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BACKGROUND: The small-diameter blood vessel prosthesis faces the problem of lumen stenosis or even occlusion due to thrombogenesis and intimal hyperplasia after transplantation. The application of stem cells as seed cells to achieve endothelialization of blood vessel prosthesis helps to improve the long-term patency rate after vascular graft transplantation. OBJECTIVE: To summarize the research progress of the application of stem cells in the endothelialization of small-diameter blood vessel prosthesis. METHODS: The relevant articles published on PubMed and WanFang databases from 2013 to 2023 were retrieved by the first author. Chinese and English search terms included “vascular graft, tissue-engineered blood vessel/vascular tissue engineering, endothelialization, stem cells, endothelial progenitor cells, mesenchymal stem cells, induced pluripotent stem cells, embryonic stem cells.” The relative articles in the domestic and overseas about the application of stem cells in the endothelialization of small-diameter blood vessel prosthesis in the past 10 years were retrieved. A total of 552 articles were initially found and we finally selected 81 articles to review according to inclusion and exclusion criteria. RESULTS AND CONCLUSION: (1) The low long-term patency rate restricts the application of small-diameter blood vessel prosthesis in clinic. The main causes of low long-term patency rate are thrombogenesis and intimal hyperplasia. The endothelium of native vessels has the function of anti-thrombogenesis and intimal hyperplasia. Endothelialization can simulate the function of native vessels, which is an effective way to improve long-term patency rate. (2) The small-diameter blood vessel prosthesis will undergo in vivo endothelialization after being implanted, but it is difficult to form complete endothelium. Stem cells have the potential to differentiate into endothelial cells. Recruiting stem cells in vivo or seeding them on the inner surface of blood vessel prosthesis in vitro is research strategy to achieve endothelialization. (3) The long-term patency rate of small-diameter blood vessel prosthesis has been improved to a certain extent through seeding endothelial progenitor cells, mesenchymal stem cells, induced pluripotent stem cells, and embryonic stem cells. Each has its own advantages. Endothelial progenitor cells are easy to obtain and can be directly used for seeding. Mesenchymal stem cells come from a wide range of sources and have the function of paracrine and immunological regulation. Induced pluripotent stem cells are rich in sources and the immunogenicity can be eliminated. Embryonic stem cells have a strong proliferative ability and can differentiate into many cells. (4) The application of stem cells in blood vessel prosthesis has not yet been transformed into clinic. Further researches are needed to promote clinical translation. [ABSTRACT FROM AUTHOR]

BACKGROUND: The self-renewal and multi-directional differentiation of pluripotent stem cells possess the potential to revolutionize people’s understanding of biology, medicine, development, and disease. Stem cells play an important role in the early stage of embryonic development, and the study of them could be beneficial to understanding of the basic principles of biological development and tissue or organ formation, exploring the potential mechanisms of various diseases, studying the repair and regeneration of damaged tissues or organs, and promoting drug discovery and personalized treatment. OBJECTIVE: To review the research progress of pluripotent stem cells, summarize and categorize the fundamental types of pluripotent stem cells, and elucidate the lineage situations of various types of pluripotent stem cells in common mammals. METHODS: PubMed, Web of Science, CNKI, and WanFang databases were searched systematically, with the keywords “pluripotent stem cells; embryonic stem cells; induced pluripotent stem cells; expanded potential stem cells; livestock pluripotent stem cells” in English and Chinese. The 99 articles related to mammalian pluripotent stem cells were systematically screened according to inclusion and exclusion criteria, and then reviewed. RESULTS AND CONCLUSION: (1) According to classical theory in mouse embryonic stem cell research, the pluripotent state of stem cells is divided into two forms: naïve and primed. Naïve state corresponds to the inner cell mass of pre-implantation embryos before attachment to the uterine wall, while primed state corresponds to the epiblast after implantation. These two states exhibit significant differences in epigenetic features, transcriptional activity, external signal dependency, and metabolic phenotype. It is later discovered that there is an intermediate state between naïve and primed called formative pluripotency. Therefore, the pluripotency of pluripotent stem cells is a continuous developmental process rather than a unique cell state. (2) In addition to obtaining pluripotent stem cells from the inner cell mass, there are various methods and lineages for acquiring pluripotent stem cells, including embryonic germ cells established using primitive germ cells from mouse embryos, induced pluripotent stem cells created by the dedifferentiation of adult mouse and human fibroblasts with four factors-Oct3/4, Sox2, c-Myc, and Klf4; embryonic stem cell-like cell lines cultured from somatic cell nuclear transfer, parthenogenesis, neonatal or adult testicular or ovarian tissue, very small embryonic-like stem cells derived from various adult tissues and expanded pluripotent stem cells derived from pre-implantation stages. These pluripotent stem cells all share the common characteristics of continuous self-renewal, expressing core pluripotency factors and possessing the ability to differentiate into the three primary germ layers. (3) Currently, pluripotent stem cells are being used for disease modeling to study the mechanisms of various diseases and develop new drugs. Simultaneously, scientists are attempting to use pluripotent stem cells to cultivate various tissues and organs, offering new possibilities for regenerative medicine and transplantation. However, the clinical application of pluripotent stem cells faces safety challenges, including issues of cell mutations and immune rejection. Continual improvement in the methods of generating pluripotent stem cells will make them safer and more efficient for clinical applications. (4) Based on the methods of obtaining and lineage establishment of pluripotent stem cells in mice and humans, various types of pluripotent stem cells have been established in livestock, including embryonic stem cells, induced pluripotent stem cells, germ lineages of pluripotent stem cells, and expanded potential stem cells. Research on livestock pluripotent stem cells opens up new avenues for animal reproduction, breeding, genetic engineering, disease modeling, drug screening, and the conservation of endangered wildlife. [ABSTRACT FROM AUTHOR]

BACKGROUND: Autoimmune regulator gene (Aire) and Wnt signaling pathway play an important role in the maintenance and differentiation of mouse embryonic stem cell pluripotency. However, whether the Wnt signal and Aire are involved in the differentiation of embryonic stem cells to thymic epithelial progenitor cells remains poorly understood. OBJECTIVE: To investigate the relationship of the Wnt signaling pathway and Aire with the differentiation of embryonic stem cells. METHODS: A two-step differentiation method was used to induce mouse embryonic stem cells to differentiate into endoderm and then into thymic epithelial progenitor cells. Mouse embryonic stem cells were infected with Aire shRNA lentivirus, and monoclonal stable strains were screened by puromycin. Mouse embryonic stem cells were collected on days 0, 3 and 10 of the directed induction of differentiation after the induced differentiation by the two-step differentiation method. Cellular immunofluorescence, flow cytometry, western blot assay, and real-time qPCR were used to detect the expression changes of related genes and proteins. RESULTS AND CONCLUSION: (1) Immunofluorescence staining showed positive expression of SSEA1 and OCT4 on day 0 of targeted induction of differentiation. (2) Immunofluorescence staining showed double-positive expression of SOX17 and FOXA2 on day 3 of targeted induction of differentiation. (3) Flow cytometry results showed positive expression of EPCAM1, K5 and K8 on day 10 of targeted induction of differentiation. (4) Compared with undifferentiated mouse embryonic stem cells, the expressions of Wnt7a, β-catenin, and Gsk-3β proteins were elevated, and the expression level of Aire protein was decreased in induced differentiated thymic epithelial progenitor cells. (5) Compared with undifferentiated mouse embryonic stem cells, the expressions of Wnt7a, β-catenin, Gsk-3β and Aire mRNA were elevated in thymic epithelial progenitor cells. (6) Compared with normal cultured mouse embryonic stem cells and their ultimately differentiated thymic epithelial progenitor cells, the expression levels of Wnt7a, β-catenin and Gsk-3β proteins were reduced in mouse embryonic stem cells with knockdown of Aire genes and their final differentiated thymic epithelial progenitor cells. In conclusion, the Wnt signaling pathway and Aire are jointly involved in the process of targeted induction of differentiation of mouse embryonic stem cells into mouse thymic epithelial progenitor cells. [ABSTRACT FROM AUTHOR]

BACKGROUND: Stem cells are self-renewing cells with differentiation potential and stem cell therapy is a promising therapeutic approach. Macrophage migration inhibitory factor is expressed in almost all mammalian cells and is essential for many physiological processes. Current studies have demonstrated that macrophage migration inhibitory factor is expressed on a variety of stem cells and regulates the proliferation, differentiation and migration of many stem cells. OBJECTIVE: To describe the basic characteristics of macrophage migration inhibitory factor and to elaborate on the role and mechanism of macrophage migration inhibitory factor on stem cells. METHODS: PubMed and CNKI databases were searched for relevant articles published in the past ten years. The key words were “macrophage migration inhibitory factor, cancer stem cell, embryonic stem cell, neural stem cell, mesenchymal stem cell, endothelial progenitor cell, stem cell therapy, tissue engineering” in Chinese and English, respectively. After removal of poorly related, outdated, and duplicate studies by reading the title and abstract, 85 articles were finally included in result analysis. RESULTS AND CONCLUSION: (1) Macrophage migration inhibitory factor protein consists of three subunits that interact on the cell surface by interacting with receptors CD74, CD44, CXCR2, CXCR4 and CXCR7. Macrophage migration inhibitory factor is stored intracellularly and secreted through non-classical pathways, and a variety of factors can affect macrophage migration inhibitory factor expression as well as secretion. (2) Macrophage migration inhibitory factor promotes migration, invasion, metastasis, escape and tumorigenicity of tumor stem cells through different signaling pathways. (3) Currently, it is relatively clear that macrophage migration inhibitory factor promotes neural differentiation of mouse embryonic stem cells. (4) Macrophage migration inhibitory factor promotes the proliferation and survival of neural stem cells through multiple signaling pathways. (5) Macrophage migration inhibitory factor acts on mesenchymal stem cells to inhibit apoptosis, promote survival, delay senescence, and increase paracrine effects of mesenchymal stem cells. In addition, macrophage migration inhibitory factor has also been shown to inhibit mesenchymal stem cell homing by binding to the receptor CD74. (6) Several studies have demonstrated that macrophage migration inhibitory factor activates downstream signaling pathways through CXCR4 receptors to promote endothelial cell migration and revascularization. (7) Macrophage migration inhibitory factor acts on tumor stem cells, embryonic stem cells, neural stem cells, mesenchymal stem cells, and endothelial progenitor cells to affect their proliferation, differentiation, migration, and apoptosis processes. Some of these effects are still controversial, and the exact mechanisms need to be further verified. [ABSTRACT FROM AUTHOR]

BACKGROUND: Embryonic stem cells have a great potential in helping to understand cell development, regenerative medicine, tissue engineering, and cell therapy due to their self-renewal and multidirectional differentiation characteristics. To have a deeper understanding and effective use of human embryonic stem cells, it is necessary to explore and discover the potential molecules that affect the proliferation and survival of human embryonic stem cells. Macrophage migration inhibitory factor is expressed in a variety of human cells. At first, it was thought to be mainly involved in inflammation, but it was later found to play an important role in cell proliferation, differentiation, angiogenesis, and tumorigenesis. However, whether human embryonic stem cells express this important factor and their functions in human embryonic stem cells remain unclear. OBJECTIVE: To determine whether human embryonic stem cells express macrophage migration inhibitory factor and its related receptors, determine which receptor interacts with macrophage migration inhibitory factor, and initially explore the effect of macrophage migration inhibitory factor on the proliferation and survival of human embryonic stem cells. METHODS: Human embryonic stem cells were cultured. The level of macrophage migration inhibitory factor in the cell culture medium was detected by enzyme-linked immunosorbent assay. The distribution of macrophage migration inhibitory factor in the cell was observed by immunofluorescence confocal microscope. Cell immunofluorescence and western blot assay were utilized to detect the expression of embryonic stem cell related factors. Immunofluorescence confocal microscopy and immunoprecipitation were applied to detect the binding of macrophage migration inhibitory factor and its receptor. Group intervention: Human embryonic stem cells were intervened with ISO-1 (0, 12, 24, 48, 96, 192 μmol/L) for 24 hours to determine the best inhibitory concentration. Different mass concentrations of macrophage migration inhibitory factor (30, 100, 300 μg/L) were incubated with the optimal inhibitory concentration of ISO-1 for 24 hours. CCK-8 assay was used to detect cell proliferation activity. TUNEL staining was used to detect cell apoptosis level. RESULTS AND CONCLUSION: (1) Macrophage migration inhibitory factor was expressed in the cytoplasm, membrane and culture medium of human embryonic stem cells. (2) Human embryonic stem cells expressed macrophage migration inhibitory factor, mainly expressing its receptors CXCR2 and CXCR7. (3) Macrophage migration inhibitory factor mainly bound to the receptor CXCR7. (4) The effect of macrophage migration inhibitory factor inhibitor ISO- 1 on cell proliferation and survival: Compared with the blank group, as the ISO-1 concentration increased, the cell gap increased significantly and the cell viability gradually decreased (P < 0.0001). TUNEL assay showed that cell apoptosis rate was significantly increased (P < 0.000 1). (5) After adding different mass concentrations of migration inhibitory factor (30, 100, 300 μg/L), it could weaken ISO-1 (192 μmol/L) induced negative effects on cells; cell viability was significantly increased (P < 0.05); and apoptosis rate was significantly reduced (P < 0.01). (6) The results have shown that human embryonic stem cells express and secrete macrophage migration inhibitory factor, which is an important factor and can be detected in the cytoplasm, cell membrane, and outside the cell. Human embryonic stem cells mainly express the receptors CXCR2 and CXCR7 of macrophage migration inhibitors, instead of the classic receptor CD74. The protein receptor that interacts with macrophage migration inhibitory factor on human embryonic stem cells is CXCR7. No evidence of interaction with CXCR2 has been found, and its role needs further study. In addition, studies have found that macrophage migration inhibitory factors play an important role in maintaining the proliferation and survival of human embryonic stem cells. [ABSTRACT FROM AUTHOR]

BACKGROUND: Transplantation therapy after the directional differentiation of induced pluripotent stem cells can solve the immune elimination and ethical problems, and directional differentiation of induced pluripotent stem cells into cardiomyocytes provides the possibility of cardiomyocyte regeneration research. OBJECTIVE: To detect the electrophysiological function of cardiomyocytes differentiated from induced pluripotent stem cells. METHODS: By culturing induced pluripotent stem cells in vitro, direct suspension culture and differentiation medium were used to induce induced pluripotent stem cells to form embryoid bodies. Cell growth was observed and formation time, number and contraction rate of cardiomyocytes were recorded. The electrophysiological characteristics of cardiomyocytes were detected by Axon Axopatch 200B single probe ultra-low noise patch clamp amplifier and MED64 multi-electrode array system. The responsiveness of differentiated cardiomyocytes to isoproterenol was observed. Cardiac troponin T expression levels in differentiated cardiomyocytes were detected by immunofluorescence. RESULTS AND CONCLUSION: (1) The growth characteristics of induced pluripotent stem cells were similar to embryonic stem cells, resembling a bird’s nest and growing like a colony. (2) In the differentiation medium culture, induced pluripotent stem cells aggregated to form embryoid bodies of different sizes after 5 days of suspension culture. During the adherent growth of embryoid bodies, up to 10% of the colonies appeared beating cardiomyocytes, and the beating rate on the 20th and 25th days was significantly higher than other time points (P < 0.05). (3) Action potentials of ventricular, atrial and sinoatrial nodular cells were observed in induced pluripotent stem cells differentiated cardiomyocytes. (4) Isoproterenol could increase the spontaneous beat frequency of cardiomyocytes differentiated from induced pluripotent stem cells. (5) Cellular immunofluorescence indicated that cardiomyocytes differentiated from induced pluripotent stem cells were positive for cardiac troponin T. (6) The results confirm that cardiomyocytes differentiated from induced pluripotent stem cells have sinoatrial-like, atrial-like and ventricular-like action potentials, and have adrenaline signal transduction. [ABSTRACT FROM AUTHOR]

Objective · To identify the RNA binding proteins of Oct4, a key factor of embryonic stem cells. Methods · Total RNA was extracted moembryonic stem cells (R1), which was reverse-transcribed into cDNA. Then PCR product of Oct4 mRNA were transcribed into Oct4 mRNA. Finally the candidate RNA-binding proteins were eluted applying the streptomycin affinity chromatography, and submitted for high-performance liquid chromatography combined with the mass spectrometry. The identified RNA binding proteins were further confirmedRNA immunoprecipitation (RIP). Results · 121 RNA binding proteins of Oct4 3 untranslated region (UTR) and Oct4 5 UTR were identified with liquid chromatography / mass spectrometry. There were 11 proteins with the number of peptide spectrum matches more or equal to 2, and 7 of them were selected for additional confirmation using RIP method. RIP results showed that Oct4 interacted with DSP, SOD1, LMNA, NPM1, PSIP1, NCL and HDGF. Among them, HDGF had the strongest binding ability to Oct4 mRNA. Conclusion · Identification of Oct4 RNA binding proteins will provide a theoretical basis for the regulation mechanism of Oct4, and will be a basis for the further study of its function. [ABSTRACT FROM AUTHOR]

Octamer-binding transcription factor 4 (OCT4) belongs to the POU-homeodomain family of transcription factors and binds to an octamer motif, ATGCAAAT. OCT4 is the key transcription factor that is involved in the maintenance of pluripotency and self-renewal in undifferentiated embryonic stem (ES) cells. OCT4 has been reported to be overexpressed in various cancers including lung, germ cell tumors, breast, cervix, prostate, gastric, liver, and ovarian cancer. MicroRNAs (miRNAs), small non-coding RNAs, modulate mRNA expression through base pairing between seed sequences in miRNA and complementary sequences of the target mRNA, thereby destabilizing mRNA and/or inhibiting protein synthesis. Several miRNAs have been demonstrated to regulate stemness factors such as OCT4, NANOG, SOX2 and KLF4 in cancer cells, thereby modulating the proliferation, apoptosis, differentiation, drug resistance and immunity of cancer cells.

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The article evaluates the effects of culture optimization to Kunming species mouse embryonic stem cells in Jiangxi, China.

Objective: To explore the role of small nuclear noncoding RNA 7SK in embryonic stem cell (ESCs) proliferation and the value of 7SK as a target for early diagnosis and treatment for primordial dwarfism (PD)., Methods: ESC line R1 was transfected with the CRISPR/Cas9 system, and sequencing of the PCR product and glycerol gradient analysis were performed to identify novel 7SK deletion mutations. A lentivirus system was used to knock down cyclin-dependent kinase 9 (CDK9) in clones with 7SK deletion mutations, and the effect of CDK9 knockdown on the protein level of cell division cycle 6 (CDC6) was analyzed with Western blotting., Results: We identified a novel deletion mutation of 7SK at 128-179 nt in the ESCs, which resulted in deficiency of cell proliferation. 7SK truncation at 128-179 nt significantly reduced the protein expressions of La-related protein 7 (LARP7) and CDC6., Conclusions: 7SK truncation at 128-179 nt can significantly impair proliferation of ESCs by downregulating CDC6. 7SK is a key regulator of proliferation and mediates the growth of ESCs through a mechanism dependent on CDK9 activity, suggesting the value of 7SK truncation at 128-179 nt as a potential target for early diagnosis and treatment of PD.

Objective: To assess the risk of male infertility in the offspring conceived through assisted reproductive technology (ART) byin vitroinductionof the differentiation of embryonic stem cells (ESCs) derived from the embryos of the couples with male asthenozoospermia and Robertsonian translocation (RT) into germ cells., Methods: We established a CCRM16ESC line with the karyotype of 46, XY, +14, rob(13; 14) (q10; q10) from the embryo donated by a patientwithasthenozoospermiaand RT and his wife by isolation of the inner cell mass of blastula, culturing, passaging, and amplification,followed by in vitro induction and differentiationof the ESCs into germ cells with ratinoic acid(RA) at 2 mol/L. Then, we analyzed the process of differentiation and the expressions of its related genes and compared them with those in the normal CCRM23ESCs., Results: CCRM16 showed the typical characteristics of ESCs, expressing the pluripotency makers of NANOG, OCT4, TRA-1-181 and SSEA4, forming embryoid bodies, and differentiating into three germlayer tissues in vitro and in vivo. Intervention with 2 mol/LRAinduced direct differentiation of the ESCs into germ cells. The expressions of the primordial germ cell marker geneDAZLand the meiosis marker geneSCP3were markedly decreased in the CCRM16 as compared with those in the normal CCRM23 ESCs., Conclusions: The CCRM16ESC linewith the karyotype of46, XY, +14, rob(13; 14) (q10; q10) has thetypical characteristics of ESCs but an abnormal process of differentiation into germ cells in the early stage. In vitroinductionof the differentiation of ESCs into germ cells can be used for assessing the risk of male infertility in the offspring conceived through ART for asthenozoospermia patients.