RESUMO
The diversity of A mating type in wild strains of Lentinula edodes was extensively analyzed to characterize and utilize them for developing new cultivars. One hundred twenty-three A mating type alleles, including 67 newly discovered alleles, were identified from 106 wild strains collected for the past four decades in Korea. Based on previous studies and current findings, a total of 130 A mating type alleles have been found, 124 of which were discovered from wild strains, indicating the hyper-variability of A mating type alleles of L. edodes. About half of the A mating type alleles in wild strains were found in more than two strains, whereas the other half of the alleles were found in only one strain. About 90% of A mating type combinations in dikaryotic wild strains showed a single occurrence. Geographically, diverse A mating type alleles were intensively located in the central region of the Korean peninsula, whereas only allele A17 was observed throughout Korea. We also found the conservation of the TCCCAC motif in addition to the previously reported motifs, including ATTGT, ACAAT, and GCGGAG, in the intergenic regions of A mating loci. Sequence comparison among some alleles indicated that accumulated mutation and recombination would contribute to the diversification of A mating type alleles in L. edodes. Our data support the rapid evolution of A mating locus in L. edodes, and would help to understand the characteristics of A mating loci of wild strains in Korea and help to utilize them for developing new cultivars.
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Many of early findings regarding intestinal stem cells (ISCs) and their niche in the human intestine have relied on colorectal cancer cell lines and labor-intensive and time-consuming mouse models. However, these models cannot accurately recapitulate the physiologically relevant aspects of human ISCs. In this study, we demonstrate a reliable and robust culture method for 3D expanding intestinal spheroids (InSexp ) mainly comprising ISCs and progenitors, which can be derived from 3D human intestinal organoids (HIOs). We did functional chararcterization of InSexp derived from 3D HIOs, differentiated from human pluripotent stem cells, and optimization culture methods. Our results indicate that InSexp can be rapidly expanded and easily passaged, and show enhanced growth rates via WNT pathway activation. InSexp are capable of exponential cell expansion and cryopreservation. Furthermore, in vitro-matured HIO-derived InSexp proliferate faster than immature HIO-derived InSexp with preservation of the parental HIO characteristics. These findings may facilitate the development of scalable culture systems for the long-term maintenance of human ISCs and provide an alternative platform for studying ISC biology.
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Innovative advances in stem cell research have resulted in the development of organoids, which are widely used as in vitro models of human organ development and for disease. The long-term goals of scientists include the generation of high-quality organoids with properties like those of native organs, and to expand their use to a variety of applications such as drug discovery and organoid-based cell therapy. In particular, the combination of human induced pluripotent stem cell (iPSC)-derived organoids with the recently developed genome engineering, biotechnology serve as an attractive platform in precision medicine. This review briefly summarizes the generation of organoids derived mostly from iPSCs without ethical issues, and describes the applications and technological advances of organoids under their differentiation and culture conditions. We also discuss the approaches to improve the organoid models, and how organoids can recapitulate mature organ systems of the human body for regenerative medicine. Finally, the future perspectives and remaining challenges in the field have been discussed to provide a better understanding of the potential applications of organoids.
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In Parkinson’s disease (PD) research, human neuroblastoma and immortalized neural cell lines have been widely used as in vitro models. The advancement in the field of reprogramming technology has provided tools for generating patient-specific induced pluripotent stem cells (hiPSCs) as well as human induced neuronal progenitor cells (hiNPCs). These cells have revolutionized the field of disease modeling, especially in neural diseases. Although the direct reprogramming to hiNPCs has several advantages over differentiation after hiPSC reprogramming, such as the time required and the simple procedure, relatively few studies have utilized hiNPCs. Here, we optimized the protocol for hiNPC reprogramming using pluripotency factors and Sendai virus. In addition, we generated hiNPCs of two healthy donors, a sporadic PD patient, and a familial patient with the LRRK2 G2019S mutation (L2GS). The four hiNPC cell lines are highly proliferative, expressed NPC markers, maintained the normal karyotype, and have the differentiation potential of dopaminergic neurons. Importantly, the patient hiNPCs show different apoptotic marker expression. Thus, these hiNPCs, in addition to hiPSCs, are a favorable option to study PD pathology.
Assuntos
Humanos , Linhagem Celular , Neurônios Dopaminérgicos , Fibroblastos , Técnicas In Vitro , Células-Tronco Pluripotentes Induzidas , Cariótipo , Neuroblastoma , Neurônios , Patologia , Vírus Sendai , Células-Tronco , Doadores de TecidosRESUMO
Autoimmune diseases (AIDs), a heterogeneous group of immune-mediated disorders, are a major and growing health problem. Although AIDs are currently treated primarily with anti-inflammatory and immunosuppressive drugs, the use of stem cell transplantation in patients with AIDs is becoming increasingly common. However, stem cell transplantation therapy has limitations, including a shortage of available stem cells and immune rejection of cells from nonautologous sources. Induced pluripotent stem cell (iPSC) technology, which allows the generation of patient-specific pluripotent stem cells, could offer an alternative source for clinical applications of stem cell therapies in AID patients. We used nonintegrating oriP/EBNA-1-based episomal vectors to reprogram dermal fibroblasts from patients with AIDs such as ankylosing spondylitis (AS), Sjögren's syndrome (SS) and systemic lupus erythematosus (SLE). The pluripotency and multilineage differentiation capacity of each patient-specific iPSC line was validated. The safety of these iPSCs for use in stem cell transplantation is indicated by the fact that all AID-specific iPSCs are integrated transgene free. Finally, all AID-specific iPSCs derived in this study could be differentiated into cells of hematopoietic and mesenchymal lineages in vitro as shown by flow cytometric analysis and induction of terminal differentiation potential. Our results demonstrate the successful generation of integration-free iPSCs from patients with AS, SS and SLE. These findings support the possibility of using iPSC technology in autologous and allogeneic cell replacement therapy for various AIDs, including AS, SS and SLE.
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Humanos , Doenças Autoimunes , Fibroblastos , Técnicas In Vitro , Células-Tronco Pluripotentes Induzidas , Lúpus Eritematoso Sistêmico , Células-Tronco Pluripotentes , Espondilite Anquilosante , Transplante de Células-Tronco , Células-Tronco , TransgenesRESUMO
Embryonic stem cells (ESCs) are an emerging source for cell-based therapies aimed at repairing damaged organ tissues; however, the efficiency of directed differentiation is low and refinement of differentiation protocols is hampered by incomplete understanding of the mechanisms involved in this process. To find new compounds which can improve the efficiency of directed differentiation of ESCs to cardiomyocytes, we screened several thousand chemical compounds and identified a promising group. All of the compounds found have a common structure of 1H-pyrrole,2,2'-(phenylmethylene)bis. Here we report the potential mechanism of action for 31002 which showed the strongest activity among the compounds selected. In the presence of 31002, 15 times more cardiomyocytes differentiated from ESCs, i.e., 3.5% to 52% of total differentiated cells. Moreover, the cardiomyocytes showed functional characteristics including rhythmic beating and marker gene expression. 31002 inhibited the down-regulation of genes related to the three germ layers in the late stage of ESCs differentiation, implying that 31002 supports a continuous fate commitment of undifferentiated ESCs to the cardiac lineage by prolonging the three germ layer stages. Therefore, compounds in this group, including 31002, might be useful as directed cardiomyogenic differentiation-inducers to produce cells for use in cell therapy aimed at restoring damaged heart tissue.
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Regulação para Baixo , Células-Tronco Embrionárias , Expressão Gênica , Camadas Germinativas , Coração , Miócitos Cardíacos , Terapia Baseada em Transplante de Células e TecidosRESUMO
Leiomyomas of the appendix are rare and most are encountered incidentally during exploration of the abdomen for some other disease, during postmortem examination, or in the course of routine pathologic examinations of surgical specimens. We report here the findings of ultrasonography, CT and surgery of a case of leiomyoma that arose from the appendix; this lesion was pathologically confirmed.
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Abdome , Apêndice , Autopsia , Leiomioma , UltrassonografiaRESUMO
BACKGROUND: Vimentin is the major intermediate-size filament in the cytoplasm of cells from mesenchymal origin. The HL-60 cell is a unique human leukemic cell line capable of terminal differentiation with several chemical inducers, and then the cell line becomes a fre#quently described model system for cell differentiation in vitro. Vimentin mRNA is reduced during all-trans retinoic acid (retinoic acid) -dependent differentication but increased by 12-0-tetradecanoylphorbol-13-acetate (TPA). In this paper, we have investigated on the mechanism of transcriptional repression of vimentin gene during retinoic acid-dependent differentication of HL-60 cell. METHODS: HL-60 cells were grown in RPMI 1640 medium supplemented with 10% heat-inactivated fetal bovine serum and antibiotics in a humidified 5% CO at 37C. Total RNA was prepared by a modification of the method of Karlinsey et al. Northern blot hybridization was performed by the method of Virca et al. EcoRI fragment of pVIM-GEM was used as probe for vimentin mRNA. DNA mobility shift assay was performed by the method of Lim et al. End labeled DNA probe (Upper strand, 5-CGCITGATGAGTCAGCCG-3) for AP-1 binding activity was mixed with nuclear extracts in a 20 pL reaction volume containing 300 mM KCI, 60 mM HEPES, pH 7.9, 25mM MgC1, 1mM EDTA, 1mM DTT, 60% glycerol, and 2 pg of poly[dI-dC]. RESULTS: The level of vimentin mRNA was decreased at 12 hours after retinoic acid treatment, and not detected at 48 hours. The level of vimentin mRNA was reduced in proportion to concentration of retinoic acid, Retinoic acid-reduced vimentin mRNA was no change in cells treated with cycloheximide. Retinoic acid-dependent decrease of vimentin mRNA was partially recovered by staurosporin pretreatment. In DNA mobility shift assay, AP-1 binding activity was reduced at 48 hr during retinoic acid-induced differentiation. CONCLUSION: These results suggest that the transcriptional repression of vimentin gene during retinoic acid-induced differentiation in HL-60 cells is correlated with reduction of DNA binding activity of AP-1.
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Humanos , Antibacterianos , Northern Blotting , Diferenciação Celular , Linhagem Celular , Cicloeximida , Citoplasma , DNA , Ácido Edético , Ensaio de Desvio de Mobilidade Eletroforética , Glicerol , HEPES , Células HL-60 , Concentração de Íons de Hidrogênio , Repressão Psicológica , RNA , RNA Mensageiro , Fator de Transcrição AP-1 , Tretinoína , VimentinaRESUMO
Computerized Tomography is now well established and important noninvasive method of diagnosting mediastinal mass lesions because of its superior imaging of their size, location and internal composition. Authors analyzed and present CT findings of 30 surgically proven mediastinal tumors and cysts that were studied and treated at the Yeungnam University Hospital during recent 6 years. The most common tumor was thymoma (9 cases), and teratoma (6 cases), lymphoma (6 cases), bronchogenic cyst (4 cases), neurogenic tumor (4 cases), pericardial cyst (1 case) were next in order of frequency. There were 5 cases of thymoma showing homogenous solid density mass, 2 cases were malignant thymoma and myasthenia gravis was present in 2 cases. A case of thymolipoma and a case of thymic carcinoma were included. All teratomas were cystic masses but pathognomonic fat, and calcified density were seen only in 4 cases. 5 cases were located in anterior mediastinum and 1 case was in posterior mediastinum. Lymphoma (3 Hodgkin's and 3 non-Hodgkin's) appeared as irregular lobulated mass in anterior mediastinum. Neurogenic tumor (2 ganglioneuroma and 2 neurilemmoma) appeared as homogenous density mass located in posterior mediastinum. Among the 4 bronchogenic cysts, 2 were located in retrotracheal area, 1 was located in subcarinal and 1 was in parathoracic area. One case of pericardial cyst was oval shaped cystic mass located in left pericardiac border.