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BACKGROUND:The microRNAs are involved in regulation of stem cel proliferation, differentiation and aging. To study the effect of Let-7c, a member of Let-7, on the neural differentiation of bone marrow mesenchymal stem cels provides new ideas for stem cel therapy. OBJECTIVE: To investigate the role of Let-7c in the neural differentiation of bone marrow mesenchymal stem cels. METHODS: The lentiviral vectors of Let-7c-up and Let-7c-inhibition were constructed and transfected into rat bone marrow mesenchymal stem cels. Optimal multiplicity of infection was screened. The cels were divided into non-transfected group, negative control group (transfected with empty virus), transfected enhancement group (transfected with LV-rno-Let-7c-up), transfected inhibition group (transfected with LV-rno-Let-7c-5p-inhibition). Bone marrow mesenchymal stem cels were treated with fasudil as an inducer for triggering the cels to differentiate into neurons. The fluorescence expressed by transfected cels was observed under inverted fluorescence microscope. The expression of neuron-specific markers, neuron-specific enolase and microtubule-associated protein 2, were measured by immunocytochemical method. The mRNA expression of microtubule-associated protein 2 was detected by RT-PCR. The cel viability was determined by MTT method. RESULTS AND CONCLUSION:Under the inverted fluorescence microscope, the cels were successfuly transfected with LV-rno-Let-7c-up and LV-rno-Let-7c-5p-inhibition. Fasudil induced bone marrow mesenchymal stem cels to differentiate into neurons. The transfection efficiency and expression levels of neuron-specific enolase and microtubule-associated protein 2 in the transfected enhancement group were significantly higher than those in the negative control group (P < 0.05), while in the transfected inhibition group, they were lower than those in the negative control group (P < 0.05). These findings indicate that the differentiation percentage of bone marrow mesenchymal stem cels is increased by fasudil after transfection with LV-rno-Let-7c-up, and Let-7c may promote the differentiation of bone marrow mesenchymal stem cels into neurons.
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BACKGROUND:Numerous studies have demonstrated that bone marrow mesenchymal stem cels can be induced to differentiate into myocardial cels under certain conditions. Dimethyl sulfoxide is one of the commonly used inducers, and its mechanism is mainly by inhibiting the c-myc gene expression, thus reducing endogenous poly(adenosine diphosphate nucleotide) level. OBJECTIVE:To study the feasibility of dimethyl sulfoxide inducing the myocardial differentiation of bone marrow mesenchymal stem cels and its optimal concentration. METHODS:Bone marrow mesenchymal stem cels from Sprague-Dawley rats were isolated and culturedin vitro, and then induced by dimethyl sulfoxide to differentiate into myocardial cels. According to the concentrations of dimethyl sulfoxide, there were three groups: 0.6%, 0.8% and 1.0% group. Additionaly, a blank control group with no induction was set up. After 72 hours of induction, induction media were removed, and cels were then cultured in normal media for 4 weeks. RESULTS AND CONCLUSION: Morphology and immunocytochemistry detection results confirmed that dimethyl sulfoxide could induce the differentiation of bone marrow mesenchymal stem cels into myocardial celsin vitro, and differentiated cels expressed desmin, α-actin, cTnT, cTnI and P38MAPK. The optimal induced concentration of dimethyl sulfoxide was 1.0%. Immunofluorescence double staining and electron microscope results further confirmed that dimethyl sulfoxide could induce the myocardial differentiation of bone marrow mesenchymal stem cels. Cite this article:Sun LY. Dimethyl sulfoxide induces differentiation of bone marrow mesenchymal stem cels into myocardial cels. Zhongguo Zuzhi Gongcheng Yanjiu. 2016;20(1):26-30.
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BACKGROUND:There is a certain cel subset in esophageal cancer tissues, with certain invasive and metastatic properties, which is closely related to the clinical therapeutic effect on tumors. OBJECTIVE:To isolate tumor stem cel spheres in human esophageal carcinoma cel lines KYSE-150 and TE-1 and to analyze their proliferation and invasion ability. METHODS:KYSE-150 and TE-1 cels were cultured in serum-free medium to observe the formation of cel spheres. Cel proliferation and invasion were detected using MTT and Transwel chamber culture. Surface markers of cels were detected using flow cytometry. RESULTS AND CONCLUSION: Cel spheres that were stably subcultured were obtained from KYSE-150 and TE-1 cels cultured in serum-free medium. The proliferation and invasion abilities of cel spheres were significantly stronger than those of parent cels (P < 0.05). The number of CD44+, CD271+ and CD44+CD271+ cels in TE-1 and KYSE-150 cel spheres was significantly higher than that in the TE-1 and KYSE-150 parent cels (P < 0.05). These experimental results show that cel spheres isolated from human esophageal carcinoma cel lines TE-1 and KYSE-150 have tumor stem cel properties as wel as strong proliferation and invasion abilities. And moreover, CD44 and CD271 can be used as important surface markers of esophageal carcinoma stem cels. Cite this article:Wang YL, Wang ZM, Wang Y, Tao YP, Han GY.Culture, differentiation, proliferation and invasion of tumor stem cels in human esophageal carcinoma cel lines KYSE-150 and TE-1. Zhongguo Zuzhi Gongcheng Yanjiu. 2016;20(1):55-59.
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BACKGROUND:Stem cels are induced to differentiate into endothelial-like cels that can be used for the treatment of diabetic lower extremity vascular disease. However, it is unclear whether these endothelial-like cels can completely replace endothelial cels to improve vascular disease and what are the differences between endothelial-like cels and endothelial cels. OBJECTIVE:To explore the differences and similarities between endothelial-like cels and human umbilical vein endothelial cels in the aspects of morphology, function, and viability. METHODS:Umbilical cord mesenchymal stem cels and umbilical vein endothelial cels were isolated, cultured and identified using flow cytometry and immunohistochemical method. Isolated umbilical cord mesenchymal stem cels were induced in DMEM-LG/F12 containing 10 μg/L vascular endothelial growth factor, 10 μg/L basic fibroblast growth factor and 2% fetal bovine serum to differentiate into endothelial-like cels folowed by immunohistochemical identification. To compare endothelial-like cels with human umbilical vein endothelial cels, cel migration detection, active substance measurement and three-dimensional angiogenesis test were performed. RESULTS AND CONCLUSION:Isolated umbilical cord mesenchymal stem cels strongly expressed the surface markers of mesenchymal stem cels, and human umbilical vein endothelial cels strongly expressed CD31 and VWF. After induction, the umbilical cord mesenchymal stem cels were identified to highly express CD31 and VWF. Through cel migration, active substance and three-dimensional angiogenesis tests, endothelial-like cels were similar to endothelial cels in the function and activity, and superior to endothelial cels. Cite this article:Hao XJ, Hao HY, Zhu MJ, Yuan Z, Li WW, Chen J, Zhu LY. Endothelial-like cels versus human umbilical vein endothelial cels. Zhongguo Zuzhi Gongcheng Yanjiu. 2016;20(1):83-88.
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BACKGROUND:Currently, there is a lack of efficient, non-invasive way to transplant stem cels to the target organ or tissue. Exploring a way to guide targeting transplantation of stem cels and to improve the efficiency of stem cel homing is now one of focuses in the field of stem cels research. OBJECTIVE: To establish a simple and feasible method to chemicaly modify the cel surface using biotin-streptavidin reaction system, and to evaluate the efficiency of this method to label bone marrow mesenchymal stem cels (BMSCs) and its effects on cel biological functions. METHODS: Passage 3 BMSCs were obtained by whole bone marrow culture method and verified by flow cytometry. Biotin, streptavidin, sulfonated biotin-N-hydroxy-succinimide were used to equip the adhesion molecule ligand, sialyated LewisX (SLeX), to the BMSCs surface. The labeling rate of BMSCs was assessed using fluorescence microscope, the vitality of BMSCs was evaluated by trypan blue staining, and the proliferation of BMSCs was evaluated by cel counting kit-8 assay. Adipogenic and osteogenic inductions were used to evaluate the effect of the method on the multi-differentiation function of BMSCs. RESULTS AND CONCLUSION: After culture for 2 weeks, passage 3 BMSCs were obtained and confirmed by expressing CD90, CD29 and lack of CD34, CD45. Biotin, streptavidin, sulfonated biotin-N-hydroxy-succinimide were successfuly used to equip sialyated LewisX (SLeX) to the BMSCs surface and had minor effects on the vitality, proliferation, and differentiation of BMSCs. This method was simple for surface modification and had a high modification rate of 88%. The homing of BMSCs modified by this method to the target organ or tissue could be greatly enhanced. Therefore, this method potentialy could have extensive and important applications.
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BACKGROUND:Stromal cel derived factor-1 is a smal molecular protein with a wide range of biological activity that can cause immune cel chemotaxis, and it also has a chemotactic effect on bone marrow stem cels and periodontal ligament cels. OBJECTIVE:To investigate the effect of stromal cel derived factor-1 with different concentrations on the proliferation of bone marrow stem cels and to probe the best concentration. METHODS:Bone marrow stem cels from beagle dogs were culturedin vitro and stimulated by different concentrations of stromal cel derived factor-1 (100, 200, 300 μg/L). MTT was used to detect the influence of stromal cel derived factor-1 on the proliferation of bone marrow stem cels so as to screen the best concentration of stromal cel derived factor-1. Then, stromal cel derived factor-1 at the best concentrations was used to intervene the bone marrow stem cels, and MTT was used again to detect the proliferation of bone marrow stem cels. RESULTS AND CONCLUSION:Stromal cel derived factor-1 at concentrations of 100, 200, 300 μg/L could promote the proliferation of bone marrow stem cels, and the effect was more notable at 200 and 300 μg/Lbut withno significant difference. Therefore, 200 μg/L was considered to be the best concentration of stromal cel derived factor-1 for intervention of bone marrow stem cels. Compared with the blank control group, 200 μg/L stromal cel derived factor-1 could significantly promote the proliferation of bone marrow stem cels. Taken together, stromal cel derived factor-1 can promote the proliferation of bone marrow stem cels, and its best concentration is 200 μg/L.
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BACKGROUND:Cel replacement therapy as an effective strategy for reconstruction of the central nervous system has very broad application prospects. OBJECTIVE:To investigate the effect of stereotactic transplantation of neural stem cels into the brain on the neuromotor function of craniocerebral trauma rats. METHODS:Twenty male Sprague-Dawley rats were equivalently randomized into study and control groups. Animal models of craniocerebral trauma were made using the improved free-fal method in the rats. Then, model rats in the study and control groups were given parenchymal transplantation of embryonic neural stem cels and the same volume of culture medium with no stem cels at 1 day after injury, respectively. Neuromotor function of rats was assessed based on the neurological severity scores. At 2 weeks after transplantation, brain tissues were taken for hematoxylin-eosin staining, anti-BrdU, glial fibrilary acidic protein, β-tubulin III and tyrosine hydroxylase immunohistochemistry staining. RESULTS AND CONCLUSION:The neurological severity scores in the study group were significantly lower than those in the control group at 1 and 2 weeks after injury (P< 0.05). In the study group, there were many BrdU-positive neural stem cels in the brain tissues, some of which were positive for glial fibrilary acidic protein, β-tubulin III and tyrosine hydroxylase; while in the control group, there was no BrdU-positive cel in the brain tissues. Experimental findings show that neural stem cels stereotacticaly transplanted into the brain can proliferate and differentiate in the brain lesion, and thereby notably improve the neuromotor function of rats with craniocerebral trauma.
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BACKGROUND:Stem cel therapy has been used for prevention and treatment of degenerative disc disease. Considering the special microenvironment in the intervertebral disc, the survival rate and differentiation ability of transplanted cels are decreased, which may lead to the poor efficacy of stem cel therapy. How to improve the survival ability and therapeutic effect of the transplanted cels is the focus of stem cel therapy for degenerative disc disease. OBJECTIVE: To investigate the effects of cobalt chloride combined with hypertonic solution pretreatment on bone marrow mesenchymal stem cels that wil be transplanted for treatment of degenerative disc disease. METHODS:(1)In vitro cel experiment: bone marrow mesenchymal stem cels were divided into three groups and subjected to normal culture medium (normal control group), 1% hypertonic mother solution (hypertonic group), 100 μmol/L cobalt chloride (hypoxia group), or 1% hypertonic mother solution plus 100 μmol/L cobalt chloride (combined group) for 1 week. Then, 2% hypertonic solution and 200 μmol/L cobalt chloride cobalt chloride were used to simulate the anaerobic and hypertonic environment intervenes in pretreated and untreated bone marrow mesenchymal stem cels for 24 hours. After that, RT-PCR was used to detect the expression of caspase-3 for apoptosis evaluation. (2)In vivo animal experiment: Sprague-Dawley rats were divided into model, cel transplantation and hypertonic plus hypoxic groups. Rat models of intervertebral disc degeneration were made in these three groups. After modeling, rats in these three groups were given no treatment, bone marrow mesenchymal stem cel transplantation or transplantation of bone marrow mesenchymal stem cels which were subjected to hypertonic and hypoxia pretreatments into the intervertebral disc. Two weeks later, immunohistochemistry and RT-PCR methods were used to detect cel distribution and related gene expression, respectively, thereby to evaluate the therapeutic effect of stem cels. RESULTS AND CONCLUSION: (1)In vitro cel experiment: caspase-3 mRNA expression was significantly reduced in pretreated bone marrow mesenchymal stem cels compared with the untreated cels (P < 0.05). (2)In vivo animal experiment: compared with the control group, the caspase-3 and interleukin-1β in the intervertebral disc and a number of degenerative indexes were decreased in the cel transplantation. Compared with the cel transplantation group, these indicators had better outcomes in the hypertonic plus hypoxic group (P < 0.05). These findings indicate that bone marrow mesenchymal stem cels have therapeutic potential for degenerative disc disease, and have better adaptability and transplantation effects by hypertonic and hypoxia pretreatments.
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BACKGROUND:The process of oxidative stress that impacts the curative effect exists in the region which accepts cel transplantation. However, there are few reports about the effects of oxidative stress on human dental pulp stem cels and relevant mechanism. OBJECTIVE:To understand the effect of hydrogen peroxide on the senescence of human dental pulp stem cels. METHODS:Human dental pulp stem cels were isolated and cultured in PBS, 100 and 200 μmol/L hydrogen peroxide for 2 hours, respectively. Cel morphology was observed under inverted microscope, degree of cel senescence monitored by β-galactosidase staining, cel proliferation ability detected by BrdU kit and cel counting method, cytoskeleton of dental pulp stem cels and expression of sirt1 tested using immunofluorescence method, and expression of sirt1 and p16 proteins measured by western blot assay. RESULTS AND CONCLUSION:Dental pulp stem cels exhibited a fibroblast-like morphology with spindle-shaped appearance. After stimulated by hydrogen peroxide, the cel volume was enlarged, theβ-galactosidase staining deepened and the proliferation of dental pulp stem cels reduced. The enhancement of senescence of dental pulp stem cels was accompanied with the increasing concentration of hydrogen peroxide, and in this process, the expression of p16 was raised while the expression of sirt1 was decreased. In conclusion, the senescence of human dental pulp stem cels can be promoted by the stimulation of hydrogen peroxide, and sirt1 and p16 are involved in this process. Our findings may provide a theoretical and experimental foundation for autologous transplantation of dental pulp stem cels.
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BACKGROUND:Isoflurane cannot only induce a wide range of large neuronal apoptosis, but also inhibit hippocampal neurogenesis in neonatal rats, thereby resulting in hippocampus-dependent learning and memory defects. OBJECTIVE:To investigate the isoflurane effect on proliferation and differentiation of the hippocampal neural stem cels. METHODS:Twenty-six Sprague-Dawley rats were randomly divided into air group and isoflurane group (n=13 per group). Rats in the isoflurane group were subjected to 2.5% isoflurane inhalation for 3 minutes folowed by 1.5% isoflurane inhalation for 4 hours. Rats in the air group only breathed in air. After the intervention, blood glucose and arterial blood gas changes were detected in the two groups. Additionaly, rats in the two groups were given intraperitoneal injection of 5-bromodeoxyuridine before and after intervention. At 24 hours after the last injection of 5-bromodeoxyuridine, brain tissues were taken to make frozen sections for immunofluorescence staining. RESULTS AND CONCLUSION:There were no significant difference in pH, PaO2, PaCO2, HCO3, BE and SaO2 levels between the two groups (P> 0.05). Compared with the air group, the number of BrdU+ cels was significantly less in the isoflurane group (P < 0.05), while the number of NeuroD+/BrdU+ cels was significantly higher in the isoflurane group (P < 0.05). The incidence of adverse reactions was 23% in the isoflurane group, which was significantly higher than that in the air group (7.7%;P < 0.05). These findings indicate that isoflurane can inhibit the proliferation of neural stem cels in the hippocampal dentate gyrus, and promote their differentiation into neurons.
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Abstract BACKGROUND: Alveolar bone deficiency wil not meet aesthetic and functional requirements for dental implants. OBJECTIVE:To observe the repair effect of passage 3 autologous bone marrow mesenchymal stem cels (BMSCs) and platelet-rich fibrin (PRF) on alveolar bone defects in rabbits. METHODS:Twenty-seven New Zealand rabbits were randomly divided into BMSCs/PRF group, PRF group and model group (n=9 per group). The left mandible incisors were extracted in al the rabbits under general anesthesia. BMSCs/PRF group was immediately implanted BMSCs/PRF composite into the alveolar socket, PRF group only implanted PRF, and model group implanted nothing. RESULTS AND CONCLUSION: In the model group, the alveolar crest and alveolar mucosa become sunken notably and narrowed. In the BMSCs/PRF and PRF groups, the thickness of alveolar bone wal, alveolar bone width, alveolar bone height difference, and bone mineral density were al increased, especialy in the former group. In addition, the trabecular arrangement was better in the BMSCs/PRF groups than the model and PRF group. Our findings indicate that alveolar socket filing with composite of BMSCs and PRF can achieve preservation of alveolar bone width and height after tooth extraction in rabbits.
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BACKGROUND:Ankylosing spondylitis is an autoimmune disease at high inflammatory state, and its pathogenesis is stil unclear. Besides, there is a lack of entirely satisfactory curative strategies. OBJECTIVE: To explore the immunoregulation capability of bone marrow mesenchymal stem cels from ankylosing spondylitis patients on macrophages and the potential therapeutic use of bone marrow mesenchymal stem cels from healthy donors on ankylosing spondylitis. METHODS: Bone marrow mesenchymal stem cels were extracted from 21 healthy donors and 25 ankylosing spondylitis patients respectively, and passage 4 cels were used in subsequent experiments. A human monocytic cel line was induced to differentiate into macrophages. The phenotypic markers of bone marrow mesenchymal stem cels and macrophages were detected by flow cytometry. Expressions of tumor necrosis factor-α and tumor necrosis factor-α-stimulated gene 6 (TSG-6) proteins in the supernatant of co-culture system were detected by ELISA. Quantitative real-time PCR was applied to detect the mRNA level of cytokines secreted by bone marrow mesenchymal stem cels and macrophages. RESULTS AND CONCLUSION:The typical mesenchymal stem cel surface markers were expressed in both bone marrow mesenchymal stem cels from healthy donors and patients with ankylosing spondylitis, and CD68 was detected positively in induced macrophages. The protein and mRNA levels of tumor necrosis factor-α secreted by macrophages co-cultured with bone marrow mesenchymal stem cels from patients with ankylosing spondylitis were obviously higher than those from healthy donors (P < 0.05). TSG-6 secreted by bone marrow mesenchymal stem cels from patients with ankylosing spondylitis was lower than that by bone marrow mesenchymal stem cels from healthy donors in both RNA transcriptional and protein levels (P < 0.05). Our study demonstrates that bone marrow mesenchymal stem cels from patients with ankylosing spondylitis shows abnormal immunoregulatory function on inhibiting the tumor necrosis factor-α secretion from macrophages, which reveals a mechanism of immune disorder in ankylosing spondylitis. The therapeutic mechanism of bone marrow mesenchymal stem cels from healthy donors may work by secreting enough TSG-6 to inhibit the activation of macrophages in patients with ankylosing spondylitis, and thereby to decrease the secretion of tumor necrosis factor-α. Cite this article:Sun SH, Wang P, Su CY, Xie ZY, Li YX, Li D, Wang S, Su HJ, Wu XH, Deng W, Wu YF, Shen HY. Bone marrow mesenchymal stem cels derived from patients with ankylosing spondylitis show abnormal immunoregulation capability on macrophages. Zhongguo Zuzhi Gongcheng Yanjiu. 2016;20(1):13-19.
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BACKGROUND:Effective sorting of prostate cancer stem cels is the basis of experimental studies in prostate cancer developing. The most common sorting method is magnetic-activated cel sorting. OBJECTIVE:To separate CD133+/CD44+ cels in prostate cancer tissues using self-designed magnetic beads folowed by culture, passage and immunological identification. METHODS:Self-designed magnetic microspheres were applied to establish immunomagnetic beads to sort CD133+/CD44+ cels in prostate cancer tissues. The sorted cels were cultured in serum-free medium. The sphere formation, cel morphology, and proliferation ability after cel passage were statisticaly compared between the sorted cels and the normal tumor cel lines. Immunofluorescence detection was performed to detect the expression of specific antibodies. RESULTS AND CONCLUSION:Self-designed immunomagnetic beads had smal diameter and a high-sorting effect. The sorted cels possessed a high capacity of microsphere formation. After cel culture and passage, the cels highly expressed CD133 and CD44 antigens. The sorted cels with no induction had varying shapes and grew vigorously. After induction with transforming growth factor-β, the cultured cels were noted to have a single shape and grow slowly. The cel proliferation ability of sorted cels in these two groups differed significantly from that of the normal cancer cel lines (bothP < 0.05). In conclusion, the CD133+/CD44+ cels sorted from prostate tumor cels possessed cel morphology and function characteristics of stem cels which can provide a basis for extraction and culture of prostate cancer stem cels. Cite this article:Gong R, Li SY, Huo ZX, Ding H, Sun EL. Extraction of prostate cancer stem cels using self-designed magnetic beads. Zhongguo Zuzhi Gongcheng Yanjiu. 2016;20(1):36-41.
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Abstract BACKGROUND:Previous studies have found that cryptotanshinone represses multiple tumors, but little is reported on its effect on renal carcinoma. OBJECTIVE:To explore the effect of cryptotanshinone on the proliferation and apoptosis of the renal carcinoma stem cels. METHODS:CD133+ renal carcinoma stem cels were separated from OS-RC-2 cels by immunomagnetic bead separation. Effects of 0, 0.2, 1, 5 mg/L cryptotanshinone on the proliferation and apoptosis of CD133+ renal carcinoma stem cels were detected by MTT and flow cytometry, respectively. Expression levels of Ki67, Bcl-2, Caspase-3 and p-Caspase-3 protein were detected by western blot assay. RESULTS AND CONCLUSION:After magnetic cel sorting, the percentage of CD133+ cels was increased from 6.32% to 82.73%, and there was a significant difference before and after cel sorting (P < 0.001). Cryptotanshinone could repress the proliferation of CD133+ renal carcinoma stem cels and promote cel apoptosis in a dose-dependent manner. The protein expression levels of Ki67 and Bcl-2 in the 5 mg/L cryptotanshinone group were significantly decreased compared with the control group, while the protein expression level of p-Caspase-3 protein was significantly increased. In addition, there was no difference in the protein expression of Caspase-3 between cryptotanshinone and control group. These findings indicate that cryptotanshinone may be a potent anticancer drug for the treatment of renal carcinoma by inhibiting expression of Ki67 and Bcl-2 and promoting protein expression of p-Caspase-3. Cite this article:Feng M, Jia MH. Cryptotanshinone effects on the proliferation and apoptosis of renal carcinoma stem cels. Zhongguo Zuzhi Gongcheng Yanjiu. 2016;20(1):49-54.
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BACKGROUND:In 2001, Zuk et al found adipose-derived stem cels (ASCs) from the aspirate of liposuction for the first time, which launched a new era of stem cel research. In recent years, stem cels have been proved to widely exist in many tissues and organs. ASCs are always in the spotlight of plastic and reconstructive surgery, tissue engineering and regenerative medicine because of extensive sources and simple isolation. OBJECTIVE: To review the fat tissue harvesting and ASCs isolation, purification, expansion, and cryopreservation, to discuss the main factors which influence the yield, proliferation capacity and differentiation potential of ASCs, and to predict the future research interests based on current issues. METHODS:On September 10th, 2015, relevant articles were searched in PubMed using the folowing format: (adipose stem cels[Title]) OR (adipose-derived stem cels[Title]) OR (adipose-derived mesenchymal stem cels[Title]) and in SinoMed using the folowing format in Chinese: (“adipose-derived stem cels” [Title])or(“adipose-derived mesenchymal stem cels”[Title]). Finaly, 81 representative articles were included according to their titles and abstracts. In this review, we also introduced relevant experience about the aforementioned procedures from the Department of Plastic Surgery and Tissue Regeneration and Molecular Cel Engineering Lab of University of Texas, MD Anderson Cancer Center, USA. RESULTS AND CONCLUSION:The widely dispersed fat tissues potentialy provide abundant stem cels for tissue engineering and regenerative medicine. Liposuction is a mini-invasive approach for harvesting fat tissues. Colagenase digestion is the major method for ASCs isolation due to its simplicity and high yield in basic research. However, clinical fat transplantation without ASCs isolation or non-colagenase isolation of stromal vascular fraction or ASCs is preferred. The phenotype, proliferation and differentiation capacity of ASCs may be affected by several factors during the fat tissue harvesting and ASCs isolation. Therefore, a standard protocol for ASCs isolation is needed.
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BACKGROUND:Studies have shown that cancer stem cels play an important role in tumor invasion and metastasis, but studies on the role of gastric cancer stem cels in invasion and metastasis of gastric cancer cels were rarely reported. OBJECTIVE:To study the effect of gastric cancer stem cels CSC-G on invasion and metastasis of gastric cancer cels. METHODS: Gastric cancer stem cels CSC-G and gastric cancer cels SGC7901 were culturedin vitro for 10 days folowed by spherical colony formation assay, western blot assay for detecting OCT4, SOX2, E-cadherin and CD44 protein expression levels in gastric cancer stem cels CSC-G and gastric cancer cels SGC7901, and Transwel assay for detecting the invasion and migration of gastric cancer stem cels and gastric cancer cels SGC7901. RESULTS AND CONCLUSION:Gastric cancer cels SGC7901 in RPMI1640 medium presented with adherent growth and were quadrilateral or polygonal after passage; gastric cancer stem cels CSC-G in serum-free medium presented with suspension growth, and adherent gastric cancer stem cels were spindle-shaped or round. Compared with gastric cancer cels SGC7901, the protein expressions of OCT4, SOX2 and CD44, the number of cancer cel spheres and the number of trans-membrane cels were significantly increased in the gastric cancer stem cels CSC-G (P < 0.05), and the expression of E-cadherin protein was significantly decreased (P < 0.05). These findings indicate that the gastric cancer stem cels CSC-G can be successfuly cultured in vitro, and have enhanced invasion and migration compared with the gastric cancer cels SGC7901, which play an important role in gastric cancer invasion and metastasis.
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BACKGROUND:Stem cels from the apical papila are a new kind of mesenchymal stem cels, and whether it can be used in root regeneration is the key to the present study. OBJECTIVE:To culture rat stem cels from the apical papila and periodontal ligament stem celsin vitro, and to compare the biology behaviors of these two kinds of cels, thereby providing experimental basis for the application of stem cels from the apical papila in root regeneration. METHODS:The apical papila, as wel as the periodontal ligament tissues from the healthy mandibular teeth of young rats were digested and cultured. Immunophenotypes of stem cels from the apical papila and periodontal ligament stem cels were detected by immunofluorescence technique. Then, cel growth curves were determined by MTT method and mineralized nodule formation was observed by alizarin red staining. RESULTS AND CONCLUSION:Stem cels from the apical papila and periodontal ligament stem cels were both positive for STRO-1. Stem cels from the apical papila were positive for CD90 and weakly positive for CD146. Periodontal ligament stem cels were positive for CD146 and weakly positive for CD90. The absorbance values of stem cels from the apical papila and periodontal ligament stem cels increased with the increasing of time and became stable at 8 days. Since the 4th day, the proliferation capacity of stem cels from the apical papila was significantly stronger than that of periodontal ligament stem cels (P < 0.05). Both of stem cels are visible to have mineralized nodule formation. Compared with the periodontal ligament stem cels, stem cels from the apical papila were stained obviously deeper and had more mineralized nodules. These results show that stem cels from the apical papila have stronger proliferation capacity and mineralization ability than periodontal ligament stem cels. Cite this article:Zhao L, Yu L, Yuan P, Zhou CM, Wu PL.Stem cels from the apical papila versus periodontal ligament stem cels: biological behaviors. Zhongguo Zuzhi Gongcheng Yanjiu. 2016;20(1):113-117.
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BACKGROUND:Isoflurane is an anesthesia drug that has a certain effect on the nervous system. It possibly causes neurologic disorders through impacting nerve stem cel function or morphology. OBJECTIVE:To investigate the effects of isoflurane on the proliferation and differentiation of neural stem cels in the hippocampus of rats. METHODS:Neural stem cels from the hippocampus of neonatal Sprague-Dawley rats, aged 7 days, were induced and differentiated. Passage 3 cels were obtained and divided into two groups: isoflurane group (a mixture gas of 2.8% isoflurane, 5% CO2 and 95% O2) and control group (a mixture of 5% CO2 and 95% O2). After intervention of 6 hours folowed by 2 hours of routine culture, anti-BrdU monoclonal antibody immunofluorescent staining was used to detect cel proliferation, and western blot assay to detect the expression of β3-tubulin and glial fibrilary acidic protein. RESULTS AND CONCLUSION:Compared with the control group, the number of BrdU positive cels in the isoflurane group reduced significantly, indicating that isoflurane inhibits the proliferation of neural stem cels. Compared with the control group, the expression of glial fibrilary acidic protein in the isoflurane group up-regulated, but the expression of β3-tubulin had no changes, indicating isoflurane promotes the differentiation of neural stem cels into astrocytes. Cite this article:Min N, Hu QF, Li XP, Nie XH, Yang LL.Isoflurane effects on the proliferation and differentiation of neural stem cels in the hippocampus of neonatal rats. Zhongguo Zuzhi Gongcheng Yanjiu. 2016;20(1):118-122.
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BACKGROUND:It wil provide a new insight into the future application of bone marrow mesenchymal stem cels in the treatment of spinal cord injury and tissue engineering by studying the effect of activation of Wnt signaling pathway in the neuronal differentiation of bone marrow mesenchymal stem cels. OBJECTIVE: To detect the expression of related genes by gene chip technology during the neuronal differentiation of bone marrow mesenchymal stem cels. METHODS:Human bone marrow mesenchymal stem cels were isolated and purified, and passage 5 cels were obtained. GatewayTM technology was used to build lentiviral vectors that was used to transfect Wnt-1 into human bone marrow mesenchymal stem cels. Control, non-transduction and transduction groups were set in this study. Human bone marrow mesenchymal stem cels were then induced to differentiate into neurons. Cel morphology was observed under inverted phase contrast microscope. Gene chip was used to detect the regulation changes and the differential expression of related genes in the Wnt signaling pathway. RESULTS AND CONCLUSION: Under the scanning electron microscope, the transfected cels were found to have the similar morphology of neuron-like cels. Analysis by the gene chip hybridization technique showed that 3 287 genes were up-regulated and 4 215 genes were down-regulated in the signal pathway. In the Wnt signaling pathway, genes related to the nervous system development and differentiation were up- or down-regulated. It is verified that the Wnt signal pathway is activated via Wnt-1 transduction, and the downstream genes appear to have genetic transcription so as to promote the neuronal differentiation of human bone marrow mesenchymal stem cels.
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BACKGROUND:Retinoic acid is the most promising inducer for neuroblastoma minimal residual lesion, and it can induce cel differentiationin vivo, accompanied by reducing tumor cel proliferation. OBJECTIVE:To study the effect of nanoparticle labeling on biological characteristics of neuroblastoma stem cels, and the role of 13-cis retinoic acid to induce differentiation of neuroblastoma stem cels. METHODS:Neuroblastoma stem cels were isolated and culturedin vitro using serum-free suspension culture method, labeled with polylysine-modified γ-Fe2O3 nanoparticles and induced in culture medium containing 13-cis retinoic acid. RT-PCR was used to detect the expression of Oct-4 before and after labeling as wel as before and after induction. Immunofluorescence method was used to detect the expression of nestin before and after labeling as wel as before and after induction. RESULTS AND CONCLUSION:Neuroblastoma stem cels were successfuly cultured in the bone marrow samples from 5 of 20 cases. Polylysine-modified γ-Fe2O3 nanoparticle labeling did no influence the viability and proliferation ability of neuroblastoma stem cels, and also had no effect on Oct-4 mRNA and nestin expression. After cultured in the culture medium containing 13-cis retinoic acid, the cel shape changed and the growth rate slowed down. Moreover, the expression of Oct-4 mRNA and nestin was gradualy reduced. These findings indicate that polylysine-modified gamma-Fe2O3 nanoparticles can be used to label neuroblastoma stem cels, and 13-cis retinoic acid can induce the differentiation of neuroblastoma stem cels.