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08 September 2018, Volume 22 Issue 25 Previous Issue    Next Issue

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Apelin combined with bone marrow mesenchymal stem cells transplantation improves cardiac function after myocardial infarction Hou Jing-ying, Long Hui-bao, Chen Xu-xiang, Wang Lei, Guo Tian-zhu, Zheng Shao-xin, Zhong Ting-ting, Wu Quan-hua, Wu Hao, Wang Tong 2018, 22 (25):  3937-3943.  doi: 10.3969/j.issn.2095-4344.0952 Abstract ( 350 )   PDF (898KB) ( 147 )   Save BACKGROUND: Our previous work demonstrated that apelin could promote bone marrow mesenchymal stem cells (BMSCs) survival and vascularization under hypoxic-ischemic conditions in vitro. OBJECTIVE: To explore the effect of apelin combined with BMSCs transplantation on cardiac function after myocardial infarction and the relevant mechanisms. METHODS: Fifty C57BL/6 mice with myocardial infarction were induced by the left anterior descending coronary artery ligation. Animal models were then randomized into five groups: BMSCs, BMSCs+apelin, sh-APJ BMSCs+apelin, apelin and PBS groups. Two weeks after modeling, the mice in the former four groups were subjected to secondary thoracotomy, BMSCs group was given intramyocardial injection of sole BMSCs. BMSCs+apelin and sh-APJ BMSCs+apelin groups were given intramyocardial injection of BMSCs and sh-APJ BMSCs respectively, in combination with intraperitoneal injection of apelin-13 for 2 weeks consecutively. Mice in the PBS group received intramyocardial injection of PBS without BMSCs. Cardiac function of mice was evaluated before and 2 weeks after treatment. MSCs survival and vascularization in the infarct border zone were examined, and the expression of relevant factors was detected thereafter. RESULTS AND CONCLUSION: (1) At 2 weeks after treatment, the cardiac function of mice was obviously improved in the former four groups in contrast with the baseline, especially in the BMSCs+apelin group; Left ventricular ejection fraction was significantly increased (P < 0.01), and left ventricular end diastolic diameter and left ventricular end systolic diameter were significantly reduced (P < 0.01). (2) Compared with BMSCs and sh-APJ BMSCs+apelin groups, the average optical density of PKH26- and CD31-positive BMSCs in the infarct border zone was significantly enhanced in the BMSCs+apelin group. Moreover, the percentage of CD31 and PKH26 double positive cells to the PKH26-positive BMSCs was considerably increased (P < 0.01). (3) The expression levels of APJ, AKT, pAKT and VEGFA were significantly increased in the BMSCs+apelin group compared with the BMSCs and sh-APJ BMSCs+apelin groups (P < 0.01). However, there was no difference in the above biomarkers between the BMSCs and sh-APJ BMSCs+apelin groups. It was concluded that apelin could combine with APJ to promote BMSCs survival and vascularization in the infarcted myocardial tissues, thereby further improving cardiac function post myocardial infarction. This effect was associated with the up-regulation of VEGFA. Activation of PI3K/AKT signaling pathway was likely to exert some functions in the above procedure. 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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Lentivirus-mediated human interleukin-12 fusion gene-transfected rat bone marrow mesenchymal stem cells inhibit CT26 tumor growth He Yang, Lin Chen, Gao Qin, Song Jing-xiang, Tu Xiao-huang 2018, 22 (25):  3944-3949.  doi: 10.3969/j.issn.2095-4344.0953 Abstract ( 307 )   PDF (797KB) ( 106 )   Save BACKGROUND: Interleukin-12 as the most potent anti-tumor factor has been a hot topic in research, but little is reported on its use in the treatment of colorectal cancer. OBJECTIVE: To construct rat bone marrow mesenchymal stem cell lines stably expressing human interleukin-12 (hIL-12-BMSCs), and to observe its effect on colorectal cancer (CT26). METHODS: Primers were designed and synthesized, and purified p40 and p35 gene fragments were amplified by PCR. The single-chain double-subunit fusion gene, hIL-12, was obtained by Overlap-ping PCR ligation, and 293T cells were co-transfected with lentiviral packaging system. Recombinant lenovirus overexpressing hIL-12 (LV-IL-12) was constructed to transfect rat BMSCs. The stable strain was then cultured via drug screening. Thirty-two nude mice were subcutaneously injected with CT26 cells to make animal models, and then randomly divided into four groups to receive peritumoral injection of hIL-12-BMSCs (0.2 mL; hIL-12-BMSCs group), 0.2 mL of PBS (PBS group), 0.2 mL of BMSCs (BMSCs group), or 0.2 mL of LV-IL-12 (LV-IL-12 group). Tumor growth was then statistically analyzed in each group. RESULTS AND CONCLUSION: There was no significant difference in tumor volume between the PBS and BMSCs groups (P > 0.05), indicating that BMSCs cannot promote or inhibit the growth of CT26 tumors. However, from the 7th day after injection, the tumor volume showed a significant difference between hIL-12-BMSCs and PBS groups as well as between PBS and LV-IL-12 groups (P < 0.01), indicating that Hil-12 has a significant inhibitory effect on CT26 tumor growth. From the 10th day after injection, a significant difference in the tumor volume was found between the hIL-12-BMSCs and LV-IL-12 groups (P < 0.05), indicating that hIL-12-BMSCs are more effective than LV-IL-12 in inhibiting the growth of CT26. To conclude, hIL-12-BMSCs considerably inhibit the growth of CT26 tumor. 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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Double transfection with lentivirus-mediated bone morphogenetic protein 2 and vascular endothelial growth factor 165 promotes osteogenesis of bone marrow mesenchymal stem cells Zhou Zhen-jie, Li Qiang, Li Shi-peng, Tao Xuan, Ma Yue-gang 2018, 22 (25):  3950-3955.  doi: 10.3969/j.issn.2095-4344.0954 Abstract ( 370 )   PDF (675KB) ( 167 )   Save BACKGROUND: Bone morphogenetic protein 2 (BMP-2) and vascular endothelial growth factor 165 (VEGF-165) are essential cytokines for bone repair. Lentiviral transfection of stem cells is important for studying the biological changes of doubly transfected cells. OBJECTIVE: To study the feasibility of human BMP-2 and human VEGF-165 dual gene transfection of bone marrow mesenchymal stem cells (BMSCs) and to explore the osteogenic capacity of doubly transfected cells. METHODS: Rabbit BMSCs were divided into four groups: untransfected group, blank vector group transfected with control lentivirus, BMP-2 group transfected with Lv-BMP-2/GFP, and BMP-2/VEGF-165 group transfected with Lv-BMP-2/GFP and Lv-VEGH-165/RFP. Expression of targeted proteins was detected using western blot at different time after transfection. MTT assay was used to detect cell proliferation in each group. Alkaline phosphatase activity was measured at 14 days after transfection. RESULTS AND CONCLUSION: (1) The corresponding green and red fluorescent protein expressions were observed under fluorescence microscope in the blank vector, BMP-2, and BMP-2/VEGF-165 groups. (2) The corresponding target proteins were highly expressed in the BMP-2 and BMP-2/VEGF-165 groups at 3 days after transfection, but no significant changes were detected at 7, 14, and 21 days after transfection (P > 0.05). (3) Compared with the BMP-2 group, the proliferation of cells was slightly faster in the BMP-2/VEGF-165 group (P < 0.05), but significantly lower in the untransfected group and blank vector group (P < 0.05). (4) The activity of alkaline phosphatase in the BMP-2, and BMP-2/VEGF-165 groups was significantly higher than that in the untransfected and blank vector groups (P < 0.05). These findings reveal that BMSCs were successfully transfected by hBMP-2 and hVEGF-165, and the dual gene transfection could promote the production of alkaline phosphatase, thereby accelerating the osteogenic differentiation of BMSCs. 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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Transfection with vascular endothelial growth factor 165, neurotrophin 3 and angiopoietin 1 induces the differentiation of bone marrow mesenchymal stem cells into neurons and vascular endothelial cells Jiang Xing-hai, Zhao Biao, Wu Kai, Xiao Ren-shun, Wang Xiao-mei 2018, 22 (25):  3956-3962.  doi: 10.3969/j.issn.2095-4344.0909 Abstract ( 351 )   PDF (942KB) ( 106 )   Save BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) are a kind of multifunctional mesenchymal stem cells that have multi-directional differentiation potential. These cells can be genetically modified by methods such as adenovirus transfection, to produce a variety of growth factors such as neurotrophin-3 (NT-3), angiopoietin-1 (Ang-1), and vascular endothelial growth factor (VEGF). BMSCs have been applied as seed cells in tissue engineering research. OBJECTIVE: To transfect VEGF165, NT-3 and Ang-1 gene into rat BMSCs and to induce them to differentiate into neuronal cells and vascular endothelial cells. METHODS: BMSCs were isolated using differentia velocity adherent method and identified by flow cytometry. Differentiation potential of BMSCs into osteogenesis and adipogenesis was also identified. We constructed two adenoviral vectors with cytomegalovirus (CMV) as a promoter. One of the two vectors was a bicistronic vector (Adv-Bic), carrying hVEGF165 and Ang-1 gene; the other vector was an AdvNT-3 vector, carrying NT-3 gene. Adv-Bic (green fluorescence) and AdvNT-3 (red fluorescence) were simultaneously transferred into rat BMSCs at various multiplicities of infection (MOIs). Two days after the transfection, the optimal MOI value was determined based on the expression of green and red fluorescent proteins under a fluorescence microscope. Then, well-grown BMSCs at passage 3 were divided into two groups. The experimental group was transfected with Adv-Bic (without fluorescence) and AdvNT-3 (without fluorescence) at the optimal MOI. The control group was transfected with blank control virus at the same MOI. The two groups were cultured in vitro for 7 days and then were detected with immunofluorescence and qPCR. RESULTS AND CONCLUSION: The expression of CD29 and CD44, which are identifications of BMSCs, was positive, while the expression of CD34 and CD45, which are identifications of hematopoietic stem cells, was negative. BMSCs could be differentiated into osteoblasts and adipocytes. After BMSCs were transfected with adenovirus for 48 hours, the expression of green and red fluorescent proteins was observed under a fluorescence microscope. With the increase of MOIs, the intensity of fluorescence gradually increased, and peaked when MOI was equal to 100. Immunofluorescence and qPCR results showed that the expression of VEGF165, NT-3 and Ang-1 in the experimental group was higher than that in the control group. The expression level of CD31 and CD34, the markers of vascular endothelium, in the experimental group was higher than that in the control group. The expression level of neuron-specific markers NSE and Nestin in the experimental group was higher than that in the control group. These results suggested that VEGF165, NT-3 and Ang-1 genes were successfully transfected into BMSCs. This transfection made BMSCs overexpress growth factors of VEGF165, NT-3 and Ang-1, which could induce BMSCs to differentiate into neuronal cells and vascular endothelial cells. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Effect of Erxian Decoction on proliferation and bone differentiation of bone marrow mesenchymal stem cells in ovariectomized rats Wu Qi, Liu Bo, Chen Zhi, Xu Peng 2018, 22 (25):  3963-3968.  doi: 10.3969/j.issn.2095-4344.0940 Abstract ( 272 )   PDF (719KB) ( 114 )   Save BACKGROUND: Some studies have shown that bone marrow mesenchymal stem cells (BMSCs) are involved in bone formation. Erxian Decoction can treat osteoporosis, but its relationship with BMSCs in ovariectomized rats is not yet clear. OBJECTIVE: To reveal the relationship between Erxian Decoction and BMSCs by exploring the effect of Erxian Decoction on proliferation and differentiation of BMSCs in rats with osteoporosis. METHODS: Sprague-Dawley rats were randomly divided into six groups: sham group, model group, estrogen group, high-dose Erxian Decoction group, middle-dose Erxian Decoction group and low-dose Erxian Decoction groups. Ovariectomy was performed to establish rat osteoporosis models in all the groups except the sham group. The rats in the sham group and model group were given normal saline via gavage, and those in the estrogen group were fed with estradiol valerate (0.08 g/L). Rats in the high-, middle- and low-dose groups were given high (1.2 g/mL), middle (0.8 g/mL) and low (0.4 g/mL) concentrations of Erxian Decoction extracts at a dose of 10 mL/kg, for 90 continuous days. BMSCs were isolated by whole bone marrow adherent method, MTT method was used to draw the growth curve of BMSCs, alkaline phosphates kit was used to detect alkaline phosphatase activity, and alizarin red staining was used to detect calcium nodules in BMSCs. RESULTS AND CONCLUSION: Compared with the model group, high-, middle -, and low-dose Erxian Decoction significantly increased the proliferative capacity of BMSCs in ovariectomized rats (P < 0.01), and significantly improved alkaline phosphatase activity in BMSCs of ovariectomized rats (P < 0.05, P < 0.0.1). The highest activity of alkaline phosphatase in BMSCs was found after treatment with middle-dose Erxian Decoction (P < 0.01). High- and middle-dose Erxian Decoction also promoted the formation of calcium nodules in BMSCs (P < 0.01). These results indicate that to improve BMSCs proliferation and osteogenic differentiation in ovariectomized rats is one of the intrinsic mechanisms of Erxian Decoction in the treatment of osteoporosis. 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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Repairing articular cartilage defects in rabbits by adipose-derived stem cells combined with chitosan/collagen scaffold and GDF-5 Wang Zheng, Yu Hui, Yu Yao, Wu Bo-lin 2018, 22 (25):  3969-3974.  doi: 10.3969/j.issn.2095-4344.0955 Abstract ( 376 )   PDF (804KB) ( 169 )   Save BACKGROUND: GDF-5 can induce the differentiation of human adipose-derived stem cells into chondrocyte-like cells, and chitosan/collagen scaffolds are widely used in cartilage tissue repair. OBJECTIVE: To repair rabbit articular cartilage defects by using rabbit autologous adipose-derived stem cells combined with chitosan/collagen scaffold and GDF-5 and to observe the repair effect. METHODS: Adipose-derived stem cells isolated from subcutaneous adipose tissues of Japanese rabbits were cultured, purified, amplified in vitro, and labeled with BrdU in vitro. The chitosan/collagen composite scaffold was prepared in a ratio of 1:3 using ultraviolet cross-linking method. Then labeled adipose-derived stem cells were seeded onto the scaffold and cultured to form complexes. A cylindrical defect (5 mm in diameter, 3 mm in depth) in the knee articular cartilage was surgically made in each rabbit, and these rabbit models were thereafter randomly divided into four groups. Composite group: adipose-derived stem cells+chitosan/collagen scaffold+GDF-5 complex implantation; stent group: simple implantation of chitosan/collagen scaffold; cell group: simple implantation of adipose-derived stem cells; and sham operation group: simple drilling without treatment. All the rabbits were intervened for 4, 8, and 12 weeks. Meanwhile, histological detection, type II collagen immunohistochemical staining and BrdU mmunofluorescence staining were performed. RESULTS AND CONCLUSION: (1) At 12 weeks after surgery, the cylindrical defect of the composite group was completely filled with the same milky white transparent substance as the surrounding normal cartilage, and the cartilage defect area was replaced by differentiated hyaline cartilage. Of all the groups, Wakitani score was the lowest in the composite group (P < 0.01). (2) Type II collagen immunohistochemical staining and BrdU immunofluorescence staining: in the composite group, the type II collagen stained cytoplasm in the repaired area was detected by immunohistochemical staining at 4, 8, and 12 weeks after surgery. The expression of BrdU in the new tissues of the repaired area was positive. These results confirm that autologous adipose-derived stem cells+chitosan/collagen scaffold+GDF-5 complex grafting has a good effect on the repair of rabbit knee cartilage defects. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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HGF/c-Met axis regulates human adipose tissue-derived stem cells in burn wound healing Li Xue-yang, Zheng Wan-ling, Yang Chao, Xia Si-zhan, Xing Xin 2018, 22 (25):  3975-3980.  doi: 10.3969/j.issn.2095-4344.0929 Abstract ( 319 )   PDF (789KB) ( 97 )   Save BACKGROUND: Studies have found that hepatocyte growth factor (HGF) can promote proliferation and migration of adipose-derived stem cells (ADSCs) and accelerate wound healing. OBJECTIVE: To investigate the mechanism of HGF/c-Met on the proliferation and migration of ADSCs and its effects on burn wound healing. METHODS: ADSCs were separated from subcutaneous adipose tissues and primary cultured. ADSCs untreated were considered as control group. Cells in experimental groups were treated with HGF (10 μg/L, 24 hours) or c-Met siRNA combined with HGF (10 μg/L, 24 hours). Then, western blot assay was used to detect expressions of c-Met, p-AKT, and AKT in the cells. The proliferation and migration abilities of ADSCs were observed by cell counting kit-8 and Transwell assay. Twelve nude mice were burned by copper bar at 99 oC for   5 seconds in order to establish animal models of skin III degree burns with the diameter of 1 cm, and then these model mice were randomly divided into experimental and control group (n=6 per group). In control group, only 1×106 ADSCs were injected around the wound, while in experimental group, 1×106 ADSCs transfected with Ad-HGF were injected around the wound. All the mice were killed at  7 days of the experiment, and fresh full-thickness skin wound tissues were taken. Wound healing rate was calculated and the thickness between dermis and epidermis was measured by hematoxylin-eosin staining. RESULTS AND CONCLUSION: Significant expression of c-Met was induced by HGF (P < 0.05). The proliferation and migration of ADSCs were promoted by HGF but suppressed by c-Met siRNA (P < 0.05). p-AKT expression could be promoted by HGF but inhibited by c-Met siRNA. Compared with the control group, the experimental group showed higher wound healing rate and thickened re-epithelium (P < 0.05). To conclude, the HGF/c-Met axis can promote the proliferation and migration of ADSCs by activating PI3K/AKT pathway, and HGF can accelerate wound healing of burn nude mice. 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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Directed differentiation of bone marrow mesenchymal stem cells under regulation by polyacrylamide with different stiffness Guo Jiang-long, He Jing, Wu Fang 2018, 22 (25):  3981-3986.  doi: 10.3969/j.issn.2095-4344.0956 Abstract ( 296 )   PDF (941KB) ( 117 )   Save BACKGROUND: The hardness of substrate materials can directly affect the differentiation fate of stem cells, but the sensitivity and regulatory mechanism of stem cells differentiating into different somatic cells are still unclear. OBJECTIVE: To explore the effect of substrate mechanical properties on the differentiation of bone marrow mesenchymal stem cells and its mechanism. METHODS: In this study, the mechanical strength of polyacrylamide was modulated to be similar to that of nerve tissue (0.5 kPa), cartilage tissue (10 kPa), myocardial tissue (20 kPa) and bone tissue (40 kPa), respectively. Differentiation of bone marrow mesenchymal stem cells on the substrate with different elastic moduli was detected using ELISA, and cytoskeleton morphology and vinculin expression were detected using cytoskeleton staining and immunofluorescence staining. RESULTS AND CONCLUSION: (1) Bone marrow mesenchymal stem cells on the substrate with low stiffness trended to differentiate into nerve and cartilage, while the cells on the substrate with high stiffness had no obvious trend to differentiate into cardiocytes and osteoblasts. This indicated that bone marrow mesenchymal stem cells were sensitive to the mechanical properties of the material during nerve and cartilage differentiation, while were insensitive to the mechanical properties of the material during cardiocyte and osteoblast differentiation. (2) The expression level of vinculin increased as the substrate stiffness increased, indicating that the mechanical signals induced by bone marrow mesenchymal stem cells on low stiffness substrates are relatively low, which can activate certain signaling pathways related to cartilage and neural 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 Figures and Tables | References | Related Articles | Metrics

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miR-93 effects on the proliferation, apoptosis and invasion of tumor stem cells in thyroid papillary carcinoma He Xin-zhi, Lu Guan-ming, Pu Jian, Wei Zhong-heng, Ma Yan-fei, Chen Yong-cheng, Qin Qiang, Huang Qian-fang, Luo Zhi-zhai 2018, 22 (25):  3987-3992.  doi: 10.3969/j.issn.2095-4344.0957 Abstract ( 285 )   PDF (743KB) ( 116 )   Save BACKGROUND: Previous studies have found that miR-93 acts as a protective factor rather than a virulence factor involved in the occurrence and development of thyroid adenomas and stress insulin resistance. OBJECTIVE: To study the effect of miR-93 on the proliferation, apoptosis and invasion of tumor stem cells in thyroid papillary carcinoma. METHODS: Flow cytometry was used to sort CD44+CD24- cells (cancer stem cells) and CD44+CD24- cells in human thyroid cancer cells TPC-1. The expression levels of miR-93 gene were detected in these two kinds of cells. CD44+CD24- cancer stem cells were transfected with miR-93 mimics and miRNA mimics. Then, cell proliferation, apoptosis and invasion were detected through cell counting kit-8, flow cytometry, Transwell assay, respectively. Western blot assay was also used to measure expression of Ki67, Bax, Bcl-2, matrix metalloproteinases 2 and 9. RESULTS AND CONCLUSION: The expression of miR-93 in CD44+CD24- cells was significantly lower than that in non-CD44+CD24- cells  (P < 0.05). CD44+CD24- tumor stem cells transfected with miR-93 mimics exhibited lower proliferation (P < 0.05) and lower Ki67 expression than those transfected with miRNA mimics (P < 0.05). Compared with the miRNA mimics group, higher apoptotic rate, higher Bax expression and lower Bcl-2 expression were detected in the miR-93 mimics group (P < 0.05). Cell invasion ability and expression levels of matrix metalloproteinases 2 and 9 were also lower in the CD44+CD24- cancer stem cells transfected with miR-93 mimics than those transfected with miRNA mimics (P < 0.05). Our experimental findings suggest that miR-93 can inhibit the proliferation and invasion of tumor stem cells in thyroid papillary carcinoma and promote cell apoptosis. 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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Transfusion with human umbilical cord mesenchymal stem cells improves liver function in hepatitis B patients with decompensated liver cirrhosis Fu Qing-chun, Jin Yin-peng, Wang Xiao-jin, Li Li, Wang Xi, Li Hong-chao, Wang Zhao-jing, Zhou Feng, Zang Zu-sheng, Shi Li-qin, Li Zhen-yu, Chen Cheng-wei 2018, 22 (25):  3993-4000.  doi: 10.3969/j.issn.2095-4344.0922 Abstract ( 255 )   PDF (893KB) ( 156 )   Save BACKGROUND: Decompensated liver cirrhosis, a life-threatening complication of chronic liver disease, is one of the major indications for liver transplantation. OBJECTIVE: To evaluate the safety and efficacy of human umbilical cord mesenchymal stem cell (hUC-MSC) transplantation in hepatitis B patients as a treatment for decompensated liver cirrhosis. METHODS: The mesenchymal stem cell (MSC) treatment group contained 17 hepatitis B patients with decompensated liver cirrhosis and the control group contained 17 matched controls. All patients received conventional treatment for hepatitis B cirrhosis, including the oral administration of antiviral nucleoside analogues. Patients in the MSC treatment group received hUC-MSC infusions into the elbow vein, while control patients received no injection. Follow-up examinations were conducted after treatment to evaluate clinical symptoms, liver function and blood coagulation function. In addition, the severity of cirrhosis was assessed based on the Child-Pugh and Model for End-Stage Liver Disease scores. RESULTS AND CONCLUSION: In this study, we revealed that hUC-MSC-treated patients had an improvement in liver function, as indicated by the recovered albumin, bilirubin, lactate dehydrogenase and alkaline phosphatase levels to some extent. Additionally, hUC-MSC-treated patients had better Child-Pugh and Model for End Stage Liver Disease scores than control patients. Therefore, our findings reveal that the transfusion with hUC-MSCs improves the liver function, which represents a safe approach for treating decompensated liver cirrhosis in hepatitis B 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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Effects of Wnt/beta-catenin signaling pathway on the proliferation, migration and invasion of ovarian cancer stem cells Yao Bin, Zhang Qing-hua 2018, 22 (25):  4001-4006.  doi: 10.3969/j.issn.2095-4344.0930 Abstract ( 379 )   PDF (704KB) ( 129 )   Save BACKGROUND: Inability of degradation makes free cytosolic β-catenin in tumor cells aggregate and bind to TCF/LEF into the nuclei, which initiates transcription of downstream genes, and eventually induces abnormal changes in Wnt/β-catenin signaling pathways. Therefore, investigations on the relationship between β-catenin and ovarian cancer stem cells can provide a new insight into the specific inhibition of ovarian cancer stem cells. OBJECTIVE: To observe the proliferation, migration and invasion abilities of ovarian cancer stem cells after β-catenin gene silencing. METHODS: CD133 positive tumor stem cells were isolated from human ovarian cancer cell line A2780 by immunomagnetic beads. CD133 positive cells with 70%-80% confluence were selected and transfected with pLKO.1-shRNA-β-catenin (experimental group) and pLKO.1-shRNA-ctrln (control group), respectively. The expression of β-catenin, E-cadherin and Vimentin were detected by western blot at 72 hours after transfection. Cell proliferation was detected by MTT, and cell migration and invasion ability was detected by Transwell chamber assay. RESULTS AND CONCLUSION: (1) The expression of β-catenin protein in the experimental group was significantly lower than that in the control group at 72 hours after transfection (P < 0.05). (2) With the extension of culture time, the proliferation ability of the two groups increased gradually, but the cell proliferation ability of the experimental group was lower than that of the control group at 2 and 3 days after transfection (P < 0.05). (3) After 72 hours of transfection, the cell migration and invasion abilities of the experimental group were significantly lower than those of the control group (P < 0.05). (4) At 72 hours after transfection, the expression of E-cadherin protein in the experimental group was significantly higher than that of the control group (P < 0.05), and the expression of Vimentin protein was lower than that of the control group (P < 0.05). To conclude, the Wnt/β-catenin signaling pathway is involved in the migration and invasion of ovarian cancer stem cells. β-catenin silencing can inhibit the proliferation, migration and invasion of ovarian cancer stem cells. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Autologous bone marrow mesenchymal stem cell transplantation for the treatment of early femoral head necrosis Chen Xiang, Wang Wei-zhuo, Xue Jian-li, Xia Li-feng, Li Tao, Qiu Ling, Song Huan-jin 2018, 22 (25):  4007-4013.  doi: 10.3969/j.issn.2095-4344.0927 Abstract ( 244 )   PDF (831KB) ( 103 )   Save BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) as ideal seed cells have been worldwide confirmed to improve the microcirculation in the necrotic area of the femoral head. OBJECTIVE: To observe the efficacy of autologous BMSCs transplantation in the treatment of early necrosis of the femoral head. METHODS: (1) In vitro experiment. Osteoblasts were co-cultured with BMSCs using Transwell system as experimental group, osteoblasts cultured normally as blank group, and osteoblasts cultured in serum-free medium as control group. MTT was used to detect cell viability at 3 days of culture, flow cytometry used to detect cell apoptosis, and ELISA used to detect levels of vascular endothelial growth factor and bone morphogenetic protein 2 in the culture medium. (2) In vivo experiment. Thirty New Zealand white rabbits were randomized into three groups: blank group, control group (core decompression plus normal saline injection), and experimental group (core decompression plus autologous BMSCs injection). Femoral head samples from white rabbits were taken for hematoxylin-eosin and immunohistochemical staining at 7 days after cell transplantation. RESULTS AND CONCLUSION: (1) Compared with the control and blank groups, the experimental group showed increased osteoblast proliferation, decreased osteoblast apoptosis, and enhanced levels of vascular endothelial growth factor and bone morphogenetic protein 2 in the culture medium (all P < 0.05). (2) In the control group, broken trabecular bone in the femoral head, evident adipocyte necrosis and a large number of empty bone lacunae were detected. In the experimental group, the number of osteoblasts and capillaries was increased as compared with the control group. The expression levels of vascular endothelial growth factor and bone morphogenetic protein 2 in the necrotic area of the femoral head were significantly increased in the experimental group compared with the control group (P < 0.05). To conclude, BMSCs transplantation can improve the microcirculation in the necrotic area of the femoral head by increasing the expression of vascular endothelial growth factor and bone morphogenetic protein 2. 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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Intravenous transplantation of human umbilical cord blood mononuclear cells in an intracerebral hemorrhage rat model: microPET-CT evaluation and anti-inflammatory mechanisms Jin Na, Yao Xing-yu, Zhang Guo-hua, Yang Li-min, Li Jian-bo, Liu Jie, Wei Lin-huan, Duan Rui 2018, 22 (25):  4014-4020.  doi: 10.3969/j.issn.2095-4344.0958 Abstract ( 378 )   PDF (912KB) ( 86 )   Save BACKGROUND: Mononuclear cells isolated from umbilical cord blood (UCB-MNCs) are rich in cell types which are of great benefit in cell replacement therapy, gene therapy and tissue engineering. OBJECTIVE: To evaluate the efficacy of human UCB-MNCs (HUCB-MNCs) in the treatment of intracerebral hemorrhage rats by microPET-CT and Longa scores, and to discuss the therapeutic mechanism. METHODS: We made a rat intracerebral hemorrhage model by autologous blood injection. After modeling, rats were randomly divided into two groups: transplantation group and control group. The HUCB-MNCs were transplanted into the rats in the transplantation group via the tail vein. Control group received no treatment. Before and at 3, 7, 14, 21 days after transplantation, brain metabolic changes and neurological function scores of rats in the two groups were evaluated by microPET-CT and Longa scores. Levels of tumor necrosis factor-α and interleukin-6 in the rat serum of two groups were measured by ELISA method. RESULTS AND CONCLUSION: (1) At 7, 14 and 21 days after HUCB-MNCs transplantation, the rates of brain metabolism in the hematoma center as well as around the hematoma in the transplantation group were significantly higher than that in the control group (P < 0.05), while the volume of hematoma and surrounding ischemic tissues as well as Longa scores were significantly reduced in the transplantation group as compared with the control group (P < 0.05). The levels of tumor necrosis factor-α and interleukin-6 in the serum in the transplantation group were significantly lower than those in the control group at 3, 7, 14 days after transplantation, and further lowered to the normal levels at 21 days after transplantation. These findings suggest that transplantation of HUCB-MNCs through the tail vein can promote the neurologic recovery of cerebral hemorrhage rats, and one of the therapeutic mechanisms may be related to the reduction of inflammatory factors and the inhibition of inflammatory response. 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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Effects of exosomes derived from canine umbilical cord mesenchymal stem cells on canine skin wound Zhan Xiao-shu, Luo Dong-zhang, Wang Bing-yun, Lu Yu, Xian Wei-hang, Lan Yi-qi, Guo Jian-bo, Luo Hui-na, Bai Yin-shan, Ji Hui-qin, Chen Sheng-feng, Chen Zhi-sheng, Liu Can-ying 2018, 22 (25):  4021-4027.  doi: 10.3969/j.issn.2095-4344.0959 Abstract ( 450 )   PDF (976KB) ( 238 )   Save BACKGROUND: Studies have found that mesenchymal stem cell-derived exosomes can inhibit inflammation,  promote skin cell proliferation and angiogenesis, and reduce myofibroblast accumulation. In other words, the injury repair by mesenchymal stem cells is mainly implemented via paracrine mechanism, providing a new method for the treatment of skin injuries. However, there are few studies on the treatment of skin wounds using canine umbilical cord mesenchymal stem cell-derived exosomes. OBJECTIVE: To investigate the repair effect of umbilical cord mesenchymal stem cell-derived exosomes on canine skin wounds and the underlying mechanisms. METHODS: Twelve healthy dogs with the same age and growth status were selected. A 4 cm×4 cm square full-thickness incision was made under the medial scapula of the dog back and these 12 dog models were randomly divided into two groups, with 6 rats in each group. The treatment group was injected with 1 mL of saline containing 300 mg/L exosomes, while the model group was injected with the same volume of normal saline as a control. Before and after the modeling, the dog’s mental status and wound healing were observed daily. The blood routine and the wound area were measured regularly and the callus tissues were taken to make pathological observation in the 1st, 2nd and 4th week after modeling. RESULTS AND CONCLUSION: In the 1st week, there was a marked inflammatory reaction in the model group, and the number of white blood cells was also increased by 30% compared with the baseline data. The inflammatory response in the treatment group was not obvious, and the number of white blood cells and neutrophils was increased slowly as compared with the model group. There was no significant difference in the number of lymphocytes and red blood cells between the two groups. The growth of granulation tissue and wound healing in the treatment group were faster compared with the model group, but no significant differences were detected. These results suggest that canine umbilical cord mesenchymal stem cell-derived exosomes, to some extent, inhibit inflammation and promote skin wound healing. 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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A comparative study on in vitro proliferation, differentiation, transportation and immunocompatibility of mesenchymal stem cells derived from human amniotic fluid, umbilical cord, and placenta Zhao Shu-can, Zheng Gui-chun, Lin Lian-peng, Mei Zhan-tu, Wang Bing-yun, Chen Sheng-feng, Chen Zhi-sheng 2018, 22 (25):  4028-4034.  doi: 10.3969/j.issn.2095-4344.0960 Abstract ( 376 )   PDF (885KB) ( 169 )   Save BACKGROUND: With the advancement of regenerative medicine, the use frequency of stem cells has gradually increased in the treatment of diseases. At present, several factors such as the number of cells, the ability of differentiation and the possibility of immunological rejection during allogeneic inoculation are taken into consideration in the clinical use of stem cells. Accordingly, it is very meaningful to screen out the more suitable type of stem cells for clinical use. OBJECTIVE: To discuss the differences of mesenchymal stem cells (MSCs) derived from human amniotic fluid, umbilical cord and placenta in terms of the ability to proliferate in vitro, the ability to differentiate, the possibility of transport and the possibility of allogeneic inoculation, thereby providing evidence for the clinical application of MSCs. METHODS: Three kinds of MSCs were isolated and cultured at first, and they were then identified by immunofluorescence and induced into osteoblasts, adipocytes and neuroblasts in vitro. qPCR was used to detect the expression of Nestin, NT-3, and MANF after neural differentiation. HLA-ABC and HLA-DR expression in these three kinds of MSCs was quantitatively analyzed by flow cytometry. Finally, the number of living cells was calculated at 24, 48 and 72 hours after they were placed in the cell transport solution at room temperature. RESULTS AND CONCLUSION: (1) The adipogenic ability of placenta-derived MSCs and the osteogenic capability of umbilical cord-derived MSCs were the best among three kinds of MSCs. All three kinds of cells expressed Nestin and GFAP, which are markers of neural stem cells. (2) NT-3 was highly expressed in umbilical cord-derived MSCs after neural induction, while MANF was highly expressed in placenta-derived MSCs after neural induction. They could be used to repair different types of nerve injuries. (3) The human amniotic fluid-derived MSCs were more suitable for allogeneic inoculation because they expressed HLA-ABC in a low level and did not express HLA-DR which showed the lowest possibility to cause immune rejection. (4) The survival rate of amniotic fluid-derived MSCs was the highest after a 72 hours induction in the cell transport fluid. In conclusion, as there is a slight difference in biological characteristics of human amniotic fluid, umbilical cord and placenta-derived MSCs, these three kinds of MSCs can be used for clinical treatment of different diseases.  中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Bone marrow mesenchymal stem cells stably expressing NICD which is controlled by Tet-On system in the treatment of renal ischemia-reperfusion injury Xi Xiao-qing, Hu Hong-lin, Zou Cong, Fang Zhou-qing, Kuang Ren-rui, Ye Zhen-feng, Huang Ya-wei 2018, 22 (25):  4035-4040.  doi: 10.3969/j.issn.2095-4344.0911 Abstract ( 368 )   PDF (796KB) ( 134 )   Save BACKGROUND: Renal ischemia-reperfusion injury (IRI) is the main cause of acute kidney injury (AKI). The treatment of renal IRI by bone marrow mesenchymal stem cells (MSC) has always been an issue of concern. OBJECTIVE: To construct BMSCs stably expressing NICD controlled by Tet-On system in vitro (NICD-Tet-BMSCs), and to explore its therapeutic effect on renal IRI. METHODS: We separated and cultured rat BMSCs by density gradient centrifugation, and constructed NICD-Tet-BMSCs in vitro. The cells were then transplanted to renal IRI rats. Serum creatinine level, the distribution of BMSCs in renal tissue and the expression of vascular endothelial growth factor and tumor necrosis factor α in the kidney were detected thereafter. RESULTS AND CONCLUSION: NICD protein stably expressed in the NICD-Tet-BMSCs which was regulated by the doxycycline. Compared with the control group, the renal IRI rats undergoing NICD-Tet-BMSCs transplantation had significantly lower serum creatinine levels (P < 0.05), sustained cell proliferation, increased vascular endothelial growth factor protein expression and decreased tumor necrosis factor α protein expression in the kidney (P < 0.05). Therefore, NICD-Tet-BMSCs can be successfully established in vitro, and promote the reparation of renal IRI 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 Figures and Tables | References | Related Articles | Metrics

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Oligodendrocyte progenitor cell transplantation in combination with miconazole for myelin repair in mice with leukodystrophy Wu Cheng-jun, Su Xue-wen, Wang Zhao-yan, Yang Yin-xiang, Luan Zuo 2018, 22 (25):  4041-4046.  doi: 10.3969/j.issn.2095-4344.0961 Abstract ( 364 )   PDF (669KB) ( 118 )   Save BACKGROUND: Oligodendrocyte progenitor cells are precursor cells of oligodendrocytes that are myelinating cells in the central nervous system. These cells are closely related to central demyelinating diseases such as leukodystrophy. Recent studies have shown that miconazole can promote oligodendrocyte progenitor cell differentiation and myelin regeneration. The combination of miconazole and oligodendrocyte progenitor cells is likely to be used for the treatment of leukodystrophy. OBJECTIVE: To investigate the effect of oligodendrocyte progenitor cell transplantation combined with miconazole on myelin reparation in mice with leukodystrophy.   METHODS: Myelin basic protein (MBP) knockout was performed to establish animal models of leukodystrophy in shiverer mice. Twenty-six newborn shi-/- mice were randomly assigned to negative control group and treatment group (n=13 per group). In addition, 13 newborn shi+/+ mice were selected as positive control group. In the treatment group, oligodendrocyte progenitor cells were transplanted within 24 hours after birth, and in the control groups, oligodendrocyte progenitor cells were replaced by PBS. At the same time, in the treatment group miconazole was injected intraperitoneally at 1-5 days after birth, and in the control groups, miconazole was replaced by normal saline. At the age of 8 weeks, MBP immunofluorescence staining and western blot were used to detect MBP protein expression. The myelin sheath structure of the corpus callosum was observed by transmission electron microscopy. RESULTS AND CONCLUSION: Compared with the negative control group, the MBP expression in the treatment group was significantly enhanced and the average fluorescence intensity was significantly increased (P < 0.05). But when compared with the positive control group, the MBP expression in the treatment group was significantly weakened and the average fluorescence intensity significantly decreased (P < 0.05). Under the transmission electron microscope, the myelin sheath structure in the corpus callosum region was significantly diverse among the groups. In the negative control group, the myelin sheath was rare and irregular, and the lamellar arrangement was loose and disordered. In the positive control group, the myelin was significantly increased in number and thickened. With the normal morphology, the lamellar layer was closely arranged and the boundary was clear. In the treatment group, the number of myelin was significantly higher than that of the negative control group, and some myelin sheaths were normal in shape, but most myelin sheaths were still slender. These findings indicate that oligodendrocyte progenitor cells combined with miconazole helps to improve demyelination in mice with leukodystrophy, but cannot completely reverse the pathological status of damaged myelin. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Biological properties of primary chondrocytes isolated by one-step digestion versus stepwise digestion in newborn rats Deng Lin-xia, Yu Mu-xue, Pan Si-nian, Guo Chu-yi, Qiu Xiao-shan, Cao Zhi-yu 2018, 22 (25):  4047-4052.  doi: 10.3969/j.issn.2095-4344.0962 Abstract ( 277 )   PDF (793KB) ( 123 )   Save BACKGROUND: In vitro culture of primary chondrocytes is one of the most important methods to study the bone growth. Currently, the main methods of primary chondrocyte separation are one-step digestion and stepwise digestion. However, the biological properties of chondrocytes isolated by these two methods have not been well documented. OBJECTIVE: To compare the biological properties of chondrocytes in newborn rats isolated using these two methods and to provide experimental basis for the programmed study of cartilage development. METHODS: Articular cartilage of newborn SD rats was removed under aseptic conditions. Chondrocytes were isolated by one-step digestion (type II collagen) and stepwise digestion (type II collagen and trypsin). Primary culture and subculture were performed. Cell morphology was observed under light microscope, cell proliferation was detected by cell counting kit-8 assay, and the expression of type II collagen of chondrocytes was analyzed by semi-quantitative immunofluorescence. RESULTS AND CONCLUSION: A large number of chondrocytes were yielded by two digestion methods, but the purity of chondrocytes obtained by stepwise digestion method was higher than that by one-step digestion method. Under the light microscope, there was no significant difference in the cell morphology between the two groups. The growth of chondrocytes obtained by one-step digestion was faster than that by stepwise digestion method. Results from the semi-quantitative immunofluorescence analysis showed that the chondrocytes obtained by these two digestion methods showed the same ability to express type II collagen. These findings indicate that compared with one-step digestion, stepwise digestion is more suitable for primary chondrocytes isolation in newborn 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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Inhibiting colorectal cancer liver metastasis by combined injection of allogeneic bone marrow cells and lymphocytes Qu Yuan-qian, Zhou Qing, Zhang Jin, Xu San-rong 2018, 22 (25):  4053-4058.  doi: 10.3969/j.issn.2095-4344.0963 Abstract ( 329 )   PDF (731KB) ( 87 )   Save BACKGROUND: Clinical and experimental studies on prevention and treatment of hepatic metastasis after colonic cancer treatment by allogeneic hematopoietic stem cell transplantation have yet to be well reported. A mixed chimera is expected to achieve certain effects in patients with low-tumor burden after colorectal cancer resection. OBJECTIVE: To investigate the inhibitory effect of allogeneic hematopoietic stem cell transplantation on colorectal cancer liver metastasis as well as the effect on chimera level and graft-versus-host disease and to study the anti-tumor mechanism.  METHODS: CB6F1 mice were hybridized by BALB/c×C57BL/6 mice and injected with CT-26 cells to make animal models of colorectal cancer liver metastasis. Then, model mice were randomized into four groups (n=8 per group): control group, cyclophosphamide group, HLA-identical spleen cells plus bone marrow cell transplantation+cyclophosphamide group (HLA-identical transplantation group), and haploidentical spleen cells plus bone marrow cell transplantation+cyclophosphamide group (haploidentical transplantation group). Mixture of spleen cells and bone marrow cells from CB6F1 and C57BL/6 mice were injected via the tail vein followed by a non-myeloablative pretreatment with intraperitoneal injection of cyclophosphamide, and thereafter, lymphocytes were given via infusion. Survival time, tumor liver metastasis and the occurrence of graft-versus-host disease were observed in mice. The chimera was analyzed by flow cytometry, and levels of plasma interleukin-2, interferon-γ, interleukin-4, and transforming growth factor β were determined by ELISA RESULTS AND CONCLUSION: The survival time and inhibition rate of liver metastasis in the haploidentical transplantation group were significantly prolonged and increased compared with cyclophosphamid and HLA-identical transplantation group, respectively. At 7 days after transplantation, the chimera level in the haploidentical transplantation group was significantly increased up to over 99% at 28 days of transplantation, indicating that donor cells were basically substituted by recipient cells. Significantly enhanced anti-tumor effect was found in the haploidentical transplantation group, while there were no signs of severe graft-versus-host disease. Moreover, the levels of plasma interleukin-2 and interferon-γ in mice were significantly higher than those in the other groups, and the level of transforming growth factor β was significantly reduced. However, there was no significant difference in the interleukin 4 level. To conclude, co-transplantation of cyclophosphamide-pretreated allogeneic bone marrow cells and lymphocytes significantly increases the chimera level, accompanied by the production of obvious anti-tumor effect of the graft that has an inhibitory effect on colorectal cancer liver metastasis. Changes in levels of plasma interleukin-2, interferon-γ, interleukin-4, and transforming growth factor β are associated with tumor growth inhibition. 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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Left ventricular transplantation of human umbilical cord blood mononuclear cells in the treatment of cerebral hemorrhage in rats: micro PET-CT evaluation and therapeutic mechanism Wei Lin-huan, Yao Xing-yu, Zhang Guo-hua, Yang Li-min, Li Jian-bo, Liu Jie, Jin Na, Duan Rui 2018, 22 (25):  4059-4064.  doi: 10.3969/j.issn.2095-4344.0928 Abstract ( 462 )   PDF (757KB) ( 134 )   Save BACKGROUND: In recent years, new stem cell transplantation therapies for cerebral hemorrhage have emerged, but little is reported on human umbilical cord blood mononuclear cells (HUCB-MNCs), especially on micro PET-CT evaluation of HUCB-MNCs effects. OBJECTIVE: To evaluate the therapeutic effect of HUCB-MNCs in a rat model of cerebral hemorrhage by Longa and micro PET-CT and to explore the therapeutic mechanism. METHODS: Models of cerebral hemorrhage were established in rats using secondary blood injection/needle retraction method. The rats in the experimental group were treated with HUCB-MNCs via left ventricular transplantation, and those in the control group received no treatment. Before and after transplantation (3, 7, 14, 21 days), the rats in the two groups were evaluated using Longa 5 scores and micro PET-CT images. ELISA was used to detect the level of angiotensin 1 in the brain homogenate. RESULTS AND CONCLUSION: Longa 5 scores in the two groups were gradually decreased with time (P < 0.05). Compared with the control group, the Longa 5 scores in the experimental group decreased more significantly (P < 0.05). The standard absorption value (SUV%) in the center of hematoma and perihematomal region increased gradually in the two groups at 7, 14, 21 days after transplantation, but it was significantly higher in the experimental group than the control group (P < 0.05). The volume of hematoma and surrounding tissues was gradually reduced in the two groups, especially in the experimental group, at 7, 14, 21 days after transplantation (P < 0.05). The level of angiotensin 1 in the control group increased gradually with time (P < 0.05), while the level of angiotensin 1 in the experimental group gradually increased, peaked at 14 days and then reduced at 21 days after transplantation (P < 0.05). The level of angiotensin 1 was higher in the experimental group than the control group (P < 0.05). Overall, these findings reveal that HUCB-MNCs via left ventricular transplantation can improve neurologic function of rats, reduce hematoma, recover hematoma metabolism, and increase the level of angiotensin 1 in the rat brain, indicating that HUCB-MNCs transplantation can promote the repair of cerebral hemorrhage through neo-angiogenesis regulation. 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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Isolation and culture of multifidus satellite cells by single muscle fiber method Liu Tong, Yu Jia-ni, Chen Yu-pei, Chen Huan, Zou De-hui, Yan Jun, Lu Zong-xiao, Liu Yue, Zhang Li, Huo Ze-jun 2018, 22 (25):  4065-4070.  doi: 10.3969/j.issn.2095-4344.0941 Abstract ( 317 )   PDF (764KB) ( 142 )   Save BACKGROUND: Atrophy and damage of multifidus muscles mainly result in various low back pains and dysfunctions. Muscle satellite cells, which belong to muscle stem cells, play an important role for muscle regeneration after injury. However, little is reported on the isolation and culture of multifidus muscle up to now. OBJECTIVE: To isolate and identify multifidus satellite cells of rats using single muscle fiber method. METHODS: Multifidus muscles were isolated in vivo and multifidus satellite cells were obtained by single muscle fiber method. Then the cells were purified by repeated differential attachment. Morphological changes of the cells were observed and the proliferation curve was made thereafter. Moreover, myogenic differentiation of the isolated cells cultured in differentiation medium was analyzed and marker proteins of multifidus satellite cells (Pax7, Myod, Desmin) were characterized by immunofluorescence. RESULTS AND CONCLUSION: A large number of fusiform or spindle cells were seen around the muscle fibers after 72 hours of culture. Round cells with high refraction were visible after being purified by repeated differential attachment, and the cells adhered after 24-48 hours of re-inoculation exhibited an “S” shaped growth curve. Large myotubes formed in the differentiation medium. Immunofluorescence findings showed that Pax7 and Myod were positively expressed in muscle satellite cell nuclei, while Desmin was positively expressed in the cytoplasm. These findings reveal that multifidus satellite cells can be successfully separated by single muscle fiber method, which can be used for regeneration of multifidus muscles in the future. 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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Embryonic hepatic stem cells from GFP transgenic mice: in vitro separation, cultivation and intra-spleen transplantation Wang Ya-ping, Kong Xiang-ping, Liu Guang-ze, Zhuo Li, Gao Hong-bo, Tang Xiao-ping, Tong Ming-hua 2018, 22 (25):  4071-4076.  doi: 10.3969/j.issn.2095-4344.0910 Abstract ( 361 )   PDF (810KB) ( 146 )   Save BACKGROUND: Recent studies have confirmed that the success of stem cell transplantation and the function of transplanted cells are closely related to the in vivo microenvironment. Under normal physiological conditions, the liver is not susceptible to exogenous cells. Therefore, a transplant animal model is usually established to damage the host liver or reduce the immune response of the host liver, creating a good condition for the transplantation and proliferation of exogenous cells. OBJECTIVE: To observe the colonization and distribution of embryonic hepatic stem cells from GFP transgenic mice in a rat model of liposome-coated dichloromethylene diphosphate/retrorsine and 1/3 liver resection (L-CL2MDP/RS/PH).  METHODS: GFP transgenic mice at gestational 15 days were taken as donors to separate and cultivate embryonic hepatic stem cells. Fluorescence microscope was used to observe the GFP expression followed by immunocytochemical identification. Thirty Sprague-Dawley rats were randomized into experimental group and control group. Rats in the experimental group were given intraperitoneal injection of retrorsine (30 mg/kg, twice, with an interval of 2 weeks). At 1 preoperative day, these rats were subjected to intravenous infusion of L-CL2MDP (1 mL) and 1/3 liver resection followed by intra-spleen transplantation of embryonic hepatic stem cells from GFP transgenic mice. Rats in the control group were only given injection of normal saline instead of L-CL2MDP and retrorsine, but no liver resection. Frozen liver sections were made in the two groups to observe the distribution of GFP-positive hepatic stem cells under the fluorescence microscope at 3, 15, 30 days after transplantation. Moreover, immunohistochemical staining was used to detect the expression of anti-mouse C-Kit antibody in the rat liver. RESULTS AND CONCLUSION: (1) The hepatic stem cells under microscope were uniform with a cobblestone shape. After 5 days of culture, the cells with increased size presented with the shape of fusiform or polygon, and the green fluorescence was observed in cells under the fluorescence microscope. Passage 3 hepatic stem cells expressed CD34 and AFP. (2) At 3 days after spleen transplantation of embryonic hepatic stem cells, GFP positive cells were scattered in the experimental group under the fluorescence microscope, and the GFP positive expression was still visible in the liver at 30 days after transplantation. (3) At 3, 15, and 30 days after transplantation, C-Kit positive cells were visible in the liver of rats in the experimental group. All these findings reveal that embryonic hepatic stem cells from GFP transgenic mice have been successfully transplanted into the spleen of the L-CL2MDP/RS/PH rat model, and the transplanted cells can survive and proliferate in the rat liver. 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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Combined effects of interleukin 1beta and mechanical stretching on Collagen I synthesis and Lumican expression in corneal fibroblasts Feng Peng-fei, Li Xiao-na, Rong Shuo, Chen Wei-yi, Wang Xiao-jun 2018, 22 (25):  4077-4082.  doi: 10.3969/j.issn.2095-4344.0924 Abstract ( 232 )   PDF (688KB) ( 122 )   Save BACKGROUND: Secondary keratoconus is one of the complications after corneal refractive surgery, but the mechanism remains unclear. OBJECTIVE: To study the effects of interleukin-1β and mechanical stretching on the synthesis of the Collagen I and Lumican in corneal fibroblasts.  METHODS: Rabbit corneal fibroblasts were cultured in vitro and were treated by different concentrations of interleukin-1β and different amplitudes of mechanical stretching (5%, 10%, 15%) for 12, 24 or 36 hours. The expression of Collagen I and Lumican was detected by real-time fluorescent quantitative PCR. RESULTS AND CONCLUSION: Interleukin-1β decreased the expression of Collagen I α1 at 12 hours and increased the expression of Lumican at 24 hours (P < 0.05). 5% stretching alone increased the expression of Collagen I α1 at 24 and 36 hours (P < 0.05), but 10% and 15% stretching alone decreased the expression of Collagen I α1 and Collagen I α2 at 12, 24 and 36 hours (P < 0.05). 5% stretching alone increased the expression of Lumican at 12 hours (P < 0.05), while 10% and 15% stretching alone decreased the expression of Lumican at 12 hours (P < 0.05). Moreover, 5%, 10% and 15% stretching alone increased the expression of Lumican at 24 and 36 hours (P < 0.05). Reduced expression of Collagen I α1 and Collagen I α2 and increased expression of Lumican in corneal fibroblasts were observed at 24 and 36 hours after combined treatment with interleukin-1β and mechanical stretching. These results reveal that interleukin-1β can inhibit the synthesis of Collagen I and promote the expression of Lumican. Low-amplitude stretching can increase collagen synthesis; however, middle-to-large- amplitude stretching can inhibit its expression in corneal fibroblasts. The synthesis of collagen can be suppressed and the expression of Lumican can be increased under the combined use of interleukin-1β and stretching. 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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Anti-tumor effects of mesenchymal stem cell derived exosomes Wu Xue, Qian Man-qing, Wu Dong-liang, Zhao Xiao-yin 2018, 22 (25):  4083-4088.  doi: 10.3969/j.issn.2095-4344.0964 Abstract ( 481 )   PDF (704KB) ( 139 )   Save BACKGROUND: Mesenchymal stem cells (MSCs) can secret a lot of exosomes in resting and stress states. What’s more, MSCs-derived exosomes (MSC-exo) have similar functions to the maternal MSCs, and play an important role in cell communication and information delivery. However, the mechanisms of MSCs-exo in tumors are still unclear and controversial. OBJECTIVE: On the basis of clarifying the source and biological characteristics of exosomes, to systematically summarize the research process of MSC-exo on tumor cells in recent years, to prospect the application of MSC-exo in the clinical treatment of tumors, and to summarize the exists problems and solutions in the clinical application of MSC-exo. METHODS: MSC-exo and tumor related literatures published from 1980 to 2018 were retrieved using the key words of “mesenchymal stem cell, exosomes, stem cells, tumor, cancer” in English and Chinese, respectively. Finally 55 articles were retrieved and analyzed. RESULTS AND CONCLUSION: Numerous studies have shown that MSC-exo have similar biological activities with parental cells, and they can also home to the tumor site, release active factors and have a great application prospect as an alternative to the cell therapy. Currently, the role of MSC-exo in tumors is still controversial. Some studies have suggested that MSC-exo can promote tumor growth and metastasis through the delivery of microRNAs and activation of Wnt pathways. However, in another studies, MSC-exo has been shown to inhibit tumor cell growth and metastasis by special cellular interactions. Many problems still need to be solved, such as elucidating the effects and mechanisms of different MSC-exo on various types of tumors, and solving the problems concerning quality control, separation and purification and therapeutic stability of MSC-exo. If the existing problems can be solved one by one, MSC-exo will be expected to be a new means for cancer therapy. 中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程 Figures and Tables | References | Related Articles | Metrics

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Wnt/beta-catenin signaling pathway regulates the differentiation of mesenchymal stem cells into epithelial cells in the treatment of chronic obstructive pulmonary disease Zhou Xiang-xiang, Yan Xing-yan, Zhong Yu-lan, Gan Xin 2018, 22 (25):  4089-4094.  doi: 10.3969/j.issn.2095-4344.0908 Abstract ( 309 )   PDF (768KB) ( 120 )   Save BACKGROUND: Wnt/β-catenin signaling pathway plays an important role in the proliferation and differentiation of mesenchymal stem cells (MSCs). Therefore, the problems encountered during MSCs transplantation, such as low differentiation rate and short survival time, are expected to find a solution by regulating the Wnt/β-catenin signaling pathway, making MSCs transplantation become a new hope of the treatment of chronic obstructive pulmonary disease (COPD). OBJECTIVE: To summarize the progress in Wnt/β-catenin signaling pathway that regulates MSCs to differentiate into epithelial cells in the treatment of COPD and the relevant mechanisms. METHODS: PubMed, Elsevier, Medline, CNKI, WanFang, and CBMdisc databases were retrieved for relevant articles published from 2007 to 2017. The keywords were “chronic obstructive pulmonary disease; Wnt/β-catenin signaling pathway; mesenchymal stem cells (MSCs); pulmonary epithelial differentiation” in Chinese and English, respectively. After literature screening, 50 eligible articles were further analyzed. RESULTS AND CONCLUSION: The Wnt/β-catenin signaling pathway is expected to reverse the low differentiation of MSCs into lung epithelial cells. COPD patients have a complex lung environment, and Wnt/β-catenin signaling pathway is extremely sensitive to the environmental factors. In addition, the regulation by Wnt/β-catenin signaling pathway is varied under different induction conditions. Therefore, enormous investigations are needed to clarify the regulation of the differentiation of MSCs into lung epithelial cells by the Wnt/β-catenin signaling pathway in the treatment of COPD. 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 salvianolic acid B on differentiation direction of bone marrow mesenchymal stem cells Li Zi-yi, Wang Na, Xue Peng, Li Yu-kun 2018, 22 (25):  4095-4100.  doi: 10.3969/j.issn.2095-4344.0428 Abstract ( 319 )   PDF (676KB) ( 75 )   Save BACKGROUND: With the development of tissue engineering, the multi-directional differentiation potential of bone marrow mesenchymal stem cells has brought a new hope for the treatment of many diseases. Salvianolic acid B, as an effective component of Salvia miltiorrhiza Bunge, has many pharmacological activities influencing the directional differentiation of bone marrow mesenchymal stem cells. OBJECTIVE: To summarize the effect of salvianolic acid B on the differentiation of bone marrow mesenchymal stem cells and to explore the underlying mechanisms. METHODS: The first author searched the CNKI, WanFang and PubMed databases using the keywords of “salvianolic acid B, directional differentiation, mesenchymal stem cells” in Chinese and English, respectively, to retrieve relevant articles published from January 2000 to September 2017. One hundred and two Chinese articles and 136 English articles were initially retrieved. Repetitive articles or those with no originality were eliminated. Finally 49 articles were included in the result analysis. RESULTS AND CONCLUSION: It has been found that salvianolic acid B can promote the osteogenic, neuronal and myocardial differentiation of bone marrow mesenchymal stem cells, but inhibit the adipogenic differentiation of the cells. Further investigations are required on whether salvianolic acid B can promote the mutli-directional differentiation of bone marrow mesenchymal stem cells and its effective concentrations.  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