Perturbations in Ataxia Telangiectasia Mutant Signaling Pathways After Drug-Induced Acute Liver Failure and Their Reversal During Rescue of Animals by Cell Therapy

Abstract

Superior insights into molecular mechanisms of liver failure, which are not fully understood, will help strategies for inducing liver regeneration. We examined hepatotoxic mechanisms in mice homozygous for the severe combined immune deficiency mutation in the protein kinase, DNA-activated, catalytic polypeptide. Mice were treated with rifampicin, phenytoin, and monocrotaline. The ensuing acute liver failure was characterized by serological, histological, and mRNA studies. Subsequently, we studied whether transplantation of hepatocytes could rescue animals with liver failure. We found extensive liver damage in these animals, with mortality over several days. The expression of multiple hepatic genes was rapidly altered, including those representing pathways in oxidative/metabolic stress, inflammation, DNA damage-repair, and ataxia telangiectasia mutant (Atm) signaling pathways. This led to liver cell growth arrest involving cyclin-dependent kinase inhibitor 1A. Transplantation of hepatocytes with microcarriers in the peritoneal cavity efficiently rescued animals with liver failure. Molecular abnormalities rapidly reversed, including in hepatic Atm and downstream signaling pathways; and residual hepatocytes overcame cyclin-dependent kinase inhibitor 1A-induced cell growth arrest. Reseeding of the liver with transplanted hepatocytes was not required for rescue because native hepatocytes overcame cell growth-arrest to regenerate the liver. This likely resulted from paracrine signaling from hepatocytes in the peritoneal cavity. We concluded that Atm signaling Drug-induced liver injury (DILI) often results in acute liver failure (ALF). 1 Acetaminophen (APAP) is the leading offender in causing ALF; however, despite extensive work, mechanisms of DILI by APAP are incompletely understood. [2][3][4][5] The identification of molecular pathways initiating or amplifying DILI will be highly significant for drug development and for preventing and/or treating ALF. After exposure to hepatotoxic drugs, perturbations in cell stress and toxicity pathways assume prominence. However, more information is required regarding intracellular signaling pathways that impart susceptibility to DILI. Similarly, more information is required regarding failure of liver regeneration in ALF. Nevertheless, establishing these mechanisms has been difficult in the available animal models of ALF. For instance, after exposure to toxic drugs or chemicals, some animals may exhibit rapid and irreversible mortality (eg, within 24 -30 hours after APAP), whereas other animals may recover spontaneously.

Spontaneous origin from human embryonic stem cells of liver cells displaying conjoint meso-endodermal phenotype with hepatic functions

“…In this system, transplanted hepatocytes adhering to microcarriers are revascularized in the peritoneal cavity, along with secretion of proteins in blood (Demetriou et al, 1986;Gupta et al, 1994;Bandi et al, 2011). Moreover, cells transplanted into the peritoneal cavity do not migrate to other organs, including the liver (Bandi et al, 2011). Furthermore, in this ALF model, reseeding of the damaged liver with healthy hepatocytes was not required for liver regeneration (Bandi et al 2011).…”

“…Previously, mice with ALF were rescued after transplantation in the peritoneal cavity of xenogeneic mature hepatocytes anchored to extracellularmatrix-coated microcarriers. In this system, transplanted hepatocytes adhering to microcarriers are revascularized in the peritoneal cavity, along with secretion of proteins in blood (Demetriou et al, 1986;Gupta et al, 1994;Bandi et al, 2011). Moreover, cells transplanted into the peritoneal cavity do not migrate to other organs, including the liver (Bandi et al, 2011).…”

“…We considered that the presence of crucial liver proteins glycogen, G-6-P and Cyp450 in hESC-MECs should permit these cells to rescue animals in ALF. We addressed this possibility in xenotolerant natural-onset diabetes-severe combined immunodeficiency (NOD/SCID) mice with ALF induced by a regimen of the hepatotoxic drugs rifampicin (Rif) and phenytoin (Phen), over 3 days, followed on day 4 by the pyrrolizidine alkaloid, monocrotaline (MCT) (Bandi et al, 2011). This produced 50-70% liver necrosis, abnormal liver tests, coagulopathy, encephalopathy and 90-100% mortality, over several days (Fig.…”

“…Here, we report our findings to indicate similarities in the conjoint mesoendodermal phenotype of hESC-derived cells and cultured fetal human liver stem cells. Despite the relative immaturity of these hESC-derived cells, as indicated by the extent of their hepatic functions in vitro, we found in intact mice with toxic-druginduced acute liver failure (ALF), that intraperitoneal transplantation of hESC-derived cells rescued various functions in vivo (Bandi et al, 2011). In this situation, hESC-derived cells provided liver support and also aided liver regeneration through secretion of multiple paracrine factors, without the requirement of reseeding the damaged liver with healthy hepatocytes.…”

Ischemic Preconditioning Affects Long-Term Cell Fate through DNA Damage–Related Molecular Signaling and Altered Proliferation

“…ajp.amjpathol.org -The American Journal of Pathology example, Western blotting confirmed greater expression in IP donor hepatocytes of JUN, phosphorylated MAPK8 (JNK), phosphorylated ERK-1/2, and phosphorylated p38 MAPK, which were observed at the mRNA level by RT-qPCR and were implicated by IPA in cell growth and proliferation. Because cell-cycle checkpoint controls are often activated by DNA damage, which is a consequence of IR 12 and is regulated by ataxia telangiectasia mutated (Atm) pathways, 23 we examined this possibility. Immunostaining of IP donor liver showed presence of 8-oxo-2 0 -deoxyguanosine (8-oxo-dG) DNA adducts, which is highly characteristic of oxidative DNA damage ( Figure 6B).…”