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Differential regulation of hepatic organic cation transporter 1, organic anion-transporting polypeptide 1a4, bile-salt export pump, and multidrug resistance-associated protein 2 transporter expression in lymphocyte-deficient mice associates with interleukin-6 production.

ABSTRACT: Cholestasis results from interrupted bile flow and is associated with immune-mediated liver diseases. It is unclear how inflammation contributes to cholestasis. The aim of this study was to determine whether T and B cells contribute to hepatic transporter expression under basal and inflammatory conditions. C57BL/6J wild-type mice or strains lacking T, B, or both T and B cells were exposed to lipopolysaccharide (LPS) or saline, and livers were collected 16 hours later. Branched DNA signal amplification was used to assess mRNA levels of organic anion-transporting polypeptides (Oatp) 1a1, 1a4, and 1b2; organic cation transporter (Oct) 1; canalicular bile-salt export pump (Bsep); multidrug resistance-associated proteins (Mrp) 2 and 3; and sodium-taurocholate cotransporting polypeptide (Ntcp). Real-time polymerase chain reaction analysis was used to correlate changes of transporter expression with interleukin-1b (IL-1b), IL-6, IL-17A, IL-17F, tumor necrosis factor-? (TNF-?), and interferon-? expression in the liver. LPS treatment inhibited Bsep and Oct1 mRNA expression, and this was abrogated with a loss of T cells, but not B cells. In addition, the absence of T cells increased Mrp2 mRNA expression, whereas B cell deficiency attenuated Oatp1a4 mRNA in LPS-treated mice. Oatp1a1, Oatp1b2, Ntcp, and Mrp3 were largely unaffected by T or B cell deficiency. Lymphocyte deficiency altered basal and inflammatory IL-6, but not TNF-? or IL-1b, mRNA expression. Taken together, these data implicate lymphocytes as regulators of basal and inflammatory hepatic transporter expression and suggest that IL-6 signaling may play a critical role.

SUBMITTER: Bodeman CE

PROVIDER: S-EPMC3781416 | biostudies-literature | 2013 Oct

REPOSITORIES: biostudies-literature

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Differential regulation of hepatic organic cation transporter 1, organic anion-transporting polypeptide 1a4, bile-salt export pump, and multidrug resistance-associated protein 2 transporter expression in lymphocyte-deficient mice associates with interleukin-6 production.

Bodeman Connor E CE Dzierlenga Anika L AL Tally Catherine M CM Mulligan Ryan M RM Lake April D AD Cherrington Nathan J NJ McKarns Susan C SC

The Journal of pharmacology and experimental therapeutics 20130808 1

Cholestasis results from interrupted bile flow and is associated with immune-mediated liver diseases. It is unclear how inflammation contributes to cholestasis. The aim of this study was to determine whether T and B cells contribute to hepatic transporter expression under basal and inflammatory conditions. C57BL/6J wild-type mice or strains lacking T, B, or both T and B cells were exposed to lipopolysaccharide (LPS) or saline, and livers were collected 16 hours later. Branched DNA signal amplifi ...[more]

PMID: 23929842

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Project description:Background & aimsOATP1B3/SLCO1B3 is a human liver-specific transporter for the clearance of endogenous compounds (eg, bile acid [BA]) and xenobiotics. The functional role of OATP1B3 in humans has not been characterized, as SLCO1B3 is poorly conserved among species without mouse orthologs.MethodsSlc10a1-knockout (Slc10a1-/-), Slc10a1hSLCO1B3 (endogenous mouse Slc10a1 promoter-driven human-SLCO1B3 expression in Slc10a1-/- mice), and human SLCO1B3 liver-specific transgenic (hSLCO1B3-LTG) mice were generated and challenged with 0.1% ursodeoxycholic-acid (UDCA), 1% cholic-acid (CA) diet, or bile duct ligation (BDL) for functional studies. Primary hepatocytes and hepatoma-PLC/RPF/5 cells were used for mechanistic studies.ResultsSerum BA levels in Slc10a1-/- mice were substantially increased with or without 0.1% UDCA feeding compared with wild-type (WT) mice. This increase was attenuated in Slc10a1hSLCO1B3-mice, indicating that OATP1B3 functions as a significant hepatic BA uptake transporter. In vitro assay using primary hepatocytes from WT, Slc10a1-/-, and Slc10a1hSLCO1B3-mice indicated that OATP1B3 has a similar capacity in taking up taurocholate/TCA as Ntcp. Furthermore, TCA-induced bile flow was significantly impaired in Slc10a1-/- mice but partially recovered in Slc10a1hSLC01B3-mice, indicating that OATP1B3 can partially compensate the NTCP function in vivo. Liver-specific overexpression of OATP1B3 markedly increased the level of hepatic conjugated BA and cholestatic liver injury in 1% CA-fed and BDL mice. Mechanistic studies revealed that conjugated BAs stimulated Ccl2 and Cxcl2 in hepatocytes to increase hepatic neutrophil infiltration and proinflammatory cytokine production (eg, IL-6), which activated STAT3 to repress OATP1B3 expression by binding to its promoter.ConclusionsHuman OATP1B3 is a significant BA uptake transporter and can partially compensate Ntcp for conjugated BA uptake in mice. Its downregulation in cholestasis is an adaptive protective response.

Project description:OBJECTIVE:Recent innovations in biology are boosting gene and cell therapy, but monitoring the response to these treatments is difficult. The purpose of this study was to find an MRI-reporter gene that can be used to monitor gene or cell therapy and that can be delivered without a viral vector, as viral vector delivery methods can result in long-term complications. MATERIALS AND METHODS:CMV promoter-human organic anion transporting polypeptide 1B3 (CMV-hOATP1B3) cDNA or CMV-blank DNA (control) was transfected into HEK293 cells using Lipofectamine. OATP1B3 expression was confirmed by western blotting and confocal microscopy. In vitro cell phantoms were made using transfected HEK293 cells cultured in various concentrations of gadoxetic acid for 24 hours, and images of the phantoms were made with a 9.4T micro-MRI. In vivo xenograft tumors were made by implanting HEK293 cells transfected with CMV-hOATP1B3 (n = 4) or CMV-blank (n = 4) in 8-week-old male nude mice, and MRI was performed before and after intravenous injection of gadoxetic acid (1.2 μL/g). RESULTS:Western blot and confocal microscopy after immunofluorescence staining revealed that only CMV-hOATP1B3-transfected HEK293 cells produced abundant OATP1B3, which localized at the cell membrane. OATP1B3 expression levels remained high through the 25th subculture cycle, but decreased substantially by the 50th subculture cycle. MRI of cell phantoms showed that only the CMV-hOATP1B3-transfected cells produced a significant contrast enhancement effect. In vivo MRI of xenograft tumors revealed that only CMV-hOATP1B3-transfected HEK293 tumors demonstrated a T1 contrast effect, which lasted for at least 5 hours. CONCLUSION:The human endogenous OATP1B3 gene can be non-virally delivered into cells to induce transient OATP1B3 expression, leading to gadoxetic acid-mediated enhancement on MRI. These results indicate that hOATP1B3 can serve as an MRI-reporter gene while minimizing the risk of long-term complications.

Dataset Information

Differential regulation of hepatic organic cation transporter 1, organic anion-transporting polypeptide 1a4, bile-salt export pump, and multidrug resistance-associated protein 2 transporter expression in lymphocyte-deficient mice associates with interleukin-6 production.

Publications

Differential regulation of hepatic organic cation transporter 1, organic anion-transporting polypeptide 1a4, bile-salt export pump, and multidrug resistance-associated protein 2 transporter expression in lymphocyte-deficient mice associates with interleukin-6 production.

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