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Chronic liver injury drives non-traditional intrahepatic fibrin(ogen) crosslinking via tissue transglutaminase.

ABSTRACT: Essentials Fibrin clots are often implicated in the progression of liver fibrosis. Liver fibrosis was induced in transgenic mice with defects in clot formation or stabilization. Liver fibrosis and fibrin(ogen) deposition do not require fibrin polymerization or factor XIIIa. Fibrin(ogen) is an in vivo substrate of tissue transglutaminase in experimental liver fibrosis. SUMMARY: Background Intravascular fibrin clots and extravascular fibrin deposits are often implicated in the progression of liver fibrosis. However, evidence supporting a pathological role of fibrin in hepatic fibrosis is indirect and based largely on studies using anticoagulant drugs that inhibit activation of the coagulation protease thrombin, which has other downstream targets that promote fibrosis. Therefore, the goal of this study was to determine the precise role of fibrin deposits in experimental hepatic fibrosis. Methods Liver fibrosis was induced in mice expressing mutant fibrinogen insensitive to thrombin-mediated proteolysis (i.e. locked in the monomeric form), termed Fib^AEK mice, and factor XIII A₂ subunit-deficient (FXIII^-/- ) mice. Female wild-type mice, FXIII^-/- mice and homozygous Fib^AEK mice were challenged with carbon tetrachloride (CCl₄ ) twice weekly for 4 weeks or 6 weeks (1 mL kg^-1 , intraperitoneal). Results Hepatic injury and fibrosis induced by CCl₄ challenge were unaffected by FXIII deficiency or inhibition of thrombin-catalyzed fibrin polymer formation (in Fib^AEK mice). Surprisingly, hepatic deposition of crosslinked fibrin(ogen) was not reduced in CCl₄ -challenged FXIII^-/- mice or Fib^AEK mice as compared with wild-type mice. Rather, deposition of crosslinked hepatic fibrin(ogen) following CCl₄ challenge was dramatically reduced in tissue transglutaminase-2 (TGM2)-deficient (TGM2^-/- ) mice. However, the reduction in crosslinked fibrin(ogen) in TGM2^-/- mice did not affect CCl₄ -induced liver fibrosis. Conclusions These results indicate that neither traditional fibrin clots, formed by the thrombin-activated FXIII pathway nor atypical TGM2-crosslinked fibrin(ogen) contribute to experimental CCl₄ -induced liver fibrosis. Collectively, the results indicate that liver fibrosis occurs independently of intrahepatic fibrin(ogen) deposition.

SUBMITTER: Poole LG

PROVIDER: S-EPMC6322974 | biostudies-literature | 2019 Jan

REPOSITORIES: biostudies-literature

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Chronic liver injury drives non-traditional intrahepatic fibrin(ogen) crosslinking via tissue transglutaminase.

Poole L G LG Pant A A Baker K S KS Kopec A K AK Cline-Fedewa H M HM Iismaa S E SE Flick M J MJ Luyendyk J P JP

Journal of thrombosis and haemostasis : JTH 20181210 1

Essentials Fibrin clots are often implicated in the progression of liver fibrosis. Liver fibrosis was induced in transgenic mice with defects in clot formation or stabilization. Liver fibrosis and fibrin(ogen) deposition do not require fibrin polymerization or factor XIIIa. Fibrin(ogen) is an in vivo substrate of tissue transglutaminase in experimental liver fibrosis. SUMMARY: Background Intravascular fibrin clots and extravascular fibrin deposits are often implicated in the progression of liver ...[more]

PMID: 30415489

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Project description:Background & aimsAcetaminophen (APAP)-induced liver injury is coupled with activation of the blood coagulation cascade and fibrin(ogen) accumulation within APAP-injured livers of experimental mice. We sought to define the role of fibrin(ogen) deposition in APAP-induced liver injury and repair.MethodsWild-type, fibrinogen-deficient mice, mutant mice with fibrin(ogen) incapable of binding leukocyte αMβ2 integrin (Fibγ390-396A mice) and matrix metalloproteinase 12 (Mmp12)-deficient mice were fasted, injected with 300mg/kg APAP i.p. and evaluated at a range of time-points. Plasma and liver tissue were analyzed. Rescue of Fibγ390-396A mice was carried out with exogenous Mmp12. To examine the effect of the allosteric leukocyte integrin αMβ2 activator leukadherin-1 (LA-1), APAP-treated mice were injected with LA-1.ResultsIn wild-type mice, APAP overdose increased intrahepatic levels of high molecular weight cross-linked fibrin(ogen). Anticoagulation reduced early APAP hepatotoxicity (6h), but increased hepatic injury at 24h, implying a protective role for coagulation at the onset of repair. Complete fibrin(ogen) deficiency delayed liver repair after APAP overdose, evidenced by a reduction of proliferating hepatocytes (24h) and unresolved hepatocellular necrosis (48 and 72h). Fibγ390-396A mice had decreased hepatocyte proliferation and increased multiple indices of liver injury, suggesting a mechanism related to fibrin(ogen)-leukocyte interaction. Induction of Mmp12, was dramatically reduced in APAP-treated Fibγ390-396A mice. Mice lacking Mmp12 displayed exacerbated APAP-induced liver injury, resembling Fibγ390-396A mice. In contrast, administration of LA-1 enhanced hepatic Mmp12 mRNA and reduced necrosis in APAP-treated mice. Further, administration of recombinant Mmp12 protein to APAP-treated Fibγ390-396A mice restored hepatocyte proliferation.ConclusionsThese studies highlight a novel pathway of liver repair after APAP overdose, mediated by fibrin(ogen)-αMβ2 integrin engagement, and demonstrate a protective role of Mmp12 expression after APAP overdose.Lay summaryAcetaminophen overdose leads to activation of coagulation cascade and deposition of high molecular weight cross-linked fibrin(ogen) species in the liver. Fibrin(ogen) is required for stimulating liver repair after acetaminophen overdose. The mechanism whereby fibrin(ogen) drives liver repair after acetaminophen overdose requires engagement of leukocyte αMβ2 integrin and subsequent induction of matrix metalloproteinase 12.

Project description:Our recent study established the NMR structure of the recombinant bAalpha406-483 fragment corresponding to the NH(2)-terminal half of the bovine fibrinogen alphaC-domain and revealed that at increasing concentrations this fragment forms oligomers (self-associates). The major goals of the study presented here were to determine the structure and self-association of the full-length human fibrinogen alphaC-domains. To accomplish these goals, we prepared a recombinant human fragment, hAalpha425-503, homologous to bovine bAalpha406-483, and demonstrated using NMR, CD, and size-exclusion chromatography that its overall fold and ability to form oligomers are similar to those of bAalpha406-483. We also prepared recombinant hAalpha392-610 and bAalpha374-568 fragments corresponding to the full-length human and bovine alphaC-domains, respectively, and tested their structure, stability, and ability to self-associate. Size-exclusion chromatography revealed that both fragments form reversible oligomers in a concentration-dependent manner. Their oligomerization was confirmed in sedimentation equilibrium experiments, which also established the self-association affinities of these fragments and revealed that the addition of each monomer to assembling alphaC-oligomers substantially increases the stabilizing free energy. In agreement, unfolding experiments monitored by CD established that self-association of both fragments results in a significant increase in their thermal stability. Analysis of CD spectra of both fragments revealed that alphaC self-association results in an increase in the level of regular structure, implying that the COOH-terminal half of the alphaC-domain adopts an ordered conformation in alphaC-oligomers and that this domain contains two independently folded subdomains. Altogether, these data further clarify the structure of the human and bovine alphaC-domains and the molecular mechanism of their self-association into alphaC-polymers in fibrin.

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Chronic liver injury drives non-traditional intrahepatic fibrin(ogen) crosslinking via tissue transglutaminase.

Publications

Chronic liver injury drives non-traditional intrahepatic fibrin(ogen) crosslinking via tissue transglutaminase.

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