Maturation of mammalian hepatocytes

Abstract

Directed differentiation and maturation of mammalian hepatocytes, such as human hepatocytes. The hepatocyte obtained show a phenotype which is more similar to that of primary hepatocytes than previously shown. In particular, exposure of mammalian hepatocytes, such as human hepatocytes, to at least one maturation factor selected from the group consisting of Src kinase inhibitors, vitamin D including precursors, metabolites and analogs thereof, hypoxia inducing compounds, sphingosine and sphingosine derivatives, activators of peroxisome proliferator-activated receptors (PPARs), platelet-activating factor (PAF), PKC inhibitors, and combinations thereof.

1 . A composition comprising: at least two maturation factors selected from the group consisting of Src kinase inhibitors, vitamin D including precursors, metabolites and analog thereof, hypoxia inducing compounds, sphingosine and sphingosine derivatives, activators of peroxisome proliferator-activated receptors (PPARs), platelet-activating factor (PAF), PKC inhibitors, and combinations thereof. 
     
     
         2 . The composition according to  claim 1 , wherein said composition comprises at least one Src kinase inhibitor and at least one further maturation factor selected from the group consisting of, vitamin D including precursors, metabolites and analog thereof, hypoxia inducing compounds, sphingosine and sphingosine derivatives, activators of peroxisome proliferator-activated receptors (PPARs), platelet-activating factor (PAF), and PKC inhibitors. 
     
     
         3 . The composition according to  claim 2 , wherein said composition comprises at least one Src kinase inhibitor selected from the group consisting of PP1, PP2, 1-NA PP1, 1-NM-PP1, Src Inhibitor-1 (Src-11), Src Kinase Inhibitor I, Src Kinase Inhibitor II, A-419529, A-770041, AZM 475271, bosutinib, CGP77675, Damnacanthal, dasatinib, dasatinib monohydrate, ER 27319 maleate, Fingolimod (FTY720), Geldanamycin, Herbimycin A, KB SRC 4, KX2-391, KX1-004, Lavendustin A, Lavendustin C, LCK inhibitor 2, Lyn peptide inhibitor, MLR-1023, MNS, N-Acetyl-O-phosphono-Tyr-Glu Dipentylamide, N-Acetyl-O-phosphono-Tyr-Glu-Glu-Ile-Glu, NVP-BHG712, PD 166285, PD173952, PD 180970, Piceatannol, pp60 c-src, quercetin, radicicol from  Diheterospora chlamydosporia  solid, saracatinib, SU 6656, TC-S 7003, TG 100572, WH-4-023, ZM 306416, and combinations thereof. 
     
     
         4 . The composition according to  claim 2 , wherein said at least one Src kinase inhibitor is PP1. 
     
     
         5 . The composition according to  claim 2 , comprises at least one vitamin D, vitamin D precursor, vitamin D metabolite or vitamin D analog. 
     
     
         6 . The composition according to  claim 5 , wherein said vitamin D is a vitamin D3, selected from the group consisting of cholecalciferol, calcifediol, calcitriol, and combinations thereof. 
     
     
         7 . The composition according to  claim 2 , wherein said composition comprises at least one hypoxia inducing compound selected from the group consisting of RAR-related orphan receptor alpha (ROR-alpha) ligand, selected from the group consisting of CGP52608, CGP52608 analogs, melatonin, melatonin analogs, cholesterol, cholesterol derivatives, and combinations thereof, CoCl 2 , and NaN 3 . 
     
     
         8 . The composition according to  claim 2 , wherein said composition comprises a sphingosine or sphingosine derivative. 
     
     
         9 . The composition according to  claim 8 , wherein said sphingosine is D-erythro-sphingosine. 
     
     
         10 . The composition according to  claim 2 , wherein said composition comprises at least one activator of peroxisome proliferator-activated receptors (PPARs), preferably selected from the group consisting of thiazolidinediones, free fatty acids (FFAs), eicosanoids including eicosanoid precursors and eicosanoid analog, and combinations thereof. 
     
     
         11 . The composition according to  claim 10 , wherein said free fatty acids (FFAs) are selected from the group consisting of dodecanoic acid, tridecanoic acid, tetradecanoic acid, pentadecanoic acid, hexadecanoic acid, heptadecanoic acid, eicosanoic acid, heneicosanoic acid, docosanoic acid, tricosanoic acid, tetracosanoic acid, pentacosanoic acid, hexacosanoic acid, and combinations thereof. 
     
     
         12 . The composition according to  claim 10 , wherein said eicosanoid, eicosanoid precursor or eicosanoid analog are selected from the group consisting of Diacylglycerol, Eicosapentaenoic acid, Dihomo-gamma-linolenic acid, Arachidonic acid, ETYA (5,8,11,14-eicosatetraynoic acid), members of the hydroxyeicosatetraenoic acid (HETE) family, including 5-HETE and 15-HETE, members of the hydroxyoctadecadieonic acid (HODE) family, including 9-HODE and 13-HODE, classic eicosanoids, and non-classic eicosanoids. 
     
     
         13 . The composition according to  claim 2 , wherein said composition comprises at least one platelet-activating factor (PAF). 
     
     
         14 . The composition according to  claim 2 , wherein said composition comprises at least one PKC inhibitor, preferably selected from the group consisting of Bisindolylmaleimide I, Bisindolylmaleimide II, Bisindolylmaleimide III, Bisindolylmaleimide V, Bisindolylmaleimide VI, Bisindolylmaleimide VII, Bisindolylmaleimide VIII, Bisindolylmaleimide X, HBDDE, Rottlerin, Palmitoyl-DL-carnitine, R-Stearoyl Carnitine Chloride, Piceatannol, H-9, H-8, 1-(5-lsoquinolinesulfonyl)-3-methylpiperazine, HA-100 dihydrochloride, HA-1004, HA-1077, 5-lodotubericidin, Ro-32-0432, Ro-31-7549, Enzastaurin (LY317615), Sotrastaurin, Dequalinium Chloride, Go 6976, Go 6983, Go 7874, Myricitrin, 4-Hydroxy-Tamoxifen, N-Desmethyltamoxifen HCl, Safingol, Phloretin, UCN-01, 7-Oxostaurosporine, K-252a, K-252b, K-252c, Melittin, Hispidin, Calphostin C, Ellagic acid, PKC Inhibitor Peptide 19-31, PKC Inhibitor Peptide 19-36, PKC epsilon Translocation Inhibitor II, EGF-R Fragment 651-658, PKC beta inhibitor (CAS 257879-35-9), PKC 20-28, PKCpII/EGFR Inhibitor (CAS 145915-60-2), PKC6 Pseudosubstrate Inhibitor, PKC6/5 Inhibitor, [Ala107]-MBP (104-118), [Ala113]-MBP (104-118), ZIP, C-1, Bryostatin 1, LY 333531 hydrochloride, CGP 53353, Chelerythrine Chloride, TCS 21311, CID 755673, Gossypol, ET-18-OCH3, 1-O-Hexadecyl-2-O-methyl-rac-glycerol, NPC-15437 dihydrochloride, NGIC-I, MDL-27,032, DAPH-7, 7-Aminoindole, 5-Amino-2-methylindole, rac-2-Methoxy-3-hexadecanamido-1-propylphosphocholine, Copperbis-3,5-diisopropylsalicylate,D,L-3,4-Dihydroxymandelic Acid, rac-3-Octadecanamido-2-Methoxypropan-1-ol Phosphocholine, KRIBB3, Ilmofosine, rac-2-Methoxy-3-hexadecanamido-1-propylphosphocholine, and combinations thereof. 
     
     
         15 . A culture medium comprising the composition according to  claim 2 . 
     
     
         16 . A kit comprising the composition according to  claim 2 , optionally further comprising at least one extracellular matrix (ECM) component or ECM component mixture. 
     
     
         17 . A kit comprising the culture medium according to  claim 14 , optionally further comprising at least one extracellular matrix (ECM) component or ECM component mixture.