Hepatocyte RIP1 Contributes To The Hepatic Homeostasis In Resistance To Fasting-induced Transient Lipotoxicity

Fasting is a common stress in the process of animal evolution.Short-term starvation can cause systematic effects and changes on different tissues and organs of animals,including but not limited to changes in hormone levels,lipolysis in adipose tissue leading to a transient rise in the level of free fatty acids in plasma.In the long-term evolution,the animals have evolved the capability to resist short-term starvation stress,and adapt to environmental changes to maintain homeostasis and health.In this study,we revealed that RIP1 in hepatocytes plays an important role in resisting the lipid toxicity induced by fasting stress or other metabolic disturbances.In this study,the mice with RIP1 deficiency in hepatocytes(Rip1^Δhep)were used to construct a short-term fasting model.The results showed that in contrast to normal liver function in WT mice after the short-term fasting,Rip1^Δhep mice exhibited transient liver injury that the serum activity of transaminase was significantly elevated.Further analysis showed that in the liver tissue of Rip1^Δhepmice,short-term fasting induced cell death upregulated expression of inflammatory molecules and expression of proliferation-related molecules（Ki67 and alpha fetoprotein）.The serum concentrations of triglyceride,free fatty acids and ketone body as well as the triglyceride content in liver tissue were similar between WT and Rip1^Δhep mice,suggesting that RIP1 deficiency in hepatocytes did not affect the metabolism processes including lipolysis in adipose tissue and fatty acid beta oxidation induced by fasting.Next,primary hepatocytes were isolated from both WT and Rip1^Δhep mice.We also used specific siRNA transfection to knockdown the expression of RIP1 in AML12hepatocytes.Then the primary hepatocytes or AML12 cells were treated with saturated fatty acid（palmitic acid）in vitro.Our results indicated that RIP1^-/-or RIP1-knockdown hepatocytes were more sensitive to the lipid toxicity by saturated fatty acid.It is known that saturated fatty acids result in lipid toxicity through inducing endoplasmic reticulum（ER）stress.Pretreatment with ER stress inhibitor can effectively alleviate the liver injury induced by fasting in Rip1^Δhepmice,suggesting that ER stress was involved in the fasting-induced liver injury in Rip1^Δhepmice.Further more,WT and Rip1^Δhepmice were fed with high-fat diet for 6 days to simulate the short-term lipid burdon in liver as in metabolic disturbance.Similar to the increase of lipid in liver induced by starvation,short-term high-fat diet feeding induced upregulation of inflammatory molecules and ER stress-related markers in liver tissues of Rip1^Δhep mice but not in WT mice.In conclusion,using both cellular and animal models,our study revealed a critical role of RIP1 in maintaining hepatic homeostasis against the transient fatty acid impulse during fasting or by high-fat diet feeding.