title: Optimized Hepatocyte-Like Cells with Functional Drug Transporters Directly-Reprogrammed from Mouse Fibroblasts and their Potential in Drug Disposition and Toxicology.
abstract: OBJECTIVE
  To develop a suitable hepatocyte-like cell model that could be a substitute for primary hepatocytes with essential transporter expression and functions. Induced hepatocyte-like (iHep) cells directly reprogrammed from mice fibroblast cells were fully characterized.
  
  
  METHODS
  Naïve iHep cells were transfected with nuclear hepatocyte factor 4 alpha (Hnf4α) and treated with selected small molecules. Sandwich cultured configuration was applied. The mRNA and protein expression of transporters were determined by Real Time PCR and confocal. The functional transporters were estimated by drug biliary excretion measurement. The inhibition of bile acid efflux transporters by cholestatic drugs were assessed.
  
  
  RESULTS
  The expression and function of p-glycoprotein (P-gp), bile salt efflux pump (Bsep), multidrug resistance-associated protein 2 (Mrp2), Na+-dependent taurocholate cotransporting polypeptide (Ntcp), and organic anion transporter polypedtides (Oatps) in iHep cells were significantly improved after transfection of hepatocyte nuclear factor 4 alpha (Hnf4α) and treatment with selected inducers. In vitro intrinsic biliary clearances (CLb,int) of optimized iHep cells for rosuvastatin, methotrexate, d8-TCA (deuterium-labeled sodium taurocholate acid) and DPDPE ([D-Pen2,5] enkephalin hydrate) correlated well with that of sandwich-cultured primary mouse hepatocytes (SCMHs) (r2 = 0.984). Cholestatic drugs were evaluated and the results were compared well with primary mice hepatocytes.
  
  
  CONCLUSIONS
  The optimized iHep cells expressed functional drug transporters and were comparable to primary mice hepatocytes. This study suggested direct reprogramming could provide a potential alternative to primary hepatocytes for drug candidate hepatobiliary disposition and hepatotoxicity screening.
authors.forename: Zhi-Tao
authors.initials: ZT
authors.lastname: Wu
authors.forename: Dan
authors.initials: D
authors.lastname: Yao
authors.forename: Shu-Yi
authors.initials: SY
authors.lastname: Ji
authors.forename: Xuan
authors.initials: X
authors.lastname: Ni
authors.forename: Yi-Meng
authors.initials: YM
authors.lastname: Gao
authors.forename: Li-Jian
authors.initials: LJ
authors.lastname: Hui
authors.forename: Guo-Yu
authors.initials: GY
authors.lastname: Pan
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics / Q000378:metabolism; D000074020:ATP Binding Cassette Transporter, Subfamily B, Member 11; D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism; D000818:Animals; D000924:Anticholesteremic Agents / Q000032:analysis / Q000378:metabolism* / Q000633:toxicity; D001647:Bile Acids and Salts / Q000378:metabolism; D018929:Cell Culture Techniques; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D065150:Cellular Reprogramming; D020881:Enkephalin, D-Penicillamine (2,5)- / Q000032:analysis / Q000378:metabolism / Q000633:toxicity; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D051557:Hepatocyte Nuclear Factor 4 / Q000235:genetics / Q000378:metabolism*; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D008727:Methotrexate / Q000032:analysis / Q000378:metabolism / Q000633:toxicity; D051379:Mice; D008813:Mice, Inbred ICR; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics / Q000378:metabolism; D027361:Organic Anion Transporters / Q000235:genetics / Q000378:metabolism; D029363:Organic Anion Transporters, Sodium-Dependent / Q000235:genetics / Q000378:metabolism; D000068718:Rosuvastatin Calcium / Q000032:analysis / Q000378:metabolism / Q000633:toxicity; D054852:Small Molecule Libraries / Q000737:chemistry / Q000494:pharmacology; D027981:Symporters / Q000235:genetics / Q000378:metabolism
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000074020:ATP Binding Cassette Transporter, Subfamily B, Member 11; D018528:ATP-Binding Cassette Transporters; D000924:Anticholesteremic Agents; D001647:Bile Acids and Salts; D020881:Enkephalin, D-Penicillamine (2,5)-; D051557:Hepatocyte Nuclear Factor 4; D026901:Membrane Transport Proteins; D008727:Methotrexate; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D027361:Organic Anion Transporters; D029363:Organic Anion Transporters, Sodium-Dependent; D000068718:Rosuvastatin Calcium; D054852:Small Molecule Libraries; D027981:Symporters; C490549:Abcb11 protein, mouse; C496593:Hnf4a protein, mouse; C071908:sodium-bile acid cotransporter