Longitudinal in vivo bioimaging of hepatocyte transcription factor activity following cholestatic liver injury in mice.
Molecular mechanisms regulating liver repair following cholestatic injury remain largely unknown. We have combined a mouse model of acute cholestatic liver injury, partial bile duct ligation (pBDL), with a novel longitudinal bioimaging methodology to quantify transcription factor activity during hepatic injury and repair. We administered lentiviral transcription factor activated luciferase/eGFP reporter (TFAR) cassettes to neonatal mice enabling longitudinal TFAR profiling by continued bioimaging throughout the lives of the animals and following pBDL in adulthood. Neonatal intravascular injection of VSV-G pseudotyped lentivirus resulted in almost exclusive transduction of hepatocytes allowing analysis of hepatocyte-specific transcription factor activity. We recorded acute but transient responses with NF-κB and Smad2/3 TFAR whilst our Notch reporter was repressed over the 40 days of evaluation post-pBDL. The bipotent hepatic progenitor cell line, HepaRG, can be directed to differentiate into hepatocytes and biliary epithelia. We found that forced expression of the Notch inhibitor NUMB in HepaRG resulted in enhanced hepatocyte differentiation and proliferation whereas over-expressing the Notch agonist JAG1 resulted in biliary epithelial differentiation. In conclusion, our data demonstrates that hepatocytes rapidly upregulate NF-κB and Smad2/3 activity, whilst repressing Notch signalling. This transcriptional response to cholestatic liver injury likely promotes partial de-differentiation to allow pro-regenerative proliferation of hepatocytes.
Stem Cell Group, Cardiovascular &Cell Sciences Research Institute, St. George's University of London, Cranmer Terrace, London SW17 0RE, UK.|Wits/SAMRC Antiviral Gene Therapy Research Unit, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa.
Juliette M K M
JM
Delhove
Gene Transfer Technology Group, Institute for Women's Health, University College London, 86-96 Chenies Mews, London WC1E 6HX, UK.
Suzanne M K
SM
Buckley
Gene Transfer Technology Group, Institute for Women's Health, University College London, 86-96 Chenies Mews, London WC1E 6HX, UK.
Dany P
DP
Perocheau
Gene Transfer Technology Group, Institute for Women's Health, University College London, 86-96 Chenies Mews, London WC1E 6HX, UK.
Rajvinder
R
Karda
Wits/SAMRC Antiviral Gene Therapy Research Unit, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa.
Patrick
P
Arbuthnot
MRC Centre for Inflammation Research, The Queens Medical Research Institute, University of Edinburgh, Edinburgh EH16 4TJ, U.K.
Neil C
NC
Henderson
Gene Transfer Technology Group, Institute for Women's Health, University College London, 86-96 Chenies Mews, London WC1E 6HX, UK.|Wits/SAMRC Antiviral Gene Therapy Research Unit, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa.
Simon N
SN
Waddington
Stem Cell Group, Cardiovascular &Cell Sciences Research Institute, St. George's University of London, Cranmer Terrace, London SW17 0RE, UK.|School of Healthcare Sciences, Manchester Metropolitan University, Chester Street, Manchester M1 5GD, U.K.
Tristan R
TR
McKay
D016475:3T3 Cells; D000818:Animals; D002454:Cell Differentiation; D049109:Cell Proliferation; D002779:Cholestasis / Q000150:complications / Q000000981:diagnostic imaging / Q000378:metabolism*; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006367:HeLa Cells; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D000072100:Jagged-1 Protein / Q000235:genetics / Q000378:metabolism; D016086:Lentivirus / Q000235:genetics; D008107:Liver Diseases / Q000000981:diagnostic imaging / Q000209:etiology / Q000378:metabolism*; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D051880:Receptors, Notch / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D051899:Smad2 Protein / Q000235:genetics / Q000378:metabolism; D051900:Smad3 Protein / Q000235:genetics / Q000378:metabolism
C000606274:Jag1 protein, mouse; D000072100:Jagged-1 Protein; D008565:Membrane Proteins; D016328:NF-kappa B; D009419:Nerve Tissue Proteins; C106293:Numb protein, mouse; D051880:Receptors, Notch; D051899:Smad2 Protein; C497783:Smad2 protein, mouse; D051900:Smad3 Protein; C497791:Smad3 protein, mouse; D049452:Green Fluorescent Proteins; D008156:Luciferases