TY  - GEN
A1  - Michaud Schjeide, Brit-Maren
A1  - Schenke, Maren
A1  - Seeger, Bettina
A1  - Püschel, Gerhard Paul
T1  - Validation of a Novel Double Control Quantitative Copy Number PCR Method to Quantify Off-Target Transgene Integration after CRISPR-Induced DNA Modification
T2  - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe
N2  - In order to improve a recently established cell-based assay to assess the potency of botulinum neurotoxin, neuroblastoma-derived SiMa cells and induced pluripotent stem-cells (iPSC) were modified to incorporate the coding sequence of a reporter luciferase into a genetic safe harbor utilizing CRISPR/Cas9. A novel method, the double-control quantitative copy number PCR (dc-qcnPCR), was developed to detect off-target integrations of donor DNA. The donor DNA insertion success rate and targeted insertion success rate were analyzed in clones of each cell type. The dc-qcnPCR reliably quantified the copy number in both cell lines. The probability of incorrect donor DNA integration was significantly increased in SiMa cells in comparison to the iPSCs. This can possibly be explained by the lower bundled relative gene expression of a number of double-strand repair genes (BRCA1, DNA2, EXO1, MCPH1, MRE11, and RAD51) in SiMa clones than in iPSC clones. The dc-qcnPCR offers an efficient and cost-effective method to detect off-target CRISPR/Cas9-induced donor DNA integrations.
T3  - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 1269 
KW  - CRISPR editing validation
KW  - copy number analyses
KW  - homology-directed repair
KW  - homologous recombination deficiency
Y1  - 2022
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-561755
SN  - 1866-8372
SP  - 1
EP  - 14
PB  - Universitätsverlag Potsdam
CY  - Potsdam
ER  - 
TY  - JOUR
A1  - Knebel, Constanze
A1  - Neeb, Jannika
A1  - Zahn, Elisabeth
A1  - Schmidt, Flavia
A1  - Carazo, Alejandro
A1  - Holas, Ondej
A1  - Pavek, Petr
A1  - Püschel, Gerhard Paul
A1  - Zanger, Ulrich M.
A1  - Süssmuth, Roderich
A1  - Lampen, Alfonso
A1  - Marx-Stoelting, Philip
A1  - Braeuning, Albert
T1  - Unexpected Effects of Propiconazole, Tebuconazole, and Their Mixture on the Receptors CAR and PXR in Human Liver Cells
JF  - Toxicological sciences
N2  - Analyzing mixture toxicity requires an in-depth understanding of the mechanisms of action of its individual components. Substances with the same target organ, same toxic effect and same mode of action (MoA) are believed to cause additive effects, whereas substances with different MoAs are assumed to act independently. Here, we tested 2 triazole fungicides, propiconazole, and tebuconazole (Te), for individual and combined effects on liver toxicity-related endpoints. Both triazoles are proposed to belong to the same cumulative assessment group and are therefore thought to display similar and additive behavior. Our data show that Te is an antagonist of the constitutive androstane receptor (CAR) in rats and humans, while propiconazole is an agonist of this receptor. Both substances activate the pregnane X-receptor (PXR) and further induce mRNA expression of CYP3A4. CYP3A4 enzyme activity, however, is inhibited by propiconazole. For common targets of PXR and CAR, the activation of PXR by Te overrides CAR inhibition. In summary, propiconazole and Te affect different hepatotoxicity-relevant cellular targets and, depending on the individual endpoint analyzed, act via similar or dissimilar mechanisms. The use of molecular data based on research in human cell systems extends the picture to refine cumulative assessment group grouping and substantially contributes to the understanding of mixture effects of chemicals in biological systems.
KW  - triazole fungicides
KW  - constitutive androstane receptor
KW  - pregnane X-receptor
KW  - enzyme induction
KW  - liver toxicity
KW  - mixtures
Y1  - 2018
U6  - https://doi.org/10.1093/toxsci/kfy026
SN  - 1096-6080
SN  - 1096-0929
VL  - 163
IS  - 1
SP  - 170
EP  - 181
PB  - Oxford Univ. Press
CY  - Oxford
ER  - 
TY  - CHAP
A1  - Henkel, Janine
A1  - Camargo, Rodolfo Gonzalez
A1  - Schanze, Nancy
A1  - Püschel, Gerhard Paul
T1  - The vicious circle of prostaglandin- and cytokine-dependent hepatic insulin resistance: a key role of prostaglandin E2
T2  - Diabetologia : journal of the European Association for the Study of Diabetes (EASD)
Y1  - 2014
SN  - 0012-186X
SN  - 1432-0428
VL  - 57
SP  - S241
EP  - S242
PB  - Springer
CY  - New York
ER  - 
TY  - JOUR
A1  - Neuschaefer-Rube, Frank
A1  - Lieske, Stefanie
A1  - Kuna, Manuela
A1  - Henkel, Janin
A1  - Perry, Rachel J.
A1  - Erion, Derek M.
A1  - Pesta, Dominik
A1  - Willmes, Diana M.
A1  - Brachs, Sebastian
A1  - von Loeffelholz, Christian
A1  - Tolkachov, Alexander
A1  - Schupp, Michael
A1  - Pathe-Neuschaefer-Rube, Andrea
A1  - Pfeiffer, Andreas F. H.
A1  - Shulman, Gerald I.
A1  - Püschel, Gerhard Paul
A1  - Birkenfeld, Andreas L.
T1  - The mammalian INDY homolog is induced by CREB in a rat model of type 2 diabetes
JF  - Diabetes : a journal of the American Diabetes Association
Y1  - 2014
SN  - 0012-1797
SN  - 1939-327X
VL  - 63
IS  - 3
SP  - 1048
EP  - 1057
PB  - American Diabetes Association
CY  - Alexandria
ER  - 
TY  - JOUR
A1  - von Loeffelholz, Christian
A1  - Lieske, Stefanie
A1  - Neuschaefer-Rube, Frank
A1  - Willmes, Diana M.
A1  - Raschzok, Nathanael
A1  - Sauer, Igor M.
A1  - König, Jörg
A1  - Fromm, Martin F.
A1  - Horn, Paul
A1  - Chatzigeorgiou, Antonios
A1  - Pathe-Neuschaefer-Rube, Andrea
A1  - Jordan, Jens
A1  - Pfeiffer, Andreas F. H.
A1  - Mingrone, Geltrude
A1  - Bornstein, Stefan R.
A1  - Stroehle, Peter
A1  - Harms, Christoph
A1  - Wunderlich, F. Thomas
A1  - Helfand, Stephen L.
A1  - Bernier, Michel
A1  - de Cabo, Rafael
A1  - Shulman, Gerald I.
A1  - Chavakis, Triantafyllos
A1  - Püschel, Gerhard Paul
A1  - Birkenfeld, Andreas L.
T1  - The human longevity gene homolog INDY and interleukin-6 interact in hepatic lipid metabolism
BT  - official journal of the American Association for the Study of Liver Diseases
JF  - Hepatology
N2  - Reduced expression of the Indy ("I am Not Dead, Yet") gene in lower organisms promotes longevity in a manner akin to caloric restriction. Deletion of the mammalian homolog of Indy (mIndy, Slc13a5) encoding for a plasma membrane-associated citrate transporter expressed highly in the liver, protects mice from high-fat diet-induced and aging-induced obesity and hepatic fat accumulation through a mechanism resembling caloric restriction. We studied a possible role of mIndy in human hepatic fat metabolism. In obese, insulin-resistant patients with nonalcoholic fatty liver disease, hepatic mIndy expression was increased and mIndy expression was also independently associated with hepatic steatosis. In nonhuman primates, a 2-year high-fat, high-sucrose diet increased hepatic mIndy expression. Liver microarray analysis showed that high mIndy expression was associated with pathways involved in hepatic lipid metabolism and immunological processes. Interleukin-6 (IL-6) was identified as a regulator of mIndy by binding to its cognate receptor. Studies in human primary hepatocytes confirmed that IL-6 markedly induced mIndy transcription through the IL-6 receptor and activation of the transcription factor signal transducer and activator of transcription 3, and a putative start site of the human mIndy promoter was determined. Activation of the IL-6-signal transducer and activator of transcription 3 pathway stimulated mIndy expression, enhanced cytoplasmic citrate influx, and augmented hepatic lipogenesis in vivo. In contrast, deletion of mIndy completely prevented the stimulating effect of IL-6 on citrate uptake and reduced hepatic lipogenesis. These data show that mIndy is increased in liver of obese humans and nonhuman primates with NALFD. Moreover, our data identify mIndy as a target gene of IL-6 and determine novel functions of IL-6 through mINDY. Conclusion: Targeting human mINDY may have therapeutic potential in obese patients with nonalcoholic fatty liver disease. German Clinical Trials Register: DRKS00005450.
Y1  - 2017
U6  - https://doi.org/10.1002/hep.29089
SN  - 0270-9139
SN  - 1527-3350
VL  - 66
IS  - 2
SP  - 616
EP  - 630
PB  - Wiley
CY  - Hoboken
ER  - 
TY  - JOUR
A1  - Wieneke, Nadine
A1  - Neuschaefer-Rube, Frank
A1  - Bode, L. M.
A1  - Kuna, Manuela
A1  - Andres, Jesus
A1  - Carnevali Junior, Luiz Carlos
A1  - Hirsch-Ernst, Karen I.
A1  - Püschel, Gerhard Paul
T1  - Synergistic acceleration of thyroid hormone degradation by phenobarbital and the PPAR alpha agonist WY14643 in rat hepatocytes
N2  - Energy balance is maintained by controlling both energy intake and energy expenditure. Thyroid hormones play a crucial role in regulating energy expenditure. Their levels are adjusted by a tight feed back-control led regulation of thyroid hormone production/incretion and by their hepatic metabolism. Thyroid hormone degradation has previously been shown to be enhanced by treatment with phenobarbital or other antiepileptic drugs due to a CAR-dependent induction of phase 11 enzymes of xenobiotic metabolism. We have recently shown, that PPAR alpha agonists synergize with phenobarbital to induce another prototypical CAR target gene, CYP2B1. Therefore, it was tested whether a PPAR alpha agonist could enhance the phenobarbital-dependent acceleration of thyroid hormone elimination. In primary cultures of rat hepatocytes the apparent half-life of T3 was reduced after induction with a combination of phenobarbital and the PPARa agonist WY14643 to a larger extent than after induction with either Compound alone. The synergistic reduction of the half-life could be attributed to a synergistic induction of CAR and the CAR target genes that code for enzymes and transporters involved in the hepatic elimination of T3, such as OATP1A1, OATP1A3, UGT1A3 and UCT1A10. The PPAR alpha-dependent CAR induction and the subsequent induction of T3-eliminating enzymes might be of physiological significance for the fasting- incluced reduction in energy expenditure by fatty acids as natural PPARa ligands. The synergism of the PPAR alpha agonist WY14643 and phenobarbital in inducing thyroid hormone breakdown might serve as a paradigm for the synergistic disruption of endocrine control by other combinations of xenobiotics.
Y1  - 2009
UR  - http://www.sciencedirect.com/science/journal/0041008X
U6  - https://doi.org/10.1016/j.taap.2009.07.014
SN  - 0041-008X
ER  - 
TY  - JOUR
A1  - Neuschäfer-Rube, Frank
A1  - Möller, Ulrike
A1  - Püschel, Gerhard Paul
T1  - Structure of the 5'-flanking region of the rat prostaglandin f(2alpha) receptor
N2  - Prostaglandin F(2alpha) (PGF(2alpha)), modulates hepatocyte functions via a heptahelical G(q)-coupled PGF(2alpha)-receptor (FP-R) which in liver is expressed exclusively in hepatocytes. The aim of the present study was to isolate the 5'-flanking region of the rat FP-R gene and to elucidate its basal and IL-6-modulated transcription control function in rat hepatocytes. The 5'-non-translated region of the rat hepatocyte FP-R mRNA differed from the corresponding region in rat fetal astrocyte or corpus luteum. It was encoded by exons 1a and 2 which were separated by a 1. 4 kb intron containing the exons 1b and 1c coding for the 5'-untranslated region of rat fetal astrocyte and corpus luteum FP-R mRNA, respectively. The transcription initiation site in hepatocytes was localized 263 bp upstream of the start ATG by 5'-RACE. A DNA-fragment covering the 5'-flanking region of the rFP-R gene from - 1 of the transcription initiation site to -2590 bp was cloned and sequenced. Its 3'-two thirds had a 65% sequence identity to the mouse FP-R promoter however no homology to the bovine FP-R promoter. In the overlapping sequence most of the putative transcription factor binding sites were conserved between mouse and rat. The rat promoter contained no classical TATA- or CAAT-boxes but putative binding sites for the transcription factors C/EBP, GATA-1, HNF-1, HNF-3beta, SP-1, and USF. Luciferase reporter gene constructs containing portions of the 5'-flanking region were transfected into rat hepatocytes. Luciferase expression ranked -181 >/= -608 < -1418 > -1821 >/= -2590. The strongest transcriptional activity was conferred by the region between -608 and -1418 containing a cluster of potential HNF-1 and HNF-3beta binding sites that might allow the exclusive expression of FP-R mRNA in hepatocytes. The amount of FP-R mRNA and the luciferase expression under control of the -2590 promoter fragment were reduced by IL-6 in hepatocytes. Copyright 2000 Academic Press.
Y1  - 2000
ER  - 
TY  - JOUR
A1  - Henkel, Janin
A1  - Frede, Katja
A1  - Schanze, Nancy
A1  - Vogel, Heike
A1  - Schürmann, Annette
A1  - Spruß, Astrid
A1  - Bergheim, Ina
A1  - Püschel, Gerhard Paul
T1  - Stimulation of fat accumulation in hepatocytes by PGE(2)-dependent repression of hepatic lipolysis, beta-oxidation and VLDL-synthesis
JF  - Laboratory investigation : the basic and translational pathology research journal ; an official journal of the United States and Canadian Academy of Pathology
N2  - Hepatic steatosis is recognized as hepatic presentation of the metabolic syndrome. Hyperinsulinaemia, which shifts fatty acid oxidation to de novo lipogenesis and lipid storage in the liver, appears to be a principal elicitor particularly in the early stages of disease development. The impact of PGE(2), which has previously been shown to attenuate insulin signaling and hence might reduce insulin-dependent lipid accumulation, on insulin-induced steatosis of hepatocytes was studied. The PGE(2)-generating capacity was enhanced in various obese mouse models by the induction of cyclooxygenase 2 and microsomal prostaglandin E-synthases (mPGES1, mPGES2). PGE(2) attenuated the insulin-dependent induction of SREBP-1c and its target genes glucokinase and fatty acid synthase. Nevertheless, PGE(2) enhanced incorporation of glucose into hepatic triglycerides synergistically with insulin. This was most likely due to a combination of a PGE(2)-dependent repression of (1) the key lipolytic enzyme adipose triglyceride lipase, (2) carnitine-palmitoyltransferase 1, a key regulator of mitochondrial beta-oxidation, and (3) microsomal transfer protein, as well as (4) apolipoprotein B, key components of the VLDL synthesis. Repression of PGC1 alpha, a common upstream regulator of these genes, was identified as a possible cause. In support of this hypothesis, overexpression of PGC1 alpha completely blunted the PGE(2)-dependent fat accumulation. PGE(2) enhanced lipid accumulation synergistically with insulin, despite attenuating insulin signaling and might thus contribute to the development of hepatic steatosis. Induction of enzymes involved in PGE(2) synthesis in in vivo models of obesity imply a potential role of prostanoids in the development of NAFLD and NASH. Laboratory Investigation (2012) 92, 1597-1606; doi:10.1038/labinvest.2012.128; published online 10 September 2012
KW  - cyclooxygenase
KW  - hepatic steatosis
KW  - mPGES
KW  - NAFLD
KW  - NASH
KW  - type 2 diabetes (T2DM)
KW  - PGC1 alpha
Y1  - 2012
U6  - https://doi.org/10.1038/labinvest.2012.128
SN  - 0023-6837
VL  - 92
IS  - 11
SP  - 1597
EP  - 1606
PB  - Nature Publ. Group
CY  - New York
ER  - 
TY  - JOUR
A1  - Henkel, Janin
A1  - Alfine, Eugenia
A1  - Saín, Juliana
A1  - Jöhrens, Korinna
A1  - Weber, Daniela
A1  - Castro, José Pedro
A1  - König, Jeannette
A1  - Stuhlmann, Christin
A1  - Vahrenbrink, Madita
A1  - Jonas, Wenke
A1  - Kleinridders, André
A1  - Püschel, Gerhard Paul
T1  - Soybean Oil-Derived Poly-Unsaturated Fatty Acids Enhance Liver Damage in NAFLD Induced by Dietary Cholesterol
JF  - Nutrients
N2  - While the impact of dietary cholesterol on the progression of atherosclerosis has probably been overestimated, increasing evidence suggests that dietary cholesterol might favor the transition from blunt steatosis to non-alcoholic steatohepatitis (NASH), especially in combination with high fat diets. It is poorly understood how cholesterol alone or in combination with other dietary lipid components contributes to the development of lipotoxicity. The current study demonstrated that liver damage caused by dietary cholesterol in mice was strongly enhanced by a high fat diet containing soybean oil-derived ω6-poly-unsaturated fatty acids (ω6-PUFA), but not by a lard-based high fat diet containing mainly saturated fatty acids. In contrast to the lard-based diet the soybean oil-based diet augmented cholesterol accumulation in hepatocytes, presumably by impairing cholesterol-eliminating pathways. The soybean oil-based diet enhanced cholesterol-induced mitochondrial damage and amplified the ensuing oxidative stress, probably by peroxidation of poly-unsaturated fatty acids. This resulted in hepatocyte death, recruitment of inflammatory cells, and fibrosis, and caused a transition from steatosis to NASH, doubling the NASH activity score. Thus, the recommendation to reduce cholesterol intake, in particular in diets rich in ω6-PUFA, although not necessary to reduce the risk of atherosclerosis, might be sensible for patients suffering from non-alcoholic fatty liver disease.
KW  - non-alcoholic fatty liver disease (NAFLD)
KW  - NASH
KW  - cholesterol
KW  - PUFA
KW  - inflammation
KW  - oxidative stress
Y1  - 2018
U6  - https://doi.org/10.3390/nu10091326
SN  - 2072-6643
VL  - 10
IS  - 9
SP  - 1
EP  - 17
PB  - Molecular Diversity Preservation International (MDPI)
CY  - Basel
ER  - 
TY  - GEN
A1  - Henkel, Janin
A1  - Alfine, Eugenia
A1  - Saín, Juliana
A1  - Jöhrens, Korinna
A1  - Weber, Daniela
A1  - Castro, José Pedro
A1  - König, Jeannette
A1  - Stuhlmann, Christin
A1  - Vahrenbrink, Madita
A1  - Jonas, Wenke
A1  - Kleinridders, André
A1  - Püschel, Gerhard Paul
T1  - Soybean Oil-Derived Poly-Unsaturated Fatty Acids Enhance Liver Damage in NAFLD Induced by Dietary Cholesterol
T2  - Nutrients
N2  - While the impact of dietary cholesterol on the progression of atherosclerosis has probably been overestimated, increasing evidence suggests that dietary cholesterol might favor the transition from blunt steatosis to non-alcoholic steatohepatitis (NASH), especially in combination with high fat diets. It is poorly understood how cholesterol alone or in combination with other dietary lipid components contributes to the development of lipotoxicity. The current study demonstrated that liver damage caused by dietary cholesterol in mice was strongly enhanced by a high fat diet containing soybean oil-derived ω6-poly-unsaturated fatty acids (ω6-PUFA), but not by a lard-based high fat diet containing mainly saturated fatty acids. In contrast to the lard-based diet the soybean oil-based diet augmented cholesterol accumulation in hepatocytes, presumably by impairing cholesterol-eliminating pathways. The soybean oil-based diet enhanced cholesterol-induced mitochondrial damage and amplified the ensuing oxidative stress, probably by peroxidation of poly-unsaturated fatty acids. This resulted in hepatocyte death, recruitment of inflammatory cells, and fibrosis, and caused a transition from steatosis to NASH, doubling the NASH activity score. Thus, the recommendation to reduce cholesterol intake, in particular in diets rich in ω6-PUFA, although not necessary to reduce the risk of atherosclerosis, might be sensible for patients suffering from non-alcoholic fatty liver disease.
T3  - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 479 
KW  - non-alcoholic fatty liver disease (NAFLD)
KW  - NASH
KW  - cholesterol
KW  - PUFA
KW  - inflammation
KW  - oxidative stress
Y1  - 2018
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-419773
ER  -