Stem cells combined with three-dimensional scaffolds for the treatment of osteoporotic bone defects北大核心

BACKGROUND: Stem cells combined with a three-dimensional scaffold have great potential for the treatment of osteoporotic bone defects. OBJECTIVE: To introduce the application of stem cells combined with the three-dimensional scaffold in repairing osteoporotic bone defects. METHODS: A computer-based search of PubMed, Web of Science, Springerlink, Medline, WanFang and CNKI databases was performed for relevant articles published from 2007 to 2017 with “stem cells, scaffold, osteoporosis, bone defects” as key words in English and Chinese, repsectively. Initially, 142 articles were retrieved, and finally 45 articles were included in result analysis. RESULTS AND CONCLUSION: Due to the potential of self-renewal and multilineage differentiation, stem cells can be used to repair or regenerate damaged tissues, which are considered as an ideal cell source for the treatment of diseases in orthopedics. The suitable scaffold can provide a favorable microenvironment for repairing the attachment and growth of the cells related to the bone defect, which can promote the healing without additional side effects. Furthermore, stem cells combined with three-dimensional scaffolds provide a promising clinical application for the treatment of osteoporotic bone defects by regulation of bone metabolism. In addition, gene-modified stem cells with three-dimensional scaffolds bring a huge potential in the treatment of osteoporotic bone defects. In conclusion, the combination of stem cells and three-dimensional scaffolds provides a new approach for the treatment of osteoporotic bone defects, which may be one of the future therapeutic strategies. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Adipose tissue engineering construction using adipose-derived stem cells and scaffolds in breast reconstruction北大核心

BACKGROUND: The development of adipose tissue engineering has solved a series of problems and brought new ideas for breast reconstruction. OBJECTIVE: To summarize the main biological characteristics of scaffolds, seed cells and adipose grafts for adipose tissue engineering construction, as well as their application prospects in breast reconstruction. METHODS: With the key words of “scaffold, mesenchymal stem cells, adipose tissue engineering, breast reconstruction, vascularization” in Chinese and English, respectively, a retrieval of CNKI and PubMed was performed for relevant articles published from 1999 to 2017. After initial screening, the eligible articles were further detailed, analyzed and summarized. RESULTS AND CONCLUSION: Scaffold materials with low immunogenicity, especially the decellularized adipose tissue, have a unique advantage in adipogenic induction that can be used to support the regeneration of human organs and tissues. Adipose-derived stem cells can also be used as the priority for adipose tissue engineering construction, which can promote tissue development and regeneration in vivo. However, much less is known about the use of adipose tissue engineering in breast reconstruction and the implementation of long-term stable growth of the adipose tissues. Further investigations are indispensable on how to realize breast reconstruction using tissue-engineered adipose so as to construct and maintain a stable structure entity throughout the entire patient’s life. Therefore, strict selection of seed cells, scaffolds and cytokines is considered pivotal in the construction of cell microenvironment. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Bone marrow mesenchymal stem cells combined with acellular dermal matrix for repair of urethral injury北大核心

BACKGROUND: In recent years, the development of tissue engineering provides more choices for the repair of urethral injury. OBJECTIVE: To investigate the effect of bone marrow mesenchymal stem cells (BMSCs) combined with acellular dermal matrix in the repair of urethral injuries. METHODS: Passage 3 BMSCs from New Zealand white rabbits were inoculated on the acellular dermal matrix to construct tissue-engineered urethra grafts. Thirty-six New Zealand rabbits were randomized into three groups (n=12 per group). Experimental group was implanted with BMSCs-acellular dermal matrix complex at urethral injury. Control group was implanted with acellular dermal matrix material at urethral injury. Normal group had neither injury nor treatment. At postoperative 4, 8 and 12 weeks, the repaired urethral tissue was subjected to hematoxylin-eosin staining. At postoperative 12 weeks, immunohistochemical staining and urodynamic study were performed. RESULTS AND CONCLUSION: At postoperative 4 weeks, thin-layer epithelial regeneration was visible in the urethra defect area of the experimental group, and the continuity was better. The urethra mucosa of the control group was discontinuous. At postoperative 8-12 weeks, the urethral epithelial layer in the experimental group became thickened, exhibiting a good continuity with the normal urethral epithelium, thickened mucosa, and smooth and continuous urethral mucosa; the regenerated urethral mucosa of the control group exhibited good continuity, but less regenerated epithelial layers. At postoperative 12 weeks, immunohistochemical results showed the repaired urethra in the experimental group was positive for uroplakin illa, CK AE1/AE3, and α-smooth muscle actm. The maximum urethral pressure in the experimental group showed no significant difference before and after operation, while the postoperative pressure in the control group showed a significant increase (P < 0.05). Overall findings indicate that the combination of BMSCs and acellular dermal matrix has better efficacy than the acellular dermal matrix alone. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Isolation and identification of human adipose-derived stem cells and its exosomes北大核心

BACKGROUND: Currently, mesenchymal stem cells have been widely explored and applied in scientific research field, and many studies suggest that the underlying mechanism of mesenchymal stem cells mainly relies on its exosomes. OBJECTIVE: To isolate and identify human adipose-derived stem cells and its exosomes, and to identify their biological characteristics. METHODS: Human adipose tissue was digested with collagenase I, and adipose-derived stem cells were isolated and purified. Immunophenotype, osteogenic and adipogenic abilities of adipose-derived stem cells were identified. Exosomes were isolated by using ultrafiltration method. Morphology of exosomes was observed by Nanosight and electron microscope. Expression of proteins in exosomes was detected by antibody array method. RESULTS AND CONCLUSION: Adipose-derived stem cells exhibited long spindle-like or fibroblast-like appearance, expressed CD73, CD44, CD90, CD105 and had the potential to differentiate into many tissues, including bone and adipose tissues. The exosomes had the similar size, with the diameter of 30-150 nm. They possessed many proteins including FLOT1, ICAM, ALIX, CD81, CD63, ANXA5, TSG101, and so on. Findings from the present study indicate the successful isolation of exosomes from human adipose-derived stem cells. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Application of graphene-based nanomaterials in stem cells北大核心

BACKGROUND: As a kind of newly-developing nanomaterial, graphene has been used in many fields. Many recent studies have found that graphene-based nanomaterials can affect the biological behaviors of stem cells. OBJECTIVE: To review the application and progress of graphene-based nanomaterials in stem cells. METHODS: We searched the articles about the application of graphene-based nanomaterials in stem cells published in PubMed, Web of Science, and CNKI databases with the search terms “graphene, nanomaterials, stem cell” in English and Chinese. Finally, 57 articles met the criteria for review. RESULTS AND CONCLUSION: Graphene-based nanomaterials have good stability and corrosion resistance, high mechanical strength, good biocompatibility, which are accepted as one of the most promising nanomaterials in biomedicine. Stem cells are undifferentiated cells that can differentiate into various mature cells in human body, which have a broad application prospect in tissue engineering, regenerative medicine and other fields. Many recent studies have applied graphene-based nanomaterials to stem cell research and found that they can affect the growth, proliferation, adhesion and differentiation of stem cells, and these nanomaterials may affect the biological behavior of stem cells by regulating the expression of related genes and various signaling pathways. However, graphene-based nanomaterials have biological toxicity, which restrict their application in biological aspects. Moreover, most researches only involved cellular level, and it needs further animal studies and in vivo experimental researches. © 2022, Publishing House of Chinese Journal of Tissue Engineering Research. All rights reserved.     

Coculture with mesenchymal stem cells facilitates the proliferation of hematopoietic stem cells under different coculture modes北大核心

BACKGROUND: Although a large number of related studies have been carried out, there is still a lack of practical methods to amplify hematopoietic stem cells (HSCs) in vitro. Mesenchymal stem cells (MSCs) secrete a variety of cytokines that promote the HSCs proliferation and inhibit their differentiation. These cytokines play an important role in maintaining the hematopoietic microenvironment and regulating HSCs function. OBJECTIVE: To investigate the effect of bone marrow MSCs on the proliferation of HSCs in vitro under different coculture modes. METHODS: Mesenchymal stem cells from the bone marrow of C57BL/6 mice were cultured in vitro using the whole bone marrow adherent culture. CD117+ cells (HSCs) were sorted from passage 3 cells by using miniMACS magnetic beads sorting. Then, CD117+ cells were co-cultured with MSCs under different coculture models, including single culture of HSCs (control group), Transwell coculture (upper chamber, HSCs; lower chamber, MSCs) and two-dimensional contact coculture (coculturing HSCs and MSCs in 24-well platts). The morphology of HSCs was observed under phase contrast microscope and fluorescence microscope, and the number of active cells of HSCs was counted at 1,3,5, and 7 days after coculture. RESULTS AND CONCLUSION: During the coculture of 1-7 days, the number of HSCs in the two groups was increased with culture time (P < 0.05). After 3 days of coculture, HSCs in each group was grown into the logarithmic growth phase, and morphological changes in some HSCs were detected at 5 days of coculture. Ait 7 days of coculture, the viabilities of HSCs in different culture models were ranked as follows: single culture model < Transwell coculture model < two-dimensional contact coculture model (P < 0.05). These findings suggest that MSCs can effectively promote the proliferation of HSCs in vitro, and the promotion effect is increased under contact coculture conditions. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Application and characteristics of bone graft materials in the treatment of spinal tuberculosis北大核心

BACKGROUND: Thorough removal of local necrotic lesions and one-stage use of bone repair materials can significantly promote local bony fusion, avoid recurrence of tuberculosis in the middle and long terms and reconstruct spinal stability in the surgical treatment of spinal tuberculosis. OBJECTIVE: To review the application of bone graft materials in the treatment of spinal tuberculosis. METHODS: The first author searched the articles related to bone graft materials of spinal tuberculosis in Bailian, CNKI, and Natures databases published from 2001 to 2020. The priority was the articles published recently or in authoritative journals. The search keywords were “bone graft materials, bone tissue engineering; spinal tuberculosis; titanium mesh; autogenous bone” in Chinese and English. RESULTS AND CONCLUSION: At present, bone graft materials have been widely used in clinic, but each has its own disadvantages. For example, the amount of autologous bone is limited, and the transplantation of autologous bone will cause bleeding and potential complications of donor site; allogeneic bone will lead to delayed healing and infection; titanium mesh has the problems of postoperative subsidence and kyphosis correction angle loss; the organic polymer materials such as polylactic acid and polymethyl methacrylate are lack of bone induction performance. Although Ca/P-based ceramic materials can be used as carrier materials of antituberculosis drugs, their biomechanical properties cannot fully meet the clinical needs. In view of the shortcomings of the above materials, it is necessary to find a composite bone tissue engineering material, which can meet the requirements of good biocompatibility, mechanical properties, degradation properties, osteogenic activity, and drug release performance. © 2022, Publishing House of Chinese Journal of Tissue Engineering Research. All rights reserved.     

Polydatin loaded collagen/heparin sulfate scaffold in the treatment of spinal cord injury in rats北大核心

BACKGROUND: At present, the research on the neuroprotective effect of polydatin has gradually become a hot spot in the field of neurology. The research direction is mostly focused on ischemic cerebrovascular disease, and there are few studies on the application of spinal cord injury. OBJECTIVE: With collagen/heparin sulfate scaffold as carrier, polydatin was applied to the injured spinal cord to observe the repair effect. METHODS: (1) Collagen/heparin sulfate scaffolds and collagen/heparin sulfate scaffolds loaded with 0.5, 1, 1.5 mmol/L polydatin were prepared. The third generation of rat neural stem cells was seeded on four kinds of scaffolds, and the proliferation of cells was detected by CCK-8 assay. During inducing neural differentiation of neural stem cells, the expression levels of glial fibrillary acidic protein, Tuj-1 and Oligo were detected by immunofluorescence staining and RTPCR. (2) The spinal cord injury model of adult male SD rats was established and divided into three groups. The model control group (n=10) was not implanted with any materials. The control group (n=10) was implanted with collagen/heparin sulfate scaffold, and the experimental group (n=10) was implanted with 1 mmol/L polydatin/collagen/heparin sulfate scaffold. Meanwhile, sham operation group (n=10) was set up. BBB score was used to test the motor function of the right hind limb within 8 weeks after operation. At 8 weeks after operation, the spinal cord tissues were taken for histological observation, immunohistochemical analysis, and western blot assay. RESULTS AND CONCLUSION: (1) At 3 and 7 days of culture, the absorbance value of cell proliferation on 1 mmol/L polydatin/collagen/heparin sulfate scaffold was higher than that on the other three scaffolds (P < 0.05). (2) Immunofluorescence staining 7 days after induction showed that the expression of glial fibrillary acidic protein in 1 and 1.5 mmol/L polydatin/collagen/heparin sulfate scaffold groups were less than those in the other two collagen/heparin sulfate scaffold groups, but the expression levels of Oligo and Tuj-1 were more than those in the other two collagen/heparin sulfate scaffold groups. (3) RT-PCR results showed that the expression of glial fibrillary acidic protein mRNA in 1 and 1.5 mmol/L polydatin/collagen/heparin sulfate scaffold groups was lower than that in the other two collagen/heparin sulfate scaffold groups (P < 0.05). There was no significant difference in the expression of Tuj-1 mRNA among the four groups. The expression of Oligo mRNA in 1 mmol/L polydatin/collagen/heparin sulfate scaffold group was higher than that in the other three groups (P < 0.05). (4) Spinal cord injury repair experiments showed that the BBB score of the experimental group was always higher than that of the model control group and the control group 2-8 weeks after operation (P < 0.05). Hematoxylin-eosin staining showed that the spinal cord defects in the control group and the observation group were filled with scaffolds, and the space between the tissues was smaller than that in the model control group. The tissue continuity and space in the observation group were better than those in the control group. Immunohistochemical analysis and western blot assay showed that the expression of neurofilament-200 protein in the experimental group was higher than that in the control group and model control group (P < 0.05). The expression of glial fibrillary acidic protein in the experimental group was lower than that in the control group and model control group (P < 0.05). (5) It is concluded that polydatin loaded collagen/heparin sulfate scaffolds can promote the repair of spinal cord injury. © 2022, Publishing House of Chinese Journal of Tissue Engineering Research. All rights reserved.     

Important role of glutathione in stemness and regulation of stem cells北大核心

BACKGROUND: Stem cells possess the capacities of self-renewal and multiple differentiation. Glutathione, an important sulfhydryl compounds, not only participates in the active oxygen metabolism of stem cells, but also plays an important role in self-renewal, proliferation, aging, stemness maintenance and differentiation regulation of stem cells. OBJECTIVE: To explore the important role of glutathione in stem cells. METHODS: In April 2020, the first author searched the title/abstract with the English keywords of “glutathione or GSH, stem cell”, and searched any field with the Chinese keywords of “glutathione, stem cell”, and searched the related articles included in CNKI, VIP, Wanfang and PubMed databases from 2000 to 2020. A total of 358 Chinese articles and 405 English articles were retrieved. Finally, 86 eligible articles were enrolled for the analysis after deleting the repetitive and non-conforming articles. RESULTS AND CONCLUSION: By reviewing the recent studies on glutathione and stem cells, we found that glutathione played an important role in maintaining stemness, regulating the differentiation of stem cells. In addition, effects of glutathione on cancer stem cells have been verified, which provides more evidence for the future treatment of cancer with reduced glutathione. Key words:. © 2022, Publishing House of Chinese Journal of Tissue Engineering Research. All rights reserved.     

Repair of rabbit radial bone defects using bone morphogenetic protein-2 combined with 3D porous silk fibroin/β-tricalcium phosphate hybrid scaffolds

Our study aimed to investigate the effect of bone morphogenetic protein-2 (BMP-2) bound to silk fibroin and β-tricalcium phosphate (SF/β-TCP) hybrid on the healing of critical-size radial defects in rabbits. A 15-mm critical-size defect was induced at mid-diaphysis in the left radius of 20 New Zealand white rabbits (average age, 3.5 months; weight, 2.5–3.0 kg). The animals were randomized into Group 1 (SF/β-TCP combined with BMP-2), Group 2 (SF/β-TCP alone), and Group 3 (nothing implanted). Radiographs were obtained every 2 weeks and euthanasia was performed after 8 weeks for visual, radiological, micro-computed tomography (micro-CT), and histological studies. Eight weeks after implantation (SF/β-TCP combined with BMP-2), radiographs showed that new bone formed on the surface of the implant and had bridged the defect in Group 1. Micro-CT imaging also confirmed the formation of new bone around the implant, and the newly formed bone was quantified. Histological examination revealed newly formed bone in the implanted area. Meanwhile, there was no formation of new bone in Group 3. Among the groups, most active formation of new bones was found in Group 1, while there was no difference between Group 2 and Group 3. Based on these results, we concluded that BMP-2-SF/β-TCP showed significant improvement in healing of critical-size defects. Therefore, the combination of BMP-2 and SF/β-TCP would be useful in the field of bone tissue engineering.     

Platelet-rich plasma combined with naringin induces osteogenic differentiation of human bone marrow mesenchymal stem cells In vitro北大核心

BACKGROUND: Platelet-rich plasma (PRP) and naringin can both promote proliferation and induce osteogenic differentiation of mesenchymal stem cells. However, their combined use is rarely reported. OBJECTIVE: To observe the effect of PRP combined with naringin on the osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) in vitro. METHODS: BMSCs at passage 3 were divided into four groups: (1) blank control group, cells were cultured in α-MEM; (2) PRP group, cells were cultured in α-MEM containing PRP; (3) naringin group, cells were cultured in α-MEM containing naringin; and (4) combined group, cells were cultured in a-MEM containing PRP and naringin. The contents oO used PRP and naringin were 12.5% and 50 µg/L respectively. Cell proliferation was detected by MTT assay. Expression oO related genes in hBMSCs was detected by RT-PCR. Alkaline phosphatase staining, collagen type I immunohistochemical staining, and alizarin red staining were used to analyze the osteogenic differentiation of hBMSCs. RESULTS AND CONCLUSION: The proliferation of hBMSCs was increased in each group, especially in the combined group. Cells in all the groups oxcept the blank control group were positive for alkaline phosphatase staining, collagen type I immunohistochemical staining, and alizarin red staining, and the positive effect was more obvious in the combined group. However, negative or weakly positive response was found in the blank control group. At 7 and 14 days, the expression of alkaline phosphatase and collagen type I was significantly higher in the PRP, naringin and combined groups than the blank control group (P < 0.05); at 14 days, the expression of alkaline phosphatase and collagen type I was significantly higher in the combined group than the PRP and naringin groups (P < 0.05). To conclude, PRP combined with naringin can promote the proliferation of hBMSCs and induce the osteogenic differentiation of hBMSCs. Moreover, there is a synergistic effect between PRP and naringin. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Complications and death causes of peripheral blood stem cell transplantation in the treatment of thalassemia major北大核心

BACKGROUND: Allogeneic hematopoietic stem cell transplantation is the only effective method to cure thalassemia major. However, the risk factors influencing the prognosis of transplantation remain unclear. OBJECTIVE: To analyze the distribution of common complications and cause of death after allogeneic hematopoietic stem cell transplantation in children with thalassemia major, and to explore the prognostic factors so as to provide references for improving the survival rate of patients. METHODS: This retrospective cohort study included 257 patients with β-thalassemia major who underwent allogeneic hematopoietic stem cell transplantation at Department of Hematology and Oncology, Guangzhou Women and Children Medical Center between September 2013 and September 2019. There were 172 males and 85 females, with a median age of 6 years at the time of transplantation. The basic clinical data before transplantation and complications after transplantation were compared between the surviving group and dead group using single factor. The overall survival rate was analyzed by the Kaplan-Meier method, and the overall survival rate was compared by the Log-rank test. Multivariate Cox regression was used to analyze factors affecting survival. RESULTS AND CONCLUSION: The median follow-up time was 29 months, and no cases were lost to follow-up. (1) Univariate analysis results showed that there were significant differences in the risk classification between surviving patients and dead patients (P=0.033). Patients with higher risk class had an increased risk of death after transplantation. Multivariate analysis showed that risk classification and severe pneumonia were independent risk factors for overall survival after thalassemia major hematopoietic stem cell transplantation (P < 0.05). (2) Among 20 dead patients, there were 13 patients with severe pneumonia and respiratory failure, 2 patients with grade IV intestinal graft-versus-host disease, 1 patient with intracranial hemorrhage due to thrombocytopenia, 1 patient with thrombocytopenia with acute pulmonary hemorrhage, 1 patient with sepsis with shock, and 1 patient with myasthenia gravis. (3) Totally 17 cases died within 1 year after transplantation, while the rest 3 patients died of severe pneumonia more than 1 year after transplantation. (4) The incidence of bronchiolitis obliterans was significantly higher in patients who died after transplantation than in those who survived (P < 0.000 1). (5) It is concluded that the important factors that affect survival rate in thalassemia major patients after hematopoietic stem cell transplantation are infection and severe graft-versus-host disease. The risk of death increased in patients with bronchiolitis obliterans after transplantation. © 2023, Chinese Journal of Tissue Engineering Research. All rights reserved.     

Bone marrow mesenchymal stem cells improve learning ability of the aging rat北大核心

BACKGROUND: At present, studies have shown that bone marrow mesenchymal stem cells (BMSCs) have self-renewal ability, which can be used as ideal seed cells for repairing tissue and organ damages caused by aging and lesions. OBJECTIVE: To study the changes in the levels of oxidation, inflammatory factors and neurotrophic factors (BDNF) in the brain of aging rats undergoing BMSCs transplantation, and to analyze the mechanism underlying the repair of learning and memory ability in the aging rats. METHODS: A total of 30 clean Sprague-Dawley rats were randomly divided into control group, model group and BMSCs group, 10 rats in each group. Aging models were made in the rats by 3-month subcutaneous injection of D-galactose. After modeling, BMSCs treatment was performed via tail vein injection in the BMSCs group. The injection was performed once a week, for 8 continuous weeks. Morris water maze was used to detect the learning and memory abilities of the rats in each group after the final injection of BMSCs. Superoxide dismutase activity in the brain tissue of rats was detected by xanthine oxidase method. Level of malondialdehyde in the rat brain tissue was detected by thiobarbituric acid method. Total antioxidant capacity of the brain tissue was detected by Fe3+ reduction method. Real-time PCR and western blot assay were used to detect the expression of brain-derived neurotrophic factor mRNA and protein in the brain tissue of the aging rat, respectively. RESULTS AND CONCLUSION: Compared with the model group, the BMSCs group exhibited significantly higher activity of superoxide dismutase, stronger total antioxidant capacity, and higher levels of brain-derived neurotrophic factor mRNA and protein (P < 0.05), but the lower malondialdehyde level in the brain (P < 0.05). Compared with the model group, there was less time and higher frequency for passing through the platform in the BMSCs group (P < 0.05). Our findings further indicate that BMSCs can improve the abilities of learning and memory in aging rats, and the underlying mechanism is likely to improve antioxidant capacity and to regulate the level of brain-derived neurotrophic factors. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Influence of Wnt/beta-catenin signal transduction pathway on the differentiation of umbilical cord mesenchymal stem cells into hepatocyte-like cells北大核心

BACKGROUND: Umbilical cord mesenchymal stem cells can be induced to differentiate into hepatocyte-like cells in vitro and in vivo. However, the exact mechanism is still unknown. [Existing studies have shown that the Wnt/β-catenin signaling pathway is closely related to this process. OBJECTIVE: To explore the effect of Wnt/β-catenin signaling pathway on the differentiation of umbilical cord mesenchymal stem cells into hepatocyte-like cells and its potential molecular mechanism. METHODS: Human umbilical cord mesenchymal stem cells were extracted from the neonatal umbilical cord by tissue adherent method. After being cultured and purified, the umbilical cord mesenchymal stem cells at passages 4-6 were divided into four groups: control group (DMEM culture group), hepatocyte-like differentiation group, activator Wnt3a group (adding 20 µg/L Wnt3a, an activator of Wnt/β-catenin signaling pathway, under the differentiation condition), ond inhibitor Dkk-1 group (adding 20 µg/L Dkk-1, on inhibitor of Wnt/β-catenin signaling pathway, under the differentiation condition). Induced cells were collected respectively on days 7,14, 21,28. Their mRNA and protem expressions of α-fetoprotein (AFP), albumin (ALB), hepatocytt nuclear factor 4α (HNF4α) and Cytokeratin-19 (CK-19) in the cells were detcted by real-time quantitative PCR and western blot respectively. (Meanwhile, Periodic Acid-Schiff staining, low-density lipoprotein uptake test and indocyanine green absorption test were applied to Oetect the function of hepatocyte-like cells. RESULTS AND CONCLUSION: Compared with the control group, expressions of AFP and HNF4a mRNA and protein as well as ALB mRNA were significantly up-regulated in the hepatocyte-like differentiation group, cctivator Wnt3a group end inhibitor Dkk-1 group (P < 0.05). Whereas, there was a decrease in the CK-19 expression at mRNA and protein levels (P < 0.01) in these three groups. Compared with the hepatocyte-like differentiation group, the mRNA and protein expressions of AFP and HNF4α, and the mRNA expression of ALB were significantly down-regulated in the activator Wnt3a group 0 < 0.05). Compared with hepatocyte-like differentiation group and activator Wnt3a group, the inhibitor Dkk-1 group had higher expression of AFP, HNF4α mRNA and their proteins as well as the mRNA expression of ALB (0 < 0.05). Findings from the Periodic Acid-Schiff staining, low-density lipoprotein uptake test and indocyanine green absorption test showed more positive cells in the inhibitor Dkk-1 group than in the hepatocyte-like differentiation group and least positive cells in the activator Wnt3a group. Overall, these findings suggest that the inhibition cf Wnt/β-catenin signaling pathway promotes the differentiation cf umbilical cord mesenchymal stem cells into hepatocyte-like cells; conversely, the cell differentiation can be inhibited cia the Wnt/β-catenin pathway. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Role of ABCG2 gene in proliferation and invasion of colorectal cancer stem cells北大核心

BACKGROUND: ABCG2 transporter can mediate multidrug resistance. ABCG2 overexpression can enhance the tolerance of stem cells to chemotherapeutic drugs. However, the mechanism of ABCG2 gene in the proliferation and invasion of colorectal cancer stem cells has not been confirmed. OBJECTIVE: To investigate the role of ABCG2 gene in the proliferation and invasion of colorectal cancer stem cells. METHODS: Colorectal cancer cell lines HCT116 were routinely isolated and cultured, and were randomly divided into four groups. CD133 positive cells were idted by immunomagnetic beads method, and were transfected with ABCG2-siRNA and ABCG2 overexpression plasmids to construct colorectal cancer stem cell modell with ABCG2 low expression and overexpression, and were set as low expression group and ovvrexpression groups, respectively. The remaining HCT116 cells were set as normal and smpty plasmid transfected control groups. The proliferation and invasion of colorectal cancer stem cell in different modell were determined by MTT assay and Transwell assay, respectively. Expressions of matrix metalloproteinase 9 (IMMP-9) mRNA snd protein were detected by enzyme linked immunosorbent sssay (ELISA) and polymerase chain reaction (PCR), respectively. RESULTS AND CONCLUSION: (1) Flow cytometry results showed that: CD133 positive cells in colorectal cancer cell line HCT116 accounted for 2.4%, while increased to 94.51 % of the cell line after being sorted by immunomagnetic beads, suggesting that isolated and cultured cell were colorectal cancer stem cell. (2) MTT assy showed no significant difference between the four groups of cell prior to the cell transfection (P > 0.05). However, MTT value in the low expression group was significantly lower than that in the overexpression group after cell transfection (P < 0.05). (3) Results from the Transwell assay showed that the cells in the low expression groups did not have the ability of migration; on the contrary, in the overexpression group, the number of cells crossing the basement membrane was relatively increased and the ability of cell migration and invasion was enhanced significantly (P <0.05). (4) The IVIMP-9 protein level was lower in the low expression group than in the overexpression group (P < 0.05). Similar results were yielded in the PCR detection. To conclude, down-regulation of ABCG2 protein expression can inhibit colorectal cancer stem cell proliferation and invasion. ABCG2 gene can change the viability of colorectal cancer stem cell by regulating the IMMP-9 expression. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.     

Analysis of the cause of pregnancy failure with combined MLPA assay for subtelomeric regions and ultrasonography北大核心CSCD

Objective To explore the value of multiplex ligation-dependent probe amplification (MLPA) for thedetection of chromosome abnormalities in miscarriage tissues, and to correlate the result with ultrasoundfindings. Methods A total of 421 cases of spontaneous abortions and fetal deaths were detected with the MLPAmethod. Results Among the 421 samples, 232 (55. 11%) had an abnormal MLPA result. For the 286 cases derivedfrom < 13 weeks pregnancy, 206 (72.03%) were abnormal. For the 49 cases from 14 ~ 19 weeks pregnancy, 14(28. 57%) were abnormal. For the 86 cases derived after 20 weeks pregnancy, 12 (13. 95%) were abnormal. Amongthe 117 cases with abnormal ultrasound findings, 33 cases (28. 21%) had an abnormal MLPA result, 28 out of the33 cases were numerical chromosome abnormality, 4 cases were chromosome microdeletion and/or micro duplication, 1 case had both numerical abnormality and microduplication. For those with abnormal ultrasound findings for the neck region, fetal edematous syndrome, multiple malformations and digestive system, the detection rates forMLPA were 71. 4% , 58. 8% , 37. 8% , and 9.1% , respectively. For those with abnormal finding of cardiacsystem, nervous system, face, skeletal system and urinary system, none was found with positive results of MLPA. Conclusion Numerical chromosomal abnormalities account for the majority of cases with spontaneous abortion.With the increase of gestational age, the occurrence of chromosomal abnormalities gradually declines. Combinedultrasound and MLPA assay can improve the detection rate and accuracy for chromosomal abormalities.     

miR-31 promotes the proliferation and migration of bone marrow mesenchymal stem cells北大核心

BACKGROUND: Bone marrow mesenchymal stem cells have been widely used in the clinical treatment of neurodegenerative-related diseases, but their efficacy is reduced by low cell survival and migration rates at the site of injury. OBJECTIVE: To investigate whether miR-31 can enhance the migration and proliferation of bone marrow mesenchymal stem cells. METHODS: Bone marrow mesenchymal stem cells from C57BL/6 mice were cultured and identified, and the cells were divided into control, miR-31 agomir, and miR-31 antagomir groups. Bone marrow mesenchymal stem cells at passage 3 were inoculated in six-well plates (1×105/well). When the cells reached 50%-80% fusion, miR-31 agomir and miR-31 antagomir were added to the six-well plates after dilution with serum-free DMEM/F12. After 24 hours of transfection, the proliferation level of cells was analyzed by CCK-8 assay as well as the migration ability of cells was analyzed by Transwell assay. Protein expression of matrix metalloproteinase 2 and CXC chemokine receptor 4 was detected by western blot assay. RESULTS AND CONCLUSION: (1) miR-31 was successfully transfected with bone marrow mesenchymal stem cells without transfection reagents and emitted red fluorescence. (2) After transfection, the proliferation ability of cells in the miR-31 agomir group was enhanced compared with the control group, which increased proportionally with time (P < 0.05). Compared with the control group, miR-31 promoted the migratory ability of bone marrow mesenchymal stem cells in the miR-31 agomir group (P < 0.05) and also upregulated protein expression of matrix metalloproteinase 2 and CXC chemokine receptor 4 (P < 0.05). (3) The results indicated that miR-31 could improve the proliferation ability of bone marrow mesenchymal stem cells and promote the migration of bone marrow mesenchymal stem cells, which provides a basic study for efficient targeting of mesenchymal stem cell migration to the site of injury. © 2023, Publishing House of Chinese Journal of Tissue Engineering Research. All rights reserved.     

Efficacy and safety of the Bushen tiaochong recipe combined with alendronate sodium for the treatment of postmenopausal senile degenerative osteoporosis: Study protocol for a randomized controlled trial北大核心

BACKGROUND: Alendronate sodium is a commonly used western medicine for the treatment of osteoporosis, but it has many adverse reactions. The main component of the traditional Chinese medicine, known as the Bushen Tiaochong recipe, is Epimedium brevicornu Maxim. (epimedium). Pharmacological studies have shown that the main active ingredient of Epimedium is icariin. Icariin has an estrogen-like effect, can prevent against bone loss and improve bone strength, and has a definite effect on the treatment of postmenopausal senile degenerative osteoporosis. OBJECTIVE: To testify the hypothesis that the Bushen Tiaochong recipe combined with alendronate sodium will be more effective, as well as safer and more reliable than alendronate sodium alone for the treatment of postmenopausal senile degenerative osteoporosis. METHODS: Two hundred patients with postmenopausal senile degenerative osteoporosis will be randomly assigned to an observation group and a control group. In the control group, patients will be given alendronate sodium tablets 10 mg/d and calcium carbonate D3 chewable tablets 0.6 g/d. In the observation group, patients will receive the same treatment as the control group and the Bushen Tiaochong recipe, simmering, twice per day (once in the morning and once in the evening). The duration of treatment will be 6 months in both groups. The primary outcome measure is the overall efficacy 6 months after treatment in both groups. The secondary outcome measures are Visual Analogue Scale scores for waist and back pain; lumbar spine (L2–4) bone mineral density; serum levels of calcium, phosphorus, alkaline phosphatase, osteocalcin, estradiol, follicle stimulating hormone, luteinizing hormone, interleukin-1, and interleukin-6 before and 6 months after treatment; and incidence of adverse reactions 6 months after treatment. This trial has been approved by the Ethics Committee of the Affiliated Hospital of Qinghai University of China (approval number: QHY1703F). The study protocol will be conducted in accordance with the Declaration of Helsinki, formulated by the World Medical Association. Written informed consent will be obtained from all participants. The recruitment of subjects will begin in January 2018. Samples and data will be collected from January to December 2018. Outcome measures will be analyzed in March 2019. This trial will be completed in April 2019. The results of the trial will be reported in a scientific conference or disseminated in a peer-reviewed journal. This trial was registered with the Chinese Clinical Trial Registry (registration number: ChiCTR-ONC-17013947). DISCUSSION: We hope to verify that the Bushen Tiaochong recipe combined with alendronate sodium provides better results than alendronate sodium alone for the treatment of postmenopausal senile degenerative osteoporosis. © 2018, Journal of Clinical Rehabilitative Tissue Engineering Research. All rights reserved.