioluminescence imaging in the stem cell transplantation for the treatment of ischemic brain injury. METHODS: The key words were “human umbilical cord mesenchymal stem cells, brain ischemia, bioluminescence imaging” in English and Chinese, respectively. The first author retrieved PubMed database, Chinese Journal Full-text Database for relevant articles published from January 2011 to December 2016. Literatures with repetitive content and poor relationship were excluded. A total of 98 literatures were initially retrieved, and finally 38 articles met the inclusion criteria. RESULTS AND CONCLUSION: Human umbilical cord mesenchymal stem cells can be transplanted via the vein, artery, lumbar puncture, stereotactic operation, intraventricular approach in the treatment of ischemic brain injury. They have their own merits and demerits, which mainly focus on stem cell migration range, survival rate, safety and so on. It has been found that bioluminescence imaging technology for the living is characterized by real-time dynamic monitoring, high sensitivity, high-time resolution and easy operation, which can be used to monitor the migration, proliferation and survival of transplanted stem cells in vivo. Therefore, further investigation on the mechanism of human umbilical cord mesenchymal stem cells that are labeled by bioluminescence imaging technology and selection of the optimal transplantation approach is warranted.    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