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Combined Activities of JNK1 and JNK2 in Hepatocytes Protect Against Toxic Liver Injury.

ABSTRACT: c-Jun N-terminal kinase (JNK) 1 and JNK2 are expressed in hepatocytes and have overlapping and distinct functions. JNK proteins are activated via phosphorylation in response to acetaminophen- or carbon tetrachloride (CCl4)-induced liver damage; the level of activation correlates with the degree of injury. SP600125, a JNK inhibitor, has been reported to block acetaminophen-induced liver injury. We investigated the role of JNK in drug-induced liver injury (DILI) in liver tissue from patients and in mice with genetic deletion of JNK in hepatocytes.We studied liver sections from patients with DILI (due to acetaminophen, phenprocoumon, nonsteroidal anti-inflammatory drugs, or autoimmune hepatitis) or patients without acute liver failure (controls) collected from a DILI Biobank in Germany. Levels of total and activated (phosphorylated) JNK were measured by immunohistochemistry and Western blotting. Mice with hepatocyte-specific deletion of Jnk1 (Jnk1(?hepa)) or combination of Jnk1 and Jnk2 (Jnk(?hepa)), as well as Jnk1-floxed C57BL/6 (control) mice, were given injections of CCl4 (to induce fibrosis) or acetaminophen (to induce toxic liver injury). We performed gene expression microarray and phosphoproteomic analyses to determine mechanisms of JNK activity in hepatocytes.Liver samples from DILI patients contained more activated JNK, predominantly in nuclei of hepatocytes and in immune cells, than healthy tissue. Administration of acetaminophen to Jnk(?hepa) mice produced a greater level of liver injury than that observed in Jnk1(?hepa) or control mice, based on levels of serum markers and microscopic and histologic analysis of liver tissues. Administration of CCl4 also induced stronger hepatic injury in Jnk(?hepa) mice, based on increased inflammation, cell proliferation, and fibrosis progression, compared with Jnk1(?hepa) or control mice. Hepatocytes from Jnk(?hepa) mice given acetaminophen had an increased oxidative stress response, leading to decreased activation of adenosine monophosphate-activated protein kinase, total protein adenosine monophosphate-activated protein kinase levels, and pJunD and subsequent necrosis. Administration of SP600125 before or with acetaminophen protected Jnk(?hepa) and control mice from liver injury.In hepatocytes, JNK1 and JNK2 appear to have combined effects in protecting mice from CCl4- and acetaminophen-induced liver injury. It is important to study the tissue-specific functions of both proteins, rather than just JNK1, in the onset of toxic liver injury. JNK inhibition with SP600125 shows off-target effects.

SUBMITTER: Cubero FJ

PROVIDER: S-EPMC5285516 | biostudies-literature | 2016 Apr

REPOSITORIES: biostudies-literature

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Combined Activities of JNK1 and JNK2 in Hepatocytes Protect Against Toxic Liver Injury.

Cubero Francisco Javier FJ Zoubek Miguel Eugenio ME Hu Wei W Peng Jin J Zhao Gang G Nevzorova Yulia A YA Al Masaoudi Malika M Bechmann Lars P LP Boekschoten Mark V MV Muller Michael M Preisinger Christian C Gassler Nikolaus N Canbay Ali E AE Luedde Tom T Davis Roger J RJ Liedtke Christian C Trautwein Christian C

Gastroenterology 20151219 4

<h4>Background & aims</h4>c-Jun N-terminal kinase (JNK) 1 and JNK2 are expressed in hepatocytes and have overlapping and distinct functions. JNK proteins are activated via phosphorylation in response to acetaminophen- or carbon tetrachloride (CCl4)-induced liver damage; the level of activation correlates with the degree of injury. SP600125, a JNK inhibitor, has been reported to block acetaminophen-induced liver injury. We investigated the role of JNK in drug-induced liver injury (DILI) in liver ...[more]

PMID: 26708719

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Project description:BACKGROUND & AIMS: c-Jun N-terminal kinase (JNK)1 and JNK2 are expressed in hepatocytes and have overlapping and distinct functions. JNK proteins are activated, via phosphorylation, in response to acetaminophen- or CCl4-induced liver damage; the level of activation correlates with the degree of injury. SP600125, a JNK inhibitor, has been reported to block acetaminophen-induced liver injury. We investigated the role of JNK in drug-induced liver injury (DILI) in liver tissues from patients and in mice with genetic deletion of JNK in hepatocytes. METHODS: We studied liver sections from patients with DILI (due to acetaminophen, phenprocoumon, non-steroidal anti-inflammatory drugs or autoimmune hepatitis), or patients without acute liver failure (controls), collected from a DILI Biobank in Germany. Levels of total and activated (phosphorylated) JNK were measured by immunohistochemistry and western blotting. Mice with hepatocyte-specific deletion of Jnk1 (Jnk1Îhepa) or combination of Jnk1 and Jnk2 (JnkÎhepa), as well as Jnk1-floxed C57BL/6 (control) mice, were given injections of CCl4 (to induce fibrosis) or acetaminophen (to induce toxic liver injury). We performed gene expression microarray, and phosphoproteomic analyses to determine mechanisms of JNK activity in hepatocytes. RESULTS: Liver samples from DILI patients contained more activated JNK, predominantly in nuclei of hepatocytes and in immune cells, than healthy tissue. Administration of acetaminophen to JnkÎhepa mice produced a greater level of liver injury than that observed in Jnk1Îhepa or control mice, based on levels of serum markers and microscopic and histologic analysis of liver tissues. Administration of CCl4 also induced stronger hepatic injury in JnkÎhepa mice, based on increased inflammation, cell proliferation, and fibrosis progression, compared to Jnk1Îhepa or control mice. Hepatocytes from JnkÎhepa mice given acetaminophen had an increased oxidative stress response, leading to decreased activation of AMPK, total protein AMPK levels, and pJunD and subsequent necrosis. Administration of SP600125 before or with acetaminophen protected JnkÎhepa and control mice from liver injury. CONCLUSIONS: In hepatocytes, JNK1 and JNK2 appear to have combined effects in protecting mice from CCl4- and acetaminophen-induced liver injury. It is important to study the tissue-specific functions of both proteins, rather than just JNK1, in the onset of toxic liver injury. JNK inhibition with SP600125 shows off-target effects. Livers and primary hepatocytes were isolated from wild type and JNKÎhepa (Jnk1Îhepa/global Jnk2-/-) double-knockout mice and subjected to gene expression profiling.

Project description:BACKGROUND & AIMS: c-Jun N-terminal kinase (JNK)1 and JNK2 are expressed in hepatocytes and have overlapping and distinct functions. JNK proteins are activated, via phosphorylation, in response to acetaminophen- or CCl4-induced liver damage; the level of activation correlates with the degree of injury. SP600125, a JNK inhibitor, has been reported to block acetaminophen-induced liver injury. We investigated the role of JNK in drug-induced liver injury (DILI) in liver tissues from patients and in mice with genetic deletion of JNK in hepatocytes. METHODS: We studied liver sections from patients with DILI (due to acetaminophen, phenprocoumon, non-steroidal anti-inflammatory drugs or autoimmune hepatitis), or patients without acute liver failure (controls), collected from a DILI Biobank in Germany. Levels of total and activated (phosphorylated) JNK were measured by immunohistochemistry and western blotting. Mice with hepatocyte-specific deletion of Jnk1 (Jnk1Δhepa) or combination of Jnk1 and Jnk2 (JnkΔhepa), as well as Jnk1-floxed C57BL/6 (control) mice, were given injections of CCl4 (to induce fibrosis) or acetaminophen (to induce toxic liver injury). We performed gene expression microarray, and phosphoproteomic analyses to determine mechanisms of JNK activity in hepatocytes. RESULTS: Liver samples from DILI patients contained more activated JNK, predominantly in nuclei of hepatocytes and in immune cells, than healthy tissue. Administration of acetaminophen to JnkΔhepa mice produced a greater level of liver injury than that observed in Jnk1Δhepa or control mice, based on levels of serum markers and microscopic and histologic analysis of liver tissues. Administration of CCl4 also induced stronger hepatic injury in JnkΔhepa mice, based on increased inflammation, cell proliferation, and fibrosis progression, compared to Jnk1Δhepa or control mice. Hepatocytes from JnkΔhepa mice given acetaminophen had an increased oxidative stress response, leading to decreased activation of AMPK, total protein AMPK levels, and pJunD and subsequent necrosis. Administration of SP600125 before or with acetaminophen protected JnkΔhepa and control mice from liver injury. CONCLUSIONS: In hepatocytes, JNK1 and JNK2 appear to have combined effects in protecting mice from CCl4- and acetaminophen-induced liver injury. It is important to study the tissue-specific functions of both proteins, rather than just JNK1, in the onset of toxic liver injury. JNK inhibition with SP600125 shows off-target effects.

Project description:ObjectiveGrowth differentiation factors (GDFs) and bone-morphogenic proteins (BMPs) are members of the transforming growth factor β (TGFβ) superfamily and are known to play a central role in the growth and differentiation of developing tissues. Accumulating evidence, however, demonstrates that many of these factors, such as BMP-2 and -4, as well as GDF15, also regulate lipid metabolism. GDF10 is a divergent member of the TGFβ superfamily with a unique structure and is abundantly expressed in brain and adipose tissue; it is also secreted by the latter into the circulation. Although previous studies have demonstrated that overexpression of GDF10 reduces adiposity in mice, the role of circulating GDF10 on other tissues known to regulate lipid, like the liver, has not yet been examined.MethodsAccordingly, GDF10-/- mice and age-matched GDF10+/+ control mice were fed either normal control diet (NCD) or high-fat diet (HFD) for 12 weeks and examined for changes in liver lipid homeostasis. Additional studies were also carried out in primary and immortalized human hepatocytes treated with recombinant human (rh)GDF10.ResultsHere, we show that circulating GDF10 levels are increased in conditions of diet-induced hepatic steatosis and, in turn, that secreted GDF10 can prevent excessive lipid accumulation in hepatocytes. We also report that GDF10-/- mice develop an obese phenotype as well as increased liver triglyceride accumulation when fed a NCD. Furthermore, HFD-fed GDF10-/- mice develop increased steatosis, endoplasmic reticulum (ER) stress, fibrosis, and injury of the liver compared to HFD-fed GDF10+/+ mice. To explain these observations, studies in cultured hepatocytes led to the observation that GDF10 attenuates nuclear peroxisome proliferator-activated receptor γ (PPARγ) activity; a transcription factor known to induce de novo lipogenesis.ConclusionOur work delineates a hepatoprotective role of GDF10 as an adipokine capable of regulating hepatic lipid levels by blocking de novo lipogenesis to protect against ER stress and liver injury.

Project description:AimFree fatty acid-induced lipotoxicity plays a crucial role in the progression of nonalcoholic fatty liver disease (NAFLD). In the present study we investigated the effects of a high-fat diet and free fatty acids on the autophagic process in hepatocytes in vivo and in vitro and the underlying mechanisms.MethodsLC3-II expression, a hallmark of autophagic flux, was detected in liver specimens from patients with non-alcoholic steatohepatitis (NASH) as well as in the livers of C57BL/6 mice fed a high-fat diet (HFD) up to 16 weeks. LC3-II expression was also analyzed in human SMMC-7721 and HepG2 hepatoma cells exposed to palmitic acid (PA), a saturated fatty acid. PA-induced apoptosis was detected by Annexin V staining and specific cleavage of PARP in the presence and absence of different agents.ResultsLC3-II expression was markedly increased in human NASH and in liver tissues of HFD-fed mice. Treatment of SMMC-7721 cells with PA increased LC3-II expression in time- and dose-dependent manners, whereas the unsaturated fatty acid oleic acid had no effect. Inhibition of autophagy with 3MA sensitized SMMC-7721 cells to PA-induced apoptosis, whereas activation of autophagy by rapamycin attenuated PA-induced PARP cleavage. The autophagy-associated proteins Beclin1 and Atg5 were essential for PA-induced autophagy in SMMC-7721 cells. Moreover, pretreatment with SP600125, an inhibitor of JNK, effectively abrogated PA-mediated autophagy and apoptosis. Specific knockdown of JNK2, but not JNK1, in SMMC-7721 cells significantly suppressed PA-induced autophagy and enhanced its pro-apoptotic activity; whereas specific knockdown of JNK1 had the converse effect. Similar results were obtained when HepG2 cells were tested.ConclusionJNK1 promotes PA-induced lipoapoptosis, whereas JNK2 activates pro-survival autophagy and inhibits PA lipotoxicity. Our results suggest that modulation of autophagy may have therapeutic benefits in the treatment of lipid-related metabolic diseases.

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Combined Activities of JNK1 and JNK2 in Hepatocytes Protect Against Toxic Liver Injury.

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Combined Activities of JNK1 and JNK2 in Hepatocytes Protect Against Toxic Liver Injury.

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