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Direct conversion of human fibroblast to hepatocytes using a single inducible polycistronic vector.

ABSTRACT: BACKGROUND:Human fibroblasts can be reprogrammed into induced hepatocyte-like cells through the expression of a set of transcription factors. Although the generation of induced hepatocyte-like cells by HNF4A, HNF1A, and FOXA3 expression has proven to be a robust experimental strategy, using multiple lentivirus results in a highly variable heterogeneous population. METHODS:We designed and implemented a novel approach based on the delivery of reprogramming factors and green fluorescent protein in a single doxycycline-inducible lentiviral vector using 2A self-cleaving peptides. RESULTS:Fibroblasts infected with the lentiviral vector can be amplified in basic fibroblast culture media in the absence of doxycycline without induction of hepatic genes. Upon switching to hepatic maturation media containing doxycycline, cells stop proliferating, activate hepatic gene transcription, and perform metabolic functions characteristic of hepatocytes. CONCLUSION:Our strategy can generate an unlimited source of homogeneously induced hepatocyte-like cells from different genetic background donors, capable of performing typical hepatic functions suitable for drug research and other in vitro applications.

SUBMITTER: Ballester M

PROVIDER: S-EPMC6829923 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Efficient conversion of astrocytes to functional midbrain dopaminergic neurons using a single polycistronic vector.

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