title: A transgenic rat hepatocyte - Kupffer cell co-culture model for evaluation of direct and macrophage-related effect of poly(amidoamine) dendrimers.
abstract: Increasing number of papers demonstrate that Kupffer cells (KCs) play a role in the development of drug induced liver injury (DILI). Furthermore, elevated intracellular Ca2+level of hepatocytes is considered as a common marker of DILI. Here we applied an in vitro model based on hepatocyte mono- and hepatocyte/KC co-cultures (H/KC) isolated from transgenic rats stably expressing the GCaMP2 fluorescent Ca2+sensor protein to investigate the effects of polycationic (G5), polyanionic (G4.5) and polyethylene-glycol coated neutral (G5 Peg) dendrimers known to accumulate in the liver, primarily in KCs. Following dendrimer exposure, hepatocyte homeostasis was measured by MTT cytotoxicity assay and by Ca2+imaging, while hepatocyte functions were studied by CYP2B1/2 inducibility, and bilirubin and taurocholate transport. G5 was significantly more cytotoxic than G4.5 for hepatocytes and induced Ca2+oscillation and sustained Ca2+signals at 1μM and10 μM, respectively both in hepatocytes and KCs. Dendrimer-induced Ca2+signals in hepatocytes were attenuated by macrophages. Activation of KCs by lipopolysaccharide and G5 decreased the inducibility of CYP2B1/2, which was restored by depleting the KCs with gadolinium-chloride and pentoxyphylline, suggesting a role of macrophages in the hindrance of CYP2B1/2 induction by G5 and lipopolysaccharide. In the H/KC, but not in the hepatocyte mono-culture, G5 reduced the canalicular efflux of bilirubin and stimulated the uptake and canalicular efflux of taurocholate. In conclusion, H/KC provides a good model for the prediction of hepatotoxic potential of drugs, especially of nanomaterials known to be trapped by macrophages, activation of which presumably contributes to DILI.
authors.affiliation: Institute of Organic Chemistry, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary. Electronic address: jemnitz.katalin@ttk.mta.hu.
authors.forename: Katalin
authors.initials: K
authors.lastname: Jemnitz
authors.affiliation: Institute of Organic Chemistry, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: Attila
authors.initials: A
authors.lastname: Bátai-Konczos
authors.affiliation: Institute of Organic Chemistry, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: Mónika
authors.initials: M
authors.lastname: Szabó
authors.affiliation: Institute of Organic Chemistry, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: Enikő
authors.initials: E
authors.lastname: Ioja
authors.affiliation: Institute of Enzymology, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: Orsolya
authors.initials: O
authors.lastname: Kolacsek
authors.affiliation: Institute of Enzymology, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: Tamás I
authors.initials: TI
authors.lastname: Orbán
authors.affiliation: Institute of Enzymology, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: György
authors.initials: G
authors.lastname: Török
authors.affiliation: Institute of Enzymology, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: László
authors.initials: L
authors.lastname: Homolya
authors.affiliation: BioTalentum Ltd, Gödöllő, Hungary.
authors.forename: Eszter
authors.initials: E
authors.lastname: Kovács
authors.affiliation: Institute of Organic Chemistry, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: István
authors.initials: I
authors.lastname: Jablonkai
authors.affiliation: Institute of Organic Chemistry, Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: Zsuzsa
authors.initials: Z
authors.lastname: Veres
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D018920:Coculture Techniques; D050091:Dendrimers / Q000633:toxicity*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D007728:Kupffer Cells / Q000187:drug effects* / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism; D008297:Male; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism; D055647:Rats, Transgenic; D017208:Rats, Wistar
chemical_list: D002147:Calmodulin; D050091:Dendrimers; C107660:M13 protein (myosin light-chain kinase); C104700:PAMAM Starburst; D010446:Peptide Fragments; D049452:Green Fluorescent Proteins; D009219:Myosin-Light-Chain Kinase; D002118:Calcium
keywords: Calcium imaging; Hepatocyte-Kupffer co-culture; Nanotoxicity; Poly(amidoamin) dendrimer; Transgenic rats