Activation of necroptosis in human and experimental cholestasis.
Cholestasis encompasses liver injury and inflammation. Necroptosis, a necrotic cell death pathway regulated by receptor-interacting protein (RIP) 3, may mediate cell death and inflammation in the liver. We aimed to investigate the role of necroptosis in mediating deleterious processes associated with cholestatic liver disease. Hallmarks of necroptosis were evaluated in liver biopsies of primary biliary cholangitis (PBC) patients and in wild-type and RIP3-deficient (RIP3-/-) mice subjected to common bile duct ligation (BDL). The functional link between RIP3, heme oxygenase-1 (HO-1) and antioxidant response was investigated in vivo after BDL and in vitro. We demonstrate increased RIP3 expression and mixed lineage kinase domain-like protein (MLKL) phosphorylation in liver samples of human PBC patients, coincident with thioflavin T labeling, suggesting activation of necroptosis. BDL resulted in evident hallmarks of necroptosis, concomitant with progressive bile duct hyperplasia, multifocal necrosis, fibrosis and inflammation. MLKL phosphorylation was increased and insoluble aggregates of RIP3, MLKL and RIP1 formed in BLD liver tissue samples. Furthermore, RIP3 deficiency blocked BDL-induced necroinflammation at 3 and 14 days post-BDL. Serum hepatic enzymes, fibrogenic liver gene expression and oxidative stress decreased in RIP3-/-mice at 3 days after BDL. However, at 14 days, cholestasis aggravated and fibrosis was not halted. RIP3 deficiency further associated with increased hepatic expression of HO-1 and accumulation of iron in BDL mice. The functional link between HO-1 activity and bile acid toxicity was established in RIP3-deficient primary hepatocytes. Necroptosis is triggered in PBC patients and mediates hepatic necroinflammation in BDL-induced acute cholestasis. Targeting necroptosis may represent a therapeutic strategy for acute cholestasis, although complementary approaches may be required to control progression of chronic cholestatic liver disease.
Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal.
Marta B
MB
Afonso
Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal.
Pedro M
PM
Rodrigues
Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal.
André L
AL
Simão
Department of Cell Biology, Harvard Medical School, Boston, MA, USA.
Dimitry
D
Ofengeim
Histology and Comparative Pathology Laboratory, Instituto de Medicina Molecular, Lisbon, Portugal.
Tânia
T
Carvalho
Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal.
Joana D
JD
Amaral
Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal.
Maria M
MM
Gaspar
Department of Gastrenterology, Hospital Santa Maria, Lisbon, Portugal.|Faculty of Medicine, Universidade de Lisboa, Lisbon, Portugal.
Helena
H
Cortez-Pinto
Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal.
Rui E
RE
Castro
Department of Cell Biology, Harvard Medical School, Boston, MA, USA.
Junying
J
Yuan
Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal.
Cecília M P
CM
Rodrigues
D000208:Acute Disease; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001647:Bile Acids and Salts / Q000633:toxicity; D001652:Bile Ducts / Q000187:drug effects / Q000473:pathology; D002779:Cholestasis / Q000150:complications / Q000473:pathology*; D002908:Chronic Disease; D004195:Disease Models, Animal; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D051547:Heme Oxygenase-1 / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007249:Inflammation / Q000150:complications / Q000473:pathology; D007501:Iron / Q000378:metabolism; D008026:Ligation; D008099:Liver / Q000473:pathology; D008103:Liver Cirrhosis / Q000150:complications / Q000473:pathology; D008810:Mice, Inbred C57BL; D009336:Necrosis; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000172:deficiency / Q000378:metabolism
D001647:Bile Acids and Salts; D007501:Iron; D051547:Heme Oxygenase-1; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; C485615:Ripk3 protein, mouse
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