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1.
Protein & Cell ; (12): 717-733, 2021.
Article in English | WPRIM | ID: wpr-888715

ABSTRACT

The coronavirus disease 2019 (COVID-19) pandemic is caused by infection with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is spread primary via respiratory droplets and infects the lungs. Currently widely used cell lines and animals are unable to accurately mimic human physiological conditions because of the abnormal status of cell lines (transformed or cancer cells) and species differences between animals and humans. Organoids are stem cell-derived self-organized three-dimensional culture in vitro and model the physiological conditions of natural organs. Here we showed that SARS-CoV-2 infected and extensively replicated in human embryonic stem cells (hESCs)-derived lung organoids, including airway and alveolar organoids which covered the complete infection and spread route for SARS-CoV-2 within lungs. The infected cells were ciliated, club, and alveolar type 2 (AT2) cells, which were sequentially located from the proximal to the distal airway and terminal alveoli, respectively. Additionally, RNA-seq revealed early cell response to virus infection including an unexpected downregulation of the metabolic processes, especially lipid metabolism, in addition to the well-known upregulation of immune response. Further, Remdesivir and a human neutralizing antibody potently inhibited SARS-CoV-2 replication in lung organoids. Therefore, human lung organoids can serve as a pathophysiological model to investigate the underlying mechanism of SARS-CoV-2 infection and to discover and test therapeutic drugs for COVID-19.


Subject(s)
Adenosine Monophosphate/therapeutic use , Alanine/therapeutic use , Alveolar Epithelial Cells/virology , Antibodies, Neutralizing/therapeutic use , COVID-19/virology , Down-Regulation , Drug Discovery , Human Embryonic Stem Cells/metabolism , Humans , Immunity , Lipid Metabolism , Lung/virology , RNA, Viral/metabolism , SARS-CoV-2/physiology , Virus Replication/drug effects
2.
Protein & Cell ; (12): 776-777, 2019.
Article in English | WPRIM | ID: wpr-757915

ABSTRACT

In the original publication the Supplementary Material and Fig. 2 are incorrect. The correct version is provided in this correction article. The text HBG2 appearing in the article should be read as HBG1.

3.
Protein & Cell ; (12): 69-79, 2014.
Article in English | WPRIM | ID: wpr-757523

ABSTRACT

With their capability to undergo unlimited self-renewal and to differentiate into all cell types in the body, human embryonic stem cells (hESCs) hold great promise in human cell therapy. However, there are limited tools for easily identifying and isolating live hESC-derived cells. To track hESC-derived neural progenitor cells (NPCs), we applied homologous recombination to knock-in the mCherry gene into the Nestin locus of hESCs. This facilitated the genetic labeling of Nestin positive neural progenitor cells with mCherry. Our reporter system enables the visualization of neural induction from hESCs both in vitro (embryoid bodies) and in vivo (teratomas). This system also permits the identification of different neural subpopulations based on the intensity of our fluorescent reporter. In this context, a high level of mCherry expression showed enrichment for neural progenitors, while lower mCherry corresponded with more committed neural states. Combination of mCherry high expression with cell surface antigen staining enabled further enrichment of hESC-derived NPCs. These mCherry(+) NPCs could be expanded in culture and their differentiation resulted in a down-regulation of mCherry consistent with the loss of Nestin expression. Therefore, we have developed a fluorescent reporter system that can be used to trace neural differentiation events of hESCs.


Subject(s)
Animals , Cell Differentiation , Cell Line , Embryonic Stem Cells , Cell Biology , Metabolism , Transplantation , Gene Knock-In Techniques , Genes, Reporter , Homologous Recombination , Humans , Luminescent Proteins , Genetics , Mice , Mice, SCID , Nestin , Genetics , Neural Stem Cells , Cell Biology , Metabolism , Neurons , Cell Biology , Metabolism , Teratoma , Pathology
4.
Article in Chinese | WPRIM | ID: wpr-407452

ABSTRACT

Sef (similar expression to fgf genes) was identified as a feedback antagonist of FGF signaling in zerbrafish, mouse and human. Sefhas been reported to function in different ways, however the regulation of Sef stability remains unknown. The possible role of c-Cbl in the regulation of Sef protein degradation was investigated. Results from coimmunoprecipitation and immunostaining assays reveal that hSef colocalizes and interacts with c-Cbl. Data suggest that the interaction between hSef and c-Cbl results in the ubiquitination and subsequent degradation of the hSef protein. It was proposed that c-Cbl may serve as a modulator to regulate Sef protein stability during FGF signal transduction.

5.
Chinese Journal of Biotechnology ; (12): 193-197, 2008.
Article in Chinese | WPRIM | ID: wpr-276141

ABSTRACT

Sef (similar expression to fgf genes) was identified as a feedback antagonist of FGF signaling in zerbrafish, mouse and human. To construct recombinant adenoviral vectors expressing hSef-L and hSef-S, the coding sequences of the two isoforms were amplified and ligated into pAdTrack-CMV, forming shuttle vectors pAdTrack-CMV/hSef-L-Myc and pAdTrack-CMV/hSef-S-Myc. After sequence confirmation, these two shuttle vector plasmids were linearized by Pme I and then co-transformed respectively with the adenoviral genome vector pAdEasy-1 into E. coli BJ5183. The successful recombinants were selected by Kanamycin and confirmed by Pac I digestion. The recombinant vectors Ad-hSef-L-Myc and Ad-hSef-S-Myc were finally digested with Pac I and transfected into HEK293 cells to pack into viral particles. The virus were amplified in 293 cells and used to infect MEF cells. Western blotting analysis was used to demonstrate the expression of hSef-L-Myc and hSef-S-Myc proteins. The inhibitory effects of the adenovirus mediated Sef expression on FGF signaling was further evaluated by Elk luciferase reporter assay. Our results indicated the constructed virus could produce effectively the proteins and then inhibit FGF signaling in MEF cells.


Subject(s)
Adenoviridae , Genetics , Metabolism , Cell Line , Cloning, Molecular , Defective Viruses , Genetics , Metabolism , Escherichia coli , Genetics , Metabolism , Genetic Vectors , Genetics , Humans , Protein Isoforms , Genetics , Receptors, Interleukin , Genetics , Recombinant Proteins , Genetics , Transfection , Virus Cultivation , Methods
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