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08 March 2021, Volume 25 Issue 7 Previous Issue    Next Issue

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Hypoxia preconditioning promotes bone marrow mesenchymal stem cells survival and vascularization through the activation of HIF-1α/MALAT1/VEGFA pathway Hou Jingying, Yu Menglei, Guo Tianzhu, Long Huibao, Wu Hao 2021, 25 (7):  985-990.  doi: 10.3969/j.issn.2095-4344.2160 Abstract ( 641 )   PDF (1563KB) ( 46 )   Save BACKGROUND: Previous study demonstrated that hypoxia preconditioning promoted mesenchymal stem cells survival and their therapeutic efficacy, and this effect was mediated by hypoxia induced factor-1α (HIF-1α). However, specific downstream mechanism remained unclear.                  OBJECTIVE: To observe the influence of hypoxia preconditioning on the survival and vascularization potential of bone marrow mesenchymal stem cells in vitro and explore the regulatory mechanism of HIF-1α/MALAT1/VEGFA pathway. METHODS: Bone marrow mesenchymal stem cells were obtained and cultured in vitro. Cells were divided into hypoxia (1% O2) and normoxia control groups (20% O2), and cultured for 24 hours. Cells proliferation, apoptosis and vascularization were evaluated. The expression of HIF-1α, MALAT1, and VEGFA was detected. HIF-1α and MALAT1 were inhibited by their siRNAs separately. HIF-1α siRNA scramble and MALAT1 siRNA scramble were used as negative controls before hypoxia preconditioning. Alterations of the molecules were examined and compared in different groups. RESULTS AND CONCLUSION: (1) Compared with the normoxia control group, cell viability was significantly enhanced; and cell apoptosis percentage was significantly declined in the hypoxia group; vascular lumen like structure was also increased significantly in the hypoxia group (P < 0.01); expression of HIF-1α, MALAT1, and VEGFA was significantly increased in the hypoxia group (P < 0.01). (2) After the inhibition of HIF-1α and hypoxia preconditioning, both MALAT1 and VEGFA expression levels were significantly reduced (P < 0.01). The expression of VEGFA was also significantly suppressed after the blockage of MALAT1 (P < 0.01). (3) This study suggested that hypoxia preconditioning effectively promoted bone marrow mesenchymal stem cell survival and vascularization through the activation of HIF-1α/MALAT1/VEGFA pathway. Figures and Tables | References | Related Articles | Metrics

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Pretreatment of placental mesenchymal stem cells to prevent bronchiolitis in mice Shi Yangyang, Qin Yingfei, Wu Fuling, He Xiao, Zhang Xuejing 2021, 25 (7):  991-995.  doi: 10.3969/j.issn.2095-4344.2161 Abstract ( 372 )   PDF (1267KB) ( 33 )   Save BACKGROUND: The recurrent attack of infantile bronchiolitis is one of the main causes of hospitalization, and its recurrent attack is closely related to the formation of asthma. In recent years, mesenchymal stem cells have been widely used in disease research. OBJECTIVE: To investigate the preventive effect of human placental mesenchymal stem cells on bronchiolitis in mice induced by respiratory syncytial virus. METHODS:  Forty Balb/c mice were randomly divided into mesenchymal stem cell pretreatment group, saline pretreatment group, model group, and normal group, with 10 in each group. The mice in the mesenchymal stem cell pretreatment group were pre-injected with 1 mL human placenta mesenchymal stem cells (containing 1 × 106 cells) in the tail vein. The mice in the saline pretreatment group were injected with the same amount of normal saline in the tail vein. The mice in the model group and the normal group were not treated. After 4 weeks of routine feeding, every week except the normal group, the model of bronchiolitis was established by nose drops, once a day, for 2 consecutive days. At 24 hours after the last nasal drip, cough and asthma, cyanosis around mouth and extremities and pathological changes of lung tissue were observed; interleukin-4 level in peripheral blood and GATA3 protein expression in lung tissue were detected.   RESULTS AND CONCLUSION:  (1) Compared with the model group, the symptoms and inflammation of lung tissue were significantly reduced in the mesenchymal stem cell pretreatment group. (2) Interleukin-4 level in peripheral blood and GATA3 protein expression in lung tissue were significantly lower in the mesenchymal stem cell pretreatment group than those in the model group (P < 0.01). (3) Interleukin-4 level in peripheral blood and GATA3 protein expression in lung tissue did not significantly change in the saline pretreatment group compared with the model group (P > 0.05). (4) Results suggest that pretreatment of placental mesenchymal stem cells can significantly reduce the content of inflammatory factors interleukin-4 and GATA3 in the peripheral blood of mice with bronchiolitis caused by respiratory syncytial virus infection in the short term, and reduce the degree of inflammation of bronchiolitis. Figures and Tables | References | Related Articles | Metrics

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Alveolar echinococcosis protoscolices inhibits the differentiation of bone marrow mesenchymal stem cells into fibroblasts Liang Xueqi, Guo Lijiao, Chen Hejie, Wu Jie, Sun Yaqi, Xing Zhikun, Zou Hailiang, Chen Xueling, Wu Xiangwei 2021, 25 (7):  996-1001.  doi: 10.3969/j.issn.2095-4344.2162 Abstract ( 401 )   PDF (2157KB) ( 113 )   Save BACKGROUND: The biological characteristics of hepatic alveolar echinococcosis are similar to cancer lesions. Its biological characteristics of invasive growth and metastasis increase the difficulty of surgery. The fibrosis of the outer capsule wall can inhibit the growth of echinococcus multilocularis and keep the disease in the quiescent stage. The role of mesenchymal stem cells in the fibrosis of the outer capsule wall of echinococcosis remains unclear.   OBJECTIVE: To investigate the effect of alveolar echinococcosis protoscolices on the differentiation of mesenchymal stem cells into fibroblasts.  METHODS:  Bone marrow mesenchymal stem cells were isolated from femur bone marrow of 4-week-old C57BL/6 mice, and cultured by adherent method. Alveolar echinococcosis protoscolices were extracted from gerbils infected with alveolar echinococcus. The experiment was divided into three groups. The alveolar echinococcosis group was co-cultured with the third generation of bone marrow mesenchymal stem cells and the protocephalus of alveolar echinococcosis protoscolices. The Echinococcus granulosus group was co-cultured with the third generation of bone marrow mesenchymal stem cells and the protocephalus of Echinococcus granulosus protoscolices. The simple control group was cultured with the third generation of bone marrow mesenchymal stem cells. At 1, 3, 5, and 7 days of cultivation, the real-time fluorescent quantitative PCR was used to detect collagen type I, collagen type III, transforming growth factor-beta 1 and Smad7 gene expression. Western blot assay was utilized to determine collagen type I, collagen type III, Smad7 and phosphorylated Smad2/3 protein expression. ELISA was applied to measure supernatant collagen type I and collagen type III contents.   RESULTS AND CONCLUSION: (1) Real-time fluorescent quantitative PCR detection displayed that transforming growth factor-beta 1, collagen type I, collagen type III mRNA relative expression levels were significantly lower in the alveolar echinococcosis group than in the Echinococcus granulosus group and simple control group (P < 0.05). Smad7 mRNA relative expression was significantly higher in the alveolar echinococcosis group than in the Echinococcus granulosus group and simple control group (P < 0.05). (2) Western blot assay showed that collagen type I, collagen type III and phosphorylated Smad2/3 protein relative expression levels were significantly lower in the alveolar echinococcosis group than in the Echinococcus granulosus group and simple control group (P < 0.05). Smad7 protein relative expression was significantly higher in the alveolar echinococcosis group than in the Echinococcus granulosus group and simple control group (P < 0.05). (3) ELISA exhibited that supernatant collagen type I and collagen type III contents were significantly lower in the alveolar echinococcosis group than in the Echinococcus granulosus group and simple control group (P < 0.05). (4) Alveolar echinococcosis protoscolices may promote bone marrow mesenchymal stem cells to secrete Smad7, inhibit the collagen type I, collagen type III and transforming growth factor-beta 1 through the transforming growth factor-beta/Smad signaling pathway, thereby inhibiting the fibrosis of bone marrow mesenchymal stem cells. Figures and Tables | References | Related Articles | Metrics

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Biological characteristics of canine adipose-derived mesenchymal stem cells cultured in hypoxia Fan Quanbao, Luo Huina, Wang Bingyun, Chen Shengfeng, Cui Lianxu, Jiang Wenkang, Zhao Mingming, Wang Jingjing, Luo Dongzhang, Chen Zhisheng, Bai Yinshan, Liu Canying, Zhang Hui 2021, 25 (7):  1002-1007.  doi: 10.3969/j.issn.2095-4344.2163 Abstract ( 364 )   PDF (2598KB) ( 177 )   Save BACKGROUND: Previous studies have found that hypoxia has different effects on the proliferation and differentiation of mesenchymal stem cells and secretion of cytokines, but the effect of hypoxia on canine adipose-derived mesenchymal stem cells has not been seen.  OBJECTIVE: To investigate the effect of hypoxic environment on the biological characteristics of canine adipose-derived mesenchymal stem cells.  METHODS:  Canine adipose-derived mesenchymal stem cells were isolated and cultured by enzyme digestion. Passage 2 adipose-derived mesenchymal stem cells could be divided into normoxia group (21% oxygen volume fraction) and hypoxia group (5% oxygen volume fraction). Morphological characteristics, proliferation speed, cell surface marker expression, differentiation capacity, and cytokine secretion level were compared between the two groups.   RESULTS AND CONCLUSION: (1) The growth of adipose-derived mesenchymal stem cells cultured in hypoxic environment was good. The cell morphology was fusiform and polyhedral, the same as that of the normoxia group. (2) The proliferation rate of cells in the hypoxia group was accelerated, and the cell doubling time was shorter than that in the normoxia group (all P < 0.05). The differentiation time of lipogenesis and osteogenesis was shortened. (3) The expression of CD90, CD44 and CD105 was high in both normoxia group and hypoxia group, while the expression of CD45 was low. (4) The mRNA expression levels of brain-derived neurotrophic factor and vascular endothelial growth factor in the hypoxia group were 2.3 times (P < 0.05) and 3.0 times (P < 0.05) higher than those in the normoxia group. (5) The results indicated that the hypoxic environment had no significant effect on the morphology and surface marker of adipose-derived mesenchymal stem cells, but promoted cell proliferation, differentiation and cytokine secretion. Figures and Tables | References | Related Articles | Metrics

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Role of hsa-miRNA-223-3p in regulating osteogenic differentiation of human bone marrow mesenchymal stem cells Geng Yao, Yin Zhiliang, Li Xingping, Xiao Dongqin, Hou Weiguang 2021, 25 (7):  1008-1013.  doi: 10.3969/j.issn.2095-4344.2164 Abstract ( 410 )   PDF (1895KB) ( 73 )   Save BACKGROUND: Studies have found that miR-223-3p is highly expressed in osteoporosis patients, but the related mechanism is rarely studied. OBJECTIVE: To investigate the regulatory effect of hsa-miR-223-3p on osteogenic differentiation of bone marrow mesenchymal stem cells. METHODS: Bone marrow mesenchymal stem cells were isolated and obtained by Percoll density gradient centrifugation. The cells were induced into osteogenic differentiation, and overexpressed hsa-miR-223-3p. RT-qPCR and western blot assay were used to detect the changes in the mRNA and protein levels of Wnt signaling pathway markers Wnt5a and its downstream signaling molecules OPG and RUNX2. PmirGLO/Wnt5a-3'UTR and pmirGLO/Wnt5a-3'UTR mut plasmids were constructed and co-transfected with hsa-miR-223-3p mimics/control plasmid into 293T cells to verify the targeting relationship between hsa-miR-223-3p and Wnt5a by dual luciferase reporter gene system. RESULTS AND CONCLUSION: (1) With the prolongation of osteogenic induction time, the level of alkaline phosphatase in cell culture fluid gradually increased. The deposition of dark red or reddish brown calcium salt nodules was gradually increased according to alizarin red staining results. (2) Overexpression of hsa-miR-223-3p decreased the mRNA and protein levels of Wnt5a, OPG and RUNX2 in cells. (3) hsa-miR-223-3p directly targeted the 3'UTR region of Wnt5a and reduced the luciferase activity of Wnt5a. This targeting effect disappeared when the binding site of Wnt5a was mutated. Blocking hsa-miR-223-3p increased the luciferase activity of wild-type Wnt5a, while the luciferase activity of mutant Wnt5a remained unchanged. (4) The results showed that during the differentiation of bone marrow mesenchymal stem cells into osteoblasts, overexpression of hsa-miR-223-3p could affect the expression of Wnt signaling pathway marker Wnt5a and its downstream signaling molecules OPG and RUNX2. hsa-miR-223-3p could directly bind to Wnt5a, which was negatively regulated as a downstream target gene of hsa-miR-223-3p.  Figures and Tables | References | Related Articles | Metrics

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Effect of peroxiredoxin 6 on proliferation and differentiation of bone marrow mesenchymal stem cells into neural lineage in vitro Lun Zhigang, Jin Jing, Wang Tianyan, Li Aimin 2021, 25 (7):  1014-1018.  doi: 10.3969/j.issn.2095-4344.2165 Abstract ( 389 )   PDF (1060KB) ( 213 )   Save BACKGROUND: It has been found that many substances can promote the differentiation of bone marrow mesenchymal stem cells into neural lineage in vitro, but the effect of peroxiredoxin 6 on bone marrow mesenchymal stem cells has not been studied.   OBJECTIVE: To explore the effects of peroxiredoxin 6 on the proliferation of rat bone marrow mesenchymal stem cells and the ability to differentiate into neural lineages in vitro.  METHODS: Bone marrow mesenchymal stem cells of Sprague-Dawley rats were cultured and passaged by whole bone marrow adherent culture in vitro into passage 3. The cells were assigned to five groups: PBS group, 1, 10, 100 μg/L and 1 mg/L peroxiredoxin 6 groups. CCK-8 assay was used to measure the absorbance at 450 nm of each group of cells for 9 consecutive days. Cell growth curves were drawn. Enzyme linked immunosorbent assay was utilized to measure absorbance at 450 nm in platform period. A relatively optimal concentration of peroxiredoxin 6 was selected to induce bone marrow mesenchymal stem cells to differentiate into neural lineages. PBS served as the control group. After 7 days of differentiation, immunofluorescence staining and western blot assay were used to detect the expression of neuronal marker protein NSE and glial cell marker protein GFAP. RESULTS AND CONCLUSION: (1) 1, 10 and 100 μg/L peroxiredoxin 6 groups complied with the general growth rule of bone marrow mesenchymal stem cells, and 10 μg/L was the optimal mass concentration. (2) NSE and GFAP immunofluorescence staining showed positive reaction after peroxiredoxin 6 induction; NSE and GFAP expression levels were higher than those in the control group. (3) The results of in vitro experiments show that the proper concentration of peroxiredoxin 6 can promote the proliferation of bone marrow mesenchymal stem cells and induce differentiation into neural lineage. Figures and Tables | References | Related Articles | Metrics

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Mechanism of bone marrow mesenchymal stem cells differentiation into functional neurons induced by glial cell line derived neurotrophic factor Zhu Xuefen, Huang Cheng, Ding Jian, Dai Yongping, Liu Yuanbing, Le Lixiang, Wang Liangliang, Yang Jiandong 2021, 25 (7):  1019-1025.  doi: 10.3969/j.issn.2095-4344.2166 Abstract ( 451 )   PDF (2961KB) ( 50 )   Save BACKGROUND: Glial cell line derived neurotrophic factor (GDNF) plays an important role in inducing the differentiation of bone marrow mesenchymal stem cells (BMSCs) in vitro and promoting the regeneration of neuron axons. OBJECTIVE: To observe BMSCs differentiation induced by over-expression of GDNF gene, detect synaptic function of cells and expression of Wnt signaling pathway components after differentiation, and preliminarily explore the mechanism of BMSCs differentiation into mature neurons. METHODS:  Rat BMSCs were isolated and cultured, and further divided into recombinant adenovirus-containing GDNF gene transfection group (Ad-GDNF-BMSCs), adenovirus transfection control group (Ad-BMSCs), and untransfected control group. The relative expression of GDNF gene in BMSCs of each group was detected by Q-PCR, and the expression of β-catenin, cyclin D1, NeuN and MAP-2 was detected by immunofluorescence technology in each group. High K+ stimulation induced cell depolarization response after differentiation, and FM4-64 marks synaptic vesicle activity of differentiated cells. RESULTS AND CONCLUSION: (1) The adenovirus-transfected gene had no significant negative effect on the proliferation of BMSCs. BMSCs could express endogenous GDNF gene continuously and at high levels. (2) Under the induction of GDNF gene, BMSCs could express neuron-specific protein NeuN after 3 days cultivation in vitro. The expression of β-catenin protein also could be detected in the cytoplasm of cells. After 7 days cultivation in vitro culture, BMSCs expressed the mature neuronal marker protein MAP-2, and the cell body contracted significantly. Neuron axon-like structures appeared around the cell body. Moreover, β-catenin and cyclin D1 were respectively detected in the cell cytoplasm and the nucleus, while the expression levels of NeuN, MAP-2, β-catenin, and cyclin D1 were not observed in Ad-BMSCs and untransfected control groups, and the cells remained spindle-shaped. (3) After 11 days of in vitro culture, the cells in the Ad-GDNF-BMSCs group showed typical neuronal processes or axons and connected to each other into a network structure, which could be labeled with FM4-64 to show red fluorescence, and induced by high K+ stimulation to induce action potentials in the cells. Synaptic vesicle activity in posterior axons showed FM4-64 red fluorescence gradually decaying. Under the same conditions, cells in the Ad-GDNF-BMSCs group and untransfected control group did not present FM4-64 fluorescently labeled synaptic vesicle activity. (4) Continuous GDNF induction can promote BMSCs differentiated into mature neurons with synaptic cycle function, and may be carried out through the classic Wnt/β-catenin signaling pathway. Figures and Tables | References | Related Articles | Metrics

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Human placenta mesenchymal stem cells-derived extracellular vesicles regulate collagen deposition in intestinal mucosa of mice with colitis Duan Liyun, Cao Xiaocang 2021, 25 (7):  1026-1031.  doi: 10.3969/j.issn.2095-4344.2167 Abstract ( 474 )   PDF (2460KB) ( 116 )   Save BACKGROUND:Intestinal fibrosis is a common complication in inflammatory bowel disease and leads to functional damage and intestinal obstruction. Intestinal fibrosis is mainly related to the imbalance of deposition and degradation of extracellular matrix components, such as collagens and fibronectins. Studies have found that mesenchymal stem cells secreted soluble bioactive substance such as extracellular vesicles via paracrine action, which exerted marked anti-fibrosis effect.  OBJECTIVE: To investigate the effect of human placenta mesenchymal stem cells-derived extracellular vesicles on collagen deposition in mice with colitis.  METHODS:  Totally 24 BALB/c mice were randomly divided into sham operation group, model group and extracellular vesicles group, with 8 mice in each group. Except the sham operation group, the remaining mice of model group and extracellular vesicles group were treated with trinitro-benzene-sulfonic acid to induce intestinal fibrosis, once a day for 6 weeks. The mice in the extracellular vesicles group and model group were administered with extracellular vesicles and phosphate-buffered saline, respectively, at 3 weeks, once a day for 6 weeks. The therapeutic effect of extracellular vesicles was evaluated by disease active index score and the colon weight/length ratio at 1-7 weeks. Diseased intestinal segment was subjected to histological staining. Western blot assay and RT-PCR were used to measure fibrosis related indicators so as to evaluate the degree of intestinal fibrosis.  RESULTS AND CONCLUSION:  (1) Compared with the model group, disease active index score and the colon weight/length ratio were significantly reduced, and colonic pathology was significantly improved in the extracellular vesicles group. (2) Compared with the model group, collagen deposition in colon mucosa of mice was significantly reduced, and the expression of collagen I, collagen III and transforming growth factor-β1 decreased significantly in the extracellular vesicles group. (3) Compared with the model group, expression levels of matrix metalloproteinase 2 and matrix metalloproteinase 9 in mouse colon tissue were significantly increased, while the expression level of tissue inhibitor of metalloproteinase 1 was decreased in the extracellular vesicles group. (4) Results suggest that human placenta mesenchymal stem cells-derived extracellular vesicles can obviously improve the severity of colon injury and reduce the collagen deposition of intestinal mucosa in mice with enteritis. Figures and Tables | References | Related Articles | Metrics

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Salvianolic acid B inhibits oxidative damage of bone marrow mesenchymal stem cells and promotes differentiation into cardiomyocytes Pei Lili, Sun Guicai, Wang Di 2021, 25 (7):  1032-1036.  doi: 10.3969/j.issn.2095-4344.2168 Abstract ( 331 )   PDF (1362KB) ( 37 )   Save BACKGROUND: In recent years, bone marrow mesenchymal stem cells have been used in clinical treatment of heart injury caused by permanent cardiomyocytopenia. However, the best inducer to promote the proliferation and differentiation of bone marrow mesenchymal stem cells into cardiomyocytes is still in the research and selection. OBJECTIVE: To investigate the protective effect of salvianolic acid B on oxidative stress injury and cardiomyocyte differentiation of bone marrow mesenchymal stem cells.  METHODS:  Bone marrow mesenchymal stem cells were used as study objects to establish oxidative stress injury model by 300 μmol/L H2O2 for 24 hours. Simultaneously, bone marrow mesenchymal stem cells were treated with salvianolic acid B for 24 hours. The contents of lactate dehydrogenase and creatine kinase in the supernatant and malondialdehyde and superoxide dismutase in the cell lysate were determined in each group using the kit. Western blot assay was used to detect the expression of nuclear factor E2 related factor 2 (Nrf2), Keap1, Bcl-2 and Bax in bone marrow mesenchymal stem cells. RT-PCT was used to detect cardiomyocyte differentiation-related markers 14 days after intervention.   RESULTS AND CONCLUSION: (1) Compared with the model group, the levels of lactate dehydrogenase and creatine kinase in the supernatant were significantly decreased in the salvianolic acid B group (P < 0.05). (2) Compared with the model group, the level of malondialdehyde in cell lysate was decreased, and the level of superoxide dismutase was increased in the salvianolic acid B group (P < 0.05). (3) Compared with the model group, expression of Nrf2 protein in cells was significantly increased, and the expression of Keap1 protein was significantly decreased in the salvianolic acid B group. (4) The expression of Bcl-2 in salvianolic acid B group was significantly higher than that in model group, and the expression of Bax in salvianolic acid B group was significantly lower than that in model group. (5) Compared with the model group, the expression of GATA4 and cTnT mRNA was significantly increased in the salvianolic acid B group. (6) Salvianolic acid B can induce the expression of proteins that activate Nrf2-ARE signaling pathway in bone marrow mesenchymal stem cells for anti-oxidation, and salvianolic acid B can inhibit apoptosis and improve the ability of bone marrow mesenchymal stem cells to differentiate into cardiomyocytes. Figures and Tables | References | Related Articles | Metrics

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Human acellular amniotic membrane scaffold promotes ligament differentiation of human amniotic mesenchymal stem cells modified by Scleraxis in vitro Zou Gang, Xu Zhi, Liu Ziming, Li Yuwan, Yang Jibin, Jin Ying, Zhang Jun, Ge Zhen, Liu Yi 2021, 25 (7):  1037-1044.  doi: 10.3969/j.issn.2095-4344.2169 Abstract ( 415 )   PDF (3498KB) ( 45 )   Save BACKGROUND: Acellular amniotic membrane is a natural biomaterial scaffold, which has been widely used in related fields of tissue engineering. Scleraxis gene plays an important regulatory role in the formation of connective tissues such as ligaments. OBJECTIVE: To verify the biocompatibility of Scleraxis modified human amniotic mesenchymal stem cells and human acellular amniotic membrane scaffold complex in rats. METHODS:  The amniotic membrane tissues of the placenta at term were taken, and the fresh human amniotic membrane was decellularized by chemical-enzymatic digestion method. Two-step enzyme digestion method was used to separate human amniotic mesenchymal stem cells from human fresh amniotic membrane. The third-generation human amniotic mesenchymal stem cells were transfected with Scleraxis lentivirus, which were then cultured with human decellularized amniotic membrane for 5, 10, and 15 days so as to detect mRNA expression of related genes. The 18 Sprague-Dawley rats were randomly divided into three groups. In the experimental group, the prepared cell scaffold complex was implanted into the subcutaneous fascia of the back of the rat. In the negative control group, an incision was made on the back of the rat, without implanting materials. In the blank control group, no treatment was performed. Hematoxylin-eosin staining was performed on the operation area tissues 1 and 4 weeks after surgery. Immunohistochemical staining of CK protein was performed on the operation area tissues 4 weeks after surgery.  RESULTS AND CONCLUSION: (1) Scleraxis gene transfection of human amniotic mesenchymal stem cells and human acellular amniotic membrane compound culture could significantly up-regulate the mRNA expression levels of ligament differentiation-related genes type I collagen, type III collagen, fibronectin and cytocontin C. (2) The neonatal granulation tissue was seen in the local tissue of the experimental group at 1 week after surgery, and the inflammatory response was heavier than that of the negative control group and the blank control group. At 4 weeks after surgery, the local tissue arrangement of the experimental group tended to be neat; the granulation tissue was reduced; and the inflammation subsided obviously. (3) The CK protein expression was positive in the experimental group at 4 weeks after the operation; the local tissues were neatly arranged; and a large number of cells were attached around the amniotic membrane tissue. (4) Results suggested that co-culture of Scleraxis gene transfected human amniotic mesenchymal stem cells with human acellular amniotic membrane scaffold can promote the differentiation of human amniotic mesenchymal stem cells into ligaments in vitro, and the cell scaffold complex shows good biocompatibility in animals. Figures and Tables | References | Related Articles | Metrics

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Morphological changes in human oligodendrocyte progenitor cells during passage Guan Qian, Luan Zuo, Ye Dou, Yang Yinxiang, Wang Zhaoyan, Wang Qian, Yao Ruiqin 2021, 25 (7):  1045-1049.  doi: 10.3969/j.issn.2095-4344.2170 Abstract ( 369 )   PDF (1386KB) ( 51 )   Save BACKGROUND: Oligodendrocyte precursor cells are seed cells for the treatment of white matter injury. The establishment of an efficient and stable in vitro culture method is an important prerequisite for clinical transformation. OBJECTIVE: To investigate the morphological changes of oligodendrocyte precursor cells during passage. METHODS: Four batches of human oligodendrocyte progenitor cells were subcultured from the second generation (P2) to the seventh generation (P7) in vitro. Five pictures of 200-fold field were taken under an optical microscope before each passage. According to the cell morphology, oligodendrocyte precursor cells were divided into three types: bipolar cells (oligodendrocyte precursor cells), multipolar cells (late oligodendrocyte precursor cells) and supra-polar bifurcated cells (immature oligodendrocyte cells). The proportion of oligodendrocyte progenitor cells to the total number of cells was calculated, so as to compare the difference of cell morphology among different generations.  RESULTS AND CONCLUSION: In the process of passage from P2 to P7, oligodendrocyte progenitor cells included three types: bipolar cells, multipolar cells and supra-polar bifurcated cells. Among them, bipolar cells and multipolar cells were the main part, and a small number of supra-polar bifurcated cells could be seen in the rest. There were no significant differences in the proportion of bipolar cells, multipolar cells and supra-polar bifurcated cells among P2-P7 (P > 0.05). The cell morphology classification and counting method can be used to preliminarily evaluate that oligodendrocyte progenitor cells have no change in morphology during culture. Figures and Tables | References | Related Articles | Metrics

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Platelet-derived growth factor-BB promotes proliferation, differentiation and migration of skeletal muscle myoblast Li Cai, Zhao Ting, Tan Ge, Zheng Yulin, Zhang Ruonan, Wu Yan, Tang Junming 2021, 25 (7):  1050-1055.  doi: 10.3969/j.issn.2095-4344.2171 Abstract ( 500 )   PDF (2429KB) ( 92 )   Save BACKGROUND: Skeletal muscle myoblasts differentiate and fuse to form polynuclear muscle tubes and muscle fibers to complete the repair of muscle injury when skeletal muscles were injured. However, the repair process is slow and incomplete. Platelet-derived growth factor-BB can stimulate the proliferation, differentiation and migration of a variety of tissue cells, and plays an important role in the process of tissue repair after various injuries. OBJECTIVE: To explore the effects of different concentrations of platelet-derived growth factor-BB on proliferation, differentiation and migration of skeletal muscle myoblast C2C12 cells and the action mechanism. METHODS:  The mouse skeletal muscle myoblast C2C12 cells were cultured with platelet-derived growth factor-BB 0, 5, 10, 20 and 40 μg/L and platelet-derived growth factor receptor inhibitor imatinib. The expression of platelet-derived growth factor receptor in C2C12 cells was detected by immunocytochemistry and western blot assay. After 1, 2, 3, 4, and 5 days of culture, CCK8 method was used to detect cell proliferation. After 4 days of induction and differentiation culture, the formation of myotubes in each group was observed by light microscope. The expression of myosin heavy chain and MyoG Gene was observed by immunofluorescence and western blot assay. After 48 hours of culture, Transwell method was used to detect cell migration.  RESULTS AND CONCLUSION: (1) Immunofluorescence chemistry and western blot assay indicated that platelet-derived growth factor receptor expression could be detected in C2C12 cells, and semi-quantitative statistical analysis showed no significant difference in platelet-derived growth factor receptor expression between groups with different platelet-derived growth factor-BB concentrations (P > 0.05). (2) CCK8 assay demonstrated that the proliferation of C2C12 cells showed no significant change in platelet-derived growth factor-BB groups compared with the control group (platelet-derived growth factor-BB 0 μg/L group). (3) Immunofluorescence chemistry showed that compared with control group, number of myosin heavy chain positive cells increased in platelet-derived growth factor-BB groups; 40 μg/L platelet-derived growth factor-BB concentration got the highest; the mature muscle tubes up to (27.00±0.76) per field of vision, and MyoG expression populations was most obviously compared with the control group (P < 0.05). (4) Transwell results showed that compared with the control group, the migration number of C2C12 cells in the platelet-derived growth factor-BB group increased, and the migration number in the 40 μg/L group was up to 144.00±13.03 (P < 0.05). (5) It is concluded that platelet-derived growth factor-BB promoted the migration, differentiation and myotube formation of C2C12 cells, and the pro-differentiation mechanism is related to its enhanced binding with platelet-derived growth factor receptor, so as to improve the expression of differentiation related gene MyoG.  Figures and Tables | References | Related Articles | Metrics

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Direct reprogramming hepatocytes into islet-like cells by efficiently targeting and activating the endogenous genes Wang Hanyue, Li Furong, Yang Xiaofei, Hu Chaofeng 2021, 25 (7):  1056-1063.  doi: 10.3969/j.issn.2095-4344.2172 Abstract ( 564 )   PDF (4125KB) ( 82 )   Save BACKGROUND: Islet cell transplantation is one of the most effective methods to treat diabetes. However, the shortage of transplanted cells has limited its clinical application. Direct reprogramming of hepatocytes to islet β cells in vitro is a new idea to solve this problem, but differentiation of hepatocytes to islet β cells is a complicated process. OBJECTIVE: Direct reprogramming hepatocytes into islet-like cells by efficient targeting and activating the endogenous genes of hepatocytes with Casilio system (constructed CRISPR/ Cas9-Pumilio system) through modifying guide RNA sequence combined with the transcriptional activator PUFa-P65-HSF1. METHODS: The endogenous PNM (Pdx1, Ngn3, MafA) genes of hepatocytes were targeted and activated by using the Casilio system, which was transfected to the HEK293T cell line by liposome transfection. The expression of endogenous PNM was detected by real-time fluorescence quantitative PCR and immunofluorescence. The Ins-EGFP cell line with stable expression of enhanced green fluorescent protein was constructed by the lentivirus carrying Ins-promoter-EGFP. The Ins-EGFP-HepG2-Cas9-PUFa-p65-HSF1 cell line (referred to as stable translocated cell lines) with stable expression of dCas9 and PUFa-P65-HSF1 in Ins-EGFP-HepG2 cell line was constructed by the PiggyBac(PB) transposon system. By using liposome, the gRNAs were transfected to stable translocated cell line, and the expression level of endogenous gene PNM was detected by real-time fluorescence quantitative PCR and immunofluorescence. Simultaneously, reprogramming efficiency was observed.  RESULTS AND CONCLUSION: (1) The activation of endogenous genes by Casilio system was verified in 293T cell line. (2) The expression of EGFP, dCas9 and PUFa-P65-HSF1 in stable cell line was detected by RT-PCR, western blot assay and immunofluorescence. (3) The real-time fluorescence quantitative PCR results confirmed that this new Casilio system could target and activate the PNM which led to the up-regulation of the endogenous PNM gene expression. The efficiency of direct reprogramming was 10%-15%. (4) Based on CRISPR/dCas9, the new Casilio system can efficiently activate the endogenous PNM gene in hepatocytes, enabling the hepatocyte line to be directly reprogrammed as islet-like cells. Figures and Tables | References | Related Articles | Metrics

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CaMKII-Smad1 promotes axonal regeneration of peripheral nerves Wang Feng, Zhou Liyu, Saijilafu, Qi Shibin, Ma Yanxia, Wei Shanwen 2021, 25 (7):  1064-1068.  doi: 10.3969/j.issn.2095-4344.2173 Abstract ( 388 )   PDF (1040KB) ( 51 )   Save BACKGROUND: Axons do not regenerate after central nervous system injury in mammals. It is mainly caused by the inhibitory microenvironment at the site of damage and the weakened self-regeneration ability. Studies have found that peripheral nervous system has certain regeneration ability after injury, so we explore the methods of central nervous system repair by studying the genes promoting peripheral nervous system regeneration. As one of the important protein kinase families of neurons, CaMKII up-regulation can improve the ability of neuron regeneration. Similarly, acute depletion of the Smad1 protein in adult mice also prevented axon regeneration in vivo. These genes can directly or indirectly regulate neuronal axon regeneration, but exactly how they regulate neuronal regeneration is still unclear.  OBJECTIVE: To study the effects of CaMKII-Smad1 signaling pathway on axon regeneration of dorsal root ganglion neurons by intraperitoneal injection of CaMKII inhibitor and activator, and explored the mechanism of CaMKII and Smad1 in regulating axon regeneration of dorsal root ganglion neurons. METHODS:  Totally 40 ICR mice were randomly divided into four groups: KN93 control group, KN93 experimental group, CdCl2 control group and CdCl2 experimental group. Dorsal root ganglion tissue was taken for in vitro culture after 7 days of continuous administration of CaMKII inhibitor KN93 and activator CdCl2. The length of axonal regeneration of dorsal root ganglion neurons was statistically analyzed after 3 days. Protein expression of p-Smad1 in dorsal root ganglion neurons was detected using western blot assay.  RESULTS AND CONCLUSION: (1) Compared with the KN93 control group, axonal regeneration of dorsal root ganglion neurons was inhibited, and the p-Smad1 protein expression was decreased in the KN93 experimental group, showing significant differences. (2) Compared with the CdCl2 control group, axonal regeneration of dorsal root ganglion neurons was promoted, and p-Smad1 protein expression was increased in the CdCl2 experimental group, showing significant differences. (3) The results showed that the CaMKII-Smad1 signaling pathway had a regulatory effect on axonal regeneration of dorsal root ganglion neurons. Figures and Tables | References | Related Articles | Metrics

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Molecular mechanism of miR-17-5p regulation of hypoxia inducible factor-1α mediated adipocyte differentiation and angiogenesis Liu Cong, Liu Su 2021, 25 (7):  1069-1074.  doi: 10.3969/j.issn.2095-4344.2174 Abstract ( 479 )   PDF (1473KB) ( 67 )   Save BACKGROUND: miR-17-5p can regulate the differentiation of adipocytes, but its action mechanism is not clear. Hypoxia inducible factor-1α can promote vascular endothelial growth factor gene transcription and promote angiogenesis, and has a regulatory effect on adipocyte formation. However, the specific mechanism of hypoxia inducible factor-1α in regulating adipocyte differentiation and angiogenesis is not clear.  OBJECTIVE: To investigate the molecular mechanism of miR-17-5p regulation of hypoxia inducible factor-1α mediated adipocyte differentiation and angiogenesis.  METHODS:  miR-17-5p expression level and adipocyte differentiation and the expression of angiogenesis markers in mature adipocytes were verified by RT-qPCR. The adipocyte differentiation and angiogenesis markers expression of adipocyte transfecting with miR-17-5p inhibitor, pri-miR-17-5p mimic and hypoxia inducible factor-1α knockdown vector were determined by western blot assay. The proliferation of adipocytes was detected by Oil Red O staining assay and MTT assay after transfection with miR-17-5p inhibitor, pri-miR-17-5p mimic and hypoxia inducible factor-1α knockdown vector. The predicted relationship between miR-17-5p and hypoxia inducible factor-1α was further verified by EGFP report gene assay.  RESULTS AND CONCLUSION: (1) Compared to immature adipocyte, miR-17-5p was highly expressed in mature adipocyte (P < 0.05); overexpression of miR-17-5p increased the adipocyte proliferation (P < 0.05), and upregulated adipocyte differentiation and angiogenesis markers level (P < 0.05). (2) The miR-17-5p directly targeted with hypoxia inducible factor-1α 3′UTR; knockdown hypoxia inducible factor-1α inhibited the adipocyte proliferation (P < 0.05), and decreased adipocyte differentiation and angiogenesis markers level (P < 0.001). (3) It is concluded that miR-17-5p regulation of hypoxia inducible factor-1α mediated adipocyte differentiation and angiogenesis.  Figures and Tables | References | Related Articles | Metrics

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Inhibitory effects of sodium butyrate on microglial activation and expression of inflammatory factors induced by fluorosis Wang Zhengdong, Huang Na, Chen Jingxian, Zheng Zuobing, Hu Xinyu, Li Mei, Su Xiao, Su Xuesen, Yan Nan 2021, 25 (7):  1075-1080.  doi: 10.3969/j.issn.2095-4344.2175 Abstract ( 492 )   PDF (1312KB) ( 71 )   Save BACKGROUND: In the diseases process of the central nervous system, pathogenic factors stimulate microglia cells to over-activate and secrete inflammatory factors, which lead to further injury of neurons. Whether the deacetylase inhibitor sodium butyrate can reduce the over-activation and inflammatory effects of microglia cells caused by fluorosis is worthy of further study.  OBJECTIVE: To investigate the effects of sodium butyrate on fluorine-induced over-activation of microglia in the brain and the expression of inflammatory factors interleukin-1β, interleukin-6 and tumor necrosis factor-α.  METHODS:  Microglia cells (BV-2) were routinely cultured in F12/DMEM medium containing 10% fetal bovine serum. CCK-8 method was used to observe the effects of sodium fluoride, sodium butyrate and their combination on the viability of BV-2 cells in logarithmic growth phase. The morphology of BV-2 cells after treatment with sodium fluoride, sodium butyrate and their combination, mRNA and protein expressions of interleukin-1β, interleukin-6 and tumor necrosis factor-α were observed by inverted microscope, RT-qPCR and western blot assay.   RESULTS AND CONCLUSION: (1) CCK8 results showed that 25×10-6 sodium fluoride could reduce the viability of BV-2 cells, and 0.25 mmol/L sodium butyrate could increase the viability of BV-2 cells. Sodium butyrate 0.25 mmol/L could reverse the decreased viability of 25×10-6 sodium fluoride on BV-2 cells. (2) RT-qPCR and western blot assay results showed that 0.25 mmol/L sodium butyrate could antagonize the over-expression of interleukin-1β, interleukin-6 and tumor necrosis factor-α induced by 25×10-6 sodium fluoride. (3) It indicates that sodium butyrate, in a certain dose, can reduce the inflammatory response mediated by fluorium-induced microglia cells. Figures and Tables | References | Related Articles | Metrics

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Strategies for improving the therapeutic efficacy of mesenchymal stem cells in the treatment of nonhealing wounds Wang Xianyao, Guan Yalin, Liu Zhongshan 2021, 25 (7):  1081-1087.  doi: 10.3969/j.issn.2095-4344.2176 Abstract ( 436 )   PDF (861KB) ( 61 )   Save BACKGROUND: Mesenchymal stem cell-based therapy is pursued aggressively with promising preclinical studies and higher future prospect in the treatment of nonhealing wounds. OBJECTIVE: To review the strategies for improving the efficacy of mesenchymal stem cells in the treatment of nonhealing wounds, which are expected to provide theoretical basis for clinical treatment of nonhealing wounds. METHODS: Wanfang, CNKI, PubMed and Web of Science were searched for articles about the application of mesenchymal stem cells in nonhealing wounds from March 2002 to March 2020. The key words were “mesenchymal stem cells, nonhealing wounds (nonhealing chronic wounds), wound repair, tissue regeneration” in Chinese and “mesenchymal stem cells, nonhealing chronic wound, wound repair, tissue regeneration” in English. After excluding outdated and repetitive viewpoints, the retrieved literature was sorted out and a total of 64 articles were included for analysis.  RESULTS AND CONCLUSION: (1) The formation mechanism of nonhealing wounds was summarized. (2) The molecular mechanisms of mesenchymal stem cells in the treatment of nonhealing wounds were summarized, such as direct differentiation, paracrine, immunomodulatory, mobilization resident stem cells to participate in wound repair. (3) This paper summarized the improvement of the efficacy of mesenchymal stem cells in the treatment of nonhealing wounds, such as gene modification and pretreatment. (4) This study can provide a theoretical basis for improving the efficacy of mesenchymal stem cells in the treatment of nonhealing wounds.  Figures and Tables | References | Related Articles | Metrics

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Research progress in the treatment of spinal cord injury with mesenchymal stem cell secretome Wan Ran, Shi Xu, Liu Jingsong, Wang Yansong 2021, 25 (7):  1088-1095.  doi: 10.3969/j.issn.2095-4344.2113 Abstract ( 564 )   PDF (1042KB) ( 106 )   Save BACKGROUND: Spinal cord injury often leads to patients’ permanent sensory, motor and autonomic dysfunction because of limited inherent regeneration capacity of the central nervous system. Spinal cord injury seriously affects patients’ quality of life but there is still no effective treatment for it. The existing studies suggest that mesenchymal stem cell secretome mediates the main therapeutic effect of cell transplantation and avoids problems such as cellular rejection, so it will become a powerful tool for the treatment of spinal cord injury.  OBJECTIVE: To summarize the research progress of mesenchymal stem cell secretome in the treatment of spinal cord injury, and analyze the current problems and future development direction.  METHODS: The PubMed database was retrieved with “spinal cord injury, secretome” as the keywords, and Chinese databases include CNKI and Wanfang were retrieved with “spinal cord injury, secretome” as the keywords in Chinese. We retrieved the articles published from January 2013 to January 2020 and excluded the articles that were irrelevant or repetitive. At last 71 articles that met the criteria were included for review.  RESULTS AND CONCLUSION: The mesenchymal stem cell secretome is rich in extracellular vesicles and soluble molecules such as growth factors and neurotrophic factors, and it plays prominent place in reducing cell apoptosis, regulating immune response, inhibiting scar formation, promoting nerve regeneration and angiogenesis. Numerous studies have shown that the mesenchymal stem cell secretome can promote nerve regeneration and function recovery after spinal cord injury, and it avoids the disadvantages of cell transplantation. The mesenchymal stem cell secretome will become a reliable method to treat spinal cord injury in the future. Figures and Tables | References | Related Articles | Metrics

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Therapeutic target and application prospects of oral squamous cell carcinoma stem cells Liao Chengcheng, An Jiaxing, Tan Zhangxue, Wang Qian, Liu Jianguo 2021, 25 (7):  1096-1103.  doi: 10.3969/j.issn.2095-4344.2126 Abstract ( 495 )   PDF (1261KB) ( 74 )   Save BACKGROUND: Tumor stem cells are a small number of types of tumor cells that have the ability to self-renew and differentiate into different types of tumor cells. Oral squamous cell carcinoma stem cells are highly tumorigenic and play a role in tumor differentiation, treatment resistance, recurrence and metastasis. Simultaneously, tumor stem cells have great similarities with normal stem cells, so it is necessary to establish effective and accurate tumor stem cell identification methods; and corresponding targeted treatment strategies are designed to help the prognosis of patients with oral squamous cell carcinoma. OBJECTIVE: To summarize the current methods used in the literature to identify and isolate oral squamous cell carcinoma stem cells, analyze potential targets for oral squamous cell carcinoma stem cells, and summarize the potential research progress on targets. METHODS: Computers were used to retrieve the CNKI and PubMed databases for relevant literature published since its establishment to 2020. The English key words were “oral squamous cell carcinoma, OSCC, cancer stem cells, HNSCC, head and neck squamous carcinoma cell”. Chinese key words were “oral squamous cell carcinoma stem cells, oral squamous cell carcinoma, tumor stem cells, head and neck squamous cell carcinoma”. Retrieval results were summarized and analyzed to exclude low-relevance, duplicate, and obsolete documents.   RESULTS AND CONCLUSION: Targeted intervention of oral squamous cell carcinoma stem cells has important clinical significance. CD44, CD133 and ALDH are currently the most suitable biomarkers for the identification and isolation of oral squamous cell carcinoma stem cells. They are the same as Oct3/4, Nanog, Sox2, Bmi1, EGFR signaling pathway, SHH signaling pathway, Notch signaling pathway, Wnt signaling pathway, Let-7 family, MicroRNA-200 family and natural compounds together as potential targets for targeted therapy of oral squamous cell carcinoma stem cells. Figures and Tables | References | Related Articles | Metrics

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Exosomes as a disease marker under hypoxic conditions Zhao Min, Feng Liuxiang, Chen Yao, Gu Xia, Wang Pingyi, Li Yimei, Li Wenhua 2021, 25 (7):  1104-1108.  doi: 10.3969/j.issn.2095-4344.2177 Abstract ( 489 )   PDF (624KB) ( 89 )   Save BACKGROUND: Hypoxia is a common physiological and pathological stress response in various diseases, and exosomes are widely involved in the mechanism of hypoxia injury and its adaptive mechanism. The two are closely related, which is the current research hotspot.  OBJECTIVE: To find a new target for the prevention and treatment of high altitude diseases and open up new ideas. METHODS: The PubMed and Medline databases from 2013 to 2020, and the CNKI and Wanfang databases from 2018 to 2019 were searched by computer. The key words were “serum exosomes, hypoxia, tumor, inflammation, cardiovascular and cerebrovascular diseases” in Chinese and English. Finally, 57 articles were included in the review.  RESULTS AND CONCLUSION: Hypoxia is a potential environmental death factor by affecting cell cycle, morphology, metabolism, proliferation, differentiation, autophagy, apoptosis and so on. Hypoxia can regulate the release and change the content of exosomes, which is strongly associated with cell specificity, the duration of hypoxia exposure and the severity of hypoxia. In the environment of hypoxia, exosomes can transmit all kinds of biological information and play an important role in biology through specific binding with receptor cells. As a biomarker of disease diagnosis, exosomes have great potential. Figures and Tables | References | Related Articles | Metrics

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Role of microglia-mediated neuronal injury in neurodegenerative diseases Xie Wenjia, Xia Tianjiao, Zhou Qingyun, Liu Yujia, Gu Xiaoping 2021, 25 (7):  1109-1115.  doi: 10.3969/j.issn.2095-4344.2178 Abstract ( 612 )   PDF (646KB) ( 528 )   Save BACKGROUND: Microglia are resident immune cells of the central nervous system that normally perform sensing, housekeeping, and defense functions. In the context of neurodegenerative diseases, the dysfunction of microglia leads to or aggravates neuronal injury. OBJECTIVE: To investigate the mechanism of microglia-mediated neuronal injury in neurodegenerative diseases. METHODS: The first author searched for relevant articles published from January 2001 to January 2020 in PubMed, CNKI, Wanfang database, and VIP database with the key words of “microglia; neurodegenerative diseases; neuronal injury”.   RESULTS AND CONCLUSION: In neurodegenerative diseases, microglia perform excessive sensing due to toxic substances during normal function, leading to increasing activation of microglia, accompanied with hyperfunction of housekeeping and intense neuroinflammation causing neuronal impairment. The dysregulation can also be manifested as dysfunction of sensing and housekeeping due to specific gene mutations, which bring about accumulation of toxic substances, aggravating the dysregulation of defense function and inducing apoptosis or necrosis of neurons as a result. Further exploration on the mechanism of microglia-mediated neuronal injury in neurodegenerative diseases may provide several targets for the treatment of neurodegenerative disease. Figures and Tables | References | Related Articles | Metrics

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Photoreceptor cell replacement therapy for retinal degeneration diseases Li Shanshan, Guo Xiaoxiao, You Ran, Yang Xiufen, Zhao Lu, Chen Xi, Wang Yanling 2021, 25 (7):  1116-1121.  doi: 10.3969/j.issn.2095-4344.2179 Abstract ( 555 )   PDF (838KB) ( 231 )   Save BACKGROUND: There is no effective treatment for retinal degenerative diseases, among which the loss of photoreceptor cells, dysfunction and loss of photoreceptor cells are the main causes of retinal degenerative diseases. Photoreceptor cell transplantation as a promising cell replacement therapy is the main research direction today. OBJECTIVE: To review the progress of photoreceptor cell replacement in the treatment of retinal degeneration. METHODS: PubMed, CNKI and Wanfang databases were searched. The Chinese keywords were “photoreceptor cells, retina, transplantation” and the English keywords were “photoreceptors, retina, transplantation”. After preliminary screening by reading the titles and abstracts, the articles with low relevance to the subject were excluded, and a total of 62 articles were finally included for review. RESULTS AND CONCLUSION: The transplantation of exogenous cells into the retinal degeneration environment to replace the photoreceptors lost in advanced retinal degeneration has shown great advantages and provided a new therapeutic strategy for the diseases of retinal degeneration. However, standardized cell screening protocols for clinical use are still not perfect, which is an important challenge for future research. Figures and Tables | References | Related Articles | Metrics

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Advances in research and application of breast cancer organoids Jiao Hui, Zhang Yining, Song Yuqing, Lin Yu, Wang Xiuli 2021, 25 (7):  1122-1128.  doi: 10.3969/j.issn.2095-4344.2180 Abstract ( 739 )   PDF (1106KB) ( 108 )   Save BACKGROUND: Breast cancer is one of the most common malignant tumors in women. Its incidence rate is increasing year by year and tends to be younger. It seriously threatens women’s health. Therefore, it is particularly important to establish an ideal breast cancer model that can accurately simulate the tumor in vivo. Organoid is a new three-dimensional cultural model in vitro, which recapitulates key aspects of in vivo tissue or organ. In recent years, researches based on organoids have covered many kinds of tumors. OBJECTIVE: To review the research progress and application of breast cancer organoids, in order to provide a new research way for personalized treatment of breast cancer.  METHODS: Using the key words of “organoid, breast cancer organoids, cancer organoids, mammosphere, three-dimensional culture” in English and Chinese, respectively, the first author retrieved relevant articles published from January 1980 to February 2020 in CNKI, Wanfang, and PubMed databases. The type of the article was not limited. After removal of the articles that were not related to the purpose of the study or repetitive, 66 articles were finally analyzed. RESULTS AND CONCLUSION: This review introduced organoid technology briefly and retraced the process of exploring suitable culture conditions to establish breast-cancer-origin organoids. Also, we concluded latest development of its applications and research progress. Breast cancer organoids have a wide range of application prospects in disease modeling, tumor pathogenesis, drug screening and other aspects, which provide a reliable model for breast cancer research and treatment, and in particular, open up a new perspective for personalized treatment of breast cancer.  Figures and Tables | References | Related Articles | Metrics

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Effects of Chinese medicine on proliferation, differentiation and aging of bone marrow mesenchymal stem cells regulating ischemia-hypoxia microenvironment Wang Shiqi, Zhang Jinsheng 2021, 25 (7):  1129-1134.  doi: 10.3969/j.issn.2095-4344.2181 Abstract ( 512 )   PDF (785KB) ( 168 )   Save BACKGROUND: Traditional Chinese medicine has certain value and significance in the treatment of ischemic cardiovascular and cerebrovascular diseases by regulating the ischemia-hypoxia microenvironment and improving the survival rate and differentiation rate of stem cells.  OBJECTIVE: To sort out and analyze the research progress of traditional Chinese medicine on regulating ischemia-hypoxia microenvironment intervention on proliferation, differentiation, aging and autophagy of bone marrow mesenchymal stem cells in recent years. METHODS: The full-text database of Chinese journals, PubMed and Wanfang were retrieved with the keywords of “bone mesenchymal stem cells, ischemia-hypoxia microenvironment, proliferation, differentiation, aging” in English or “bone marrow mesenchymal stem cells, ischemia and hypoxia, proliferation, differentiation, aging” in Chinese for articles regarding effects of ischemia and hypoxia microenvironment on survival rate and differentiation rate of bone marrow mesenchymal stem cells published from 2002 to 2019. Fifty-five articles were selected for review, including 22 Chinese articles and 33 English articles. RESULTS AND CONCLUSION: The ischemia-hypoxia microenvironment is the important reason for the low survival rate and differentiation rate of bone marrow mesenchymal stem cells. There are many adverse reactions in the intervention of bone marrow mesenchymal stem cells with gene modification or cell molecules and drugs, which have become difficult problems to be solved in modern medicine. Exploring the internal relationship between microenvironment and stem cells using single or active components of traditional Chinese medicine combined with RNA transcriptomics is a new way to improve the viability of stem cells.  Figures and Tables | References | Related Articles | Metrics

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In vitro culture and purification of Schwann cells: a systematic review Zeng Yanhua, Hao Yanlei 2021, 25 (7):  1135-1141.  doi: 10.3969/j.issn.2095-4344.2182 Abstract ( 481 )   PDF (705KB) ( 58 )   Save OBJECTIVE: Schwann cells can promote the regeneration of damaged peripheral nerves and serve as seed cells in the engineering repair of peripheral nerve tissue. The effective culture and purification of Schwann cells are the basis of clinical treatment for peripheral nerve injury. This paper summarized the literature of culture of Schwann cells in vitro in recent ten years and made a descriptive review on the research progress of Schwann cell culture and purification, in order to provide references for culture of Schwann cells in vitro. METHODS: Literature related to Schwann cell isolation, culture and purification was retrieved from PubMed, Web of Science, Medline databases, CNKI, Wanfang, VIP and other databases. The keywords included “Schwann cells; isolation; culture; purification”. All the papers obtained from the search were read, analyzed and judged, and finally 62 papers meeting the standards were included. RESULTS: The classical methods of Schwann cell culture include tissue block culture and enzyme digestion. Purification methods include mitosis resistance, immune selection, specific adhesion, pre degeneration, cold jet, differential adherent, laminin package, low serum, stimulating factor, fluorescence activated cell sorting or magnetic activated cell sorting, immunopanning, and extracorporeal shock wave treatment.   CONCLUSION: Schwann cells can be cultured and purified in various ways in vitro, and each method has its advantages and disadvantages. How to obtain high-purity Schwann cells quickly and efficiently is still a challenge. Multiple methods can be combined for purification and affective factors can be controlled for Schwann cell proliferation as much as possible.  Figures and Tables | References | Related Articles | Metrics

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Efficacy of mesenchymal stem cells in the spinal cord injury of large animal models: a meta-analysis Kong Desheng, He Jingjing, Feng Baofeng, Guo Ruiyun, Asiamah Ernest Amponsah, Lü Fei, Zhang Shuhan, Zhang Xiaolin, Ma Jun, Cui Huixian 2021, 25 (7):  1142-1148.  doi: 10.3969/j.issn.2095-4344.2183 Abstract ( 524 )   PDF (1506KB) ( 80 )   Save OBJECTIVE: Mesenchymal stem cells transplantation is a promising treatment for spinal cord injury. Most of studies focused on small animal models. In large animal experiments, there are still controversies in selection of stem cells and therapeutic effect. This article analyzed the effects of mesenchymal stem cells on related indicators of spinal cord injury in large animal models and evaluated their effects on spinal cord injury repair. METHODS: PubMed, Cochrane, OVID, Web of Science and CNKI databases were retrieved before December 2019. A series of studies on the treatment of spinal cord injury in large animal models by mesenchymal stem cells were collected. According to the inclusion criteria, two researchers independently completed literature screening, data extraction and methodological quality evaluation, and meta-analysis was conducted with Stata16.0.   RESULTS:  A total of 10 articles were included. The results of meta-analysis showed that: (1) Mesenchymal stem cells could significantly improve motor function after spinal cord injury [I2=97.73%, MD=3.94, 95%CI (2.15, 5.72), P < 0.01]. Based on cell source, observation time, intervention phase, transplantation mode and graft type subgroup analysis showed that motor scores of bone marrow mesenchymal stem cells group, non-bone marrow mesenchymal stem cells group, short-term observation (< 2 months) group, long-term observation (≥ 2 months) group, acute stage group, non-acute stage group, cells group and allograft group were significantly higher than those of control group (P < 0.01). There was no significant difference in motor score between scaffold group and control group (P > 0.05). (2) The injury size in mesenchymal stem cells treatment group was significantly smaller than that in the control group [I2=98.05%, MD=-1.00, 95%CI (-1.95, -0.04), P=0.04]. (3) There was no significant difference in the relative expression of glial fibrillary acidic protein between the mesenchymal stem cells treatment group and the control group [I2=99.48%, MD=80.61, 95%CI (-27.48, 188.70), P=0.14].  CONCLUSION: Mesenchymal stem cells transplantation has a significant improvement on the motor function and injury repair of spinal cord injury in large animals, and the security is high. Due to the limitation of the quality of the included literature, the above conclusions need to be validated by high-quality and large-sample randomized controlled trials. Figures and Tables | References | Related Articles | Metrics