Derivation of hepatic stem cells and mature liver cell types and uses thereof
AssigneesAgency for Science, Technology and Research, National University of Singapore
InventorsYuanyu HU, Huck Hui NG, Yock Young DAN
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Abstract
This application describes liver stem cells (LSC), and differentiated hepatocytes, cholangiocytes, and 3D cellular structures derived therefrom. Methods for producing LSC and mature, differentiated hepatocytes and cholangiocytes in culture are provided. Also provided are cell culture systems and cell culture media for producing a homogenous population of liver stem cells that remain in an undifferentiated state over multiple passages in culture. The LSC and methods are useful for producing homogenous populations of hepatocytes and cholangiocytes for downstream applications. The LSC can be transplanted into subjects to treat liver diseases.
Claims20
A liver stem cell comprising the following characteristics:
- (i) expresses at least 1, 2, 3 or 4 of the protein markers selected from SOX9, HNF4α, HNF3β, and EPCAM;
- (ii) expresses low levels of KRT19 protein,
- (iii) does not express detectable protein levels of KRT7 or AFP;
- and
- (iv) does not have a polarized cell phenotype.
A differentiated hepatocyte comprising at least one of the following characteristics:
- (i) expresses the markers HNF4α and ALB;
- (ii) expresses at least 1, 2, 3,4,5,6,7 or 8 of the CYP enzymes selected from CYP3A4, CYP2C9, CYP2B6, CYP1A2, CYP1A1, CYP2D6, CYP3A7, and CYP2E1;
- (iii) have at least 1, 2 or 3 of the following functions:
- a) functional glucose metabolism;
- b) functional lipid metabolism;
- c) functional albumin secretion;
- (vi) indocyanine green (ICG) uptake;
- and
- (vii) forms tight junctions when cultured with other differentiated hepatocytes under conditions sufficient to form an epithelium.
A differentiated cholangiocyte comprising at least one of the following characteristics:
- (i) expresses the markers KRT19 (Accession no:
- P08727.4) and KRT7 (Accession no:
- P08729.5) and ECAD;
- (ii) has columnar epithelial cell polarity comprising an apical region and a basolateral region when cultured with other differentiated cholangiocytes under conditions sufficient to form an epithelium;
- (iii) comprises microvilli on the apical region;
- (iii) forms biliary duct-like 3 dimensional (3D) structure when cultured with other differentiated cholangiocytes under conditions sufficient to form 3D structures.
-132. (canceled)
Dependencies are inferred from the claim text.