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    08 January 2024, Volume 28 Issue 1 Previous Issue   
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    Effect of CD34+ cell dose on haploidentical hematopoietic stem cell transplantation for treating malignant hematological diseases
    Peng Yingnan, Bian Zhilei, Zhang Suping, Li Li, Cao Weijie, Wan Dingming
    2024, 28 (1):  1-6.  doi: 10.12307/2023.749
    Abstract ( 296 )   PDF (2540KB) ( 68 )   Save
    BACKGROUND: Haploidentical hematopoietic stem cell transplantation is associated with a higher rate of graft rejection and therefore often requires a higher CD34+ cell dose, but the findings reported in existing studies regarding the relationship between CD34+ cell dose and study endpoints after allogeneic hematopoietic stem cell transplantation are controversial.  
    OBJECTIVE: To investigate the effect of CD34+ cell dose on clinical outcomes of haploidentical hematopoietic stem cell transplantation for malignant hematological diseases.
    METHODS: 135 patients who underwent haploidentical hematopoietic stem cell transplantation at Hematopoietic Stem Cell Transplantation Center, Department of Hematology, First Affiliated Hospital of Zhengzhou University between January 2019 and December 2021 were included. Combining the results of previous studies and our center’s experience, the cohort was divided into two groups using a CD34+ cell count of 5.0×106/kg as the cut-off point. Clinical outcomes related to graft implantation, relapse incidence, non-relapse mortality, overall survival and progression-free survival were evaluated in both groups.  
    RESULTS AND CONCLUSION: (1) CD34+ cell dose correlated with platelet engraftment, with platelets implanted earlier in the high-dose group than in the low-dose group (14 days vs. 16 days, P=0.013). (2) There was no significant difference in 3-year overall survival between the two groups (67.5% vs. 53.8%, P=0.257); nor was there a significant difference in progression-free survival between the two groups (65.6% vs. 44.2%, P=0.106), but stratified analysis based on disease risk index revealed an association with elevated 3-year progression-free survival in the high-dose group among low-risk patients (72.0% vs. 49.3%, P=0.036). (3) The cumulative 3-year relapse incidence was smaller in the high-dose group than in the low-dose group (16.0% vs. 33.5%, P=0.05). (4) The rate of non-relapse mortality within 100 days was greater in the high-dose group than in the low-dose group, but there was no significant difference (17.3% vs. 6.7%, P=0.070); stratified analysis revealed that non-relapse mortality within 100 days was significantly higher in the high-dose group than in the low-dose group (20.0% vs. 3.3%, P=0.046). (5) In conclusion, CD34+ cell doses >5.0×106/kg promote early platelet implantation, improve 3-year progression-free survival in low-risk patients at transplantation and reduce the cumulative relapse incidence. However, in high-risk patients, high-dose CD34+ cells result in increased non-relapse mortality within 100 days after transplantation, which is considered to be possibly associated with an increased occurrence of severe acute graft versus host disease in the early post-transplantation period. Therefore, it is considered that graft versus host disease monitoring should be enhanced in patients who transfused high-dose CD34+ cells.
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    Removal of the selective marker gene in the fusion gene expression vector of plant anti-caries vaccine
    Lang Yaoling, Wang Qian, Chen Bin, Bai Guohui, Guan Xiaoyan, Liu Jianguo
    2024, 28 (1):  7-11.  doi: 10.12307/2023.767
    Abstract ( 229 )   PDF (1675KB) ( 91 )   Save
    BACKGROUND: In consideration of the food safety and ecological safety of transgenic plants, the retention of marker genes is the primary safety issue affecting transgenic plants.  
    OBJECTIVE: Based on the principle of immune caries prevention, our research team successfully constructed the plant anti-caries vaccine fusion gene expression vector pCAMBIA-E8-APB-DOCK8 for these two caries causing virulence factors surface protein and glucosyltransferase, which provides a basis for the research and development of transgenic plant vaccine.
    METHODS: In this study, the selective marker genes Km and GUS in the plant caries vaccine fusion gene expression vector pCAMBIA-E8-APB-DOCK8 were removed by DNA recombination technology through a series of steps such as DNA fragment separation, connection, transformation, clone detection, and sequencing.  
    RESULTS AND CONCLUSION: The efficiency of marker gene removal was 99%. This study has laid a good experimental foundation for the safe production of transgenic plant vaccine against dental caries, and also provided ideas for the construction of other plant vaccine vectors.
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    Effect of lentiviral silencing of Piezo1 on osteogenic differentiation and TAZ expression in human bone marrow mesenchymal stem cells
    Wei Yurou, Tian Jiaqing, He Xianshun, Zhan Zhiwei, Wei Tengfei, Lin Tianye, He Wei, Wei Qiushi
    2024, 28 (1):  12-19.  doi: 10.12307/2023.901
    Abstract ( 318 )   PDF (2134KB) ( 64 )   Save
    BACKGROUND: Piezo1, a mechanosensitive protein, is tightly connected to osteogenic differentiation, and it has been demonstrated that TAZ has a role in regulating osteogenic differentiation. It is unclear whether TAZ participates in the regulation of osteogenic differentiation of human bone marrow mesenchymal stem cells by Piezo1, so it is crucial to investigate its unique mechanism to prevent osteonecrosis of the femoral head.
    OBJECTIVE: To elucidate what function Piezo1 plays in osteogenic differentiation and TAZ expression in human bone marrow mesenchymal stem cells. 
    METHODS: The siRNA targeting Piezo1 was constructed and transfected into 293T cells. The silencing efficiency was detected by RT-qPCR. The selected Piezo1-Home-2337 was packaged according to the silencing efficiency, and its optimal multiplicity of infection value was assayed by immunofluorescence staining. The packaged Piezo1 silencing recombinant lentivirus was transfected into human bone marrow mesenchymal stem cells, and its silencing effect was detected by RT-qPCR and western blot assay. Alizarin red staining, alkaline phosphatase activity analysis, immunofluorescence staining, RT-qPCR and western blot assay were utilized to analyze the effect of silencing Piezo1 on the osteogenic differentiation of human bone marrow mesenchymal stem cells.
    RESULTS AND CONCLUSION: (1) The mRNA and protein levels of Piezo1 in human bone marrow mesenchymal stem cells transfected by si-Piezo1 were decreased significantly, with a statistically significant difference compared with normal and negative control groups. (2) The alkaline phosphatase activity in the si-Piezo1 group was much lower and the calcium deposition in the si-Piezo1 group was significantly reduced compared with the negative control group. (3) The mRNA levels of osteogenesis-related genes including Runt-related transcription factor 2 (Runx2), osteopontin (OPN), distal-less homeobox 5 (DLX5), osteocalcin, β-catenin and Tafazzin (TAZ) in the si-Piezo1 group were significantly decreased compared with the negative control group. Afterward, the expression levels of TAZ and β-catenin protein in the si-Piezo1 group were down-regulated significantly compared with the negative control group, whereas the expression levels of p-TAZ and p-β-catenin protein in the si-Piezo1 group had the opposite condition. (4) The results of immunofluorescence staining showed that the expression of TAZ and β-catenin in human bone marrow mesenchymal stem cells in the si-Piezo1 group was less compared with the negative control group. (5) These findings indicate that Piezo1 can promote the osteogenic differentiation of human bone marrow mesenchymal stem cells. The osteogenic ability of human bone marrow mesenchymal stem cells is significantly reduced after silencing Piezo1, and the expression of TAZ is also reduced.
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    Mechanism underlying exosomal lncRNA H19 derived from umbilical cord mesenchymal stem cells promotes cartilage injury repair
    Wang Xianfeng, Wang Kun, Sun Han, Sun Xiaoliang, Yan Litao
    2024, 28 (1):  20-25.  doi: 10.12307/2023.917
    Abstract ( 313 )   PDF (2109KB) ( 39 )   Save
    BACKGROUND: Umbilical cord mesenchymal stem cells (UMSCs) have been proven to have therapeutic effects on cartilage injury, and exosomes are the main carriers for UMSCs to exert therapeutic effects in vivo. Our research group previously found that lncRNA H19 is an important active molecule that mediates the activity of UMSCs-derived exosomes regulating chondrocytes. LncRNA H19 could adsorb miR-29b-3p to promote the proliferation and regeneration of chondrocytes, but its downstream mechanism is still unclear. 
    OBJECTIVE: To reveal the specific mechanism of UMSCs in the treatment of cartilage injury from the perspective of exosomes and lncRNAs, so as to provide a new target for the treatment of cartilage injury. 
    METHODS: UMSCs stably overexpressing lncRNA H19 were constructed. H19-Exos were extracted by ultra-centrifugation. The exosomes were identified by transmission electron microscopy, Nanosight, western blot assay and exosome uptake assay. The effect of miR-29b-3p overexpression and silencing on the TGF-β1/Smad3 pathway was detected by western blot assay, qPCR and dual luciferase reporter gene system. The biological effect of H19-Exos on cartilage regeneration was verified by the specific TGF-β1/Smad3 inhibitor in vitro and in vivo. 
    RESULTS AND CONCLUSION: (1) H19-Exos showed a typical cup shape under an electron microscope, and the particle size was approximately 130 nm. H19-Exos expressed CD63, CD81 and TSG1010. (2) Overexpression of miR-29b-3p could down-regulate the mRNA and protein levels of TGF-β1 and Smad3, while silencing miR-29b-3p could up-regulate the mRNA and protein levels of TGF-β1/Smad3. (3) Dual-luciferase reporter gene system showed that miR-29b-3p had significant differences in the activities of downstream target genes TGF-β1 and Smad3. (4) The osteoarthritis models of rats were successfully established by injection of type II collagenase into the knee joint. H19-Exos significantly promoted cartilage regeneration. The specific TGF-β1/Smad3 inhibitor SB-431542 could block the biological effect of H19-Exos on cartilage regeneration in vitro and in vivo. (5) This study systematically demonstrated the promotion effect of UMSCs-derived exosomes highly expressing lncRNA H19 on cartilage regeneration, and the specific mechanism is that lncRNA H19 promotes cartilage regeneration by targeting miR-29b-3p/TGF-β1/Smad3 pathway. 
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    Irisin alleviates palmitic acid-induced osteogenic inhibition in bone marrow mesenchymal stem cells
    Zhang Yuanshu, He Xu, Xue Yuan, Jin Yesheng, Wang Kai, Shi Qin, Rui Yongjun
    2024, 28 (1):  26-31.  doi: 10.12307/2023.772
    Abstract ( 279 )   PDF (2003KB) ( 37 )   Save
    BACKGROUND: Irisin, a myokine isolated from the transmembrane protein FNDC5 by muscle cells during exercise, has the function of inducing the browning of white adipose tissue, but its effect on lipotoxicity-induced osteogenic differentiation and the mechanism is unclear.
    OBJECTIVE: To investigate the effect of irisin on the osteogenic ability of palmitic acid-induced bone marrow mesenchymal stem cells and the mechanism of action. 
    METHODS: CCK-8 assay was used to detect the effect of different concentrations of palmitic acid on the proliferation of mouse bone marrow mesenchymal stem cells and the effect of irisin on the proliferation of mouse bone marrow mesenchymal stem cells in the presence of palmitic acid. After pretreatment with irisin and palmitic acid for 24 hours, osteogenic differentiation of mouse bone marrow mesenchymal stem cells was induced by alkaline phosphatase staining as well as qRT-PCR was performed to detect the expression of osteogenesis-related genes on day 7 of osteogenic induction culture. The expression of proteins related to the AMPK/BMP2/SMAD signaling pathway was detected by western blot assay. Alizarin red staining was conducted on day 21 to detect osteogenic differences. 
    RESULTS AND CONCLUSION: (1) The CCK-8 assay results suggested that the amplification of bone marrow mesenchymal stem cells was inversely proportional to the concentration of palmitic acid, but at 0.02 mmol/L concentration, palmitic acid had no significant effect on the amplification of bone marrow mesenchymal stem cells, and irisin did not affect the proliferation of bone marrow mesenchymal stem cells when its mass concentration was in the range of 0.1-20 μg/L. (2) Alkaline phosphatase staining and alizarin red staining showed that palmitic acid inhibited the osteogenic differentiation ability of bone marrow mesenchymal stem cells. Irisin improved palmitic acid-induced osteogenic inhibition of bone marrow mesenchymal stem cells. qRT-PCR results showed that palmitic acid could cause the downregulation of osteogenic-related genes, and irisin could inhibit this trend. (3) Western blot assay results showed that compared with the palmitic acid intervention group, irisin treatment enhanced AMPK/BMP2/SMAD signal transduction in bone marrow mesenchymal stem cells. It is found that irisin can improve the osteogenic differentiation ability of bone marrow mesenchymal stem cells pretreated with palmitic acid, and proposed that the specific mechanism might be mediated by AMPK/BMP/SMAD signaling pathway.
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    Adipose-derived mesenchymal stem cells overexpressing bone morphogenetic protein 2 promote alveolar bone defect repair in osteoporosis rats
    He Lijun, Qi Xiaojuan
    2024, 28 (1):  32-37.  doi: 10.12307/2024.105
    Abstract ( 227 )   PDF (3407KB) ( 64 )   Save
    BACKGROUND: Jaws are most vulnerable to osteoporosis. Adipose-derived mesenchymal stem cells and bone morphogenetic protein 2 have the effect of promoting bone regeneration in osteoporosis. However, the repair effect of bone morphogenetic protein 2 modified adipose-derived mesenchymal stem cells on alveolar bone defects in osteoporosis is rarely reported.  
    OBJECTIVE: To explore the repair effect of adipose-derived mesenchymal stem cells overexpressing bone morphogenetic protein 2 on alveolar bone defects in osteoporosis rats.
    METHODS: (1) The rat adipose-derived mesenchymal stem cells were infected with lentivirus overexpressing bone morphogenetic protein 2 gene, and identified by detecting the expression of green fluorescent protein and bone morphogenetic protein 2. (2) Osteoporosis rat model was established by ovariectomy. A 3 mm × 3 mm × 3 mm cylindrical defect was prepared at the first molar position on both sides of the upper jaw. (3) Gelatin sponge was implanted in rats of the sham operation group and osteoporosis group. In the adipose-derived stem cell group, the adipose-derived mesenchymal stem cells infected with empty vector lentivirus and gelatin sponge complex were implanted. In the adipose-derived mesenchymal mesenchymal stem cell group overexpressing bone morphogenetic protein 2, a complex of adipose-derived mesenchymal stem cells overexpressing bone morphogenetic protein 2 and gelatin sponge was implanted. Relevant indexes were tested one month later.  
    RESULTS AND CONCLUSION: (1) The transfection efficiency of the adipose-derived mesenchymal stem cell group and adipose-derived mesenchymal stem cell group overexpressing bone morphogenetic protein 2 reached more than 70%. Compared with the adipose-derived mesenchymal stem cell group, the level of bone morphogenetic protein-2 protein in the adipose-derived mesenchymal stem cell group overexpressing bone morphogenetic protein-2 was significantly higher (P < 0.05). (2) A large amount of new bone could be seen in the bone defect area of the sham operation group. Compared with the sham operation group, the osteoporotic group had a small amount of new bone formation; the new bone area was significantly reduced, and alkaline phosphatase, osteocalcin, and bone morphogenetic protein 2 mRNA and protein levels were significantly reduced. Compared with the osteoporosis group, the adipose-derived mesenchymal stem cell group and the adipose-derived mesenchymal stem cell group overexpressing bone morphogenetic protein 2 had a large number of new bone formation; the area of new bone was significantly increased, and the levels of alkaline phosphatase, osteocalcin, and bone morphogenetic protein 2 mRNA and protein were significantly increased. Moreover, the adipose-derived mesenchymal stem cell group overexpressing bone morphogenetic protein 2 was superior to the adipose-derived mesenchymal stem cell group (all P < 0.05). (3) The results showed that bone morphogenetic protein 2 was less expressed in the alveolar bone of osteoporosis rats, and adipose-derived mesenchymal stem cells overexpressing bone morphogenetic protein 2 could promote osteogenesis and regeneration of alveolar bone defects in osteoporosis rats.
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    LncRNA SNHG4 regulates miR-152-3p during osteoblastic differentiation of periodontal ligament stem cells
    Zhou Minghua, Hu Xiaoyu
    2024, 28 (1):  38-43.  doi: 10.12307/2023.791
    Abstract ( 266 )   PDF (2639KB) ( 18 )   Save
    BACKGROUND: Studies have shown that long non-coding RNA small nucleolar RNA host gene 4 (lncRNA SNHG4) is involved in the progress of many inflammatory diseases, but the effect of lncRNA SNHG4 on the osteogenic differentiation of human periodontal ligament stem cells during the treatment of periodontitis is still unclear.  
    OBJECTIVE: To investigate the effect of lncRNA SNHG4 on the osteogenic differentiation of human periodontal ligament stem cells by regulating miR-152-3p.
    METHODS: Human periodontal ligament stem cells were isolated from periodontal membranes of premolars extracted for orthodontic purposes. After human periodontal ligament stem cells were induced to differentiate into osteoblasts for 0, 7, and 14 days, the expression levels of RUNX family transcription factor 2, osteocalcin mRNA, lncRNA SNHG4 and miR-152-3p in human periodontal ligament stem cells were detected by qRT-PCR. The third-generation human periodontal ligament stem cells were divided into the NC group, pcDNA group, pcDNA-SNHG4 group, inhibitor NC group, miR-152-3p inhibitor group, pcDNA-SNHG4+mimic NC group, and pcDNA-SNHG4+miR-152-3p mimic group. The expression of lncRNA SNHG4 and miR-152-3p in human periodontal ligament stem cells was detected by qRT-PCR. The proliferation of human periodontal ligament stem cells was detected by CCK-8 assay. Alkaline phosphatase activity was detected by colorimetry. The formation of mineralized nodules was detected by alizarin red staining. Western blot assay was used to detect the expression of RUNX family transcription factor 2, osteocalcin and alkaline phosphatase proteins. A double luciferase reporter gene experiment was applied to verify the relationship between lncRNA SNHG4 and miR-152-3p.  
    RESULTS AND CONCLUSION:  (1) The expression of RUNX family transcription factor 2, osteocalcin mRNA and lncRNA SNHG4 in human periodontal ligament stem cells after 7 and 14 days of osteogenic induction was higher than that after 0 days of osteogenic induction, while the expression of miR-152-3p was lower (P < 0.05). (2) Overexpression of lncRNA SNHG4 or inhibition of miR-152-3p was able to enhance the proliferation of human periodontal ligament stem cells, the alkaline phosphatase activity, mineralized nodule formation, the expression of RUNX family transcription factor 2, osteocalcin, and alkaline phosphatase proteins (P < 0.05). miR-152-3p mimic attenuated the promoting effect of overexpression of lncRNA SNHG4 on osteogenic differentiation of human periodontal ligament stem cells. LncRNA SNHG4 had a targeting relationship with miR-152-3p. (3) These findings indicate that overexpression of lncRNA SNHG4 may promote the osteogenic differentiation of human periodontal ligament stem cells by inhibiting miR-152-3p.
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    Mechanism underlying rat hepatocyte apoptosis regulated by exosomes derived from bone marrow mesenchymal stem cells
    Zheng Rongjiong, Deng Zerun, Han Dan, Sun Lihua
    2024, 28 (1):  44-49.  doi: 10.12307/2023.916
    Abstract ( 396 )   PDF (2651KB) ( 74 )   Save
    BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) can release a large number of exosomes (Exos). The effect of Exos derived from BMSCs on hepatocyte apoptosis and the specific mechanism has not been fully clarified.
    OBJECTIVE: To explore the effect of miR-21-5p carried by Exos derived from BMSCs on apoptosis of rat liver cells and its mechanism. 
    METHODS: Rat BMSCs were isolated and miR-21-5p NC or miR-21-5p inhibitor was transfected into BMSCs. The Exos were extracted by ultracentrifugation and named (BMSCs+miR-21-5p NC)-Exos and (BMSCs+miR-21-5p inhibitor)-Exos. BMSCs-derived Exos were co-cultured with rat hepatocytes to observe the effect of inhibiting miR-21-5p expression on the apoptosis of rat hepatocytes. The targeting relationship between miR-21-5p and PIK3R1 was verified by double luciferase reporter gene detection. TUNEL was used to detect the effect of miR-21-5p directly targeting PIK3R1 in Exos to activate the PI3K/AKT signaling pathway on hepatocyte apoptosis in BRL rats.
    RESULTS AND CONCLUSION: (1) The double luciferase reporting system confirmed that when PI3KR1 wild type vector and miR-21-5p mimics co-transfected 293T cells, the luciferase activity decreased significantly compared with the PI3KR1 mutant vector co-transfected group, indicating that miR-21-5p could target PIK3R1. (2) TUNEL test results showed that compared with (BMSCs+miR-21-5p NC)-Exos group, (BMSCs+miR-21-5p inhibitor)-Exos treatment significantly increased the apoptosis rate. Compared with the (BMSCs+miR-21-5p NC)-Exos group, after the addition of AKT inhibitor LY294002, the apoptosis rate was significantly increased. (3) The results indicate that Exos may inhibit the apoptosis of BRL rat hepatocytes through miR-21-5p, in which miR-21-5p directly targets PIK3R1 to activate PI3K/AKT signaling pathway. 
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    Human umbilical cord mesenchymal stem cell-derived exosomes reduce the permeability of blood-spinal cord barrier after spinal cord injury
    Zheng Mingkui, Xue Chenhui, Guan Xiaoming, Ma Xun
    2024, 28 (1):  50-55.  doi: 10.12307/2023.908
    Abstract ( 299 )   PDF (2077KB) ( 116 )   Save
    BACKGROUND: Endothelin has been found to be involved in the breakdown of the blood-spinal cord barrier after spinal cord injury, and stem cell-derived exosomes can reduce the permeability of the blood-spinal cord barrier and repair spinal cord injury.  
    OBJECTIVE: To investigate whether exosomes produced by human umbilical cord mesenchymal stem cells can reduce the permeability of the blood-spinal cord barrier by inhibiting endothelin-1 expression, thus repairing spinal cord injury.
    METHODS: Exosomes were extracted from the cultured supernatant by the hyperspeed centrifugation method. The morphology of exosomes was observed by transmission electron microscope. The expression levels of tsg101 and CD63 were detected by western blot assay. Eighty SD rats were randomly divided into sham operation group, model group, exosome group, and endothelin-1 group (n=20). The modified Allen’s method was used to create the rat model of spinal cord injury. In the endothelin-1 group, 10 μL (1 μg/mL) endothelin-1 was injected directly into the injured area with a microsyringe. Immediately, 1 day, 2 days after operation, sham operation group and model group were injected with 200 μL PBS solution through the tail vein; the exosome group and endothelin-1 group were injected with 200 μL exosome (200 μg/mL) solution through the tail vein, respectively. Hind limb motor function scores were performed on days 1, 3, 7, 14 and 21 after spinal cord injury. The blood-spinal cord barrier permeability was observed by Evans blue staining on day 7 after injury. The expression levels of tight junction proteins β-Catenin, ZO-1, Occludin and endothelin-1 in the spinal cord were detected by western blot assay.  
    RESULTS AND CONCLUSION: (1) Basso-Beattie-Bresnahan score in the exosome group was significantly higher than that in the model group at 3-21 days after injury (P < 0.05). Hematoxylin-eosin staining showed that spinal cord injury was greatly reduced in the exosome group compared with the model group. Basso-Beattie-Bresnahan score in the endothelin-1 group was significantly decreased compared with the exosome group (P < 0.05). Spinal cord injury was more severe in the endothelin-1 group than that in the exosome group. (2) The expression of endothelin-1 in the model group was significantly increased compared with the sham operation group (P < 0.05), and the expression of endothelin-1 in the exosome group was significantly decreased compared with the model group (P < 0.05). (3) The blood-spinal cord barrier Evans blue exudate in the exosome group was significantly decreased compared with the model group (P < 0.05). The expression levels of the tight junction proteins β-Catenin, Occludin and ZO-1 in the exosome group were increased (P < 0.05); the Evans blue exudate in the endothelin-1 group was significantly increased compared with the exosome group (P < 0.05). The expression level of tight junction protein was significantly decreased compared with the exosome group (P <0.05). (4) The results show that human umbilical cord mesenchymal cell-derived exosomes protect the permeability of the blood-spinal cord barrier by down-regulating the expression of endothelin-1 and play a role in the repair of spinal cord injury.
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    Recombinant human growth hormone promotes osteogenic differentiation of human dental pulp stem cells
    Sun Jing, Liao Jian, Sun Jiangling, Cheng Ping, Feng Hongchao
    2024, 28 (1):  56-61.  doi: 10.12307/2024.103
    Abstract ( 284 )   PDF (4229KB) ( 23 )   Save
    BACKGROUND: Previous studies have shown that human dental pulp stem cells have good osteogenic differentiation potential and are potential seed cells in bone tissue engineering, and the effect of recombinant human growth hormone on the proliferative osteogenic differentiation of human dental pulp stem cells is still unclear.
    OBJECTIVE: To explore the effect of recombinant human growth hormone on the proliferation and osteogenic differentiation of human dental pulp stem cells.
    METHODS: Human dental pulp stem cells were isolated and cultured by tissue block culture method. After screening according to the drug concentration gradient, recombinant human growth hormone containing 10, 100, 250, 500, 1 000 μg/L was selected as the experimental group, and 0 μg/L without recombinant human growth hormone was selected as the control group. CCK-8 detection reagents were used on days 1, 3, 5, and 7 after the drug intervention to detect the proliferation of human dental pulp stem cells. Different concentrations (10, 100, 250, 500, and 1 000 μg/L) of recombinant human growth hormone were added to the osteogenesis induction solution to intervene in human dental pulp stem cells. Alkaline phosphatase activity was detected by alkaline phosphatase staining and semi-quantitative analysis on day 7 of mineralization induction. The mRNA expression levels of osteogenic gene type I collagen, osteocalcin and Runt-related transcription factor 2 were detected by fluorescence quantitative RT-qPCR. Alizarin red staining was performed on day 14 of mineralization induction to detect osteogenic mineralized nodules.  
    RESULTS AND CONCLUSION: (1) CCK-8 assay results showed that from the third day of intervention, the 100, 250, 500, 1 000 μg/L recombinant human growth hormone group could promote the proliferation of human dental pulp stem cells compared with the control group (P < 0.01). (2) The alkaline phosphatase activity of human dental pulp stem cells in the 100, 250, and 500 μg/L recombinant human growth hormone group was significantly increased compared with the control group (P < 0.01). The number of alizarin-stained mineralized nodules in human dental pulp stem cells in the 100, 250 μg/L recombinant human growth hormone group was significantly increased compared with the control group (P < 0.01). Compared with the control group, the mRNA expression of type I collagen and osteocalcin increased in the 250 μg/L recombinant human growth hormone group (P < 0.05, P < 0.01). mRNA expression of Runt-associated transcription factor 2 increased in the 100 and 250 μg/L recombinant human growth hormone groups (P < 0.01). (3) According to the above results, recombinant human growth hormone at a concentration of 250 μg/L is a more suitable concentration to promote the proliferation and osteogenic differentiation of human dental pulp stem cells.
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    Effect of heme oxygenase-1-mediated atorvastatin on macrophage polarization and cholesterol accumulation
    Deng Rui, Huang Keming, Luo Jian, Chen Gong, Feng Jian, Huang Weiyi, Wei Gang
    2024, 28 (1):  62-67.  doi: 10.12307/2023.770
    Abstract ( 235 )   PDF (1790KB) ( 23 )   Save
    BACKGROUND: Studies have shown that atorvastatin can up-regulate the expression of heme oxygenase-1 and enhance the anti-inflammation and anti-oxidative damage ability of cells. However, whether atorvastatin can regulate macrophage polarization, inhibit inflammation and reduce cholesterol accumulation by inducing heme oxygenase-1 expression remains unclear. 
    OBJECTIVE: To investigate the effect of atorvastatin on polarization, inflammation and cholesterol content of oxidized low-density lipoprotein stimulated RAW264.7 macrophages by inducing heme oxygenase-1 expression and its related mechanism.
    METHODS: Firstly, RAW264.7 cells were randomly divided into six groups and incubated with different concentrations of atorvastatin for 24 hours. The expression of heme oxygenase-1 protein and cell activity were detected to explore the optimal dose of atorvastatin for subsequent studies. RAW264.7 cells were randomly divided into control group, atorvastatin group and heme oxygenase-1 inhibition group. Cells were preincubated with pure medium, atorvastatin 20 μmol/L and atorvastatin 20 μmol/L + zinc protoporphyrin IX 10 μmol /L for 24 hours, and then oxidized low-density lipoprotein 50 mg/L was added for 48 hours. The polarization of macrophages was detected by flow cytometry. The secretion of inflammatory factors such as transforming growth factor β, interleukin 10, interleukin 1β, and tumor necrosis factor α was detected by ELISA. The expression levels of heme oxygenase-1, LC3II, LC3I, P62, PPARγ and ABCA1 were detected by western blot assay. The intracellular cholesterol content was measured with the oxidose method and the accumulation degree of intracellular lipid droplets was evaluated by oil red O staining.
    RESULTS AND CONCLUSION: (1) Atorvastatin could induce the expression of heme oxygenase-1 protein in macrophages in a dose-dependent manner. (2) Oxidized low-density lipoprotein could induce macrophages to polarize towards M1, secrete proinflammatory factors, and increase the accumulation of intracellular cholesterol. (3) Compared with the control group, the heme oxygenase-1 protein expression of macrophages was increased after atorvastatin intervention, and the cells turned to M2-type polarization and mainly secreted anti-inflammatory factors such as transforming growth factor-β and interleukin-10. PPARγ, ABCA1, LC3II/I and other signal molecules reflecting cholesterol efflux and autophagy increased, and the contents of intracellular cholesterol and lipid droplets decreased significantly (P < 0.05). (4) The heme oxygenase-1 inhibition group treated with zinc protoporphyrin IX significantly reversed the above changes in the atorvastatin group. (5) The results have shown that atorvastatin may promote the polarization of macrophages stimulated by oxidized low-density lipoprotein to M2 type and inhibit inflammation by up-regulating the expression of heme oxygenase-1 and by up-regulating PPARγ/ABCA1 signaling pathway and enhancing autophagy. Atorvastatin can increase the outflow of intracellular cholesterol and reduce the accumulation of intracellular lipids. 
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    Effect of overexpression of protein phosphatase 2Cm on transcriptome of human renal tubular epithelial cells
    Zhang Li, Yang Wenjun, Sang Xiaohong, Han Yuanyuan, Mao Zhijie, Wang Shun, Lu Chen
    2024, 28 (1):  68-73.  doi: 10.12307/2023.915
    Abstract ( 252 )   PDF (6771KB) ( 39 )   Save
    BACKGROUND: A previous study by our group found that protein phosphatase 2Cm (PP2Cm) null mice developed significantly fewer symptoms of renal failure relative to wild-type mice, and thus it was speculated that PP2Cm may play an important protective role in the development of renal fibrosis, however, the molecular mechanisms remain undefined.
    OBJECTIVE: To investigate the effect of the PP2Cm gene on the transcriptome of human renal tubular epithelial cells.
    METHODS: Cultured human renal tubular epithelial cells were transfected with the PP2Cm gene into human renal tubular epithelial cells using plasmids. The expression of PP2Cm in the cells was detected by fluorescence quantitative PCR assay and western blot assay, and subsequently, cell RNA was separately extracted for transcriptome sequencing to look for differentially expressed genes between transfected and control groups. The resulting differential genes were further subjected to GO analysis and KEGG analysis using bioinformatics methods.
    RESULTS AND CONCLUSION: There were 796 differentially expressed genes, 553 of which were downregulated genes and 243 upregulated genes, in human renal tubular epithelial cells transfected with the PP2Cm gene compared with untransfected blank cells by sequencing analysis. GO analysis results showed that the upregulated genes were significantly enriched in cellular biosynthetic processes, protein translation, intrinsic apoptotic signaling pathways, and so on. The downregulated expressed genes were significantly enriched in endothelial cell proliferation, cell adhesion and other signaling pathways. KEGG analysis results showed that the significantly up-regulated genes were enriched in metabolism-related signaling pathways such as amino acid metabolism and biosynthesis. The downregulated expressed genes were significantly enriched in signaling pathways such as pantothenate and coenzyme A biosynthesis. Our results show that PP2Cm overexpression can affect a number of signaling pathways related to a range of biological processes in renal tubular epithelial cells, which may be important in metabolism-related signaling pathways such as amino acid metabolism and biosynthesis. 
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    Reversal effect of Lycium barbarum polysaccharide in combination with oxaliplatin on drug resistance of colon cancer stem cells
    Ai Fangfang, Xiao Hongyan, Wang Fang, Zhu Yongzhao, Ma Lijun
    2024, 28 (1):  74-79.  doi: 10.12307/2024.106
    Abstract ( 282 )   PDF (2209KB) ( 40 )   Save
    BACKGROUND: Clinical treatment for colon cancer mainly includes fluorouracil, irinotecan and oxaliplatin-based therapy. Studies have shown that membrane transport proteins such as ATP-binding cassette transport protein of G2 (ABCG2) mediate the transport of these drugs. However, when patients develop resistance to these chemotherapeutic drugs, the high expression of ABCG2 leads to a significant decrease in the therapeutic effect and raises the problem of drug resistance in colon cancer. New drugs and treatments are urgently needed to improve the efficacy. Lycium barbarum polysaccharide has a wide range of biological activities. It can be used as anti-tumor drug to overcome the damage to normal cells in the process of chemotherapy and radiotherapy in tumor patients.  
    OBJECTIVE: To explore the reversal effect of Lycium barbarum polysaccharide in combination with oxaliplatin on colon cancer drug-resistant cells through in vitro experiments to investigate the possible molecular mechanism of Lycium barbarum polysaccharide reversal on colon cancer drug-resistant cells.  
    METHODS: Colon cancer cell line HCT116 and oxaliplatin-resistant cell line HCT116-OXR were selected for in vitro experiments. The optimal intervention concentration and intervention time of Lycium barbarum polysaccharide and oxaliplatin were determined by CCK8 assay of cell proliferation. Samples were further divided into the HCT116 control group, HCT116-OXR blank treatment group, Lycium barbarum polysaccharide group (2.5 mg/mL Lycium barbarum polysaccharide), and oxaliplatin group (10 μmol/L oxaliplatin), and Lycium barbarum polysaccharide + oxaliplatin group (2.5 mg/mL Lycium barbarum polysaccharide +10 μmol/L oxaliplatin). Cell apoptosis was detected by flow cytometry. The protein expression levels of phosphomannose isomerase (PMI) and ABCG2 were detected by immunofluorescence and western blot assay. Phosphatidylinositol3-kinase (PI3K), protein kinase B (AKT), B-cell lymphoma 2 (Bcl-2) and BCL2-Associated X (Bax) were detected by western blot assay. 
    RESULTS AND CONCLUSION: (1) HCT116-OXR was more sensitive to Lycium barbarum polysaccharide compared to HCT116 (P < 0.05). (2) Compared with the HCT116-OXR blank group, Lycium barbarum polysaccharide + oxaliplatin could promote apoptosis of HCT116-OXR cells (P < 0.05). The protein expression of Bcl-2 was significantly down-regulated (P < 0.05); the protein expression of Bax was significantly up-regulated (P < 0.05); the protein expression of ABCG2, PMI, PI3K and AKT was significantly down-regulated (P < 0.05). (3) These results indicate that Lycium barbarum polysaccharide reverses drug resistance in colon cancer by inhibiting PMI/PI3K/AKT signaling pathway, which lays the foundation for studying the molecular mechanism of Lycium barbarum polysaccharide’s sensitizing chemotherapeutic effects.
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    Dapagliflozin attenuates endothelial cell pyroptosis and dysfunction induced by oxidized low-density lipoprotein
    Zhao Quanwei, Li Hui, Liu Danan, Gong Caiwei, Chen Long
    2024, 28 (1):  80-85.  doi: 10.12307/2023.773
    Abstract ( 303 )   PDF (1797KB) ( 53 )   Save
    BACKGROUND: Dapagliflozin, an inhibitor of sodium-glucose cotransporter 2, can delay the progression of atherosclerosis by regulating glucose metabolism, inhibiting inflammation and improving endothelial cell function.
    OBJECTIVE: To study the effect of dapagliflozin on cell pyroptosis and endothelial dysfunction induced by oxidized low-density lipoprotein.
    METHODS: Human umbilical vein endothelial cells were divided into a control group (no intervention), a model group (treated with oxidized low-density lipoprotein for 24 hours), and a dapagliflozin group (treated with oxidized low-density lipoprotein + dapagliflozin for 24 hours). Endothelial cell proliferation activity was measured by cell counting kit-8 assay. The levels of intercellular adhesion molecule 1, vascular cell adhesion molecule 1, and monocyte chemotactic protein-1 in cell supernatant were detected using ELISA. Nitric oxide level in the cells was detected by nitrate reductase assay. The pyroptosis rate and characteristics of endothelial cells were detected by Hoechst 33342/PI fluorescence co-staining and lactate dehydrogenase release assay. The protein expression levels of NLRP3, caspase-1, GSDMD, interleukin-1β, and interleukin-18 were detected by western blot assay.  
    RESULTS AND CONCLUSION: (1) Oxidized low-density lipoprotein could cause pyroptosis and dysfunction of endothelial cells. (2) Compared with the control group, the level of nitric oxide and cell activity were decreased (P < 0.05), while lactate dehydrogenase, intercellular adhesion molecule 1, vascular cell adhesion molecule 1, and monocyte chemotactic protein-1 levels were significantly increased in the model group (P < 0.05). Compared with the model group, cell activity and nitric oxide levels significantly increased (P < 0.05), but lactate dehydrogenase, intercellular adhesion molecule 1, vascular cell adhesion molecule 1, and monocyte chemotactic protein-1 levels were significantly diminished in the dapagliflozin group (P < 0.05). (3) Compared with the model group, cell pyroptosis rate and the protein expression of pyroptosis factor NLRP3, caspase-1, GSDMD, interleukin-18 and interleukin-1β significantly reduced in the dapagliflozin group (P < 0.05). (4) The results indicate that dapagliflozin inhibits oxidized low-density lipoprotein-induced endothelial pyroptosis and ameliorates endothelial cell dysfunction.
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    Research hot spots and trends of exosomes in theranostic application for chronic kidney disease
    Chen Guanting, Zhang Linqi, Li Qingru
    2024, 28 (1):  86-92.  doi: 10.12307/2023.765
    Abstract ( 423 )   PDF (15745KB) ( 39 )   Save
    BACKGROUND: To investigate the research focus and follow-up research trend of exosomes in the diagnosis and treatment of chronic kidney disease, in order to provide a corresponding reference basis for the future research of exosomes in the diagnosis and treatment of chronic kidney disease, and promote the development of this field.  
    OBJECTIVE: To conduct a bibliometric analysis of relevant studies in each database painstakingly until now for public publication on exosome diagnosis and treatment of chronic kidney disease, to explore the current state and trend of the field in this discipline, and to predict future research directions.
    METHODS: A computerized search was performed on WanFang, CNKI, CBM, VIP, Web of Science, Cochrane Library, PubMed, and Embase databases from inception to December 2022 for published literature related to the diagnosis and treatment of chronic kidney disease by exosomes. The literature transcripts were screened by NoteExpress for co-occurrence, clustering and mutational analysis among authors, institutions, and keywords through CiteSpace 6.1R4 and VOSviewer software, and the visual knowledge map was plotted.  
    RESULTS AND CONCLUSION: (1) A total of 804 articles, including 133 in Chinese and 671 in English, were included, and the volume of publications climbed year by year with a rapid trend. We included 3 649 literature authors, including 326 Chinese authors and 3 323 English authors, and the field has formed a core team centered on scholars such as Liu Bicheng, Wang Bin, Lyu LinLi, Wang Xiaonan and Wang Haidong, and has formed a stable multicenter collaboration platform among institutions. Research focuses on the three functions of exosomes: carrier, diagnosis and therapy. (2) As a form of extracellular vesicles, exosomes have important mechanisms for carrying, transferring molecular mediators and signal transduction, and have an important role in the physiopathological development of chronic kidney disease, which can provide important health surveillance data for epidemiological studies and clinical decision-making. In recent years, the development of relevant studies on exosome-based diagnosis of chronic kidney disease has expanded dramatically, forming a development layout of collaborative cooperation among multiple institutions worldwide, led by our scientific research institutions. However, at present, the study of the specific function and mechanism of action of exosomes and contents in the disease process has not been fully validated. Their significance for the early diagnosis and prognosis evaluation of chronic kidney disease is not very clear. The intrinsic mechanism of action-related research is still relatively poor. Isolation and purification techniques still need to be improved, and high-quality evidence-based clinical trials with multicenter and large samples have not yet appeared, which still need to be verified by further studies.
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    Single-cell RNA sequencing and the pathogenesis of intervertebral disc degeneration
    Cheng Haotian, Zhao Xiaofeng, Lu Xiangdong, Zhao Yibo, Fan Zhifeng, Qi Detai, Wang Xiaonan, Zhou Runtian, Jin Xinjie, Zhao Bin
    2024, 28 (1):  93-99.  doi: 10.12307/2023.768
    Abstract ( 301 )   PDF (1770KB) ( 60 )   Save
    BACKGROUND: Intervertebral disc degeneration is clinically considered to be the main cause of low back pain, but due to the unclear pathogenesis of intervertebral disc degeneration, there is still a lack of effective means to delay the progression of the disease. Single-cell RNA sequencing technology can amplify and sequence mRNA at the single-cell level, reveal the gene expression intensity of a single cell, discover different cell subsets in tissues according to the heterogeneity of cells, study the pathogenesis of intervertebral disc degeneration at the molecular level, and provide a new theoretical basis for its early diagnosis and treatment.
    OBJECTIVE: To introduce the basic principles of single-cell RNA sequencing technology and review the research progress of single-cell RNA sequencing technology in intervertebral disc degeneration in recent years.
    METHODS: A computer was used to search PubMed, Web of Science, CNKI and WanFang databases for the literature published from 2012 to 2022. Key words were “single-cell RNA sequencing, intervertebral disc degeneration, sequencing Technology” in Chinese and English. Duplicate, poor-quality and irrelevant articles were excluded; a total of 70 articles were eventually included. 
    RESULTS AND CONCLUSION: (1) We identified new cell subsets such as homeostatic chondrocytes, hypertrophy chondrocyte-like nucleus pulposus cells and fibrous nucleus pulposus cells, identified the marker genes and transcription factors of these cell subsets, and described the functions, differentiation paths and cell fate of these cell subsets during the development and progression of intervertebral disc degeneration, and proposed the concept of progenitor nucleus pulposus cells. A cell subpopulation with progenitor nucleus pulposus cells properties was identified and its effectiveness in treating intervertebral disc degeneration was verified in mice. (2) Fibro chondrocyte-like annulus fibrosus cells and annulus fibrosus stem cells with both cartilage and fiber properties were identified, and a new type of composite hydrogel was prepared by combining fibrous cartilage inducers silk fibroin and hyaluronic acid in vitro. Experiments in mice demonstrated that this hydrogel could repair both annulus fibrosus tissue and cartilage matrix, and was remarkably effective in the treatment of intervertebral disc degeneration. (3) Regulatory chondrocytes were found in endplate cartilage. Two distinct fates in the progression of intervertebral disc degeneration were analyzed and the differential genes in the two fates were identified. Intercellular communication analysis indicated that regulatory chondrocytes interact with endothelial cells to promote angiogenesis. (4) Immune cells such as macrophages, T cells, myeloid progenitor cells and neutrophils were identified in the degenerated intervertebral disc tissues, demonstrating the existence of immune response during intervertebral disc degeneration. It was found that apolipoprotein induced the polarization of macrophages M1 and M2 subtypes, and this polarization process affected the activity of progenitor nucleus pulposus cells by amplifying the inflammatory response through the MIF signaling pathway.
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    Characteristics, advantages and application of osteogenic differentiation of jaw bone marrow mesenchymal stem cells
    Fan Yongjing, Wang Shu, Jin Wulong
    2024, 28 (1):  100-106.  doi: 10.12307/2023.790
    Abstract ( 306 )   PDF (1596KB) ( 31 )   Save
    BACKGROUND: The repair of maxillofacial bone tissue defects is a hot and difficult point in current research and the selection of seed cells is the key. Jaw bone marrow mesenchymal stem cells are adult mesenchymal stem cells that exist in the jaw bone. They have advantages in the application of maxillofacial tissue regeneration.
    OBJECTIVE: To summarize the biological characteristics, osteogenic differentiation advantages of jaw bone marrow mesenchymal stem cells, and the effects of drugs, in vivo environment, and microRNAs on the osteogenic differentiation of jaw bone marrow mesenchymal stem cells.
    METHODS: Computers were used to perform literature retrieval in PubMed and CNKI. Chinese and English search terms were “oral, bone tissue engineering, stem cells”. 405 articles were retrieved and downloaded. The articles were screened according to the inclusion and exclusion criteria and 70 articles were finally included for literature review.
    RESULTS AND CONCLUSION: Jaw bone marrow mesenchymal stem cells were excellent seed cells for oral bone tissue engineering, and had good proliferation and osteogenic differentiation potential. Drugs, in vivo environment and microRNAs could regulate the osteogenic differentiation of jaw bone marrow mesenchymal stem cells. However, the research on jaw bone marrow mesenchymal stem cells was still in the initial stage, so more research with strong demonstration is needed to confirm that jaw bone marrow mesenchymal stem cells have more advantages in the application of maxillofacial bone tissue regeneration.  
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    Application prospect of mesenchymal stem cells in promoting muscle tissue repair
    Huang Yongbin, Wang Tao, Lou Yuanyi, Pang Jingqun, Chen Guanghua
    2024, 28 (1):  107-112.  doi: 10.12307/2023.769
    Abstract ( 181 )   PDF (1632KB) ( 45 )   Save
    BACKGROUND: Mesenchymal stem cells are multipotent stromal cells isolated from bone marrow, fat, umbilical cord and other tissues. It can differentiate into different cell types and secrete a variety of proteins with therapeutic potential, which has a good application prospect in the repair of muscle tissue.
    OBJECTIVE: To review the research progress of mesenchymal stem cells in promoting muscle tissue repair and provide a theoretical basis for further clinical application.

    METHODS: Relevant articles published from inception to 2022 were retrieved from CNKI, VIP, WanFang, PubMed, Embase and Web of Science databases. The keywords were “mesenchymal stem cells, muscle tissue, muscle injury, muscle atrophy, exosomes, scaffolds” in Chinese and English. The literature about mesenchymal stem cell migration promoting muscle fiber proliferation and repair was screened. Finally, 98 articles were included for review and analysis.

    RESULTS AND CONCLUSION: (1) The related mechanisms of mesenchymal stem cell migration promoting muscle fiber proliferation and repair are complex, mostly by anti-inflammatory, inhibiting interstitial fibrosis, inhibiting the fat formation and other ways to promote muscle fiber proliferation and repair. (2) The related biological scaffolds and cell co-culture based on mesenchymal stem cells can significantly compensate for the low survival rate of mesenchymal stem cells after colonization. (3) At present, mesenchymal stem cell therapy still has apparent limitations. In the future, mesenchymal stem cells combined with other therapies should become the primary development trend. 

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    Construction methods and application of assembloids
    Liu Chunlei, , , Yao Xi, Wei Zhengbo, Xie Ying
    2024, 28 (1):  113-120.  doi: 10.12307/2023.921
    Abstract ( 284 )   PDF (1823KB) ( 65 )   Save
    BACKGROUND: In recent years, many studies have confirmed that assembloids can make up for the shortcomings of organoids, which cannot fully reproduce the interaction between cell and cell and between cell and matrix. Since the assembloids construction methods are in the early stage of development, there is no unified standard. 
    OBJECTIVE: To review the current construction methods, applications, advantages, and disadvantages of assembloids, guide the development and improvement of vitro cell models.
    METHODS: PubMed, CNKI, and WanFang databases were searched with English search terms “assembloids, organoids, tumor microenvironment, organoids AND assemble, organoids AND microenvironment” and Chinese search terms “assembloids, organoids, tumor microenvironment, organoid reorganization, multicellular model”. Totally 94 articles were screened out for review after excluding irrelevant articles and deduplication. 
    RESULTS AND CONCLUSION: (1) According to the different sources of cells, the construction of assembloids can be divided into three methods: self-assembly, direct-assembly, and mixed-assembly. According to the differences of cell culture methods, it can be divided into suspension culture method, matrix culture method, organ chip culture method, and 3D bio-printing. (2) The process of self-assembly covers early stages of cell and tissue development, so it has broad prospects in the fields of organ development and developmental disorders. The function of differentiated mature cells is relatively perfect, and the assembloids directly assembled by them have more potential in the study of functional disorders and cell-damaging diseases. Self-assembly may be better in organ transplantation, and direct-assembly will be more suitable for the repair of tissue damage. Mixed-assembly combines the advantages of the former two and is mostly used to explore the physiological and pathological mechanisms of cells in the microenvironment, as well as drug screening. (3) Although different assembloids have their own advantages, they all face the problem of imperfect vasculature system, then, each method has its own limitations, for example, the degree of cell differentiation in self-assembly assembloids may still be different from that in vivo, and the fixed cell types in direct-assembly models cannot simulate complex microenvironments in vivo. These are urgent problems to be solved. (4) In the future, with the continuous improvement of assembloids culture technology, scientists can assemble biomimetic organoids with more complex tissues in vitro, providing infinitely realistic models for the study of physiological and pathological processes of human tissue and organ.
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    Effect of traditional Chinese medicine and compounds for supplementing qi and activating blood circulation and inducing resuscitation on regulating stem cells to promote nerve repair of acute ischemic stroke
    Ying Chunmiao, Pan Xiaolong, Liu Feixiang, Chen Na, Fan Feiyan, Zhang Yunke
    2024, 28 (1):  121-130.  doi: 10.12307/2023.781
    Abstract ( 256 )   PDF (1768KB) ( 49 )   Save
    BACKGROUND: Endogenous neurogenesis and exogenous stem cell transplantation in the brain show great therapeutic potential for neurological diseases including ischemic stroke, repairing and replacing lost neurons, promoting synaptic remodeling, and inhibiting apoptosis. Traditional Chinese medicine and compound therapy for supplementing qi, activating blood circulation and inducing resuscitation for the treatment of neurological dysfunction after ischemia have certain advantages, targeting nerve repair through a variety of ways, including promoting endogenous neurogenesis and exogenous stem cell survival, proliferation, homing, and inducing neuronal differentiation.
    OBJECTIVE: To summarize the mechanism of traditional Chinese medicine and compound for supplementing qi, activating blood circulation and inducing resuscitation to promote nerve repair in the acute phase of ischemic stroke, in order to provide a reference for the research and treatment of new drugs in ischemic stroke.
    METHODS: The articles from CNKI and PubMed databases about traditional Chinese medicine and compound for supplementing qi, activating blood circulation and inducing resuscitation in promotion of nerve repair in the acute phase of ischemic stroke from 2010 to 2022 were searched, with “supplementing qi and activating blood circulation; inducing resuscitation; traditional Chinese medicine (TCM); compounds; ischemic stroke; nerve repair; stem cells” as Chinese and English search terms. After excluding old and duplicate views, the retrieved literature was analyzed and collated, and a total of 124 articles were included for analysis.
    RESULTS AND CONCLUSION: (1) The definition of stem cells, ischemic stroke and the nerve repair pathway in the acute phase of ischemic stroke were sorted out. (2) The mechanism of action of traditional Chinese medicine and compound for supplementing qi, activating blood circulation and inducing resuscitation to promote nerve repair in the acute phase of ischemic stroke was summarized, mainly including promoting stem cell proliferation, improving stem cell viability and survival rate, promoting nerve cell homing, inducing stem cell differentiation to neurons, inhibiting apoptosis of nerve cells, promoting axon regeneration, regulating angiogenesis and remodeling, improving the level of neurotrophic factors and repairing the integrity of the blood-brain barrier. (3) Through the existing research, the relevant factors and signaling pathways of traditional Chinese medicines and compounds for supplementing qi, activating blood circulation and inducing resuscitation to promote nerve repair in the acute phase of ischemic stroke were summarized, such as Nestin protein expression, DCX protein expression, brain-derived neurotrophic factor, vascular endothelial growth factor and Wnt/β-catenin signaling pathway, Notch signaling pathway, PI3k/Akt signaling pathway, BDNF/TrkB signaling pathway and ERK/MAPK signaling pathway. It provides a relevant reference for future research on ischemic stroke-specific drugs and new clinical treatment methods.
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    Astrocytes regulate glial scar formation in cerebral ischemic stroke
    Yang Ting, Ding Zhibin, Jiang Nan, Han Hongxia, Hou Miaomiao, Ma Cungen, Song Lijuan, Li Xinyi
    2024, 28 (1):  131-138.  doi: 10.12307/2023.742
    Abstract ( 351 )   PDF (1728KB) ( 52 )   Save
    BACKGROUND: Cerebral ischemic stroke is one of the main fatal and disabling diseases in the clinic, but only a few patients benefit from vascular recanalization in time, so it is urgent to explore new and effective therapy. As one of the critical pathological changes of ischemic stroke, the glial scar formed mainly by astrocytes is one major cause that hinders axonal regeneration and neurological recovery at the late stage of stroke.  
    OBJECTIVE: To elucidate the pathological process and crucial signal regulatory mechanism of astrocytes in the formation of glial scar after ischemic stroke, as well as the potential therapeutic targets, to provide a theoretical reference for intervening astrocytic scar formation against ischemic stroke effectively, and novel strategies for promoting post-stroke rehabilitation.
    METHODS: The relevant articles published in CNKI, PubMed and Web of Science databases from 2010 to 2022 were retrieved. The search terms were “Ischemic stroke, Brain ischemi*, Cerebral ischemi*, Astrocyt*, Astroglia*, Glial scar, Gliosis, Astrogliosis” in Chinese and English. Finally, 78 articles were included after screening and summarized.  
    RESULTS AND CONCLUSION:  (1) Astrocytes play an important role in the maintenance of central nervous system homeostasis. After ischemic stroke, astrocytes change from a resting state to an active state. According to the different severities of cerebral ischemic injury, astrocyte activation changes dynamically from swelling and proliferation to glial scar formation. (2) Mature astrocytes are stimulated to restart the cell cycle, then proliferate and migrate to lesions, which is the main source of the glial scar. Neural stem cells in the subventricular zone, neuron-glial antigen 2 precursor cells and ependymal precursor cells in the brain parenchyma can also differentiate into astrocytes. Endothelin-1, aquaporin 4, ciliary neurotrophic factor and connexins are involved in this process. In addition, chondroitin sulfate proteoglycan, as the main component of the extracellular matrix, forms the dense glial scar barrier with proliferated astrocytes, which hinders the polarization and extension of axons. (3) Activation or inhibition of crucial signal molecules involved in astrocyte activation, proliferation, migration and pro-inflammation functions regulate the glial scar formation. Transforming growth factor beta 1/Smad and Janus kinase/signal transducer and activator of transcription 3 are classical pathways related to astrogliosis, while receptor-interacting protein 1 kinase and glycogen synthase kinase 3β are significant molecules regulating the inflammatory response. However, there are relatively few studies on Smad ubiquitination regulatory factor 2 and Interleukin-17 and their downstream signaling pathways in glial scar formation, which are worthy of further exploration. (4) Drugs targeting astrogliosis-related signaling pathways, cell proliferation regulatory proteins and inflammatory factors effectively inhibit the formation of glial scar after cerebral ischemic stroke. Among them, the role of commonly used clinical drugs such as melatonin and valproic acid in regulating glial scar formation has been verified, which makes it possible to use drugs that inhibit glial scar formation to promote the recovery of neurological function in patients with stroke. (5) Considering the protective effects of glial scar in the acute phase, how to choose the appropriate intervention chance of drugs to maintain the protective effect of the glial scar while promoting nerve regeneration and repair in the local microenvironment is the direction of future efforts. 
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    Single-cell RNA sequencing reveals the heterogeneity of astrocytes
    Long Qingxi, Zhang Pingshu, Liu Qing, Ou Ya, Zhang Lili, Yuan Xiaodong
    2024, 28 (1):  139-146.  doi: 10.12307/2023.743
    Abstract ( 323 )   PDF (2585KB) ( 106 )   Save
    BACKGROUND: Astrocytes are the most abundant cells in the central nervous system, and various subsets of astrocytes are heterogeneous, performing a variety of special functions. Single-cell RNA sequencing (scRNA-seq) technology developed in recent years has extended our understanding of astrocyte heterogeneity from the perspective of transcriptome profiling.  
    OBJECTIVE: To summarize the heterogeneity of scRNA-seq technology in different time and space, and pathological states and expand our knowledge of astrocyte heterogeneity on both molecular and functional levels.
    METHODS: The relevant articles on astrocyte heterogeneity and scRNA-seq were searched on PubMed, Elsevier, and CNKI databases. The search terms were “astrocytes, scRNA-seq, heterogeneity, Alzheimer disease, spinal cord injury, multiple sclerosis” in Chinese and English. Finally, 74 articles were selected for viewing after screening according to inclusion criteria.  
    RESULTS AND CONCLUSION: scRNA-seq studies related to the heterogeneity of astrocytes have shown that astrocyte is significantly heterogeneous across four aspects: species, developmental stage, central nervous system region, and pathological state. (1) Unique expression of certain genes occurs in astrocytes of different species, and the discovery of species-specific genes is beneficial for the translation of clinical studies. (2) During astrocyte development, differential gene expression emerged in the cellular subtypes identified at each stage, which further refined the cellular lineage of astrocytes and laid the foundation for the study of astrocyte developmental trajectories and mechanisms. (3) The discovery of differential gene expression allows regional localization of different astrocyte subpopulations and assists in the diagnosis and treatment of neurological diseases. (4) Astrocyte heterogeneity revealed by scRNA-seq can provide specific markers at the time of disease diagnosis and identify potential therapeutic targets. (5) The heterogeneity of astrocytes exists in many aspects, interacts with each other and is complex. The mechanisms of its generation, maintenance and transformation remain unclear. At present, molecular research on the single-cell level is still lacking. Linking transcriptionally defined astrocyte subpopulations to cellular activity, behavior and disease markers in real time remains one of the great challenges in the field.
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    Effects of different exercise types on human DNA damage, DNA methylation and telomere length
    Yang Pei
    2024, 28 (1):  147-152.  doi: 10.12307/2023.906
    Abstract ( 443 )   PDF (1614KB) ( 44 )   Save
    BACKGROUND: Exercise is not only an effective means to improve physical and mental health, but also has a good intervention effect on the occurrence and development of metabolic, cardiovascular and cerebrovascular diseases. The reason is related to the epigenetic factors.  
    OBJECTIVE: To summarize the effects of different exercise types on human DNA damage, DNA methylation, and telomere length, and analyze the possible mechanism of exercise regulation epigenetic modification, in order to provide a reference for exercise to improve body function.
    METHODS: “Exercise, aerobic training, acute exercise, anaerobic training, resistance training, DNA damage, DNA methylation, telomere” were used as the Chinese search terms, and “exercise, sport, aerobic exercise, anaerobic exercise, resistance training, acute exercise, DNA metabolism, DNA damage, telomere” were used as the English search terms. We searched PubMed, Embase, Web of Science, and CNKI databases, and screened articles according to inclusion and exclusion criteria, and finally included 70 articles.
    RESULTS AND CONCLUSION: (1) Long-term aerobic, resistance, and anaerobic exercises can improve DNA damage. The reason is that exercise can improve the body’s antioxidant capacity. Acute exercise can aggravate the degree of DNA damage by up-regulating the expression of reactive oxygen species and reactive nitrogen oxides. (2) Acute exercise, long-term resistance exercise, and anaerobic exercise play a positive role in reducing DNA methylation. The key mechanism may be that exercise-induced reactive oxygen species changes the expression of glutathione oxidized/glutathione, DNA methyltransferase, and 10-11 translocation enzyme. Then it can regulate DNA methylation. (3) Compared with other types of exercise, long-term aerobic exercise may have more potential value in increasing telomere length, and its biological mechanism involves inflammation, oxidative stress, DNA methylation, and regulation of microRNAs (miRNAs) expression. (4) Based on the current literature, aerobic exercise lasting at least 2 years can increase telomere length, and future research should further clarify the optimal exercise duration.
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    Traditional Chinese medicine monomer in the prevention and treatment of flap necrosis by regulating “autophagy”
    Ma Suilu, He Zhijun, Liu Tao, Li Yan, He Yuanxu, He Bo, Wang Weiwei, Wei Xiaotao
    2024, 28 (1):  153-158.  doi: 10.12307/2023.755
    Abstract ( 296 )   PDF (1732KB) ( 29 )   Save
    BACKGROUND: In recent years, it has been found that some traditional Chinese medicine monomers can alleviate oxidative stress and apoptosis of the skin flap, promote vascular regeneration of the skin flap and prevent skin flap necrosis by activating autophagy. 
    OBJECTIVE: To review the research progress of traditional Chinese medicine monomer regulating autophagy in preventing flap necrosis. 
    METHODS: The Chinese and English key words were “traditional Chinese medicine (TCM), autophagy, skin flaps”. The first author searched the relevant articles published in CNKI and PubMed databases from January 2010 to October 2022. A total of 196 articles were retrieved in the preliminary screening and then screened according to the inclusion and exclusion criteria. The quality assessment was conducted by reading the literature titles and abstracts. Finally, 55 articles were summarized. 
    RESULTS AND CONCLUSION: (1) The regulation of autophagy is mediated by AMPK/mTOR, PI3K/AKT and other signaling pathways. Activation of autophagy can alleviate the oxidative stress and apoptosis of the flap, promote the regeneration of blood vessels in the flap, and prevent flap necrosis. (2) Terpenoids (Betulinic acid, Andrographolide, Notoginseng Triterpenes, Catalpa), phenolic compounds (Resveratrol, Curcumin, Gastrodin), phenolic acids (Salvianolic acid B) and steroid compounds (Pseudoginsenoside F11) in traditional Chinese medicine monomers can alleviate oxidative stress and apoptosis of skin flap by regulating related signaling pathways to activate autophagy, promote skin flap angiogenesis and promote skin flap survival. (3) Studying the research progress of traditional Chinese medicine monomer to prevent flap necrosis by regulating autophagy can provide a reference and theoretical basis for traditional Chinese medicine to prevent flap necrosis and promote flap healing in the clinic. 
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