Mark J. Czaja

We have all learned in textbooks that “lysosomes contain hydrolases able to degrade all types of intracellular molecules which include proteases, glycosidases, nucleotidases and lipases.” To date, ...

The intracellular storage and utilization of lipids are critical to maintain cellular energy homeostasis. During nutrient deprivation, cellular lipids stored as triglycerides in lipid droplets are hydrolysed into fatty acids for energy. A second cellular response to starvation is the induction of autophagy, which delivers intracellular proteins and organelles sequestered in double-membrane vesicles (autophagosomes) to lysosomes for degradation and use as an energy source. Lipolysis and autophagy share similarities in regulation and function but are not known to be interrelated. Here we show a previously unknown function for autophagy in regulating intracellular lipid stores (macrolipophagy). Lipid droplets and autophagic components associated during nutrient deprivation, and inhibition of autophagy in cultured hepatocytes and mouse liver increased triglyceride storage in lipid droplets. This study identifies a critical function for autophagy in lipid metabolism that could have important implications for human diseases with lipid over-accumulation such as those that comprise the metabolic syndrome.

Activation of c-Jun N-terminal kinase (JNK) has been implicated as a mechanism in the development of steatohepatitis. This finding, together with the reported role of JNK signaling in the development of obesity and insulin resistance, two components of the metabolic syndrome and predisposing factors for fatty liver disease, suggests that JNK may be a central mediator of the metabolic syndrome and an important therapeutic target in steatohepatitis. To define the isoform-specific functions of JNK in steatohepatitis associated with obesity and insulin resistance, the effects of JNK1 or JNK2 ablation were determined in developing and established steatohepatitis induced by a high-fat diet (HFD). HFD-fed jnk1 null mice failed to develop excessive weight gain, insulin resistance, or steatohepatitis. In contrast, jnk2^−/− mice fed a HFD were obese and insulin-resistant, similar to wild-type mice, and had increased liver injury. In mice with established steatohepatitis, an antisense oligonucleotide knockdown of jnk1 decreased the amount of steatohepatitis in concert with a normalization of insulin sensitivity. Knockdown of jnk2 improved insulin sensitivity but had no effect on hepatic steatosis and markedly increased liver injury. A jnk2 knockdown increased hepatic expression of the proapoptotic Bcl-2 family members Bim and Bax and the increase in liver injury resulted in part from a Bim-dependent activation of the mitochondrial death pathway. Conclusion: JNK1 and JNK2 both mediate insulin resistance in HFD-fed mice, but the JNK isoforms have distinct effects on steatohepatitis, with JNK1 promoting steatosis and hepatitis and JNK2 inhibiting hepatocyte cell death by blocking the mitochondrial death pathway. (HEPATOLOGY > 2009;49:87-96.)

Oxidative stress is a common mechanism of liver injury. Recent investigations have demonstrated that oxidant-induced liver injury is mediated by the direct effects of reactive oxygen species on signal transduction pathways. Although the function of cell signaling in this form of injury is complex and likely variable depending on the type and duration of oxidative stress, common regulatory pathways of hepatocyte oxidant injury have been identified that include the mitogen-activated protein kinases extracellular signal-regulated kinase 1/2 (ERK1/2) c-Jun N-terminal kinase (JNK), and the nuclear factor κB (NF-κB) pathway. Studies in cultured hepatocyte and rodent models of oxidative stress have demonstrated that ERK1/2 typically induces resistance to oxidant stress, whereas JNK promotes cell death. The effects of NF-κB activation are more complex and cell-type specific. A further understanding of the signaling pathways that regulate oxidant-induced liver injury may suggest new therapies for hepatic diseases resulting from oxidative stress.

Oxidative stress has been implicated as the mechanism of hepatocyte injury from numerous agents. Although reactive oxygen species injure cells by the modification of critical cellular macromolecules, recent studies have demonstrated the mechanistic involvement of oxidant stress‐induced alterations in signal transduction cascades. Studies in menadione‐treated hepatocytes have demonstrated differential effects of mitogen‐activated protein kinase activation on hepatocyte death from acute oxidative stress. Activation of the extracellular signal‐regulated kinase pathway 1/2 (ERK1/2) confers hepatocyte resistance to death whereas sustained c‐Jun N‐terminal kinase (JNK)/c‐Jun/AP‐1 activation promotes apoptosis. Redundant protective signals such as the protein kinase C/protein kinase D pathways also downregulate the JNK/c‐Jun/AP‐1 cascade and provide resistance to cell death. Although ERK1/2 overactivation also acts as a protective response to chronic oxidative stress, enhanced activation of this kinase sensitizes hepatocytes to death from free fatty acids in this setting. The outcome from challenge with an oxidative stress, therefore, depends on the integration of a series of signaling cascades that both protect against and promote hepatocyte apoptosis.

Nonalcoholic fatty liver disease (NAFLD) is characterized by hepatic steatosis and varying degrees of necroinflammation. Although chronic oxidative stress, inflammatory cytokines, and insulin resistance have been implicated in the pathogenesis of NAFLD, the mechanisms that underlie the initiation and progression of this disease remain unknown. c-Jun N-terminal kinase (JNK) is activated by oxidants and cytokines and regulates hepatocellular injury and insulin resistance, suggesting that this kinase may mediate the development of steatohepatitis. The presence and function of JNK activation were therefore examined in the murine methionine- and choline-deficient (MCD) diet model of steatohepatitis. Activation of hepatic JNK, c-Jun, and AP-1 signaling occurred in parallel with the development of steatohepatitis in MCD diet–fed mice. Investigations in jnk1 and jnk2 knockout mice demonstrated that jnk1, but not jnk2, was critical for MCD diet–induced JNK activation. JNK promoted the development of steatohepatitis as MCD diet–fed jnk1 null mice had significantly reduced levels of hepatic triglyceride accumulation, inflammation, lipid peroxidation, liver injury, and apoptosis compared with wild-type and jnk2 −/− mice. Ablation of jnk1 led to an increase in serum adiponectin but had no effect on serum levels of tumor necrosis factor-α. In conclusion, JNK1 is responsible for JNK activation that promotes the development of steatohepatitis in the MCD diet model. These findings also provide additional support for the critical mechanistic involvement of JNK1 overactivation in conditions associated with insulin resistance and the metabolic syndrome. (HEPATOLOGY 2006;43:163–172.)